BR0209033A

Core-glycosylated hcv envelope proteins

Abstract

"CONSTRUCTIONS AND METHODS FOR THE EXPRESSION OF RECOMBINANT HCV WRAP PROTEINS". The present invention relates to vectors and methods for the efficient expression of HCV envelope proteins in eukaryotic cells. More particularly, said vectors comprise the coding sequence for an avian lysozyme signal peptide, or a functional equivalent thereof, attached to an HCV envelope protein, or a part thereof. Said avian lysozyme signal peptide is efficiently removed when the protein comprising said avian lysozyme signal peptide attached to an HCV envelope protein or a part thereof is expressed in a eukaryotic cell. Suitable eukaryotic cells include yeast cells, such as Saccharomyces or Hansenula cells.

BR0209033A, drawing sheet 1 — Sheet 1 of 54

Term

Term ended

Projected expiry passed 24 April 2022, 4.4 years ago.

23 claims: 14 independent, 9 dependent

1
REIVINDICAÇÕES 1. Ácido nucléico recombinante, CARACTERIZADO por compreender uma sequência de nucleotídeos codificando uma proteína compreendendo um peptídeo líder de lisozima aviário ou um seu equivalente funcional unido a uma proteína de envoltório de HCV ou uma parte da mesma.
2
Ácido nucléico recombinante, de acordo com a reivindicação 1, CARACTERIZADO pelo fato de que a dita proteína é distinguida pela estrutura CL-[(Al) a - (PSI) b (A2 ) c ]-HCVENV-[ (A3 ) d - (PS2) e - (A4) f ] em que:CL é um peptídeo líder de lisozima aviário ou um seu equivalente funcional, Al, A2, A3 e A4 são peptídeos adaptadores que podem ser diferentes ou iguais, PS1 e PS2 são sítios de processamento que podem ser diferentes ou iguais, HCVENV é uma proteína de envoltório de HCV ou uma parte da mesma, a, b, c, d, e e f são 0 ou 1, e em que, opcionalmente, Al e/ou A2 são parte de PS1 e/ou em que A3 e/ou A4 são parte de PS2.
3
Ácidos nucléicos recombinantes, de acordo com a reivindicação 1 ou 2, CARACTERIZADOS por adicionalmente compreender elementos reguladores que permitem a expressão da dita proteína em uma célula hospedeira eucariótica.
4
Ácido nucléico recombinante, de acordo com quaisquer das reivindicações 1 a 3, CARACTERIZADO pelo fato • · de que o peptídeo líder de lisozima aviário CL tem uma seqüência de aminoácidos definida por SEQ ID NO:1.
5
Ácido nucléico recombinante, de acordo com as reivindicações 2 ou 3, CARACTERIZADO pelo fato de que A tem uma sequência de aminoácidos escolhida a partir de SEQ ID NOs :63-55, 70-72 e 74-82, em que PS tem uma sequência de aminoácidos escolhida a partir de SEQ ID NOs:66-68 e 83-84 ou em que PS é um sítio dibásico tal como Lys-Lys, Arg-Arg, Lys-Arg e Arg-Lys ou um sítio monobásico tal como Lys, e em que HCVENV é escolhida a partir de SEQ ID NOs :85-98 e seus fragmentos.
6
Vetor, CARACTERIZADO por compreender o ácido nucléico recombinante de acordo com quaisquer das reivindicações 1 a 5.
7
Vetor, de acordo com a reivindicação 6, CARACTERIZADO pelo fato de que é um vetor de expressão.
8
Vetor, de acordo com a reivindicação 6 ou 7, CARACTERIZADO pelo fato de que é um vetor que se replica autonomamente ou um vetor integrativo.
9
Vetor, de acordo com quaisquer das reivindicações 6 a 8, CARACTERIZADO pelo fato de que é escolhido a partir de SEQ ID NOs:20, 21, 32, 35, 36, 39, 40, 49 e 50.
10
Célula hospedeira, CARACTERIZADA por compreender o ácido nucléico recombinante de acordo com quaisquer das reivindicações 1 a 5 ou o vetor de acordo com quaisquer das reivindicações 6 a 9.
11
Célula hospedeira, de acordo com a reivindicação 10, CARACTERIZADA pelo fato de que é capaz de expressar • · a proteína compreendendo um peptídeo líder de lisozima aviário ou um equivalente funcional do mesmo unido a uma proteína de envoltório de HCV ou uma parte da mesma.
12
Célula hospedeira, de acordo com a reivindicação 10 ou 11, CARACTERIZADA pelo fato de que é capaz de expressar a proteína distinguida pela estrutura CL-[(Al) a (PSI) b - (A2 ) c ]-HCVENV-[ (A3) d - (PS2) e - (A4) f ] em que:CL é um peptídeo líder de lisozima aviário ou um seu equivalente funcional, Al, A2, A3 e A4 são peptídeos adaptadores que podem ser diferentes ou iguais, PS1 e PS2 são sítios de processamento que podem ser diferentes ou iguais, HCVENV é uma proteína de envoltório de HCV ou uma parte da mesma, a, b, c, d, e e f são 0 ou 1, e em que, opcionalmente, Al e/ou A2 são parte de PS1 e/ou em que A3 e/ou A4 são parte de PS2.
13
Célula hospedeira, de acordo com quaisquer das reivindicações 10 a 12, CARACTERIZADA pelo fato de que é capaz de translocar a proteína CL-[(Al) a - (PSI) b - (A2) c ]HCVENV-[ (A3 ) d - (PS2) e - (A4) f ] para o retículo endoplasmático com a remoção do peptídeo CL em que a dita proteína e o dito peptídeo CL são derivados da proteína distinguida pela estrutura CL-[(Al) a - (PSI) b - (A2 )J-HCVENV-[(A3) d - (PS2) e (A2) J-HCVENV-[ (A3) d (A4) f ] em que:• · ···· ···· ·· ·· CL é um peptídeo líder de lisozima aviário ou um seu equivalente funcional, Al, A2, A3 e A4 são peptídeos adaptadores que podem ser diferentes ou iguais, PS1 e PS2 são sítios de processamento que podem ser diferentes ou iguais, HCVENV é uma proteína de envoltório de HCV ou uma parte da mesma, a, b, c, d, e e f são 0 ou 1, e em que, opcionalmente, Al e/ou A2 são parte de PS1 e/ou em que A3 e/ou A4 são parte de PS2.
14
Célula hospedeira, de acordo com quaisquer das reivindicações 10 a 13, CARACTERIZADA pelo fato de que é capaz de processar os sítios de processamento PS1 e/ou PS2 na dita proteína translocada para o retículo endoplasmático.
15
Célula hospedeira, de acordo com quaisquer das reivindicações 10 a 13, CARACTERIZADA pelo fato de que é capaz de N-glicosilar a dita proteína translocada para o retículo endoplasmático.
16
Célula hospedeira, de acordo com a reivindicação 14, CARACTERIZADA pelo fato de que é capaz de Nglicosilar a dita proteína translocada para o retículo endoplasmático e processada nos ditos sítios PS1 e/ou PS2.
17
Célula hospedeira, de acordo com quaisquer das reivindicações 10 a 16, CARACTERIZADA pelo fato de que é uma célula eucariótica.
18
Célula hospedeira, de acordo com quaisquer das reivindicações 10 a 16, CARACTERIZADA pelo fato de que é uma célula fúngica.
19
Célula hospedeira, de acordo com quaisquer das 5 reivindicações 17, CARACTERIZADA pelo fato de que é uma célula de levedura.
20
Célula hospedeira, de acordo com a reivindicação 19, CARACTERIZADA pelo fato de que é uma célula de Saccharomyces , tal como uma célula de Saccharomyces cerevisiae, 10 uma célula de Saccharomyces kluyveri, ou uma célula de Saccharomyces uvarum, uma célula de Schizosaccharomyces, tal como uma célula de Schizosaccharomyces pombe, uma célula de Kluyveromyces, tal como uma célula de Kluyveromyces lactis, uma célula de Yarrowia, tal como uma célula de Yarrowia li15 polytica, uma célula de Hansenula, tal como uma célula de Hansenula polymorpha, uma célula de Pichia, tal como uma célula de Pichia pastoris, uma célula de Aspergillus, uma célula de Neurospora, tal como uma célula de Neurospora crassa, ou uma célula de Schwanniomyces, tal como uma célula de 20 Schwanniomyces occidentalis, ou uma célula mutante derivada de quaisquer das mesmas.
21
Método para produzir uma proteína de envoltório de HCV ou parte da mesma em uma célula hospedeira, o dito método sendo CARACTERIZADO pelo fato de que compreende 25 transformar a dita célula hospedeira com o ácido nucléico recombinante de acordo com quaisquer das reivindicações 1 a 5 ou com o vetor de acordo com quaisquer das reivindicações 6 a 9, e em que a dita célula hospedeira é capaz de expres6 • · sar uma proteína compreendendo o peptídeo líder de lisozima aviário ou um equivalente funcional do mesmo unido a uma proteína de envoltório de HCV ou uma parte da mesma.
22
Método para produzir uma proteína de envoltó5 rio de HCV ou parte da mesma em uma célula hospedeira, o dito método sendo CARACTERIZADO pelo fato de que compreende transformar a dita célula hospedeira com o ácido nucléico recombinante de acordo com quaisquer das reivindicações 1 a 5 ou com o vetor de acordo com quaisquer das reivindicações 10 6 a 9, e em que a dita célula hospedeira é capaz de expressar a proteína distinguida pela estrutura CL-[(Al) a - (PSI) b - (A2 ) c ]-HCVENV-[ (A3 ) d - (PS2) e - (A4) f ] em que:CL é um peptídeo líder de lisozima aviário ou um 15 seu equivalente funcional, Al, A2, A3 e A4 são peptídeos adaptadores que podem ser diferentes ou iguais, PS1 e PS2 são sítios de processamento que podem ser diferentes ou iguais, 22, CARACTERIZADO pelo fato de que a dita célula hospedeira é capaz de translocar a proteína CL-[(Al) a - (PSI) b - (A2)JHCVENV-[ (A3) d (PS2) (A4) f ] para o retículo endoplasmático com a remoção do peptídeo CL em que a dita proteína e o dito peptídeo CL são derivados da proteína distinguida pela estrutura CL-[(Al) a - (PSI) b - (A2 ) c ]-HCVENV-[(A3) d - (PS2) e (A4) f ] 5 em que: CL é um peptídeo líder de lisozima aviário ou um seu equivalente funcional, Al, A2, A3 e A4 são peptídeos adaptadores que podem ser diferentes ou iguais, 10 PS1 e PS2 são sítios de processamento que podem ser diferentes ou iguais, HCVENV é uma proteína de envoltório de HCV ou uma parte da mesma, a, b, c, d, e e f são 0 ou 1, e 15 em que, opcionalmente, Al e/ou A2 são parte de PS1 e/ou em que A3 e/ou A4 são parte de PS2. 24. Método, de acordo com quaisquer das reivindicações 21 a 23, CARACTERIZADO pelo fato de que a dita célula hospedeira é capaz de processar os sítios de processamento 20 PS1 e/ou PS2 na dita proteína translocada para o retículo endoplasmático. 25. Método, de acordo com quaisquer das reivindicações 21 a 23, CARACTERIZADO por adicionalmente compreender o processamento in vitro dos sítios de processamento PS1 25 e/ou PS2. 26. Método, de acordo com quaisquer das reivindicações 21 a 23, CARACTERIZADO pelo fato de que a dita célula hospedeira é capaz de N-glicosilar a dita proteína translocada para o retículo endoplasmático. 27. Método, de acordo com a reivindicação 24, CARACTERIZADO pelo fato de que a dita célula hospedeira é capaz de N-glicosilar a dita proteína translocada para o retículo endoplasmático e processada nos ditos sítios PS1 e/ou PS2 . 28. Método, de acordo com quaisquer das reivindicações 21 a 27, CARACTERIZADO pelo fato de que a dita célula hospedeira é uma célula eucariótica. 29. Método, de acordo com quaisquer das reivindicações 21 a 27, CARACTERIZADO pelo fato de que a dita célula hospedeira é uma célula fúngica. 30. Método, de acordo com quaisquer das reivindicações 21 a 27, CARACTERIZADO pelo fato de que a dita célula hospedeira é uma célula de levedura. 31. Método, de acordo com quaisquer das reivindicações 21 a 27, CARACTERIZADO pelo fato de que a dita célula hospedeira é uma célula de Saccharomyces, tal como uma célula de Saccharomyces cerevisiae, uma célula de Saccharomyces kluyverí, ou uma célula de Saccharomyces uvarum, uma célula de Schizosaccharomyces, tal como uma célula de Schizosaccharomyces pombe, uma célula de Kluyveromyces, tal como uma célula de Kluyveromyces lactis, uma célula de Yarrowia, tal como uma célula de Yarrowia lipolytica, uma célula de Hansenula, tal como uma célula de Hansenula polymorpha, uma célula de Pichia, tal como uma célula de Pichia pastoris, uma célula de Aspergillus, uma célula de Neurospora, tal como uma célula de Neurospora crassa, ou uma célula de Schwanniomyces, tal como uma célula de Schwanniomyces occidentalis, ou uma célula mutante derivada de quaisquer das mesmas. 32. Método, de acordo com quaisquer das reivindicações 21 a 27, CARACTERIZADO por adicionalmente compreender a cultivação das ditas células hospedeira em um meio adequado para obter a expressão da dita proteína. 33. Método, de acordo com a reivindicação 32, CARACTERIZADO por adicionalmente compreender o isolamento da proteína expressada a partir de uma cultura das ditas células hospedeiras, ou a partir das ditas células hospedeiras. 34. Método, de acordo com a reivindicação 33, CARACTERIZADO pelo fato de que a dita etapa de isolamento envolve a lise das ditas células hospedeiras na presença de um agente caotrópico. 35. Método, de acordo com a reivindicação 33 ou 34, CARACTERIZADO pelo fato de que os grupos tiol da cisteína nas proteínas isoladas são quimicamente modificados e em que a dita modificação química é reversível ou irreversível. 36. Método, de acordo com quaisquer das reivindicações 32 a 35, CARACTERIZADO por envolver a cromatografia por afinidade de heparina. Spel Notl Pstl Accl Sall Ndel Saci BstXl Nsil FIGURA 1 FIGURA 2 FIGURA 3 Tth111! FIGURA 4 FIGURA 5 Ecl136ll Saci BamHI FIGURA 6 ECI136II Saci BamHI Muni Bsgl Nael NgoMIV BsaAI Aatll EcoO109l BseRI Hincll Sall Sphl Hindlll PshAI BstAPI Ndel Btgl Sacll EcoRV Xcml Apal PspOMI Psil FIGURA 7 FIGURA 8 FIGURA 9 FIGURA 10 TthWI FIGURA 11 FIGURA 12 FIGURA 13 Tth111l BamHI Ncol Sbfl FIGURA 14 Ecl136ll Hindlll BstAPI Ndel FIGURA 15 Nael NgoMIV BamHI BsaAI TthWI Muni Bsgl Nael Ncol Sbfl FIGURA 16 Ecl136ll Saci BspLUHI Muni Bsgl Nael NgoMIV BsaAI PshAI Aatll EcoO109l BseRI Hincll Sall Sphl Hindlll BstAPI Ndel Dsal Sacll EcoRV Xcml Apal PspOMI Psil FIGURA 17 FIGURA 18 Xbal Stul Accl Sall Kpnl Bglll FIGURA 19 FIGURA 20 Pvull FIGURA 21 Clal Hindlll Pstl Ncol Seal Stul Hindlll EcoRI EcoRI Smal Pvull BstEII Bglll Hindlll Smal Hindlll BamHI Xbal Xbal Xbal Hindlll Stul Pvul Pvull Pvull Hindlll EcoRI Stul EcoRI BstEII Pvul FIGURA 22 Kpnl Apal Aval Xhol Accl Hincll Sall Clal Hindlll Pstl Pacl FIGURA 23 Kpnl Apal A\a! Xhol Accl Hincll Sall Clal Hindlll EcoRV EcoRI Pstl Aval Smal BamHI Spel Xbal Notl BstXI i EcoRI I BamHI .Xbal Xbal Hindlll BamHI Accl Hindlll AlwNI pYIG5HCCL-22aH6 ICCG2424 6104 bps Notl Apal Apal Sphl Xhol Accl FIGURA 24 FIGURA 25 FIGURA 26 FIGURA 27 Clal Hindlll Pstl Ncol Aval Accl Aval++ Eco47lll++ Accl Eco47lll ECO47III Xbal Eco47lll Eco47lll Xbal Eco47lll Eco47lll Accl Hindlll Stul Pvul Spel Pvull ECO47III Pvull Spel Hindlll Accl ++ Accl Stul EcoRI Kpnl Clal ++Pvul ++ Eco47lll ++Aval Eco47lll Spel Accl Hindlll EcoRI Clal EcoRI Ncol Seal Accl Apal Stul Hindlll Bglll++ Pvull++ Aval++ Spel Saci PAGXbal LER Xbal BamHI ECO47III Accl Sall Eco47lll Aval Eco47lll Pvull Accl FIGURA 28 FIGURA 29 FIGURA 30 FIGURA 31 FIGURA 32 FIGURA 33 Pacl Pstl Hindlll Clal Accl Hincll Sall Notl FIGURA 34 FIGURA 35 FIGURA 36 mAU FIGURA 37 FIGURA 38 kDa FIGURA 39 I FIGURA 40 kDa
23
2 3 4 5 6 7 • rx 2 3 4 5 6 7 kDa 18 — _ 14 —'-¾¾ FIGURA 41 1000 1200 1400 1600 mL FIGURA 42 FIGURA 43 FIGURA 44 Μ 1 2 3 4 kDa 18 _ 14— FIGURA 45 FIGURA 46 Ο 20 40 60 mL FIGURA 47 FIGURA 48 mAU 50,0 40,0 30,0 20,0 10,0 ο,ο Frações de VLP-reunidas -10,0 FIGURA 49 FIGURA 50 I 1000000η 10000010000 1000000η 100000▼ ▼ ▼ ▼ 10000-J-Μ FIGURA 51 1000000η 100000- ▼ 10000- 1 S 1000000η 100000▼ ▼ ▼ ▼ ▼ ▼ 10000-1A FIGURA 52 100000 10000 1000 100 100000 10000 1000 100 FIGURA 53 FIGURA 54

Independent claims23

4,280 paragraphs in 66 sections, as filed

(54) Title: CONSTRUCTIONS AND METHODS FOR THE EXPRESSION OF RECOMBINANT HCV WRAP PROTEINS (30) Unionist Priority: 24/04/2001 ep O187OO88.O; 07/17/2001 US 60 / 305,604 (71) Depositor (s): Innogenetics NV (BE) (72) Inventor (s): Erwin Sablon, Annie Van Broekhoven, Alfons Bosman, Erik Depla, Geert Deschamps (74) Attorney: Nellie Anne Daniel Shores (86) International Order: pct BE02 / 00062 of 04/24 / 2002 (87) International Publication: wo 02/085932 of 10/31/2002 (57) Abstract: CONSTRUCTIONS AND METHODS FOR THE EXPRESSION OF RECOMBINANT HCV WRAP PROTEINS. The present invention relates to vectors and methods for the efficient expression of HCV envelope proteins in eukaryotic cells. More particularly, said vectors comprise the coding sequence for an avian lysozyme desinalde peptide, or an equivalent equivalent thereof, attached to an HCV envelope protein, or a part thereof. Said avian lysozyme signal peptide is efficiently removed when the protein comprising said avian lysozyme signal peptide attached to an HCV envelope protein or a part thereof is expressed in a eukaryotic cell. Suitable eukaryotic cells include yeast cells, such as Saccharomyces or Hansenula cells.

CONSTRUCTIONS, AND METHODS 'FOR THE EXPRESSION OF RECOMBINANT HCV WRAP PROTEINS

FIELD OF. INVENTION

The present invention relates to the general field of expression of recombinant proteins. More particularly, the present invention relates to the expression of hepatitis C virus envelope proteins in a eukaryote, such as yeast. Constructions and methods for the expression of viral envelope proteins, glycosylated in the yeast nucleus, are disclosed.

BACKGROUND GIVES INVENTION

Hepatitis C virus (HCV) infection is a major health problem in both developed and developing countries. It is estimated that about 1 to 5% of the world's population is affected by the virus. HCV infection appears to be the most important cause of hepatitis associated with transfusion and often progresses to chronic liver damage. In addition, there is evidence implicating the. HCV in the induction of hepatocellular carcinoma. Consequently, there is a high demand for safe diagnostic methods and effective therapeutic agents. Sensitive and specific methods of examining blood products contaminated with HCV and improved methods for cultivating HCV are also needed.

HCV is a positive 9,600 base RNA strand virus, which encodes a single polyprotein precursor of about 3000 amino acids. The proteolytic divage of the precursor linked to joint and subsequent translation modifications has been shown to result in at least three structural and six non-structural proteins. Based on sequence homology, structural proteins have been functionally designated as a single-core protein and two envelope glycoproteins: E1 and E2. The El protein consists of 192

<td>amino acids</td><td>and contains 5 to</td><td>6 sites</td><td>in</td><td>N-glycosylation,</td>

<td>depending on</td><td>of the HCV genotype</td><td>. The protein</td><td>E2</td><td>consists of 363</td>

<td colspan="2">to 370 amino acids and contains 9</td><td>to 11 sites</td><td>in</td><td>N-glycosylation,</td>

<td>depending on</td><td>of the genotype of</td><td>HCV (how much</td><td>The</td><td>reviews, see:</td>

Major, ME and Feinstone, SM 1997, Maertens, G. and Stuyver, L. 1997). The El protein contains several variable domains. (Maertens, G. and Stuyver, L. 1997). The E2 protein contains three hypervariable domains, of which the main domain is located at the N end of the protein (Maertens, G. and Stuyver, L. 1997). HCV glycoproteins are located predominantly in the ER, where they are modified and grouped into oligomeric complexes.

In eukaryotes, sugar residues are commonly linked to four different amino acid residues. These amino acid residues are classified as linked to O (serine, threonine, and hydroxylysine) and linked to N (asparagine). O-linked sugars are synthesized in the Golgi or simple Endoplasmic Reticulum (ER) from nucleotide sugars. N-linked sugars are synthesized from a common precursor, and subsequently processed. H.CV wrap proteins are believed to be glycosylated in N. It is known in the art that the addition of N-linked carbohydrate chains is important for stabilizing the duplication of intermediates and thus for efficient duplication, the prevention of bad duplication and degradation in the endoplasmic reticulum, oligomerization, biological activity, and transport of glycoproteins (see reviews by Rose, JK and Doms, RW 1988, Doms, RW et al. 1993, Helenius, A. 1994)). The tripeptide sequences Asn-X-Ser and Asn-X-Thr (where X can be any amino acid) on the polypeptides are the sites. of consensus for the binding of N-linked oligosaccharides After the addition of N-linked oligosaccharides to the polypeptide, the oligosaccharide is further processed to the complex type (containing N-acetylglycosamine, mannose, fucose, galactose and sialic acid) or even to the type with high mannose content (containing N-acetylglycosamine and mannose). HCV wrap proteins are believed to be of the high mannose type. The biosynthesis of the N-linked oligosaccharide in yeast is very different from biosynthesis in mammalian cells. In yeast, the oligosaccharide chains are elongated in the Golgi through the addition, in stages, of mannose, resulting in elaborated structures with a high mannose content, referred to as hyperglycosylation. In contrast, proteins expressed in prokaryotes are never glycosylated.

So far, vaccination against the disease has been proven to be the most cost-effective and efficient method for disease control. Despite the promising results, efforts to develop an effective vaccine against

HCV, however, have been plagued with difficulties. A sine qua non condition for vaccines is the induction of an immune response in patients. Consequently, HCV antigenic determinants must be identified, and administered to patients in an appropriate setting. Antigenic determinants can be divided into at least two forms, ie linear and conformational epitopes. Conformational epitopes result from the duplication of a molecule in a three-dimensional space, including joint and subsequent translation modifications, such as glycosylation. In general, it is believed that conformational epitopes will make the most effective vaccines, since they represent epitopes that resemble HCV epitopes similar to native ones, and which can be better conserved than the true linear amino acid sequence. Therefore, the final degree of glycosylation of HCV envelope proteins is of the utmost importance to generate HCV antigenic determinants similar to native ones. However, there are apparently insurmountable problems with the cultivation of HCV, which result in only small amounts of virions. In addition, there are vast problems with the expression and purification of recombinant proteins, which result in low amounts of proteins, hyperglycosylated proteins, or proteins that are not glycosylated.

In order to obtain the glycosylation of an expressed protein, said protein needs to be targeted to the endoplasmic reticulum (ER). This process requires the presence of a pre-pro- or post-sequence, also known as a signal peptide or leader peptide, at the amino-terminus of the expressed protein. With the translocation of the protein to the ER lumen, the pre-sequence is removed by means of a signal peptidase complex. A large number of pre-pro and pre-sequences are currently known in the art. These include the S.cerevisiae α crossing factor leader (pre-pro; aMF or MFa), the leading sequence of Carcinus maenas · hyperglycemic hormone (pre; CHH), the leading amylase sequence of S. occidentalis (pre ; Amyl), the leading sequence of S. occidentalis Gaml glycoamylase (pre; Gaml), the leading sequence of fungal phytase (pre; Phy5),. the Pichia pastoris' acid phosphatase leader sequence (pre; phol), the yeast 3 protease aspartic signal peptide (pre; YAP3), the mouse salivary amylase signal peptide (pre) and the leader sequence of chicken lysozyme (pre; CL).

The CHH leader has been linked with hirudin and G-CSF (granulocyte colony stimulating factor) and the expression of the CHH-hirudin and CHH-G-CSF proteins in Hansenula polymorpha results in the correct removal of the leader sequence (Weydemann, U. et al 1995, Fischer et al WO00 / 40727). The leading sequence of chicken lysozyme has been fused to human interferon a2b (IFNa2b), human serum albumin and human lysozyme or 1,4-β-Νacetylmuramidase and has been expressed in S. cerevisiae (Rapp at GenBank accession number AF405538, Okabayashi, K. et al. 1991, by Baetselier et al. in EP0362183, Oberto and Davison in

ΕΡ0184575). Mustilli and collaborators (Mustilli, AC et al. 1999) have used the leading Kluyveromyces lactis exterminating toxin peptide for HCV E2 expression in S. cerevisiae and K. lactis.

HCV wrap proteins have been produced by recombinant techniques in Escherichia coli, insect cells, yeast cells and mammalian cells. However, expression in higher eukaryotes has been characterized by the difficulty of obtaining large amounts of antigens for the final production of the vaccine. Expression in prokaryotes, such as E. coli, results in HCV envelope proteins that are not glycosylated. The expression of HCV wrap proteins in yeast resulted in hyperglycosylation. As already demonstrated in WO

96/04385, the expression of HCV E2 wrap protein in

Saccharomyces cerevisiae results in proteins that are densely glycosylated. This hyperglycosylation results in the shielding of protein epitopes. Although Mustilli and collaborators (Mustilli, AC 1999) claim that the expression of HCV E2 in S. cerevisiae results in the glycosylation of the nucleus, the analysis of the intracellularly expressed material demonstrates that part of it is at least hyperglycosylated, although the correct processing of the rest of this material has not been shown. The need for HCV wrap proteins derived from an intracellular source is widely accepted (WO 96/04385 for Maertens et al. And Heile,

JM et al. 2000). This need is further exemplified by the unsatisfactory reactivity of E2 derived from yeast secreted with chimpanzee sera immunized with E2 proteins derived from mammalian cell cultures, as shown in Figure 5 by Mustilli et al. (Mustilli, ACe col. 1999). This is further documented by Rosa and colleagues (Rosa, D. and col. 1996) who show that immunization. with yeast-derived HCV wrap proteins cannot protect itself from provocation.

Consequently, there is a need for efficient expression systems that result in large and cost-effective amounts of proteins, and in particular, such systems are needed for the production of HCV wrap proteins. If a pre- or pre-pre-sequence is used to target the protein of interest to the ER, then the efficiency of the expression system is, among others, dependent on the efficiency and fidelity with which the pre- and pre-sequences are removed from the protein of interest.

SUMMARY OF THE INVENTION

A first aspect of the present invention relates to recombinant nucleic acids comprising a nucleotide sequence encoding a protein comprising an avian lysozyme leader peptide, or a functional equivalent, attached to an HCV envelope protein, or a part thereof . More specifically, said protein is distinguished by the CL - [(Al) structureThe - (PSl)B ~ (A2)ç] -HCVENV- [(A3)d - (PS2)and (A4)f] on what:

CL is a leading avian lysozyme peptide or functional equivalent,

A1, A2, A3 and A4 are adapter peptides that can be different or the same, '5 PS1 and PS2 are processing sites that can be different or the same,

HCVENV is an HCV wrap protein or a part thereof, az b, c, d, e and f are 0 or 1, and where, optionally, Al and / or A2 are part of PS1 and / or where A3 and / or A4 are part of PS2.

The recombinant nucleic acids according to the invention can additionally comprise regulatory elements that allow the expression of said protein in a eukaryotic host cell.

Another aspect of the invention relates to a recombinant nucleic acid according to the invention, which is comprised in a vector. The dictum vector can be an expression vector and / or an autonomously replicating vector or an integrative vector.

A further aspect of the invention relates to a host cell harboring a nucleic acid. recombinant according to the invention or a vector according to the invention. More particularly, said host cell is capable of expressing the protein comprising an avian lysozyme leader peptide, or a functional equivalent thereof, attached to an HCV envelope protein, or a part thereof. More specifically, said protein is distinguished by the structure 'CL - [(Al)The - (PSl) b (A2) ç] -HCVENV- [(A3) d (PS2)and (Á4) £] on what:

CL is. a leading avian lysozyme peptide or functional equivalent,

A1, A2, A3 and A4 are adapter peptides that can be different or the same,

PS1 and PS2 are processing sites that can be different or the same.

HCVENV is an HCV wrap protein or a part thereof, a, b, c, d, and f are 0 or 1, and where, optionally, Al and / or A2 are part of PS1 and / or where A3 and / or A4 are part of PS2.

The host cell according to the invention may be able to remove the avian lysozyme leading peptide with high efficiency and fidelity and may be able to process the PS1 and PS2 processing sites on said translocated protein to the endoplasmic reticulum. Said host cell may additionally be able to Nglycosylate said translocated protein to the endoplasmic reticulum or said translocated protein to the endoplasmic reticulum and processed at said PS1 and / or PS2 sites. The host cell can be a eukaryotic cell, such as a yeast cell.

A further aspect of the invention relates to a method for producing an HCV envelope protein, or part of it, in a host cell, said method comprising transforming said host cell with a recombinant nucleic acid according to invention or a vector according to the invention, and wherein said host cell is capable of expressing a protein comprising the avian lysozyme leader peptide, or a functional equivalent thereof, attached to an HCV wrap protein, or a part of it. More particularly, said protein is distinguished by the CL - [(Al) structureThe - (PSl)B (A2)ç] -HCVENV - [(A3)d - (PS2)and (A4)f] io where:

CL is a leading avian lysozyme peptide or functional equivalent,

A1, A2, A3 and A4 are adapter peptides that can be different or the same,

PS1 and PS2 are processing sites that can be different or the same,

HCVENV is an HCV wrap protein or a part thereof, a, b, c, d, and f are 0 or 1, and where, optionally, Al and / or A2 are part of PS1 and / or where A3 and / or A4 are part of PS2.

The method according to the invention may further comprise culturing said host cells in a suitable medium to obtain expression of said protein, isolating the expressed protein from a culture of said host cells, or from said host cells . Said isolation may include one or more of (i) lysis of said host cells in the presence of a chaotropic agent, (ii.) Modification. chemistry of the thiol groups of cysteine in isolated proteins, wherein said chemical modification can be. reversible or irreversible and (iii) heparin affinity chromatography.

PICTURE'S DESCRIPTION

Figure 1. Schematic map of the vector pGEMT-ElsH6RB, which has the sequence as defined in SEQ ID N0: 6.

'· Figure 2. Schematic map of the vector pCHH-Hir, which has the sequence as defined in SEQ ID NO: 9.

Figure 3. Schematic map of the vector pFPMTl21, which has the sequence as defined in SEQ ID NO: 12.

Figure 4. Schematic map of the vector pFPMT-CHH-ElH6, which has the sequence as defined in SEQ ID NO: 13.

Figure 5. Schematic map of the vector pFPMT-MFa-El15 H6, which has the sequence as defined in SEQ ID NO: 16.

Figure 6. Schematic map of the vector pUC18-FMD-MFaE1-H6, which has the sequence as defined in SEQ ID NO: 17.

Figure 7. Schematic map of the vector pUC18-FMD-CLE1-H6, which has the sequence as defined in SEQ ID NO: 20.

Figure 8. Schematic map of the vector pFPMT-CL-ElH6, which has the sequence as defined in SEQ ID NO: 21.

Figure 9. Schematic map of the vector pSP72E2H6, which has the sequence as defined in SEQ ID NO: 22.

Figure 10. Schematic map of the vector pMPT121, which 25 has the sequence as defined in SEQ ID NO: 23.

Figure 11. Schematic map of the vector pFPMT-MFa-E2H6, which has the sequence as defined in SEQ ID NO: 24.

Figure 12. Schematic map of the vector pMPT-MFa-E2H6, which has the sequence as defined in SEQ ID NO: 25.

Figure 13. Schematic map of the vector pMF30, which has the sequence as defined in SEQ ID NO: 28.

'5 Figure 14. Schematic map of the vector pFPMT-CL-E2H6, which has the sequence as defined in SEQ ID NO: 32.

Figure 15. Schematic map of the vector pUC18-FMD-CLEl, which has the sequence as defined in SEQ ID NO: 35.

Figure 16. Schematic map of the vector pFPMT-CL-El, io that has the sequence as defined in SEQ ID NO: 36.

Figure 17. Schematic map of the vector pUC18-FMD-CLH6-E1-K-H6, which has the sequence as defined in SEQ ID.

NO: 39.

Figure 18. Schematic map of the vector pFPMT-CL-H615 K-El, which has the sequence as defined in SEQ ID NO: 40.

Figure 19. Schematic map of the vector pYIG5, which has the sequence as defined in SEQ ID NO: 41.

Figure 20. Schematic map of the vector pYIG5ElH6, which has the sequence as defined in SEQ ID NO: 42.

Figure 21. Schematic map of the vector pSYl, which has the sequence as defined in SEQ ID NO: 43.

Figure 22. Schematic map of the vector pSYlaMFElsH6a, which has the sequence as defined in SEQ ID NO: 44.

Figure 23. Schematic map of the vector pBSK-E2sH6, 25 that has the sequence as defined in SEQ ID NO: 45.

Figure 24. Schematic map of the vector pYIG5HCCL22aH6, which has the sequence as defined in SEQ ID NO: 46.

Figure '25. Schematic map of the vector pYYIGSE2H6, which has the sequence as defined, in SEQ ID NO: 47.

Figure 26. Schematic map of the pYIG7 vector, which has the sequence as defined in SEQ ID NO: 48.

Figure 27. Schematic map of the vector pYIG7El, which has the sequence as defined in SEQ ID NO: 49.

Figure 28. Schematic map of the vector pSYlYIG7El, which has the sequence as defined in SEQ ID NO: 50.

Figure 29. Map, schematic of the vector pPICZalfaA, io it has. the sequence as defined in SEQ ID NO: 51.

Figure 30. Schematic map of the vector pPICZalfaD ',. which has sequence as defined in SEQ ID NO: 52.

Figure 31. Schematic map of the vector pPICZalfaE ', which has the sequence as defined in SEQ ID NO: 53.

Figure 32. Schematic map of the vector pPICZalfaD'ElsH6, which has the sequence as defined in SEQ ID NO: 58.

Figure 33. Schematic map of the vector pPICZalfaE'ElsH6, which has the sequence as defined in SEQ ID NO: 59.

Figure 34. Schematic map of the vector pPICZalfaD'E2sH6, which has the sequence as defined in SEQ ID NO: 60.

Figure 35. Schematic map of the vector pPICZalfaE'E2sH6, which has the sequence as defined in SEQ ID NO: 61.

Figure 36. Schematic map of the vector pUC18MFa, which has the sequence as defined in SEQ ID NO: 62.

Figure 37. Chromatography elution profile by size exclusion of the IMAC-purified E2-H6 protein, expressed from Hansenula polymorpha expressing MFaE2-H6 (see Example 15). The X axis indicates the elution volume (in ml). The vertical lines through the elution profile indicate the fractions collected. Pl = fractions pooled 4 to 9, P2 = fractions pooled 30 to 35, and P3 = fractions pooled 37 to 44. The Y-axis indicates the absorbance given in mAU (units of milli absorbance). 0 X-axis indicates the elution volume in ml.

Figure 38. The different assemblies and fractions collected after size exclusion chromatography (see Figure 37) were analyzed by SDS-PAGE under non-reduction, followed by silver staining of the polyacrylamide gel. The meetings (Pl, P2, and P3) and the fractions (16 to 26) analyzed are indicated on the top of the gel figure. dyed with silver. On the left (path M) the sizes of the molecular mass markers are indicated.

Figure 39. Fractions 17 to 23 of the size exclusion chromatographic step, as shown in Figure 37, were assembled and alkylated. After that, the protein material was subjected to treatment with Endo H for deglycosylation. The untreated material and the Endo H-treated material were separated on an SDS-PAGE gel and stained on a PVDF membrane. The stain was dyed with amide black.

Pathway 1: E2-H6 alkylated before treatment with

Endo H

Path 2: E2-H6 alkylated after treatment with Endo H

Figure 40. Western blot analysis of El cell lysates expressed in Saccharomyces cerevisiae. Western blotting was developed using the specific monoclonal antibody. for El IGH 201.

Pathways 1-4: 'product of expression after 2, 3, 5 or 7 days of expression, respectively, in a Saccharomyces clone. transformed with ..pSYlYIG7Els (SEQ ID NO: 50, Figure 28) comprising the nucleotide sequence encoding the chicken lysozyme leader peptide attached to

E1-H6. ·,

Paths 5-7: product of the expression after. 2, 3 or 5 days of expression, respectively, in a Saccharomyces clone transformed with pSYlaMFElsH6aYIG7 (SEQ ID NO: 44, Figure 22) comprising the nucleotide sequence encoding the ct crossing factor leader peptide attached to E1-H6.

Path 8: molecular weight markers sized as indicated.

Path 9: Here are purified, produced by mammalian cells infected with HCV-recombinant vaccinia virus.

Figure 41. Analysis of the purified E2-H6 protein using immobilized metal ion affinity chromatography (IMAC), expressed by and processed from CL-E2-H6 to E2-H6 by H. polymorpha (see Example 17) . Proteins in different wash fractions (paths 2 to 4) and elution fractions (paths 5 to 7) were analyzed by SDS-PAGE under reduction, followed by pretreatment with silver gel (A, top figure) or western transfer using a specific monoclonal antibody, directed against E2 (B, bottom figure). The sizes of the molecular weight markers are indicated on the left.

Figure 42. Elution profile of the first IMAC chromatography step on a Ni-IDA column (Ni-loaded chelating Sepharose FF2+, Pharmacia) for the purification of the sulfonated H6-K-E1 protein, produced by H. polymorpha

<td>(to see</td><td>Example 18)</td><td>. THE</td><td>column was balanced</td><td>with buffer</td><td>' A</td>

<td> (50</td><td>mM phosphate,</td><td>6 M</td><td>of GuHCl, 1,% of Empigen</td><td>BB (v / v),</td><td>PH</td>

<td> 7,2)</td><td>supplemented</td><td>with</td><td>20 mM imidazole. After</td><td>application</td><td>gives</td>

<td colspan="2">10 sample, the column</td><td>yo</td><td>sequentially washed</td><td>with buffer</td><td>THE</td>

containing 20 mM and 50 mM imidazole, respectively (as indicated on the chromatogram). A washing and cleaning step. additional elution of His-labeled products was performed by sequential application of buffer B (PBS, 1%

BB, pH 7.2) supplemented with 50 mM imidazole and 200 mM imidazole, respectively (as indicated on the chromatogram). The following fractions were combined: wash 1 (fractions 8 to 11, wash with 50 mM imidazole). The eluted material was collected as separate fractions 63 to

72 or an elution meeting was held (fractions 63 to 69). The Y axis indicates the absorbance given in mAU (units of milli absorbance). The X-axis indicates the elution volume in ml.

Figure 43. Analysis of the HAC-K-E1 protein purified by IMAC (see Figure 42), expressed by and processed in

CL-H6-K-E1 to H6-K-E1 by H. polymorpha. Proteins in wash 1 (path 12) and elution fractions 63 to 72 (paths 2 to 11) were analyzed by SDS-PAGE under reduction, followed by simulating the gel with silver (A, top figure). The proteins present in the sample before IMAC (path 2), in the flow meeting, from end to end (path 4), in the washing meeting 1. (path 5) and the elution pool (path 6) were analyzed by western blot using a specific monoclonal antibody, directed against El (IGH201) (B, background figure; no samples were loaded on path 3). The sizes of the molecular weight markers (M paths) are indicated on the left.

Figure 44. Elution profile of the second IMAC chromatography step on a Ni-IDA column (Ni-loaded chelating sepharose FF2+, Pharmacia) for the purification of E1 resulting from the in vitro processing of H6-K-E1 (purification: see Figure 42) with Endo Lys-C. The flow from end to end was collected in different fractions (1 to 40), which were examined for the · presence of Eis products. The fractions (7 to 28), containing the intact El processed from H6-K-E1, were pooled. The Y axis indicates the absorbance given in mAU (units of milli absorbance). The X-axis indicates the elution volume in ml.

Figure 45. Western blot analysis indicating specific Eis protein bands reacting with biotinylated heparin (see also Example 19). Eis preparations purified from cultured mammalian cells infected with HCV-recombinant vaccinia virus or expressed by H. polymorpha were analyzed. The right of the panel from the vertical line shows a Western blot developed with the monoclonal specific for biotinylated IGH 200. The left of the panel from the vertical line shows a Western blot developed with biotinylated heparin. From these results it is concluded that mainly the less glycosylated Eis has a high affinity for heparin.

'5 Paths M: molecular weight marker (molecular weights shown on the left).

Pathways 1: This is from mammalian cells and alkylated during isolation.

Pathways 2: Els-H6 expressed by H. polymorpha and sulfonated during isolation.

Pathways 3: Els-H6 expressed by H. polymorpha and alkylated during isolation.

Paths 4: same material as loaded in path 2, but treated with dithiothreitol to convert the sulfonated Cys-thiol groups into Cys-thiol.

Figure 46. E2-H6 size exclusion chromatography (SEC) profile expressed by purified H. polymorpha in its sulfonated form, subjected to a run in PBS, 3% betaine, to force the formation of virus-like particles by exchange of betaine for Empigen BB. The combined fractions containing the VLPs, used for further study, are indicated by the Y axis indicating the absorbance given in mAU (units of milli absorbance). The X-axis indicates the elution volume in ml. See also

Example 20.

Figure 47. E2-H6 size exclusion chromatography (SEC) profile expressed by H. polymorpha purified in its alkylated form, subjected to a run in PBS, 3% betaine ,, to force the formation of particles similar to virus through exchange of · betaine by Empigen BB. The combined fractions containing the VLPs are indicated by the Y axis indicates the absorbance given in mAU (units of milli absorbance). The X-axis indicates the elution volume in ml. See also Example 20.

Figure 48. El 'size exclusion' (SEC) chromatography profile expressed by purified H. polymorpha in its sulfonated form, subjected to a run in PBS, 3% betaine, to force the formation of virus-like particles by exchange of betaine by Empigen BB. The combined fractions containing the VLPs are indicated by The Y-axis indicates the absorbance given in mAU (units of milli absorbance). The X-axis indicates the elution volume in ml. See also Example 20.

Figure 49. Profile of the size exclusion chromatography (SEC) of El expressed by H. polymorpha purified in its alkylated form, subjected to a run in PBS, 3% betaine, to force the formation of virus-like particles by exchange of betaine by Empigen BB. The combined fractions containing the VLPs are indicated by The Y-axis indicates the absorbance given in mAU (units of milli absorbance). The X-axis indicates the elution volume in ml. See also Example 20.

Figure 50. SDS-PAGE (under reducing conditions) and western blot analysis of VLPs as isolated after size exclusion chromatography (SEC), as described in Figures 48 and 49. Left panel: gel

Silver-dyed SDS-PAGE. Right panel: western blot using a specific monoclonal antibody, directed against El (IGH201). Paths 1: molecular weight markers (molecular weights shown on the left);

paths 2: assembly of VLPs containing El sulfonada (see Figure 48); paths 3: assembly of VLPs containing El alkylated (see Figure 49). See also Example 20.

Figure 51. The E1 produced in mammalian cells (M) or the El produced in Hansenula (H) were coated on a solid ELISA support to determine the endpoint titer of the antibodies present in the sera after vaccination of mice with El produced in mammalian cells (top panel), or after vaccination of mice with El produced in Hansenula (bottom panel). The horizontal bar represents the mean antibody titer. Endpoint titles (multiple dilution) are indicated on the Y axis. See also Example 22.

Figure 52. The El produced in Hansenula was alkylated (A) or sulfonated (S) and coated on a solid ELISA support to determine the endpoint titer of antibodies present in the sera after vaccination of mice with El produced in Hansenula which was alkylated (top panel), or after vaccination of mice with El produced in Hansenula that was sulfonated (bottom panel). The horizontal bar represents the mean antibody titer. Endpoint titles (multiple dilution) are indicated on the Y axis. See also Example 23.

. Figure '53. The El dè HCV produced by mammalian cells infected with HCV-recombinant vaccinia virus and the El de HCV produced' by H. polymorpha were coated directly on ELISA plates. Endpoint titers of antibodies were determined in sera from chimpanzees vaccinated with El produced by mammalian cells (top panel) and murine monoclonal antibodies produced against El produced by. mammalian cells (bottom panel). Chimpanzees Yoran and Marti were vaccinated prophylactically. The chimpanzees Ton, .Phil, Marcei, Peggy and Femma were vaccinated therapeutically. Bars filled with black: EL-coated ELISA plate produced by mammalian cells. Open bars: EL coated ELISA plate produced by Hansenula. Endpoint titles (multiple dilution) are indicated on the Y axis. See also Example 24.

Figure 54. Carbohydrate gel electrophoresis aided by oligosaccharide fluorophore released from E1 produced by mammalian cells infected with recombinant vaccine virus and E1-H6 protein produced by

Hansenula.

Path 1: Glucose ladder pattern with indication on the left of the number of monosaccharides (3 to 10, indicated by G3 to G10)

Path 2: 25 μρ of N-linked oligosaccharides released from El (alkylated) produced by mammalian cells.

Path 3: 25 μς of N-linked oligosaccharides released from E1-H6 (alkylated) produced by Hansenula.

Way 4: 100 pmoles of maltotetraose.

EXAMPLES

EXAMPLE 1

VECTOR CONSTRUCTION IN SHUTTLE pFPMT-MFa ~ El-H6

The plasmids for the transformation of Hansenula polymorpha were constructed as follows. The pFPMT-MFa-El-H6 shuttle vector was constructed in a multi-step procedure. Initially, the nucleic acid sequence encoding, HCV El's protein (SEQ ID NO: 2) was cloned after a CHH leader sequence (CHH = Carcinus maenas hyperglycemic hormone), which was subsequently replaced by an MFa leader sequence ( MFa = Saccharomyces cerevisíae α crossing factor).

Firstly, a pUC18 derivative was constructed housing the CHH-E1-H6 unit as a FcoRI / BamHI fragment by the seamless cloning method (Padgett, K. Ά. And Sorge, JA 1996). In addition, the DNA fragment encoding Els-H6 and the receptor plasmid derived from pCHHHir were generated by PCR, as described below.

Generation of DNA fragment encoding Els-H6

I ....... 11

The DNA fragment for Els-H6 (encoding the Eis type HCV protein lb consisting of amino acids S 192 to 326 of the Eis elongated with 6 His residues; SEQ ID NO: 5) was isolated by PCR from plasmid pGEMTElsH6 (SEQ ID NO: 6; Figure 1). The following primers were used for this purpose:

CHHE1-F:

5 '-agttactcttca.aggtatgaggtgcgcaacgtgtccg-3' (SEQ ID NO: 7);

The amllO4I site is underlined, the dot marks the dividing site. The bases printed in bold are complementary to those of the primer CHH-links. The unmarked bases ring within the initiation region of El (192— 326) in the direction with direction; and

CHHE1-R:

5 '-agttactcttca.cagçrgatcctccttaatggtgatggtggtggtgcc-3' (SEQ ID NO: 8);

BamllO4I site is underlined, the dot marks the dividing site. The bases printed in bold are complementary to those of the MF30-rechts primer. The bases forming the BamHI sites, useful for subsequent cloning procedures, are printed in italics. The unmarked bases ring in the nonsense direction, inside the end of the E1-H6 unit, including the interruption codon and three additional bases between the interruption codon and the BamHI site.

io The reaction mixture was constituted as follows:

total volume of 50 μΐ containing 20 ng of pGEMTElsH6 linearized with Bco31I, each -0.2 μΜ of CHHE1-F and CHHE1-R primers, dNTP's (each at 0.2 μΜ), 1 x buffer 2 (Mold PCR system Long to Expand; Boehringer; Cat. No. 1681

834), 2.5 U of polymerase mix (PCR system of

Long Expanding Mold; Boehringer; Cat. No. 1681 834).

Program 1 was used, said program consisting of the following steps:

1. denaturation: 5 min 95 ° C;

2. 10 cycles of 30 sec denaturation at 95 ° C, annealing at 65 ° C, and 130 sec elongation at 68 ° C

3. termination at 4 ° C.

Then 5 μΐ of 10 x buffer 2 (Long Expanding Mold PCR System; Boehringer; Cat. No. 1681 834),

40 μΐ of H2O, and 5 μΐ of [dATP, dGTP, and dTTP (2 mM each); 10 mM 5-methyl-dCTP] were added to the sample derived from program 1, and further amplification was performed following program 2, which consists of the following steps:

. 1. denaturation: 5 min at 95 ° C

2'. 5 cycles of 45 sec denaturation at 95 ° C, 30 sec annealing at 65 ° C, and 130 sec at 68 ° C ·

3. . termination at 4 ° C.

Generation of the receptor plasmid derived from pCHH-Hir

The receptor fragment was made by PCR from plasmid pCHH-Hir (SEQ ID NO: 9; Figure 2) and consists of almost the entire plasmid pCHH-Hir, except that the Hir coding sequence is not. is present in the PCR product. The following primers were used for this PCR:

CHH-links: 5'-agttactcttca.cctcttttccaacgggtgtgtag-3 '(SEQ ID NO: 10); . The £ amll04I site is underlined, the dot marks the dividing site. The bases printed in bold are complementary to those of the CHHE1-F primer. The unmarked bases ring in the end of the CHH sequence in the meaningless direction; and

MF30-rechts: 5'-agtcactcttca.ctgcaggcatgcaagcttggcg-3 '(SEQ ID NO: 11);

The amllO4I site is underlined, the dot marks the dividing site. The bases printed in bold are complementary to those of the CHHE1-R primer. Unmarked bases anneal within the pUC18 sequences behind the CHHHirudin HL20 cloned from pCHH-Hir, moving away from the insert.

The reaction mixture was constituted as follows: total volume of 50 μ1 containing 20 ng of pCHH-Hir linearized with Asp718I, each 0.2 μΜ of CHH-links and MF30-rechts primers, dNTP's (each at 0.2 μΜ), 1 x buffer 2 (f PCR system

Long Expanding Mold; Boehringer; Cat. No. 1681 834),

2.5 U polymerase mix (Mold PCR System, Long Expand; Boehringer; N2 1681 834).

program 1 as described above was used.

Then, 5 μΐ of 10 x buffer 2 (PCR system of

Long Expanding Mold; Boehringer; N2 1681 834), 40 μΐ of H2O, and 5 μΐ of [dATP, dGTP, and dTTP (2 mM each); 10 mM 5-methyl-dCTP] were added to the sample derived from program 1, and further amplification was performed following program 2, as described above.

Generation of the pCHHEl vector

The DNA fragment encoding Els-H6 and o. receptor plasmid derived from pCHH-Hir generated by PCR, as described above, were purified using the PCR product purification kit (Qiagen) according to the \ specifications of the supplier. Subsequently, the purified fragments were digested separately with EamllO4I. Subsequently, the DNA fragment for Els-H6 was ligated into the receptor plasmid derived from pCHH-Hir using T4 ligase (Boehringer) following the supplier's specifications.

E. coli XL-Gold cells were transformed with the ligation mixture and plasmid DNA from several ampicillin resistant colonies was analyzed by digestion with EcoRI and BamHI. A positive clone was selected and named as pCHHEl.

Generation jdo vector pFPMT-CHH-E! H6

The BcoRI / BamHI fragment of pCHHE1 was ligated with the BcoRI / BamHI digested vector pFPMT121 (SEQ ID NO: 12; Figure 3). T41igase (Boehringer) was used according to the supplier's instructions. The ligation mixture was used to transform E. coli DH5aF 'cells. Several transformants were analyzed on restriction pattern of plasmid DNA and a positive clone was retained, which was called pFPMT-CHH-ElH6 (SEQ ID NO: 13; Figure 4) ,.

Generation of pFPMT-MFa-El-H6

Finally, the shuttle vector pFPMT-MFa-El-Hõ was generated by connecting three fragments, the said fragments being:

1. the 6,961 kb BcoRI / BamHI digested pFPMT121 (SEQ IDN0: 12; Figure 3),

2. the BcoRI / Hindllp payment of 0.245 pUC18-MFa (SEQ ID NO: 62; Figure 36), and

3. the 0.442 kb BindIII / BamHI fragment of a 0.454 kb PCR product derived from pFPMT-CHH-ElH6.

The 0.454 kb PCR product, giving rise to the N-3 fragment, was obtained by PCR using the following primers:

1. MFa-El f-Hi primer:

5 '-aggggtaagcttggataaaaggtatgaggtgcgcaacgtgtccgggatgt-3' (SEQ ID NO: 14); and

2. primer El back-Bam:

5'-agttacggatccttaatggtgatggtggtggtgccagttcat-3 '(SEQ ID NO: 15).

The reaction mixture was constituted as follows: Volume of the reaction mixture 50 μΐ, pFPMT-CHH-El-H6 (linearized with EcoRI; 15 ng / μΐ), 0.5 μΐ; MFa-El fHi primer (50 μΜ), 0.25 μΐ; primer El back-Bam (50 μΜ), 0.25 μΐ;

dNTP's (all at 2 mM), 5 μΐ; DMSO, 5 μΐ; H2O, 33.5 μΐ; Buffer 2 (10 x concentrate) of Long Expanding Mold PCR System (Boeringer Mannheim; Cat. No. 1681 834), 5 μΐ; Polymerase mixture of the Long Expanding Mold PCR System (1 U / μΙ), 0.5 μΐ.

The PCR program was used, consisting of the following steps:

1. denaturation: 5 min at 95 ° C

2. 29 cycles of 45 sec denaturation at 95 ° C, 45 sec annealing at 55 ° C, and 40 sec elongation at 68 ° C

3. termination at 4 ° C.

Based on the primers used, the PCR product of

The resulting 0.454 kb contained the codons of El (192-326) followed by six histidine codons and a taa interruption codon, upstream flanked by the 22 '3-terminal base pairs of the MFa pre-pro sequence (including the Cloning relevant Hindlll plus a six base pair projection) and downstream flanked by a BamHI site (cloning relevant) and a six base pair projection.

For the ligation reaction, T4 DNA ligase (Boehringer Mannheim) was used according to the supplier's conditions (sample volume 20 μΐ) ·

E. çoli HB101 cells were transformed with the ligation mixture and the positive clones retained after restriction analysis of plasmids isolated from several transformants, · A positive plasmid was selected and named as pFPMT-MFa-El-H6 (SEQ ID NO : 16; Figure 5).

EXAMPLE 2

CONSTRUCTION OF SHUTTLE VECTOR pFPMT-CL ~ El-H6

·. The plasmids for the transformation of Hansenula polymorpha were constructed as follows. The pFPMT-CL-El-H6 shuttle vector was constructed in three stages, starting from pFPMT-MFct-El-H6 (SEQ ID NO: 16, Figure 5).

In a first step, the MFaE1-H6 reading frame of pFPMT-MFa-El-H6 was. subcloned into the pUC18 vector. Then, a 1,798 kb SalF / BamHI fragment of pFPMT-MFaE1-H6 (containing the FMD'more MFa-El-H6 promoter) was ligated to the yector fragment by Sall / BamHI of pUC18 with T4 ligase (Boehringer) according to the supplier's conditions. This resulted in a plasmid that is represented in Figure 6 (SEQ ID NO: 17), and 'further named as pMal2-1 (pUCl8-FMD-MFa-El-H6). The ligation mixture was used to transform E. coli DH5aF 'cells. Several ampicillin resistant colonies were collected and analyzed by digestion with plasmid DNA restriction enzyme isolated from the selected clones. A positive clone was further analyzed by determining the DNA sequence of the MFot-El-H6 coding sequence. A correct clone was used for PCR-oriented mutagenesis to replace the MFa pre-pro-sequence with the codons from the avian lysozyme pre97 sequence (CL; corresponding to amino acids 1 to 18 of avian lysozyme; SEQ ID NO: 1). The principle of the applied method of PCR-oriented mutagenesis is based on the amplification of an entire plasmid with the '5 desired changes located at the 5' ends of the primers. In the downstream steps, the ends of the linear PCR product are modified before autolinking which results in the desired altered plasmid.

The following primers were used for the io PCR reaction:

1. CL hin primer:

5'-tgttcctaccactagcagcactaggatatgaggtgcgcaacgtgtccggg-3 '(SEQ ID NO: 18);

2. CL her neu primer:

5 '-tagtactagtattagtaggcttcgcatgaattcccgatgaaggcagagagcg-3' (SEQ ID NO: 19).

The underlined 5 'regions of the primers contain codons from approximately half of the avian lysozyme pre-sequence. The CL herneu primer includes a Spel restriction site (italics). The non-underlined regions of the primers ring with the codons for amino acid residues 192 to 199 from El (CL hin) or with the initiation codon atg on the EcoRI site to position -19 (counted from the EcoRI site ) of the FMD promoter. The primers are designed to amplify the complete pMal2-l, thereby replacing the codons of the MFa pre-pro-sequence with the codons of the avian lysozyme pre-sequence.

. The reaction mixture was constituted as follows: pUC18-FMD-Mfa-El-H6 (pMal2-l; 1.3 ng / μΐ), 1 μΐ; primer CL hin (100 μΜ), 2 μΐ; CL .her neu primer. (100 μΜ), 2 μΐ; dNTP's (all at 2.5. mM), 8 μΐ; H2O, 76 μΐ; Buffer 2 (10 x concentrate) of. Long Expanding Mold PCR System

<td>(Boeringer; N- from Cat.</td><td> 1681</td><td> 834) ,</td><td>10 μΐ; Mixture</td><td>in</td>

<td>polymerase of the</td><td>PCR of</td><td>Mold</td><td>Long to Expand</td><td> (1</td>

<td>The</td><td colspan="2">program</td><td>PCR consisting</td><td>in</td>

following steps:

1. denaturation: 15 min at 95 ° C

2. 35 cycles of 30 sec denaturation at 95 ° C, 1 min of annealing at 60 ° C, and 1 min of elongation at 72 ° C

3. termination at 4 ° C.

The resulting PCR product was checked by agarose gel electrophoresis for its correct size (3.5 kb). After that, the 3'-A projections of the PCR product were removed by a reaction with T4 polymerase, resulting in blunt ends with 3 'and 5' OH groups. Then the PCR product was treated with T4 polymerase (Boehringer; 1 U / μΙ): to the remaining 95 μΐ of the PCR reaction mixture were added 1 μΐ of T4 polymerase and 4 μΐ of dNTP 's (all at 2.5 mM). The sample was incubated for 20 min at 37 ° C. Subsequently, the DNA was precipitated with ethanol and absorbed in 16 μΐ H2O.

Subsequently, the 5'-phosphates were added to the blunt-ended PCR product by a reaction with kinase. Then, to the 16 μΐ of blunt-ended PCR product, 1 μΐ of T4 polynucleotide kinase (Boehringer; 1 U / μΙ), 2 μΐ of T4 polynucleotide kinase reaction buffer concentrated 10 times (Boehringer), and 1 μΐ of ATP (10 mM). The sample was incubated for 30 min at

37 ° C. Subsequently, the DNA was applied to a 1% agarose gel and the correct product band was isolated using the gel extraction kit (Qiagen) according to the supplier's conditions. Fifty (50) ng of the purified product was then self-ligated using T4 ligase (Boehringer) according to the supplier's conditions. After incubation for 72 h at 16 ° C, the DNA in the ligation mixture was precipitated with ethanol and dissolved in 20 μΐ of water.

E.coli DH5a-F 'cells were subsequently transformed with 10 μ of the ligation sample. Plasmid DNA from several ampicillin-resistant clones was checked by restriction enzyme digestion. One positive clone was retained and named p27d-3 (pUC18-FMD-CLE1-H6, SEQ ID NO: 20, Figure 7). Subsequently, the. reading frame of CL-E1-H6 was verified by sequencing of

DNA.

In a final step, the shuttle vector pFPMT-CLE1-H6 was constructed as described below. The 0.486 kb EcoRI / BamHI fragment of p27d-3 (harboring CL-E1 (19225 326) -H6) was ligated with BcoRI / BamHI digested pFPMT121 (SEQ ID NO: 12, Figure 3). For the reaction, T4 ligase (Boehringer) was used according to the supplier's recommendations. The DNA in the ligation sample was precipitated with

100 ethanol e.g. dissolved eip 10 μΐ of H20. The E. coli DH5a F 'cells were transformed with 10' μΐ of the binding sample, and the plasmid DNÀ 'of several ampicillin resistant colonies was analyzed by digestion with FcoRI and BamHI. Plasmid clone p37-5 (pFPMT-CL-El-H6; SEQ ID NO: 21, Figure 8) showed the desired fragment sizes of 0.486 kb and 6.961 kb. The correct sequence of CL-E1-H6 from p375 was verified by sequencing.

EXAMPLE 3

CONSTRUCTION OF SHUTTLE VECTORS pFPMT-MFa ~ E2-H6 AND pMPT-MFgE2-H6

The plasmids for the transformation of Hansenula polymorpha were constructed as follows. The DNA sequence encoding MFa-E2s (HCV E2 amino acids 384-673) -VIEGR-His6 (SEQ ID NO: 5) was isolated as a fragment by FcoRI / BgJII d 1.331 kb from plasmid pSP72E2H6 (SEQ ID NO: 22, Figure 9). This fragment was ligated with the FcoRI / BgTII digested vectors pFPMT121 (SEQ ID NO: 12, Figure C + 2) or pMPT121 (SEQ'ID NO: 23, Figure 10) using the T4 DNA ligase (Boehringer Mannheim) according to supplier recommendations. After the transformation of E. coli and checking plasmid DNA isolated from different transformants by restriction enzyme digestion, positive clones were retained and the resulting shuttle vectors are called pFPMT-MFa-E2-H6 (SEQ ID NO: 22, Figure 11) and pMPT-MFa-E2-H6 (SEQ ID NO: 23, Figure 12), respectively.

101

EXAMPLE 4

VECTOR CONSTRUCTION IN SHUTTLE pFPMT-CL-E2-H6

The shuttle vector pFPMT-CL-E2-H6 was assembled in a three-step procedure. An. intermediate construction • 5 was prepared, in which the 'E2 coding sequence was cloned behind the α-amylase signal sequence from Schwanniomyces accídentalis. This was done using the seamless cloning method (Padgett, KA, and Sorge, JA 1996).

Generation of the DNA fragment encoding E2s-H6

First, the DNA sequence encoding E2-H6 (amino acids 384 to 673 of HCV E2 extended with the VIEGR linker peptide and with 6 His residues, SEQ ID NO: 5) was amplified from plasmid pSP72E2H6 (SEQ ID NO: 24 , Figure 11) by PCR. The primers used were named MF30E2 / F and MF30E2 / R and have the following sequences:

MF30E2 / F primer:

5'-agtcactcttca.aggcatacccgcgtgtcaggaggg-3 '(SEQ ID NO: 26); the amllO4I site is underlined, the dot marks the enzyme dividing site; the last codon of the S. occidentalis signal sequence is printed in bold; unmarked bases ring with E2 codons (HCV E2 amino acids 384-390);

MF30E2 / R primer:

5 '-agtcactcttca. caggrgratccttagtgatggtggtgatg-3 '(SEQ ID

NO: 27); the EamllO4I site is underlined, the dot marks the enzyme dividing site; the bases printed in bold are complementary to the bases printed in bold of the primer

102

MF30-rechts (see below); a BamHI site to be introduced in the construction is, 'printed in italics; the unmarked sequence rings with the interruption codon and - the six His codons · E2 terminals (384-673) -VIEGR-H6 (SEQ ID

NO: 5).

The reaction mixture was constituted as follows: total volume of 50 μΐ containing 20 ng of the fragment by EcoRI / BglII of 1.33 kb of pSP7.2E2H6, 'each 0.2 μΜ of MF30E2 / F and MF30E2 / R primers, dNTP's (each at 0.2 μΜ), 1 x buffer 2 (Long Mold PCR System. Expand; Boehringer; N£ No. 1681 834), 2.5 U of polymerase mixture (Long Expanding Mold PCR System; Boehringer; Cat. No. 1681 834).

The PCR 3 program was used, consisting of the following steps:

1. denaturation: 5 min at 95 ° C;

2. 10 cycles of 30 sec denaturation at 95 ° C, 30 sec annealing at 65 ° C, and 1 min elongation at 68 ° C

3. termination at 4 ° C.

Then 10 μΐ of 10 x buffer 2 (PCR system of

Long Mold 'to Expand; Boehringer; N£ 1681 834), 40 μΐ of H2O, and 5 μΐ of [dATP, dGTP, and dTTP (2 mM each); 10 mM 5-methyl-dCTP] were added to the sample derived from the PCR 3 program, and it was continued with the

PCR 4 consisting of the following steps:

1. denaturation: 5 min at 95 ° C

2. 5 cycles of 45 sec denaturation at 95 ° C, 30 sec annealing at 65 ° C, and 1 min elongation at 68 ° C

103

3. termination at 4 ° C.

Generation of the pMF30-derived receptor plasmid

The second fragment originated from the plasmid pMF30 (SEQ ID NO: 28, Figure 13), the amplicon was almost the complete plasmid '5 pMF30, excluding the codons from the mature S. occidentalis α-amylase, the relevant modifications for cloning were introduced by primer design. The following group of primers was used:

MF30-Links primer:

5'-agtcactcttcá.cctcttgtcaaaaataatcggttgag-3 '(SEQ ID

NO: 29; the FamllO4I site is underlined, the dot marks the enzyme dividing site, ctt printed in bold is. complement the agg printed in bold in the MF30E2 / F primer (see above), the unmarked bases and those printed in bold ring with the 26 terminal bases of the codons of the S. occidentalis aMylase in pMF30);

MF30-Rechts primer:

5'-agtcactcttca.ctgcaggcatgçaagcttggcg-3 '(SEQ ID

NO: 11; the Eamll04I site is underlined, the dot marks the enzyme dividing site, bold printed ctg is complementary to bold printed primer MF30E2 / R (see above); the unmarked bases ring with the pUC18 sequences upstream of the S. occidentalis α-Amylase interruption codon in pMF30);

The reaction mixture was constituted as follows:

total volume of 50 μΐ containing 20 ng of pMF30 linearized with BglII, each 0.2 μΜ of MF30-Links and MF30-Rechts primers, dNTP's (each at 0.2 μΜ), lx buffer 1 (Mold PCR system

104

Long to Expand; Boehringer; No. Cat. 1681 834), 2.5 U of polymerase mix (Expandable Long Mold PCR System; 'Boehringer;' No. Cat. 1681- 834). The same PCR programs (programs 3 and 4) as described above were used. except for the stretching times that were increased from 1 minute to 4 minutes in both programs. Generation of the vector pAMY-E2 ·, The DNA fragment encoding E2s-H6 and the receptor plasmid derived from pMF30 obtained by. PCR were monitored in their respective size by gel electrophoresis on a 1% agarose gel. PCR products were purified with a PCR product purification kit (Qiagen) according to the supplier's instructions. Subsequently, the purified fragments were digested separately with Eamll004I. The ligation of the E2sH6 fragment with the receptor plasmid derived from pMF30 was performed using T4 ligase (Boehringer) according to the supplier's recommendations. The ligation mixture was used to transform E. coli DH5aF 'cells and plasmid DNA from several clones was analyzed by digestion with FcoRI / BamHI. A positive clone was selected, its plasmid additionally called pAMY-E2, and used for further modifications, as described below.

Generation of the vector pUC18-CL-E2-H6 0 pAMY-E2 was subjected to mutagenesis guided by

PCR in order to replace the codons of the α-amylase signal sequence with the codons of the avian lysozyme pre-sequence.

105

This is additionally referred to as CL, corresponding to the first 18 amino acids of the avian lysozyme ORF (SEQ ID NO: 1). For this mutagenesis, the following primers were used:

hin CL2 primer:

5 '-tgcttcctaccactagcagcactaggacatacccgcgtgtcaggaggggcag-3' (SEQ ID NO: 30); and primer CL2 her:

5 '-1agtactagtattagtaggcttcgcatgqaa t tcactggccgtcgttttacaacgtc3' (SEQ ID NO: 31).

The underlined 5 'regions of the primers contain the DNA sequence of approximately half of the avian lysozyme pre-sequence. The CL2 her primer includes the Spel (italic) and EcoRI (italic, double underlined) restriction sites. The non-underlined regions of the primers ring with the amino acid residue codons 384 to 392 of E2 (CL2 hin) or with the initiation codon atg on the EcoRI site until apposition -19 (counted from the EcoRI site) of the promoter FMD. Primers are designed to amplify the complete pAMY-E2 vector, thereby replacing the codons in the α-amylase signal sequence with the codons in the avian lysozyme pre-sequence.

The PCR reaction was carried out according to the following program:

1. denaturation: 15 min at 95 ° C

2. 35 cycles of 30 sec denaturation at 95 ° C, 1 min of annealing at 60 ° C, and 1 min of elongation at 72 ° C

3. termination at 4 ° C.

106

The following reaction mixture was used: pAMY-E2 (1 ng / μΐ), 1 μΐ; primer CL2 hin (100 μΜ), 2 μΐ; primer CL2 her (100 μΜ), 2 μΐ; dNTP's' (all. at 2.5 mM each), 8 μΐ; Η2Ο, · 76 μΐ; Buffer 2 (10 x concentrate) of the Mold PCR System

Long to Expand (Boeringer; Cat. No. 1681 834), 10 μΐ;

Polymerase mixture of the Long Expanding Mold PCR System (1 U / μΙ), 0.75 μΐ.

The resulting PCR product was checked by gel electrophoresis on a 1% agarose gel. Binding dyes, the PCR fragment was modified as follows. The 3'-A projections were removed by T4 polymerase, resulting in blunt ends with 3 'and 5' OH groups. Then, 1 μΐ of T4 polymerase (Boehringer, 1 U / μΙ) was added to the residual 95 μΐ of PCR reaction mix along with 4 μΐ of dNTP's (2.5 mM each). The sample was incubated for 20 min at 37 ° C. Subsequently, the DNA was precipitated with ethanol and dissolved in 16 μΐ of water

<td rowspan="2">deionized. This was followed by to add the 5'-phosphates</td><td rowspan="2">one to</td><td colspan="4">kinase treatment</td>

<td>product</td><td>in</td><td>PCR</td><td>in</td>

<td>blunt end. At 16 μΐ</td><td>in</td><td>product</td><td>in</td><td>PCR</td><td>in</td>

<td colspan="3">dissolved blunt end were added</td><td> 1</td><td>μΐ of</td><td>T4</td>

polynucleotide kinase (Boehringer, 1 υ / μΐ), 2 μΐ of T4 reaction buffer 10 times concentrated polynucleotide kinase (Boehringer) and 1 μΐ of ATP (10 mM). The sample was incubated for 30 min at 37 ° C.

The kinase-treated sample was subsequently separated on a 1% agarose gel. The product band was isolated. DNA was extracted from the agarose slice using

107 of the Gel Extraction kit (Qiagen) according to the supplier's recommendations. Fifty (50) ng of the purified product was then self-ligated using T4 ligase (Boehringer) according to the supplier's conditions. After incubation for 16 h at 16 ° C, the DNA in the ligation mixture was precipitated with ethanol and dissolved in 20 μΐ H2O (connecting sample).

The DH5aF 'cells of E. .coli were transformed with 10 μΐ of the ligation sample. Several ampicillin-resistant clones have been further characterized by restriction analysis of isolated plasmid DNA. A positive clone was termed as pUC18-CL-E2-H6 and was used for further modifications as described below.

Generation of shuttle vector pFPMT-CL-E2-H6 A 0.966 kb EcoRI / BamHI fragment was isolated from pUC18-CL-E2-H6 (harboring CL-E2 (384 - 673) -VIEGRH6) and was linked to pFPMT121 digested with FcoRI / BamHI (SEQ ID NO: 12, Figure 3). For the reaction, T4 ligase (Boehringer) was used according to the supplier's conditions. The binding sample was precipitated with ethanol and dissolved in 10 μΐ of water. This was used to transform DH5aF 'E cells. coli, a positive clone was retained after restriction analysis and the respective plasmid is called PFPMT-CL-E2-H6 (SEQ ID NO: 32, Figure 14).

EXAMPLE 5

VECTOR CONSTRUCTION IN SHUTTLE pFPMT-CL ~ K-H6-El

The construction of the shuttle vector was comprised of two stages.

108

In a 'first step, the construction of pUC18-FMDCL-H6-K-E1-H6 was constructed by site-oriented mutagenesis. PUClô-FMD-CL-El-Hó was used as a template (SEQ · ID NO: 20; Figure 7) ·. The following primers were used:

Pr.imer H6K hin neu:

5'-catcacaaatatgaggtgcgcaacgtgtccgggatgtac-3 '(SEQ ID

NO: 37).

Prirner H6KRK her neu:

5 '-gtgatggtggtgtcctagtgctgctagtggtaggaagcatag-3' (SEQ ID NO: 3.8). '(The bases that provide additional codons are underlined.)

The PCR reaction mixture was constituted as follows: pUC18-FMD-CL-El-H6 (2 ng / μΐ), 1 μΐ; prirner H6K hin neu (100 μΜ), 2 μΐ; prirner H6KRK her neu (100 μΜ), 2 μΐ;

dNTP 's (2.5 mM each), θ μΐ; H2O, 7 6 μΐ; Buffer 2 (10 x concentrate) of the Long Expanding Mold PCR System (Boeringer; N£ Cat. 1681 834), 10 μΐ; Polymerase mixture of the Long Expanding Mold PCR System (1 U / μΙ), 0.75 μΐ.

The PCR program used consisted of the following steps:

1. denaturation stage: 15 min at 95 ° C

2. 35 cycles of 30 sec denaturation at 95 ° C, 1 min of annealing at 60 ° C, and 5 min of elongation at 72 ° C

3. termination at 4 ° C.

109

An aliquot of the PCR sample was analyzed on a 1% agarose gel to check its size, which was correct (~ 4.2 kb).

After that, the 3'-A projections of the PCR 5 product were removed by a reaction with T4 polymerase, resulting in blunt ends with 3 'and 5' OH groups. Then, to the remaining 95 μΐ of the PCR reaction, 1 μΐ of T4 polymerase (Boehringer; 1 U / μΙ) was added, and 4 μΐ of dNTP 's (2.5 mM each). The sample was incubated for 20 min at 37 ° C.

Subsequently, the DNA in the sample was precipitated with ethanol and dissolved in 16 μΐ H20.

Subsequently, the 5'-phosphates were added to the blunt-ended PCR product by a kinase reaction. Then, to the 16 μΐ of PCR product with dissolved blunt ends, 1 μΐ of T4 polynucleotide kinase (Boehringer; 1 U / μΙ), 2 μΐ of T4 reaction buffer concentrated 10 times (Boehringer), and 1 μΐ of ATP (10 mM). The sample was incubated for 30 min at 37 ° C.

Subsequently, the sample was applied on a 1% agarose gel and the correct product band was isolated, using the gel extraction kit (Qiagen) according to the supplier's conditions. Fifty (50) ng of the purified product was then self-ligated using

T4 ligase (Boehringer) according to the supplier's recommendations. After incubation for 72 h at 16 ° C, the DNA in the ligation sample was precipitated with ethanol and dissolved in 10 μΐ of water.

110

DH5aF 'cells of E .. coli were transformed with 5 μΐ of the ligation sample. Plasmid DNA from several ampicillin-resistant colonies1 was analyzed by restriction enzyme digestion, a positive clone was retained. and the corresponding plasmid called: pUC18-FMDCL-H6-E1-K-H6 (SEQ ID NO: 39, Figure 17).

In a second step, the transfer vector was constructed by connecting two fragments. In the construction that follows, the fragments with cohesive ends Bell were involved. Since Bell can cleave its site only on unmethylated DNA, a strain of E. coli darei has been transformed with the plasmids involved pUC18-FMD-CL-H6-KE1-H6 (SEQ ID NO: 39, Figure 17). and pFPMT-CL-El (SEQ ID NO: 36, Figure 16). From each transformation, an ampicillin-resistant colony was collected, developed in a liquid culture, and the DNAs of non-methylated plasmids were prepared for further use. The 1,273 kb Bcll / Hindlll fragment of the non-methylated plasmid pUC18FMD-CL-H6-K-E1-H6 (harboring the 'FMD promoter, the codons of the CL-H6-K unit, and the beginning of E1) and the 6.057 kb Bcll / Hindlll fragment of plasmid pFPMT-CL-El (harboring the missing part of the El reading frame starting from the Bell site, without the His C-terminal mark, as well as the elements located in pFPMT121, except for the FMD promoter) were prepared and connected together for 72 h at 16 ° C, by using T4 ligase (Boehringer) in a total volume of 20 μΐ, according to the supplier's specifications. Subsequently, the ligation mixture was

111 placed on a piece of nitrocellulose membrane floating over sterile deionized water, in order to desalify the binding mixture (incubation for 30 min at room temperature). The cells. E. coli TOPIO were '5 transformed by electroporation with 5 μΐ of the desalified sample. Plasmid DNA from the resulting resulting ampicillin-resistant colonies was analyzed by restriction enzyme digestion. A positive clone was retained and named pFPMT-CL-H6-K-El (SEQ ID NO: 40, Figure

18) .

EXAMPLE 6

TRANSFORMATION OF HANSENULA POLYMORPHA AND SELECTION OF

TRANSFORMANTS

The H. polymorpha RB11 strain was transformed (PEG-mediated DNA uptake protocol essentially as described by (Klebe, RJ et al. 1983) with the modification of (Roggenkamp, R. et al. 1986) with the shuttle vectors different parents as described in Examples 1 to 5. For each transformation, 72 prototrophic uracil colonies were selected and used for strain generation by the following procedure. For each colony, a 2 ml liquid culture was inoculated and developed in test tubes for 48 h (37 ° C; 160 rpm; 45 ° angle) in selective medium (YNB / glucose, Difco). This stage is defined as the first passing stage. An aliquot of 150 μΐ of the cultures from the first pass was used to inoculate 2 ml of new YNB / glucose medium. Again, the cultures were incubated as described above (second stage

112 passing). Together, eight of these passages have been completed. ' The culture aliquots after the third and eighth passages were used to inoculate 2 ml- of non-selective YPD medium (Difco). After 48 h of incubation at 37 ° C (160 rpm; 45 ° angle; the so-called first stabilization step), the 150 μΐ aliquots of these YPD cultures were used to inoculate new 2 ml YPD cultures, the • which were incubated as described above (second stabilization step). The aliquots of the cultures from the second stabilization step were then streaked onto plates containing YNB / selective agar. These plates were incubated for four days until macroscopic colonies were visible. A single, well-defined colony from each separation was defined as a strain and used for further expression analysis.

Expression analysis was performed on small-scale flask cultures. One colony was collected from the aforementioned YNB / agar plate and inoculated in 2 ml of YPD and incubated for 48 h, as mentioned above. This 2 ml aliquot was used as a seed culture for 20 ml of culture in a shake flask. YPGlycerol (1%) was used as the medium and the shake flask was incubated on a rotary shaker (200 rpm, 37 ° C). After 48 h of development, 1% MeOH was added to the culture to induce the expression cassette. At different time intervals, cell pellets from 1 ml aliquots were collected and stored at -20 ° C until further analysis. Specific protein expression was analyzed by SDS-PAGE /

113 Western transfer. Then, the cell pellets were solubilized in sample buffer (TrisHCl - SDS) and incubated for> 15 minutes at 95 ° C. The proteins were separated on a 15% polyacrylamide gel and stained (wet spot; bicarbonate buffer) on nitrocellulose membranes. The spots were developed using a murine anti-E1 (IGH 201) or murine anti-E2 (IGH 216, described by Maertens et al. in W096 / 04385), specific as the first antibody, Rabbit Mouse Anti-AP was used as the second antibody. Dyeing was carried out with NBT-BCIP. Positive strains were retained for further investigation.

Five of these positive clones were used in a shake flask experiment. A colony of the respective strain was collected from the YNB plate and used to inoculate 2 ml of YPD. These cultures were incubated as described above. This cell suspension was used to inoculate a second 100 ml seed culture of YPD medium in a 500 ml shake flask. This shaking flask was incubated on a rotary shaker for 48 h at 37 ° C and 200 rpm. A 25 ml aliquot of this seed culture was used to inoculate 250 ml of YPGlycerol medium (1%) and was incubated in a 2 1 shake bottle with baffles, under the conditions described above. 48 h after inoculation, 1% MeOH (promoter induction) was added and the shake flasks were further incubated under the conditions described above. 24 h after induction, the experiment was stopped and the pellets of

114 cells collected by, centrifugation · The expression level of the five different clones. was analyzed by SDSPAGE / Western blot (conditions as described above). A series of titrations for each clone was loaded onto the gel and the most productive strain was selected for further fermentation and purification tests.

Surprisingly, H. polymorpha, a yeast strain · closely related to Pichia pastoris (Gellissen, G. 2000), is capable of expressing HCV proteins essentially without hyperglycosylation and thus with portions of sugar comparable in size to proteins of HCV envelope expressed by mammalian cells infected with HCV-recombinant vaccinia virus.

The Hansenula polymorpha RB11 strain was deposited on April 19, 2002, under the conditions of the Budapest Treaty, at the Mycothèque de I'UCL (MUCL), Université Catholique de Louvain, Laboratoire de mycologie, Place Croix du Sud 3 bte 6 , B-1348 Louvain-la-Neuve, Belgium, and has the access number of MUCL MUCL43805.

EXAMPLE 7

CONSTRUCTION OF THE pSYlaMFElsH6a VECTOR

The S. cerevisiae expression plasmid was constructed as follows. An El coding sequence was isolated as a fragment by Nsll / Eco521 from pGEMT-ElsH6 (SEQ ID NO: 6, Figure 1), which was thickened at the end (using T4 DNA polymerase) and cloned into the pYIG5 vector (SEQ ID NO: 41, Figure 19) using T4 DNA ligase (Boehringer) according to the supplier's specifications.

115

The cloning was such that the Els-H6 coding fragment was joined directly and in frame to the aMF coding sequence. The ligation mixture was transformed into E.coli DH5aF 'cells. Subsequently, plasmid DNA from several ampicillin resistant clones was analyzed by restriction digestion and a positive clone was retained and named pYIG5ElH6 (ICCG3470; SEQ ID NO: 42, Figure 20).

expression cassette (containing the aMF sequence and the Eis coding region with a His tag) io was transferred, as a fragment by BamHI (2790 bp) from pYIG5ElH6, to the E. coli / pSYl shuttle vector S.

cerevisiae digested with BamHI (SEQ ID NO: 21, Figure 43). The connection was made with T4 DNA ligase (Boehringer) according to the supplier's conditions. The ligation mixture was transformed into E. coli DH5aF 'cells, and plasmid DNA from several ampicillin resistant colonies was analyzed by restriction enzyme digestion. One positive clone was retained and named pSYlAMFElsH6a (ICCG3479; SEQ ID NO: 44, Figure 22).

EXAMPLE 8

CONSTRUCTION OF THE pSYYIGSE2H6 VECTOR The S. cerevisiae pSYYIGSE2H6 expression plasmid was constructed as follows. An E2 coding sequence was isolated as a fragment by

Sall / Kpnl from pBSK-E2sH6 (SEQ ID NO: 45, Figure 23), which was thickened at the end (using T4 DNA polymerase) and subsequently cloned into the vector pYIG5 (SEQ ID NO: 41, Figure 19) using T4 DNA ligase (Boehringer) from

116 according to the supplier's specifications. The cloning was such that the E2-H6 coding fragment was joined directly and in frame to the otMF coding sequence. The ligation mixture was then transformed into E. coli DH5aF 'cells, the plasmid DNA of several ampicillin resistant clones was analyzed by restriction digestion and a positive clone retained and named as pYIG5HCCL-22aH6 (ICCG2424; SEQ. ID NO : 46, Figure 24).

The expression cassette (containing the aMF sequence and the E2 coding region (384 - 673) with a His tag) was transferred, 'as a fragment by BamHl (3281 bp) from pYIG5HCCL-22aH6, to the vector in E. coli / S pSYl shuttle. cerevisiae opened by BamHl (SEQ ID NO: 43, Figure 21). The connection was made with T4 DNA ligase (Boehringer) according to the supplier's conditions. The ligation mixture was transformed into E. DH5otF cells. coli and plasmid DNA from several ampicillin-resistant colonies were analyzed by restriction enzyme digestion. One restriction positive clone was retained and named pSYYIGSE2H6 (ICCG2466; SEQ ID NO: 47, Figure 25).

EXAMPLE 9

PSY! YIG7Els VECTOR CONSTRUCTION

The S. cerevisiae pSYlYIG7Els expression plasmid was constructed as follows. An El coding sequence was isolated as a fragment by Nsll / Eco521 from pGEMT-Els (SEQ ID NO: 6, Figure 1), which was thickened at the end and cloned into the vector pYIG7 (SEQ ID NO: 48, Figure 26) using T4 DNA ligase

117 (Boehringer) according to the supplier's specifications. The cloning was such that the E1 coding fragment was joined directly and in frame to the ocMF coding sequence. The ligation mixture was transformed into E. coli DH5aF 'cells, the plasmid DNA from several ampicillin resistant clones analyzed by restriction digestion and a positive clone retained and named as pYIG7El (SEQ ID NO: 49, Figure 27). , expression cassette (containing the CL leader sequence and the coding region of El (192-326)) was transferred, as a fragment by BamHl (2790 bp) from pYIG7El, to the shuttle vector pSYl of E. coli / S. cerevisiae digested with BamHl (SEQ ID NO: 43, Figure 21). The connection was made with T4 DNA ligase (Boehringer) according to the supplier's conditions. The ligation mixture was transformed into E DH5aF 'cells. coli, and plasmid DNA from several ampicillin-resistant colonies was analyzed by restriction enzyme digestion. A positive clone was retained and named pSYlYIG7Els (SEQ ID NO: 50,

Figure 28).

EXAMPLE 10

TRANSFORMATION OF SACCHAROMYCES CEREVISIAE AND SELECTION OF

TRANSFORMANTS

In order to overcome the problems of hyper25 glycosylation, often described for proteins overexpressed in Saccharomyces cerevisiae, a mutant test was constructed. This examination was based on the

Bailou (Bailou, L. et al. 1991), with which they were

118 spontaneous, recessive, orthogonadate-resistant mutants were selected. The initial selection of the strain was made based on the glycosylation pattern of the invert, as noted, after native gel electrophoresis. One strain, reduced in glycosylation capacities, was retained for additional recombinant protein expression experiments and called strain IYCC155. The nature of the mutation has not been further studied.

Said glycosylation deficient strain IYCC155 was transformed with the plasmids as described in Examples 7 to 9 essentially by the lithium acetate method, as described by Elble (Elble, R. 1992). Several complemented strains of Ura were collected from a YNB + plate

<td> 2 %</td><td>of agar</td><td>selective</td><td>(Difco)</td><td>and used for</td><td>inoculate 2 ml</td><td>in</td>

<td>YNB</td><td>+ 2%</td><td>glucose.</td><td>These</td><td>cultures were</td><td>incubated for</td><td> 72</td>

<td>H,</td><td>37 ° C,</td><td>200 rpm,</td><td>on</td><td>a stirrer</td><td>orbital, and</td><td>O</td>

culture supernatant and intracellular fractions were analyzed for El expression by western blot revealed with a murine monoclonal antibody specific for El (IGH 201). A high production clone was retained for further experiments.

Protein expression in the glycosylation-deficient mutant of S. cerivisiae, used here, is hampered by the suboptimal growth characteristics of such strains, which results in a lower biomass yield and thus a lower yield of the desired proteins in comparison with S. cerivisiae strains of the type

119 wild. The yield of the desired proteins was still substantially greater than in mammalian cells.

EXAMPLE 11

CONSTRUCTION OF THE pPICZalf aD ', ElsH6 AND pPICZalfaE'ElsH6' VECTORS The shuttle vector pPICZalfaE'ElsH6 was constructed starting from the vector pPICZalfaA (Invitrogen; SEQ ID NO: 51, Figure 29). In a first step, said vector was adapted in order to enable the cloning of the El coding sequence directly behind the dividing site of the KEX2 or STE13 processing proteases, respectively. Then, pPICZalfaA was digested with Xhol and Notl. The digestion was separated on a 1% agarose gel and the. 3519 kb (largest part of the vector) was isolated and purified using a gel extraction kit (Qiagen). This fragment was then ligated using T4 polymerase (Boehringer) according to the supplier's conditions, in the presence of specific oligonucleotides, producing either pPICZalfaD '(SEQ ID NO: 52, Figure 30) or pPICZalfaE' (SEQ ID NO: 53 , Figure

31).

The following oligonucleotides were used:

for the construction of pPICZalfaD ':

8822: 5'-TCGAGAAAAGGGGCCCGAATTCGCATGC-3 '(SEQ ID.NO:54); and

8823: 5'-GGCCGCATGCGAATTCGGGCCCCTTTTC-3 '(SEQ IDNO: 55) which produce, after annealing, the linker oligonucleotide:

TCGAGAAAAGGGGCCCGAATTCGCATGC (SEQ ID NO: 54)

CTTTTCCCCGGGCTTAAGCGTACGCCGG (SEQ ID NO: 55) for the construction of pPICZalfaE '

120

8649: 5'-TCGAGAAAAGAGAGGCTGAAGÇCTGCAGCATATGC-3 '(SEQ ID

NO: 56)

650: '5'-GGCCGCATATGCTGCAGGCTTCAGCCTCTCTTTTC-3' (SEQ ID

NO: 57) that produce, after annealing, the linker oligonucleotide:

TCGAGAAAAGAGAGGCTGAAGCCTGCAGCATATGC (SEQ ID NO: 56)

CTTTTCTCTCCGACTTCGGACGTCGTATACGCCGG (SEQ ID NO: 57)

These shuttle vectors pPICZalfaD 'and pPICZalfaE' recently introduced cloning sites directly behind the dividing site of the respective processing proteases, KEX2 and STE13. The E1-H6 coding sequence was isolated as a fragment by NsII / Eco52I from pGEMT-ElsH6 (SEQ ID NO: 6, Figure 1). The fragment was purified using a gel extraction kit (Qiagen) after separating the digestion on a 1% agarose gel. The resulting fragment was thickened at the end (using T4 DNA polymerase) and ligated into pPICZalfaD 'or pPICZalfaE' directly behind the respective processing protease divider site.

The ligation mixtures were transformed into E. coli TOPIOF 'cells and the plasmid DNA from several zeocin-resistant colonies analyzed by restriction enzyme digestion. Positive clones were retained and named pPICZalfaD'ElsH6 (ICCG3694; SEQ ID NO: 58, Figure 32) and pPICZalfaE'ElsH6 (ICCG3475; SEQ ID NO: 59, Figure 33), respectively.

121

EXAMPLE 12

CONSTRUCTION OF THE pPICZalfaD'E2sH6 AND pPICZalfaE'E2sH6 VECTORS

The shuttle vectors pPICZalfaD 'and pPICZalfaE' were constructed as described in Example 11.

The E2-H6 coding sequence was isolated as a fragment by Sall / Kpnl from pGEMT-E2sH6 (SEQ ID NO: 45, Figure 23). The fragment was purified with a gel extraction kit (Qiagen) after separating the digestion on a 1% agarose gel. The resulting fragment was thickened at the end (using T4 DNA polymerase) and ligated into pPICZalfaD 'or pPICZalfaE' directly behind the respective processing protease divider site. The ligation mixture was transformed into E. coli TOPIOF 'cells and the plasmid DNA from several zeocin-resistant colonies was analyzed by restriction enzyme digestion. Positive clones were retained and named pPICZalfaD'E2sH6 (ICCG3692; SEQ ID NO: 60, Figure 34) and pPICZalfaE'E2sH6 (ICCG3476; SEQ ID NO: 61, Figure 35), respectively.

EXAMPLE 13

PICHIA PASTORIS TRANSFORMATION AND TRANSFORMANT SELECTION

<td>The</td><td>plasmids</td><td>in</td><td>shuttle</td><td>of P.</td><td>pastoris, as</td>

<td colspan="2">described in the Examples</td><td> 11</td><td>and 12,</td><td>were</td><td>transformed into</td>

<td>cells of</td><td>P. pastoris</td><td>in</td><td>wake up</td><td>with</td><td>the conditions of the</td>

<td>provider (</td><td>Invitrogen).</td><td>An</td><td colspan="3">producing strain of El and one of</td>

E2 were retained for further characterization.

HCV wrap proteins were expressed in P. pastoris, a yeast strain well known for

122 the fact that hyperglycosylation is normally absent (Gellissen, G. 2000) and previously used to express the dengue E virus protein as a GST fusion (Sugrue, RJ et al. 1997). Notably, protein wrap

HCV expressed in the resulting P. pastoris showed comparable glycosylation as seen in wild-type Saccharomyces strains. More specifically, the HCV envelope proteins produced by P. pastoris are hyperglycosylated (based on the molecular weight of the expression products detected in western transfers of proteins isolated from transformed P. pastoris cells).

EXAMPLE 14

CULTURE CONDITIONS FOR SACCHAROMYCES CEREVISIAE,

HANSENULA POLYMORPHA AND PI CHIA PASTORIS

Saccharomyces cerevisiae

Cell Bank

From the selected recombinant clone, a main cell bank and a working cell bank were prepared. The small cryovials were prepared from a semi-exponentially developed flask culture (incubation conditions as for fermentation seed cultures, see below). Glycerol was added (50% final conc.) As a cryoprotectant.

Fermentation

Seed cultures were initiated from a small vial from the cryopreserved working cell bank and grown in 500 ml of medium (YNB

123 supplemented with 2% sucrose,, 'Difco) in a 2 1 Erlenmeyer shaking bottle at 37 ° C,' 200 rpm, for 48 h.

Fermentations were typically carried out in Çiostat C fermenters with a working volume of 15 l (B. Braun Int.,. Melsungen, Germany). The fermentation medium contained 1% Yeast Extract, 2% Peptone and 2% sucrose as the carbon source. Polyethylene glycol was used as a defoaming agent.

The temperature, pH and dissolved oxygen were typically controlled during the fermentation, the applicable set points are summarized in Table 1. Dissolved oxygen was controlled in the cascade by agitation / aeration. The pH was controlled by adding NaOH (0.5 M) or H3PO4 solution (8.5%).

Table 1: Typical parameter adjustments for S. cerevisiae fermentations

<td>Parameter</td><td>set point</td>

<td>Temperature</td><td>33 - 37 ° C</td>

<td>DO (growth phase)</td><td>10 - 40% air saturation</td>

<td>DO (induction)</td><td> 0-5%</td>

<td>aeration</td><td>0.5 - 1.8 vvm *</td>

<td>agitation</td><td>150 - 900 rpm</td>

* volume replacement per minute

Fermentation was initiated by the addition of 10% seed culture. During the growth phase, the sucrose concentration was monitored off-line 20 by HPLC analysis (Polysphere OAKC Merck column).

124

During the growth phase, dissolved oxygen was controlled by cascade control (agitation / aeration). After complete sucrose metabolization, the production of heterologous protein was guided by endogenous ethanol produced, supplemented with the addition, in stages, of EtOH, in order to maintain the concentration at approximately 0.5% (HPLC analysis offline, OAKC polyspher column). During this induction phase, dissolved oxygen was controlled below 5% air saturation by manually adjusting the air flow and agitator speed.

Typically, fermentation was collected 48 to 72 h after concentration induced via flow filtration. tangential followed by centrifugation of the concentrated cell suspension to obtain the cell pellets. If not analyzed immediately, the cell pellets were stored at -70 ° C.

Hansenula polymorpha

Cell Bank

From the selected recombinant clone, a main cell bank and a working cell bank were prepared. The small cryovials were prepared from a semi-exponenially developed flask culture (incubation conditions as for fermentation seed cultures, see below). Glycerol was added (50% final conc.) As a cryoprotectant.

Fermentation

Seed cultures were initiated from a small flask in the working cell bank

125 cryopreserved (-70 ° C), and developed in 500 ml of medium (YNB, Difcó) in a 2 1 Erlenmeyer shaking flask, at 37 ° C, 200 rpm, for 48 h.

Fermentations were typically carried out in Biostat C fermenters with a working volume of 15 l (B. Braun Int., Melsungen, Germany). The fermentation medium contained 1% Yeast Extract, 2% Peptone and 1% glycerol as the carbon source. Polyethylene glycol was. used as a defoaming agent.

ίο Temperature, pH and oxygen in air and dissolved were typically controlled during fermentation, the applicable set points are summarized in Table 2. Dissolved oxygen was controlled by stirring. The pH was controlled by adding NaOH · (0.5 M) or H solution3POWDER4 (8,5 %) .

Table 2: Typical parameter adjustments for H polymorpha fermentations

<td>Parameter</td><td>set point</td>

<td>Temperature</td><td>30 - 40 ° C</td>

<td>OF</td><td>10 - 40% air saturation</td>

<td>aeration</td><td>0.5 - 1.8 vvm *</td>

<td>agitation</td><td>150 - 900 rpm</td>

* volume replacement per minute

Fermentation was initiated by the addition of 10% seed culture. During the growth phase, the glycerol concentration was monitored offline (Polysphere OAKC Merck Column) and, 24 h after the complete consumption of glycerol, 1% methanol was added in order to

126 induce the expression of heterologous protein. The fermentation was collected 24 h after concentration induction via tangential flow filtration, followed by centrifugation of the concentrated cell suspension to obtain the cell pellets. If not analyzed immediately, the cell pellets were stored at -70 ° C.

Pichia pastoris

Small-scale protein production experiments with recombinant Pichia pastoris were established in shake flask cultures. Seed cultures were grown overnight, in YPD (Difco) medium. The initial pH of the medium was corrected to 4.5. The shake flasks were incubated on a rotary shaker at 200 250 rpm, 37 ° C.

Small scale production was typically carried out on a 500 ml scale in 2 1 shake flasks and started with a 10% inoculation in expression medium, containing 1% yeast extract, 2% Peptone (both from Difco), and 2% glycerol as the carbon source. The incubation conditions were as for the seed culture. Induction was initiated by adding 1% MeOH approximately 72 h after inoculation. The cells were collected 24 h after induction, by centrifugation. If not analyzed immediately, the cell pellets were stored at -70 ° C.

127

EXAMPLE 15

REMOVAL OF THE 'LEADING PEPTIDE FROM THE MFQ-El-H6 AND MFg-E2H6 PRQTINES EXPRESSED IN SELECTED YEAST CELLS ·

The expression products in Hansenula polymorpha and a negative glycosylation strain of Saccharomyces cerevisiae from the HCV E1 and E2 protein constructs with the S. cerevisiae α crossing factor (aMF) leader sequence were further analyzed. Since both HCV Eis (aa 192-326) and HCV E2s (aa 383-673 extended 'by the VIEGR sequence (SEQ ID NO: 69)) of lb genotype were expressed as his C-terminal (H6-labeled proteins) , HHHHHH, SEQ ID NO: 63; said HCV proteins are additionally in this Example designated as aMF-ElH6 and (XMF-E2-H6), a quick and efficient purification of the products expressed after solubilization with guanidium chloride (GuHCl) of the yeast cells on Ni- IDA (iminodiacetic Ni-acid). In summary, the cell pellets were resuspended in 50 mM phosphate, 6 M GuHCl, pH 7.4 (9 vol / g cells). The proteins were sulfonated overnight, at room temperature (RT), in the presence of 320 mM (4% w / v) sodium sulfite and 65 mM (2% w / v) sodium tetrathione. The lysate was purified, after a freeze-thaw cycle, by centrifugation (10,000 g, 30 min, 4 ° C) and Empigen (Albright & Wilson, UK) and imidazole were added to the supernatant to final concentrations of 1 % (w / v) and 20 mM, respectively. The sample was filtered (0.22 μΜ) and loaded onto a Ni-IDA Sepharose FF column, which was equilibrated with 50 mM of

128 phosphate, 6M GuHCl, 1% Empigen (buffer A) supplemented with 20 mM imidazole. The column was washed sequentially with buffer A containing 20 mM and 50 mM imidazole, respectively, until the absorbance at 280 nm reached the baseline level. The products marked with his were eluted by applying buffer D, 50 mM phosphate, 6 M GuHCl, 0.2% (for E1) or 1% (for E2) Empigen, 200 mM imidazole. The eluted materials were analyzed by SDSPAGE and western blot using a specific monoclonal antibodies directed against El (IGH201), or E2 (IGH212).

El products were immediately analyzed for Edman degradation.

Since at this stage the SDS-PAGE already revealed a very complex picture of protein bands for HCV E2, an additional fractionation was carried out through size exclusion chromatography. The Ni-IDA eluate was concentrated by ultrafiltration (MWCO 10 kDa, centriplus, Amicron, Millipore) and loaded onto Superdex G200 (10/30 or 16/60; Pharmacia) in PBS, 1% Empigen or PBS, 3% Empigen. The elution fractions, containing E2 products, with a Mr between ~ 80 kDa and ~ 45 kDa, ie fractions 17-23 of the elution profile in Figure 37 based on migration on SDS-PAGE (Figure 38), were pooled and alkylated (incubation with 10 mM DTT, 3 h, at RT, followed by incubation with 30 mM of iodine-acetamide for 3 hours at RT). The samples for amino-terminal sequencing were treated with Endo H (Roche Biochemicals) or left untreated. The products

129

Ε2 glycosylates 'and deglycosylates.' were stained on PVDF membranes for amino-terminal sequencing. A stain streaked with ida2 glycosylated and deglycosylated amide black is shown in Figure 39.

The sequencing of both purified E1 and E2 products resulted in the disappointing observation that removal of the signal sequence from HCV wrap proteins is only partially occurring (see Table 3). (degradation products and products still containing the leader sequence or part of it) is glycosylated. This glycosylation is present, even in part. in the non-cleaved fragment of the signal sequence, which also contains a Nglycosylation site. These sites can be mutated to result in less glycosylated side products. However, even more problematic is the verification that some alternatively cleaved products are only 1 to 4 amino acids apart, compared to the desired intact wrap protein. Consequently, purification of the correctly processed product is virtually impossible due to the lack of sufficiently discriminating biochemical characteristics between the different expression products. Several of the degradation products can be a result of a Kex-2-like degradation (e.g. the divage observed after the aa 196 of El, which is a divage after an arginine), which is also required for divination of the α crossing factor leader and which can thus not be blocked without disturbing this essential process.

130

A high production clone of El, derived from the transformation of S. cerevisiae IYCC155 with pSYlYIG7Els (SEQ ID NO: 50; Figure 28), was compared with a high production clone derived from the transformation, of S. cerevisiae IYCC155 with pSYlaMFElsH6aYIGlEls (SEQ ID NO: 44, Figure 22). The intracellular expression of the E1 protein was assessed after 2 to 7 days after induction, and this was by Western blot using the E1-specific monoclonal antibody (IGH 201). As can be judged from Figure 40, 'maximum expression was observed after 2 days for both strains, but the expression patterns for both strains are completely different. The expression with the crossover factor α leader results in a very complex band pattern, which is a consequence of the fact that the leader processing is not efficient. This results in several expression products with a different amino terminus and some of which are modified by 1 to 5 Nglycosylations. However, for El expressed with the CL leader, a limited number of distinct bands are visible, which reflects the high level of correct removal of the CL leader and the fact that only this properly processed material can be modified by N-glycosylation (1 to 5 chains), as observed for El derived from Hansenula expressed with the

<td>same leader</td><td>from CL</td><td>(to see</td><td>the example</td><td> 16) .</td><td></td><td></td>

<td>THE</td><td>line</td><td>in</td><td>cells</td><td>hybridoma</td><td>producing</td><td>O</td>

<td>antibody</td><td colspan="2">monoclonal</td><td>directed</td><td>against El</td><td>(IGH201)</td><td>was</td>

<td>deposited</td><td>in 12</td><td>in</td><td>March of</td><td>1998, under the</td><td>conditions</td><td>of</td>

Budapest Treaty, at the European Collection of Cultures of

131

Cells (European Coilectión of Cell Cultures), Center for Applied Microbiology and Research, Salisbury, Wiltshire SP4 OJG, UK, and has accession number ECACC 98031216. The monoclonal antibody directed against E2 (IGH212) has been described as- the 12D11F2 antibody in Example 7.4 by Maertens et al., WO96 / 04385.

Table 3. Identification of the N ends of the proteins. OMF-E1-H6 and OMF-E2-H6 expressed in S. cerevisiae. or H. polymorpha. Based on the N-terminal sequencing, the number of N ends of the mature Ei-Ηβ and E2-H6 proteins could be estimated (mature indicating the correct removal of the oeMF signal sequence). The total amount of protein products was calculated as pmol of protein based on the intensity of the peaks recovered by Edman degradation. Subsequently, for each specific protein (ie for each 'N-tip detected'), molar% versus total was estimated.

132

<td>Yeast</td><td>aMF-El-H6</td><td>aMF-E2-VIEGR-H6</td>

<td>S. cerevisiae</td><td>Experience 1:</td><td> /</td>

<td></td><td>- 16% of the proteins still containing OtMF sequences - 18% of proteins cleaved between aa 195 and 196 of El - 66% of proteins with aMF correctly removed Experience 2: - 18% of proteins still containing aMF sequences - 33% of the proteins cleaved between aa 195 and 196 of El - 8% of other proteins in other El divage products - 44% of proteins with aMF correctly removed</td><td> /</td>

<td>H.polymorpha</td><td>- 64% of proteins still containing aMF sequences - 6% of cleaved proteins</td><td>- 75% of proteins still containing aMF strings</td>

<td></td><td>between aa 192 and 193 of El - 30% of proteins with aMF correctly removed</td><td>- 25% of proteins with aMF correctly removed)</td>

EXAMPLE 16 'EXPRESSION OF A YEAST CONSTRUCTION SUITABLE FOR LARGE-SCALE PRODUCTION AND PURIFICATION Several other leading sequences were used to replace the leading S. cerevisiae aMF peptide, including CHH (leading sequence of Carcinum maenas hyperglycemic hormone ), Amyl (leading sequence of amylase from S. occidentalis), Gaml (leading sequence of glycoamylase from S. occidentalis), Phy5 (lead sequence of fungal phytase), phol (lead sequence from Pichia pastoris acid phosphatase) and CL (leader of avian lysozyme C, 1,4-beta-N-acetylmuramidase C), and connected to E1-H6 (ie, El with his C-terminal mark).

All constructions were expressed in Hansenula polymorpha

133 and each of the resulting cell lysates was subjected to western blot analysis. This has already led to the conclusion that the degree of removal of the leading sequence or peptide or signal was extremely low, except for the construction in which CL is used as the peptide, leader. This was confirmed for the construction of CHH-E1-H6 by Edman's degradation of material purified by Ni-IDA: no properly cleaved product could be detected. although several different sequences were recovered (see Table 4).

Table 4. Identification of the N ends of the CHH-E1-H6 proteins expressed 'in H. polymorpha, based on the sequencing of N-terminal amino acids from different protein bands after separation by SDS-PAGE and staining on a membrane of PVDF.

<td>Molecular size</td><td>N ends identified</td>

<td>45kD</td><td>starts at amino acid 27 of the CHH leader = only the 'cleaved pre-sequence, the pro-sequence still linked</td>

<td>26kD</td><td>- starts partially at amino acid 1 of the CHH leader = no removal of the pre-pro-sequence - starts partially at amino acid 9 of the CHH leader = alternative translation product starting at the second AUG codon</td>

<td>24kD</td><td>- starts partially at amino acid 1 of the CHH leader = no removal of the pre-pro-sequence - starts partially at amino acid 9 of the CHH leader = alternative translation product starting at the second AUG codon)</td>

As already mentioned, western transfers of cell lysates revealed a pattern of specific protein bands for E1, indicative to a greater degree. correct removal of the leading CL peptide. This is surprising, since this leader is not derived from a

134 yeast. The sequencing of amino acids by Edman degradation of the material solubilized in GuHCl and purified by NiIDA in fact confirmed that 84% of the El proteins are correctly cleaved and the material is essentially free from degradation products. Still 16% of the unprocessed material is present, however, since this material is non-glycosylated, it can be easily removed from the mixture, allowing the specific enrichment of El to be properly cleaved and glycosylated. Such an enrichment method may be an affinity chromatography on lectins, the other alternatives are also given in Example 19. Alternatively, the greater hydrophobic character of the non-glycosylated material can be used to select and optimize other enrichment procedures. The correct removal of the CL leader peptide from the CL-E1-H6 protein was further confirmed by mass spectrometry, which also confirmed that up to 4 out of 5 Eis N-glycosylation sites of the lb genotype can be occupied, with which the sequence NNSS (amino acids 233 to 236; SEQ ID NO: 73) is considered to be a unique N-glycosylation site.

EXAMPLE 17

PURIFICATION AND BIOCHEMICAL CHARACTERIZATION OF PROTEIN E2 FROM

HCV EXPRESSED IN HANSENULA POLYMORPHA FROM THE

CONSTRUCTION OF CL-E2-H6 CODIFICATION The efficiency of removing the leading CL peptide from the CL-E2-VIEGR-H6 protein (additionally in this Example referred to as CL-E2-H6), expressed in Hansenula polymorpha, was analyzed. Since HCV E2s (aa 383-673) were

135 expressed as a protein with, 'his mark, a rapid and efficient' purification of the protein expressed after solubilization with GuHCl of the collected cells was carried out on Ni-IDA. In summary, the cell pellets were resuspended in 30 mM. phosphate, 6 M GuHCl, pH 7.2 (9 ml buffer / g cells). The protein was sulfonated overnight, at room temperature, in the presence of 320 mM (4% w / v) of sulfite. sodium and 65 mM (2% w / v) of sodium. sodium tetrathionate. The lysate was purified, after a freeze-thaw cycle, by centrifugation (10,000 g, 30 min, 4 ° C). Empigen BB (Albright & Wilson) and imidazole were added to a final concentration of 1% (w / v) and 20 mM, respectively. All additional chromatographic steps were performed on an Àkta FPLC workstation (Pharmacia). The sample was filtered through a 0.22 µm pore size membrane (cellulose acetate) and loaded onto a NiIDA column (Ni-loaded chelating Sepharose FF2+, Pharmacia), which was equilibrated with 50 mM phosphate, 6 M GuHCl, 1% Empigen BB, pH 7.2 (buffer A) supplemented with 20 mM imidazole. The column was washed sequentially with buffer A containing 20 mM and 50 mM imidazole, respectively, until the absorbance at 280 nm reached the baseline level. The products marked with his were eluted by application of buffer D, 50 mM phosphate, 6 M GuHCl, 0.2% Empigen BB (pH 7.2), 200 mM imidazole. The purified materials were analyzed by SDS-PAGE and western blot using a specific monoclonal antibody directed against E2

136 (IGH212) (Figure 41). The E2-H6 protein purified by IMAC was also subjected to N-terminal sequencing by Edman degradation. Then, the proteins were treated with N-glycosidase F (Roche) (0.2 U / pg E2, 1 h incubation at 37 ° C in PBS / 3% empigen BB) or left untreated. The glycosylated and deglycosylated E2-H6 proteins were subjected to SDS-PAGE and stained on a PVDF membrane for amino acid sequencing (the analysis was carried out on a PROCISE® 492 protein sequencer, Applied Biosystems). Since at this stage the SDS-PAGE revealed some degradation products, an additional fractionation was carried out using size exclusion chromatography. Then, the Ni-IDA eluate was concentrated by ultrafiltration (MWCO 10 kDa, centriplus, Amicon, Millipore) and loaded onto a Superdex G200 (Pharmacia) in PBS, 1% Empigen BB. The elution fractions, containing mainly products related to intact E2s, with a Mr between ~ 30 kDa and ~ 70 kDa based on migration on SDS-PAGE, were pooled and finally alkylated (incubation with 5 mM DTT for 30 min, at 37 ° C, followed by incubation with 20 mM iodoacetamide for 30 minutes, at 37 ° C). The possible presence of degradation products after purification by IMAC can thus be overcome by additional fractionation of the intact product by means of size exclusion chromatography. An unexpectedly good result was obtained. Based on the N-terminal sequencing, it was possible to estimate the amount of E2 product from which the CL leader peptide is removed. The total amount of protein products is

137 calculated as pmol of, protein based on peak intensity recovered by Edman degradation. Subsequently, for each specific protein. (ie, for each 'N-tip detected') molar% versus total is estimated. In the present experiment, only the correct N end of

E2-H6 was detected and the other E2-H6 variants lacking the E2 protein amino acid or containing Nterminal amino acids. not included in the E2 protein were absent., In conclusion, the E2-H6 protein, expressed by H. .polymorpha as CL-E2-H6 protein, was isolated, without any additional in vitro processing, as a> 95% correctly cleaved protein . This is in sharp contrast to the fidelity of the removal of the leader peptide by H. polymorpha from the OMF-E2-H6 protein to the E2-H6 protein, which was estimated to occur in -25% of the isolated proteins (see Table 3).

EXAMPLE 18

PURIFICATION AND BIOCHEMICAL CHARACTERIZATION OF THE PROTEIN El DE

HCV EXPRESSED IN HANSENULA POLYMORPHA FROM THE

CONSTRUCTION OF CL-H6-K-E1 CODING AND IN PROCESSING

HIT-CONTAINING PROTEIN VITRO

The efficiency of removing the leading CL peptide from the CL-H6-K-E1 protein expressed in H. polymorpha was analyzed, as well as the efficiency of subsequent in vitro processing to remove the H6 adapter peptide (his-brand) and the processing site by Endo Lys-C. Since HCV Eis (aa 192-326) was expressed as a N-terminal His-K branded protein CL-H6-K-E1, rapid and efficient purification could be performed, as described in

138

Example 17. The elution profile of the IMAC chromatographic purification of the H6-K-E1 proteins (and possibly the residual CL-H6-K-E1) is shown in Figure 42. After SDS-PAGE and silver staining of the gel and western blot analysis using a specific monoclonal antibody, directed against El (IGH201) (Figure 43), the elution fractions (63—69) containing the recombinant Eis products were pooled (^ meeting by IMAC ') and subjected to a treatment with Endoproteinase Lys-C (Roche) (enzyme / substrate ratio of 1/50 (w / w), 37 ° C) in order to remove the H6- K. The removal of the unprocessed melt was carried out by a negative IMAC chromatography step on. a Ni-IDA column, with which the proteins processed by Endo-Lys-C are collected in the flow fraction from end to end. With reference to this, the protein sample digested with Endoproteinase Lys-C was applied on a Ni-IDA column after a 10-fold dilution with 10 mM NaH2POWDER4.3H20.1% (v / v) Empigen B, pH 7.2 (buffer B), followed by washing with buffer B until the absorbance at 280 nm reached the baseline level. The end-to-end flow was collected in different fractions (1-40) that were examined for the presence of Eis products (Figure 44). Fractions (7-28), containing intact El from which the N-terminal H6-K end (and possibly the residual CL-H6-K) are removed (with a Mr between ~ 15 kDa and ~ 30 kDa with based on migration on SDS-PAGE, followed by silver staining or western blot analysis using a specific monoclonal antibody, directed against El

139 (IGH201))., Were1 pooled and alkylaclas (incubation with 5 mM DTT for 30 minutes at 37 ° C, followed by incubation with 20 mM iodoacetamide for 30 minutes at 37 ° C);

This material was subjected to N5 terminal sequencing (Edman degradation). Then, the protein samples were treated with N-glycosidase F (Roche) (0.2 U / gg of El, 1 h of incubation at 37 ° C in PBS / 3% empigen BB) or left untreated. Glycosylated proteins e. Deglycosylated cells were then separated by SDS-PAGE and stained on a .PVDF membrane for further analysis by Edman degradation (the analysis was carried out on a PROCISE® 492 protein sequencer, Applied Biosystems). Based on the Ν-terminal sequencing, the amount of correctly processed El product could be estimated (the processing includes the correct dividing of the H6K sequence). The total amount of protein products is calculated as pmol of protein based on the intensity of the peaks recovered by Edman degradation. Subsequently, for each specific protein (ie, for each molar 'N-tip detected' versus the total is estimated. In the present experiment, only the correct N-tip of El was detected and not the N-ends of other processing variants of H6-K-E1. Based on this, in vitro Endo Lys-C processing of the H6-K-E1 El protein (and possibly the residual CL-H6-K-E1) until the El protein was estimated to occur with a fidelity of more than 95% .

140

EXAMPLE 19

SPECIFIC REMOVAL OF LOW GLYCOSILATED FORMS OF HCV

BY HEPARINA

In order to find. specific purification steps for HCV wrap proteins from yeast cells, heparin binding was assessed. Heparin is known to bind to several viruses and, consequently, binding to the HCV envelope has already been suggested (Garson, JA et al. 1999). In order to analyze this potential link, heparin was biotinylated and the interaction

<td>with El</td><td>in</td><td>HCV</td><td>analyzed on plates</td><td>microtitre</td>

<td>coated</td><td>with</td><td>El</td><td>sulfonated HCV</td><td>from H.</td>

<td>polymorpha,</td><td>El</td><td colspan="2">alkylated HCV from</td><td>from H. polymorfa</td>

(both produced as described in Example 16) and alkylated HCV E1 from a culture of mammalian cells transfected with a vaccinia expression vector. Surprisingly, a strong bond could only be seen with the sulfonated HCV El from H. polymorpha, while the HCV El binding from the mammalian cell culture was completely absent. Through western transfer, we were able to show that this binding was specific to the lower molecular weight bands of the HCV E1 protein mix (Figure 45), corresponding to the low glycosylated mature HCV Eis. Figure 45 also reveals that sulfonation is not essential for heparin binding, since with the removal of this sulfonation, binding is still observed for low molecular weight El (path 4). Alternatively, the

141 alkylation is 'reduced, this bond going substantially, however, this may be caused by the specific alkylating agent' (iodine-acetamide) used in this example. This verification additionally demonstrated the industrial applicability of the. CL-HCV envelope expression cassettes for yeast, since we can specifically enrich HCV El preparations with respect to a preparation with HCV El proteins with a higher degree of glycosylation (ie, more occupied glycosylation sites) .

EXAMPLE. 20

FORMATION AND ANALYSIS OF VIRUS-LIKE PARTICLES (VLPs), The conversion of HCV El and E2 envelope proteins, expressed in H. polymç> rpha (Examples 16 to 18), in

VLPs were performed essentially as described by Depla et al. in WO99 / 67285 and by Bosmán et al. in W001 / 30815. Briefly, after culturing the transformed H. polymorpha cells, during which the HCV envelope proteins were expressed, the cells were collected, lysed in GuHCl 'and sulfonated as described in Example 17. The His-tagged proteins were subsequently purified by IMAC and concentrated by ultrafiltration, as described in Example 17.

Formation of VLPs from HCV wrap proteins with sulfonated Cys-thiol groups The concentrated HCV wrap proteins, sulfonated during the isolation procedure, were not subjected to a reducing treatment and were loaded onto a size exclusion chromatography column.

142 (Superdex G200, Pharmacia), balanced with PBS, 1% (v / v) Empigen. The eluted fractions were analyzed by SDS-PAGE and western blot. Fractions with a relative Mr ~ 29 ~ 25 kD (based on migration over.SDS-PAGE) were pooled, concentrated and loaded on Süperdex G200, balanced with PBS, 3% (w / v) betaine, to force formation virus-like particles (VLPs). The fractions were combined, concentrated and desalted to PBS, 0.5% (w / v) betaine.

Formation of VLPs from HCV wrap proteins with irreversibly modified Cys-thiol groups

The concentrated HCV wrap proteins, sulfonated during the isolation procedure, were subjected to a reducing treatment (incubation in the presence of

5 mM DTT in PBS) to convert the sulfonated Cys-thiol groups to the free Cys-thiol groups. The irreversible modification of Cys-thiol was carried out by (i) incubation for 30 min in the presence of 20 mM iodoacetamide, or (ii) incubation for 30 min in the presence of 5 mM N-ethylmaleimide (NEM) and 15 mM of biotin-N-ethylmaleimide. The proteins were subsequently loaded onto a size exclusion chromatography column (Superdex G200, Pharmacia), balanced with PBS, 1% (v / v) Empigen, in the case of iodoacetamide blocking, or with 0.2% PBS of CHAPS, in case of blockade with NEM and biotin-NEM. The eluted fractions were analyzed by SDS-PAGE and Western blot. Fractions with a relative Mr ~ 29—15 kD (based on migration over SDS-PAGE) were pooled, concentrated and, to force

143 the formation of virus-like particles, loaded on a Supérdex G200 column, balanced with PBS, 3% (w / v) betaine. The fractions were combined, concentrated and desalified, for PBS, 0.5% (w / v) betaine, in the case of blockade with iodoacetamide, or with PBS, 0.05% CHAPS, in the case of blockage with NEM and biotin -NOR.

Formation of VLPs from HCV wrap proteins with • reversibly modified Cys-thiol groups The concentrated, sulphonated 'HCV wrap proteins during the isolation procedure were subjected to a reducing treatment (incubation in the presence of 5 mM DTT in PBS) to convert the sulfonated Cys-thiol groups to the free Cys-thiol groups. The reversible modification of Cys-thiol was performed by incubation for 30 min in the presence of dithiodipyridine · (DTDP), dithiocarbamate (DTC) or cysteine. The ppoteins were subsequently loaded onto a size exclusion chromatography column (Superdex G200, Pharmacia), equilibrated with PBS, 1% (v / v) Empigen. The eluted fractions were analyzed by SDS-PAGE and Western blot. Fractions with a relative Mr ~ 29 ~ 15 kD (based on migration on SDS-PAGE) were pooled, concentrated and loaded onto Superdex G200, equilibrated with PBS, 3% (w / v) betaine, to force the formation of virus-like particles (VLPs). The fractions were combined, concentrated and desalified for PBS, 0.5% (w / v) betaine.

The size exclusion chromatography elution profiles in PBS, 3% (w / v) betaine, to obtain VLPs of

144

Ε2-Η6 expressed in H. polymorpha, are shown in Figure 46 (sulfonated) and Figure 47 (alkylated with iodoacetamide).

The size exclusion chromatography elution profiles in PBS, 3% (w / v) betaine, to obtain VLPs of

El expressed in H. polymorpha, are shown in Figure 48 (sulfonated) and Figure 49 (alkylated with iodoacetamide). The resulting VLPs were analyzed by SDS-PAGE 'and western blot, as shown in Figure 50.

Analysis of the size of VLPs formed by HCV envelope proteins expressed in H. polymorpha

The particle size of the VLPs was determined by Dynamic Light Diffusion. For the light diffusion experiments, a particle size analyzer (Model Zetasizer 1000 HS, Malvern Instruments Ltd.,

Malvern, Worcester UK), which was controlled by photon correlation spectroscopy (PCS) software. Photon correlation spectroscopy or dynamic light scattering (DLS) is an optical method that measures Brownian motion and relates it to particle size. The light from a visible, continuous laser beam is directed through a set of macromolecules or particles in suspension that move under Brownian motion. Something from the laser light is scattered by the particles and this diffused light is measured by a photomultiplier. Fluctuations in the intensity of diffused light are converted into electrical pulses, which are fed to a correlator. This generates the autocorrelation function, which is passed to a computer, where the appropriate data analysis is performed. The laser used was a

145 coherent 10 mW monochrome He-Ne 'laser with a fixed wavelength of 633 nm. For each sample, three to six consecutive measurements were taken1.

The results of these experiments are summarized in Table 5. 5..

Table 5: Results of the analysis by dynamic light diffusion on the indicated VLP compositions of HCV envelope proteins expressed by H. polymorpha. The particle sizes of the VLPs are given as the average particle diameter.

<td>Modification of Cys-thiol</td><td>E1-H6</td><td>E2-VIEGR-H6</td><td>El</td>

<td>sulfonation</td><td>25-45 nm</td><td>. 20 nm</td><td>20-26 nm</td>

<td>alkylation (iodoacetamide)</td><td>23-56 nm</td><td>20-56 nm</td><td>21-25 nm</td>

The observation that the sulfonated HCV El, derived from H. polymorpha, still forms particles with a size in the same range as the alkylated HCV El from Hansenula is surprising. Such an effect was not expected, since a high net increase (up to 8 Cys-thiol groups can be modified on the HCV E1) of negative charges as a consequence of the sulfonation should induce an ionic repulsion between the subunits. The other reversible cysteine modifying agents tested also allowed particle formation, the HCV E1 produced in this mode, however, proved to be less stable than the sulfonated material, resulting in aggregation of the disulfide-based HCV E1. In order to use these other blockers

146 reversible, additional optimization of conditions is required.

EXAMPLE 21

ANTIGENIC EQUIVALENCE OF HC1 E1-H.6 PRODUCED BY

HANSENULA AND HCV PRODUCED 'BY MAMMALIAN CELLS

INFECTED WITH VACCINIA

The reactivity of HCV E1-H6 produced by

Hansenula with chronic HCV carrier sera was compared to HCV El reactivity produced by mammalian cells infected with HCV-virus- of recombinant vaccinia, as described by Depla et al. in WO99 / 67285. Both HCV El preparations tested consisted of VLP's in which. HCV E1 proteins were alkylated with NEM and biotinNEM. The reactivities of both HCV El VLP preparations with HCV serum carriers were determined by ELISA. The results are summarized in Table 6. As can be derived from Table 6, no difference in reactivity was observed between HCV E1 expressed in mammalian cells infected with HCV20 recombinant vaccinia virus and HCV E1 expressed in H. polymorpha.

Table 6: The antigenicity of El produced in a culture of mammalian cells or produced in H. polymorpha was evaluated on a panel of sera from chronic human HCV carriers. For this purpose, biotinylated El was linked to streptavidin-coated ELISA plates. After that, human sera were added in a 1/20 dilution and immunoglobulins

147 of the serum bound to El were detected with. a specific secondary anti-human IgG-Fc antibody in rabbit, killed with peroxidase. The results are expressed as OD values. The mean values are the mean values, OD of all tested serum samples.

<td>Serum</td><td>Hansenula</td><td>mammal</td><td>Serum</td><td>Hansenula</td><td>mammal</td>

<td> 55333</td><td> 1,217</td><td> 1,168</td><td>average</td><td> 1,313</td><td> 1,245</td>

EXAMPLE 22

IMMUNOGENIC EQUIVALENCE OF HC1 E1-H6 PRODUCED BY

HANSENULA AND HCV ELECTED BY VACCINIA-INFECTED MAMMALIAN CELLS

The immunogenicity of HCV E1-H6 produced by

Hansenula was compared to HCV E1 immunogenicity produced by mammalian cells infected with HCV-recombinant vaccinia virus, as described by Depla et al. in WO99 / 67285. Both HCV El preparations tested

148 consisted of VLP 's in which HCV E1 proteins were alkylated with iodoacetamide. Both VLP preparations were formulated with alum and injected into Balb / c mice (3 intramuscular / subcutaneous injections with an interval of three weeks between each and each 'consisting of 5 pg of El in 125 μΐ containing 0.13% of Alhydrogel, Superfos, Denmark). The mice were bled ten days after the third immunization.

The results of this experiment are shown in Figure 51. For the top part of Figure 51, the antibodies produced following immunization with VLPs of E1 produced in mammalian cells were determined. Antibody titers were determined by ELISA (see Example 21), in which the El produced in mammalian cells (M) or the El produced in Hansenula (H) were coated directly on the solid ELISA support, after which the plates ELISA strains were blocked with casein. For the bottom part of Figure 51, antibodies produced following immunization with VLPs of El produced in Hansenula were determined. Antibody titers were determined by ELISA (see Example 21), in which the El produced in mammalian cells (M) or the El produced in Hansenula (H) were coated directly on the solid ELISA support, after which the plates ELISA strains were blocked with casein.

The antibody titers determined were the end point titers. The end point titer is determined as the dilution of the serum that results in an OD

149 (as determined by ELISA) equal to twice the average of the assay base.

Figure 51 shows. that no significant difference was observed between the immunogenic properties of both El compositions and that the titers of the antibodies determined are independent of the antigen used in the ELISA to perform the endpoint titration.

Yeast-derived HCV E1 induced a protective response similar to the protective response obtained in vaccination with alkylated HCV E1 derived from mammalian cell culture in vaccination. This response was able to prevent the chronic evolution of HCV after an acute infection.

EXAMPLE 23

ANTIGENIC AND IMMUNOGENIC PROFILE OF HC1 E1-H6 PRODUCED

BY 'HANSENULA THAT IS SULPHONATED The reactivity of HCV E1-H6 produced by

Hansenula with chronic HCV carrier sera was compared to HCV El reactivity produced by mammalian cells infected with HCV-recombinant vaccinia virus, as described by Depla et al. in WO99 / 67285. Both of the tested HCV El preparations consisted of VLP's in which the HCV El proteins produced by Hansenula were sulfonated and the HCV El produced by mammalian cells was alkylated. The results are given in Table 7. Although the overall (mean) reactivity was identical, some major differences were noted for the individual sera. This implies that the sulfonated material presents

150 at least some of its epitopes in a mode other than the alkylated HCV El.,

The immunogenicity of HCV E1-H6 produced by Hansenula, which was sulfonated, was compared to the immunogenicity of HCV E1-H6 produced by Hansenula, which was alkylated. Both HCV El preparations tested consisted of VLP's. Both VLP preparations were formulated with alum and injected into Balb / c mice (3 intramuscular / subcutaneous injections with an interval of 1 to 3 weeks between each and each consisting of 5 pg of El in 125 μΐ containing 0.13% of Alhydrogel, Superfos, Denmark). The mice were bled ten days after the third immunization.

Antibody titers were determined similarly as described in Example 22. Surprisingly, immunization with the sulfonated material resulted in higher antibody titers, independent of the antigen used in the ELISA to evaluate these titers (Figure 51; top panel: titration of antibodies produced against Al alkylated; bottom panel: titration of antibodies produced against Sulphonated el; A: Alkylated el coated on ELISA plate; S: El sulphonated coated on ELISA plate). However, in this experiment, the individual titers are different, depending on the antigen used for the analysis, which confirms the observation noted with the sera of HCV patients. Consequently, HCV El, in which the thiol groups of cysteine are reversibly modified, may be more immunogenic and thus have a

151 · increased potency of the vaccine 'protecting against HCV (chronic infection). In addition, the induction of a response to the neo - epitopes induced by. irreversible blockage is less likely to occur.

Table 7: The antigenicity of El alkylated (produced in cultured mammalian cells) or sulfonated E1-H6 (produced in H. polymorpha) was evaluated on a panel of sera from, chronic carriers, of human HCV (sera from patients). and a panel of control sera (serums from blood donors). For this purpose, El was linked to ELISA plates, after which the plates were further saturated with casein. Human sera were added at a 1/20 dilution and bound immunoglobulins were detected with a specific secondary anti-human IgG15 Fc antibody in rabbit, labeled with peroxidase. The results are expressed as OD values. The mean values are the mean OD values for all serum samples tested.

serum from patients_ 'blood donor serum

<td>serum number</td><td>Hansenula</td><td>mammal</td><td></td><td>serum number</td><td>Hansenula</td><td>mammal</td>

<td> 55333</td><td> 0, 69</td><td> 0, 534</td><td></td><td>average</td><td> 0,052</td><td> 0,054</td>

<td>average</td><td> 0,706</td><td> 0,691</td><td colspan="2"></td><td></td><td></td>

152

EXAMPLE 24

HCV E1-H6 IDENTIC ANTIGENIC REACTIVIPAPE PRODUCED

BY HANSENULA AND EL HCV PRODUCED BY MAMMALIAN CELLS

INFECTED WITH VACCINATION WITH SERUM, FROM VACCINATED CHIMPANZES '5

The reactivities of E1 produced by mammalian cells infected with HCV-vaccinia virus and E1-H6 produced by Hansenula (both alkylated) were compared with the sera of vaccinated chimpanzees and with monoclonal antibodies. For that purpose, said El proteins were coated directly on ELISA plates, followed by saturation of the plates with casein. Endpoint titers of antibodies binding coated E1 proteins to ELISA plates were determined for chimpanzee sera and specific murine monoclonal antibodies, all obtained from animals immunized with E1 produced by mammalian cells. The determination of the end point titer was made as described in Example 22. The monoclonal antibodies used were IGH201 (see Example 15), IGH198 (IGH198 = 23C12 in Maertens et al. in WO96 / 04385), IGH203 (IGH203 = 15G6 in Maertens et al. in WO96 / 04385) and IGH202 (IGH202 = 3F3 in Maertens et al. in W099 / 50301).

As can be derived from Figure 53, the reactivities of 7 different chimpanzees are identical when tested with the protein E1 produced by Hansenula or by mammalian cells. The reactivities of monoclonal antibodies against HCV E1 are also almost equal. Two of the chimpanzees (Yoran and Marti) were involved in a prophylactic vaccine study and were able to remove a

153 infection, acute with, provocation, while an animal, control did not remove the infection. .The five other chimpanzees (Ton, Phil, Marcei, Peggy, Femma) were involved in therapeutic vaccination studies and showed a reduction in liver damage, as measured by. ALT in serum and / or histological activity index on liver biopsy, with immunizations with HCV El.

The results obtained in this experiment are clearly different from the findings of Mustilli et al. (Mustilli, ACe, col. 1999), who expressed HCV E2 protein in both Saccharomyces cerevisiae and Kluyveromyces lactis. The E2 produced by purified yeast was, however, different from HCV E2 produced by mammalian cells (CHO) in that less reactivity was observed with 'chimpanzee sera immunized with HCV E2 produced by mammalian cells, whereas reactivity with monoclonal antibodies was higher for HCV E2 produced by yeast.

EXAMPLE 25

HCV EL GLYCOPERFIL BY CARBOHYDRATE ELECTROPHORESIS

AIDED BY FLUOROPHORE (FACE)

Glycosylation profiles were compared between HCV E1 produced by Hansenula and HCV E1 produced by mammalian cells infected with HCV-recombinant vaccinia virus, as described by Depla et al. WO99 / 67285. This was done using fluorophore-assisted carbohydrate electrophoresis (FACE). To this end, oligosaccharides were released from Eis produced by

154 mammalian cells or by Hansenula by peptide-Nglycosidase (PNGase F) and labeled with ANTS (proteins E1 were alkylated with iodoacetamide before digestion with PNGase F). The ANTS-labeled oligosaccharides were '5 separated by PAGE on a 21% polyacrylamide gel, in a current of 15 mA, at 4 ° C, for 2-3 h. From Figure 54, it was concluded that the oligosaccharides in El produced by mammalian cells and E1-H6 produced by Hansenula migrate like oligomaltose, with a degree of polymerization between 7 and

11 monosaccharides. ' This indicates that Hansenula's expression system surprisingly results in an E1 protein that is not hyperglycosylated and that has sugar chains of similar length to the sugar chains added to E1 proteins produced in mammalian cells.

155

List of References

Agaphonov, MO, Beburov, MY, Ter Avanesyan, MD, è Smirnov, VN (1995) A disruption-replacement approach for the targeted integration of foreign genes in Hansenula polymorpha. Yeast 11: 1241-1247.

Agaphonov, MO, Trushkina, PM, Sohn, JH, Choi, ES, Rhee, SK, and Ter Avanesyan, MD (1999) Vectors for rapid selection of integrants with different plasmid copy numbers in the yeast Hansenula polymorpha DL1. Yeast

15:541-551)

Alber, T. and Kawasacki, G. (1982). Nucleotide sequence of the triose phosphate isomerase gene of Saccharomyces cerevisiae. J. Mol. Appl. Genet 1: 419-434.

Ammerer, G. (1983) Expression of genes in yeast using the ADCI promoter. Methods Enzymol. 101: 192-201.

Bailou, L., Hitzeman, RA, Lewis, MS, and Bailou,

CE (1991) Vanadate-resistant yeast mutants are defective in protein glycosylation. Proc. Natl. Acad. USA 88: 32093212.

Beekman, NJ, Schaaper, WM, Tesser, GI, Dalsgaard, K., Kamstrup, S., Langeveld, JP, Boshuizen, RS, and Meloen, RH (1997) Synthetic peptide vaccines: palmitoylation of peptide antigens by a thioester bond increases immunogenicity, J. Pept. Res. 50: 357-364.

Burns, J., Butler, J., and Whitesides, G. (1991) Selective reduction of disulfides by Tris (2carboxyethyl) phosphine. J. Org. Chem. 56: 2648-2650.

156

Cox, H., Mead, D., Sudbery, P., Eland, RM, Mannazzu, I., and Evans, L. (2000) Constitutive expression of recombinant proteins in the methylotrophic yeast Hansenula polymorpha using the PMA1 promoter. Yeast 16: 1191-1203.

• 5 Gregg, JM (1999) Expression in the methylotrophic yeast Pichia pastoris. In Gene expression systems: using nature for the art of expression, JM Fernandez and JP Hoeffler, ed (San Diego: Academic Press), p. 157-191.

Darbre, Al (1986) Practical protein chemistry: a handbook. Whiley & Sons Ltd.

Doms, RW, Lamb, RA, Rose, JK, and Helenius, A. (1993) Folding and assembly of viral membrane proteins. Virology 193: 545-562.

Elble, R. (1992) A simple and efficient procedure for transformation of yeast. Biotechniques 13: 18-20.

Gailit, J. (1993) Restoring free sulfhydryl groups in synthetic peptides. Anal. Biochem. 214: 334-335.

Garson, JA, Lubach, D., Passas, J., Whitby, K., 20 and Grant, PR (1999) Suramin blocks hepatitis C binding to human hepatoma cells in vitro. J. Med. Virol. 57: 238-242.

Gatzke, R., Weydemann, Ü., Janowicz, ZA, and Hollenberg, CP (1995) Stable multicopy integration of vector sequences in Hansenula polymorpha. Appl. Microbiol.

Biotechnol. 43: 844-849.

Gellissen, G. (2000) Heterologous protein production in methylotrophic yeasts. Appl. Microbiol. Biotechnol. 54: 741-750.

157

Grakoui, A., Wychowski, C., Lin, C., Feinstone, SM, and Rice, CM (1993) Expression and Identification of hepatitis C virus polypròtein cleavage products. J. Virol.

67:1385-1395.

Heile, JM, Fong, YL, Rosa, D., Berger, K., Saletti, G., Campagnoli, S., Bensi, G., Capo, S., Coates, S., Crawford, K., Dong, C., Wininger, M., Baker, G., Cousens, L., Chien, D., Ng, P., Archangel, P., Grandi, G., Houghton, M., and Abrignani, S. (2000 ) Evaluation for hepatitis C virus glycoproteins E2 for vaccine design: an endoplasmic reticulum-retained recombinant protein is superior to secreted recombinant protein and DNA-based vaccine candidates. J. Virol. 74: 6885-6892.

Helenius, A. (1994) How N-linked oligosaccharides affect glycoprotein folding in the endoplasmic reticulum.

Mol. Biol. Cell 5: 253-265.

Hermanson, GT (1996) Bioconj ugat.e techniques. San Diego: Academic Press. Herscovics, A. and Orlean, P. (1993) Glycoprotein biosynthesis in yeast. FASEB J. 7: 540-550.

Hijikata, M., Kato, N., Ootsuyama, Y., Nakagawa, M., and Shimotohno, K. (1991) Gene mapping of the putative structural region of the hepatitis C virus genome by in vitro Processing analysis. Proc. Natl. Acad. Know. USA

88:5547-5551.

Hitzeman, RA, Clarke, L., and Carbon, J. (1980) Isolation and characterization of the yeast 3m

158 phosphoglycerokinase gene (PGK) by an immunological screening technique. J. Biol. Chem. 255: 12073-12080.

Hollenberg, CP and Gellissen, G. (1997) Production of recombinant proteins by methylotrophic yeasts. Curr. Opin. Biotechnol. 8: 554-560.

Holmgren, A. (1979) Thioredoxin catalyses the reduction of insulin disulfides by dithiothreitol and dihydrolipoamide. J. Biol. Chem. 254: 9627-9632.

Jayabaskaran, C., Davison, PF, and Paulus, H. (1987) Facile preparation and some applications of an affinity matrix with a cleavable connector arm containing a disulfide bond. Prep. Biochem. 17: 121-141.

Julius, D., Brake, A., Blair, L., Kunisawà, R., and Thorner, J. (1984) Isolation of the putative structural gene for the lysine-arginine-cleaving endopeptidase required for Processing of yeast prepro-alpha -factor. Cell 37: 1075-1089.

Kalef, E., Walfish, PG, and Gitler, C. (1993) Arsenical-based affinity chromatrography of vicinal dithiolcontaining proteins: purification. of L1210 leukemia cytoplasmic proteins and the recombinant rat c-erb A beta 1 T3 receptor. Anal. Biochem. 212: 325-334.

Kato, N., Ootsuyama, Y., Tanaka, T., Nakagawa, M., Nakazawa, T., Muraiso, K., Ohkoshi, S., Hijikata, M., and Shimotohno, K. (1992) Marked sequence diversity in the putative envelope proteins of hepatitis C viruses. Virus Res. 22: 107-123.

159

Kawasaki, G., and Fráenkel, DG (1982) Cloning. of yeast glycolysis genes'by complementation. Biochem. Biophys. Res. Commuri. 108: 1107-1122. '

Klebe, RJ, Harriss, JV, Sharp, ZD, and Douglas., MG. (1983) A general method for polyethyleneglycol-induced genetic transformation of bacteria and yeast. Gene 25: 333-341.

Rumar, N., Kella, D., and Kinsella, JE (1985) A method for the controlled cleavage of disulfide bonds in proteins in the absense of denaturants. J. Biochem. Biophys.

Methods 11: 251-263.

Kumar, N., Kella, D. and Kinsella, JE (1986) Anomalous effects of denaturants on sulfitolysis of protein disulfide bonds. J. Peptide Prot. Res. 28: 586-592.

Maertens, G. and Stuyver, L. (1997) Genotypes and genetic variation, of hepatitis C virus. In The molecular medicine of viral hepatitis, TJ Harrison and AJ Zuckerman, eds John Wiley & Sons), p. 183-233.

Major, ME and Feinstone, SM (1997) The molecular virology of hepatitis C. Hepatology 25: 1527-1538.

Mustilli, AC, Izzo, E., Houghton, M. and Galeotti, CL (1999) Comparison of secretion of a hepatitis C virus glycoprotein in Saccharomyces cerevisiae and Kluyvermyces lactis. Res. Microbiol. 150: 179-187.

Negai, K. and Thogersen, HC (1984) Generation of beta-globin by sequence-specific proteolysis of a hybrid protein produced in Escherichia coli. Nature 309: 810-812.

160

Nielsen, PE (2001) Targenting double stranded DNA with peptide nucleic acid (PNA). Curr. Med. Chem. 8: 545550.

Okabayashi, K., Nakagawa, Y., Hayasuke, N., Ohi, H., Miura, M., Ishida, Y., Shimizu, M., Murakami, K., Hirabayashi, K., Minamino, H. and , (1991) Secretory expression of the human serum albumin gene in the yeast, Saccharomyces cerevisiae. J. Biochem. (Tokyo) 110: 103-110.

Orum, H. and Wengel, J. (2001) Locked nucleic acids: a promising molecular family for gene-function analysis and antisense drug development. Curr.Opin.Mol.Ther. 3: 239-243.

Padgett, KA and Sorge, JA (1996) Creating seamless junctions independent of restriction sites in PCR cloning. Gene 168: 31-35.

Pedersen, J., Lauritzen, C., Madsen, MT, and Weis,

DS (1999) Removal of N-terminal polyhistidine tags from recombinant proteins using engineered aminopeptidases. Protein Expr. Purif. 15: 389-400.

Pomroy, NC and Deber, CM (1998) Solubilization of hydrophobic peptides by reversible cysteine PEGylation. Biochem. Biophys. Commun. 245: 618-621.

Raymond, CK (1999) Recombinant protein expression in Pichia methanolica. In Gene expression systems: using nature for the art of expression, JM Fernandez and JP Hoeffler, eds. (San Diego: Academic Press), p. 193-209.

Rein, A., Ott, DE, Mirro, J., Arthur, LO,

Rice, W., and Henderson, LE (1996) Inactivation of murine

161 , ι leukemia virus by compounds that react with the zinc finger in the viral nucleocapsid protein. J. Virol. 70: 4966-4972.

Roggenkamp, R., Hansen, H., Eckart, M. Janowicz, Z. and Hollenberg, CP (1986) Transformation of the methylo.trophic. yeast Hansenula. · polymorpha by autonomous replication and integration vectors. Mol. Gen. Genet. 202: 302-308.

• Rosa, D., CampagnoliS., Moretto, C., Guenzi, E., Cousens, L., Chin, M., Dong, C., Weiner, AJ, chest, JY, Choo, Q.LÍ, Chien. , D., Piíeri, P., Houghton, M., and Abrignani, S. (1996) A quantitative test to estimate neutralizing antibodies to the hepatitis C virus: cytofluorimetric assessment of envelope glycoprotein 2 binding to target cells. Proc. Natl. Acad. Know. USA

93:1759-1763.

Rose, J, K. and Doms, RW (1988) Regulation of protein export from the endoplasmic reticulum. Annu. Rev.

Cell. Biol. 4: 257-288.

Russell, DW, Smith, M. Williamson, VM, and Young, ET (1983) Nucleotide sequence of the yeast alcohol dehydrogenase II gene. J. Biol. Chem. 258: 2674-2682.

Russel, PR (1983) Evolutionary divergence of the mRNA transcription initiation mechanism in yeast. Nature

301:167-169.

Russel, PR (1985) Transcription of the triosephosphate-isomerase gene of Schizosaccharomyces pombe initiates from a start point different from that in Saccharomyces cerevisiae. Gene 40: 125-130.

162

Russell, PR and Hall, BD (1983) The primary structure of the alcohol dehydrogenase gene from the fission yeast Schizosaccharomyces pombe. J. Biol. Chem. 258: 143-149.

Sambrook, J., Fritsch, EF, and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press.

Singh, R. and Kats, L. (1995) Catalysis of reductión de disulfide by selenol. Anal. Biochem. 232: 86-91.

Sohn, JH, Choi, ES, Kang, HA, Rhee, JS, and Rhee, SK (1999) A family of telomere-associated autonomously replicating sequences and their functions in targeted recombination in Hansenula polymorpha DL-1. J. Bacteriol. 181: 1005-1013.

Stevens, RC (2000) Design of high-throughput methods of protein production for. structural biology.

Structure. Fold. Des 8: R177-R185.

Stuyver, L., van Arnhem, W., Wyseur, A., Hernandez, F., Delaporte, E., and Maertens, G. (1994) Classification of hepatitis C viruses based on phylogenetic analysis of the envelope 1 and nonstructural 5B regions and identification of five additional subtypes. Proc. Natl. Acad. Know. USA 91: 10134-10138.

Sugrue, RJ, Cui, T., Xu, Q., Fu, J., and Chan,

YC (1997) The production of recombinant dengue virus and protein using Escherichia coli and Pichia pastoris. J.

Virol. Methods 69: 159-169.

Thakur, ML, DeFulvio, J., Richard, MD, and Park,

CH (1991) Technetium-99m labeled monoclonal antibodies:

163 evaluation of reducing .agents. Int .J. Rad. Appl. Instrum. B 18: 227-233. '

Vingerhoeds, MH, Haisma, HJ Belliot, SO, Smit, RH,, Crommelin, DJ, and Storm, G. (1996)

Immunoliposomes as' enzyme-carriers (immuno-enzymosomes) for antibody-directed enzyme prodrug therapy (ADEPT):

<td>optimization</td><td>of prodrug</td><td>activating</td><td>capacity. Pharm. Res.</td>

<td colspan="2">Wahlestedt, C.,</td><td>Smil, P.,</td><td>Good, L., Kela, J.</td>

<td>10 Johnsson, ¢.</td><td>Hokfelt, T.,</td><td>Broberger,</td><td>C., Porreca, F., Lai,</td>

<td>J., Ren, K.,</td><td>Ossipov, M.,</td><td>Koshkin, A.</td><td>, Jakobsen, N., Skouv,</td>

J. Oerum, H., Jacobsen, MH, and Wengel, J. (2000) Potent and nontoxic antisense oligonucleotides containing locked nucleic acids. Proc. Natl. Acad. Know. USA 97: 5633-5638.

Weydemann, U., Keup, P., Piontek, M., Strasser,

AW, Schweden, J. ,, Gellissen, G., and Janowicz, ZA (1995) High-level secretion of hirudin by Hansenula polymorphaauthentic Processing of three different preprohirudins. Appl. Microbiol. Biotechnol.'44: 377-385.

SEQUENCE LISTING <110> Innogenetics NV

<120> Constructions and methods for the expression of recurrent HCV envelope proteins <130> 134 PCT

<td> <17 0></td><td>Patentln version 3.1</td>

<td> <210> <211> <212> <213></td><td>1 18 PRT poultry lysozyme signal peptide</td>

<td> <220> <221> <222> <223></td><td>MISCELLANEOUS (2) . . (2) Xaa is Arg, Lys or Vai</td>

<td> <220> <221> <222> <223></td><td>MISCELLANEOUS (3) .. (3) Xaa is Ser, Ala, Vai, Arg or Met</td>

<td> <22 0> <221> <222> <22 3></td><td>MISCELLANEOUS (4). . (4) Xaa is Leu or Phe</td>

<td> <22 0> <221> <222> <22 3></td><td>MISCELLANEOUS (5). . (5) Xaa is Leu or Ala</td>

<td> <220> <221> <222> <223></td><td>MISCELLANEOUS (6). . (6) Xaa is Ile, Thr, Phe or Vai</td>

<220>

<221> MISCELLANEOUS <222> (7). . (7) <223> Xaa is Leu, Phe or Ala

<td> <220> <221> <222> <22 3></td><td>MISCELLANEOUS (8) -. (8) Xaa is Vai, Ile, Ala, Leu or Cys</td>

<22O>

<221> MISCELLANEOUS <222> (9) .. (9) <223> Xaa is Leu, Phe, Ala or Ile <220>

<221> MISCELLANEOUS <222> (10) .. (10) <223> Xaa is Phe, Leu, Ser or Pro <220>

<221> MISCELLANEOUS <222> (11) .. (11) <223> Xaa is Phe, Leu, Ser or Pro

<td rowspan="2"> <220> <221> <222> <223></td><td colspan="3">MISCELLANEOUS</td>

<td> <220> <221></td><td>MISCELLANEOUS</td><td></td><td></td>

<td> <223></td><td>Xaa is Pro,</td><td>Allah</td><td>or Ile</td>

<td> <221></td><td>MISCELLANEOUS</td>

<td> <221></td><td>MISCELLANEOUS</td>

<td> <221></td><td>MISCELLANEOUS</td><td></td>

<td> <221></td><td>MISCELLANEOUS</td><td></td>

Met Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa 1 5 10 15

Xaa Gly <210> 2 <211> 135 <212> PRT <213> hepatitis virus Ç <400> 2

Tyr Glu Vai Arg Asn Gonna Be Gly Met Tyr His Vai Thr Asn Asp Cys 15 10 15

Ser Asn Ser Ser Ile Vai Tyr Glu Ala Ala Asp Met Ile Met His Thr 20 25 30

Pro Gly Cys Vai Pro Cys Vai Arg Glu Asn Asn Ser Be Arg Cys Trp

<td>Go</td><td>Allah 50</td><td>Read</td><td>Thr</td><td>Pro</td><td>Thr</td><td>Read 55</td><td>Allah</td><td>Allah</td><td>Arg</td><td>Asn</td><td>Allah 50</td><td>To be</td><td>Go</td><td>Pro</td><td>Thr</td>

<td>Thr 65</td><td>Thr</td><td>Ile</td><td>Arg</td><td>Arg</td><td>His 70</td><td>Go</td><td>Asp</td><td>Read</td><td>Read</td><td>Go 75</td><td>Gly</td><td>Allah</td><td>Allah</td><td>Allah</td><td>Phe 80</td>

<td>Cys</td><td>To be</td><td>Allah</td><td>Met</td><td>Tyr 85</td><td>Go</td><td colspan="2">Gly Asp</td><td>Read</td><td>Cys 90</td><td>Gly</td><td>To be</td><td>Go</td><td>Phe</td><td>Read 95</td><td>Go</td>

<td>Cys</td><td>Asn</td><td>Cys 115</td><td>To be</td><td>Ile</td><td>Tyr</td><td>Pro</td><td>Gly 120</td><td>His</td><td>Ile</td><td>Thr</td><td>Gly</td><td>His 125</td><td>Arg</td><td>Met</td><td>Allah</td>

130 135 <210> 3 <211> 290 <212> PRT

<td colspan="2"><213> virus</td><td>gives</td><td colspan="2">hepatitis C</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

<td>His 1</td><td>Thr Arg</td><td>Go</td><td>To be 5</td><td>Gly Gly</td><td>Allah</td><td>Allah</td><td>Allah 10</td><td>To be</td><td>Asp</td><td>Thr</td><td>Arg</td><td>Gly 15</td><td>Read</td>

<td>Go</td><td>Be Leu</td><td>Phe 20</td><td>To be</td><td>Pro Gly</td><td>To be</td><td>Allah 25</td><td>Gin</td><td>Lys</td><td>Ile</td><td>Gin</td><td>Read 30</td><td>Go</td><td>Asn</td>

<td>Thr</td><td>Asn Gly</td><td>To be</td><td>Trp</td><td>His Ile</td><td>Asn</td><td>Arg</td><td>Thr</td><td>Allah</td><td>Read</td><td>Asn</td><td>Cys</td><td>Asn</td><td>Asp</td>

40 45

Ser Leu Gin Thr Gly Phe Phe Ala Ala Leu Phe Tyr Lys His Lys Phe 50. ES 60

Aen Ser Ser Gly Cys Pro Glu Arg Leu Ala Ser Cys Arg Ser Ile Asp € 5 70 75 80

Lys Phe Ala Gin Gly Trp Gly Pro Leu Thr Tyr Thr Glu Pro Asn Ser 85 90 95

Ser Asp Gin Arg Pro Tyr Cys Trp His Tyr Wing pro Arg Pro Cys Gly ·. 100 105 110

Ile Vai Pro Ala Ser Gin Vai Çys Gly Pro Vai Tyr Cys Phe Thr Pro 115 '120 125

Be Pro Go Go Go Gly Thr Thr Asp Arg Phe Gly Go Pro Thr Tyr 130 135 140

Asn Trp Gly Ala Asn. Asp Ser Asp Vai Leu Ile Leu Asn Asn Thr Arg 145 150 155 160

Pro Pro Arg Gly Asn Trp Phe Gly Cys Thr Trp Met Asn Gly Thr Gly 16S 170 17S

Phe Thr Lys Thr Cys Gly Gly Pro Pro Cys Asn Ile Gly Gly Ala Gly 1B0 185 190

Asn Asn Thr Leu Thr Cys Pro Thr Asp Cys Phe Arg Lys His Pro Glu 195 200 205

Thr Tyr Wing Arg Cys Gly Wing Ser Gly Pro Trp Leu Thr Pro Arg Cys 210 215 220

Met Vai His Tyr Pro Tyr Arg Leu Trp His Tyr Pro Cys Thr Vai Asn 225 230 235 240

Phe Thr Ile Phe Lys Vai Arg Met Tyr Vai Gly Gly Vai Glu His Arg 245 250 255

Phe Glu Ala Ala Cys Asn Trp Thr Arg Gly Glu Arg Cys Asp Leu Glu 260 265 270

Asp Arg Asp Arg Ser Glu Leu Ser Pro Leu Leu Leu Ser Thr Thr Glu 275 280 285

Trp Gin 290 <210> 4 <211> 141 <212> PRT <213> hepatitis C virus <400> 4

Tyr Glu Vai Arg Asn Gonna Be Gly Met Tyr His Vai Thr Asn Asp Cys 1 5 10 15

Ser Αεη Ser Ser Xle Vai Tyr Glu Wing Wing Asp Met Ile Met His Thr 20 2S 30

Pro Gly Cys Vai Pro Cys Vai Arg Glu Asn Asn Ser Be Arg Cys Trp 35 40 45

Vai Ala Leu Thr Pro Thr Leu Ala Ala Arg As »Ala Ser Vai Pro Thr 50 55 60

Thr Thr Ile Arg Arg Bis Vai Asp Leu Leu Vai Gly Ala Ala Ala Phe 65 70 75 80

Cys Ser Ala Met Tyr Vai Gly Asp Leu Cys Gly Ser Vai Phe Leu Vai 85 90 95

Ser Gin Leu Phe Thr Ile Ser Pro Arg Arg His Glu Thr Vai Gin Asp 100 105 110

Cys Asn Cys Ser lie Tyr Pro Gly His Ile Thr Gly His Arg Met Ala 115 120 125

Trp Asp Met Met Met Asn Trp His His His His His 130 135 140 <210> 5 <211> 301 <212> PRT <213> hepatitis C virus <400> 5

His Thr Arg Will Be Gly Gly Wing Wing Wing Asp Thr Arg Gly Leu 15 10 15

Gonna Be Leu Phe Ser Pro Gly Ser Ala Gin Lys Ile Gin Leu Vai Asn

25 , -30

Thr Asn Gly Ser Trp His Ile Asa Arg Thr Ala Leu Asa Cys Asa Asp 35 40 45

Ser Leu Gla Thr Gly Phe Phe Ala Ala Leu Phe Tyr Lys His Lys Phe 50 55 60

Wing Ser Sèr Gly Cys Pro Glu Arg Leu Wing Ser Cys Arg Ser Ile Asp 65 70 75 80

Lys Phe Ala Gin Gly Trp Gly Pro Leu Thr Tyr. Thr Glu Pro Asa Ser 85 90 95

Ser Asp Gla Arg Pro Tyr Cys, Trp His Tyr Ala Pro Arg Pro Cys Gly 100 105 110

Ile Vai Pro Ala Ser Gla Vai Cys Gly Pro Vai Tyr Cys Phe Thr Pro 115 120 125

Be Pro Go Go Go Gly Thr Thr Asp Arg Phe Gly Go Pro Thr Tyr 130 135 140

Wing Trp Gly Wing Wing Asp Ser Asp Vai Leu Ile Leu Asn Wing Thr Arg 145 150 155 l € 0

Pro Pro Arg Gly Asn Trp Phe Gly Cys Thr Trp Met Asn Gly Thr Gly 165 170 175

Phe Thr Lys Thr Cys Gly Gly Pro Pro Cys Wing Ile Gly Gly Wing Gly 180 185 190

Asn Asn Thr Leu Thr Cys Pro Thr Asp Cys Phe Arg Lys His Pro Glu 195 200 205

Thr Tyr Ward Arg Cys Gly Ser Gly Pro Trp Leu Thr pro Arg Cys 210 21S 220

Met Vai His Tyr Pro Tyr Arg Leu Trp His Tyr Pro Cys Thr Vai Asn 22S 230 235 240

Phe Thr Ile Phe Lys Vai Arg Met Tyr Vai Gly <3ly vai Glu His Arg 245 250 255

Phe Glu Ala Ala Cys Asn Trp Thr Arg Gly Glu Arg Cys Asp Leu Glu 260 265 270

Αερ Arg Aep Arg Ser Glu Leu Ser Pro Leu Leu Leu Ser Thr Thr Glu 275 280 285

Trp Gin will Ile Glu Gly Arg His Hib Hib His Kis His 230 295 300 <210> 6 <211> 3448 <212> DNA <213> Vector pGEMTElsH6 <400> 6 aatcactagt catagettga tttcctgtgt aagtgtaaag ctgcccgctt gcggggagag cgçteggtcg tccacagaat aggaaccgta catcacaaaa • caggcgtttc ggatacctgt aggtatctca gttcagcccg cacgacttat ggcggtgcta tttggtatct tccggcaaac cgcagaaaaa tggaacgaaa tagatccttt tggtctgaca cgttcatcca gcggccgcct gtattctata gaaattgtta cctggggtgc tccagtcggg gcggtttgcg ttcggctgcg caggggataa aaaaggccge atcgacgctc cccctggaag ccgcctttct gttcggtgta accgctgcgc cgccactggc cagagttctt gcgctctgct aaaccaccgc aaggatctca actcacgtta taaattaaaa gttaocaatg tagttgcctg gcaggtcgac gtgtcaccta tccgctcaca ctaatgagtg aaacctgtcg tattgggcgc gcgagcggta cgcaggaaag gttgctggcg aagtcagagg ctecctogtg cccttcggga ggtcgttcgc cttatccggt agcagccact gaagtggtgg gaagccagtt tggtagcggt agaagatcct agggattttg atgaagtttt cttaatcagt actccccgtc catatgggag aatagcttgg attccacaca agctaactca tgccagctgc tcttccgctt tcagctcact aacatgtgag tttttegata tggcgaaacc cgctctcctg agcgtggcgc tocaagctgg aactatcgtc ggtaacagga cctaactacg accttcggaa ggtttttttg ttgatctttt gtcatgagat aaatcaatct gaggcaccta gtgtagataa agctcccaac cgtaatcatg acataogagc cattaattgc attaatgaat cctcgctcac caaaggcggt caaaaggcca ggctccgccc cgacaggact ttccgaccct tttcfccatag gctgtgtgca ttgagtccaa ttagcagagc gctacactag aaagagttgg tttgcaagca ctacggggtc tatcaaaaag aaagtatata tctcagcgat ctacgatacg gcgttggatg gtcatagctg cggaagcata gttgcgctca cggccaacgc tgactcgctg aatacggtta gcaaaaggcc ccctgacgag ataaagatac gccgcttacc ctcacgctgt cgaacccccc cccggtaaga gaggtatgta aaggacagta tagctcttga gcagattacg tgacgctcag gatcttcacc tgagtaaact ctgtctattt ggagggctta

120

180

240

300

360

420

480

540 €00 €60

720

780

840

900

960

1020

1080

1140

1200

1260

1320

1380 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta 1440 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc 1500 gcctccatcc agtctattas ttgttgccgg gaagctagag taagtagttc gceagttaat 1560 agtttgcgca acgttgttgg cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt 1520 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ecccatgttg 16 BO tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca 1740 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatocgta 1800

<td>agatgctttt</td><td>ctgtgactgg</td><td>tgagtactca</td><td>accaagtcat</td><td>tctgagaata</td><td>ccgcgcccgg</td><td> 1860</td>

<td colspan="2">cgaccgagtt gctcttgccc</td><td>ggcgtcaata</td><td colspan="2">cgggataata gtgtatgaca</td><td>fcagcagaact</td><td> 1920</td>

<td>ttaaaagtgc</td><td>tcatcattgg</td><td>aaaacgttct</td><td>tcggggcgaa</td><td>aactctcaag</td><td>gatcttaccg</td><td> 1980</td>

<td>ctgttgagat</td><td>ccagttcgat</td><td>gtaacccact</td><td>cgtgcaccca</td><td>actgatcttc</td><td>agcatctttt</td><td> 2040</td>

<td>actttcacca</td><td>gcgtttctgg</td><td>gtgagcaaaa</td><td>acaggaaggc</td><td>aaaatgccgc</td><td>aaaaaaggga</td><td> 2100</td>

<td>ataagggcga</td><td>cacggaaatg</td><td>ttgaatactc</td><td>atactcttcc</td><td>tttttcaata</td><td>ttattgaagc</td><td> 2160</td>

<td>atttatcagg</td><td>gttattgtct</td><td>catgagcgga</td><td>tacatatttg</td><td>aatgtattta</td><td>gaaaaataaa</td><td> 2220</td>

<td>caaatagggg</td><td>ttccgcgcac</td><td>atttccccga</td><td>aaágtgccac</td><td>ctgtatgcgg</td><td>tgtgaaatac</td><td> 2280</td>

<td>cgcacagatg</td><td>cgtaaggaga</td><td>aaataccgca</td><td>tcaggcgaaa</td><td>ttgtaaacgt</td><td>taatattttg</td><td> 2340</td>

<td>ttaaaattcg</td><td>cgfctaaatat</td><td>ttgttaaatc</td><td>agctcatttt</td><td>ttaaccaata</td><td>ggcCgaaatc</td><td> 2400</td>

<td>ggcaaaatcc</td><td>ctt.ata.aatc '</td><td>aaaagaatag</td><td>accgagatag</td><td>ggttgagtgt</td><td>tgttccagtt</td><td> 2460</td>

<td>tggaacaaga</td><td>gtccactatt</td><td>aaagaacgtg</td><td>gactccaacg</td><td>tcaaagggog</td><td>aaaaaccgtc</td><td> 2520</td>

<td>tatcagggcg</td><td>atggcccact</td><td>acgtgaacca</td><td>tcacccaaat</td><td>caagtttttt</td><td>gcggtcgagg</td><td> 2580</td>

<td>tgccgtaaag</td><td>ctctaaatcg</td><td>gaaccctaaa</td><td>gggagccccc</td><td>gatttagagc</td><td>ttgacgggga</td><td> 2640</td>

<td>aagccggcga</td><td>acgtggcgag</td><td>aaaggaaggg</td><td>aagaaagcga</td><td>aaggagcggg</td><td>cgctagggcg</td><td> 2700</td>

<td>ctggcaagtg</td><td>tagcggtcac</td><td>gctgcgcgta</td><td>accaccacac</td><td>ccgccgcgct</td><td>taatgcgccg</td><td> 2760</td>

<td colspan="2">ctacagggcg cgtccattcg</td><td>ccattcaggc</td><td>tgcgcaactg</td><td>ttgggaaggg</td><td>cgatcggtgc</td><td> 2820</td>

<td>gggcctcttc</td><td>gctattacgc</td><td>cagctggcga</td><td>aagggggatg</td><td>tgctgcaagg</td><td>cgattaagtt</td><td> 2880</td>

<td>gggtaacgcc</td><td>agggttttcc</td><td>cagtcacgac</td><td>gttgtaaaac</td><td>gacggccagt</td><td>gaattgtaat</td><td> 2940</td>

<td>acgactcact</td><td>atagggcgaa</td><td>ttgggcccga</td><td>cgtcgcatgc</td><td>tcccggccgc</td><td>catggccg-cg</td><td> 3000</td>

<td>ggattccaat</td><td>gcatatgagg</td><td>tgcgcaacgt</td><td>çrtccgggatg</td><td>taccatgtca</td><td>cgaacgactg</td><td> 3060</td>

<td>ctccaactca</td><td>agcattgtgt</td><td>atgaggcagc</td><td>ggacatgafcc</td><td>atgcacaccc</td><td>cogggtgcgt</td><td> 3120</td>

<td>gccctgcgtt</td><td>cgggagaaca</td><td>actcttceeg</td><td>ctgctgggta</td><td>gcgcfccaocc</td><td>ccacgctcgc</td><td> 3180</td>

agctaggaac gccagcgtcc ccactacgac aatacgacgc cacgtcgatt tgctcgttgg 3240 ggcggctgct ttctgttccg ctatgtacgt gggggatctc tgcggatctg tcttcctcgt 3300 ctcccagctg ttcaccatct cgcctcgccg gcatgagacg gtgcaggact gcaattgctc 3360 aatctatccc ggccacataa caggtcaccg tatggcttgg gatatgatga tgaactggca 3420 ccaccaccat caccattaag gatccaag 3448 <210> 7 <211> 37 <212> DNA <213> primer or probe synthetic <400> 7 agttactctt caaggtatga ggtgcgcaac gtgtccg 37 <210> 8 <211> 47 <212> DNA <213> synthetic primer or probe <400> 8 agttactctt cacagggatc ctccttaatg gtgatggtgg tggtgcc 47 <210> 9 <211> 3067 <212> DNA <213> vector pCHH-Hir <400> 9

<td>gcgcccaata</td><td>cgcaaaccgc</td><td>ctctccccgc</td><td>gcgttggccg attcattaat</td><td>gcagctggca</td><td> 60</td>

<td>cgacaggttt</td><td>cccgactgga</td><td>aagcgggcag</td><td>tgagcgcaac gcaattaatg</td><td>tgagttagct</td><td> 120</td>

<td>cactcattag</td><td>gcaccccagg</td><td>ctttacactt</td><td>tatgcttccg gctcgtatgt</td><td>tgtgtggaat</td><td> 180</td>

<td>tgtgagcgga</td><td>taacaatttc</td><td>acacaggaaa</td><td>cagctatgac catgattacg</td><td>ccaagcttgc</td><td> 240</td>

<td>atgcctgcag</td><td colspan="2">gtcgacccta gatctctatt</td><td>actgcaggta ttcttccggg</td><td>atttctfccga</td><td> 300</td>

<td>agtcgccgtc</td><td>gttgtgagac</td><td>tgcggacgcg</td><td>gggtaccttc gccagtaacg</td><td>cactggttac</td><td> 360</td>

<td>gttcgccttt</td><td>agagcccagg</td><td>atgcatttgt</td><td>tgccctggcc gcaaacgtta</td><td>gagccttegc</td><td> 420</td>

<td>acaggcacag</td><td>gttctgaccg</td><td>gattcagtgc</td><td>agtcagtgta aacaaccctc</td><td>ttttccaacg</td><td> 480</td>

<td>ggtgtgtagt</td><td>tccattctcc</td><td>accgctaggg</td><td>ctgcgctggg ctccattggc</td><td>gaggttttca</td><td> 540</td>

<td>aggccgctag</td><td>gatgcgatcc</td><td>atgcgtccgt</td><td>agccttgcgt ggagcgtgcg</td><td>tgtgcgtgcg</td><td> 600</td>

<td>ggagtgcgca</td><td>taggtaggct</td><td>acggtgatga</td><td>ttgctagcat ggcgggaata</td><td>gttttgctat</td><td> 660</td>

<td>acatgaattc</td><td>actggccgtc</td><td>gttttacaae</td><td>gtogtgactg ggaaaaccct</td><td>ggcgttaocc</td><td> 720</td>

<td>aacttaatcg</td><td>ccttgcagca</td><td>catcccoctt</td><td>tcgccagctg gcgtaatagc</td><td>gaagaggccc</td><td> 780</td>

, ι

<td>gcaccgatcg</td><td>cccttcccaa</td><td>cagttgcgca</td><td>gcctgaatgg</td><td>cgaatggcgc i</td><td>ctgatgcggt</td><td>B40</td>

<td>attttctcct</td><td>tacgcatctg</td><td>tgcggtattt</td><td>cacaccgcat,</td><td>atggtgcact</td><td>ctcagtacaa</td><td> 900</td>

<td colspan="2">tctgctctga tgccgcatag</td><td>ttaagçcagc</td><td>cccgacaccc</td><td>gccaacaccc</td><td>gctgacgcgc</td><td> 960</td>

<td>cctgacgggc</td><td>ttgtctgctc</td><td>ccggcatccg</td><td>cttacagaca</td><td>agctgtgacc</td><td>gtctccggga</td><td> 1020</td>

<td>gctgcatgtg</td><td>tcagaggttt</td><td>tcaccgtcat</td><td>caccgaaacg</td><td>cgcgaga cga</td><td>aagggccteg</td><td> 1080</td>

<td>tgatacgcct</td><td>atttttatag</td><td>gttaatgtca</td><td>tgataàtaat</td><td>ggtttcttag</td><td>acgtcaggtg</td><td> 1140</td>

<td>gcactttVcg</td><td>gggaaatgtg</td><td>cgcggaaccc</td><td>ctatttgttt</td><td>atttttctaa</td><td>wholesale</td><td> 1200</td>

<td>atatgtatcc</td><td>gctcatgaga</td><td>caataaccct</td><td>gataaatgct</td><td>tcaataatat</td><td>tgaaaaagga</td><td> 1260</td>

<td>agagtatgag,</td><td>tattcaacat</td><td>ttccgtgtcg</td><td>cccttattcc</td><td>cttttttgcg</td><td>gcattttgcc</td><td> 1320</td>

<td>ttcctgtttt</td><td>tgctcaccca</td><td>gaaacgctgg</td><td>tgaaagtaaa</td><td>agatgctgaa</td><td>triggergg</td><td> 1380</td>

<td>gtgcacgagt</td><td>gggtfcacate</td><td>gaactggatc</td><td>tcaacagcgg</td><td>taagatcctt</td><td>gagagttttc</td><td> 1440</td>

<td>gccccgaaga</td><td>acgttttcca</td><td>atgatgagca</td><td>cttttaaagt</td><td>tctgctatgt</td><td>ggcgcggtat</td><td> 1500</td>

<td>tateccgtat</td><td>tgacgccggg</td><td>caagagcaac</td><td>teggtcgccg</td><td>catacactat</td><td>tctcagaatg</td><td> 1560</td>

<td>acttggttga</td><td>gtactcacca</td><td>gtcacagaaa</td><td>agcatcttac</td><td>ggatggcatg</td><td>acagtaagag</td><td> 1620</td>

<td>aattatgcag</td><td>tgctgccata</td><td>accatgagtg</td><td>ataacacfcge</td><td>ggccaactta</td><td>cttctgacaa</td><td> 1680</td>

<td>egatcggagg</td><td>accgaaggag</td><td>ctaacegctt</td><td>ttttgcacaa</td><td>catgggggat</td><td>catgtaactc</td><td> 1740</td>

<td>gccttgatcg</td><td>ttgggaaecg</td><td>gagctgaatg</td><td>aagccatacc</td><td>aaacgacgag</td><td>cgtgacacca</td><td> 1800</td>

<td>cgatgcctgt</td><td>agcaatggcá</td><td>acaacgttgc</td><td>gcaaactatt</td><td>aactggcgaa</td><td>ctacttactc</td><td> 1860</td>

<td>tagcttcccg</td><td>gcaacaatta</td><td>atagactgga</td><td>tggaggcgga</td><td>taaagttgca</td><td>ggaccacttc</td><td> 1920</td>

<td>tgcgctcggc</td><td>ccttccggct</td><td>ggctggttta</td><td>ttgctgataa</td><td>atctggagcc</td><td>ggtgagcgtg</td><td> 1980</td>

<td>ggtctcgcgg</td><td>tatcattgca</td><td>gcactggggc</td><td>cagatggtaa</td><td>gccctcccgt</td><td>atcgtagtta</td><td> 2040</td>

tctacacgac ggggagtcag gcaactatgg atgaaçgaaa tagacagatc gctgagatag 2100 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga 2160 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc 2220 tcatgaccaa aatcocttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa 2280 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa 2340 aaaaaccacc gctaccagcg gtggtttgtt tgccggafcca agagctacca actctttttc 2400 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta 2460 gtgtagccgt agttaggcca ccacttcaag aaofcctgtag caccgcctac atacctcgct ctgctaatcc 2S20 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccaggttg gactcaagac 2580 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca 2640 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg g-cgaggaag cggaaga <210> 10 <212> 35 <212> DNA <213> primer or synthetic probe <400> 10 agttactctt cacctctttt eeaacgggtg tgtag <210> 11 <211> 34 <212> DNA <213> primer or synthetic probe

2700

2760

2820.

2880

2940

3000

3060

3067 <400> 11 agtcactctt cactgcaggc atgcaagctt <210> 12 <211> 6973 <212> DNA <213> vector pFPMT121 <400> 12 ggtaccctgc tcaatctccg gaatggtgat tccctcctga cacaggtact cgtacaggtt gacaacgttc ttagaagcgg ccggcatttt aatttaattt tcattaaatt ttcctatact ataaacgata taaaccagaa aaagaactat ccttccacgt ctccttagaa tctggcaagt atgaagcaga gagcgcagga ggcggtattt gatggtagtc gagtgtatcg gagacagctt tattggogga caccagtgag acaccccgga accagtggga gcctatttgc acacacgagc ctgatcgttc ctgaaaacct cgacattggc ccaggtaaac gagtcgtagt tgtcgatcat gaaggtgact aatagcctaa gaaaatattt cgctatttca gcttttcatc tcatcacttc tttcaaacac gcttctcaaa agcggtatgt ccgcgagggg gatccagatc tgaattcccg atagtgccat tcccctctct gagagaccog gatgtagact ccgtgcctgc cggctcctct acttgctgtt tttctgcaaa atccggggtg gggacacccc actctggtga agagtgccaa

120

180

240

300

360

420

480

540

600 ágtcattctt tttcccgttg oggggcagcc gattgcatgt tttaggaaaa tattaccttt 660 gctacaccct gtçagattta ccctccacac atafataftc cgtcacctcc agggactatt 720 attcgtcgtt gcgccgccag cggaagatat acagaagctg ttttccgaga gactcggttg 780 g gcgcctggta tatttgatgg atgtcgcgct-cctcacgtc ccggtaccca ggaacgcggt 840 gggatctcgg gcccatcgaa gactgtgctc cagactgctc gcccagcagg tgtttcttga 900 tcgccgcctc taaattgtcc gcgcatcgcc ggtaacattt ttccagctcg gagtttgcgt 960

<td>ttagatacag tttctgcgat gccaaaggag cctgcagatt ataacctcgg </td><td>atgctgtcat</td><td> 1020</td>

<td>tcagcgcttt taatttgacc tccagatagt tgctgtattt ctgtteccat</td><td>tggctgctgc</td><td> 1080</td>

<td>gcagcttcgtataactcgag ttattgttgc gctctgcctc ggcgtactgg</td><td>ctcatgatct</td><td> 1140</td>

<td>ggatcttgtc cgtgtcgctt ttcttcgagt gtttctcgca aacgatgtgc</td><td>açggcctgca</td><td> 1200</td>

<td>gtgtccaatç ggagtcgagc tggcgccgaa actggcggat ctgagcctcc</td><td>acactgccct</td><td> 1260</td>

<td>gtttctctat ccacggcgga accgcctcct gccgtttcag aatgttgttc</td><td>aagtggtact</td><td> 1320</td>

<td>ctgtgcggtc aatgaaggcg ttattgccgg tgaaatcttt gggaagcggt</td><td>tttcctcggg</td><td> 1380</td>

<td>gaagattacg aaattccccg cgtcgttgcg cttcctggat ctcgaggaga</td><td>tcgttctccg</td><td> 1440</td>

<td>cgtcgaggag atcgttctcc gcgtcgacac cattccttgc Çgcggcggtg</td><td>ctcaacggcc</td><td> 1500</td>

<td>tcaacctact actgggctgc ttcctaatgc àggagtçgca taagggagag</td><td>cgtcgacaaa</td><td> 1560</td>

<td>cccgcgtttg agaacttgct caagcttctg gtaaacgttg tagtactctg</td><td>aaacaaggcc</td><td> 1620</td>

<td>ctagcactct gatctgtttc tcttgggtag oggtgagtgg tttattggag</td><td>ttcactggtt</td><td> 1680</td>

<td>tcagcacatc tgtcatctag acaatattgt tactaaattt ttttgaacta</td><td>caattgttcg</td><td> 1740</td>

<td>taattcatct attattatac atcctcgtea gcaatttctg gcagacggag</td><td>tttactaacg</td><td> 1800</td>

<td>tcttgagtat gaggccgaga atccagctct gtggccatac tcagtcttga</td><td>cagcctgctg</td><td> 1860</td>

<td>atgtggctgc gttcaacgca ataagcgtgt cctccgactc cgagttgtgc</td><td>tcgttatcgt</td><td> 1920</td>

<td>cgttctcatc ctcggaaaaa tcacacgaaa gaacatactc accagtaggc</td><td>tttctggtcc</td><td> 1980</td>

<td>ctggggcacg gctgtttctg acgtattccg gcgttgataa tagctcgaaa</td><td>gtgaacgccg</td><td> 2040</td>

<td>agtcgcggga gtcgaccgat gcccttgaga gccttcaacc cagtcagctc</td><td>cttccggtgg</td><td> 2100</td>

<td>ffcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat</td><td>gcaactcgta</td><td> 2160</td>

<td>ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg aocgctttcg</td><td>ctggagcgcg</td><td> 2220</td>

<td>acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc acgccctcgc</td><td>tcaagccttc</td><td> 2280</td>

<td>gtcactggtc ccgccaccaa acgtttcggc gagaagcagg ccattatcgc</td><td>cggcatggcg</td><td> 2340</td>

<td>gccgacgcgc tgggctacgt cttgctggcg ttcgcgacgc gaggctggat</td><td>ggccttcccc</td><td> 2400</td>

I

<td>attatgattc</td><td>ttctcgcttc</td><td>cggcggcatc</td><td>gggatgcccg</td><td>cgttgcaggc</td><td>catgctgtcc</td><td> 2460</td>

<td>aggcaggtag</td><td>atgacgaoca</td><td>tcagggacag</td><td>cttcaaggat</td><td>cgctcgcggc</td><td>tcttaccagc</td><td> 2520</td>

<td>ctaacttcga</td><td>tcactggacc</td><td>gctgatcgtc</td><td>acggcgattt</td><td>atgccgcctc</td><td>ggcgagcaca</td><td> 2580.</td>

<td>tggaacgggt</td><td>tggcatggat</td><td>tgtaggcgcc</td><td>gccctatacc</td><td>ttgtctgcct</td><td>ccccgcgttg</td><td> 2640</td>

<td>cgtcgcggtg</td><td>catggagccg</td><td>ggccacctcg</td><td>acctgaatgg</td><td>aagccggcgg</td><td>cacctcgcta</td><td> 2700</td>

<td>acggattcac</td><td>cactccaaga</td><td>attggagcca</td><td>atcaattctt</td><td>gcggagaact</td><td>gtgaatgcgc</td><td> 2760</td>

<td>aaaccaaccc</td><td>ttggcagaac</td><td>atatccatcg</td><td>cgtocgccat</td><td>ctccagcagc</td><td>cgcacgcggc</td><td> 2820</td>

<td>gcatcggggg</td><td>gggggggggg</td><td>gggggggggc</td><td>aaacaattca</td><td>tcattttttt</td><td>tttattcttt</td><td> 2880</td>

<td>tttttgattt</td><td>cggtttcttt</td><td>gaaatttttt</td><td>tgattcggta</td><td>afcctcegaac;</td><td>agaaggaaga</td><td> 2940</td>

<td>acgaaggaag</td><td>gagcacagac</td><td>ttagattggt</td><td>atatata-cgc</td><td>atatgtagtg</td><td>ttgaagaaac</td><td> 3000</td>

<td>atgaaattgc</td><td>ccagtattct</td><td>taacccaact</td><td>gcacagaaça</td><td>aaaacctgca</td><td>ggaaacgaag</td><td> 3060</td>

<td>ataaatcatg</td><td>tcgaaagcta</td><td>catataagga</td><td>acgtgctgct</td><td>actcatccta</td><td>gtcctgttgc</td><td> 3120</td>

<td>tgccaagcta</td><td>tttaatatca</td><td>tgcacgaaaa</td><td>gcaaacaaac</td><td>ttgtgtgctt</td><td>cattggatgt</td><td> 3180</td>

<td>tcgtaccacc</td><td>aaggaattac</td><td>tggagttagt</td><td>tgaagcatta</td><td>ggtcccaaaa</td><td>tttgtttaet</td><td> 3240</td>

<td>aaaaacacat</td><td>gtggatatct</td><td>tgactgattt</td><td>ttocatggag</td><td>ggcacagtta</td><td>agccgctaaa</td><td> 3300</td>

<td>ggcattatcc</td><td>gccaagtaca</td><td>attttttact</td><td>cttcgaagac</td><td>agaaaatttg</td><td>ctgacattgg</td><td> 3360</td>

<td>taatacagtc</td><td>aaattgcagt</td><td>actctgcggg</td><td>tgtatacaga</td><td>atagcagaat</td><td>gggcagacat</td><td> 3420</td>

<td>tacgaatgca</td><td>cacggtgtgg</td><td>tgggcccagg</td><td>tattgttagc</td><td>ggtttgaagc</td><td>aggcggcaga</td><td> 3480</td>

<td>agaagtaaca</td><td>aàggaaccta</td><td>gaggcctttt</td><td>gatgttagca</td><td>gaattgtcat</td><td>gcaagggctc</td><td> 3540</td>

<td>cctatctact</td><td>ggagaatata</td><td>ctaagggtac</td><td>tgttgacatt</td><td>gcgaagagcg</td><td>acaaagattt</td><td> 3600</td>

<td>tgttatcggc</td><td>tttattgctc</td><td>aaagagacat</td><td>gggtggaaga</td><td>gatgaaggtt</td><td>acgattggtt</td><td> 3660</td>

<td>gattatgaca</td><td>cccggtgtgg</td><td>gtttagatga</td><td>caagggagac</td><td>gcattgggtc</td><td>aacagtatag</td><td> 3720</td>

<td>aaccgtggat</td><td>gatgtggtct</td><td>ctacaggatc</td><td>tgacattatt</td><td>attgttggaa</td><td>gaggactatt</td><td> 3780</td>

<td>tgcaaaggga</td><td>agggatgcta</td><td>aggtagaggg</td><td>tgaacgttac</td><td>agaaaagcag</td><td>gctgggaagc</td><td> 3840</td>

<td>atatttgaga</td><td>agatgcggcc</td><td>agcaaaacta</td><td>aaaaactgta</td><td>ttataagtaa</td><td>atgcatgtat</td><td> 3900</td>

<td>actaaactca</td><td>caaattagag</td><td>cttcaattta</td><td>attatatcag</td><td>ttattaoccg</td><td>ggaatctcgg</td><td> 3960</td>

<td>tcgtaatgat</td><td>ttttataatg</td><td>acgaaaaaaa</td><td>aaaaattgga</td><td>aagaaaagcc</td><td>cccccccccc</td><td> 4020</td>

<td>cccccccccc</td><td>CCCCCCCCCC</td><td>ccgcagcgtt</td><td>gggtcctggc</td><td>chacgggtgcg</td><td>catgatcgtg</td><td> 4080</td>

<td>ctcctgtcgt</td><td>tgaagacccg</td><td>gctagg-ctgg</td><td>cggggttgcc</td><td>ttactggtta</td><td>gcagaatgaa</td><td> 4140</td>

<td>tcaccgatac</td><td>gcgagcgaac</td><td>gtgaagcgac</td><td>tgctgctgca</td><td>aaacgtctgc</td><td>gacctgagca</td><td> 4200</td>

<td>acaacatgaa</td><td>tggtcttcgg</td><td>tttcogtgtt</td><td>tcgtaaagtc</td><td>tggaaacgcg</td><td>gaagtcagcg</td><td> 4260</td>

.1 ccctgcacca ttatgttccg gatctgcatc gcaggatgct gctggctacc ctgtggaaca 4320 cctacatctg tattaacgaa gcgctggcat tgaccctgag tgatttttct ctggtcccgc 4380 cgcatccata. ccgccagttg tttacçctca caaçgttcca gtaaccgggc atgttcatca 4440 teagtaaccc gtatcgtgag catcctctct cgtttcatcg gtat.catt.ac ccccatgaac 4500 agaaattccc ccttacacgg aggcatcaag tgaccaaaca ggaaaaaacc gcccttáaca 4560 tggcccgctt tateagaagc cagacattaa cgcttctgga gaaactcaac gagctggacg 4620 cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca cgctgatgag ctttaccgca 4680 gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga 4740 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca ágcccgtcag ggcgcgtcag 4800 cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat agcggagtgt 4860 atactggctt aactatgcgg catcagagca gattgtactg agagtgcacc atatgcggtg 4920 tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt ccgcttcctc 4980 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa 5040 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa 5100 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct 5160 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac 5220 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc 5280 gaccctgccg cttaccggat 'acctgtccgc ctttctccct tcgggaagcg tggcgetttc 5340 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg 5400 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga 5460 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag 5520 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta 5580 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag 5640 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg 5700 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac 5760 ggggrtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc 5820 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag 5880 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc 5940 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac 6000 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc 6060 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg 6120 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag 6180 tagttcgcca gttaatagtt tgcgcaacgt tgcgggggtggggggtg

acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac 6300 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag 6360 aagtaagttg gccgcagtgt tatcacfccat ggttatggca gcactgeata attctcttac 6420 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg 6480 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaacacggg ataataccgc 6S40 gçcacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact 6600 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactegtg 6660 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa 6720

<td>tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt</td><td> €780</td>

<td>tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg</td><td> 6840</td>

<td>tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga</td><td> 6900</td>

<td>cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc</td><td> €960</td>

<td>ctttcgtctt caa</td><td> 6973</td>

<td><213> pPPMT-CHH-ElH6 vector</td><td></td>

<td>ggtaccctgc tcaatctccg gaatggtgat ctgatcgttc ctgaaaacct cgacattggc</td><td> 60</td>

<td>tccctcctga cacaggtact cgtacaggtt ccaggtaaac gagtcgtagt tgtcgatcat</td><td> 120</td>

<td>gacaacgttc ttagaagcgg ccggcatttt gaaggtgact aatagcctaa gaaaatattt</td><td> 180</td>

<td>aatttaattt tcattaaatt ttcctafcact cgctatttca gcttttcatc tcatcacttc</td><td> 240</td>

<td>ataaacgata taaaccagaa aaagaactat tttcaaacac gcttctcaaa agcggtatgt</td><td> 300</td>

<td>ccttccacgt ctccttagaa tctggcaagt ccgcgagggg gatccttaat ggtgatggtg</td><td> 360</td>

<td>gtggtgccag ttcatcatca tatoccaagc catacggtga cctgttatgt ggccgggata</td><td> 420</td>

<td>gattgagcaa ttgcagtcct gcaccgtctc atgccggoga ggcgagatgg tgaacagctg</td><td> 480</td>

<td>ggagacgagg aagacagatc cgcagagatc ccccacgtac atagcggaac agaaagcagc</td><td> 540</td>

<td>cgcccc & acc agcaaatcga cgtggcgtcg tattgtcgta gtggggacgc tggcgttoct</td><td> 600</td>

<td>agctgcgagc gtgggggtga gcgctaccca gcagcgggaa gagttgttct cccgaacgca</td><td> 660</td>

gggcacgcac ccgggggtgt gcatgatca-t-gtccg ctgcc tcatacacaa tgcttgagtt 720 ggagcagtcg ttcgtgacat ggtacatccc ggacacgttg cgcacctcat acctcttttc 780 caacgggtgt gtagttccat tctccaccgc tagggctgcg ctgggctcca ttggcgaggt 840 tttcaaggcc gctaggatgc gatccatgcg tccgtagcct tgcgtggagc gtgcgtgtgc 900 çtgcgggagt gcgcatággt aggctacggt gatgattgct agcatggcgg gaatagtttt 960 gctatacatg aattcccgat gaagcagaga gcgcaggagg cggtatttat 1020 agtgccattc

<td>ccctctctga</td><td>gagacccgga</td><td>tggtagtcga</td><td>gtgtatcgga.</td><td>gacagcttga '</td><td>tgtagactcc</td><td> 1080</td>

<td>gtgcctgccg</td><td>gctcctctta</td><td>ttggcggaca</td><td>ccagtgagac</td><td>accccggaac</td><td>ttgctgtttt</td><td> 1140</td>

<td>tctgcaaaat,</td><td>ccggggtgac</td><td>cagtgggagc</td><td>ctatttgcac</td><td>acacgagcgg</td><td>gacaccccac</td><td> 1200</td>

<td>tctggtgaag</td><td>agtgccaaag</td><td>tcattctttt</td><td>tcccgttgcg</td><td>gggcagccga</td><td>ttgcatgttt</td><td> 1260</td>

<td>taggaaaata</td><td>ttacctttgc</td><td>tacaccctgt</td><td>cagatttacc</td><td>ctccacacat</td><td>atatattccg</td><td> 1320</td>

<td>tcacctccag</td><td>ggactattat</td><td>tcgtcgttgc</td><td>gccgccagcg</td><td>gaagatatcc</td><td>agaagctgtt</td><td> 1380</td>

<td>ttccgagaga</td><td>ctcggttggc</td><td>gcctggtata</td><td>tttgatggat</td><td>gtcgcgctgc</td><td>Ctcacgtccc</td><td> 1440</td>

<td>ggtacccagg</td><td>aacgcggtgg</td><td>gatctcgggc</td><td>ccatcgaaga</td><td>ctgtgctcca</td><td>gactgctcgc</td><td> 1500</td>

<td>ccagcaggtg</td><td>tttcttgatc</td><td>gccgcctcta</td><td>aattgtccgc</td><td>gcatcgccgg</td><td>taacattttt</td><td> 1560</td>

<td>ccagctcgga</td><td>gtttgcgttt</td><td>agatacagtt</td><td>tctgcgatgc</td><td>caaaggagcc</td><td>tgcagattat</td><td> 1620</td>

<td>aacctcggat</td><td>gctgtcattc</td><td>agcgctttta</td><td colspan="2">atttgacctc -cagatagttg</td><td>ctgtatttct</td><td> 1680</td>

<td>gttcccattg</td><td>gctgctgcgc '</td><td>agcttcgtat</td><td>aactcgagtt</td><td>attgttgcgc</td><td>tctgcctcgg</td><td> 1740</td>

<td>cgtactggct</td><td>catgatctgg</td><td>atcttgtccg</td><td>tgtcgctttt</td><td>cttcgagtgt</td><td>ttctcgcaaa</td><td> 1800</td>

<td>cgatgtgcac</td><td colspan="2">ggcctgcagt · gtccaatcgg</td><td>agtcgagctg</td><td>gcgccgaaac</td><td>tggcggatct</td><td> 1860</td>

<td>gagcctccac</td><td>actgccctgt</td><td>ttctctatcc</td><td>acggcggaac</td><td>cgcctoctgc</td><td>cgtttcagaa</td><td> 1920</td>

<td>tgttgttcaa</td><td>gtggtactct</td><td>gtgcggtcaa</td><td>tgaaggcgtt</td><td>attgccggtg</td><td>aaatctttgg</td><td> 1980</td>

<td>gaagcggttt</td><td>tcetcgggga</td><td>agattacgaa</td><td>attccecgcg</td><td>tcgttgcgct</td><td>tcctggatct</td><td> 2040</td>

<td>cgaggagatc</td><td>gttctccgcg</td><td>tcgaggagat</td><td>cgttctccgc</td><td>gtcgacacca</td><td>ttccttgcgg</td><td> 2100</td>

<td>cggcggtgct</td><td>caacggcctc</td><td>aacctactac</td><td>tgggctgctt</td><td>cctaafcgcag</td><td>gagfccgcata</td><td> 2160</td>

<td>agggagagcg</td><td>tcgacaaacc</td><td>cgcgtttgag</td><td>aacttgctca</td><td>agcttctggt</td><td>aaacgttgta</td><td> 2220</td>

<td>gtactctgaa</td><td>acaaggccct</td><td>agcactctga</td><td>tctgtttctc</td><td>ttgggtagcg</td><td>gtgagtggtt</td><td> 2280</td>

<td>tattggagtt</td><td>cactggtttc</td><td>agcacatctg</td><td>tcatctagac</td><td>aatattgtta</td><td>ctaaattttt</td><td> 2340</td>

<td>ttgaactaca</td><td>attgttcgta</td><td>attcatctat</td><td>tattatacat</td><td>cctcgtcagc</td><td>aatttctggc</td><td> 2400</td>

agacggagtt tactaacgtc ttgagtatga ggccgagaat ccagctctgt ggccatactc 2460 agtcttgaca gcctgctgat gtggctgcgrt tcaacgcaat aagcgtgtcc tccgactccg 2520

<td>agttgtgctc</td><td>gttatcgtcg ttctcatcct cggaaaaatc acacgaaaga acatactcac</td><td> 2580</td>

<td>cagtaggctt</td><td>tctggtccct ggggcacggc tgtttctgac gtattceggc gttgataata</td><td> 2640</td>

<td>gctcgaaagt</td><td>gaacgccgag tcgcgggagt cgaccgatgc ccttgagagc cttcaaccca</td><td> 2700</td>

<td>gtcagctcct</td><td>tccggtgggc gcggggcatg actatcgtcg ccgcacttat gactgtcttc</td><td> 2760</td>

<td>tttatcatgc</td><td>aactcgtagg acaggtgccg gcagcgctct gggtcatttt cggcgaggac</td><td> 2820</td>

<td>cgctttcgct</td><td>agagcgcgac gatgatcggc ctgtcgcttg cggtattcgg aatcttgcac</td><td> 2880</td>

<td>gccctcgctc</td><td>aagccttcgt cactggtccc gccaccaaac gtttcggcga gaagcaggcc</td><td> 2940</td>

<td>attatcgccg</td><td>gcatggcggc ogacgcgctg ggctacgtct tgctggcgtt cgcgacgcga</td><td> 3000</td>

<td>ggctggatgg</td><td>ecttccccat tatgattctt ctcgcttccg gcggcatcgg gatgcccgcg</td><td> 3060</td>

<td>ttgcaggcca</td><td>tgctgtccag gcaggtagat gaogaccatc agggacagct tcaaggatcg</td><td> 3120</td>

<td>ctcgcggctc</td><td>ttaccagcct áacttcgatc actggaccgc tgatcgtcac ggcgatttat</td><td> 3180</td>

<td>gccgcctcgg</td><td>cgagcacatg gaacgggttg gcatggattg taggcgccgc cctatacctt</td><td> 3240</td>

<td>gtctgcctcc</td><td>ccgcgttgcg tcgcggtgca tggagccggg c-cacctcgac ctgaatggaa</td><td> 3300</td>

<td>gccggcggca</td><td>cctcgctaac ggattcacca ctccaagaat tggagccaat caattcttgc</td><td> 3360</td>

<td>ggagaactgt</td><td>gaatgcgcaa accaaccctt ggcagaacat atccatcgcg tccgccatct</td><td> 3420</td>

<td>ccagcagccg</td><td>cacgcggcgc atcggggggg 9999999999 9999999 ^33 acaattcatc</td><td> 3480</td>

<td>attttttttt</td><td>tattcttttt tttgattteg gtttctttga aatttttttg attcggtaat</td><td> 3540</td>

<td>ctccgaacag</td><td>aaggaagaac gaaggaagga gcacagactt agattggtat atatacgcat</td><td> 3600</td>

<td>atgtagtgtt</td><td>gaagaaacat gaaattgccc agtattctta acccaactgc acagaacaaa</td><td> 3660</td>

<td>aacctgcagg</td><td>aaaogaagat * aaatcatgtc gaaagctaca tataaggaac gtgctgctac</td><td> 3720</td>

<td>tcatcctagt</td><td>cctgttgctg ccaagctatt taatatcatg cacgaaaagc aaacaaactt</td><td> 3 780</td>

gtgtgcttca ttggatgttc gtaccaccaa ggaattactg gagttagttg aagca-ttagg 3840 tcccaaaatt tgtttactaa aaacacatgt ggatatcttg actgattttt ccatggaggg 3900 cacagttaag ccgctaaagg cattateegc caagtacaat tttttactct tcgaagacag 3960 aaaatttgct gacattggta atacagtcaa attgeagtac tctgcgggtg tatacagaat 4020 agcagaatgg gcagacatta cgaatgcaca cggtgtggtg ggoocaggta ttgttagcgg 4080 tttgaagcag gcggcagaag aagtaa-RACH ggaacctaga ggccttttga tgttagcaga 4140 attgtcatgc aagggctccc tatctactgg agaatatact aagggtactg ttgacattgc 4,200 gaagagcgac aaagattttg ttatcggctt tafctgctcaa agagacatgg gtggaagaga 4260 tgaaggttac gattggttça ttatgacacc cggtgtgggt ttagatgaca agggagacge 4320 attgggtcaa cagtggggga cagtggggga acaggatctg acattattat 4380 tgttggaaga ggactatttg. caaagggaag ggatgctaag gtagagggtg aa-4440 cgttacag aaaagcaggc tgggaagcat atttgagaag atgcggocag caaaactaaa aaactgtatt 4500 ataagtaaat gcatgtatac taaactcaca aattagagct tcaatttaat tatatcagtt 4560 attacccggg aatctcggtc gtaatgattt ttataatgac gaaaaaaaaa aaattggaaa 4620 cccccccccc cccccccccc cccccccccc gaaaagcccc gcagcgttgg 4680 gtcctggcca

<td>cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt</td><td> 4740</td>

<td>actggttagc agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa</td><td> 4800</td>

<td>acgtctgcga · cctgagcaaç aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg</td><td> 4860</td>

<td>gaaacgcgga agtcagcgcc ctgcaccatt étgttccgga tctgcatcgc aggatgctgc</td><td> 4920</td>

<td>tggctaccct gtggaacacc tacatctgta ttaacgaagc gctggcattg accctgagtg</td><td> 4980</td>

<td>atttttctct ggtcccgccg catccatacc gccagttgtt taccctcaca acgttccagt</td><td> 5040</td>

<td>aaccgggcat gttcatcatc agtaacccgt atcgtgagca tcctctctcg tttcatcggt</td><td> 5100</td>

<td>atcattaccc ccatgaacag aaattccccc ttacacggag gcatcaagtg accaaacagg</td><td> 5160</td>

<td>aaaaaaccgc ccttaacatg gcccgcttta tcagaagcÇa gacattaacg cttctggaga</td><td> 5220</td>

<td>aactcaàcga gctggacgcg gatgaacagg cagacatçtg tgaatcgctt cacgaccacg</td><td> 5280</td>

<td>ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga tgaeggtgaa aacctctgac</td><td> 5340</td>

<td>acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag</td><td> 5400</td>

<td>cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac</td><td> 5460</td>

<td>gtaacgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag</td><td> 5520</td>

<td>agtgcaccat atgcggtgtg aaataccgca cagatgcgtâ aggagaaaat accgcatoag</td><td> 5580</td>

<td>gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc</td><td> 5640</td>

<td>ggtatcagct eactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg</td><td> 5700</td>

<td>aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct</td><td> 5760</td>

<td>ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca</td><td> 5820</td>

<td>gaggtggcga aacccgacag gactataaag Ataccaggcg tttccccctg gaagctccct</td><td> 5880</td>

<td>cgtgcgctct cctgttccga ccctgocgct taccggatac ctgtccgcct ttctcccttc</td><td> 5940</td>

<td>gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt</td><td> 6000</td>

<td>tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc</td><td> 6060</td>

<td>cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc</td><td> 6120</td>

<td>cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg</td><td> 6180</td>

gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc 6240 agttaccttc ggaaaaagag ttggtagctc ttgatccggç aaacaaacca ccgctggtag 63 00 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga 6360 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat 6420

<td>tttggtcatg agattatcaa aaaggat-ctt cacctagatc cttttaaatt aaaaatgaag</td><td> 6480</td>

<td>ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat</td><td> «540</td>

<td>cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc</td><td> 6600</td>

<td>cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat</td><td> 6660</td>

<td>accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag</td><td> 6720</td>

<td>ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg</td><td> 6780</td>

<td>• ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc</td><td> 6840</td>

<td>tgcaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca</td><td> 6900</td>

<td>acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg</td><td> 6960</td>

<td>tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc</td><td> 7020</td>

<td>actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta</td><td> 7080</td>

<td>ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc</td><td> 7140</td>

<td>aacacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg</td><td> 7200</td>

<td>ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaaçc</td><td> 7260</td>

<td>cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt cfcgggtgagç</td><td> 7320</td>

<td>aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat</td><td>73B0</td>

<td>actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag</td><td> 7440</td>

<td>cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttce</td><td>7E00</td>

<td>ccgaaaagtg ceacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa</td><td> 7560</td>

<td>taggcgtatc acgaggccct ttcgtcttca a</td><td> 7591</td>

<td><213> synthetic primer or probe</td><td></td>

<td>aggggtaagc ttggataaaa ggtatgaggt gcgcaacgtg tccgggatgt</td><td> 50</td>

<210> 15 <211> 42 <212> DMA <213> Synthesizer or probe <400> 15 agttacggat ccttaatggt gatggtggtg gtgecagttc afc 42

<td colspan="3"><212> 7648 <213> pFPMT-Mfalfa-El-H6 vector</td><td> . ·</td><td></td><td></td><td></td>

<td>ggtaccctgc</td><td>tcaatctccg</td><td>gaatçgtgat</td><td>etgatcgttc</td><td>ctgaaaacct 1</td><td>cgacattggc</td><td> 60</td>

<td>tccctcctgja</td><td>cacaggtact</td><td>cgtacaggtt</td><td>ccaggtaaac</td><td>gagtcgtagt</td><td>tgtcgatcat</td><td> 120</td>

<td>gacaacgttc</td><td>ttagaagcgg</td><td>ccggcatttt</td><td>gaaggtgact</td><td>aatagcctaa</td><td>gaaaatattt</td><td> 180</td>

<td>aatttaattt</td><td>tcattaatt</td><td>ttcctàtact</td><td>cgefcatttca</td><td>gcttttcatc</td><td>tcatcacttc</td><td> 240</td>

<td>ataaacgata</td><td>taaaccagaa</td><td>aaagaactat</td><td>tttcaaacac</td><td>gcttcteaaa</td><td>agcagtatgt</td><td> 300</td>

<td>ccttccacgt</td><td>ctccttagaa</td><td>tctggcaagt</td><td>ccgcgagggg</td><td>gatccttaat</td><td>ggtgatggtg</td><td> 360</td>

<td>gtggtgccag</td><td>ttcatcàtca</td><td>tatcccaagc</td><td>catacggtga</td><td>cctgttatgt</td><td>ggccgggata</td><td> 420</td>

<td>gattgagcaa</td><td>ttgcagtcet</td><td>gcaccgtctc</td><td>atgccggcga</td><td>ggcgagatgg</td><td>tgaacagctg</td><td> 480</td>

<td>ggagacgagg</td><td>aagacagatc</td><td>cgcagagatc</td><td>pcccacgtac</td><td>atagcggaac</td><td>agaaagcagc</td><td> 540</td>

<td>cgccccaacg</td><td>agcaaatcga</td><td>cgtggcgtcg</td><td>tattgtcgta</td><td>gtggggacgc</td><td>tggcgttcct</td><td> 600</td>

<td>agctgcgagc</td><td>gtgggggtga</td><td>g-cgctacçca</td><td>gcagcgggaa</td><td>gagttgttct</td><td>cccgaacgca</td><td> £60</td>

<td>gggcacgcac</td><td>ccgggggtgt</td><td>gcatgatcat</td><td>gtccgctgcc</td><td>tcatacacaa</td><td>tgcttgagtt</td><td> 720</td>

<td>ggagcagtcg</td><td>ttcgtgacat</td><td>ggtacatccc</td><td>ggacacgttg</td><td>cgcacctcat</td><td>accttttatc</td><td> 780</td>

<td>caagcttacc</td><td>ccttcttctt</td><td>tagcagcaat</td><td>gctggcaata</td><td>gtagtattta</td><td>taaacaataa</td><td> 840</td>

<td>cccgttattt</td><td>gtgctgttgg</td><td>aaaatggcaa</td><td>aacagcaaca</td><td>tcgaaatccc</td><td>cttctaaatc</td><td> 900</td>

<td>tgagtaaccg</td><td>atgacagctt</td><td>cagccggaat</td><td>ttgtgccgtt</td><td>teatcttctg</td><td>ttgtagtgtt</td><td> 960</td>

<td>gactggagca</td><td>gctaatgcgg</td><td>aggatgctgc</td><td>gaataaaact</td><td>gcagtaaaaa</td><td>ttgaaggaaa</td><td> 1020</td>

<td>tctcatgaat</td><td>tcccgatgaa</td><td>gcagagagcg</td><td>caggaggcgg</td><td>tatttatagt</td><td>gccattcccc</td><td> 1080</td>

<td>tctctgagag</td><td>acccggatgg</td><td>tagtcgagtg</td><td>tatcggagac</td><td>agcttgatgt</td><td>agactccgtg</td><td> 1140</td>

<td>cctgccggct</td><td>cctcttattg</td><td>gcggacacca</td><td>gtgagacacc</td><td>ccggaacttg</td><td>ctgtttttct</td><td> 1200</td>

<td>gcaaaatccg</td><td>SgÇftgaccag</td><td>tgggagccta</td><td>tttgcacaca</td><td>cgagcgggac</td><td>accccactct</td><td> 1260</td>

<td>ggtgaagagt</td><td>gccaaagtca</td><td>ttctttttcc</td><td>cgttgcgggg</td><td>cagccgattg</td><td>catgttttag</td><td> 1320</td>

<td>gaaaatatta</td><td>• cctttgctac</td><td>accctgtcag</td><td>atttaccctc</td><td>cockatoo</td><td>tattccgtca</td><td> 1380</td>

<td>cctccaggga</td><td>ctattattcg</td><td>tcgttgcgcc</td><td>gccagcggaa</td><td>gatatccaga</td><td>agctgtttt-c</td><td> 1440</td>

cgagagactc ggttggcgcc tggtatattt gatggatgtc gcgctgeetc acgtcccggt 1500 acccaggaac gcggtgggat ctcgggccca tcgaagactg tgctccagac tgctcgccca 1560 gcaggtgttt cttgatcgcc gcctctaaat tgtccgcgca tcgccagtaa catttttcea 1620 gctcggactt tgcgtttaga tacagtttct gcgatgccaa aggagcctgc agattataac 1680 ctcggatgct gtcattcagc gcttttaatt tgacctccág atagttgctg tatttctgtt 1740 cccattggct gctgcgcagc ttcgtataac tcgagttatt gttgcgctct gcctcggcgt 1800 actggctcat gatctggatc ttgtccgtgt cgcttttctt cgagtgtttc 1860 tcgcaaacga tgtgcacggc ctgcagtgtc caatcggagt cgagctggcg ccgaaactgg cggatctgag 1920

<td>cçtccacaet gccctgtttc tctatccacg gcggaaccgc ctcctgccgt ttcagaatgt</td><td> 1980</td>

<td>tgttcaagtg gtactctgtg cggtcaatga aggcgttatt gccggtgaaa tctttgggaa</td><td> 2040</td>

<td>gcggttttcc tcggggaaga ttacgaaatt ccccgcgtcg ttgcgcttcc tggatctcga</td><td> 2100</td>

<td>ggagatcgtt ctccgcgtcg aggagatcgt tctccgcgtc gacaccattc cttacggcgg</td><td> 2160</td>

<td>cggtgctcaa cggcctcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg</td><td> 2220</td>

<td>gagagcgtcg acaaacccgc gtttgagaae ttgctcaagc ttctggtaaa cgttgtagta</td><td> 2280</td>

<td>ctctgaaaca aggccctagc actctgatct gtttctcttg ggtagcgçtg agtggtttat</td><td> 2340</td>

<td>tggagttcac tggttteagc acatctgtca tctagacaat attgttacta aatttttttg</td><td> 2400</td>

<td>aactacaatt gttcgtaatt catctattat tatacatcct cgtcagcaat ttctggcaga</td><td> 2460</td>

<td>cggagtttac taacgtcttg agtatgaggc cgagaatcca gctctgtggc catactcagt</td><td> 2520</td>

<td>cttgacagcc tgctgatgtg gctgcgttca acgcaataag cgtgtcctcc gactccgagt</td><td> 2580</td>

<td>tgtgctcgtt atcgtcgttc tcatcctcgg aaaaatcaca cgaaagaaca tactcaccag</td><td> 2640</td>

<td>taggctttct ggt-coctggg gcacggctgt ttctgacgta ttccggcgtt gataatagct</td><td> 2700</td>

<td>cgaaagtgaa cgccgagtcg cgggagtcga ccgatgocct tgagagcctt caacccagtc</td><td> 2760</td>

<td>agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt</td><td> 2820</td>

<td>atcatgcaac tcgtaggaca ggtgccggca gcgctctggg tcattttcgg çgaggacçgc</td><td> 2880</td>

<td>tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc</td><td> 2940</td>

<td>ctcgctcaag ccttcgtcac tggtcccg-cc accaaacgtt tcggcgagaa gcaggcçatt</td><td> 3000</td>

<td>atcgccggca tggcggccga cgcgctgggc tacgtcttgc tggcgttcgc gacgcgaggc</td><td> 3060</td>

<td>tggatggcct tccccattat gattcttctc gcttccggcg gcatcgggat gcccgcgttg</td><td> 3120</td>

<td>caggccatgc tgtccaggca ggtagatgac gaccatcagg gacagcttca aggatcgctc</td><td> 3180</td>

<td>gcggctctta ccagoctaac ttcgatcact ggaccgctga tcgtcacggc gatttatgcc</td><td> 3240</td>

<td>gcctcggcga gcacatggaa cgggttggca tggattgtag gcgccgccct Ataccttgtc</td><td> 3300</td>

<td>tgcctccccg</td><td>cgttgcgtcg</td><td>cggtgcatgg</td><td>agccgggcca</td><td>cctcgacctg j</td><td>aatggaagcc</td><td> 3360</td>

<td>ggcggcacçt</td><td>cgctaacgga</td><td>ttcaccaete</td><td>caagaatt ^ g</td><td>agccaatcaa</td><td>ttcttgcgga</td><td> 3420</td>

<td>gaactgtgaa</td><td>tgcgcaaacc</td><td>aacccttggc</td><td>agaacatatc</td><td>catcgcgtcc</td><td>gccatctcca</td><td> 3480</td>

<td>gcagccgcac</td><td>gcggcgcatc</td><td>gggggggggg</td><td>gggggggggg</td><td>ggggc & aaca</td><td>attcatcatt</td><td> 3540</td>

<td>ttttttttat</td><td>tctttttttt</td><td>gatttcggtt</td><td>tctttgaaat</td><td>ttttttgatt</td><td>cggtaatctc</td><td> 3600</td>

<td>cgaacagaag</td><td>gaagaacgaa</td><td>ggaaggagca</td><td>cagacttaga</td><td>ttggtatata</td><td>tacgcatatg</td><td> 3660</td>

<td>tagtgttgaa</td><td>gaaacatgaa</td><td>attgcccagt</td><td>attcttaacc</td><td>caactgcaca</td><td>gaacaaaaac</td><td> 3720</td>

<td>ctgcaggaaa</td><td>cgaagataaa</td><td>tcatgtcgaa</td><td>agctacatat</td><td>aaggaacgtg</td><td>ctgctactca</td><td> 3780</td>

<td>tcctagtcct</td><td>gttgctgcca</td><td>agctatttaa</td><td>tatcatgcac</td><td>gaaaagcaaa</td><td>caaacttgtg</td><td> 3840</td>

<td>tgcttcattg</td><td>gatgttcgta</td><td>ccaccaagga</td><td>attactggag</td><td colspan="2">ttagttgaag cattaggt.cc</td><td> 3900</td>

<td>caaaatttgt</td><td>ttactaaaaa</td><td>cacatgtgga</td><td>tatcttgact</td><td>gatttttcca</td><td>tggagggcac</td><td> 3960</td>

<td>agttaagccg</td><td>ctaaaggcat</td><td>tatccgccaa</td><td>gtacaatttt</td><td>ttactcttcg</td><td>aagacagaaa</td><td> 4020</td>

<td>atttgctgac</td><td>attggtaata</td><td>cagtcaaatt</td><td>gcagtactct</td><td>gcgggtgtat</td><td>acagaatagc</td><td> 4080</td>

<td>agaatgggca</td><td>gacattacga</td><td>atgcacacgg</td><td>tgtggtgggc</td><td>ccaggtattg</td><td>ttagcggttt</td><td> 4140</td>

<td>gaagcaggcg</td><td>gcagaagaag</td><td>taacaaagga</td><td>acctagaggc</td><td>cttttgatgt</td><td>tagcagaatt</td><td> 4200</td>

<td>gtcatgcaag</td><td>ggctccctat</td><td>ctactggaga</td><td>àtataçtaag</td><td>ggtactgttg</td><td>acattgcgaa</td><td> 4260</td>

<td>gagcga-caaa</td><td>gattttgtta</td><td>tcggctttat</td><td>tgctcaaaga</td><td>gacatgggtg</td><td>gaagagatga</td><td> 4320</td>

<td>aggttacgat</td><td>tggttgattà</td><td>tgacacccgg</td><td>tgtgggttta</td><td>gatgacaagg</td><td>gagacgcatt</td><td> 4380</td>

<td>gggtcaacag</td><td colspan="2">tatagaaccg tggatgatgt</td><td>ggtctctaca</td><td>ggatctgaca</td><td>ttattattgt</td><td> 4440</td>

tggaagagga ctatttgcaa agggaaggga tgctaaggta gagggtgaac gttacagaaa 4S00 agcaggctgg gaagcatatt tgagaagatg cggccagcaa aacta & yyyy ctgtattata 4560 agtaaatgca tgtatactaa actcacaaat tagagcttca atttaattat atcagttatt 4620 acccgggaat ctcggtcgta atgattttta taatgacgaa aaaaaaaaaa ttggaaagaa 4680 aagccccccc cccccccccc cccccccccc cccccccgca gcgttgggtc ctggccacgg 4740 gtgcgcatga tcgtgctcct gtcgttgagg acccggctag gctggcgggg ttgccttact 4800 ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg ctgcaaaacg 4860 tctgcgacct gagcaacaac atgaatggtc ttcggtttcc gtgtttcgta aagtctggaa 4920 acgcggaagt cagcgccetg caccattatg ttccggatct gcatcgcagg atgctgctgg 4980 ctaccctgtg gaacacctac atctgtatta acgaagcgct ggcattgacc ctgagtgatt 5040 tttctctggt cccgccgcat -ccataccgcc agttgtttac octcacaacg 5100 ttccagtaac

<td>cgggcatgtt</td><td>catcatcagt</td><td>aacccgtatc</td><td>gtgagcatcc</td><td>tctctcgttt</td><td>catcggtat-c</td><td> 5160</td>

<td>attaccccca</td><td>tgaacagaaa</td><td>ttccccctta</td><td>cacggaggca</td><td>tcaagtgacc</td><td>aaacaggaaa</td><td> 5220</td>

<td>aaaccgccct</td><td>taacatggcc</td><td>cgctttatca</td><td>gaagccagac</td><td>attaacgctt</td><td>ctggagaaac</td><td> 5260</td>

<td>tcaacgagct</td><td>ggacgcggat</td><td>gaacaggcag</td><td>acatctgtga</td><td>atcgcttcac</td><td>gaccacgctg</td><td> 5340</td>

<td>atgagcttta</td><td>ccgcagctgc</td><td>ctcgcgcgtt</td><td>tcggtgatga</td><td>cggtgaaaac</td><td>ctctgacaca</td><td> 5400</td>

<td>tgcagctccc</td><td>ggagacggtc</td><td>acagcttgtc</td><td>tgtaagçgga</td><td>tgccgggagc</td><td>agacaagccc</td><td> 5460</td>

<td>gtcagggcgc</td><td>gtcagcgggt</td><td>gttggcgggt</td><td>gtcggggcgc</td><td>agccatgacc</td><td>cagtcacgta</td><td> 5520</td>

<td>gcgatagcgg</td><td>agtgtatact</td><td>ggcttaacta</td><td>tgcggcatca</td><td>gagcagattg</td><td>tactgagagt</td><td> 5580</td>

<td>gcaccatatg</td><td>cggtgtgaaa</td><td>taccgcacag</td><td>atgcgtaagg</td><td>agaaaatacc</td><td>gcatcaggcg</td><td> 5640</td>

<td>ctcttccgct</td><td>tcctcgctca</td><td>ctgactcgct</td><td>gcgctcggtc</td><td>gttcggctgc</td><td>ggcgagcggt</td><td> 5700</td>

<td>atcagctcac</td><td>tcaaaggcgg</td><td>taatacggtt</td><td>atccacagaa</td><td>tcaggggata</td><td>acgcagga & a</td><td> 5760</td>

<td>gaacatgtga</td><td>gcaaaaggcc</td><td>agcaaaaggc</td><td>caggaaccgt</td><td>aaaaaggccg</td><td>cgttgctggc</td><td> 5820</td>

<td>gtttttccat</td><td>aggctccgcc</td><td>cccctgacga</td><td>gcatcacaaa</td><td>aatcgacgct</td><td>caagtcagag</td><td> 5880</td>

<td>gtggcgaaac</td><td>ccgacaggac</td><td>tataaagata</td><td>ccaggcgttt</td><td>ccccctggaa</td><td>gctccctcgt</td><td> 5940</td>

<td>gcgctctcct</td><td>gttccgacce</td><td>tgecgcttaC</td><td>cggatacctg</td><td>tccgcctttc</td><td>tcccttcggg</td><td> 6000</td>

<td>aagcgtggcg</td><td>ctttctcata</td><td>gctcacgctg</td><td>taggtatctc</td><td>agttcggtgt</td><td>aggtcgttcg</td><td> 6060</td>

<td>ctccaagctg</td><td>ggctgtgtgc</td><td>acgaaccccc</td><td>cgttcagccc</td><td>gaccgctgcg</td><td>ccttatccgg</td><td> €120</td>

<td>taactatcgt</td><td>cttgagtcca</td><td>acccggtaag</td><td>acacgactta</td><td>tcgccactgg</td><td>cagcagccac</td><td> €180</td>

<td>tggtaacagg</td><td>attagcagag</td><td>cgaggtatgt</td><td>aggcggtgct</td><td>acagagttct</td><td>tgaagtggtg</td><td> 6240</td>

<td>gcctaactac</td><td>ggctacacta</td><td>gaaggacagt</td><td>atttggtatc</td><td>tgcgctctgc</td><td>tgaagccagt</td><td> 6300</td>

taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg € 360 tggttttttt gtttg-caagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc € 420 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt € 4 80 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt € 54 0 taaatcaatc taaagtatat atgagtaaac ttggtetgac agttaccaat gcttaatcag € 600 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt 6660 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc 6720 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc € 780 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtçtatta attgttgccg 6840 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctgc € 900 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg 6960

<td>atcaaggcga</td><td>gttacatgat cccccatgtt</td><td>gtgcaaaaaa</td><td>gcggttagct</td><td>ccttcggtcc</td><td> 7020</td>

<td>tccgatcgtt</td><td>gtcagaàgta.agttggccgc</td><td>agtgttatcá</td><td>ctcatggtta</td><td>tggcagcact</td><td> 7000</td>

<td>gcataattct</td><td>cttactgtca tgccatccgt</td><td>aagatgcttt</td><td>tctgtgactg</td><td>gtgagtactc</td><td> 7140</td>

<td>aaccaagtca</td><td>ttctgagaat agtgtatgcg</td><td>gegaccgagt</td><td>tgctcttgcc</td><td>cggcgtcaac</td><td> 7200</td>

<td>acgggataat</td><td>accgcgccac atagcagaac</td><td>tttaaaagtg</td><td>ctcatcattg</td><td>gaaaacgttc</td><td> 7260</td>

<td>ttcggggcga</td><td>aaactctcaa ggatcttapc</td><td>gctgttgaga</td><td>tccagttcga</td><td>tgtaacccac</td><td> 7320</td>

<td>tcgtgcaccc</td><td>aactgatctt cagcatcttt</td><td>tactttcacc</td><td>agcgtttctg</td><td>ggtgagcaaa</td><td> 7380</td>

<td>aacaggaagg</td><td>caaaatgccg caaaaaaggg</td><td>aataagggcg</td><td>acacggaaat</td><td>gttgaatact</td><td> 7440</td>

<td>catactcttC</td><td>ctttttcaat attattgaag</td><td>catttatcag</td><td>ggttattgtc</td><td>tcatgagcgg</td><td> 7500</td>

<td>attackattt</td><td>gaatgtattt agaaaaataa</td><td>acaaataggg</td><td>gttccgcgca</td><td>catttccccg</td><td> 7560</td>

<td>aaaagtgcca gcgtatcacg</td><td>cctgacgtct aagaaaccat aggccctttc gtcttcaa</td><td>tatvatcatg</td><td>acattaacct</td><td>ataaaaatag</td><td> 7620 7648</td>

<td> <213></td><td>vector pUC18-FMD-MFalf a-El- '</td>

<td> <221></td><td>hodgepodge</td>

<td> <223></td><td>N is any nucleotide</td>

<td> <22 0> <221> <222> <22 3></td><td>MISCELLANEOUS (1386) .. (1387) N is any nucleotide</td>

<td>gcgcccaata</td><td>cgcaaaccgc</td><td>ctctcccogc</td><td>gcgttggccg</td><td>attcattaat</td><td>gcagctggca</td><td> 60</td>

<td>cgacaggttt</td><td>cccgactgga</td><td>aagcgggcag</td><td>tgagcgcaac</td><td>gcaattaatg</td><td>tgagttagct</td><td> 120</td>

<td>cactcattag</td><td>gcaccccagg</td><td>ctttacactt</td><td>tatgcttccg</td><td>gctcgtatgt</td><td>tgtgtggaat</td><td> 180</td>

<td>tgtgagcgga</td><td>taacaatttc</td><td>acacaggaaa</td><td>cagctatgac</td><td>catgattacg</td><td>aattcgagct</td><td> 240</td>

<td>cggtacccgg</td><td>ggatccttaa</td><td>tggtgatggt</td><td>ggtggtgcca</td><td>gttcafccatc</td><td>atatcccaag</td><td> 300</td>

<td>ccatacggtg</td><td>acctgttatg</td><td>tggccgggat</td><td>agattgagca</td><td>attgcagtcc</td><td>tgcaccgtct</td><td> 360</td>

<td>catgccggcg</td><td>aggcgagatg</td><td>gtgaacagct</td><td>gggagacgag</td><td>gaagacagat</td><td>ccgcagagat</td><td> 420</td>

<td colspan="2">cccccacgta catagcggaa</td><td>cagaaagcag</td><td>ccgccocaac</td><td>gagcaaatcg</td><td>acgtggcgtc</td><td> 480</td>

<td>gtattgtcgt</td><td>agtggggacg</td><td>ctggcgttcc</td><td>tagctgcgag</td><td>cgtgggggtg</td><td>agcgctaccc</td><td> 540</td>

agcagcggga agagttgttc tcccgaacgc agggcacgca cccgggggtg tgcatgatca 600 tgtcçgctgc ctcatacaca atgcttgagt tggagcagtc gttcgtgaca tggtacatcc € 60 cggacacgtt gcgcacctca taccttttat ccaagcttac cccttcttct ttaçrcagcaa 720 tgctggcaat agtagtattt ataaacaata acccgttatt tgtgetgttg gaaaatggca 760 aaacãgcaac atcgaaatcc ccttctaaat ctgagtaacc gatgacagct tcagccggaa 840 tttgtgccgt ttcatcttct gttgtagtgt tgactggagc agctaatgcg gaggatgctg 900 cgaataaaac tgcagtaaaa attgaaggaa atctcatgaa ttcccgatga aggcagagag 960 cgcaaggagg cggtatttat agtgccattc ccctctctga gagacccgga tggtagtcga 1020

<td>gtgttatcgg</td><td>agacagcttg</td><td>atgtagactc</td><td>cgtgcctgcc</td><td>ggtcctctta</td><td>ttggcggaca</td><td> 1080</td>

<td>ccagtgagac</td><td>accccggaac</td><td>ttgctgtttt</td><td>tctgcaaaat</td><td>ccggggtgac</td><td>cagtgggagc</td><td> 1140</td>

<td>ctatttgcac</td><td>acacgagcgg</td><td>gacaccccac</td><td>tctggtgaag</td><td>agtgccaaag</td><td>tcattctttt</td><td> 1200</td>

<td>tcccgtnncg</td><td>gggcagccga</td><td>ttgcatgttt</td><td>taggaaaata</td><td>ttacctttgc</td><td>tacaccctgt</td><td> 1260</td>

<td>cagatttacc</td><td>ctccacacat</td><td>atatattccg</td><td>tcacctccag</td><td>ggactattct</td><td>tggctcgttg</td><td> 1320</td>

<td>cgccgccgcg</td><td>gaagatatçç</td><td>agaagctgtg</td><td>ttttccgaga</td><td>gactcggttg</td><td>gcgcctggta</td><td> 1380</td>

<td>tatttnnagg</td><td>atgtcgcgct</td><td>gcctcacgtc</td><td>ccggtaccca</td><td>ggaacgcggt</td><td>gggatctcgg</td><td> 1440</td>

<td>gcccatcgaa</td><td>gactgtgctc</td><td>cagactgctc</td><td>gcccagcagg</td><td>tgtttcttga</td><td>ttgccgcctc</td><td> 1500</td>

<td>taaatagtcc</td><td>gcgcatcgcc</td><td>ggtaacattt</td><td>ttccagctcg</td><td>gagtttgcgt</td><td>ttagatacat</td><td>1H60</td>

<td>ttctgcgatg</td><td>ccaaaggagc</td><td>ctgcagatta</td><td>taacctcgga</td><td>tgctgtcatt</td><td>Cagcgctttt</td><td> 1620</td>

<td>aatttgacct</td><td>ccagatagtt</td><td>gctgtatttc</td><td>tgttccattg</td><td>gctgctggac</td><td>grttcgtataa</td><td> 1660</td>

<td>ctcgagttat</td><td>tgttgcgctc</td><td>tgcctcggcg</td><td>tactggctca</td><td>tgactgactg</td><td>cggtcgcttc</td><td> 1740</td>

<td colspan="2">tcgagtgttc tcgcaacagg</td><td>acgcctgcag</td><td>gtcatcgagt</td><td>cgagctggcg</td><td>ccgaaactgg</td><td> 1800</td>

<td>cggatctgac</td><td>ctccacactg</td><td>ccctgtatct</td><td>ctatccaccg</td><td>ggaaccgcct</td><td>cctgccgttc</td><td> 1860</td>

<td>cagaatgttg</td><td>ttcaagtggt</td><td>agctctgtgc</td><td>ggtcaatgaa</td><td>ggcgttattg</td><td>ccggtgaaat</td><td> 1920</td>

<td>ctttgggaag</td><td>cggtttatcc</td><td>teggggaaga</td><td>ttacgaaatt</td><td>cccgcgcgtc</td><td>gttgcgcttc</td><td> 1980</td>

<td>ctggatctcg</td><td>aggaagatcg</td><td>ttctccgcgt</td><td>cgaggagatc</td><td>gttctccgcg</td><td>tcgacctgca</td><td> 2040</td>

<td colspan="2">ggcatgcaag cttggcactg</td><td>gccgtcgttt</td><td>tacaacgtcg</td><td>tgactgggaa</td><td>aaccctggcg</td><td> 2100</td>

<td>ttacccaact</td><td>taatcgcctt</td><td>gcagcacatc</td><td>cccctttcge</td><td>cagctggcgt</td><td>aatagcgaag</td><td> 2160</td>

<td>aggcccgcac</td><td>cgatcgccct</td><td>tcccaacagt</td><td>tgcgcagcct</td><td>gaatggcgaa</td><td>tggcgcctga</td><td> 2220</td>

<td>tgcggtattt</td><td>tctccttacg</td><td>catctgtgog</td><td>gtafcttcaca</td><td>oegcatatgg</td><td>tgcactctca</td><td> 2280</td>

<td>gtacaatctg</td><td>ctctgatgcc</td><td>gcatagttaa</td><td>gccag-ccccg</td><td>acacccgcca</td><td>acacccgctg</td><td> 2340</td>

<td>acgcgccctg</td><td>acgggcttgt</td><td>ctgctcccgg</td><td>catccgctta</td><td>cagácaagct 1</td><td>gtgaccgtct</td><td> 2400</td>

<td>ccgggagctg</td><td>çatgtgtcag.</td><td>aggttttcac</td><td>Cgtcatcacc ·</td><td>gaaacgcgcg</td><td>agacgaaagg</td><td> 2460</td>

<td>gcctcgtgat</td><td>acgcctatt.t</td><td>ttataggtta</td><td>atgtcatgat</td><td>aataatggtt</td><td>tettagacgt</td><td> 2520</td>

<td>caggtggcac</td><td>tttfccgggga</td><td>aatgtgcgcg</td><td>gaacccctat</td><td>ttgtttattt</td><td>ttctaaatac</td><td> 2580</td>

<td>attcaaatat</td><td>gtatcegctc</td><td>atgagacaat</td><td>aaccctgata</td><td>aatgcttcaa</td><td>taatatfcgaa</td><td> 2640</td>

<td>aaaggaagag</td><td>tatgagtatt</td><td>caacatttcc</td><td>gtgtcgccct</td><td>tattcccttt</td><td>tttgcggcat</td><td> 2700</td>

<td>tttgccttcc</td><td>tgtttttgct</td><td>cacccagaaa</td><td>cgctggtgaa</td><td>agtaa & agat</td><td>gctgaagatc</td><td> 2760</td>

<td>agttgggtgc</td><td>acgagtgggt</td><td>tacatcgaac</td><td>tggatctcaa</td><td>cagcggtaag</td><td>atccttgaga</td><td> 2820</td>

<td>gttttcgccc</td><td>cgaagaacgt</td><td>tttccaatga</td><td>tgágcacttt</td><td>taaagttctg</td><td>ctatgtggcg</td><td> 2880</td>

<td>cggtattatc</td><td>ccgtattgac</td><td>gccgggcaag</td><td>agcaactcgg</td><td>tcgccgcata</td><td>cactattctc</td><td> 2940</td>

<td>agaatgaçtt</td><td>ggttgagtac</td><td>tcaccagtca</td><td>cagaaaagca</td><td>tcttacggat</td><td>ggcatgacag</td><td> 3000</td>

<td>taagagaatt</td><td>atgcagtgct</td><td>gccataacca</td><td>tgagtgataa</td><td>cactgcggcc</td><td>aacttacttc</td><td> 3060</td>

<td>tgacaacgat</td><td>cggaggaccg</td><td>aaggagctaa</td><td>ccgctttttt</td><td>gcacaacatg</td><td>ggggatcatg</td><td> 3120</td>

<td>taactcgcct</td><td>tgatcgttgg</td><td>gaaccggagc</td><td>tgaatgaagc</td><td>cataccaaac</td><td>gacgagcgtg</td><td> 3180</td>

<td>acaccacgat</td><td>gcctgtagca</td><td>atggcaacaa</td><td>cgttgcgcaa</td><td>actattaact</td><td>ggcgaactac</td><td> 3240</td>

<td>ttactctagc</td><td>ttcccggcaa</td><td>caattaatag</td><td>acidggatgga</td><td>ggcggataaa</td><td>gttgcaggac</td><td> 3300</td>

<td>cacttctgcg</td><td>ctcggccctt</td><td>ccggctgget</td><td>ggtttattgc</td><td>tgataaatct</td><td>ggagccggtg</td><td> 3360</td>

<td>agcgtgggtc</td><td>tcgcggtatç</td><td>attgcagcac</td><td>tggggccaga</td><td>tggtaagccc</td><td>tcccgtatcg</td><td> 3420</td>

<td>tagttatcta</td><td>cacgacgggg</td><td>agtcaggcaa</td><td>ctatggatga</td><td>acgaaataga</td><td>cagatcgctg</td><td> 3480</td>

<td>agataggtgc</td><td>ctcactgatt</td><td>aagcattggt</td><td>aactgtcaga</td><td>ccaagtttac</td><td>: tcatatatae</td><td> 3540</td>

tttagattga tttaaaactt -catttttaat ttaaaaggat ctaggtgaag atcctttttg 3600 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagçg tcagaccccg 3660 fcagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc 3720 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc 3780 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt 3B40 agccgtagtt aggccaccac fctcaagaact ctgtagcacc gcctacatac ctcgctctgc 3900 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact 3960 caagacgata gttaccggat aaggegcagc ggtcgggctg aaeggggggt tcgtgcacac 4020 agcccagctt ggagcgaacg acctaeacog aactgagata cctacagcgt gagctatgag 4080 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg 4140 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtoctg 4200 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga 4260 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt 4320 ttgctcacat gttctttcct gogttatccc ctgattctgt ggataaccgt attaccgcct 4380 ttgagtgagc tgataocgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg 4440 aggaagcgga aga 4453 <210> 18 <211> 51 <212> DMA <213> primer or synthetic probe <400> 18 gt > DNA <213> primer or synthetic probe <400> 19 tagtactagt attagtaggc ttcgcatgaa ttcccgatga aggcsgagag cg 52 <210> 20 <211> 4252 <212> DNA <213> vector pUC18-FMD-CL-El-H6 <220>

<221> MISCELLANEOUS <222> (1006) .. (1007) <223> N is any nucleotide <220>

<221> MISCELLANEOUS <222> (1185) .. (1186) <223> N is any nucleotide <400> 20

<td>gcgcccaata</td><td>cgcaaaccgc</td><td>ctctccccgc</td><td>gcgttggccg</td><td>attcattaat</td><td>gcagctggca</td><td> 60</td>

<td>cgacaggttt</td><td>ccogactgga</td><td>aagcgggcag</td><td>tgagcgcaac</td><td>gcaattaatg</td><td>tgagttagct</td><td> 120</td>

<td>cactcattag</td><td>gcaccccagg</td><td>ctttacactt</td><td>tatgcttccg</td><td>gctcgtatgt</td><td>tgtgtggaat</td><td> 180</td>

<td>tgtgagcgga</td><td>taacaatttc</td><td>acacaggaaa</td><td>cagctatgac</td><td>catgattacg</td><td>aattcgagct</td><td> 240</td>

<td>cggtacccgg</td><td>ggatccttaa</td><td>tggtgatggt</td><td>ggtggtgcca</td><td>gttcatcatc</td><td>atatcccaag</td><td> 300</td>

<td>ccatacggtg</td><td>acctgttatg</td><td>tggccgggat</td><td>agattgagca</td><td>attgcagtcc</td><td>tgcaccgtct</td><td> 360</td>

<td>catgccggcg</td><td>aggcgagatg</td><td>gtgaacagct</td><td>gggagacgag</td><td>gaagacagat</td><td>ocgcagagat</td><td> 420</td>

<td>cccccacgta</td><td>catagcggaa</td><td>cagaaagcag</td><td>ccgccccaac</td><td>gagcaaatcg</td><td>acgtggcgtc</td><td> 480</td>

<td>gtattgtcgt</td><td>agtggggacg</td><td>ctggcgttcc</td><td>tagctgcgag</td><td colspan="2">cgtaggggtg agcgctaccc</td><td> 540</td>

<td>agcagcgggâ</td><td>agagttgttc</td><td>tcccgaacgc</td><td>agggcacgca</td><td colspan="2">cccgggggtg tgcatgatca</td><td> 600</td>

<td>tgtccgctgc</td><td>ctcatacaca</td><td>atgcttgagt</td><td>tggagcagtc</td><td>gttcgtgaca</td><td>tggtacatcc</td><td> 660</td>

<td>cggacacgtt</td><td>gcgcacctca</td><td>tatcctagtg</td><td>ctgctagtgg</td><td>taggaagcat</td><td>agtactagta</td><td> 720</td>

<td>ttagtaggct</td><td>tcgcatgaat</td><td>tcccgatgaa</td><td>ggcagagagc</td><td>gcaaggaggc</td><td>ggtatttata</td><td> 780</td>

<td>gtgccattcc</td><td>cctctctgag</td><td>agacccggat</td><td>ggtagtcgag</td><td>tgttatcgga</td><td>gacagcttga</td><td> 840</td>

<td>tgtagactcc</td><td>gtgcctgccg</td><td>gtcctcttat</td><td>tggcggacac</td><td>cagtgagaca</td><td>ccccggaact</td><td> 900</td>

<td>tgctgttttt</td><td>ctgcaaaatc</td><td>cggggtgacc</td><td>agtgggagcc</td><td>tatttgcaca</td><td>cacgagcggg</td><td> 960</td>

<td>acaccccact</td><td>ctggtgaaga</td><td>gtgccaaagt</td><td>cattcttttt</td><td>cccgtnncgg</td><td>ggcagccgat</td><td> 1020</td>

<td>tgcatgtttt</td><td>aggaaaatat</td><td>tacctttgct</td><td>acaccctgtc</td><td>agatttaccc</td><td>tccacacata</td><td> 1080</td>

<td>tatattccgt</td><td>cacctccagg</td><td>gactattctt</td><td>ggctcgttgc</td><td>gccgccgcgg</td><td>aagatatcca</td><td> 1140</td>

<td>gaagctgtgt</td><td>tttccgagag</td><td>actcggttgg</td><td>cgcctggtat</td><td>atttxmagga</td><td>tgtcgcgctg</td><td> 1200</td>

<td>cctcacgtcc</td><td>cggtacccag</td><td>gaacgcggtg</td><td>ggatctcggg</td><td>cccatcgaag</td><td>actgtgctcc</td><td> 1260</td>

agactgctcg cccagcaggt gtttcttgat tgccgcctct aaatagtccg cgcatcgccg 1320 gtaacatttt tccagctcgg agtttgcgtt tagatacatt tctgcgatgc caaayyagcc 1380 tgcagattat aacctcggat gctgtcattc agcgctttta atttgacctc cagatagttg 1440 ctgtatttct gttccattgg ctgctggacg ttcgtataac tcgagttatt gttgcgctct 15 OO gcctcggcgt actggctcat gactgactgc ggtcgcttct cgagtgttct cgcaacagga 1560 cgcctgcagg tcatcgagtc gagctggcgc cgaaactggc ggatctgacc tccacactgc 1620 cctgtatctc tatccaccgg gaaccgcctc ctgecgttcc agaatgttgt tcaagtggta 1680 gctctgtgcg gtcaatgaag gcgttattgc cggrtgaaatc tttgggaagc ggtttatcct 1740 cggggaagat tacgaaattc ccgcgcgtcg ttgcgcttcc tggatctcga ggaagatcgt 1800 tctccgcgtc gaggagatcg ttctccgcgt cgacctgcag gcatgcaagc ttggcactgg 1860 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacocaactt aatcgccttg 1920 cagcacatcc ccctttcgcc agctgg-cgtA atagcgaaga ggcccgcacc gatcgccctt 1980 eccaacagtt gcgcagcctg aatggcgaat ggcgectgat gcggtatttt ctccttacgc 2040 atctgtgcgg tatttcacac cgcatatggt g-cactctcag tacaatctgc tctgatgccg 2100 catagttaag ccagccccga cacccgccaa eaeccgctga cgcgccctga cgggcttgtc 2160 tgctcccggc atccgcttac agacaagctg tgaccgtctc egggagcfcgc atgtgtcaga 2220

<td>ggttttcacc</td><td>gtcatcaccg</td><td>aaacgcgcga</td><td>gacgaaaggg</td><td>cctcgtgata</td><td>cgcctatttt</td><td> 2280</td>

<td>tataggttaa</td><td>tgtcatgata</td><td>ataatggttt</td><td>cttagacgtc</td><td>aggtggcact</td><td>tttcggggaa</td><td> 2340</td>

<td>atgtgcgcgg</td><td>aacccctatt</td><td>tgtttatttt</td><td>tctaaataca</td><td>ttcaaatatg</td><td>tatccgctca</td><td> 2400</td>

<td>tgagacaata</td><td>accctgataa</td><td>atgcttcaat</td><td>aatattgaaa</td><td>aaggaagagt</td><td>atgagtattc</td><td> 2460</td>

<td>aacatttccg</td><td>tgtcgccctt</td><td>attccctttt</td><td>ttgcggcatt</td><td>ttgccttcct</td><td>gtttttgctc</td><td> 2520</td>

<td>acccagaaac</td><td>gctggtgaaa</td><td>gtaaaagatg</td><td>ctgaagatca</td><td colspan="2">gttgggtgca cgagtgggtt</td><td> 2580</td>

<td>acatcgaact</td><td>ggatctcaac</td><td>agcggtaaga</td><td>tccttgagag</td><td>ttttcgcece</td><td>gaag & acgtt</td><td> 2640</td>

<td>ttccaatgat</td><td>gagcactttt</td><td>aaagttctgc</td><td>tatgtggcgc</td><td>ggtattatcc</td><td>cgtattgacg</td><td> 2700</td>

<td>ccgggcaaga</td><td>gcaactcggt</td><td>cgccgcatac</td><td>actattctca</td><td>gaatgacttg</td><td>gttgagtact</td><td> 2760</td>

<td>caccagtcac</td><td>agaaaagcat</td><td>cttacggatg</td><td>gcatgacagt</td><td>aagagaatta</td><td>tgcagtgctg</td><td> 2820</td>

<td>ccataaccat</td><td>gagtgataac</td><td>actgcggcca</td><td>acttacttct</td><td>gacaacgatc</td><td>ggaggaccga</td><td> 2880</td>

<td>aggagctaac</td><td>cgcttttttg</td><td>cacaacatgg</td><td>gggateatgt</td><td>aactcgcctt</td><td>gatcgttggg</td><td> 2540</td>

<td>aaccggagct</td><td>gaatgaagcc</td><td>attackcaaacg</td><td>acgagcgtga</td><td>caccacgatg</td><td>cctgtagcaa</td><td> 3000</td>

<td>tggcaacaac</td><td>gttgcgcaaa</td><td>ctattaactg</td><td>gcgaactact</td><td>tactctagct</td><td>tcccggcaac</td><td> 3060</td>

<td>aattaataga</td><td>ctggatggag</td><td>gcggataaag</td><td>ttgcaggacc</td><td>acttctgcgc</td><td>tcggcccttc</td><td> 3120</td>

<td>cggctggctg</td><td>gtttattgct</td><td colspan="2">gataaatctg gagccggtga</td><td>gçgtgggtct</td><td>cgcggtat ^ A</td><td> 3180</td>

<td>ttgcagcact</td><td>ggggccagat</td><td>ggtaagccct</td><td>cccgtatcgt</td><td>agttatctac</td><td>acgacgggga</td><td> 3240</td>

<td>gtcaggcaac</td><td>tatggatgaa</td><td>cgaaatagac</td><td>agatcgctga</td><td>gataggtgcc</td><td>tcactgatta</td><td> 3300</td>

<td>agcattggta</td><td>actgtcagac</td><td>caagtttact</td><td>catatatact</td><td>ttagattgat</td><td>ttaaaacttc</td><td> 3360</td>

<td>atttttaatt</td><td>taaaaggatc</td><td>taggtgaaga</td><td>tcctttttga</td><td>taatctcatg</td><td>accaaaatcc</td><td> 3420</td>

<td>cttaacgtga</td><td>gttttcgttc</td><td>cactgagcgt</td><td>cagaccccgt</td><td>agaaaagatc</td><td>aaaggatctt</td><td> 3480</td>

<td>cttgagatcc</td><td>tttttttctg</td><td>cgcgtaatct</td><td>gctgcttgca</td><td>aacaaaaaaa</td><td>ccaccgctac</td><td> 3540</td>

<td>cagcggtggt</td><td>ttgtttgccg</td><td>gatcaagagc</td><td>taccaactct</td><td>ttttcogaag</td><td>gtaactggct</td><td> 3600</td>

<td>tcagcagagc</td><td>gcagatacca</td><td>aatactgtcc</td><td>ttctagtgta</td><td>gccgtagtta</td><td>ggccaccact</td><td> 3660</td>

<td>tcaagaactc</td><td>tgtagcaccg</td><td>cctacatacc</td><td>tcgctctgct</td><td>aatcctgtta</td><td>ccagtggctg</td><td> 3720</td>

<td>ctgccagtgg</td><td>cgataagtcg</td><td>tgtcttaccg</td><td>ggttggactc</td><td>aagacgatag</td><td>ttaccggata</td><td> 3780</td>

<td>aggcgcagcg</td><td>gtcgggctga</td><td>acggggggtt</td><td>cgtgcacaca</td><td>gcccagcttg</td><td>gagcgaacga</td><td> 3840</td>

<td>cctacaccga</td><td>actgagatac</td><td>ctacagcgtg</td><td>agctatgaga</td><td>aagcgccacg</td><td>cttcccgaag</td><td> 3900</td>

<td>ggagaaaggc</td><td>ggacaggtat</td><td>ccggtaagcg</td><td>gcagggtcgg</td><td>aacaggagag</td><td>cgcacgaggg</td><td> 3960</td>

<td>agcttccagg</td><td>gggaaacgcc</td><td>tggtatcttt</td><td>atagtcctgt</td><td>cgggtttcgc</td><td>cacctctgac</td><td> 4020</td>

<td>ttgagcgtcg</td><td>atttttgtga</td><td>tgctcgfccag</td><td>SSgggcggag</td><td>cctatggaaa</td><td>aacgccagca</td><td> 4080</td>

acgcggcctt tttacggttc gqtggccttt ctagcctttt, 4140 tgctcacatg ttctttcctg cgttatcccc tgattctgtg.gataaccgta ttaccgcctfc tgagtg-AGCT gataccgctc 4200 gccgcagcçg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa ga 4252 <210> 21 <211> 7447 <212> DNA <213> vector pFPMT-CL-El-K6 <400 > 21

<td>ggtaccctgc</td><td>tcaatctccg</td><td>gaatggtgat</td><td>ctgatcgttc</td><td>ctgaaaacct </td><td>cgacattggc</td><td> 60</td>

<td>tccctcctga</td><td>cacaggtact</td><td>cgtacaggtt</td><td>ccãggtaaac</td><td>gagtcgtagt</td><td>tgtcgatcat</td><td> 120</td>

<td>gacaacgttc</td><td>ttagaagcgg</td><td>ccggcatttt</td><td>gaaggtgact</td><td>aatagcctaa.</td><td>gaaaatattt</td><td> 180</td>

<td>aatttaattt</td><td>tcattaaatt</td><td>ttcctatact</td><td>cgctatttca</td><td>gcttttcatc</td><td>tcatcacttc</td><td> 240·</td>

<td>ataaacgata</td><td>taaaccagaa</td><td>aaagaactat</td><td>tttcaaacac</td><td>gcttctcaaa</td><td>agcggtatgt</td><td> 300</td>

<td>ccttccacgt</td><td>ctccttagaa</td><td>tctggcaagt</td><td>ccgcgagggg</td><td>gatccttaat</td><td>ggtgatggtg</td><td> 360</td>

<td>gtggtgccag</td><td>ttcatcatca</td><td>tatcccaagc</td><td>catacggtga</td><td>cctgttatgt</td><td>ggccgggata</td><td> 420</td>

<td>gattgagcaa</td><td>ttgcagtcct</td><td>gcaccgtctc</td><td>atgccggcga</td><td>ggcgagatgg</td><td>tgaacagctg</td><td> 480</td>

<td>ggagacgagg</td><td>aagacagatc</td><td>cgcagagatc</td><td>ccccacgtac</td><td>atagcggaac</td><td>agaaagcagc</td><td> 540</td>

<td>cgccccaacg</td><td>agcaaatcgà</td><td>cgtggcgtcg</td><td>tattgtcgta</td><td>gtggggacgc</td><td>tggcgttcct</td><td> 600</td>

<td>agctgcgagc</td><td>gtgggggtga</td><td>gcgctaccca</td><td>gcagcgggaa</td><td>gagttgttct</td><td>cccgaacgca</td><td> 660</td>

<td>gggcacgcac</td><td>ccgggggtgt</td><td>gcatgatcat</td><td>gtccgctgcc</td><td>tcatacacaa</td><td>tgcttgagtt</td><td> 720</td>

<td>ggagcagtcg</td><td>ttcgtgacat</td><td>ggtacatccc</td><td>ggacacgttg</td><td>cgcacctcat</td><td>atcctagtgc</td><td> 780</td>

<td>tgctagtggt</td><td>aggaagcata</td><td colspan="2">gtactagtat tagtaggctt</td><td>cgcatgaatt</td><td>cccgatgaag</td><td> 840</td>

<td>cagagagcgc</td><td>aggaggcggt</td><td>atttatagtg</td><td>ccattcccct</td><td>ctctgagaga</td><td>cccggatggt</td><td> 900</td>

<td>agtcgagtgt</td><td>atcggagaca</td><td>gcttgatgta</td><td>gactccgtgc</td><td>ctgccggctc</td><td>ctcttattgg</td><td> 960</td>

<td>cggacaccag</td><td>tgagacaccc</td><td>cggaacttgc</td><td>tgtttttctg</td><td>caaaatccgg</td><td>ggtgaccagt</td><td> 1020</td>

<td>gogagcctat</td><td>ttgcacacac</td><td>gagcgggaca</td><td>ccccactctg</td><td>gtgaagagtg</td><td>ecaaagtcat</td><td> 1080</td>

<td>tctttttccc</td><td>gttgcggggc</td><td>agccgattgc</td><td>atgttttagg</td><td>aaaatattac</td><td>ctttgctaca</td><td> 1140</td>

<td>ccctgtcaga</td><td>tttaccctcc</td><td>acacatatat</td><td>attccgtcac</td><td>ctccagggac</td><td>tattattcgt</td><td> 1200</td>

<td>cgttgcgccg</td><td>ccagcggaag</td><td>atatccagaa</td><td>gctgttttcc</td><td>gagagactcg</td><td>gttggcgcct</td><td> 1260</td>

<td>ggtatatttg</td><td>atggatgtcg</td><td>cgctgcctca</td><td>cgtcccggta</td><td>cccaggaacg</td><td>eggtgggatc</td><td> 1320</td>

<td>tcgggcccat</td><td>cgaagactgt</td><td>gctccagact</td><td>gçtcgcccag</td><td>caggtgtttc</td><td>ttgatcgccg</td><td> 1380</td>

<td>cctctaaatt</td><td>gtccgcgcat</td><td>cgccggtaac</td><td>atttttccag</td><td>ctcggagttt</td><td>gcgtttagat</td><td> 1440</td>

<td>acagtttctg</td><td>cgatgccaaa</td><td>ggagcctgca</td><td>gattataacc</td><td>tcggatgctg '</td><td>tcattcagcg</td><td> 1500</td>

<td>cttttaattt</td><td>gacctccaga</td><td>tagttgctgt</td><td>atttctgtte</td><td colspan="2">ecattggctg ctgcgcagct</td><td> 1560</td>

<td>tcgtataact</td><td>cgagttattg</td><td>ttgcgctctg</td><td>cctcggcgta</td><td>ctggctcatg</td><td>atctggatct</td><td> 1620</td>

<td>tgtccgtgtc</td><td>gcttttcttc</td><td>gagtgtttct</td><td>cgcaaacgat</td><td>gtgcacggcc</td><td>tgcagtgtcc</td><td> 1680</td>

<td>aatcggagtc</td><td>gagctggcgc</td><td>cgaaactgge</td><td>ggatctgagc</td><td>ctccacactg</td><td>ccctgtttct</td><td> 1740</td>

<td>ctatccacgg</td><td>cggaaccgcc</td><td>tcctgccgtt</td><td>tcagaatgtt</td><td>gttcaagtgg</td><td>tactctgtgc</td><td> 1800</td>

<td>ggtcaatgaa</td><td>ggcgttattg</td><td>ccggtgaaat</td><td>ctttgggaag</td><td>cggttttcct</td><td>cggggaagat</td><td> 1860</td>

<td>tacgaaattc</td><td>cccgcgtcgt</td><td>tgcgcttcct</td><td>ggatctcgag</td><td>gagatcgttc</td><td>tccgcgtcga</td><td> 1920</td>

<td>ggagatcgtt</td><td>ctccgcgtcg</td><td>acaceattcc</td><td>ttgcggcggc</td><td>ggtgctcaac</td><td>ggcctcaacc</td><td> 1980</td>

<td>tactactggg</td><td>ctgcttccta</td><td>atgcaggagt</td><td>cgcataaggg</td><td>agagcgtcga</td><td>caaacccgcg</td><td> 2040</td>

<td>tttgagaact</td><td>tgctcaagct</td><td>tctggtaaac</td><td>gttgtagtac</td><td>tctgaaacaa</td><td>ggccctagca</td><td> 2100</td>

<td>ctctgatctg</td><td>tttctcttgg</td><td>gtagcggtga</td><td>gtggtttatt</td><td>ggagttcact</td><td>ggtttcagca</td><td> 2160</td>

<td>catctgtcat</td><td>ctagacaata</td><td>ttgttactaa</td><td>atttttttga</td><td>actacaattg</td><td>ttcgtaattc</td><td> 2220</td>

<td>atctattatt</td><td>attackatcctc</td><td>gtcagcaatt</td><td>tctggcagac</td><td>ggagtttact</td><td>aacgtcttga</td><td> 2280</td>

<td>gtatgaggcc</td><td>gagaatccag</td><td>ctctgtggcc</td><td>atactcagtc</td><td>ttgacagcct</td><td>gctgatgtgg</td><td> 2340</td>

<td>ctgcgttcaa</td><td>cgcaataagc</td><td>gtgtcctccg</td><td>actccgagtt</td><td>gtgctcgtta</td><td>tcgtcgttct</td><td> 2400</td>

<td>catcctcgga</td><td>aaaatcacac</td><td>gaaagaacat</td><td>actcaccagt</td><td>aggctttctg</td><td>gtccctgggg</td><td> 2460</td>

<td>cacggctgtt</td><td>tctgacgtat</td><td>tccggcgttg</td><td>ataatagctc</td><td>gaaagtgaac</td><td>gccgagtcgc</td><td> 2520</td>

<td>gggagtcgac</td><td>cgatgccctt</td><td>gagagccttc</td><td>aacccagtca</td><td>gctccttccg</td><td>gtgggcgcgg</td><td> 2580</td>

<td>ggcatgacta</td><td>tcgtcgccgc</td><td>acttatgact</td><td>gtcttcttta</td><td>tcatgcaact</td><td>cgtaggacag</td><td> 2640</td>

<td>gtgccggcag</td><td>cgctctgggt</td><td>cattttcggc</td><td>gaggaccgct</td><td>ttcgctggag</td><td>cgcgacgatg</td><td> 2700</td>

atcggcctgt cgcttgcggt attcggaatc ttgcacgccc tcgctcaagc cttcgtcact 2760 ggtcccgcca ccaaacgttt cggcgagaag caggccatta tcgccggcat ggcggccgac 2820 gcgctgggct acgtcttgct ggcgttcgcg aogcgaggct ggatggcctt ccccattatg 2860 attcttctcg çttccggcgg categggatg cccgcgttgc aggccatgct gtccaggcag 2940 gtagatgacg accatcaggg acagcttcaa ggatcgctcg cggctcttac cagcctaact 3000 tcgatcactg gaccgctgat cgtcacggcg atttatgccg cctcggcgag cacatggaac 3060 gggttggcat ggattgtagg cgccgcccta taccttgtct gcctccccgc 3120 gttgcgtcgc ggtgcatgga gccgggccac ctcgacctga atggaagccg gcggcacctc gctaacggat 3180 tcaccactcc aagaattgga gccaatcaat tcttgcggag aactgtgaat gcgcaaacca 3240 acccttggggacggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggggg

9999999999 9999999999 gggcaaacaa ttcatcattt tttttttatt cttttttttg 3360 atttcggttt ctttgaaatt tttfctgatfcc ggtaatetce gaacagaagg aagaacgaag 3420 gaaggagcac agacttagat tggtatatat acgcatatgt agtgttçaag aaacatgaaa 3480 ttgcocagta ttcttaaccc aactgcacag aacaaaaacc tgcaggaaac gaagataaat 3540 catgtcgaaa gctacatata aggaacgtgc tgctáctcat cctagtcctg ttgctgccaa 3600 gctatttaat atcatgcacg aaaagcaaac aaacttgtgt gcttcattgg 3660 atgttcgtac

<td>caccaaggaa</td><td>ttactggagt</td><td>tagttgaagc</td><td>attaggtccc</td><td>aaaatttgtt 1</td><td>tactaaaaac</td><td> 3720</td>

<td>acatgtggat</td><td>atcttgactg</td><td>atttttccat</td><td>ggagggcaca</td><td>gttaagccgc</td><td>taaaggcatt</td><td> 3780</td>

<td>atccgccaag</td><td>tacaattttt</td><td>tactcttçga</td><td>agacagaaaa</td><td>tttgctgaca</td><td>ttggtaatac</td><td> 3840</td>

<td>agtcaaattg</td><td>cagtactctg</td><td>cgggtgtata</td><td>cagaatagca</td><td>gaatgggcag</td><td>acattacgaa</td><td> 3900</td>

<td>tgcacacggt</td><td>gtggtgggcc</td><td>caggtattgt</td><td>tagcggtttg</td><td>aagcaggcgg</td><td>cagaagaagt</td><td> 3960</td>

<td>aacaaaggaa</td><td>cctagaggcc</td><td>ttttgatgtt</td><td>agcagaattg</td><td>tcatgcaagg</td><td>gctccctatc</td><td> 4020</td>

<td>tactggagaa</td><td>tatactaagg</td><td>gtactgttga</td><td>cattgcgaag</td><td>agcgacaaag</td><td>attttgttat</td><td> 4080</td>

<td>cggctttatt</td><td>gctcaaagag</td><td>acatgggtgg</td><td>aagagatgaa</td><td>ggttacgatt</td><td>ggttgattat</td><td> 4140</td>

<td>gacacccggt</td><td>gtgggtttag</td><td>atgacaaggg</td><td>agacgçattg</td><td>ggtcaacagt</td><td>atagaaccgt</td><td> 4200</td>

<td>ggatgatgtg</td><td>gtctctacag</td><td>gatctgácat</td><td>tattattgtt</td><td>ggaagaggac</td><td>tatttgcaaa</td><td> 4260</td>

<td>gggaagggat</td><td>gct.aaggt.ag</td><td>agggtgaacg</td><td>ttacagaaaa</td><td>gcaggctggg</td><td>aagcatattt</td><td> 4320</td>

<td>gagaagatgc</td><td>ggccagcaaa</td><td>actaaaaaac</td><td>tgtattataa</td><td>gtaaatgcat</td><td>gtatactaaa</td><td> 4380</td>

<td>ctcacaaatt</td><td>agagcttcaa</td><td>tttaattata</td><td>tcagttatta</td><td>cocgggaatc</td><td>tcggtcgtaa</td><td> 4440</td>

<td>tgatttttat</td><td>aatgacgaaa</td><td>aaaaaaaaat</td><td>tggaaagaaa</td><td>agcccccccc</td><td>cccccccccc</td><td> 4500</td>

<td>cccccccccc</td><td>ccccccgcag</td><td>cgttgggtcc</td><td>tggccacggg</td><td>tgcgcatgat</td><td>cgtgctcctg</td><td> 4560</td>

<td>tcgttgagga</td><td>cccggctagg</td><td>ctggcggggt</td><td>tgccttactg</td><td>gttagcagaa</td><td>tgaatcaccg</td><td> 4620</td>

<td>attackgcgagc</td><td>gaacgtgaag</td><td>cgactgctgc</td><td>tgcaaaacgt</td><td>ctgcgacctg</td><td>agcaacaaca</td><td> 4680</td>

<td>tgaatggtct</td><td>tcggtttccg</td><td>tgtttcgtaa</td><td>agtctggaaa</td><td>cgcggaagtc</td><td>agcgccctgc</td><td> 4740</td>

<td>accattatgt</td><td>tccggatctg</td><td>catcgcagga</td><td>tgctgctggc</td><td>taccctgtgg</td><td>aacacctaca</td><td> 4800</td>

<td>tctgtattaa</td><td>cgaagcgctg</td><td>gcattgaccc</td><td>tgagtgattt</td><td>ttctctggtc</td><td>ccgccgcatc</td><td> 4860</td>

<td>cataccgcca</td><td>gttgtttacc</td><td>ctcacaacgt</td><td>tccagtaacc</td><td>gggcatgttc</td><td>atcatcagta</td><td> 4920</td>

<td>acccgtatcg</td><td>tgagcatcct</td><td>ctctcgtttc</td><td>atcggtatca</td><td>ttacccccat</td><td>gaacagaaat</td><td> 4980</td>

<td>teeeecttae</td><td>acggaggcat</td><td>caagtgacca</td><td>aacaggaaaa</td><td>aaocgccctt</td><td>aacatggccc</td><td> 5040</td>

<td>gctttatcag</td><td>aagccagaca</td><td>ttaacgcttc</td><td>tggagaaact</td><td>caacgagctg</td><td>gacgcggatg</td><td> 5100</td>

<td>aacaggcaga</td><td>catctgtgaa</td><td>tcgcttcacg</td><td>aceacgctga</td><td>tgagctttac</td><td>cgcagctgcc</td><td> 5160</td>

<td>tcgcgcgttt</td><td>cggtgatgac</td><td>ggtgaaaacc</td><td>tctgacacat</td><td>gcagctccçg</td><td>gagacggtca</td><td> 5220</td>

<td>cagcttgtct</td><td>gtaagcggat</td><td>gccgggagca</td><td>gacaagcccg</td><td>tcagggcgcg</td><td>tcagcgggtg</td><td> 5280</td>

<td>ttggcgggtg</td><td>tcggggcgca</td><td>gccatgaccc</td><td>agtcacgtag</td><td>cgatagcgga</td><td>gtgtatactg</td><td> 5340</td>

<td>gcttaactat</td><td>gcggcatcag</td><td>agcagattgt</td><td colspan="2">actgagagtg caccatatgc</td><td>ggtgtgaaat</td><td> 5400</td>

<td>accgcacaga</td><td>tgcgtaagga</td><td>gaaaataccg</td><td>catcaggcgc</td><td>tcttccgctt</td><td>cctcgctcac</td><td> 5460</td>

<td>tgactcgctg</td><td>cgcfccggtcg</td><td>ttcggctgcg</td><td>gcgagcggta</td><td>tcagctcact</td><td>caaaggcggt</td><td> 5520</td>

<td>aatacggtta</td><td>tccacagaat</td><td>caggggataa</td><td>cgcaggaaag</td><td>aacatgtgag</td><td>caaaaggcca</td><td> 5580</td>

<td>gcaaaaggcc</td><td>aggaaccgta</td><td>aaaaggccgc</td><td>gttgctggcg</td><td>tttttccata</td><td>ggctccgccc</td><td> 5640</td>

<td>coctgacgag</td><td>catcacaaaa</td><td>atcgacgctc</td><td>aagtcagagg</td><td>tggcgaaacc</td><td>cgacaggact</td><td> 5700</td>

<td>ataaagatac</td><td>caggcgtttc</td><td>cccctggaag</td><td>ctccctcgtg</td><td>cgctctcctg</td><td>ttccgaccct</td><td> 5760</td>

<td>gccgcttacc</td><td>ggatacctgt</td><td>ccgcctttct</td><td>cccttcggga</td><td>agcgtggcgc</td><td>tttctcatag</td><td> 5820</td>

<td>ctcacgctgt</td><td>aggtatctca</td><td>gttcggtgta</td><td>ggtcgttcgc</td><td>tccaagctgg</td><td>gctgtgtgca</td><td>S880</td>

<td>cgaacccccc</td><td>gttcagcecg</td><td>accgctgcgc</td><td>cttatccggt</td><td>aactatcgtc</td><td>ttgagtecaa</td><td> 5940</td>

<td>cocggtaaga</td><td>cacgacttat</td><td>cgccactggc</td><td>agcagccact</td><td>ggtaacagga</td><td>ttagcagagc</td><td> 6000</td>

<td>fcã.</td><td>ggcggtgcta</td><td>cagagttctt</td><td>gaagtggtgg</td><td>cctaactacg</td><td>gctacactag</td><td> 6060</td>

<td>aaggacagta</td><td>tttggtatct</td><td>gcgctctgct</td><td>gaagccagtt</td><td>accttcggaa</td><td>aaagagttgg</td><td> 6120</td>

<td>tagctcttga</td><td>tccggcaaac</td><td>aaaccaccgc</td><td>tggtagcggt</td><td>çgtttttttg</td><td>tttgcaagca</td><td> 6180</td>

gcagattacg aaggatctca agaagatcct ttgatctttt cgcagaaaaa ctacggggtc 6240 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag 6300 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata 6360 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat 6420 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg 6480 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc 6540 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa 6600 gtggtcctgc aactttatcc goctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc 6660 gccagttaat agtttgcgca acgttgttgc cattgctgca ggcatcgtgg tgtcacgctc 6720 gtcgtttggt atggettcat tcagctocgg ttcccaacga tcaaggcgag ttacatgatc 6780 ccccatgttg tgcaaaaaag cggttagctc cttoggtcct ccgat-cgttg tcagaagtaa 6840 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat 6900 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata 6960 gtgtatgcgg çgaccgagtt gctcttgocc ggcgtcaaca cgggataata ccgcgccaca 7020 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag 7080 gatcttaccg ctgttgagat ccagtfccgat gtaacccact cgtgcaocca actgatcttc 7140 agcatctttt açtttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaãtgccgc 7200 aaaaaaggga ataagggcga cacggaaatg ttgaatacfcc atactcttcc ttttfceaata 7260 ttattgaagc atttatcagg gttattgtçt catgagcgga tacatatttg aatgtattta 7320 gaaaaataaa caaatagggg ttccgcgcac atttceccga aaagtgcc & c ctgacgtcta 7380 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg 744Ò tcttcaa 7447 <210> 22 <211> 3730 <212> DNA <213> vector pSP72E2H6 <400> 22

<td>gaactcgagc</td><td>agctgaagct</td><td>tgaattcatg</td><td>agatttcctt</td><td>caatttttac</td><td>tgcagtttta</td><td> 60</td>

<td>ttcgcagcat</td><td>cctccgcatt</td><td>agctgctcca</td><td>gtcaacacta</td><td>caacagaaga</td><td>tgaaacggca</td><td> 120</td>

<td>caaattccgg</td><td>ctgaagctgt</td><td>catcggttac</td><td>tcagatttag</td><td>aaggggattt</td><td>cgatgttgct</td><td> 180</td>

<td>gttttgccat</td><td>tttccaacag</td><td>eacaaataae</td><td>gggttattgt</td><td>ttataaatac</td><td>tactattgcc</td><td> 240</td>

<td>agcattgctg</td><td>ctaaagaaga</td><td>aggggtatet</td><td colspan="2">ctagataaaá ggcatacccg</td><td>cgtgtcagga</td><td> 300</td>

<td>ggggcagcag</td><td>cctccgatac</td><td>caggggcctt</td><td>gtgtccctct</td><td colspan="2">ttagccecgg gteggctcag</td><td> 360</td>

<td>aaaatccagc</td><td>tcgtaaacac</td><td>caacggcagt</td><td>tggcacatca</td><td>acaggactgc</td><td>cctgaactge</td><td> 420</td>

<td>aacgactccc</td><td>tccaaacagg</td><td>gttctttgcc</td><td>gcactattct</td><td>acaaacacaa</td><td>attcaactcg</td><td> 480</td>

<td>tctggatgcc</td><td>cagagcgctt</td><td>ggccagctgt</td><td>cgctccatcg</td><td>acaagttcgc</td><td>tcaggggtgg</td><td>S40</td>

<td>ggteccctca</td><td>cttacactga</td><td>gcctaacagc</td><td>tcggaccaga</td><td>ggccctactg</td><td>ctggcactac</td><td> 600</td>

<td>gcgcctcgac</td><td>cgtgtggtat</td><td>tgtaccegcg</td><td>tctcaggtgt</td><td>gcggtccagt</td><td>gtattgcttc</td><td> 660</td>

<td>accccgagcc</td><td>ctgttgtggt</td><td>ggggacgacc</td><td>gatcggtttg</td><td>gtgtccccac</td><td>gtataactgg</td><td> 720</td>

<td>ggggcgaacg</td><td>actcggatgt</td><td>gctgattctc</td><td>aacaacacgc</td><td>ggccgccgcg</td><td>aggcaactgg</td><td> 780</td>

<td>ttcggctgta</td><td>catggatgaa</td><td>tggcactggg</td><td>ttcaccaaga</td><td>cgtgtggggg</td><td>ccccccgtgc</td><td> 840</td>

<td>aacatcgggg</td><td>gggccggcaa</td><td>caacaccttg</td><td>acctgcccca</td><td>ctgaetgttt</td><td>tcggaagcac</td><td> 900</td>

<td>cccgaggcca</td><td>cttacgccag</td><td>atgcggttct</td><td>gggccctggc</td><td>tgacacctag</td><td>gtgtatggtt</td><td> 960</td>

cattacccat afcaggctctg gcactacocc tgcactgtca acttcaocat cttcaaggtt 1020

<td>aggatgtacg</td><td>tggggagcgt</td><td>ggagcacagg</td><td>ttcgaagecg</td><td>catgcaattg</td><td>gactcgagga</td><td> 1080</td>

<td>gagcgttgtg</td><td>acttggagga</td><td>cagggataga</td><td>teagagetta</td><td>gctcgctgct</td><td>gctgtctaca</td><td> 1140</td>

<td>aeagagtggc</td><td>aggtgatega</td><td>gggcagacac</td><td>catcaceacc</td><td>ateactaata</td><td>gttaattaae</td><td> 1200.</td>

<td>gatctcgact</td><td>tggttgaaca</td><td>cgttgccaag</td><td>gcttaagtga</td><td>atttacttta</td><td>aagtcttgca</td><td> 1260</td>

<td>tttaaataaa</td><td>ttttettttt</td><td>atagctttat</td><td>çactfcagttt</td><td>caatttatat</td><td>actattttaa</td><td> 1320</td>

<td>tgacattttc</td><td>gattcattga</td><td>ttgaaagcta</td><td>tcagatctgc</td><td>cggtctccct</td><td>atagtgagtc</td><td> 1380</td>

<td>gtattaattt</td><td>cgataagcca</td><td>ggttaacctg</td><td>eattaatgaa</td><td>tcggccaacg</td><td>cgcggggaga</td><td> 1440</td>

<td>ggcggtttgc</td><td>gtattgggcg</td><td>ctcttccgct</td><td>tcctcgctca</td><td>ctgactcgct</td><td>gcgctcggtc</td><td> 1500</td>

<td>gttcggctgc</td><td>ggcgagcggt</td><td>atcagctcac</td><td>tcaaaggcgg</td><td>taatacggtt</td><td>atccacagaa</td><td> 1560</td>

<td>tcaggggata</td><td>acgcaggaaa</td><td>gaacatgtga</td><td>gcaaaaggec</td><td>agcaaaaggc</td><td>caggaaccgt</td><td> 1620</td>

<td>aaaaaggccg</td><td>cgttgctggc</td><td>gtttttccat</td><td>aggctccgcc</td><td>cccctgacga</td><td>gcatcacaaa</td><td> 1680</td>

<td>aatcgacgct</td><td>caagtcagag</td><td>gtggcgaaac</td><td>cegacaggae</td><td>tataaagata</td><td>ecaggcgttt</td><td> 1740</td>

<td>ccccctggaa</td><td>gctccctcgt</td><td>gcgctctcct</td><td>gttccgaocc</td><td>tgccgcttac</td><td>cggataectg</td><td> 1800</td>

<td>tccgcctttc</td><td>tcccttcggg</td><td>aagcgtggcg</td><td>ctttcteaat</td><td>gctcacgctg</td><td>taggtatetc</td><td> 1860</td>

<td>agttcggtgt</td><td>aggtcgttcg</td><td>ctceaagctg</td><td>ggetgtgtgc</td><td>acgaacccec</td><td>cgttcagccc</td><td> 1920</td>

<td>gaccgctgcg</td><td>ccttatccgg</td><td>taactatcgt</td><td>cttgagtcca</td><td>acccggtaag</td><td>acacgactta</td><td> 1980</td>

<td>tcgceactgg</td><td>cagcagccac</td><td>tggtaaeagg</td><td>attagcagag</td><td>cgaggtatgt</td><td>aggcggtgct</td><td> 2040</td>

<td>acagagttct</td><td>tgaagtggtg</td><td>gcctaactac</td><td>ggctacacta</td><td>gaaggacagt</td><td>atttggtatc</td><td> 2100</td>

<td>tgcgctctgc</td><td>tgaagecagt</td><td>taccttcgga</td><td>aaaagagttg</td><td>gtagctettg</td><td>atccggcaaa</td><td> 2160</td>

<td>caaaccaccg</td><td>ctggtagcgg</td><td>tggttttttt</td><td>gtttgeaagc</td><td>agcagattac</td><td>gcgcagaaaa</td><td> 2220</td>

<td>aaaggatctc</td><td>aagaagatcc</td><td>tttgatcttt</td><td>tctacggggt</td><td>ctgacgctca</td><td>gtggaacgaa</td><td> 2280</td>

aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcett 2340 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac 2400 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc 2460 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc 2S20 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata 2580 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcct-ccatc 2640 cagtctatta attgttgocg ggaagctaga gtaagtagtt cgccagttaa tagtttg -cgc 2700 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca 2760 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa 2820 gcggggtgtcggggtcgcggggtcgcggggtcggcgggtggcggggtcggcggggtcggggtggcggggtgt

<td>ctcatggtta</td><td>tggcagcact</td><td>gcataattct</td><td>çttactgtca</td><td>tgccatccgt</td><td>aagatgcttt</td><td> 2940</td>

<td>tctgtgactg</td><td>gtgagtactc</td><td>aaccaagtca</td><td>ttctgagçat</td><td>agtgtatgcg</td><td>gçgaccgagt</td><td> 3000</td>

<td>tgctcttgcc</td><td>cggcgtcaat</td><td>acgggataat</td><td>accgcgccac</td><td>atagcagaac</td><td>tttaaaagtg</td><td> 3060</td>

<td>ctcatcattg</td><td>gaaaacgttc</td><td>ttcggggçga</td><td>aaactctcaa</td><td>ggatcttacc</td><td>gctgttgaga</td><td> 3120</td>

<td>tccagttcga</td><td>tgtaacccac</td><td>tcgtgcaccc</td><td>aactgatctt</td><td>cagcatcttt</td><td>tactttcacc</td><td> 3180</td>

<td>agcgtttctg</td><td>ggtgàgcaaa</td><td>aacaggaagg</td><td>caaaatgccg</td><td>caaaaaaggg</td><td>aataagggcg</td><td> 3240</td>

<td>acacggaaat</td><td>gttgaatact</td><td>catactcttc</td><td>ctttttcaat</td><td>attattgaag</td><td>catttatcag</td><td> 3300</td>

<td>ggttattgtc</td><td>tcatgagcgg</td><td>attackattt</td><td>gaatgtattt</td><td>agaaaaataa</td><td>acaaataggg</td><td> 3360</td>

<td>gttccgcgca</td><td>catttccccg</td><td>aaaagtgcca</td><td>cctgacgtct</td><td>aagaaaccat</td><td>tattatcatg</td><td> 3420</td>

<td>acattaacct</td><td>ataaaaatag</td><td>gcgtatcacg</td><td>aggccctttc</td><td>gtctcgcgcg</td><td>tttcggtgat</td><td> 3480</td>

<td>gacggtgaaa</td><td>aicctctgaca</td><td>catgçagctc</td><td>ccggagacgg</td><td>tcacagcttg</td><td>tctgtaagcg</td><td> 3540</td>

<td>gatgccggga</td><td>gcagacaagc</td><td>ccgtcagggc</td><td>gcgtcagegg</td><td>gtgttggcgg</td><td>gtgtcggggc</td><td> 3600</td>

<td>tggcttaact</td><td>atgeggeatc</td><td>agagcagatt</td><td>gtactgagag</td><td>tg-caccatat</td><td>ggacatattg</td><td> 3660</td>

<td>tcgttagaac</td><td>gcggctacaa</td><td>ttaatacata</td><td>accttatgta</td><td>tcatacacat</td><td>acgatttagg</td><td> 3720</td>

tgacactata 3730 <211> 7370 <212> DNA <213> vector pMPT121 '<22 0>

<221> MISCELLANEOUS <222> (778) .. (778) <223> N is any nucleotide <400> 23

<td>ggtaccctgc</td><td>tcaatctocg</td><td>gaatggtgat</td><td>ctgatcgttc</td><td>ctgaaaacct</td><td>cgacattggc</td><td> €0</td>

<td>tccctcctga</td><td>cacaggtact</td><td>cgtacaggtt</td><td>ceaggtaaac</td><td>gagtcgtagt</td><td>tgtcgatcat</td><td> 120</td>

<td>gacaacgttc</td><td>ttagaagcgg</td><td>ccggcatttt</td><td>gaaggtgact</td><td>aatagcctaa</td><td>gaaaatattt</td><td> 180</td>

<td>aatttaattt</td><td>tcattaaatt</td><td>ttcctatact</td><td>cgctatttca</td><td>gcttttcatc</td><td>tcatcacttc</td><td> 240</td>

<td>ataaacgata</td><td>taaaccagaa</td><td>aaagaactat</td><td>tttcaaacac</td><td>gcttctcaaa</td><td>agcggtatgt</td><td> 300</td>

<td>ccttccacgt</td><td>ctccttagaa</td><td>tctggcaagt</td><td>ccgcgagggg</td><td>gatccagatc</td><td>tgaattcgtt</td><td> 360</td>

<td>tttgtacttt</td><td>agattgatgt</td><td>caccaccgtg</td><td>cactggcagc</td><td>agtatttata</td><td>gatggaccgt</td><td> 420</td>

<td>gtggggacgg</td><td>ttgggtacac</td><td>ttagoggçag</td><td>cgctgacccc</td><td>atctgtgatc</td><td>aagtagggca</td><td> 480</td>

<td>aaaactgggg</td><td>atgtcggagt</td><td>cgctgcacgg</td><td>tagcataaga</td><td>atttactttc</td><td>tggecggttc</td><td> 540</td>

<td>acocgcattt</td><td>gcactgtgga</td><td>gaaacagcct</td><td>gtccgacacc</td><td>ccaccagttg</td><td>ccacatcggc</td><td> 600</td>

<td>cctctgctgc</td><td>tctggtgatt</td><td>ttctggtagc</td><td>aggeacagac</td><td>agcagtgggt</td><td>agcgccgtcc</td><td> 660</td>

<td>ggttaggcaa</td><td>ggtcacgttg</td><td>taggctaccc</td><td>cagcaaacag</td><td>agcctcacat</td><td>gacaceatcc</td><td> 720</td>

<td>agctgcgtcc</td><td>tcgaagcgaa</td><td>aagttcggtt</td><td>gcggetgcag</td><td>aaccccctca</td><td>gttgccanat</td><td> 780</td>

<td>teacáagttt</td><td>tacgcgacgg</td><td>ctaaagegag</td><td colspan="2">tgggttttaa aaacttgcgg</td><td>tgcaaggatg</td><td> 840</td>

<td>catgcggcaa</td><td>caattaattg</td><td>gtgcatecag</td><td>cacagcaagc</td><td>ccagtctcga</td><td>gatgtccagt</td><td> 900</td>

<td>cgctacagag</td><td>tggagtacgc</td><td>actcaaggaa</td><td>caccgtcgag</td><td>atggcctcat</td><td>agaatggatc</td><td> 960</td>

<td>aagggcctgc</td><td>tggccacgcc</td><td>gttcgtcctg</td><td>tacgcggtga</td><td>agagcaacgg</td><td>catctctgca</td><td> 1020</td>

<td>gtggacgacc</td><td>tcatggtaaa</td><td>ctctgaggca</td><td>aaacgccgct</td><td>acgcggaaat</td><td>cttccacgac</td><td> 1080</td>

<td>ctcgaactcc</td><td>tcatcgacga</td><td>caacattgaa</td><td>atgaceaaag</td><td>ccggcacccc</td><td>cgaattgtct</td><td> 1140</td>

<td>cggctcgtgc</td><td>agctggttcc</td><td>gagcgttggc</td><td>agettettea</td><td>cgagactgcc</td><td>tctggaaaag</td><td> 1200</td>

<td>gccttctaca</td><td>tcgaggacga</td><td>gcgccgcgcc</td><td>atcagcaaac</td><td>gccggcttgt</td><td>ggccccctcg</td><td> 1260</td>

<td>ttcaacgacg</td><td>tccggctcat</td><td>tctcaacacg</td><td>gcccagctgt</td><td>tggagatgtc</td><td>gcggttcttc</td><td> 1320</td>

<td>cattccaaaa</td><td>ccatecgaga</td><td>tcgcaagctg</td><td>eagctcatta</td><td>cattcgatgg</td><td>tgacatcaea</td><td> 1380</td>

<td>ctgtacgacg</td><td>acggcaaaaa</td><td>tttcgatgcc</td><td>gagtcgccca</td><td>tcctgcccca</td><td>cctcatcaaa</td><td> 1440</td>

<td>etaatggeca</td><td>aggacctcta</td><td>fcgfegggtatc</td><td>at rarmrcrt st - ———</td><td>ccggctacag</td><td>cgacggaaca</td><td> 1500</td>

<td>agtactacga</td><td>gcgccteaag</td><td>ggccteatcg</td><td>aegcegtcca</td><td>gacgtccccg</td><td>ctgctcacag</td><td> 1560</td>

<td>gceaccagaa</td><td>agagaacctg</td><td>ttcattatgg</td><td>gcggcgaggc</td><td>aaactacctc</td><td>ttccggtaea</td><td> 1620</td>

gtaacgagga gcagagatta cgcttctact eeaaagacag atggctgctc gagaacafcgc 1660 tgaattggte cgaggaggac atteatctga cactggactt tgegcaggac gttctaaacg 1740 acctcgttca caaactgggc tegccagcca ccgtggtceg caaggagcgt cgcgtcggcc 1800 tggttccatt accgggccac aagctgatcc gcgagcagct cgaggagatc gttctccgcg 1860 tcgacaccat tccttgcggc ggcggtgctc aacggcctca acctactact gggctgcttc 1920 ctaatgcagg agtcgcataa gggagagcgt cgactcccgc gactcggcgt tcactttcga 1980 gctattatea acgccggaat acgteagaaa cagccgtgcc ccagggacca gaaagcctac 2040 tggtgagtat gttctttcgt gtgatttttc cgaggatgag aaegacgata aegagcacaa 2100 ctcggagtcg gaggacacge ttattgcgtt gaacgcagcc acatcagcag gctgtcaaga 2160 ctgagtatgg ccacagagct ggattctcgg octcatactc aagacgttag taaactccgt 2220 ctgccagaaa ttgctgacga ggatgtataa taatagatga attacgaaca attgtagttc 2280 aaaaaaattt agtaacaata fctgfcctagat gacagatgtg ctgaaaccag tgaactccaa 2340, (taaaccactc accgctaçcc aagagaaaca gatcagagtg ctagggcctt gtttcagagt 2400 actacaacgt ttaccágaag cttgagcaag ttctcaaacg cgggtttgtc gaccgatgcc 2460 cttgagagcc ttcaacceag tcagctcctt ccggtgggcg cggggcatga ctatcgtcgc 2520 cgcacttatg actgtcttct ttatcatgca actcgtagga caggtgccgg cagcgctctg 2560 ggcgaggacc gctttcgctg gagcgcgacg ggtcattttc atgatcggcc tgtcgcttgc 2640 ggtattcgga atcttgcacg ccctcgctca agccttcgtc actggtcccg ccaccaaacg 2700 tttcggcgag aagcaggcca ttatcgccgg catggcggcc gacgcgctgg gctacgtctt 2760 gctggcgttc gcgacgcgag gctggatggc cttccccatt atgattcttc tcgcttccgg 2820 cggcatcggg atgcccgcgt tgcaggccat gctgtccagg caggtagatg acgaccatca 2880 gggacagctt caaggatcgc tcgcggctct taccagccta acttcgatca ctggaccgct 2940 gatcgtcacg gcgatttatg ccgcctcggc gagcacatgg aacgggttgg catggattgt 3000 aggcgccgcc ctataccttg tctgcctccc cgcgttgcgt cgcggtgcat ggagccgggc 3060 cacctcgacc tgaatggaag ccggcggcac ctcgctaacg gattcaccac tccaagaatt 3120 ggagccaatc aattcttgcg gagaactgtg aatgcgcaaa ccaacccttg gcagaacata 3180 tccatcgcgt ccgccatctc cagcagccgc acgcggcgca tcgggggggg gggggggggg 3240 ggggsgca ^ a caattcatca tttttttttt attctttttt ttgatttcgg tttcfcttgaa 3300 atttttttga ttcggtaatc tccgaacaga aggaagaacg aaggaaggag cacagactta 3360 gattggtata tatacgcata tgtagtgttg aagaaacatg aaattgccca gtattcttaa 3420 cccaactgca cagaacaaaa acctgcagga aacgaagata aatcatgtcg aaagctacat 3480 ataaggaacg tgctgctact catcctagtc ctgttgctgc caagctattt aatatcatgc 3540 acgaaaagca aacaaacttg tgtgcttcat tggatgttcg taccaccaag gaattactgg 3600 agttagttga agcattaggt cccaaaattt gtttactaaa aacacatgtg gatatcttga 3660 ctgatttttc catggagggc acagttaagc cgctaaaggc attatccgcc aagtacaatt 3720 ttttactctt cgaagacaga aaatttgctg acattggtaa tacagtcaaa ttgcagtact 3780 ctgcgggtgt atacagaata gcagaatggg cagacattac gaatgeacac ggtgtggtgg 3840 gcccaggtat tgttagcggt ttgaagcagg cggcagaaga agtaacaaag gaacctagag 3900 gccttttgat gttagcagaa ttgtcatgca agggctccct atctactgga gaatatacta 3960 agggtactgt tgacattgcg aagagcgaca aagattttgt tatcggcttt attgctcaaa 4020 gagacatggg tggaagagat gaaggttacg attggttgat tatgacaccc ggtgtgggtt 4080 tagatgacaa gggagacgca ttgggtcaac agtatagaac cgtggatgat gtggtctcta 4140 caggatctga cattattatt gttggaagag gactafcttgc aaagggaagg gatgctaagg 4200

<td>tagagggtga</td><td>acgttacaga</td><td>aaagcaggct</td><td>gggaagcata</td><td>tttgagaaga</td><td>tgcggccagc</td><td> 4260</td>

<td>aaaaçtaaaa</td><td>aactgtatta</td><td>taagtaaatg</td><td>catgtatact</td><td>aaactcacaa</td><td>attagagctt</td><td> 4320</td>

<td>caatttaatt</td><td>atatcagtta</td><td>ttacccggga</td><td>atcfccggtcg</td><td>taatgatttt</td><td>tataatgacg</td><td> 4380</td>

<td>aaaaaaaaaa</td><td>aattggaaag</td><td>aaaagooccc</td><td>cccccccccc</td><td>cccccccccc</td><td>cccccccccg</td><td> 4440</td>

<td>cagcgttggg</td><td>tcctggccac</td><td>gggtgcgcat</td><td>gatcgtgctc</td><td>ctgtegttga</td><td>ggacccgget</td><td> 4500</td>

<td>aggctggcgg</td><td>ggttgcctta</td><td>ctagttagca</td><td>gaatgaatca</td><td>ccgatacgcg</td><td>agcgaacgtg</td><td> 4560</td>

<td>aagcgactgc</td><td>tgctgcaaaa</td><td>cgtctgcgac</td><td>ctgagcaaca</td><td>acatgaatgg</td><td>tcttcggttt</td><td> 4620</td>

<td>ccgtgtttcg</td><td>taaagtctgg</td><td>aaacgcggaa</td><td>gtcagcgccc</td><td>tgcaccatta</td><td>tgttccggat</td><td> 4680</td>

<td>ctgcatcgca</td><td>ggatgctgct</td><td>ggctaccctg</td><td>tggaacacct</td><td>acatctgtat</td><td>taacgaagcg</td><td> 4740</td>

<td>ctggcattga</td><td>ccctgagtga</td><td>tttttctctg</td><td>gtcccgccgc</td><td>atccataccg</td><td>ccagttgttt</td><td> 4800</td>

<td>accctcacaa</td><td>cgttccagta àccgggcatg ttcatcatca gtaacccgta tcgtgagcat</td><td> 4860</td>

<td>cctctctcgt</td><td>ttcatcggta tcattacccc cataaacaga aattccccct tacacggagg</td><td> 4520</td>

<td>catcaagtga</td><td>ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag</td><td> 4580</td>

<td>acattaacgc</td><td>ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt</td><td> 5040</td>

<td>gaatcgcttc</td><td>acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat</td><td>5X00</td>

<td>gacggtgaaa</td><td>acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg</td><td>S160</td>

<td>gatgccggga</td><td>gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc</td><td> 5220</td>

<td>gcagccatga</td><td>cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat</td><td> 5280</td>

<td>cagagcagat</td><td>tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa</td><td> 5340</td>

<td>ggagaaaata</td><td>ccgcatcagg cgctcttccg ettcctcgct cactgactcg ctgcgctcgg</td><td> 5400</td>

<td>tcgttcggct</td><td>gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag</td><td> 5460</td>

<td>aatcagggga</td><td>taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc</td><td> 5520</td>

<td>gtaaaaaggc</td><td>cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca</td><td> 5580</td>

<td>aaaatcgacg</td><td>ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt</td><td> 5640</td>

<td>ttccccctgg</td><td>aagctccctc gtgcgctctc ctgttccgac cctgccgctt aecggatacc</td><td> 5700</td>

<td>tgtccgcctt</td><td>tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc</td><td> 5760</td>

<td>tcaç _tcggt</td><td>gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc eccgttcagc</td><td> 5820</td>

<td>ccgaccgctg</td><td>cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact</td><td> 5880</td>

<td>tatcgccact</td><td>ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg</td><td> 5940</td>

<td>ctacagagtt</td><td>cttgaagtgg tggcotaact acggctacac tagaaggaca gtatttggta</td><td> 6000</td>

<td>tctgcgctct gctgaagcca gttaocttcg gaaaaagagt tggtagctct</td><td>tgatccggca</td><td> 6060</td>

<td>aacaaaccaç cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt</td><td>acacgcagaa</td><td> 6120</td>

<td>aaaaaggatc tcaagaagat octttgatct ttt-ctacggg gtctgacgct</td><td>cagtggaacg</td><td> 6180</td>

<td>aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc</td><td>acctagatcc</td><td> €240</td>

<td>ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa</td><td>acttggtctg</td><td> 6300</td>

<td>acagttacca atgcttaatc agtgaggçac ctatctcagc gatetgtcta</td><td>tttcgttcat</td><td> 6360</td>

<td>ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc</td><td>ttaccatctg</td><td> 6420</td>

<td>gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat</td><td>ttatcagcaa</td><td> 6480</td>

<td>taaaccagct agccggaagg gccgagcgca gaagtggtcc tgcaacttta</td><td>tccgcctcca</td><td>6S40</td>

<td>tccagtctat taattgttgc cgggaagçta gagtaagtag ttcgccagtt</td><td>aatagtttgc</td><td> £600</td>

<td>gcaacgttgt tgccattgct gcaggcat-cg tggtgtcacg ctcgtcgttt</td><td>ggtatggctt</td><td> 6660</td>

<td>cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg</td><td>ttgtgcaaaa</td><td> 6720</td>

<td>aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc</td><td>gcagtgttat</td><td> 6780</td>

<td>cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc</td><td>gtaagatgct</td><td> 6840</td>

<td>tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg</td><td>cggcgaccga</td><td> 6900</td>

<td>gttyctcfctg cccggcgtca acacgggata Ataccgcgcc acatagcaga</td><td>actttaaaag</td><td> 6960</td>

<td>tgctcatcat tggaaaacgt tcttcggggc gaaaactctç aaggatctta</td><td>ccgctgttga</td><td> 7020</td>

<td>gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct</td><td>tttactttca</td><td> 7080</td>

ecagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg 7140 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc 7200 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag 7260 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca 7320 tgacattaac ctataaaaat aggcgtatca ogaggccctt tcgtcttcaa 7370 <210> 24 <211> 8298 <212> DNA <213> Vector pFMPT-MFalfa-E2-H6 <400> 24 ggtaccctgc tcaatctccg gaatggtgat ctgatcgttc ctgaasacct cgacattggc -60 tccctcctga cacaggtact cgtacaggtt ccaggtaaac gagtcgtagt tgtcgatcat 120 gacaacgttc ttagaagcgg oçggcatttt gaaggtgact aatagcctaa gaaaatattt 180 aatttaattt tcattaaatt ttcctatact cgctatcctc

<td>ataaacgata</td><td>taaaccagaa</td><td>aaagaactat</td><td>tttcaaacac</td><td>gcttctcaaa</td><td>agcggtatgt</td><td> 300</td>

<td>ccttçcacgt</td><td>ctccttagaa</td><td>tctggcaagt</td><td>ccgccagggg</td><td>gatccagatc</td><td>tgatagcttt</td><td> 360</td>

<td>caatcaatga</td><td>atcgaaaatg</td><td>tcattaaaat</td><td>agtatataaa</td><td>ttgaaactaa</td><td>gtcataaagc</td><td> 420</td>

<td>tataaaaaga</td><td>aaatttattt</td><td>aaatgcaaga</td><td>ctttaaagta</td><td>aattcactta</td><td>agccttggca</td><td> 480</td>

<td>acgtgttcaa</td><td>ccaagtcgag</td><td>atcgttaatt</td><td>aactattagt</td><td>gatggtggtg</td><td>atggtgtctg</td><td> 540</td>

<td>ccctcgatca</td><td>cctgccactc</td><td>tgttgtagac</td><td>agcagcagcg</td><td>agctaagctc</td><td>tgatctatcc</td><td> $00</td>

<td>ctgtcctcca</td><td>agtcacaacg</td><td>ctctcctcga</td><td>gtccaattgc</td><td>atgcggcttc</td><td>gaacctgtgc</td><td> 660</td>

<td>tccacgcccc</td><td>ccacgtacat</td><td>cctaaccttg</td><td>aagatggtga</td><td>agttgacagt</td><td>gcaggggtag</td><td> 720</td>

<td>tgccagagcc</td><td>tatatgggta</td><td>atgaaccata</td><td>cacctaggtg</td><td>tcagccaggg</td><td>cccagaaccg</td><td> 780</td>

<td>catctggcgt</td><td>aagtggcctc</td><td>ggggtgcttc</td><td>cgaaaacagt</td><td>cagtggggca</td><td>ggtcaaggtg</td><td> 840</td>

<td>ttgttgccgg</td><td>cccccccgat</td><td>gttgcacggg</td><td>gggcccccac</td><td>acgtcttggt</td><td>gaacccagtg</td><td> 900</td>

<td>ccattcatcc</td><td>atgtacagcc</td><td>gaaccagttg</td><td>cctcgcggcg</td><td>gccgcgtgtt</td><td>gttgagaatc</td><td> 960</td>

<td>agcacatccg</td><td>agtcgttcgc</td><td>cccccagtta</td><td>tacgtgggga</td><td>caccaaaccg</td><td>atcggtcgtc</td><td> 1020</td>

<td>cccaccacaa</td><td>cagggctcgg</td><td>ggtgaagcaa</td><td>tacactggac</td><td>cgcacacctg</td><td>agacgcgggt</td><td> 1080</td>

<td>acaataccac</td><td>acggtcgagg</td><td>cgegtagtgc</td><td>cagcagtagg</td><td>gcctctggtc</td><td>cgagctgtta</td><td> 1140</td>

<td>ggctcag tgt</td><td>ãagtgagggg</td><td>aeceeaccec</td><td>tgagcgaact</td><td>tgtcgatgga</td><td>gcgacagctg</td><td> 1200</td>

<td>gccaagcgct</td><td>ctgggcatcc</td><td>agacgagttg</td><td>aatttgtgtt</td><td>tgtagaatag</td><td>tgcggcaaag</td><td> 1260</td>

<td>aaccctgttt</td><td>ggagggagtc</td><td>gttgcagttc</td><td>agggcagtcc</td><td>tgttgatgtg</td><td>ccaactgccg</td><td> 1320</td>

<td>ttggtgttta</td><td>cgagctggat</td><td>tttctgagcc</td><td>gacccggggc</td><td>taaagaggga</td><td>cacaaggccc</td><td> 1380</td>

<td>ctggtatcgg</td><td>aggctgctgc</td><td>ccctcctgac</td><td>acgcgggtat</td><td>gccttttatc</td><td>tagagatacc</td><td> 1440</td>

<td>ccttcttctt</td><td>tagcagcaat</td><td>gctggcaata</td><td>gtagtattta</td><td>taaacaataa</td><td>cccgttattt</td><td> 1500</td>

gtgctgttgg aaaatggcaa aacagcaaca tcgaaatccc cttctaaatc tgagtaaccg 1560 atgacagctt cagccggaat ttgtgccgtt tcatcttctg ttgtagtgtt gactggagca 1620 gctaatgcgg aggatgctgc gaataaaact gcagtaaaaa ttgaaggaaa tctcatgaat 1680 tcccgatgaa gcagagagcg caggaggcgg tatttatagt gccattcccc tctctgagag 1740 acccggatgg tagtcgagtg tatcggagac agcttgatgt agactocgtg cctgccggct 1800 cctcttattg gcggacacca gtgagacacc ccggaacttg ctgtttttct gcaaaatccg 1860 gggtgaccag tgggagccta tttgcacaca cgagcgggac aocccactct ggtgaagagt 1920 gccaaagtca ttctttttcc cgttgcgggg cagccgattg catgttttag gaaaatatta 1980 cctttgctac acoctgtcag atttaccctc cacacatata tattccgtca cctccaggga 2040 ctattattcg tcgttgcgcc gccagcgggccccatag

<td>ggttggcgcc tggtatàttt gatggatgtc gcgctgcctç acgtcccggt acccaggaac</td><td> 2160</td>

<td>gcggtgggat ctcgggccca tcgaagactg tgctccagãc tgctcgccca gcaggtgttt</td><td> 2220</td>

<td>cttgatcgcc gcctctaaat tgtccgcgca tcgccggtaa catttttcca gctcggagtt</td><td> 2280</td>

<td>tgcgtttaga tacagtttct gcgatgcçaa aggagcctgc agattataac ctcggatgct</td><td> 2340</td>

<td>çtcattcagc gcttttaatt tgacctccag atagttgctg tatttctgtt cccattggct</td><td> 2400</td>

<td>gctgcgcagc ttcgtataac tcgagttatt gttgcgctct gcctcggcgt actggctcat</td><td> 2460</td>

<td>gatctggatc ttgtccgtgt cgcttttctt cgagtgtttc tcgcaaacga tgtgcacggc</td><td> 2520</td>

<td>ctgcagtgtc caatçggagt cgagctggcg ccgaaactgg cggatctgag cctccacact</td><td> 2580</td>

<td>gccctgtttc tctatccacg gcggaaccgc ctcctgccgt ttcagaatgt tgttcaagtg</td><td> 2640</td>

<td>gtactctgtg cggtcaatga aggcgttatt gccggtgaaá tctttgggaa gcggttttcc</td><td> 2700</td>

<td>tcggggaaga ttacgaaatt ccccgcgtcg ttgcgcttcc tggatctcga ggagatcgtt</td><td> 2760</td>

<td>ctccgcgtcg aggagatcgt tctccgcgtc gacaccattc cttgcggcgg cggtgctcaa</td><td> 2820</td>

<td>cggeetcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg</td><td> 2880</td>

<td>acaaacccgc gtttgagaac ttgctcaagc ttctggtaaa cgttgtagta ctctgaaaca</td><td> 2940</td>

<td>aggccctagc actctgatct gtttctcttg ggtagcggtg agtggtttat tggagttcac</td><td> 3000</td>

<td>tggtttcagç acatctgteâ tctagacaat attgttacta aatttttttg aactacaatt</td><td> 3 060</td>

<td>gttcgtaatt catctattat tatacatcct cgtcagcaat ttctggcaga cggagtttac</td><td> 3120</td>

<td>taacgtcttg agtatgaggc cgagaatcca gctctgtggc catactcagt cttgacagcc</td><td> 3180</td>

<td>tgctgatgtg gctgcgttca acgcaataag cgtgtcctcc gactccgagt tgtgctcgtt</td><td> 3240</td>

<td>atcgtcgttc tcatcctcgg aaaaatcaca cgaaagaaca tactcaccag taggctttct</td><td> 3300</td>

<td>ggtccctggg gcacggctgt ttctgacgta ttccggcgtt gataatagct cgaaagtgaa</td><td> 3360</td>

<td>cgccgagtcg cgggagtcga ccgatgccct tgagagcctt caacccagtc agctccttcc</td><td> 3420</td>

<td>ggtgggcgcg gggcatgact atcgtçgccg cacttatgac tgtcttcttt atcatgcaac</td><td> 3480</td>

<td>tcgtaggaca ggtgccggca gcgctctggg tcattttcgg cgaggaccgc tttcgctgga</td><td> 3540</td>

<td>gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc ctcgctcaag</td><td> 3600</td>

<td>ccttcgtcac tggtcccgcç accaaacgtt tcggcgagaa gcaggccatt atcgccggca</td><td> 3660</td>

<td>tggcggccga ogcgctgggc tacgtcttgc tggcgttcgc gacgcgaggc tggatggcct</td><td> 3720</td>

<td>tccccattat gattcttctc gcttccggcg gcatcgggat gcccgcgttg caggccatgc</td><td> 3780</td>

<td>tgtccaggca ggtagatgac gaccatcagg gacagcttca aggatcgctc gcggctctta</td><td> 3840</td>

ccagcctaac ttcgatcact ggaocgctga tcgtcacggc gatttatgcc gcctcggcga 3900

<td>gcacatggaa</td><td>cgggttggca</td><td colspan="3">tggattgtag gcgcccccct Ataccttgtc</td><td>tgcctccccg</td><td> 3960</td>

<td>cgttgcgtcg</td><td>cggtgcatgg</td><td>agccgggcca</td><td>cctcgaectg</td><td>aatggaagcc</td><td>ggcggcacct</td><td> 4020</td>

<td>cgctaacgga</td><td>ttcaccactc</td><td>caagaattgg</td><td>agccaatcaa</td><td>ttcttgcgga</td><td>gaactgtgaa</td><td> 4080</td>

<td>tgcgcaaacc</td><td>aacccttggc</td><td>agaacatatc</td><td>catcgcgtcc</td><td>gccatctcca</td><td>gcagccgcac</td><td> 4140</td>

<td>gcggcgcatc</td><td>gggggggggg</td><td>gggggggggg</td><td>ggggcaaáca</td><td>attcatcatt</td><td>ttttttttat</td><td> 4200</td>

<td>tctttttttt</td><td>gatttcggtt</td><td>tctttgaaat</td><td>ttttttgatt</td><td>cggtaatctc</td><td>cgaacagaag</td><td> 4260</td>

<td>gaagaacgaa</td><td>ggaaggagca</td><td>cagacttaga</td><td>ttggtatata</td><td>fcacgcatatg</td><td>tagtgttgaa</td><td> 4320</td>

<td>gaaacatgaa</td><td>attgcccagt</td><td>attcttaaçc</td><td>caactgcaca</td><td>gaa-caaaaac</td><td>ctgcaggaaa</td><td> 4380</td>

<td>cgaagataaa</td><td>tcatgtcgaa</td><td>agctacatat</td><td>aaggaacgtg</td><td>ctgctactca</td><td>tcctagtcct</td><td> 4440</td>

<td>gttgctgcca</td><td>agctatttaa</td><td>tatcatgcac</td><td>gaaaagcaaa</td><td>caaacttgtg</td><td>tgcttcattg</td><td> 4500</td>

<td>gatgttcgta</td><td>ccaccaagga</td><td>attactggag</td><td>ttagttgaag</td><td>cattaggtcc</td><td>caaaafettgt</td><td> 4560</td>

<td>ttactaaaaa</td><td>cacatgtgga</td><td>tatcttgact</td><td>gatttttcca</td><td>tggagggcac</td><td>agttaagccg</td><td> 4620</td>

<td>ctaaaggcat</td><td>tatccgccaa</td><td>gtacaatttt</td><td>ttactcttcg</td><td>aagacagaaa</td><td>atttgctgac</td><td> 4680</td>

<td>attggtaata</td><td>cagtcaaatt</td><td>gcagtactct</td><td>gcgggtgtat</td><td>acagaatags</td><td>agaatgggca</td><td> 4740</td>

<td>gacattacga</td><td>atgcacacgg</td><td>tgtggtgggc</td><td>ccaggtattg</td><td>ttagcggttt</td><td>gaagcaggcg</td><td> 4800</td>

gcagaagaag taacaaagga acctagaggc cttttgatgt tageag & att. gtcatgcaag 4860 ggctccctat ctactggaga atatactaag ggtactgttg acattgcgaa gagcgacaaa 4920 gattttgtta tcggctttat tgctcaaaga gacatgggtg gaagagatga aggttacgat 4980 tggttgatta tgacacccgg tgtgggttta gatgacaagg gagacgcatt gggtcaacag 5040 tatágaaccg tggatgatgt ggtctctaca ggafcctgaca ttattattgt tggaagagga 5100 ctatttgcaa agggaaggga tgctaaggta gagggtgaac gttacagaaa agcaggctgg 5150 gaagcatatt tgagaagatg cggccagcaa aactaaaaaa ctgtattata agtaaatgca 5220 tgtatactaa actcacaaat tagagcttca atttaattat atcagttatt acccgggaat 5280 ctcggtcgta atgattttta taatgacgaa aaaaaaaaaa 5340 ttggaaagaa aagccccccc cccccccccc cccccccccc cccccccgca gcgttgggtc ctggecacgg gtgcgcatga 5400 tcgtgctcct gtcgttgagg acccggctag gctggcgggg ttgccttact ggttagcaga 5460 atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg ctgcaaaacg tctgcgacct 5520 gagcaacaac atgaatggtc ttcggtttcc gtgtttcgta aagtctggaa acgcggaagt 5580 cagcgccctg caccattatg ttccggatct gcatcgcagg atgctgctgg ctaccctgtg 5640 gaacacctac atctgtatta acgaagcgct ggcattgacc ctgagtgatt tttctctggt 5700 cccgccgcat ccataccgcc agttgtttac cctcacaacg ttccagtaac cgggcatgtt 5760 catcatcagt aacccgtatc gtgagcatcc tctctcgttt catcggtatc attaccccca 5820 tgaacagaaa ttccccctta cacggaggca tcaagtgacc aaacaggaaa aaaccgccct 58B0 taacatggcc cgctttatca gaagccagac attaacgctt ctggagaaac tcaacgagct 5940 ggacgcggat gaaeaggcag acatctgtga atcgcttcac gaccacgctg 6000 atgagcttta

<td>ccgcagctgc</td><td>ctcgcgcgtt</td><td>tcggtgatga</td><td>cggtgaaaac <</td><td>etctgacaca *</td><td>tgcagctccc</td><td> 6060</td>

<td>ggagacggtc</td><td>acagcttgtc</td><td>tgtaagcgga</td><td>tgccgggagc i</td><td>agacaagccc <</td><td>gtcagggcgc</td><td> 6120</td>

<td>gtcagcgggt</td><td>gttggcgggt</td><td>gtcggggcgc</td><td>agccatgacc</td><td>cagtcacgta </td><td>gcgatagcgg</td><td> 6180</td>

<td>agtgtatact</td><td>ggcttaacta</td><td>tgcggcatca</td><td>gagcagattg</td><td>tactgagagt</td><td>gcaccatatg</td><td> 6240</td>

<td colspan="2">cggtgtgaaâ 'taccgcacag</td><td>atgcgtaagg</td><td>agaaaatacc</td><td>gcatcaggcg</td><td>ctcttccgct</td><td> 6300</td>

<td>tcctcgctca</td><td>ctgactcgct</td><td>gcgctcggtc</td><td>gttcggctgc</td><td>ggcgagcggt</td><td>atcagctcac</td><td> 6360</td>

<td>tcaaaggcgg</td><td>tàataeggtt</td><td>atccacagaa</td><td>tcaggggata</td><td>acgcaggaaa</td><td>gaacatgtga</td><td> 6420</td>

<td>gcaaaaggcc</td><td>agcaaaaggc</td><td>caggaaccgt</td><td>aaaaaggccg</td><td>cgttgctggc</td><td>gtttttccat</td><td> 64 80</td>

<td>aggctccgcc</td><td>cccctgacga</td><td>gcatcacaaa</td><td>aatcgacgct</td><td>caagtcagag</td><td>gtggcgaaac</td><td> 6540</td>

<td>ccgacaggac</td><td>tataaagata</td><td>ccaggegttt</td><td>eccçctggaa</td><td>gctccctcgt</td><td>gcgctctcct</td><td> 6600</td>

<td>gttccgaccc</td><td>tgccgcttac</td><td>cggatacctg</td><td>tocgcctttc</td><td>tcccttcggg</td><td>aagcgtggcg</td><td> 6660</td>

<td>ctttctcata</td><td>gctcaogctg</td><td>taggtatctc</td><td>àgttcggtgt</td><td>aggtcgttcg</td><td>ctccaagctg</td><td> 6720</td>

<td>ggctgtgtgc</td><td>acgaaccccc</td><td>cgttcagccc</td><td colspan="2">gaccgctgcg ccttatccgg</td><td>taactatcgt</td><td> 6780</td>

<td>cttgagtcca</td><td>acccggtaag</td><td>acacgactta</td><td>tcgccactgg</td><td>cagcagceac</td><td>tggtaacagg</td><td> 6840</td>

<td>attagcagag</td><td>cgaggtatgt</td><td>aggcggtgct</td><td>acagagttct</td><td>tgaagtggtg</td><td>gcctaactac</td><td> 6900</td>

<td>ggctacacta</td><td>gaaggacagt</td><td>atttggtatc</td><td>tgcgctctgc</td><td>tgaagccagt</td><td>taccttcgga</td><td> 6960</td>

<td>aaaagagttg</td><td>gtagctcttg</td><td>atccggcaaa</td><td>caaaccaccg</td><td>ctggtagcgg</td><td>tggttttttt</td><td> 7020</td>

<td>gtttgcaagc</td><td>agcagattac</td><td>gcgcagaaaa</td><td>aaaagatctc</td><td>aagaagatcc</td><td>tttgatcttt</td><td> 7080</td>

<td>tctacggggt</td><td>ctgacgctca</td><td>gtggaacgaa</td><td>aactcacgtt</td><td>aagggatttt</td><td>ggtcatgaga</td><td> 7140</td>

<td>ttatcaaaaa</td><td>ggatcttcac</td><td>ctagatcctt</td><td>ttaaattaaa</td><td>aatgaagttt</td><td>taaatcaatc</td><td> 7200</td>

<td>taaagtatat</td><td>atgagtaaac</td><td>ttggtctgac</td><td>agttaccaat</td><td>gcttaatcag</td><td>tgaggcacct</td><td> 7260</td>

<td>atctcagcga</td><td>tctgtctatt</td><td>tcgttcatcc</td><td>atagttgcct</td><td>gactocccgt</td><td>cgtgtagata</td><td> 7320</td>

<td>actacgatac</td><td>gggagggctt</td><td>accafcctggc</td><td>cccagtgctg</td><td>caatgatacc</td><td>gcgagaccca</td><td> 7380</td>

<td>cgctcaccgg</td><td>ctccagattt</td><td>àtcagcaata</td><td>aaccagccag</td><td>ccggaagggc</td><td>cgagcgcaga</td><td> 7440</td>

<td>agtggtcctg</td><td>caactttatc</td><td>cgcctccatc</td><td>cagtctatta</td><td>attgttgccg</td><td>ggaagctaga</td><td> 7500</td>

<td>gtaagtagtt</td><td>cgccagttaa</td><td>tagtttgcgc</td><td>aacgttgttg</td><td>ocattgctgc</td><td>aggcatcgtg</td><td> 7560</td>

<td>gtgtcacgct</td><td>cgtcgtttgg</td><td>tatggcttca</td><td>ttcagctccg</td><td>gttcccaacg</td><td>atcaaggcga</td><td> 7620</td>

<td>gttacatgat</td><td>cccccatgtt</td><td>gtgcaaaaaa</td><td>gcggttagct</td><td>ccttcggtce</td><td>tccgatcgtt</td><td> 7680</td>

<td>gtcagaagta '</td><td>agttggccgc</td><td>agtgttatca</td><td>ctcatggtta</td><td>tggcagcact</td><td>gcataattct</td><td> 7740</td>

<td>cttactgtca</td><td>tgccatccgt</td><td>aagatgcttt</td><td>-tctgtgactg</td><td>gtgagtactc</td><td>aaceaagtca</td><td> 7800</td>

<td>ttctgagaat</td><td>agtgtatgcg</td><td>gcgaccaagt</td><td>tgctcttgcc</td><td>cggcgtcaac</td><td>acgggataat</td><td> 7860</td>

<td>accgcgccac</td><td>atagcagaac</td><td>tttaaaagtg</td><td>ctcatcattg</td><td>gaaaacgttc</td><td>ttcggggcga</td><td> 7920</td>

<td>aaactetcaa</td><td>ggatcttacc</td><td>gctgttgaga</td><td>tccagttcga</td><td>tgtaaeccac</td><td>tcgtgcaccc</td><td> 7980</td>

<td>aactgatctt</td><td>cagcatcttt</td><td>tactttcacc</td><td>agcgtttctg</td><td>ggtgagcaaa</td><td>aacaggaagg</td><td> 8040</td>

<td>caaaatgccg</td><td>caaaaaaggg</td><td>aataagggcg</td><td>acacggaaat</td><td>gttgaatact</td><td>catactcttc</td><td> 8100</td>

<td>ctttttcaat</td><td>attattgaag</td><td>catttatcag</td><td>ggttattgtc</td><td>tcatgagcgg</td><td>attackattt</td><td> 8160</td>

<td>gaatgtattt</td><td>agaaaaa taa</td><td>acaaataggg</td><td>gttccgcgca</td><td>catttccccg</td><td>aaaagtgcca</td><td> 8220</td>

<td>cctgacgtct</td><td>aagaaaccat</td><td>tattatcatg</td><td>acattaacct</td><td>ataaaaatag</td><td>gcgtatcacg</td><td> 8280</td>

<td>aggccctttc</td><td>gtcttcaa</td><td></td><td></td><td></td><td></td><td> 8298</td>

<210> 25 <211> 8695 <212> DNA <213> pMPT-M? Al-a2-H6 vector <220>

<221> MISCELLANEOUS <222> (2103). . (2103) <223> Ν is any nueleotideo <400> 25

<td>ggtacectgc</td><td>tcaatctccg- gaatggtgat ctgatcgttc ctgaaaacct cgaeattggc</td><td> 60</td>

<td>tccctcctga</td><td>cacaggtact cgtacaggtt ccaggtaaac gagtçgtagt tgtcgatcat</td><td> 120</td>

<td>gacaacgttc</td><td>ttagaag-cgg ccggcatttt gaaggtgact aatagcctaa gaaaatattt</td><td> 180</td>

<td>aatttaattt</td><td>tcattaaatt ttcctatact egctattbca gcttttcatc tcatcacttc</td><td> 240</td>

<td>ataaacgata</td><td>taaaccagaa aaagaactat tttcaaacac gcttctcaaa agcggtatgt</td><td> 300</td>

<td>ccttccacgt</td><td>ctccttagaa tctggcaagt ccgcgagggg gatccagatc tgatagcttt</td><td> 360</td>

<td>caatcaatga</td><td>atcgaaaatg tcattaaaat agtatataaa ttgaaactaa gtcataaagc</td><td> 420</td>

<td>tataaaaaga</td><td>aaatttattt aaatgcaaga ctttaaagta aattcactta agccttggca</td><td> 480</td>

<td>acgtgttcaa</td><td>ccaagtcgag atcgttaatt aactattagt gatggtggtg atggtgtctg</td><td> 540</td>

<td>ccctcgatca</td><td>cctgccactc tgttgtagae agcagcagcg agctaagctc tgatctatcc</td><td> 600</td>

<td>ctgtcctcca</td><td>agtcacaaeg ctctcctcga gtccaattgc atgcggcttc gaacctgtgc</td><td> 660</td>

tccacgcccc ccacgtacat cctaaccttg aagatggtga agttgacagt gcaggggtag 720 tatatgggta tgccagagcc, atgaaccata cacctaggtg tcagccaggg cçcagaaccg 780 catctggcgt aagtggcctc ggggtgcttc cgaaaacagt cagtggggca ggtcaaggtg 840 ttgttgccgg cccccccgat gttgcacggg gggcccccac acgtcttggt gaacccagtg 900 ccattcatcc atgtacagcc gaaccagttg cctcgcggcg gccgcgtgtt gttgagaatc 960 agcacatccg agtcgttcgc cccccagtta tacgtgggga caccaaaccg 1020 atcggtcgtc

<td>cccaccacaa</td><td>cagggctcgg</td><td>agtgaagcaa</td><td colspan="2">tacactggae cgcacacctg;</td><td>agacgcgggt</td><td> 1080</td>

<td>acaataccac</td><td>acggtcgagg</td><td>cgcgtagtgc</td><td>cagcagtagg -</td><td>gcctctggtc i</td><td>cgagctgtta</td><td> 1140</td>

<td>ggctcagtgt</td><td>aagtgagggg</td><td>accccacccc</td><td>tgagcgaact</td><td>tgtcgatgga </td><td>gcgacagctg</td><td> 1200</td>

<td>gccaagcgct</td><td>ctgggeatcc</td><td>agacgag-ttg</td><td>aatttgtgtt</td><td>tgtagaatag</td><td>tgcggcaaag</td><td> 1260</td>

<td>aacoctgttt</td><td>ggagggagtc</td><td>gttgcagttc</td><td>agggcagtcc</td><td>tgttgatgtg</td><td>ccaactgccg</td><td> 1320</td>

<td>ttggtgttta</td><td>cgagctggat</td><td>tttctgagce</td><td>gacccggggc</td><td>taaagaggga</td><td>cacaaggccc</td><td> 1380</td>

<td>ctggtatcgg</td><td>aggctgctgc</td><td>ccctcctgac</td><td>acgcgggtat</td><td>gccttttatc</td><td>tagagatacc</td><td> 1440</td>

<td>ccttcttctt</td><td>tagcagcaat</td><td>gctggcaata</td><td>gtagtattta</td><td>taaacaataa</td><td>cccgttattt</td><td> 1500</td>

<td>gtgctgttgg</td><td>aaaâtggcaa</td><td>aacagcaaca</td><td>tcgaaatccc</td><td>cttctaaatc</td><td>tgagtaaccg</td><td> 1560</td>

<td>atgacagett</td><td>cagecggaat</td><td></td><td></td><td>s '-"-The"··""</td><td>gactggagea</td><td> 1620</td>

<td>gctaatgcgg</td><td>aggatgctgc</td><td>gaataaaact</td><td>gcagtaaaaa</td><td>ttgaaggaaa</td><td>tctcatgaat</td><td> 1680</td>

<td>tcgtttttgt</td><td>actttagatt</td><td>gatgtcaoca</td><td>cegtgcactg</td><td>gcag-cagtat</td><td>tfcatagatgg</td><td> 1740</td>

<td>accgtgtggg</td><td>gacggttggg</td><td>tacacttagc</td><td>ggcagcgctg</td><td>accccatctg</td><td>tgatcaagta</td><td> 1800</td>

<td>gggcaaaaac</td><td>tgggsatgte</td><td>ggagtcgctg</td><td>cacggtagca</td><td>taagaattta</td><td>ctttctggcc</td><td> 1860</td>

<td>ggttcacccg</td><td>catttgcact</td><td>gtggagaaac</td><td>agcctgtccg</td><td>acaccccacc</td><td>agttgccaca</td><td> 1920</td>

<td>.tcggccctct</td><td>gctgctctgg</td><td>tgatçttctg</td><td>gtagcaggca</td><td>cagacagcag</td><td>tgggtagcgc</td><td> 1980</td>

<td>cgtccggtta</td><td>ggcaaggtca</td><td>cgttgtaggc</td><td>taccccagca</td><td>aacagagcct</td><td>cacatgacac</td><td> 2040</td>

<td>catccagctg</td><td>cgtcctcgaa</td><td>gcgaaaagtt</td><td>cggttgcggc</td><td>tgcagaaccc</td><td>cctcagttgc</td><td> 2100</td>

<td>canattcaca</td><td>agttttacgc</td><td>gacggctaaa</td><td>gcgagtgggt</td><td>tttaaaaact</td><td>tgcggtgcaa</td><td> 2160</td>

<td>ggatgcatgc</td><td>ggcaacaatt</td><td>aattggtgca</td><td>tccagcacag</td><td>caagcccagt</td><td>ctcgagatgt</td><td> 2220</td>

<td>ccagtcgcta</td><td>cagagtggag</td><td>tacgcactca</td><td>aggaacaccg</td><td>tcgagatggc</td><td>ctcatagaat</td><td> 2280</td>

<td>ggatcaaggg</td><td>cctgctggcc</td><td>acgccgttcg</td><td>tcctgtacgc</td><td>ggtgaagagc</td><td>aacggcatct</td><td> 2340</td>

<td>ctgcagtgga</td><td>cgacctcatg</td><td>gtaaactctg</td><td>aggcaaaacg</td><td>ccgctacgcg</td><td>gaaatcttcc</td><td> 2400</td>

<td>acgacctcga</td><td>actcctcatc</td><td>gacgacaaca</td><td>ttgaaatgac</td><td>caaagccggc</td><td>acccccgaat</td><td> 2460</td>

<td>tgtctcggct</td><td>cgtgcagctg</td><td>gtfcocgagcg</td><td>ttggcagctt</td><td>ctfccacgaga</td><td>. ctgcctctgg</td><td> 2520</td>

aaaaggcctt ctacatcgag gacgagcgcc gcgccatcag caaacgccgg cttgtggcoc 2580 cctcgttcaa cgacgtccgg ctcattctca acacggccca gctgttggag atgtcgcggt 2640 tcttccatte caaaaccatc cgagatcgca agctgcagct cattacattc gatggtgaca 2700 fccacactgta cgacgacggc aaaaatttcg atgccgagtc gcccatcctg ccccacctca 2760 tcaaactaat ggecaaggac ctctatgtgg gtatcgtcac cgcggccggc tacagcgacg 2820 gaacaagtac tacgagcgcc tcaagggcct catcgacgcc gtccagacgt cccegctgct 2β80 cacaggccac cagaaagaga acctgttcat tatgggcggc gaggcaaact 2940 acctcttccg gtacagtaac gaggagcaga gattacgctt ctactccaaa gacagatggc tgctcgagaa 3000 eatgctgaat tggtccgagg aggacattea tctgacactg gactttgcgc aggacgttct 3060 aaacgacctc gttcacaggggggggggggggggggggggggggggggggtg

<td>cggcctggtt</td><td>ccattacegg</td><td>gccacaagct.</td><td>gatccgcgag</td><td>cagctcgagg</td><td>agatcgttct</td><td> 3180</td>

<td>ccgcgtcgac</td><td>accattcctt</td><td>gcggcggcgg</td><td>tgctcaacgg</td><td>cctcaaccta</td><td>ctactgggct</td><td> 3240</td>

<td>gcttcctaat</td><td>gcaggagtcg</td><td>cataagggag</td><td>agcgtcgact</td><td>cccgcgactc</td><td>ggcgttcact</td><td> 3300</td>

<td>ttcgagctat</td><td>tatcaacgcc</td><td>ggaatacgtc</td><td>agaaacagcc</td><td>gtgccccagg</td><td>gaccagaaag</td><td> 3360</td>

<td>cctactggtg</td><td>agtatgttct</td><td>ttcgtgtgat</td><td>ttttccgagg</td><td>atgagaacga</td><td>cgataacgag</td><td> 3420</td>

<td>cacaactcgg</td><td>agtcggagga</td><td>caegcttatt</td><td>gcgttgaaeg</td><td>cagecaeate</td><td>“3 ^ 33« - ^^ u.</td><td> 3430</td>

<td>caagactgag</td><td>tatggccaca</td><td>gagctggatt</td><td>ctcggcctca</td><td>tactcaagac</td><td>gttagtaaac</td><td> 3540</td>

<td>tccgtctgcc</td><td>agaaattgct</td><td>gacgaggatg</td><td>tataataata</td><td>gatgaattac</td><td>gaacaattgt</td><td> 3600</td>

<td>agttcaaaaa</td><td>aatttagtaa</td><td>caatattgtc</td><td>tagatgacag</td><td>atgtgctgaa</td><td>accagtgaac</td><td> 3660</td>

<td>tccaataaac</td><td>cactcaocgc</td><td>tacccaagag</td><td>aaacagatca</td><td>gagtgctagg</td><td>gccttgtttc</td><td> 3720</td>

<td>agagtactac</td><td>aacgtttacc</td><td>agaagcttga</td><td>gcaagttctc</td><td>aaacgcgggt</td><td>ttgtcgaccg</td><td> 3780</td>

<td>atgcccttga</td><td>gagccttcaa</td><td>cccagtcagc</td><td>tccttccggt</td><td>gggcgcgggg</td><td>catgactatc</td><td>3B40</td>

<td>gtcgccgcac</td><td>ttatgactgt</td><td>cttctttatc</td><td>atgcaactcg</td><td>taggacaggt</td><td>gccggcagcg</td><td> 3900</td>

<td>ctctgggtca</td><td>ttttcggcga</td><td>ggaecgcttt</td><td>cgetggagcg</td><td>cgacgatgat</td><td>cggcctgtcg</td><td> 3960</td>

<td>cttgcggtat</td><td>tcggaatctt</td><td>gcacgccctc</td><td>gctcaagcct</td><td>tcgtcactgg</td><td>tcccgccacc</td><td> 4020</td>

<td>aaacgtttcg</td><td>gcgagaagca</td><td>ggccattatc</td><td>gccggcatgg</td><td>cggccgacgc</td><td>gctgggctac</td><td> 4080</td>

<td>gtcttgctgg</td><td>cgttcgcgac</td><td>gcgaggctgg</td><td>afcggocttcc</td><td>ccattatgat</td><td>tcttctcgct</td><td> 4140</td>

<td>tccggcggca</td><td>fccgggatgcc</td><td>cgcgttgcag</td><td>gccatgctgt</td><td>ccaagcaggt</td><td>agatgacgac</td><td> 4200</td>

<td>catcagggac</td><td>agcttcaagg</td><td>atcgctcgcg</td><td>gctcttacca</td><td>gcctaacttc</td><td>gatcactgga</td><td> 4260</td>

<td>ccgctgatcg</td><td>tcacggcgat</td><td>ttatgccgcc</td><td>tcggcgagca</td><td>catggaacgg</td><td>gttggcatgg</td><td> 4320</td>

, ι

<td>àttgtaggcg ccgccctàta ccttgtctgc ctccccgcgt tgcgtcgcgg tgcatggagc</td><td> 4380</td>

<td>cgggccacct cgacctgaat ggaagccggc ggcacct-cgc taacggattc accactccaa</td><td> 4440</td>

<td>gaattggagc caatcaattc ttgcggagaa ctgtgaatgc gcaaaccaac ccttggcaga</td><td> 4500</td>

<td>acatatccat cgcgtccgcc atctccagca gccgca-cgcg gcgcatcggg gggggggggg</td><td> 4560</td>

<td>gggggggggg gcaaacaatt catcattttt tttttattct tttttttgat ttcggtttct</td><td> 4620</td>

<td>ttgaaatttt tttgattcgg taatctccga acagaággaa gaacgaagga aggagcacag</td><td> 4680</td>

<td>acttagattg gtatatatac gcatatgtag tgttgaagaa acatgaaatt gcccagtatt</td><td> 4740</td>

<td>cttaacccaa ctgcacagaa caaaaacctg caggaaacga agataaatca tgtcgaaagc</td><td> 4800</td>

<td>tacatataag, gaacgtgctg ctactcatcc tagtcctgtt gctgccaagc tatttaatat</td><td> 4860</td>

<td>catgcacgaa aagcaaacaa acttgtgtgc ttcattggat gttcgtacca ccaaggaatt</td><td> 4920</td>

<td>actggagtta gttgaagcat taggtcccaa aatttgttta ctaaaaacac atgtggatat</td><td> 4980</td>

<td>cttgactgat ttttccatgg agggcacagt taagccgcta aaggcattat ccgccaagta</td><td> 5040</td>

<td>caatttttta ctcttcgasg acagaaaatt tgctgacatt ggtaatacag tcaaattgca</td><td> 5100</td>

<td>gtactctgcg ggtgtataca gaatagcaga atgggcagac attacgaatg cacacggtgt</td><td> 5160</td>

<td>ggtgggccca ggtattgtta gcggtttgaa gcaggcggca gaagaagtaa caaaggaacc</td><td> 5220</td>

<td>Lagaggcctt ttgatgttag cagaattgtc atgçaagggc fceçctatcta ctggagaata</td><td>cna OR</td>

<td>tactaagggt actgttgaca ttgcgaagag cgacaaagat tttgttatcg gctttattgc</td><td> 5340</td>

<td>tcaaagagac atgggtggaa'gagatgaagg ttacgattgg ttgattatga cacccggtgt</td><td> 5400</td>

<td>gggtttagat gacaagggag acgcattggg tcaacagtat agaaccgtgg atgatgtggt</td><td> 5460</td>

<td>ctctacagga tctgaçatta ttattgttgg aagaggacta tttgcaaagg gaagggatgc</td><td> 5520</td>

<td>taaggtagag ggtgaacgtt acagaaaagc aggctgggaa gcatatttga gaagatgcgg</td><td> 5580</td>

<td>ccagcaaaac taaaaaactg tattataagt aaatgcatgt Atactaaa-ct cacaaattag</td><td> 5640</td>

<td>agcttcaatt taattatatc agttattacc cgggaatctc ggtcgtaatg atttttataa</td><td> 5700</td>

<td>tgacgaaaaa aaaaaaattg gaaagaaaag cccccccccc cccccccccc cccccccccc</td><td> 5760</td>

<td>ccccgçagcg ttgggtcctg gccacgggtg cgcatgatcg tgctcctgtc gttgaggacc</td><td> 5820</td>

<td>cggctaggct ggcggggttg ccttactggt tagcagaatg aatcaccgat acgcgagcga</td><td> 5880</td>

<td>acgtgaagcg actgctgctg caaaacgtct gcgacctgag caacaacatg aatggtcttc</td><td> 5940</td>

<td>ggtttccgtg tttcgtaaag tctggaaacg cggaagtcag cgocctgcac cattatgttc</td><td> 6000</td>

<td>cggatctgca tcgcaggatg ctgctggcta ccctgtggaa cacctacatc tgtattaacg</td><td> 6060</td>

<td>aagcgctggc attgaccctg agtgattttt ctctggtccc gccgcatcca taccgccagt</td><td> 6120</td>

<td>tgtttaccct cacaacgttc cagtaaccgg gcatgttcat catcagtaac ccgtatcgtg</td><td> 6180</td>

<td>agcatcctct</td><td>ctcgtttcat</td><td>cggtatcatt</td><td>acccccatga</td><td>acagaaattc</td><td>ccccttacac</td><td> 6240</td>

<td>ggaggcatca</td><td>agtgaccaaa</td><td>caggaaaaaa</td><td>ccgcccttaa</td><td>catggcocgc</td><td>tttatcagaa</td><td> 6300</td>

<td>gccagacatt</td><td>aacgcttctg</td><td>gagaaactca</td><td>acgagctgga</td><td>cgcggatgaa</td><td>caggcagaca</td><td> 6360</td>

<td>tctgtgaatc</td><td>gcttcacgac</td><td>cacg-ctgatg</td><td>agctttaccg</td><td>eagetgcctc</td><td>gcgcgtttcg</td><td> €420</td>

<td>gtgatgacgg</td><td>tgaaaacctc</td><td>tgacacatgc</td><td>agctcccgga</td><td>gacggtcaca</td><td>gcttgtctgt</td><td> 6480</td>

<td>aagcggatgc</td><td>cgggagcaga</td><td>caagcccgtc</td><td>agggcgcgtc</td><td>agcgggtçtt</td><td>ggegggtgtc</td><td> 6540</td>

<td>ggçgcgcagc</td><td>catgacccag</td><td>tcacgtagcg</td><td>atagcggagt</td><td>gtatactggc</td><td>ttaaetatgc</td><td> 6600</td>

<td>ggcatcagag</td><td>cagattgtac</td><td>tgagagtgca</td><td>ccatatgcgg</td><td>tgtgaaatac</td><td>cgcacagatg</td><td> 6660</td>

<td>cgtaaggaga</td><td>aaataccgca</td><td>tcaggcgctc</td><td>ttccgcttcc</td><td>tcgctcactg</td><td>actcgctgcg</td><td> 6720</td>

<td>ctcggtcgtt</td><td>cggctgcggc</td><td>Sagcggtatc</td><td>agctcactca</td><td>aaggcggtaa</td><td>tacggttatc</td><td> 6780</td>

<td>oacagaatca</td><td>ggggataacg</td><td>caggaaagaa</td><td>catgtgagca</td><td>aaaggccagc</td><td>aaaaggccag</td><td> 6840</td>

<td>gaaccgtaaa</td><td>aaggccgcgt</td><td>tgctggcgtt</td><td>tttccatagg</td><td>otccgccccc</td><td>ctgacgagça</td><td> 6900</td>

<td>tcacaaaaat</td><td>cgacgctcaa</td><td>gtcagaggtg</td><td>gcgaaacccg</td><td>acaggactat</td><td>aaagatacca</td><td> 6960</td>

<td>ggcgtttccc</td><td>cctggaagct</td><td>ccctcgtgcg</td><td>cfcctcctgtt</td><td>ccgaccctgc</td><td>cgcttaccgg</td><td> 7020</td>

<td>Atacctgtcc</td><td>gcetttctcc</td><td>cttcgggaag</td><td>cgtggcgctt</td><td>tctcatagct</td><td>cacgctgtag</td><td> 7080</td>

<td>gtatctcagt</td><td>tcggtgtagg</td><td>tcgttcgctc</td><td>caagctgggc</td><td>tgtgtgcacg</td><td></td><td> 7140</td>

<td>tcagcccgac</td><td>cgctgcgcct</td><td>tatccggtaa</td><td>ctatcgtctt</td><td>gagtccaacc</td><td>cggtaagaca</td><td> 7200</td>

<td>cgacttatcg</td><td>ccactggcag</td><td>cagccactgg</td><td>taacaggatt</td><td>agcagagcga</td><td>ggtatgtagg</td><td> 7260</td>

cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt 7320 tggtatctge gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc 73B0 cggcaaacaa accaccgctg gtagcggtgg ttfcttttgtt tgcaagcagc agattacgcg 7440 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg 7500 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaceta 7S60 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg 7620 gtctgacagt taccaatgct taatcagtga ggcacctatc fccagcgatct 7680 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc 7740 atctggcocc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc 7800 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc 7860 ctccatccag tctattaatt gttgccggga agctagagta agtagttegc cagttaatag 7920 tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat 7980 ggcttcattc agctocggtt cccaacgatc aaggcgagtt acatgatcoc ccatgttgtg 8040 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt 8100 tggccgcagt gttatcactc atggttatgg cagcaçtgca taattctctt actgteatgc catccgtaag 8160 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg 8220 accgagttgc tcttgcccgg cgbcaacacg ggataatacc gcgccacata gcagaacttt 8280 aaaagtgctc atcattggaa aacgttcttc ggggegaaaa ctctcaagga tcttaccgct 8340 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac 8400 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat 8460 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt 8520 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca 8580 aataggggtt ôcgcgcacat ttccccgaaa agtgçcacct gacgtctaag aaaccattat 8640 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc ttcaa 8695 <21 0> 26 <211> 36 <212> DNA <213> primer or synthetic probe <400> 26 agtcactctt caaggcatac ccgcgtgtca ggaggg 36 <210> 27 <211> 39 <212> DNA <213> primer or synthetic probe <400> 27

<td>agtcactctt cacagggatc cttagtgatg gtggtgatg</td><td> 39</td>

<td><210> 28 <211> 4190 <212> DNA <213> pMF30 vector <400> 28 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca</td><td> 60</td>

<td>cgacaggttt cccgactgga aagcgggcag tgagegcaac gcaattaatg tgagttagct</td><td> 120</td>

<td>cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat</td><td> 180</td>

<td>tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattaog ccaagcttgc</td><td> 240</td>

<td>atgcctgcag ttgattgcag atgccagatc ccgaaagaac agaggacgga gcgtaaactt</td><td> 300</td>

<td>gtggcattcc accagaaatt gatacagata agcttccgga gtcaccagct aaaacggaat</td><td> 360</td>

tgcaagaaat aatatcgata actttatcac cactagaata gccggtgttg ctgacagtaa 420 tatcetgtga cccgtttgaa cctaaattat taaaaatgga aatcaattga ttagcatcgc 48o tacccttcct agtgtctgtagtgtgtgtgtgtgtgtgtgtg ctgattagct 600 cgtagtattc tgaatcggta ttatatccac ttaaccataa agcttctcta ttggcaggat 660 cggaaccacc attgagacct tgttcttggc cataataaat aattgggata ccatcaccca 720 aaattataaa agccatgtca ttcttaatca aggatgtgtc tgsggtaact aatggaaatc 780 taacttggtc atggttttca ataaagtttc ccaacaaaga gacgtccgaa caagatgact 840 gtaacgtgga gatcattgaa gttaactcac tggaagtcgc cgaagtatca 900 ctgaagaatc

<td>tatatactgg atagtataat ggatagttgg taactccttt catataattc tgatatggac</td><td> 960</td>

<td>sagtataagt tggatctcct tgataaactt cacctaagtt ataaacacca gaagcgtcct</td><td> 1020</td>

<td>caaacttcgt taatgaagcg gtatctacgt gctttgcact atcaattctt aaaccatcga</td><td> 1080</td>

<td>ttgaatagtt ttgaacaaaa tctgacaccc aagtttgaaa tactcctata acttcattat</td><td> 1140</td>

<td>cctcggtact taaatctgga agggagactt cagtatcacc ttcccaacaa tcttcaacat</td><td> 1200</td>

<td>tggtttgatc attataattt gtaat-caaac aataatcgtg gaagtaagat tgttgattga</td><td> 1260</td>

<td>atggagtgaa actagaataa tctacgcttg aaccatctcc gttccaagca taatggttgt</td><td> 1320</td>

<td>aaacaacgtc gaccatcaat aacatgcttc tggaatgcaa ttcgctagct aattgtttca</td><td> 1380</td>

<td>attcatcagc ggtaccaaaa ttagtgttca attcatcaat atttttcatc caataaccat</td><td> 1440</td>

<td>ggtaagcata accataagca gtattgtcag gaatttgctc aacaactggg gagatccaga</td><td> 1500</td>

<td>tcgcagtgaa acccatacct tgaatataat ccaacttgtc gataatccct ttataagatc</td><td> 1560</td>

<td>caccacagta cttgcgatca ctcactaaac agtcagctgt ggfccgagoca tcagatctgg</td><td> 1620</td>

<td>caaacctatc agtaacgatt tgataaatcg attggtcttt ccat.ttatca gctgacgagc</td><td> 1680</td>

<td>taacatccct cttgtcaaaa ataatcggtt gagcagatac caatcttgag aatgctaaaa</td><td> 1740</td>

<td>ttgctgcaac aactttactt gtaaatcctt cagttgaaaa tctcattgaa ttcactggcc</td><td> 1800</td>

<td>gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgocttgca</td><td> 1860</td>

<td>gcacatcccc ctttcgocag ctggcgtaat agcgaagagg cccgcaecga tcgcccttcc</td><td> 1920</td>

<td>caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat</td><td> 1980</td>

<td>ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca</td><td> 2040</td>

<td>tagttaagcc aaccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg</td><td> 2100</td>

<td>ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg</td><td> 2160</td>

<td>ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta</td><td> 2220</td>

taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat 2280 gtgcgcggaa cccctatttg tttatttttc taaataqatt caaatatgta tccgctcatg 2340 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa 2400 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac 2460 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac 2520 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgceccga agaacgtttt 2580 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg 2640 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattetcaga atgacttggt tgagtactca 2700

<td>ccagtcacag</td><td>aaaagcatct</td><td>tacggatggc</td><td>atgacagtaa</td><td>gagaattatg</td><td>cagtgctgcc</td><td> 2760</td>

<td>ataaccatga</td><td>gtgataacac</td><td>tgcggccaac</td><td>ttacttctgà</td><td>caacgatçgg</td><td>aggaccgaag</td><td> 2820</td>

<td>gagctaaccg</td><td>cttttttgca</td><td>caacatgggg</td><td>gatcatgtaa</td><td>ctcgccttga</td><td>tcgttgggaa</td><td> 2880</td>

<td>ccggagctga</td><td>atgaagccat</td><td>accaaacgac</td><td>gagcgtgaca</td><td>ocacgatgcc</td><td>tgtagcaatg</td><td> 2940</td>

<td>gcaacaacgt</td><td>tgcgcaaact</td><td>attaactggc</td><td>gaactactta</td><td>ctctagcttc</td><td>ccggcaacaa</td><td> 3000</td>

<td>ttaatagact</td><td>ggatggaggc</td><td>ggataaagtt</td><td>gcaggaccac</td><td>ttctgcgctc</td><td>ggcccttccg</td><td> 3060</td>

<td>gctggctggt</td><td>ttattgctga</td><td colspan="2">taaatctgga gcçggtgagc</td><td>gtgggtctcg</td><td>cggtatcatt</td><td> 3120</td>

<td>gcagcactgg</td><td>ggccagatgg</td><td>taagcçcteç '</td><td></td><td>ttatctàcac</td><td>gacggyyagt</td><td> 3160</td>

<td>caggcaacta</td><td>tggatgaacg</td><td>aaatagàcag</td><td>atcgctgaga</td><td>taggtgcctc</td><td>actgattaag</td><td> 3240</td>

<td>cattggtaac</td><td>tgtcagacca</td><td>agtttactca</td><td>tatatacttt</td><td>agattgattt</td><td>aaaacttcat</td><td> 3300</td>

<td>ttttaattta</td><td>aaaggatcta</td><td>ggtgaagatc</td><td>ctttttgata</td><td>atctcatgac</td><td>caaaatccct</td><td> 3360</td>

<td>taacgtgagt</td><td>tttcgttcca</td><td>ctgagcgtca</td><td>gaccccgtag</td><td>aaaagatcaa</td><td>aggatcttet</td><td> 3420</td>

<td>tgagatcctt</td><td>tttttctgcg</td><td>cgtaatctgc</td><td>tgcttgcaaa</td><td>caaaaaaacc</td><td>accgctacca</td><td> 3480</td>

<td>gcggtggttt</td><td>gtttgccgga</td><td>tcaagagcta</td><td>• ccaactcttt</td><td>ttccgaaggt</td><td>aactggcttc</td><td> 3540</td>

<td>agcagagcgc</td><td>agataccaaa</td><td>tactgtcctt</td><td>otagtgtagc</td><td>cgtagttagg</td><td>• ccaccacttc</td><td> 3600</td>

<td>aagaactctg</td><td>tagcaocgcc</td><td>tacatacctc</td><td>gctctgctaa</td><td>tcctgttacc</td><td>agtggctgct</td><td> 3660</td>

<td>gccagtggcg</td><td>ataagtcgtg</td><td>tcttaccggg</td><td>ttggactcaa</td><td>gacgatagtt</td><td>accggataag</td><td> 3720</td>

<td>gcgcagcggt</td><td>egggctgaac</td><td>ggggggttcg</td><td>tgcacacagc</td><td>ccagcttgga</td><td>gcgaacgacc</td><td> 3780</td>

<td>tacaccgaac</td><td>tgagatacct</td><td>acagcgtgag</td><td>ctatgagaaa</td><td>gcgccacgct</td><td>tcccgaaggg</td><td> 3840</td>

<td>agaaaggcgg</td><td>acaggtat.cc</td><td>ggtaagcggc</td><td>agggtcggaa</td><td>caggagagcg</td><td>cacgagggag</td><td> 3900</td>

<td>cttoeagggg</td><td>gaaacgcctg</td><td>gtatctttat</td><td>agtcctgtcg</td><td>ggtttcgcca</td><td>cctctgactt</td><td> 3960</td>

<td>gagcgtcgat</td><td>ttttgtgatg</td><td>ctcgtcaggg</td><td>gggcggagoc</td><td>tatggaaaaa</td><td>cgccagcaac</td><td> 4020</td>

gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg 4080 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc 4140 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga 4190.

<210> 29 <211> 38 <212> DNA <213> primer or synthetic probe <400> 29 agtcactctt cacctcttgt caaaaataat cggttgag 38 <210> 30 <211> 52 <212> DNA <213> primer or synthetic probe <400> 30 tgcttectac cactagcagc actaggacat aecegcgtgt caggaggggc ag 52 <210> 31 <211> 57 <212> DNA <213> primer or synthetic probe <400> 31 tãgtacfcagt attagtaggc ttcgcatgga attcactggc c227 <213> vector pFMPT-CL-E2-H6 <400> 32

<td>ggtaccctgc</td><td>tcaatctccg</td><td>gaatggtgat</td><td>ctgatcgttc</td><td>ctgaaaacct</td><td>cgacattggc</td><td> 60</td>

<td>tccctcctga</td><td>cacaggtact</td><td>cgtacaggtt</td><td>ccaggtaaac</td><td>gagtcgtagt</td><td>tgtcgatcat</td><td> 120</td>

<td>gacaacgttc</td><td>ttagaagcgg</td><td>ccggcatttt</td><td>gaaggtgact</td><td>aatagcctaa</td><td>gaaaatattt</td><td> 180</td>

<td>aatttaattt</td><td>tcattaaatt</td><td>ttcctatact</td><td>cgctatttca</td><td>gcttttcatc</td><td>tcatcacttc</td><td> 240</td>

<td>ataaacgata</td><td>taaaccagaa</td><td>aaagaactat</td><td>tttcaaacac</td><td>gcttctcaaa</td><td>agcggtatgt</td><td> 300</td>

<td>ccttccacgt</td><td>ctccttagaa</td><td>tctggcaagt</td><td>ocgogagggg</td><td>gatccttagt</td><td>gatggtggtg</td><td> 360</td>

<td>atggtgtctg</td><td>ccctcgatca</td><td>cctgccactc</td><td>tgttgtagac</td><td>agcagc & gcg</td><td>agctaagctc</td><td> 420</td>

<td colspan="2">tgatctatcc ctgtcctcca</td><td>agtcacaacg</td><td>ctctcctcga</td><td>gtccaattgc</td><td>atgçggcttc</td><td> 480</td>

<td>gaacctgtgc</td><td>tccacgcccc</td><td>ccacgtacat</td><td>cctaaccttg</td><td>aagatggtga</td><td>agttgacagt</td><td> 540</td>

<td>gcaggggtag</td><td>tgccagagcc</td><td>tatatgggta</td><td>atgaaccata</td><td>cacctaggtg</td><td>tcagccaggg</td><td> 600</td>

<td>cccagaaccg</td><td>catctggcgt</td><td>aagtggcctc</td><td>ggggtgcttc</td><td>cgaaaacagt</td><td>cagtggggca</td><td> 660</td>

<td>ggtcaaggtg ttgttgccgg cccccccgat</td><td>gttgcacggg</td><td>gagcccccac acgtcttggt</td><td> 720</td>

<td>gaacccagtS çcattcatcc. atgtacagcc</td><td>gaaccagttg</td><td>cctcgcggcg gccgcgtgtt</td><td> 780</td>

<td>gttgagaatç agcacatcçg agtcgttcgc</td><td>cccccagtta</td><td>tacgtgggga caceaaaccg</td><td> 840</td>

<td>atcggtcgtc cccaccacaa cagggctçgg</td><td>ggtgaagcaa</td><td>tacactggac cgcacacctg</td><td> 900</td>

<td>agacgcgggt acaataccac acggtcgagg</td><td>cgcgtegtgc</td><td>cagcagtagg gcctctggtc</td><td> 960</td>

<td>cgagctgtta ggctcagtgt aagtgagggg</td><td>accccacccc</td><td>tgagcgaact tgtcgatgga</td><td> 1020</td>

<td>gcgacagctg gccaagcgct. ctgggcatcc</td><td>agacgagttg</td><td>aatttgtgtt tgtagaatag</td><td> 1080</td>

<td>tgcggcaaag aaccctgttt ggagggagtc</td><td>gttgcagttc</td><td>agggcagtcc tgttgatgtg</td><td> 1140</td>

<td>ccaactgcdg ttggtgttta cgagctggat</td><td>tttctgagcc</td><td>gacccggggc taaagaggga</td><td> 1200</td>

<td>cacaaggccc ctggtatcgg aggctgctgc</td><td>ccctcctgac</td><td>acgcgggtat gtcctagtgc</td><td> 1260</td>

<td>tgctagtggt ãggaagcata gtactagtat</td><td>tagtaggctg</td><td>cgcatgaatt cccgatgaag</td><td> 1320</td>

<td>cagagagcgc aggaggcggt atttatagtg</td><td>ccafctcccct</td><td>ctctgagaga cccggatggt</td><td> 1380</td>

<td>agtcgagtgt atcggagaca gcttgatgta</td><td>gactccgtgc</td><td>ctgccggctc ctcttattgg</td><td> 1440</td>

<td>cggacaccag tgagacaccc cggaacttgc</td><td>tgtttttctg</td><td>caaaatccgg ggtgaccagt</td><td> 1500</td>

<td>gggagcctat ttgcacacac gagcgggaca</td><td>ccccactctg</td><td>gtgaagagtg ccaaagfccat</td><td> 1560</td>

<td>tctttttccc gttgcggggc agcegattgc</td><td>gttttagg</td><td>cttt gc ta ca</td><td> 1620</td>

<td>occtgtcaga tttaccctcc acacatatat</td><td>attccgtcac</td><td>ctccagggac tattattcgt</td><td> 1680</td>

<td>cgttgcgccg ccagcggaag atatccagaa</td><td>gctgttttcc</td><td>gagagactcg gttggcgcct</td><td> 1740</td>

<td>ggtatatttg atggatgtcg cgctgcctca</td><td>cgtcccggta</td><td>cccaggaacg cggtgggatc</td><td> 1800</td>

<td>tcgggcccat cgaagactgt gctccagact</td><td>gctcgcccag</td><td>caggtgtttc ttgatcgccg</td><td> 1860</td>

<td>cctctaaatt gtccgcgcat cgccggtaac</td><td>atttttccag</td><td>ctcggagttt gcgtttagat</td><td> 1920</td>

<td>acagtttctg cgatgccaaa ggagcctgca</td><td>gattataacc</td><td>tcggatgctg tcattcagcg</td><td>19B0</td>

<td>cttttaattt gacctccaga tagttgctgt</td><td>atttctgttc</td><td>ccattggctg ctgcgcagct</td><td> 2040</td>

<td>tcgtataact cgagttattg ttgcgctctg</td><td>cctcggcgta</td><td>ctggctcatg atetggatct</td><td> 2100</td>

tgtccgtgtc gçttttcttc gagtgtttct cgcaaacgat gtgcaoggcc tgcagtgtcc 2160 aatcggagtc gagctggcgc cgaaactggc ggatctgagc ctccacactg ccctgtttct 2220 ctatccacgg cggaaccgcc tcctgccgtt fccagaatgtt gttcaagtgg tactctgtgc ggcgttattg ccggtgaaat ggtcaatgaa 2280 ctttgggaag cggfctttcct cggggaagat 2340 tacgaaattc cccgcgt-CGT tgcgcttcct ggatctcgag gagatcgttc tccgcgtcga 2400 ggagatcgtt ctccgcgtcg acaccattcc ttgcggcggc ggtgctcaac 2460 ggcctcaacc

I tactactggg ctgcttccfca atgcaggagt cgcataaggg agagcgtcga caaacccgcg 2520 tttgagaact tgctcaagct tctggtaaac gttgtagtac tctgaaacaa ggccctagca 2580 ctctgatctg tttctcttgg gtagcggtga gtggtttatt ggagttcact agtttcagca 2640 catctgtcat ctagacaata ttgtfcactaa atttttttga actacaattg ttcgtaattc 2700 atctattatt atacatcctc gtcagcaatt tctggcagac ggagtttact aacgtcttga 2760 gtatgaggcc gagaatccag ctctgtggcc atactcagtc ttgacagcct gctgatgtgg 2020 ctgcgttcaa cgcaataagc gtgtcctccg actccgagtt gtgctcgtta tcgtcgttct 2880

<td>catcctcgga</td><td>aaaatcacac</td><td>gaaagaacat</td><td>actcaccagt</td><td>aggctttctg i</td><td>gtccctgggg</td><td> 2940</td>

<td>cacggctgtt</td><td>tctgaegtat</td><td>tccggcgttg</td><td>ataatagctc</td><td>gaaagtgaac</td><td>gccgagtcgc</td><td> 3000</td>

<td>gggagtcgac</td><td>cgatgccctt</td><td>gagagccttc</td><td>aacccagtca</td><td>gctccttocg</td><td>gtgggcgcgg</td><td> 3060</td>

<td>ggcatgacta</td><td>tcgtcgccgc</td><td>acttatgact</td><td>gtcttcttta</td><td>tcatgcaact</td><td>cgtaggacag</td><td> 3120</td>

<td>gtgccggcag</td><td>cgctctgggt</td><td>cattttcggc</td><td>gaggaccgct</td><td>ttcgctggag</td><td>cgcgacgatg</td><td> 3180</td>

<td>atcggcctgt</td><td>cgcttgcggt</td><td>attcggaatc</td><td>ttgcacgccc</td><td>tcgctcaagc</td><td>cttcgtcact</td><td> 3240</td>

<td>ggtcccgcca</td><td>ccaaacgttt</td><td>cggcgagaag</td><td>caggccatta</td><td>tcgccggcat</td><td>ggcggccgac</td><td> 3300</td>

<td>gcgctgggct</td><td>acgtcttgct</td><td>ggcgttcgcg</td><td>acgcgaggct</td><td>ggatggcctt</td><td>ccccattatg</td><td> 3360</td>

<td>attcttctcg</td><td>Cttccggcgg</td><td>catcgggatg</td><td>cccgcgttgc</td><td>aggccatgct</td><td>gtccaggcag</td><td> 3420</td>

<td>gtagatgacg</td><td>accatcaggg</td><td>acagcttcaa</td><td>ggatcgctcg</td><td>cggctcttac</td><td>cagcctaact</td><td> 3480</td>

<td>tcgatcactg</td><td>gaccgctgat</td><td>cgtcacggcg</td><td>atttatgccg</td><td>cctcggcgag</td><td>cacatggaac</td><td>3H40</td>

<td>gggttggcat</td><td>ggattgtagg</td><td>cgccgcccta</td><td>taccttgtct</td><td>gcctcoccgc</td><td>gttgcgtcgc</td><td> 3600</td>

<td>ggtgcatgga</td><td>gccgggccac</td><td>ctcgacctga</td><td>atggaagccg</td><td>gcggcacctc</td><td>gctaacggat</td><td> 3660</td>

<td>tcaccactcc</td><td>aagaattgga</td><td>gccaatcaat</td><td>tcttgcggag</td><td>aactgtgaat</td><td>gcgcaaacca</td><td> 3720</td>

<td>acccttggca</td><td>gaacatatcc</td><td>atcgcgtccg</td><td>ccatctccag</td><td>cagccgcacg</td><td>cggcgcatcg</td><td> 3780</td>

<td>gggggggggg</td><td>gggggggggg</td><td>gggcaaacaa</td><td>ttcatcattt</td><td>tttttttatt</td><td>ottttttttg</td><td> 3840</td>

<td>atttcggttt</td><td>ctttgaaatt</td><td>tttttgattc</td><td>ggtaatctcc</td><td>gaacagaagg</td><td>aagaacgaag</td><td> 3900</td>

<td>gaaggagcac</td><td>agácttagat</td><td>tggtatatat</td><td>acgcatatgt</td><td>agtgttgaag</td><td>aaacatgaaa</td><td> 3960</td>

<td>ttgcccagta</td><td>ttcttaaccc</td><td>aactgcacag</td><td>aacaaaaacc</td><td>tgcaggaaac</td><td>gaagataaat</td><td> 4020</td>

<td>catgtcgaaa</td><td>gctacatata</td><td>aggaacgtgc</td><td>tgctactcat</td><td>cctagtcctg</td><td>ttgctgccaa</td><td> 4080</td>

<td>gctatttaat</td><td>atcatgcacg</td><td>aaaagcaaac</td><td>aaacttgtgt</td><td>gettcattgg</td><td>atgttcgtac</td><td> 4140</td>

<td>caccaaggaa</td><td>ttactagagt</td><td>tagttgaagc</td><td>attaggtccc</td><td>aaaatttgtt</td><td>tactaaaaac</td><td> 4200</td>

<td>acatgtggat</td><td>atctfcgaetg</td><td>atttttccat</td><td>ggagggcaca</td><td>gttaagccgc</td><td>taaaggcatt</td><td> 4260</td>

<td>atccgccaag</td><td>tacaattttt</td><td>tactcttcga</td><td>agacagaaaa</td><td>tttgctgaca</td><td>ttggtaatac</td><td> 4320</td>

agtcsaattg cagtactctg cgggtgtata çagaatagc ^. gaatggacag acattacgaa 4380 tgcacacggt gtggtgggcc caggtattgt feagcggtfitg aagcaggcgg cagaagaagt 4440 aacaaaggaa cctagaggcc ttttgatgtt agcagaattg tcatgcaagg gctccctatc 4500 tactggagaa tatactaagg gtactgttga cattgcgaag agcgacaaag attttgttat 4550 cggctttatt gctcaàagag acatgggtgg aagagatgaa ggttacgatt 4620 ggttgattat

<td>gacacccggt gtgggtttag atgacaaggg agacgcattg ggtcaacagt atagaaccgt</td><td> 4680</td>

<td>ggatgatgtg gtctctacag, gatctgacat tattattgtt ggaagaggac tatttgcaaa</td><td> 4740</td>

<td>gggaagggat gctaaggtag agggtgaacg ttacagaaaa gcaggctggg aagcatattt</td><td> 4800</td>

<td>gagaagatgc ggccagcaaa actaaaaaac tgtattataa gtaaatgcat gtatactaaa</td><td> 4860</td>

<td>ctcacaaatt agagcttcaa tttaattata tcagttatta cccgggaa.tc tcggtcgtaa</td><td> 4920</td>

<td>tgatttttat àatgacgaaa aaaaaaaaat tggaaagaaa agcccccccc cccccccccc</td><td> 4980</td>

<td>cccccccccc ccccccgcag cgttgggtcc tggccacggg tgcgcatgat cgtgctcctg</td><td> 5040</td>

<td>tcgttgagga cccggctagg ctggcggggt tgccttactg gttagcagaa tgaatcaccg</td><td> 5100</td>

<td>atacgcgagc gaacgtgaag cgactgctgc tgcaaaacgt ctgcgacctg agcaacaaca</td><td> 5160</td>

<td>tgaatggtct tcggtttccg tgtttcgtaa agtctggaaa cgcggaagtc agcgocctgc</td><td> 5220</td>

<td>accattatgü tccggatctg catcgcagga tgctgctggc taccctgtgg aacaectaca</td><td>528Õ</td>

<td>tctgtattaa cgaagcgctg gcattgaccc tgagtgattt ttctctggtc ccgccgcatc</td><td> 5340</td>

<td>cataccgcca gttgtttacc ctcacaacgt tccagtaacc gggçatgttc atcatcagta</td><td> 5400</td>

<td>acccgtatcg tgagcatcct ctctcgtttc atcggtatca ttacccccat gaacagaaat</td><td> 5460</td>

<td>tcccccttac acggaggcat caagtgacca aacaggaaaa aaccgccctt aacatggccc</td><td>5S20</td>

<td>gctttatcag aagccagaca ttaacgcttc tggagaaact caacgagctg gacgcggatg</td><td> 5580</td>

<td>aacaggcaga catctgtgaa tcgcttcacg accacgctga tgagctttac cgcagctgcc</td><td> 5640</td>

<td>tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca</td><td> 5700</td>

<td>cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg</td><td> 5760</td>

<td>ttggcgggtg tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtatactg</td><td> 5820</td>

<td>gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat</td><td> 5880</td>

<td>accgcacaga tgcgtaagga gaaaataccg catcaggcgc tcttccgctt cctcgcfccac</td><td> 5940</td>

<td>tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt</td><td> 6000</td>

<td>aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca</td><td> 6060</td>

<td>gcaaaaggcc aggaaocgta aaaaggccgc gttgctggcg ttttt-ceata ggctccgccc</td><td> 6120</td>

<td>ccctgacgag</td><td>catcacaaaa</td><td>atcgacgctc</td><td>aagtcagagg</td><td>tggcgaaacc '</td><td>cgacaggact</td><td> 6180</td>

<td>ataaagatac</td><td>caggcgtttc</td><td>cccctggaag</td><td>ctccctcgtg</td><td>cgctctcctg</td><td>ttccgaccct</td><td> 6240</td>

<td colspan="2">gccgcttacc ggatacctgt</td><td>ccgcctttct</td><td>cccttcggga</td><td>agcgtggcgc</td><td>tttctcatag</td><td> 630 0</td>

<td>ctcacgctgt</td><td>aggtatctca</td><td>gttcggtgta</td><td>ggtcgttcgc</td><td>tccaagctgg</td><td>gctgtgtgca</td><td> 6360</td>

<td>cgaacccccc</td><td>gttcagcccg</td><td>accgctgcgc</td><td>cttatccggt</td><td>aactatcgtc</td><td>ttgagtccaa</td><td> 6420</td>

<td>cccggtaaga</td><td>cacgacttat</td><td>cgccactggc</td><td>agcagccact</td><td>ggtaacagga</td><td>ttagcagagc</td><td> 6480</td>

<td>gaagtatgta</td><td>ggcggtgcta</td><td>cagagttctt</td><td>gaagtggtgg</td><td>cctaactacg</td><td>gctacactag</td><td> 6540</td>

<td>aaggacagta</td><td>tttggtatct</td><td>gcgctctgct</td><td>gaagccagtt</td><td>accttcggaa</td><td>aaagagttgg</td><td> 6600</td>

<td>tagctcttga</td><td>tccggcaaac</td><td>aaaccáccgc</td><td>tggtagcggt</td><td>ggtttttttg</td><td>tttgcaagca</td><td> 6660</td>

<td>gcagattacg</td><td>cgcagaaaaa</td><td>aaggatctca</td><td>agaagatcct</td><td>ttgatctttt</td><td>ctacggggtc</td><td> 672 0</td>

<td>tgacgctcag</td><td>tggaacgaaa</td><td>actcacgtta</td><td>agggattttg</td><td>gtcatgagat</td><td>tatcaaaaag</td><td> 6780</td>

<td>gatcttcacc</td><td>tagatccttt</td><td>taaattaaaa</td><td>atgaagtttt</td><td>aaatcaatct</td><td>aaagtatata</td><td> 6840</td>

<td>tgagtaaact</td><td>tggtctgaca</td><td>gttaccaatg</td><td>cttaatcagt</td><td>gaggcaccta</td><td>tctcagcgat</td><td> 6900</td>

<td>ctgtctattt</td><td>cgttcatcca</td><td>tagttgcctg</td><td>actccccgtc</td><td>gtgtagataa</td><td>ctacgatacg</td><td> 6960</td>

<td>ggagggctta</td><td>ccatctggcc</td><td>ccagtgctgc</td><td>aatgataccg</td><td>cgagacccac</td><td>gctcaccggc</td><td> 7020</td>

<td>tccagattta</td><td>tcagcaataa</td><td>accagccagc</td><td>cggaagggcc</td><td>gagcgcagaa</td><td>gtagtcctgc</td><td> 7080</td>

<td>aactttatcc</td><td>gcctccatcc</td><td>agtctattaa</td><td>ttgttgccgg</td><td>gaagctagag</td><td>taagtagttc</td><td> 7140</td>

<td>gccagttaat</td><td>agtttgcgca</td><td>acgttgttgc</td><td>cattgctgca</td><td>ggcatcgtgg</td><td>tgtcacgctc</td><td> 7200</td>

<td>gtcgtttggt</td><td>atggettcat</td><td>tcagctccgg</td><td>ttcccaacga</td><td>tcaaggcgag</td><td>ttacatgatc</td><td> 7260</td>

<td>ccccatgttg</td><td>tgcaaaaaag</td><td>cggttagctc</td><td>cttcggtcct</td><td>ccgatcgttg</td><td>tcagaagtaa</td><td> 7320</td>

gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat 7380 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata 7440 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaaca cgggataata cogcgccaca 7S00 tagcagaact ttaaaagtgc tcatcattgg aaaacgttet tcagggcgaa aactctcaag 7560 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca 7620 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc 7680 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc 7740 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta 7800 gaaaaataaa caaataaggg ttcegcgcac atttccccga aaagtgccac ctgacgtcta 7860 agaaaccatt attatcatga ctaaccaa

7927 , ι

<td> <213></td><td>synthetic primer or probe</td>

<td colspan="2">taaggatccc cgggtaccga gctc</td>

<td> <213></td><td>synthetic primer or probe</td>

<td colspan="2">ccagttcatc atcatatccc aagcc</td>

<td> <213></td><td>pUC18-FMD-CL-El vector</td>

<td> <221></td><td>hodgepodge</td>

<td> <223></td><td>N is any nucleotide</td>

<220>

<221> MISCELLANEOUS <222> (1167) .. (1168) <223> N is any nucleotide <400> 35

<td>acgcccaata</td><td>cgcaaaccgc</td><td>ctctccccgc</td><td>gcgttggccg</td><td>attcattaat</td><td>gcagctggca</td><td> 60</td>

<td>cgacaggttt</td><td>cccgactgga</td><td>aagcgggcâg</td><td>tgagcgcaac</td><td>gcaattaatg</td><td>tgagttagct</td><td> 120</td>

<td>cactcattag</td><td>gcaccccagg</td><td>ctttacactt</td><td>tatgcttccg</td><td>gctcgtatgt</td><td>tgtgtggaat</td><td> 180</td>

<td>tgtgagcgga</td><td>taacaatttc</td><td>acacaggaaa</td><td>cagctatgac</td><td>catgattacg</td><td>aattcgagct</td><td> 240</td>

<td>cggtacccgg</td><td>ggatccttac</td><td>cagttcatca</td><td>tcatatccca</td><td>agccat & cgg</td><td>tgacctgtta</td><td> 300</td>

<td>tgtggccggg</td><td>atagattgag</td><td>caattgcagt</td><td>cctgcaccgt</td><td>ctcatgccgg</td><td>cgaggcgaga</td><td> 360</td>

<td>tggtgaacag</td><td>ctgggagacg</td><td>aggaagacag</td><td>atccgcagag</td><td>afccccccacg</td><td>tacatagcgg</td><td> 420</td>

<td>aacagaaagc</td><td>agccgcccca</td><td>acgagcaaat</td><td>cgacgtggcg</td><td>tcgtattgtc</td><td>gtagtgggga</td><td> 480</td>

<td>cgctggcgtt</td><td>cctagctgcg</td><td>agcgtggggg</td><td>tgagcgctac</td><td>ccagcagcgg</td><td>gaagagttgt</td><td> 540</td>

<td>tctcccgaac</td><td>gcagggcacg</td><td>cacccggggg</td><td>tgtgcatgat</td><td>catgtccgct</td><td>gcctcataca</td><td> 600</td>

<td>caatgcttga</td><td>gttggagcag</td><td>tcgttcgtga</td><td>catggtacat</td><td>cccggacacg</td><td>ttgcgcacet</td><td> 660</td>

<td>catatcctag</td><td>tgctgctagt</td><td>ggtaggaagc</td><td>atagtactag</td><td>tattagtagg</td><td>cttcgcatga</td><td> 720</td>

attcccgatg aaggcagaga gcgcaaggag gcggtattta tagtgccatt JSTCC ctctg-780 agagacccgg atggtagtcg agtgttatcg gagacagctt gatgtagact ccgtgcctgc 840 cggtcctctt attogcggac accagtgaga caccccggaa cttgctgttt ttctgcaaaa 900 tccggggtga ccagtgggag cctatttgca caeacgagcg ggacacecca ctctggtgaa 960 gagtgccaaa gtcattcttt ttcccgtimc ggggcagccg attgcatgtt ttaggaaaat 1020 attacctttg ctacaccctg tcagatttac cctccacaca tatatattcc gtcacctcca 1.080

<td>gggactattc</td><td>ttggctcgtt</td><td>gcgccgccgc</td><td>ggaagatatc</td><td>cagaagctgt 1</td><td>gtttfcccgag</td><td> 1140</td>

<td>agactcggtt</td><td>ggcgcctggt</td><td>atatttnnag</td><td>gatgtcgcgc</td><td>tgcctcacgt</td><td>cccggtaccc</td><td> 1200</td>

<td>aggaacgcgg</td><td>tgggatctcg</td><td>ggcccatcga</td><td>agactgtgct</td><td>ccagactgct</td><td>cgcccagcag</td><td> 1260</td>

<td>gtgtttcttg</td><td>attgccgcct</td><td>ctaaatagtc</td><td>cgcgcatcgc</td><td>cggtaacatt</td><td>tttccagctc</td><td> 1320</td>

<td>ggagtttgcg</td><td>tttagataca</td><td>tttctgcgat</td><td>gccaaaggag</td><td>cctgcagatt</td><td>ataacctcgg</td><td> 1380</td>

<td>atgctgtcat</td><td>tcagcgcttt</td><td>taatttgacc</td><td>tccagatagt</td><td>tgctgtattt</td><td>ctgttccafct</td><td> 1440</td>

<td>ggctgctgga</td><td>cgttcgtata</td><td>actcgagtta</td><td>ttgttgcgct</td><td>ctgcctcggc</td><td>gtactggctc</td><td>1SOO</td>

<td>atgactgact</td><td>gcggtcgctt</td><td>ctcgagtgtt</td><td>ctcgcaacag</td><td>gacgcctgca</td><td>ggtcatcgag</td><td> 1560</td>

<td>tcgagctggc</td><td>gccgaaactg</td><td>gcggatctga</td><td>cctccacact</td><td>gccctgtatc</td><td>tctatccacc</td><td> 1620</td>

<td>gggaaccgcc</td><td>tcctgccgtt</td><td>ccagaatgtt</td><td>gttcaagtgg</td><td>t agetetgtg</td><td>cggtcaatga</td><td> 1680</td>

<td>aggcgttatt</td><td>gccggtgaaa</td><td>tctttgggaa</td><td>gcggtttatc</td><td>ctcggggaag</td><td>attacgaaat</td><td> 1740</td>

<td>tcccgcgcgt</td><td>cgttgcgctt</td><td>cctggatctc</td><td>gaggaagatc</td><td>gttctccgcg</td><td>tcgaggagat</td><td> 1800</td>

<td>cgttctccgc</td><td>gtcgacctgc</td><td>aggcatgcaa</td><td>gcttggcact</td><td>ggccgtcgtt</td><td>ttacaacgtc</td><td> 1860</td>

<td>gtgactggga</td><td>aaaccctggc</td><td>gttacccaac</td><td>ttaatcgcct</td><td>tgeagcacat</td><td>ccccctttcg</td><td> 1920</td>

<td>ccagctggcg</td><td>taatagcgaa</td><td>gaggcccgca</td><td>ccgatcgccc</td><td>ttcccaacag</td><td>ttgcgcagcc</td><td> 1980</td>

<td>tgaatggcga</td><td>atggcgcctg</td><td>atgcggtatt</td><td>ttctccttac</td><td>gcatctgtgc</td><td>ggtatttcac</td><td> 2040</td>

<td>accgcatatg</td><td>gtgcactctc</td><td>agtacaatct</td><td>gctctgatgc</td><td>cgcatagtta</td><td>agccagcccc</td><td> 2100</td>

<td>gacacccgcc</td><td>aacacccgct</td><td>gacgcgccct</td><td>gacgggcttg</td><td>tctgctcccg</td><td>gcatccgctt</td><td> 2160</td>

<td>acagacaagc</td><td>tgtgaccgtc</td><td>tccgggagct</td><td>gcatgtgtca</td><td>gaggttttca</td><td>ccgtcatcac</td><td> 2220</td>

<td>cgaaacgcgc</td><td>gagacgaaag</td><td>ggcctcgtga</td><td>tacgcctatt</td><td>tttataggtt</td><td>aatgtcatga</td><td> 2280</td>

<td>taataatggt</td><td>ttcttagacg</td><td>t-caggtggca</td><td>cttttcgggg</td><td>aaatgtgcgc</td><td>ggaaccccta</td><td> 2340</td>

<td>tttgtttatt</td><td>tttctaaata</td><td>cattcaaata</td><td>tgtatccgct</td><td>catgagacaa</td><td>taaccctgat</td><td> 2400</td>

<td>aaatgcttca</td><td>ataatattga</td><td>aaaaggaaga</td><td>gtatgagtat</td><td>tcaacatttc</td><td>cgtgtcgccc</td><td> 2460</td>

<td>ttattccctt</td><td>ttttgcggca</td><td>ttttgccttc</td><td>ctgtttttgc</td><td>tcacccagaa</td><td>acgctggtga</td><td>2H20</td>

, ι

<td>aagtaaaaga tgctgaagat cagttgggtg çacgagtggg ttacatcgaa i</td><td>ctggatctca</td><td> 2580</td>

<td>aeagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg i</td><td>atgagcactt</td><td> 2640</td>

<td>ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa ·</td><td>gagcaactcg</td><td> 2700</td>

<td>gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc</td><td>acagaaaagc</td><td> 2760</td>

<td>atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc</td><td>atgagtgata</td><td> 2820</td>

<td>acactgcggc caacttactt ctgacaaçga tcggaggacc gaaggagcta</td><td>accgcttttt</td><td> 2880</td>

<td>tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag</td><td>ctgaatgaag</td><td> 2940</td>

<td>ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca</td><td>acgttgcgca</td><td> 3000</td>

<td>aactattaac tggcgaacta cttactctag cttcccggca acaattaata</td><td>gactggatgg</td><td> 3060</td>

<td>aggcggataa agttgcagga ccacttctge gctcggccct tccggctggc</td><td>tggtttattg</td><td>3X2 0</td>

<td>ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca</td><td>ctggggccag</td><td> 3180</td>

<td>atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca</td><td>actatggatg</td><td> 3240</td>

<td>aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg</td><td>taactgteag</td><td> 3300</td>

<td>accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa</td><td>tttaaaagga</td><td> 3360</td>

<td>tctaggtgaa gatCcttttt gataatctca tgaccaaa & t ceettaacgt</td><td>gagttttcgt</td><td> 3420</td>

<td>tccactgagc gtcagacçcc gtagaaaaga tcaaaggatc ttcttgagat</td><td>cctttttttc</td><td> 3480</td>

<td>tgcgcgtaat ctgctgcttg caaaeaaaaa aaccaccgct accagcggtg</td><td>gtttgtttgc</td><td> 3540</td>

<td>cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga</td><td>gcgcagatac</td><td> 3600</td>

<td>caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac</td><td>tctgtagcac</td><td> 3660</td>

<td>cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt</td><td>ggcgataagt</td><td> 3720</td>

<td>Cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag</td><td>çggtcgggct</td><td> 3780</td>

<td>gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc</td><td>gaactgagat</td><td> 3840</td>

<td>acctacagcg tgágctatga gaaagcgcca cgcttcccga agggagaaag</td><td>gcggacaggt</td><td> 3900</td>

<td>atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca</td><td>gggggaaacg</td><td> 3960</td>

<td>cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt</td><td>cgatttttgt</td><td> 4020</td>

<td>gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc</td><td>tttttacggt</td><td> 4060</td>

<td>tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc</td><td>cctgattctg</td><td>4X40</td>

<td>tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc</td><td>cgaacgaccg</td><td> 4200</td>

<td>agcgcagcga gtcagtgagc gaggaagcgg aaga</td><td></td><td> 4234</td>

. <210> 36 <211> 7429 <212> DNA <213> vector pFPMT-CL-El <400> 36 ggtaccctgc tcaatctccg gaatggtgat ctgatcgttc ctgaaaacct cgacattggc 60 tcoctcctga cacaggtact cgtacaggtt ccaggtaaac gagtcgtagt tgtcgatcat 120 gacaacgttc ttagaagcgg ccggcatttt gaaggtgact aatagcctaa gaaaatattt 180 aatttaattt tcattaaatt ttcctatact egctatttca gcttttcatc tcatcacttc 240

<td>ataaacgáta</td><td>taaaccagaa</td><td>aaagaactat</td><td>tttcaaacaç <</td><td>gcttctcaaa j</td><td>agcggtatgt</td><td> 300</td>

<td>ccttccacgt</td><td>ctccttagaa</td><td>tctggcaagt</td><td>ccgcgagggg.</td><td>gatccttacc <</td><td>agttcatcat</td><td> 360</td>

<td>catatcccaa</td><td>gccatacggt</td><td>gacctgttat</td><td>gtggccggga</td><td>tagattgagc</td><td>aattgcagtc</td><td> 420</td>

<td>ctgcaccgtc</td><td>tcatgccggc</td><td>gaggcgagat</td><td>ggtgaacagc</td><td>tgggagacga</td><td>ggaagacaga</td><td> 480</td>

<td>tccgcagaga</td><td>tcccccacgt</td><td>acatagcgga</td><td>aoagaaagca</td><td>gccgccccaa</td><td>cgagcaaatc</td><td> 540</td>

<td>gacgtggcgt</td><td>cgtattgtcg</td><td>tagtggggac</td><td>gctggcgttc</td><td>ctagctgcga</td><td>gogtgggggt</td><td> 600</td>

<td>gagcgctacc</td><td>cagcagcggg</td><td>aagagttgtt</td><td>ctcccgaacg</td><td>cagggcacgc</td><td>acccgggggt</td><td> 660</td>

<td>gtgcatgatc</td><td>atgtcegctg</td><td>cctcatacac</td><td>aatgcttgag</td><td>ttggagcagt</td><td>cgtfccgtgac</td><td> 720</td>

<td>atggtacatc</td><td>ccggacacgt</td><td>tgcgcacctc</td><td>atatcctagt</td><td>gctgctagtg</td><td>gtaggaagca</td><td> 780</td>

<td>tagtactagt</td><td>attagtaggc</td><td>ttcgcatgaa</td><td>ttcccgatga</td><td>agcagagagc</td><td>gcaggaggcg</td><td> 840</td>

<td>gtatttatag</td><td>tgccattccc</td><td>ctctctgaga</td><td>gacccggatg</td><td>gtagtcgagt</td><td>gtatcggaga</td><td> 900</td>

<td>cagcttgatg</td><td>tagactecgt</td><td>gcctgccggc</td><td>tcctcttatt</td><td>ggcggacacc</td><td>agtgagacac</td><td> 960</td>

<td>cccggaactt</td><td>gctgtttttc</td><td>tgcaaaatcc</td><td>ggggtgacca</td><td>gtgggagcct</td><td>atttgcacac</td><td> 1020</td>

<td>acgagcggga</td><td>caccccactc</td><td>tggtgaagag</td><td>tgccaaagte</td><td>attctttttc</td><td>ccgttgcggg</td><td> 1080</td>

<td>gcagccgatt</td><td>gcatgtttta</td><td>ggaaaatatt</td><td>acctttgcta</td><td>caccctgtca</td><td>gatttaccct</td><td> 1140</td>

<td>ccacacatat</td><td>atattccgtc</td><td>acctccaggg</td><td>actattattc</td><td>gtcgttgcgc</td><td>cgccagcgga</td><td> 1200</td>

<td>agatatccag</td><td>aagctgtttt</td><td>ccgagagact</td><td>cggttggcgc</td><td>ctggtatatt</td><td>tgatggatgt</td><td> 1260</td>

<td>cgcgctgcct</td><td>cacgtcccgg</td><td>tacccaggaa</td><td>cgcggtggga</td><td>tctogggccc</td><td>atcgaagact</td><td> 1320</td>

<td>gtgctccaga</td><td>ctgctcgccc</td><td>agcaggtgtt</td><td>tcttgatcgc</td><td colspan="2">cgcctctaaa ttgtccgcgc</td><td> 1380</td>

<td>atcgccggta</td><td>acatttttcc</td><td>agctcggagt</td><td>ttgcgtttag</td><td>attackagtttc</td><td>tgcgatgcca</td><td> 1440</td>

<td>aaggagcctg</td><td>cagattataa</td><td>cctcggatgc</td><td>tgtcattcag</td><td>cgcttttaat</td><td>ttgacctoca</td><td> 1500</td>

<td>gatagttgct</td><td>gtatttctgt</td><td>tcccattggc</td><td>tgctgcgcag</td><td>cttcgtataa</td><td>ctcgagttat</td><td> 1560</td>

<td>tgttgcgctc</td><td>tgcctcggcg</td><td>tactggctea</td><td>tgatctggat</td><td>cttgtccgtg</td><td>fccgcttttct</td><td> 1620</td>

<td>tcgagtgttt</td><td>ctcgcaaacg</td><td>atgtgcacgg</td><td>cctgcagtgt</td><td>ccaatcggag</td><td>tcgagctggc</td><td> 1680</td>

<td>gccgaaactg</td><td>gcggatctga</td><td>gcctccacac</td><td>tgccctgttt</td><td>ctctatccac</td><td>ggcggaaccg</td><td> 1740</td>

cctcctgccg tttcagaatg ttgttcaagt ggtactctgt gcggtcaatg aaggcgttat 1800 tgccggtgaa atçtttggga agcggttttc ctcggggaág attacgaaat tccccgcgtc 1860 gttgcgcttc ctggatctcg aggagatcgt tctccgcgtc gaggagatcg ttctccgcgt 1920 cgacaccatt ccttgcggcg gcggtgctca acggcctcaa cctactactg ggctgcttcc 1980 taatgcagga gtcgcátaag ggagagcgtc gacaaacccg cgtttgagaa cttgctcaag 2040 cttctggtaa acgttgtagt actctgaaac aaggccctag cactctgatc tgtttctctt 2100 gggtagcggt gagtggttta ttggagttca ctggtttcag cacatctgtc atctagacaa 2160

<td>tattgttact</td><td>aaattttttt</td><td>gaactacaat</td><td>tgttcgtaat</td><td>tcatctatta</td><td>ttatacatcc</td><td> 2220</td>

<td>tcgtcagcaa</td><td>tttctggeag</td><td>acggagttta</td><td>ctaacgtctt</td><td>gagtatgagg</td><td>ccgagaatcc</td><td> 2280</td>

<td>agctctgtgg</td><td>ccatactcag</td><td>tcttgacagc</td><td colspan="2">ctgctgatgt ggctgcgttc</td><td>aacgcaataa</td><td> 2340</td>

<td>gcgtgtcctc</td><td>cgactccgag</td><td>ttgtgctcgt</td><td>tategtcgtt</td><td>ctcatcctcg</td><td>gaaaaatcac</td><td> 2400</td>

<td>acgaaagaac</td><td>atactcacca</td><td>gtaggcttte</td><td>tggtccctgg</td><td>ggcaeggctg</td><td>tttctgacgt</td><td> 2460</td>

<td>attccggcgt</td><td>tgataatagc</td><td>tcgaaagtga</td><td>acgccgagtc</td><td>gogggagtcg</td><td>accgatgccc</td><td> 2520</td>

<td>ttgagagcct</td><td>tcaacccagt</td><td>cagctccttc</td><td>cggtgggcgc</td><td>ggggcatgac</td><td>tatcgtegcc</td><td> 2580</td>

<td>gcacttatga</td><td>ctgtcttctt</td><td>tatcatgcaa</td><td>ctcgtaggae</td><td>aggtgccggc</td><td>agcgctctgg</td><td> 2640</td>

<td>gtcattttcg</td><td>gcgaggaçcg</td><td>ctfctcgctgg</td><td>ágcgcgacga</td><td>tgatcggcct</td><td>gtcgcttgcg</td><td> 2700</td>

<td>gtattcggaa</td><td colspan="2">tcttgcacgc cctcgctcaa</td><td>gccttcgtca</td><td>ctggtcccgc</td><td>caccaaacgt</td><td> 2760</td>

<td>ttcggcgaga</td><td>agcaggccat</td><td>tatcgccggc</td><td>atggcggccg</td><td>acgcgctggg</td><td>ctacgtcttg</td><td> 2820</td>

<td>ctggcgttcg</td><td>cgacgcgagg</td><td>ctggatggcc</td><td>ttccccatta</td><td>tgattcttct</td><td>cgcttccggc</td><td> 2880</td>

<td>ggcatcggga</td><td>tgcccgcgtt</td><td>gcaggccatg</td><td>ctgtccaggc</td><td>aggtagatga</td><td>cgaccatcag</td><td> 2940</td>

<td>ggacagcttc</td><td>aaggatcgct</td><td>cgcggctctt</td><td>accagcctaa</td><td>cttcgatcac</td><td>tggaccgctg</td><td> 3000</td>

<td>atcgtcacgg</td><td>cgatttatgc</td><td>cgcctcggcg</td><td>agcacatgga</td><td>acgggttggc</td><td>atggattgta</td><td> 3050</td>

<td>ggcgocgccc</td><td>tataccttgt</td><td>ctgcctcccç</td><td>gcgttgcgtc</td><td>gcggtgcatg</td><td>gagccgggec</td><td> 3120</td>

<td>acctcgacct</td><td>gaatggaagc</td><td>cggcggcace</td><td>tcgctaacgg</td><td>attcaccact</td><td>ocaagaattg</td><td> 3180</td>

<td>gagccaatca</td><td>attcttgcgg</td><td>agaactgtga</td><td>atgcgcaaac</td><td>caacccttgg</td><td>cagaacatat</td><td> 3240</td>

<td>ccatcgcgtc</td><td>cgccatctcc</td><td>agcagccgca</td><td>cgcggcgcat</td><td>cggggggggg</td><td>gggggggggg</td><td> 3300</td>

<td>gggggcaaac</td><td>aatteatcat</td><td>ttttttttta</td><td>ttcttttttt</td><td>tgatttcggt</td><td>ttctttgaaa</td><td> 3360</td>

<td>tttttttgat</td><td>tcggtaatct</td><td>cogaacagaa</td><td>ggaagaaoga</td><td>aggaaggagc</td><td>acagacttag</td><td> 3420</td>

<td>attggtatat</td><td>Atacgcatat</td><td>gtagtgttga</td><td>agaaacatga</td><td>aattgcccag</td><td>tattcttaac</td><td> 3480</td>

ccaactgcac agaacaaaaa cctgcaggaa acgaagataa atcatgtcga aagctacata 3540

I

<td>taaggaacgt</td><td>gctgctactc</td><td>atcctagtcc</td><td>tgttgctgcc</td><td>aagctattta</td><td>atatcatgca</td><td> 3600</td>

<td>cgaaaagcaa</td><td>acaaacttgt</td><td>gtgcttcatt</td><td>ggatgttcgt</td><td>accaccaagg</td><td>aattactgga</td><td> 3660</td>

<td>gttagttgaa '</td><td>gcattaggtc</td><td>ccaaaatttg</td><td>tttactaaaa</td><td>acacatgtgg</td><td>atatcttgac</td><td> 3720.</td>

<td>tgatttttcc</td><td>atggagggca</td><td>cagttaagcc</td><td>gctaaaggca</td><td>ttatccgcca</td><td>agtacaattt</td><td> 3780</td>

<td>tttactcttc</td><td>gaagacagaa</td><td>aatttgctga</td><td>cattggtaát</td><td>acagtcaaat</td><td>tgcagtactc</td><td> 3840</td>

<td>tgcgggtgta</td><td>tacagaatag</td><td>cagaatgggc</td><td>agacattacg</td><td>aatgcacacg</td><td>gtgtggtggg</td><td> 3900</td>

<td>cccaggtatt</td><td>gttagcggtt</td><td>tgaagcaggc</td><td>cgcagaagaa</td><td>gtaacaaagg</td><td>aacctagagg</td><td> 3960</td>

<td>ccttttgatg</td><td>ttagcagaat</td><td>tgtcatgcaa</td><td>gggctcccta</td><td>tctactggag</td><td>aatata-ctaa</td><td> 4020</td>

<td>gggtactgtt</td><td>gacattgcga</td><td>agagcgacaa</td><td>agattttgtt</td><td>atcggcttüa</td><td>ttgctcaaag</td><td> 4080</td>

<td>agacatgggt</td><td>ggaagagatg</td><td>aaggt.tacga</td><td>ttggttgatt</td><td>atgacacccg</td><td>gtgtgggttt</td><td> 4140</td>

<td>agatgacaag</td><td>ggagacgcat</td><td>tgggtcaaca</td><td>gtatagaacc</td><td>gtggatgatg</td><td>tggtctctac</td><td> 4200</td>

<td>aggatctgac</td><td>attattattg</td><td>ttggaagagg</td><td>aetatttgca</td><td>aagggaaggg</td><td>atgctaaggt</td><td> 4260</td>

<td>agagggtgaa</td><td>cgttacagaa</td><td>aagcaggctg</td><td>ggaagcatat</td><td>ttgagaagat</td><td>gcggccagca</td><td> 4320</td>

<td>aaactaaasa</td><td>actgtattat</td><td>aagtaaatgc</td><td>atgtatacta</td><td>aactcacaaa</td><td>ttagagcttc</td><td> 4380</td>

<td>aatttaatta</td><td>tatcagttat</td><td>tacccgggaa</td><td>tctcggtcgt</td><td>aatgattttt</td><td>ataatgacga</td><td> -4440</td>

<td>aaaaaaaaaa</td><td>attggaaaga</td><td>aaagcccccc</td><td>cccccccccc</td><td>cccccccccc</td><td>ccccccccgc</td><td> 4500</td>

<td>agcgttgggt</td><td>cctggccacg</td><td>ggtgcgcatg</td><td>atcgtgctcc</td><td>tgtcgttgag</td><td>gaoccggcta</td><td> 4560</td>

<td>ggctggcggg</td><td>gttgccttac</td><td>tggttagcag</td><td>aatgaatcac</td><td>cgatacgcga</td><td>gcgaacgtga</td><td> 4620</td>

<td>agcgactgct</td><td>gctgcaaaac</td><td>gtctgcgacc</td><td>tgagcaacaa</td><td>catgaatggt</td><td>ctteggtttc</td><td> 4-680</td>

<td>cgtgtttcgt</td><td>aaagtctgga</td><td>aacgcggaag</td><td>tcagcgccct</td><td>gcaccattat</td><td>gttocggatc</td><td> 4740</td>

tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt aacgaagcgc 4-800 tggcattgac cctgagtgat ttttctctgg tcccgccgca tccataccgc cagttgttta 4860 ccctcacaac gttccagtaa ccgggcatgt tcatcatcag taacccgtat cgtgagcatc 4320 ctctctcgtt tcatcggtat cattaccccc atgaacagaa attccccctt acacggaggc 4380 ateaagtgac caaacaggaa aaaaccgccc ttaacatggc ccgctttatc agaagccaga 5040 cattaacgct tctggagaaa ctcaacgagc tggacgcgga tgaacaggca gacatctgtg 5100 aatcgcttca cgaccacgct gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg 5160 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg 5220 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg 5280 cagccatgac ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc 5340 agagcagatt gtactgagag tgcaccatat gcggtgtgaa the tac CGCA ca gatgcgtaag 5400 gagaasatac cgcatcaggc gctcttccgç ttcctcgctç actgactcgc tg-cgctcggt 5460 cgttcggçtg cggcgagcgg tatcagctca ctcaaaggfcg gtaatacggt tatccacaga 5520 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg 5580 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ecccctgacg agcatcacaa 5640 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat aceaggcgtt 5700 tccccctgga agctccctcg tgcgctçtcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg 5760, 5820 gaagcgtggc gctttcteat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc 5B80 ccgttcagcc

<td>cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt</td><td> 5940</td>

<td>atcgccactg gcagcagcca ctggtaaçag gattagcagâ gcgaggtatg taggcggtgc</td><td> 6000</td>

<td>tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat</td><td> 6060</td>

<td>ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa</td><td> 6120</td>

<td>acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa</td><td> 6180</td>

<td>aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga</td><td> 6240</td>

<td>aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct</td><td> 6300</td>

<td>tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga</td><td> 6360</td>

<td>cagttaccaa tgcttaatca gtgaggcàcc tatctcagcg atctgtctat ttcgttcatc</td><td> 6420</td>

<td>catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg</td><td> 6480</td>

<td>ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat</td><td> €540</td>

<td>aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat</td><td> €600</td>

<td>ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg</td><td> 6660</td>

<td>caacgttgtt gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc</td><td> €720</td>

<td>atteagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa</td><td> 6780</td>

<td>agcggttagc tccttcggtc ctccgatcgt tgtcaçaagt aagttggccg cagtgttatc</td><td> 6840</td>

<td>actcatggtt atggcagcac tgcataattc tcttactgtc atgocatccg taagatgctt</td><td> 6900</td>

<td>ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag</td><td> 6960</td>

<td>ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt</td><td> 7020</td>

<td>gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag</td><td> 7080</td>

<td>atccagtteg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac</td><td> 7140</td>

<td>cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc</td><td> 7200</td>

I gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca 7260 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg 7320 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat 7380 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcaa 7429 <210> 37 <211> 39 <212> DMA <213> primer or synthetic probe <400> 37 catcacaaat atgaggtgcg caacgtgtcc gggatgtac 39 <210> 38 <211> 42 <212> DNA <213> primer or synthetic probe <400> 38 gtgatggtgg tgtcctagtg ctgctagtgg taggaagcat ag 42 <210> 39 <211> 4273 <212> DNA <213> vector PUC18-FMD-CL-E1-H-K6 <220>

<221> MISCELLANEOUS <222> (1027) .. (1028) <223> N is any nucleotide <220>

<221> MISCELLANEOUS <222> (1206) .. (1207) <223> N is any nucleotide <400> 39

<td>gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca</td><td> €0</td>

<td>cgacaggttt ccogactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct</td><td> 120</td>

<td>cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat</td><td> 180</td>

<td>tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg aattcgagct</td><td> 240</td>

<td>cggtacccgg ggatccttaa tggtgatggt ggtggtgcca gtbcatcatc atatcccaag</td><td> 300</td>

<td>ccatacggtg acctgttatg tggccgggat agattgagca attgcagtcc tgcaccgtct</td><td> 360</td>

<td>catgceggcg aggcgagatg gtgaacagct gggagacgag gaagacagat ccgcagagat</td><td> 420</td>

<td>cccccacgta catagcggaa cagaaagcag ccgccccaac gagcaaatcg acgtggcgtc</td><td> 480</td>

<td>gtattgtcgt</td><td>agtggggacg</td><td>ctggcgttcc</td><td>tagctgcgag.</td><td>cgtggsgstg</td><td>agcgctaccc</td><td> 540</td>

<td>agcagcggga</td><td>agagttgttc.</td><td>tcocgaacgc</td><td>agggcacgçá</td><td>cccgggggtg</td><td>tgçatgatca</td><td> 600</td>

<td>tgtccgctgc.</td><td>ctcatacaca</td><td>atgcttgagt</td><td>tggagcagtc</td><td>gttcgtgaca</td><td>tggtacatcc</td><td> 660</td>

<td>cggacacgtt</td><td>gcgcacctca</td><td>tatttgtgat</td><td>ggtgatggtg</td><td>gtgtcctagt</td><td>gctgctagtg</td><td> 720</td>

<td>gtaggaagca</td><td>tagtactagt</td><td>attagtaggc</td><td>tfccgcatgaa</td><td>ttcccgatga</td><td>aggcagagag</td><td> 780</td>

<td>cgcaaggágg</td><td>cggtatttat</td><td>agtgccattc</td><td>ccctctctga</td><td>gagacccgga</td><td>tggtagtcga</td><td> 840</td>

<td>gtgttatcgg</td><td>agacag-cttg</td><td>atatagactc</td><td>cgtgcctgcc</td><td>ggtcctctta</td><td>ttggcggaca</td><td> 900</td>

<td>ccagtgagac</td><td>accccggaac</td><td>ttgctgtttt</td><td>tctgcaaaat</td><td>ccggggtgac</td><td>cagtgggagc</td><td> 960</td>

<td>ctatttgcac</td><td>acacgagcgg</td><td>gacaccccac</td><td>tctggtgaag</td><td>agtgccaaag</td><td>tcattctttt</td><td> 1020</td>

<td>tcccgtnncg</td><td>gggcagccga</td><td>ttgcatgttt</td><td>taggaaaata</td><td>ttacctttgc</td><td>tacaccctgt</td><td> 1080</td>

<td>cagatttacc</td><td>ctccacacat</td><td>atatattccg</td><td>tcacctccag</td><td>ggactattct</td><td>tggctcgttg</td><td> 1140</td>

<td>cgccgccgcg</td><td>gaagatatcc</td><td>agaagctgtg</td><td>ttttccgaga</td><td>gactcggttg</td><td>gcgcctggta</td><td> 1200</td>

<td>tatttimagg</td><td>atgtcgcgct</td><td>gcctcacgtc</td><td>coggtaccca</td><td>ggaacgcggt</td><td>gggatctcgg</td><td> 1260</td>

<td>gcccatcgaa</td><td>gactgtgctc</td><td>cagactgctc</td><td>gcccagcagg</td><td>tgtttcttga</td><td>ttgccgcctc</td><td> 1320</td>

<td>taaatagt.cc</td><td>gcgcatcgcc</td><td>ggtaacattt</td><td>ttccagctcg</td><td>gagtttgcgt</td><td>ttagatacat</td><td> 1380</td>

<td>ttctgcgatg</td><td>ccaaaggagc</td><td>ctgcagatta</td><td>taacctcgga</td><td>tgctgtcatt</td><td>cagcgctttt</td><td> 1440</td>

<td>aatttgacct</td><td>ccagatagtt</td><td colspan="2">gctgtatttc tgttccattg</td><td>gctgctggac</td><td>gttcgtataa</td><td> 1500</td>

<td>ctcgagttat</td><td>tgttgcgcte</td><td>tgcctcggcg</td><td>tactggctca</td><td>tgactgactg</td><td>cggtcgcttc</td><td> 1560</td>

tcgagtgttc tcgcaacagg acgcctgcag gtcatcgagt cgagctggcg ccgaaactgg 1620 cggatctgac ctccacaçtg ccctgtatct ctatccaccg ggaaccgcct cctgccgttc 1680 cagaatgttg ttcaagtggt agctctgtgc ggtcaatgaa ggcgttattg eeggtgaaat 1740 ctttaggaag cggtttatcc tcggggaaga ttacgaaatt cccgcg-TCGC gttgcgcttc 1800 ctggatctcg aggaagatcg ttctccgcgt cgaggagatc gttctccgcg tcgacctgca 1860 ggcatgcaag cttggcactg gecgtcgttt tacaacgtcg tgactgggaa aaccctggcg 1920 ttacccaact taatcgcctt gcagcacatc cccctttogc cagctggcgt aatagcgaag 1980 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga 2040 tgcggtattt tctccttaeg catctgtgcg gtatttcaca ocgcatatgg tgcactctca 2100 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg 2160 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct 2220 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agaogaaagg 2280 gcctcgtgat acgcctattt ttataggtta atgtcataat aataatggtt 2340 tcttagacgt

<td>caggtgg-cac</td><td>ttttcgggga</td><td>aatgtgcgcg</td><td>gaacccctat</td><td>ttgtttattt '</td><td>ttctaaatac</td><td> 2400</td>

<td>attcaaatat</td><td>gtatccgctc</td><td>atgagacaat</td><td>aaccctgata</td><td>aatgcttcaa:</td><td>taatattgaa</td><td> 2460</td>

<td>aaaggaagag</td><td>tatgagtatt</td><td>caacatttcc</td><td>gtgtcgccct</td><td>tattcccttt</td><td>tttgcggcat</td><td> 2520</td>

<td>tttgccttcc</td><td>tgtttttgct</td><td>cacccagaaa</td><td>cgctggtgaa</td><td>agtaaaagat</td><td>gctgaagatc</td><td>2H80</td>

<td>agttgggtgc</td><td>acgagtgggt</td><td>tacatcgaac</td><td>tggatctcaa</td><td>cagcggtaag</td><td>atccttgaga</td><td> 2640</td>

<td>gttttcgccc</td><td>cgaagaacgt</td><td>tttccaatga</td><td>tgagcacttt</td><td>taaagttctg</td><td>ctatgtggcg</td><td> 2700</td>

<td>cggtattatc</td><td>ccgtattgac</td><td>gccgggcaag</td><td>agcaactcgg</td><td>tcgccgcata</td><td>cactattctc</td><td> 2760</td>

<td>agaatgactt</td><td>ggttgagt.ac</td><td>tcaccagtca</td><td>cagaaaagca</td><td>tcttacggat</td><td>ggcatgacag</td><td> 2820</td>

<td>taagagaatt</td><td>atgcagtgct</td><td>gccataacca</td><td>tgagtgataa</td><td>cactgcggcc</td><td>aacttacttC</td><td> 2880</td>

<td>tgacaacgat</td><td>cggaggaccg</td><td>aaggagctaa</td><td>ccgctttttt</td><td>gcacaacatg</td><td>ggggatcatg</td><td> 2940</td>

<td>taactcgcct</td><td>tgatcgttgg</td><td>gaaccggagc</td><td>tgaatgaagc</td><td>cataccaaac</td><td>gacgagcgtg</td><td> 3000</td>

<td>acaccacgat</td><td>gcctgtagca</td><td>atggcaacaa</td><td>cgttgcgcaa</td><td>actattaact</td><td>ggcgaactac</td><td> 3060</td>

<td>ttactctagc</td><td>ttcccggcaa</td><td>caattaatag</td><td>actggatgga</td><td>ggcggataaa</td><td>gttgcaggac</td><td> 3120</td>

<td>cacttctgcg</td><td>ctcggccctt</td><td>ccggctggct</td><td>ggtttattgc</td><td>tgataaatct</td><td>ggagccggtg</td><td> 3180</td>

<td>agcgtgggtc</td><td>tcgcggtatc</td><td>attgcagcac</td><td>tSSSSecaga</td><td>tggtaagccc</td><td>tcccgtatcg</td><td> 3240</td>

<td>tagttatcta</td><td>cacgacgggg</td><td>agtcaggcaa</td><td>ctatggatga</td><td>acgaaataga</td><td>cagatcgctg</td><td> 3300</td>

<td>agataggtgc</td><td>ctcactgatt</td><td>aagcattggt</td><td>aactgtcaga</td><td>ccaagtttac</td><td>tcatatatac</td><td> 3360</td>

<td>tttagattga</td><td>tttaaaactt</td><td>catttttaat</td><td>ttaaaaggat</td><td>ctaggtgaag</td><td>atcctttttg</td><td> 3420</td>

<td>ataatctcat</td><td>gaccaaaatc</td><td>ccttaaegtg</td><td>agttttcgtt</td><td>ccactgagcg</td><td>tcagaccceg</td><td> 3480</td>

tagaaaagat caaaggatct tcttgagatc ctttttttct gcgçgtaatç tgctgcttgc 3540 aaacaaaaaa accaccgcta ecagcggtgg tttgtttgcc ggatcaagag ctaccaactc 3600 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt 3660 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc 3720 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact 3760 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tegtgcacac 3840 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt 3900 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg 3960 gaacaggaça gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcetg 402 0 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga 4060 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt 4140 ttgctcacat gttctttcct gcgttatccc ctgattctgt · ggataaccgt attaccgcct 4200 ttgagtgagc tgataccgct cgccgcagcc gaacgaccgá gcgcagcgag tcagtgagcg 4260 aggaagcgga AGA 4273

<td> <213></td><td>vector pFPMT-CL-H6-K-El.</td>

<td> <221></td><td>hodgepodge</td>

<td> <22 3></td><td>No nucleotide</td>

<td> <220> <221> <222> <22 3></td><td>hodgepodge (1277) .. (1278) N is any nucleotide</td>

<td>ggtaccctgc tcaatctccg gaatggtgat ctgatcgttc</td><td>ctgaaaaect cgacattggc</td><td> 60</td>

<td>tccctcctga cacaggtact cgtacaggtt ccaggtaaac</td><td>gagtcgtagt tgtcgatcat</td><td> 120</td>

<td>gacaacgttc ttagaagcgg ccggcatttt gaaggtgact</td><td>aatagcctaa gaaaatattt</td><td> 180</td>

<td>aatttaattt tCattaaatt ttcctataçt cgctatttca</td><td>gcttttcatc tcatcacttc</td><td> 240</td>

<td>ataaacgata taaaccagaa, aaagaactat tttcaaacac</td><td>gctfcctcaaa agcggtatgt</td><td> 300</td>

<td>ccttccacgt ctccttagaa tctggcaagt ccgcgagggg</td><td>gatccttacc agttcatcat</td><td> 360</td>

<td>catatcccaa gccatacggt gacctgttat gtggccggga</td><td>tagattgagc aattgcagtc</td><td> 420</td>

<td>ctgcaccgtc tcatgccggc gaggcgagat ggtgaacagc</td><td>tgggagacga ggaagacaga</td><td> 480</td>

<td>tccgcagaga tcccccacgt acatagcgga acagaaagca</td><td>gccgccccaa cgagcaaatc</td><td> 540</td>

<td>gacgtggcgt cgtattgtcg tagtggggac gctggcgttc</td><td>ctagctgcga gcgtgggggt</td><td> 600</td>

<td>gagcgctacc cagcagcggg aagagttgtt ctcccgaacg</td><td>cagggcacgc acccgggggt</td><td> 660</td>

<td>gtgcatgatc atgtccgctg cctcatacac aatgcttgag</td><td>ttggagcagt cgttcgtgac</td><td> 720</td>

<td>atggtacatc ccggacacgt fcgcgcacctc atatttgtga</td><td>tggtgatggt ggtgtcctag</td><td> 780</td>

<td>tgctgctagt ggtaggaagc atagtactag tattagtagg</td><td>cttcgcatga attcccgatg</td><td> 840</td>

<td>aaggcagaga gcgcaaggag gcggtattta tagtgccatt</td><td>cccctctctg agagacccgg</td><td> 900</td>

<td>atggtagtcg agtgttafccg cagacagctt gatgtagact</td><td>ocgtgcctgc cggtcctctt</td><td> 960</td>

<td>attggcggac accagtgaga caccccggaa cttgctgttt</td><td>ttctgcaaaa tccggggtga</td><td> 1020</td>

<td>ccagtgggag cctatfctgca cacacgagcg ggacaoccca</td><td>cfcctggtgaa gagtgccaaa</td><td> 1080</td>

.X '.: «« · L · .69

<td>gtcattcttt</td><td>ttcccgtnnc</td><td>ggggcagccg</td><td>attgcatgtt</td><td>ttaggaaaat</td><td>attacctttg</td><td> 1140</td>

<td>ctacaccctg</td><td>tcagatttac</td><td>cctccacaca</td><td>tatatattcc</td><td>gtcacctcca</td><td>gggactattc</td><td> 1200</td>

<td>ttggctcgtt</td><td>gcgccgccgc</td><td>ggaagatatc</td><td>cagaagctgt</td><td>gttttccgag</td><td>agactcggtt</td><td> 1260</td>

<td>ggcgcctggt</td><td colspan="2">atatttnnag gatgtcgcgc</td><td>tgcctcacgt</td><td>cccggtaccc</td><td>aggaacgcgg</td><td> 1320</td>

<td>tgggátctcg</td><td>ggcccatcga</td><td>agactgtgct</td><td>ccagactgct</td><td>cgcccagcag</td><td>gtgtttcttg</td><td> 1380</td>

<td>attgccgcct</td><td>ctaaatagtc</td><td>cgcgcatcgc</td><td>cggtaacatt</td><td>tttccagctc</td><td>ggagtttgcg</td><td> 1440</td>

<td>tttagataca</td><td>tttctgogat</td><td>gccaaaggag</td><td>cctgcagatt</td><td>ataacctcgg</td><td>atgctgtcat</td><td> 1500</td>

<td>tcagcgcttt</td><td>taatttgacc</td><td>tccagatagt</td><td>tgctgtattt</td><td>ctgttccatt</td><td>ggctgctgga</td><td> 1560</td>

<td>cgttcgtata</td><td>actcgagtta</td><td>ttgttgcgct</td><td>ctgcctcggc</td><td>gtactggctc</td><td>atgactgact</td><td> 1620</td>

<td>gcggtcgctt</td><td>ctcgagtgtt</td><td colspan="2">ctcgcaacag gacgcctgca</td><td>ggtcatcgag</td><td>tcgagctggc</td><td> 1680</td>

<td>gccgaaactg</td><td>gcggatctga</td><td>cctccacact</td><td>gccctgtatc</td><td>tctatccacc</td><td>gggaaccgcc</td><td> 1740</td>

<td>tcctgccgtt</td><td>ccagaatgtt</td><td>gttcaagtgg</td><td>tagctctgtg</td><td>cggtcaatga</td><td>aggcgttatt</td><td> 1800</td>

<td>gccggtgaaa</td><td>tctttgggaa</td><td>gcggtttatc</td><td>ctcggggaag</td><td>attacgaaat</td><td>tcccgcgcgt</td><td> 1860</td>

<td>cgttgcgctt cctggatctc gaggaagatc gttctccgcg</td><td>tcgaggagat cgttctccgc</td><td> 1920</td>

<td>gtcgacctgc aggcatgcaa gcttctggta aacgttgtag</td><td>tactctgaaa caaggcccta</td><td> 1980</td>

<td>gcactctgat ctgtttctct tgggtagcgg tgagtggttt</td><td>attggagttc actggtttca</td><td> 2040</td>

<td>gcacatctgt catctagaca atattgttac taaatttttt</td><td>tgaactacaa ttgttcgtaa</td><td> 2100</td>

<td>ttcatctatt attatacatc ctcgtcagca atttctggca</td><td>gaoggagttt actaaçgtct</td><td> 2160</td>

<td>tgagtatgag gccgagaatc cagctctgtg gccatactca</td><td>gtcttgacag cctgctgatg</td><td> 2220</td>

<td>tggctgcgtt caacgcaata agcgtgtcct ccgactccga</td><td>gttgtgctcg ttatcgtcgt</td><td> 2280</td>

<td>tctcatcctc ggaaaaatca caogaaagaa cat.act.cacc</td><td>agtaggcttt ctggtccctg</td><td> 2340</td>

<td>gggcacggct gtttctgacg tattccggcg ttgataatag</td><td>ctcgaaagtg aacgcogagt</td><td> 2400</td>

<td>cgcgggagtc gaccgatgcc cttgagagcc ttcaaeceag</td><td>tcagctcctt ccggtgggcg</td><td> 2460</td>

<td>cggggcatga ctatcgtcgc cgcacttatg actgtcttct</td><td>ttatcatgca actcgtagga</td><td> 2520</td>

<td>caggtgccgg cagcgctctg ggtcattttc ggcgaggacc</td><td>gctttcgctg gagcgogacg</td><td> 2580</td>

<td>atgatcggcc tgtcgcttgc ggtattcgga atcttgcacg</td><td>occtcgctca agccttcgtc</td><td> 2640</td>

<td>actggtcccg ccaccaaacg tttcggcgag aagcaggcca</td><td>ttatcgccgg catggcggcc</td><td> 2700</td>

<td>gacgcgctgg gctacgtctt gctggcgttc gcgacgcgag</td><td>gctggatggc cttccccatt</td><td> 2760</td>

<td>atgattcttc tcgcttcogg cggcatcggg atgcccgcgt</td><td>tgcaggccat gctgtccagg</td><td> 2820</td>

<td>caggtagatg aogaccatca gggacagctt caaggafccgc</td><td>tcgcggctct taccagccta</td><td> 2880</td>

acttcgatca ctggaccgct gategtcacg gcgatttatg ccgcctcggc gagcacatgg 2940

ΊΟ .1 catggattgt aggcgccgcc çtataccttg tetgcctccc cgcgttgcgt 3000 ggagccgggc cacctcgacc tgaatggaag ccggcggcac ctcgctaacg 3060 tccaagaatt ggagcçaatc aattcttgcg gagaactgtg aatgcgcaaa 3120 gcagaacata tccatcgcgt ccgccatctc cagcagccgc acgcggcgca 3180

9999999999 ggggggcaaa caattcatca tttttttttt attctttttt 3240 aacgggttgg cgcggtgcat gattcaccaç ecaacccttg tcgggggggg ttgatttegg aaggaaggag aaattgccca aatcatgtcíg caagctattt taccaccaag aacacatgtg attatccgcc tacagtcaaa gaatgcacac agtaacaaag atctactgga tatcggcttt tatgacaccc cgtggatgat aaagggaagg tttgagaaga aaactcacaa taatgatttt cccccccccc ctgtcgttga ccgatacgcg acatgaatgg tgcaccatta acatctgtat

<td>tctctttgaa atttttttga</td><td>ttcggtaatc tccgaacaga</td><td>aggaagaacg</td><td> 3300</td>

<td>cacagactta gattggtata</td><td>tatacgcata tgtagtgttg</td><td>aagaaacatg</td><td> 3360</td>

<td>gtattcttaa cccaactgca</td><td>cagaacaaaa acctgcagga</td><td>aacgaagata</td><td> 3420</td>

<td>aaagctacat ataaggaacg</td><td>tgctgctact catcctagtc</td><td>ctgttgctgc</td><td> 3480</td>

<td>aatatcatgc acgaaaagca</td><td>aacaaacttg tgtgcttcat</td><td>tggatgttcg</td><td>3H40</td>

<td>gaattactgg agttagttga</td><td>agcattaggt cccaaaattt</td><td>gtttactaaa</td><td> 3600</td>

<td>gatatcttga ctgatttttc</td><td>catggagggc acagttaagc</td><td>cgctaaaggc</td><td> 3660</td>

<td>aagtacaatt ttttactctfc</td><td>cgaagacaga aaatttgctg</td><td>acattggtaa</td><td> 3720</td>

<td>ttgcagtact ctgcgggtgt</td><td>Atacagaata gcagaatggg</td><td>cagacattac</td><td> 3780</td>

<td>ggtgtggtgg gcccaggtat</td><td>tgttagcggt ttgaagcagg</td><td>cggcagaaga</td><td> 3840</td>

<td>gaacctagag gccttttgat</td><td>gttagcagaa ttgtcatgca</td><td>agggetcoct</td><td> 3900</td>

<td>gaatatacta agggtactgt</td><td>tgacattgcg aagagcgaca</td><td>aagattttgt</td><td> 3960</td>

<td>attgctcaaa gagacatggg</td><td>tggaagagat gaaggttacg</td><td>attggttgat</td><td> 4020</td>

<td>ggtgtgggtt tagatgacaa</td><td>gggagacgca ttgggtcaac</td><td>agtatag & ac</td><td> 4080</td>

<td>gtggtctcta caggatctga</td><td colspan="2">cattattatt gttggaagag gaetatttgc</td><td> 4140</td>

<td colspan="2">gatgctaagg tagagggtga acgttacaga asagcaggct</td><td>gggaagcata</td><td> 4200</td>

<td>tgcggccagc aaaactaaaa</td><td>aactgtatta taagtaaatg</td><td>catgtatact</td><td> 4260</td>

<td>attagagctt caatttaatt</td><td>atatcagtta ttacccggga</td><td>atctcggtcg</td><td> 4320</td>

tataatgacg aaaaaaaaaa 4380 aattggaaag aaaagccccc cccccccccc cccccccccg cagcgttggg tcctggccac gggtgcgcat gatcgtgctc 4440 ggacccggct aggctggcgg ggttgcctta ctggttagca gaatgaatca 4500 agcgaacgtg aagcgactgc tgctgcaaaa cgtctgcgac ctgagcaaca 4560 tcttcggttt ccgtgtttcg taaagtctgg aaacgcggaa gtcagcgccc 4620 tgttccggat ctgcatcgca ggatgctgct ggctaccctg tggaacacct 4680 taacgaagcg ctggcattga ccctgagtga ttttfcctctg gtcccgccgc 4740 at.ccat.accg ccagttgttt accctcacaa cgttccagta accgggcatg ttcatcatca 4800 gtaaçccgta tcgtgagcat cctctctcgt tfccatcggta tcattacccc catgaacaga 4860 aattccccct tacacggagg catcaagtga ccaaacagga aaaaaccgcc cttaacatgg 4920.

cccgctttat cagaagccag acattaacgc ttctggagaa actcaacgag ctggacgcgg 4980 atgaacaggc agacatctgt gaatcgcttc acgaccacgc tgatgagctt taccgcagct 5040 GCCT-cgcgcg tttcggtgat gacggtgaaa aectctgaca catgcagctc ccggagacgg 5100 tcacagcttg tctgtaageg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg 5160 gtgttggcgg GTGT-cggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata 5220 ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga 528Θ

<td>aataccgcac</td><td>agatgcgtaa</td><td>ggagaaaata</td><td>ccgcatcagg i</td><td>cgctcttccg i</td><td>cttcctcgct</td><td> 5340</td>

<td>cactgactcg</td><td>ctgcgctcgg</td><td>tcgttcggct</td><td>gcggcgagcg</td><td>gtatcagctc</td><td>actcaaaggc</td><td> 5400</td>

<td>ggtaatacgg</td><td>ttatccacag</td><td>aatcagggga</td><td>taacgcagga</td><td>aagaacatgt</td><td>gagcaaaagg</td><td> 5460</td>

<td>ccagcaaaag</td><td>gccaggaacc</td><td>gtaaaaaggc</td><td>cgcgttgctg</td><td>gcgtttttcc</td><td>ataggctccg</td><td> 5520</td>

<td>cccccctgac</td><td>gagcatcaca</td><td>aaaatcgacg</td><td>ctcaagtcag</td><td>aggtggcgaa</td><td>acccgacagg</td><td> 5580</td>

<td>actataaaga</td><td>taccaggcgt</td><td>ttccccctgg</td><td>aagctccctc</td><td>gtgcgctctc</td><td>ctgttccgac</td><td> 5640</td>

<td>cctgccgctt</td><td>accggatacc</td><td>tgtccgcctt</td><td>tctcccttcg</td><td>ggaagcgtgg</td><td>cgctttctca</td><td>S700</td>

<td>tagctcacgc</td><td>tgtaggtatc</td><td>tcagttcggt</td><td>gtaggtegtt</td><td>cgctccaagc</td><td>tgggctgtgt</td><td> 5760</td>

<td>gcacgaaccc</td><td>cccgttcagc</td><td>ccgaccgctg</td><td colspan="2">cgccttatee ggtaactatc</td><td>gtcttgagtc</td><td> 5820</td>

<td>• caacccggta</td><td>agacacgact</td><td>tatcgccact</td><td>ggcagcagcc</td><td>actggtaaca</td><td>ggattagcag</td><td> 5880</td>

<td>agcgaggtat</td><td>gtaggcggtg</td><td>ctacagagtt</td><td>cttgaagtgg</td><td>tggcctaact</td><td>acggctacac</td><td> 5940</td>

<td>tagaaggaca</td><td>gtatttggta</td><td>tctgcgctct</td><td>gctgaagcca</td><td>gttaccttcg</td><td>gaaaaagagt</td><td> 6000</td>

<td>tggtagctct</td><td>tgatccggca</td><td>aacaaaccac</td><td>cgctggtagc</td><td>ggtggttttt</td><td>ttgtttgcaa</td><td> 6060</td>

<td>gcagcagatt</td><td>acgcgcagaa</td><td>aaaaaggatc</td><td>tcaagaagat</td><td>cctttgatct</td><td>tttctacggg</td><td> 6120</td>

<td>gtctgacgct</td><td>cagtggaacg</td><td>aaaactcacg</td><td>ttaagggatt</td><td>ttggtcatga</td><td>gattatcaaa</td><td> 6180</td>

<td>aaggatcttc</td><td>acctagatcc</td><td>ttttaaatta</td><td>aaaatgaagt</td><td>tttaaatcaa</td><td>tctaaagtat</td><td> 6240</td>

<td>atatgagtaa</td><td>acttggtctg</td><td>acagttacca</td><td>atgcttaatc</td><td>agtgaggcac</td><td>ctatctcagc</td><td> 6300</td>

<td>gatctgtcta</td><td>tfctcgttcat</td><td>ecatagttgc</td><td>ctgactecec</td><td>gtcgtgtaga</td><td>taactacgat</td><td> 6360</td>

<td>acgggagggc</td><td>ttaccatctg</td><td>gccccagtgc</td><td>tgcaatgata</td><td>ccgcgagaec</td><td>cacgctcacc</td><td> 6420</td>

<td>ggctccagat</td><td>ttatcagcaa.</td><td>taaaccagcc</td><td>agccggaagg</td><td>gccgagcgca</td><td>gaagtggtcc</td><td> 6480</td>

<td>tgcaacttta</td><td>tccgcctcca</td><td>tccagtctat</td><td>taattgttgc</td><td>cgggaagcta</td><td>gagtaagtag</td><td> 6540</td>

<td>ttcgccagtt</td><td>aatagtttgc</td><td>gcaaogttgt</td><td>tgccattgct</td><td>gcaggcatcg</td><td>tggtgtcacg</td><td> 6600</td>

ctcgtcgttt ggtatggctt csttcagctc çggttcccaa cgatcaaggc gagttacatg 6660 atcccccatg ttgtgcaaaa aagcggttag ckccttcggt cctccgatcg ttgtcagaag 6720 taagttggcç gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt 6780 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga 6840 atagtgtatg cggogaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc 6900 acatagcaga actttaaaag tgctcatçat tggaaaacgt tcttcggggc gaaaactctc 6960 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac 7020 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc 7080 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca 7140 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat 7200 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt 7260 ctaagaaacc attattatca tgacattaac çtataaaaat aggcgtatca cgaggccctt 7320 tcgtcttcaa 7330 <210> 41 <211> 5202 <212> DNA < 213> pYUG5 vector <400> 41

<td>agcgcccaat</td><td>acgeaaaccg</td><td>cctctccccg</td><td>cgcgttggcc</td><td>gattcattaa</td><td>tgcagetggc</td><td> 60</td>

<td>acgacaggtt</td><td>tcccgactgg</td><td>aaagcgggca</td><td>gtgagcgcaa</td><td>cgcaattaat</td><td>gtgagttagc</td><td> 120</td>

<td>tcactcatta</td><td>ggcaccccag</td><td>gctttacact</td><td>ttatgcttcc</td><td>ggctcgtatg</td><td>ttgtgtggaa</td><td> 180</td>

<td>ttgtgagcgg</td><td>ataacaattt</td><td>cacacaggaa.</td><td>acagctatga</td><td>ccatgattac</td><td>gaatttaata</td><td> 240</td>

<td>cgactcacta</td><td>tagggaattc</td><td>gaggatcctt</td><td>caatatgcgc</td><td>acatacgctg</td><td>ttatgttcaa</td><td> 300</td>

<td>ggrtcccttcg</td><td>tttaagaacg</td><td>aaagcggtct</td><td>tccttttgag</td><td>ggatgtttca</td><td>agttgttcaa</td><td> 360</td>

<td>atctatcaaa</td><td>tttgcaaatc</td><td>cccagtcfcgt</td><td>atctagagcg</td><td>ttgaatoggt</td><td>gatgcgattt</td><td> 420</td>

<td>gttaattaaa</td><td>ttgatggtgt</td><td>caccattacc</td><td>aggtctagat</td><td>attackcaatgg</td><td>caaactgagc</td><td> 480</td>

<td>acaacaatac</td><td>eagtccggat</td><td>caactggcac</td><td>catctctccc</td><td>gtagtctcat</td><td>ctaatttttc</td><td> 540</td>

<td>ttccggatga</td><td>ggttecagat</td><td>Ataccgcaac</td><td>acctttatta</td><td>tggtttccct</td><td>gagggaataa</td><td> 600</td>

<td colspan="2">tagaatgtcc cattcgaaat</td><td>caccaattct</td><td>aaacctgggc</td><td>gaattgtatt</td><td>tcgggtttgt</td><td> 660</td>

<td>taactcgttc</td><td>cagtcaggaa</td><td>tgttccacgt</td><td>gaagctatct</td><td>tccagcaaag</td><td>tctccacttc</td><td> 720</td>

<td>ttcatcaaat</td><td>tgtggagaat</td><td>actcccaatg</td><td>ctcttatcta</td><td>tgggacttcc</td><td>gggaaacaca</td><td> 780</td>

<td>gtaccgatac</td><td>ttcccaattc</td><td>gtcttcagag</td><td>ctcattgttt</td><td>gtttgaagag</td><td>theretaatcaaa</td><td> 840</td>

Ί3

<td>gaatcgtttt ctcaaaaaaa ttaatatctt aactgatagt ttgatcaaag</td><td>gggcaaaacg</td><td> 900</td>

<td>taggggcaaa caaacggaaa aatcgtttct caaattttct gatgccaaga</td><td>actetaacca</td><td> 960</td>

<td>gtcttatctá aaaattgcct tatgatccgt ctctccggtt acagcctgtg</td><td>taactgatta</td><td> 1020</td>

<td>atcctgcctt tctaatcacc attctaatgt tttaattaag ggattttgtc</td><td>ttcattaacg</td><td> 1080</td>

<td>gctttcgctc ataaaaatgt tatgacgttt tgcccgcagg cgggaaacca</td><td>tccacttcac</td><td> 1140</td>

<td>gagactgatc tcctctgccg gaacaccggg catctccaac ttataagttg</td><td>gagaaataag</td><td>l? 00</td>

<td>agaatttcag attgagagaa tgaaaaaaaa aaaccctgaa aaaaaaggtt</td><td>gaaaccagtt</td><td> 1260</td>

<td>ccctgaaatt attcccctac ttgactaata agtatataaa gacggtaggt</td><td>attgattgta</td><td> 1320</td>

<td>attctgtaaa tctatttctt aaacttctta aattctactt ttatagttag</td><td>tctttttttt</td><td> 1380</td>

<td>agttttaaaa caccaagaac ttagtttcga ataaacacac ataaacaaac</td><td>accatgagat</td><td> 1440</td>

<td>ttccttcaat ttttactgca gttttattcg cagcatcctc cgcattagct</td><td>gctccagtca</td><td> 1500</td>

<td>acactacaac agaagatgaa acggcacaaa ttccggctga agctgtcatc</td><td>ggttactcag</td><td> 1560</td>

<td>atttagaagg ggatttcgat gttgctgttt tgccattttc caacagcaca</td><td>aataacgggt</td><td> 1620</td>

<td>tattgtttat aaatactact attgccagca ttgctgctaa agaagaaggg</td><td>gtatctctag</td><td> 1680</td>

<td>ataaaaggcç tgtcgacggt accagatctc gacttggttg aacacgttgc</td><td>caaggcttaa</td><td> 1740</td>

<td>gtgaatttac tttaaagtct tgcatttaaa taaattttct ttttatagct</td><td>ttatgactta</td><td> 1800</td>

<td>gtttcaattt atatactatt ttaatgacat tttcgattca ttgattgaaa</td><td>gctttgtgtt</td><td> 1860</td>

<td>ttttcttgat gcgctattgc attgttcttg tctttttcgc cacatgtaat</td><td>atctgtagta</td><td> 1920</td>

gatacctgat acattgtgga tgctgagtga aattttagtt aataatggag gcgctcttaa 1980 taattttggg gatattggct ttttttttta aagtttacaa atgaattttt tcegccagga 2040 taacgattct gaagttactc ttagcgttcc tatcggtaca gccatcaaat çatgcctata 2100 aatcatgcct atatttgcgt gcagtcagta tcatctacat gaaaaaaact cccgcaattt 2160 cttatagaat acgttgaaaa ttaaatgtac gcgccaagat aagataacat atatctagct 2220 agatgcagta atatacacag attcccgcgg acgtgggaag gaaaaaatta gataacaaaa 2280 tctgagtgat atggaaattc ogctgtatag ctcatatctt tcccttcaac 2340 accagaaatg taaaaatctt gttacgaagg atctttttgc taatgtttct cgctcaatcc tcatttcttc 2400 cctacgaaga gtcaaatcta cttgttttct gccggtatca agatccatat cttctagttt 2460 caccatcaaa gtccaatttc tagtatacag tttatgtccc aacgtaacag acaatcaaaa 2S20 ttggaaagga taagtatcct tcaaagaatg attctgcgct ggctcctgaa ccgcctaatg 2580 ggaacagaga agtccaaaac gatgctataa gaaccagaaa taaaacgata aaaccatacc 2640 aggatccaag cttggcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg 2700 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt 2760 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgggaaattg 2B20 taaacgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta 2B80 accaataggc cgaaatcggc aaaatccctt ataaafccaaa agaatagacc gagatagggt 2940 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca 3000 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa 3060 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat 3120 ttagagcttg acggggaaag ccggcgaaçg tggcgagaaa ggaagggaag 3180 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg 3240

<td>ccgcgcttaa</td><td>tgcgccgcta</td><td>cagggcgcgt</td><td>caggtggcac</td><td>ttttcgggga;</td><td>aatgtgcgcg</td><td> 3300</td>

<td>gaacccctat</td><td>ttgtttattü</td><td>ttctaaatac</td><td>attcaaatat</td><td>gtatccgctc</td><td>atgagacaat</td><td> 3360</td>

<td>aaccctgata</td><td>aatgcttcaa</td><td>taatattgaa</td><td>aaaggaagag</td><td>tatgagtatt</td><td>caacatttcc</td><td> 3420</td>

<td>gtgtcgccct</td><td>tattcccttt</td><td>tttgcggcat</td><td>tttgccttcc</td><td>tgtttttgct</td><td>cacccagaaa</td><td> 3480</td>

<td>cgctggtgaa</td><td>agtaaaagat</td><td>gctgaagatc</td><td>agttgggtgc</td><td>acgagtgggt</td><td>tacatcgaac</td><td> 3540</td>

<td>3. t C t C3.5</td><td>ra Γτ / ^ ίΤΓτΙ 'aan ss 3</td><td>atccttgaga</td><td>gfctttcgocc</td><td>cgaagaacgt</td><td>tttccaatga</td><td> 3600</td>

<td>tgagcacttt</td><td>taaagttctg</td><td>ctatgtggcg</td><td>cggtattatc</td><td>• ccgtattgac</td><td>gccgggcaag</td><td> 3660</td>

<td>agcaactcgg</td><td>tcgccgcata</td><td>cactattctc</td><td>agaatgactt</td><td>ggttgagtac</td><td>tcaccagtca</td><td> 3720</td>

<td>cagaaaagca</td><td>tcttacggat</td><td>ggcatgacag</td><td>taagagaatt</td><td>atgcagtgct</td><td>g-cçataacca</td><td> 3780</td>

<td>tgagt.gat.aa</td><td>cactgcggcc</td><td>aacttacttc</td><td>tgacaacgat</td><td>cggaggaccg</td><td>aaggagctaa</td><td> 3840</td>

<td>ccgctttttt</td><td>gcacaaeatg</td><td>ggggatcatg</td><td>taactcgccfc</td><td>tgatcgttgg</td><td>gaaccggagc</td><td> 3900</td>

<td>tgaatgaagc</td><td>cataccaaac</td><td>gacgagcgtg</td><td>acaccacgat</td><td>gcctgtagca</td><td>atggcaacaa</td><td> 3960</td>

<td>cgttgcgcaa</td><td>actattaact</td><td>ggcgaactac</td><td>ttactctagc</td><td>ttcccggcaa</td><td>caattaatag</td><td> 4020</td>

<td>actggatgga</td><td>ggcggataaa</td><td>gttgcaggac</td><td>cacttctgcg</td><td>ctcggccctt</td><td>ccggctggct</td><td> 4080</td>

<td>ggtttattgc</td><td>tgataaatct</td><td>ggagccggtg</td><td>agcgtgggtc</td><td>tcgcggtatc</td><td>attgcagcac</td><td> 4140</td>

<td>tggggccaga</td><td>tggtaagccc</td><td>tcccgtatcg</td><td>tagttatcta</td><td>cacgacgggg</td><td>agtcaggcaa</td><td> 4200</td>

<td>ctatggatga</td><td>acgaaataga</td><td>cagatcgctg</td><td>agataggtgc</td><td>ctcactgatt</td><td>aagcattggt</td><td> 4260</td>

<td>aactgtcaga</td><td>ccaagtttac</td><td>tcatatatac</td><td>tttagattga</td><td>tttaaaactt</td><td>catttttaat</td><td> 4320</td>

<td>ttaaaaggat</td><td>ctaggtgaag</td><td>atectttttg</td><td>ataatctcat</td><td>gaceaaaatc</td><td>ccttaacgtg</td><td> 4360</td>

<td>agttttcgtt</td><td>ccactgagcg</td><td>tcagaccccg</td><td>tagaaaagat</td><td>caaaggatct</td><td>tcttgagatc</td><td> 4440</td>

<td>ctttttttct</td><td>gcgogtaafcc</td><td>tgctgcttgc</td><td>aaacaaaaaa</td><td>accaccgcta</td><td>. ecagcggtgg</td><td> 4500</td>

<td>tttgtttgcc</td><td>ggatcaagag</td><td>ctaccaactc</td><td>tttttccgaa</td><td>ggtaactggc</td><td>ttcagcagag</td><td> 4560</td>

<td>cgcagatacc</td><td>aa & tactgtc</td><td>cttctagtgt</td><td>agccgtagtt</td><td>aggccaccac</td><td>ttcaagaact</td><td> 4620</td>

<td>ctgtagcacc</td><td>gcctacatac</td><td>ctcgctctgc</td><td>taatcctgtt</td><td>accagtggct</td><td>gctgccagtg</td><td> 4660</td>

<td>gcgataagtc</td><td>gtgtcttacc</td><td>gggttggact</td><td>caagacgata</td><td>gttaccggat</td><td>aaggcgcagc</td><td> 4740</td>

<td>ggtcgggctg</td><td>aaeggggggt</td><td>tcgtgcacac</td><td>agcccagctt</td><td>ggagcgaacg</td><td>acctacaccg</td><td> 4800</td>

<td>aactgagata</td><td>cctacagcgt</td><td>gagcattgag</td><td>aaagcgccac</td><td>gcttcocgaa</td><td>gggagaaagg</td><td> 4860</td>

<td>cggacaggta</td><td>tccggtaagc</td><td>ggcagggtcg</td><td>gaacaggaga</td><td>gcgcacgagg</td><td>gagcttccag</td><td> 4920</td>

<td>ggggaaacgc</td><td>ctggtatctt</td><td>tatagtcetg</td><td>tcgggtttcg</td><td>ccacctetga</td><td>cttgagcgtc</td><td> 4980</td>

<td>gatttttgtg</td><td>atgctcgtca</td><td>ggggggcgga</td><td>gcctatggaa</td><td>aaacgccagc</td><td>aacgcggcct</td><td> 5040</td>

<td>ttttacggtt</td><td>cctggccttt</td><td>tgctggcctt</td><td>ttgctcacat</td><td>gttctttcct</td><td>gcgttatccc</td><td> 5100</td>

<td>ctgattctgt</td><td>ggataaccgt</td><td>attaccgcct</td><td>ttgagtgagc</td><td>tgataccgct</td><td>cgccgcagcc</td><td> 5160</td>

<td>gaacgaccga</td><td>gcgcagcgag</td><td>tcagtgagcg</td><td>aggaagcgga</td><td>ag</td><td></td><td> 5202</td>

<210> 42 <211> 5613 <212> DNA <213> vetor pYIG5ElH6 <400> 42

<td>ggatccttca</td><td>atatgcgcac</td><td>atacgctgtt</td><td>atgttcaagg</td><td>tcccttcgtt</td><td>taagaacgaa</td><td> 60</td>

<td>agcggtcttc</td><td>cttttgaggg</td><td>atgtttcaag</td><td>ttgttcaaat</td><td>ctatcaaatt</td><td>tgcaaatccc</td><td> 120</td>

<td>cagtctgtat</td><td>ctagagegtt</td><td>gaatcggtga</td><td>tgcgatttgt</td><td>taattaaatt</td><td>gatggtgtca</td><td> 180</td>

<td>ccattaccag</td><td>gtctagatat</td><td>accaatggca</td><td>aactgagcac</td><td>aacaatacca</td><td>gtccggatca</td><td> 240</td>

<td colspan="2">actggcacca -tctctcccgt</td><td>agtctcatct</td><td>aattttbctt</td><td>ccggatgagg</td><td>ttccagatat</td><td> 300</td>

<td>accgcaacac</td><td>ctttattafcg</td><td>gttfcceçfcga</td><td>gggaataata</td><td>gaatgtccca</td><td>ttcgaaatca</td><td> 360</td>

<td>ccaattctaa</td><td>acctgggcga</td><td>attgtatttc</td><td>gggtttgtta</td><td>actcgttcca</td><td>gtcaggaatg</td><td> 420</td>

<td>ttecaegtga</td><td>agcfcafccttc</td><td>cagcaaagbe</td><td>tccacttctt</td><td>catcaaattg</td><td>tggagaatac</td><td> 480</td>

<td>tcccaatgct</td><td>cttatetatg</td><td>ggacttccgg</td><td>gaaacacagt</td><td>aocgataett</td><td>cocaattcgt</td><td> 540</td>

<td>cttcagagct</td><td>cafctgtfctgt</td><td>ttgaagagac</td><td>taatcaaaga</td><td>atcgttttct</td><td>caaaaaaatt</td><td> 600</td>

<td>aatatcttaa</td><td>ctgatagttt</td><td colspan="2">gatcaaaggg gcaaaacgta</td><td>ggggcaaaca</td><td>aacggaaaaa</td><td> 660</td>

<td>tcgtttctca</td><td>aafctttctga</td><td>tgccaagaac</td><td>tctaaccagt</td><td>cttatctaaa</td><td>aattgcctta</td><td> 720</td>

<td>tgatcegtct</td><td>ctccggttac</td><td colspan="2">agcctgtgta actgattaat</td><td>cctgcctttc</td><td>taatcaccat</td><td> 780</td>

<td>tctaatgttt</td><td>taattaaggg</td><td colspan="2">attttgbctt -cattaacggc</td><td>fcttcgefccat</td><td>aaaaatgtta</td><td> 840</td>

<td>tgacgttttg</td><td>cccgcaggcg</td><td colspan="2">ggaaaccatc cacttcaega</td><td>gactgatctc</td><td>ctctgccgga</td><td> 900</td>

acaccgggca tctccaactt ataagttgga gaaataagag aatttcagat tgagagaatg 960 aaaaaaaaaa accctgaaaa aaaaggttga aaccagttcc ctgaaattat tcccctactt 1020 gactaataag tatataaaga cggtaggtat tgattgtaat tctgtaaatc tatttcttaa 1080 a-ctt.ctt.aaa ttctactttt atagttagtc ttttttttag ttttaaaaca ccaagaactt 1140 agtttcgaat aaacacacat aaacaaacac catgagattt ccttcaattt ttactgcagt 1200 tttattcgca gcatcctccg cattagctgc tccagtcaac actacaacag aagatgaaac 1260 ggcacaaatt ccggctgaag ctgtcatcgg ttacttagat ttagaagggg atttcgatgt 1320

<td>tgctgttttg</td><td>ccattttcca</td><td>acagcacaaa</td><td>taacgggtta ’</td><td>ttgtttataa ;</td><td>atactactat</td><td> 1380</td>

<td>tgccagcatt</td><td>gctgctaaag</td><td>aagaaggggt</td><td>atctctagat .</td><td>aaaaggtatg </td><td>aggtgcgcaa</td><td> 1440</td>

<td>cgtgtccggg</td><td>atgtaccatg</td><td>tcacgaacga</td><td>ctgctccaac</td><td>tcaagcattg</td><td>tgtatgaggc</td><td> 1500</td>

<td>agcggacatg</td><td>atcatgcaca</td><td>cccccgggtg</td><td>cgtgccctgc</td><td>gttcgggaga</td><td>acaactcttc</td><td>1S60</td>

<td>ccgctgctgg</td><td>gtagcgctca</td><td>cccccacgct</td><td>cgcagctagg</td><td>aacgccagcg</td><td>tccccactac</td><td> 1620</td>

<td>gacaatacga</td><td>cgccacgtcg</td><td>atttgctcgt</td><td>tggggcggct</td><td>gctttctgtt</td><td>ccgctatgta</td><td> 1680</td>

<td>cgtgggggat</td><td>ctctgcggat</td><td>ctgtcttcct</td><td>cgtctcccag</td><td>ctgttcacca</td><td>tctcgcctcg</td><td> 1740</td>

<td>ccggcatgag</td><td>acggtgcagg</td><td>actgcaattg</td><td>ctcaatctat</td><td>cccggccaca</td><td>taacaggtca</td><td> 1800</td>

<td>ccgtatggct</td><td>tgggatatga</td><td>tgatgaactg</td><td>gcaccaccac</td><td>catcaccatt</td><td>aaagatctcg</td><td> 1860</td>

<td>acttggttga</td><td>acacgttgcc</td><td>aaggcttaag</td><td>tgaatttact</td><td>ttaaagtctt</td><td>gcatttaaat</td><td> 1920</td>

<td>aaattttctt</td><td>tttatagctt</td><td>tatgacttag</td><td>tttcaattta</td><td>tatactattt</td><td>taatgacatt</td><td> 1986</td>

<td>ttcgattcat</td><td>tgattgaaag</td><td>ctttgtgttt</td><td>tttcttgatg</td><td>cgctattgca</td><td>ttgttcttgt</td><td> 2040</td>

<td>ctttttçgcc</td><td>acatgtaata</td><td>tctgtagtag</td><td>atacctgata</td><td>cattgtggat</td><td>gctgagtgaa</td><td> 2100</td>

<td>attttagtta</td><td>ataatggagg</td><td>cgct-cttaat</td><td>aattttgggg</td><td>atattggctt</td><td>ttttttttaa</td><td> 2160</td>

<td>agtttacaaa</td><td>tgaatttttt</td><td>ccgocaggat</td><td>aacgattctg</td><td>aagttactct</td><td>tagcgttcct</td><td> 2220</td>

<td>atcggtacag</td><td>ccatcaaatc</td><td>atgcctataa</td><td>atcatgccta</td><td>tatttgcgtg</td><td>cagtcagtat</td><td> 2286</td>

<td>catctacatg</td><td>aaaaaaactc</td><td>ccgcaatttc</td><td>ttatagaata</td><td>cgttgaaaat</td><td>taaatgtacg</td><td> 2340</td>

<td>cgccaagata</td><td>agataacata</td><td>tatctagcta</td><td>gatgcagtaa</td><td>tatacacaga</td><td>ttcccgcgga</td><td> 2400</td>

<td>cgtgggaagg</td><td>aaaaaattag</td><td>ataacaaaat</td><td>ctgagtgata</td><td>tggaaattcc</td><td>gctgtatagc</td><td> 2460</td>

<td>tcatatcttt</td><td>cccttcaaca</td><td>ccagaaatgt</td><td>aaaaatcttg</td><td>ttacgaagga</td><td>tctttttgct</td><td> 2520</td>

<td>aatgtttctc</td><td>gctcaatcct</td><td>cattfcctbcc</td><td>ctacgaagag</td><td>tcaaatctac</td><td>ttgttttctg</td><td> 2580</td>

<td>ccggtatcaa</td><td>gatccatatc</td><td>ttctagtttc</td><td>accatcaaag</td><td>tccaatttct</td><td>agtatacagt</td><td> 2640</td>

<td>ttatgtccca</td><td>acgtaacaga</td><td>caatcaaaat</td><td>tggaaaggat</td><td>aagtatcctt</td><td>caaagaatga</td><td> 2700</td>

I

<td>ttctgcgctg gctcctgaac cgcctaatgg gaacagagaa gtccaaaacg atgctataag</td><td> 2760</td>

<td>aaccagaaat aaaacgataa aaccatacca ggatccaagc ttggcactgg ccgtcgtttt</td><td> 2820</td>

<td>acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc</td><td> 2880</td>

<td>ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt occaacagtt</td><td> 2940</td>

<td>gcgcagcctg aatggcgaat gggaaattgt aaacgttaat attttgttaa aattcgcgtt</td><td> 3000</td>

<td>aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta</td><td> 3060</td>

<td>taaãtcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc</td><td> 3120</td>

<td>actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg</td><td> 3180</td>

<td>cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact</td><td> 3240</td>

<td>aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt</td><td> 3300</td>

<td>ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc</td><td> 3360</td>

<td>ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc</td><td> 3420</td>

<td>aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca</td><td> 3480</td>

<td>ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa</td><td> 3540</td>

<td>aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt</td><td> 3600</td>

<td>fetgccfetcct gttttfegctc acccagaaac gctggtgaaa gtãaaagatg ctgaagatca</td><td> 3660</td>

<td>gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccfctgagag</td><td> 3720</td>

<td>ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc</td><td> 3780</td>

ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca 3840 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt 3900 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct 3360 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt 4020 aaetcgeett gategttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga 4060 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact 4140 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc 4200 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga 4260 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt 4320 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga 4380 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact 4440 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga 4SO0 taatctcatg aocaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt 4S60

<td>agaaaagatc</td><td>aaaggatctt</td><td>cttgagatcc</td><td>tttttttctg</td><td>cgcgtaatct gctgcttgca</td><td> 4620</td>

<td>aacaaaaaaa</td><td>ccaccgctac</td><td>cagcggtggt</td><td>ttgtttgccg</td><td>gatcaagagc taccaactct</td><td> 4680</td>

<td>ttttccgaag</td><td>gtaactggct</td><td>tcagcagagc</td><td>gcacatacca</td><td>aatactgtcc ttctagtgta</td><td> 4740</td>

<td>gccgtagtta</td><td>ggccaocact</td><td>tcaagaactc</td><td>tgtagcaccg</td><td>cctacatacc tcgctctgct</td><td> 4800</td>

<td>aatcctgtta</td><td>ccagtggctg</td><td>ctgccagtgg</td><td>cgataagtcg</td><td>tgtcttaccg ggttggactc</td><td> 4860</td>

<td>aagacgatag</td><td>ttaccggata</td><td>aggcgcagcg</td><td>gtcgggctga</td><td>acggggggtt cgtgcacaca</td><td> 4320</td>

<td>gcccagcttg</td><td>gagcgaacga</td><td>cctacaccga</td><td>actgagatac</td><td>ctacagcgtg agcattgaga</td><td> 4980</td>

<td>aagcgccacg</td><td>cttcccgaag</td><td>ggagaaaggc</td><td>ggacaggtat</td><td>ccggtaagcg gcagggtcgg</td><td> 5040</td>

<td>aacaggagag</td><td>cgcacgaggg</td><td>agcttccagg</td><td>gggaaacgcc</td><td>tggtatcttt atagtcctgt</td><td> 5100</td>

<td>cgggtttcgc</td><td>cacctctgac</td><td>ttgagcgtcg</td><td>atttttgtga</td><td>tgctcgtcag gggggcggag</td><td> 5160</td>

<td>cctatggaaa</td><td>aacgccagca</td><td>acgcggcctt</td><td>tttacggttc</td><td>ctggcctttt gctggccttt</td><td> 5220</td>

<td>tgctcacatg</td><td>ttctttcctg</td><td>cgttatcccc</td><td>tgattctgtg</td><td>gataaccgta ttaccgoctt</td><td> 5280</td>

<td>tgagtgagct</td><td>gataccgctc</td><td>gccgcagccg</td><td>aacgaccgag</td><td>cgcagcgagt cagtgagcga</td><td> 5340</td>

<td>ggaagcggaa</td><td>gagcgcccaa</td><td>tacgcaaacc</td><td>gcctctcccc</td><td>gcgcgttggc cgattcatta</td><td> 5400</td>

<td>atgcagctgg</td><td>cacgacaggt</td><td>ttcccgactg</td><td>gaaagcgggc</td><td>agtgagcgca acgcaattaa</td><td> 5460</td>

<td>tgtgagttag</td><td>ctcactcatt</td><td>aggcacccca</td><td>ggctttacac</td><td>tttatgefcfcc eggctcgtat</td><td>Kton</td>

<td>gttgtgtgga</td><td>attgtgagcg</td><td>gataacaatt</td><td>tcacacagga</td><td>aacagctatg accatgatta</td><td> 5580</td>

<td>cgaatttaat</td><td>acgactcact</td><td>atagggaatt</td><td>cga</td><td></td><td> 5613</td>

<210> 43 <211> 13020 <212> DNA <213> vetor pSYl

<td>atcgataagc</td><td>ttttcaattc</td><td>aattcatcat</td><td>ttttttttta</td><td>ttcttttttt</td><td>tgatttcggt</td><td> 60</td>

<td>ttctttgaaa</td><td>ttttttfcgat</td><td>tcggtaatct</td><td>ccgaacagaa</td><td>ggaagaacga</td><td>aggaaggagc</td><td> 120</td>

<td>acagacttag</td><td>attggtatat</td><td>atacgcatat</td><td>gtagtgttga</td><td>agaaacatga</td><td>aattgcccag</td><td> 180</td>

<td>tattcttaac</td><td>ccaactgcac</td><td>agaacaaaaa</td><td>cctgcaggaa</td><td>acgaagataa</td><td>atcatgtcga</td><td> 240</td>

<td>aagctacata</td><td>taaggaacgt</td><td>gctgctactc</td><td>atcctagtcc</td><td>tgttgctgcc</td><td>aagctattta</td><td> 300</td>

<td>atatcatgca</td><td>cgaaaagcaa</td><td>acaaacttgt</td><td>gtgcttcatt</td><td>ggatgttcgt</td><td>accaccaagg</td><td> 360</td>

<td colspan="2">aattactgga gttagttgaa</td><td>gcattaggtc</td><td>ccaaaatttg</td><td>tttactaaaa</td><td>acacatgtgg</td><td> 420</td>

<td colspan="2">atatcttgac tgatttttcc</td><td>atggagggca</td><td>cagttaagcc</td><td colspan="2">gctaaaggca ttatccgcca</td><td> 480</td>

<td colspan="2">agtacaattt tttactcttc</td><td>gaagacagaa</td><td>aatttgctga</td><td colspan="2">cattggtaat acagtcaaat</td><td> 540</td>

tgcagtactc tgcgggtgta tacagaatag cagaatgggc agacattacg aatgcacacg 600 gtgtggtggg cccaggtatt gttagcggtt tgaagcaggc ggcagaagaa gtaacaaagg 660 aacctagagg ccttttgatg ttagcagaat tgtcatgcaa gggctcccta tctactggag 720 aatatactaa gggtactgtt gacattgega agagcgacaa agattttgtt atcggcttta 780 ttgctcaaag agacatgggt ggaagagatg aaggttacga ttggttgatt atgacacccg 840 gtgtgagttt agatgacaag ggagaogcat tgggtcaaca gtatagaacc gtggatgatg 900 tggtctctac aggatctgac attattattg ttggaagagg actatttgca aagggaaggg 960 atgctaaggt agagggtgaa egttacagaa aagcaggctg ggaagcatat ttgagaagat 1020

<td>gcggccagca</td><td>aaactaaaaa</td><td>actgtattat</td><td>aagtaaatgc</td><td>atgtatacta i</td><td>aactcacaaa</td><td> 1080</td>

<td>ttagagcttc</td><td>aatttaatta</td><td>tatcagttat</td><td>tacccgggaa</td><td>tctcggtcgt</td><td>aatçattttt</td><td> 1140</td>

<td>ataatgacga</td><td>aaaaaaaaaa</td><td>attggaaaga</td><td>aaaagcttta</td><td>atgcggtagt</td><td>ttatcacagt</td><td> 1200</td>

<td>taaattgcta</td><td>acgcagtcag</td><td>gcaccgtgta</td><td>tgaaatctaa</td><td>caatgcgctc</td><td>atcgtcatcc</td><td> 1260</td>

<td>tcggcaccgt</td><td>caccctggat</td><td>getgtaggca</td><td>taggcttggt</td><td>tatgccggta</td><td>etgccgggcc</td><td> 1320</td>

<td>tcttgcggga</td><td>tatcgtccat</td><td>tccgacagca</td><td>tcgccagtca</td><td>ctatggcgtg</td><td>ctgctagcgc</td><td> 1380</td>

<td>tatatgcgtt</td><td>gatgcaattt</td><td>ctatgcgcac</td><td>ccgttctcgg</td><td>agcactgtcc</td><td>gaccgctttg</td><td> 1440</td>

<td>gcegccgccc</td><td>agtcctgetc</td><td>gcttcgctac</td><td>ttggagccac</td><td>tatcgactac</td><td>gcgatcatgg</td><td> 1500</td>

<td>cgaccacacc</td><td>cgtcctgtgg</td><td>atcctctacg</td><td>ccggacgcat</td><td>cgtggccggc</td><td>atcaccggcg</td><td> 1560</td>

<td>ccacaggtgc</td><td>ggttgctggc</td><td>ccctatatcg</td><td>ccgacatcac</td><td>cgatggggaa</td><td>gatcgggctc</td><td> 1620</td>

<td>gccacttcgg</td><td>gctcatgagc</td><td>gcttgtttcg</td><td>gcgtgggtat</td><td>ggtggcaggc</td><td>cccgtggccg</td><td> 1680</td>

<td>ggggactgtt</td><td>gggcgccatc</td><td>tccttgcatg</td><td>caccattcct</td><td>tgcggcggcg</td><td>gtgctcaacg</td><td> 1740</td>

<td>gcctcaacct</td><td>actactgggc</td><td>tgcttcctaa</td><td>tgcaggagtc</td><td>gcataaggga</td><td>gagcgfccgac</td><td> 1800</td>

<td>cgatgccctt</td><td>gagagccttc</td><td>aacccagtca</td><td>gctccttccg</td><td>gtgggcgcgg</td><td>ggcatgacta</td><td> 1860</td>

<td>tcgtcgccgc</td><td>acttatgact</td><td>gtcttcttta</td><td>tcatgcaact</td><td>cgtaggacag</td><td>gtgccggcag</td><td> 1920</td>

<td>cgctctgggt</td><td>cattttcggc</td><td>gaggaccgct</td><td>ttcgctggag</td><td>cgcgacgatg</td><td>atcggectgt</td><td> 1980</td>

<td>cgcttgcggt</td><td>attcggaatc</td><td>ttgcacgccc</td><td>tcgctcaagc</td><td>cttcgtcact</td><td>ggtcccgcca</td><td> 2040</td>

<td>ccaaacgttt</td><td>cggcgagaag</td><td>caggccatta</td><td>tcgccggcat</td><td>ggcggccgac</td><td>gcgctgggct</td><td> 2100</td>

<td>acgtcttget</td><td>ggcgttcgcg</td><td>acgcgagg-ct</td><td>ggatggcctt</td><td>ccccattatg</td><td>attcttcfccg</td><td> 2160</td>

<td>cttccggcgg</td><td>catcgggafcg</td><td>cccgcgttgc</td><td>aggccatgct</td><td>gtccaggcag</td><td>gtagatgacg</td><td> 2220</td>

<td>accatcaggg</td><td>acagcttcaa</td><td>ggatcgctcg</td><td>cggctcttac</td><td>cagcctaact</td><td>togateactg</td><td> 2280</td>

<td>gaccgctgat</td><td>cgtcacggcg</td><td>atttatgccg</td><td>cctcggcgag</td><td>cacatggaac</td><td>gggttggcat</td><td> 2340</td>

<td>ggattgtagg</td><td>cgccgcccta</td><td>taccttgtct</td><td>gcctccccgc</td><td>gttgcgtcgc</td><td>ggtgçatgga</td><td> 2400</td>

<td>gccgggccac</td><td>ctcgacctga</td><td>atggaagccg</td><td>gcggcacctc</td><td>gctaacggat</td><td>tcaccactcc</td><td> 2460</td>

<td>aagaattgga</td><td>gccaatcaat</td><td>tcttgcggag</td><td>aactgtgaat</td><td>gcgcaaacca</td><td>acccttggca</td><td> 2520</td>

<td>gaacatatcc</td><td>atcgcgtccg</td><td>ccatctccag</td><td>cagccgcacg</td><td>cggcgcatct</td><td>cgggcagcgt</td><td> 2580</td>

<td>tgggtcctgg</td><td>ccacgggtgc</td><td>gcatgatcgt</td><td>gctcctgtcg</td><td>ttgaggaccc</td><td>ggctaggctg</td><td> 2640</td>

<td>gcggggttgc</td><td>cttactggtt</td><td>agcagaatga</td><td>atcaccgata</td><td>cgcgagcgaa</td><td>cgtgaagcga</td><td> 2700</td>

<td>ctgctgctgc</td><td>aaaacgtctg</td><td>cgacctgagc</td><td>aacaacatga</td><td>atggtcttcg</td><td>gtttccgtgt</td><td> 2760</td>

<td>ttcgtaaagt</td><td>ctggaaacgc</td><td>ggaagtcagc</td><td>gccctgcacc</td><td>attatgttcc</td><td>ggatctgcat</td><td> 2820</td>

<td>cgcaggatgc</td><td>tgctggctac</td><td>cctgtggaac</td><td>acctacatct</td><td>gtattaacga</td><td>agcgctggca</td><td> 2880</td>

<td>ttgaccctga</td><td>gtgatttttc</td><td>tctggtcccg</td><td>ccgcatçcat</td><td>accgccagtt</td><td>gtttaccctc</td><td> 2940</td>

<td>acaacgttcc</td><td>agtaaccggg</td><td>catgttcatc</td><td>atcagtaacc</td><td>cgtatcgtga</td><td>gcatcctctc</td><td> 3000</td>

<td>tcgtttcatc</td><td>ggtatcatta</td><td>cccccatgaa</td><td>cagaaattcc</td><td>cccktacacg</td><td>gaggcatcaa</td><td> 3060</td>

<td>gtgaccaaac</td><td>aggaaaaaac</td><td>cgcccttaac</td><td>atggcccgct</td><td>ttatcagaag</td><td>ccagacatta</td><td> 3120</td>

<td>acgcttctgg</td><td>agaaactcaa</td><td>cgagctggac</td><td>gcggatgaac</td><td>aggcagacat</td><td>ctgtgaatcg</td><td> 3180</td>

<td>cttcacgacc</td><td>acgctgatga</td><td>gctttaccgc</td><td>agctgcctcg</td><td>cgcgtttcgg</td><td>tgatgacggt</td><td> 3240</td>

<td>gaaaacctct</td><td>gacacatgca</td><td>gcteecggag</td><td></td><td>Cfct^fcCtCjtS</td><td>agcggtgccg</td><td> 3300</td>

<td>ggagcagaca</td><td>agcccgtcag</td><td>ggcgcgtcag</td><td>cgggtgttgg</td><td>cgggtgtcgg</td><td>ggcgcagcca</td><td> 3360</td>

<td>tgacccagtc</td><td>acgtagcgat</td><td>agcggagtgt</td><td>atactggctt</td><td>aactatg-cgg</td><td>catcagagca</td><td> 3420</td>

<td>gattgtactg</td><td>agagtgcacc</td><td>atatgcggtg</td><td>tgaaataccg</td><td>cacagatgcg</td><td>taaggagaaa</td><td> 3460</td>

<td>ataccgcatc</td><td>aggcgctctt</td><td>ccgcttcctc</td><td>gctcactgac</td><td>tcgctgcgct</td><td>cggtcgttcg</td><td> 3540</td>

gctgcggoga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg 3600 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa 3660 ggccgogttg ctggcgtttt tccataggct ccgccccoct gacgagcatc acaaaaatcg 3720 acgctcaagt cagaggtgge gaaacccgac aggactataa agataceagg cgtttccccc 3780 tggaagctcc ctegtgcgct ctcctgttcc gaocctgccg cttaocggat acctgtccgc 3840 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc 3900 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgtfcc agcccgaccg 3960 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc 4020 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga 4080 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc 414 0 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac 4200

<td>caccgctggt</td><td>agcggtggtt</td><td>tttttgtttg ·</td><td>caagcagcag</td><td>attacgcgca</td><td>gaaaaaaagg</td><td> 4260</td>

<td>atctcaagaa</td><td>gatcctttga</td><td>tcttttctac</td><td>ggggtctgac</td><td>gctcagtgga</td><td>acgaaaactc</td><td> 4320</td>

<td>acgttaaggg</td><td>attttggtca</td><td>tgagattatc</td><td>aaaaaggatc</td><td>ttcacctaga</td><td>tccfctttaaa</td><td> 4380</td>

<td>ttaaaaatga</td><td>agttttaaat</td><td>caatctaaag</td><td>tatatatgag</td><td>taaacttggt</td><td>ctgacagtta</td><td> 444 0</td>

<td>ccaatgctta</td><td>atcagtgagg</td><td>cacctatctc</td><td>agcgatctgt</td><td>ctatttegtt</td><td>catccatagt</td><td> 4500</td>

<td>tgcctgactc</td><td>cccgtcgtgt</td><td>agataactac</td><td>gatacgggag</td><td>ggcttaccat</td><td>ctggccccag</td><td> 4560</td>

<td>tgctgcaatg</td><td>ataccgcgag</td><td>acccacgctc</td><td>accggctcca</td><td>gatttatcag</td><td>caataaacca</td><td> 4620</td>

<td>gccagccgga</td><td>agggccgagc</td><td>gcagaagtgg</td><td>tcctgcaact</td><td>ttatccgcct</td><td>ccatccagtc</td><td> 4680</td>

<td>tattaattgt</td><td>tgccgggaag</td><td>ctagagtaag</td><td>tagttcgcca</td><td>gttaatagtt</td><td>tgcgcaacgt</td><td> 4740</td>

<td>tgttgccatt</td><td>gctgcaggca</td><td>tegtggtgtc</td><td>acgctcgtcg</td><td>tttggtatgg</td><td>cttcattcag</td><td> 4800</td>

<td>ctccggttcc</td><td>caacgatcaa</td><td>ggcgagttac</td><td>atgatccccc</td><td>atgttgtgca</td><td>aaaaagcggt</td><td> 4860</td>

<td>tagctccttc</td><td>ggtcctccga</td><td>tcgttgtcag</td><td>aagtaagttg</td><td>gccgcagtgt</td><td>tateact-cat</td><td> 4920</td>

<td>ggttatggca</td><td>gcactgcata</td><td>attctcttac</td><td>tgtcatgcca</td><td>tccgtaagat</td><td>gcttttctgt</td><td> 4980</td>

<td>gactggtgag</td><td>tactcaacca</td><td>agtcattctg</td><td>agaatagtgt</td><td>atgcggcgac</td><td>cgagttgctc</td><td> 5040</td>

<td>ttgcccggeg</td><td>tcaacacggg</td><td>ataataccgc</td><td>gccacatagc</td><td>agaactttaa</td><td>aagtgctcat</td><td> 5100</td>

<td>cattggaaaa</td><td>cgttcttcgg</td><td>ggcgaaaact</td><td>ctcaaggatc</td><td>ttaccgctgt</td><td>tgagatccag</td><td> 5160</td>

<td>ttegatgtaa</td><td>cccactcgtg</td><td>cacccaactg</td><td>atetteagea</td><td>tcttttactt</td><td>tcaccagcgt</td><td> 5220</td>

<td>ttctgggtga</td><td>geaaaaacag</td><td>gaaggcaaaa</td><td>tgccgcaaaa</td><td>aagggaataa</td><td>gggcgacacg</td><td> 5280</td>

gaaatgttga atactcatac tcttcctttt fccaatattat tgaagcattt atcagggtta 5340 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc 5400 gcgcacattt ccccgaaaag tgocacctga cgtctaagaa accattatta tcatgacatt 5460 aacctataaa aaataggcgt atcacgaggc cctttcgtct tcaagaatfcc tcatgtttga 5520 cagcttatea tcgatccact tgtatatttg gatgaatttt tgaggaattc tgaaccagtc 5S80 ctaaaacgag taaataggae eggcaattct tcaageaata aacaggaata ccaattatta 5640 aaagataact tagtcagatc gtacaataaa gctttgaaga aaaatgcgcc ttattcaatc 5700 tttgcataaa aaaatggccc aaaatctcac attggaagac atttgatgac ctcatttctt 5760 tcaatgaagg gcctaacgga gttgactaat gttgtgggaa attggaccga taagcgtgct 5820 tctgccgtgg ccaggacaac gtatactcat cagataacag caatacctga tcactacttc 5880 gcactagttt ctcggtacta tgcatatgat ccaatatcaa aggaaatgat agcattgaag 5940 gatgagacta atccaattga ggagtggcag catatagaac agctaaaggg tag-tgetgaa €000

<td>ggaagcatac</td><td>gataccccgc</td><td>atggaatggg</td><td>ataatatcac</td><td>aggaggtact</td><td>agactacctt</td><td> 6060</td>

<td>tcatcctaca</td><td>taaatagacg</td><td>catataagta</td><td>cgcattfcaag</td><td>cataaacacg</td><td>cactatgccg</td><td> 6120</td>

<td>ttcttctcat</td><td>gtatatatat</td><td>atacaggcaa</td><td>cacgcagata</td><td>taggtgcgac</td><td>gtgaacagtg</td><td> 6180</td>

<td>agctgtatgt</td><td>gcgcagctcg</td><td>cgttgcattt</td><td>tcggaagcgc</td><td>tcgttttcgg</td><td>aaacgctttg</td><td> 6240</td>

<td>aagttcctat</td><td>tccgaagttc</td><td>ctattctcta</td><td>gaaagtatag</td><td>gaacttcaga</td><td>gcgcttttga</td><td> 6300</td>

<td>aaaccaaaag</td><td>cgctctgaag</td><td>acgcactttc</td><td>aaaaaaccaa</td><td>aaacgcaccg</td><td>gactgtaacg</td><td> 6360</td>

<td>agctactaaa</td><td>atattgcgaa</td><td>taccgcttcc</td><td>acaaacattg</td><td>ctcaaaagta</td><td>tctctttgct</td><td> 6420</td>

<td>atatatctct</td><td>gtgctatatc</td><td>cctatataac</td><td>catcccatcc</td><td>acctttcgct</td><td>ccttgaactt</td><td> 6480</td>

<td>gcatctaaac</td><td>tcgacctcta</td><td>•cattttttat</td><td>gtttatctct</td><td>agtattacct</td><td>cttagacaaa</td><td> 6540</td>

<td>aaaattgtag</td><td>taagaactat</td><td>tcatagagtt</td><td>aatcgaaaac</td><td>aatacgaaaa</td><td>tgtaaacatt</td><td> 6600</td>

<td>tcctatacgt</td><td colspan="2">agtatataga gacaaaatag</td><td>aagaaaccgt</td><td>tcataatttt</td><td>ctgaccaatg</td><td> 6660</td>

<td>aagaatcatc</td><td>aacgctatca</td><td>ctttctgttc</td><td>acaaagtatg</td><td>cgcaatccac</td><td>afccggtatag</td><td> 6720</td>

<td>aatataatcg</td><td>gggatgcctt</td><td>tatcttgaaa</td><td>aaatgcaccc</td><td>gcagcttcgc</td><td>tagtaatcag</td><td> 6780</td>

<td>taaacgcggg</td><td>aagtggagtc</td><td>aggctttttt</td><td>tatggaagag</td><td>aaaatagaca</td><td>ccaaagtagc</td><td> 6840</td>

<td>cttcttctaa</td><td>ccttaacgga</td><td>cctacagtgc</td><td>aaaaagttat</td><td>caagagactg</td><td>cattatagag</td><td> 6800</td>

<td>cgcacaaagg</td><td>agaaaaaaag</td><td>taatctaaga</td><td>tgctttgtta</td><td>aaaaaatanr</td><td>CT C-f ΓΊΤ r*rrrrrre</td><td> 6860</td>

tgcatttttg tagaacaaaa aagaagtata gattcttgtt ggtaaaatag cgctctcg-cg 7020

-ttgcatttct gttctgtaaa aatgcagctc agattcrtttg tttgaaaaat tagcgctctc 7060 gcgttgcatt tttgttttac aaaaatgaag cacagattct tcgttggtaa aatagcgctt 7140 tcgcgttgca tttctgttct gtaaaaatgc agctcagatt ctttgtttga aaaattagcg 7200 ctctcgcgtt gcatttttgt tctacaaaat gaagcacaga tgcttcgtta acaaagatat 7260 gctattgaag tg-caagatgg aaacgcagaa aatgaaccgg ggatgcgacg tgcaagatta 7320 cetatgcaafc agatgcaata gtttctccag gaaccgaaat acatacattg tcttccgtaa 7380 agcgctagac tatatattat tatacaggtt caaatatact atctgtttca gggaaaactc 7440 ccaggttcgg atgttcaaaa ttcaatgatg ggtaacaagt acgatcgtaa atctgtaaaa 7500 cagtttgtcg gatattaggc tgtatctoct caaagcgtat tcgaatatca ttgagaagct 7S60 gcattttttt tttttttttt tttttttttt tttttatata tatttcaagg atataccatt 7620 gtaatgtetg cccctaagaa gatcgtcgtt ttgccaggtg accacgttgg tcaagaaatc 7680 acagccgaag ccattaaggt tcttaaagct atttctgatg ttcgttccaa tgtcaagttc 7740 gatttcgaaa atcatttaat tggtggtgct gctatcgatg ctacaggtgt cccacttcca 7800 gatgaggcgc tggaagcctc caagaaggtt gatgccgttt tgttaggtgc tgtgggtggt 7860

<td>cctaaatggg</td><td>gtaccggtag</td><td>tgttagacct</td><td>gaacaaggtt</td><td>tactaaaaat</td><td>ccgtaaagaa</td><td> 7920</td>

<td>cttcaattgt</td><td>acgccaactt</td><td>aagaccatgt</td><td>aactttgcat</td><td>ccgactctct</td><td>tttagactta</td><td> 7980</td>

<td>tctccaatca</td><td>agccacaatt</td><td>tgctaaaggt</td><td>actgacttcg</td><td>ttgttgtcag</td><td>agaattagtg</td><td> 8040</td>

<td>ggaggtattt</td><td>actttggtaa</td><td>gagaaaggaa</td><td>gacgatggtg</td><td>atggtgtcgc</td><td>ttgggatagt</td><td> 8100</td>

<td>gaacaataca</td><td>ccgttccaga</td><td>agtgcaaaga</td><td>atcacaagaa</td><td>tggccgcttt</td><td>catggcccta</td><td> 8160</td>

<td>caacatgagc</td><td>caccattgcc</td><td>tatttggtcc</td><td>-tt.ggat.aaag</td><td>ctaatgtttt</td><td>ggcctcttca</td><td> 8220</td>

<td>agattatgga</td><td>gaaaaactgt</td><td>ggaggaaacc</td><td>atcaagaacg</td><td>aattccctac</td><td>attgaaggtt</td><td> 8280</td>

<td>caacatcaat</td><td>tgattgattc</td><td>tgccgccatg</td><td>atcctagtta</td><td>agaacccaac</td><td>ccacctaaat</td><td> 8340</td>

<td>ggtattataa</td><td colspan="2">tcaccagcaa catgtttggt</td><td>gatatcatct</td><td>ccgatgaagc</td><td>ctccgttatc</td><td> 8400</td>

<td>ccaggttcct</td><td>tgggtttgtt</td><td>gccatctgcg</td><td>tccttggcct</td><td>ctttgccaga</td><td>caagaacacc</td><td> 8460</td>

<td>gcatttggtt</td><td>tgtacgaacc</td><td>atgccacggt</td><td>tctgctccag</td><td>atttgccaaa</td><td>gaataaggtt</td><td> 8520</td>

<td>gaccctatcg</td><td>ccactatctt</td><td>gtctgctgca</td><td>atgatgttga</td><td>aattgtcatt</td><td>gaacttgcct</td><td> 8580</td>

<td>gaagaaggta</td><td>aggccattga</td><td>agatgcagtt</td><td>aaaaaggttt</td><td>tggatgcagg</td><td>tatcagaact</td><td> 8640</td>

<td>ggtgatttag</td><td>gtggttccaa</td><td>caçtaccacc</td><td>gaagtcggtg</td><td>atgctgtcgc</td><td>çgaagaagtt</td><td> 8700</td>

<td>aagaaaatcc</td><td>ttgcttaaaa</td><td>agattctctt</td><td>tttttatgat</td><td>atttgtacaa</td><td>aaaaaaaaaa</td><td> 8760</td>

<td>aaaaaaaaaa</td><td>aaaaaaaaaa</td><td>aaaaaaaaaa</td><td>aaaatgcagc</td><td>gtcacatcgg</td><td>ataataatga</td><td> 8820</td>

<td>tggcagccat</td><td>tgtagaagtg</td><td>ccttttgcat</td><td>ttctagtctc</td><td>tttctcggtc</td><td>tagctagttt</td><td> 8880</td>

tactacatcg cgaagataga atcttagatc acactgcctt tgctgagctg gatcatatga 8940 gtaacaaaag agtggtaagg cctcgttaaa ggacaaggac ctgagcggaa gtgtatcgta 9000 aagtagacgg agtatactag tatagtctat agtccgtgga attctaagtg ccagctttat 9060 aatgtcattc tccttactac agacccgcct gaaagtagac acatcatcat cagtaagctt 9120 tgacaaaaag cattgagtag ctaactcttc tatgcaatct atagctgttt tataaggcat 9180 tcaatggaca gattgaggtt tttgaaacat actagtgaaa ttagccttaa tcccttctcg 9240 aagttaatca tgcattatgg tgtaaaaaat gcaactcgcg ttgctctact ttttcccgaa 9300 tttccaaata cgcagctggg gtgattgctc gatttcgtaa cgaaagtttt gtttataaaa 9360 accgcgaaaa ccttctgtaa cagatagatt tttacagcgc tgatatacaa tgacatcagc 9420 tgtaatggaa aataactgaa atatgaatgg cgagagactg cttgcttgta ttaagcaatg 9480 tattatgcag cacttccaac ctatggtgta cgatgaaagt aggtgtgtaa tcgagacgac 9540 a-agggggact tttccagttc ctgatcatta taagaaatac aaaacgttag catttgcatt 9600 tgttggacat gtactgaata cagacgacac accggtaatt gaaaaagaac tggattggcc 9660 tgatcetgca ctagtgtaca atacaattgt cgatcgaatc ataaatcacc cagaattatc 9720 acagtttata tcggttgcat ttattagtca gttaaaggcc accatcggag agggtttaga 97B0 fcattaatgta aaaggcacgc taaaocgcag gggaaagggt afccagaaggc ctaaaggcgt 9B40 attttttaga tacatggaat ctccatttgt caatacaaag gtcactgcat tcttctctta 9900 tcttcgagat tataataaaa ttgcctcaga atatcacaat aatactaaat tcattctcac 9960 gttttcatgt caagcatatt gggcatctgg cccaaacttc tccgccttga agaatgttat 10020 ttggtg-ctcc ataattcatg aatacatttc taagtttgtg gaaagagaac aggataaagg 20080 tcatatagga gatcaggagc taccgcctga agaggaccct tctcgtgaac taaacaatgt 10140

<td>acaacatgaa</td><td>gtcaatagtt</td><td>taacggaaca</td><td>agatgcggag</td><td>gcggatgaag 1</td><td>gattgtgggg</td><td> 10200</td>

<td>tgaaatagat</td><td>tcattatgtg</td><td>aaaaatggca</td><td>gtctgaagcg</td><td>gagagtcaaa</td><td>ctgaggcgga</td><td> 10280</td>

<td>gataatagcc</td><td>gacaggataa</td><td>ttggaaatag</td><td>ccagaggatg</td><td>gcgaacctca</td><td>aaattcgtcg</td><td> 10320</td>

<td>tacaaagttc</td><td>aaaagtgtct</td><td>tgtatcatat</td><td>actaaaggaa</td><td>ctaattcaat</td><td>ctcagggaac</td><td> 10380</td>

<td>cgtaaaggtt</td><td>tatcgcggta</td><td>gtagtttttc</td><td>acacgattcg</td><td>ataaagataa</td><td>gcttacatta</td><td> 10440</td>

<td>tgaagagcag</td><td>catattacag</td><td>ccgtatgggt</td><td>etacttgata</td><td>gtaaaatttg</td><td>aagagcattg</td><td> 10500</td>

<td>gaagcctgtt</td><td>gatgtagagg</td><td>tcgagtttag</td><td>atgcaagttc</td><td>aaggagcgaa</td><td>aggtggatgg</td><td> 10580</td>

<td>gtaggttata</td><td>tagggatata</td><td>gcacagagat</td><td>atatagcaaa</td><td>gagatacttt</td><td>tgaggcaatg</td><td> 10620</td>

<td>tttgtggaag</td><td>cggtattcgc</td><td>aatattttag</td><td>tagctcgtta</td><td>cagtccggtg</td><td>cgtttttggt</td><td> 10680</td>

<td>tttttgaaag</td><td>tgcgtcttca</td><td>gagcgctttt</td><td>ggttttcaaa</td><td>agcgctctga</td><td>agttcctata</td><td> 10740</td>

<td>ctttctagag</td><td>aataggaact</td><td>tcggaatagg</td><td>aacttcaaag</td><td>cgtttccgaa</td><td>aacgagcgct</td><td> 10800</td>

<td>tccgaaaatg</td><td>caacgcgagc</td><td>tgcgcacata</td><td>cagctcactg</td><td>ttcacgtcgc</td><td>acctatatct</td><td> 10860</td>

<td>gcgtgttgcc</td><td>tgtatatata</td><td>tatacatgag</td><td>aagaaçggca</td><td>tagtgcgtgt</td><td>ttatgcttaa</td><td> 1092.0</td>

<td>atgcgtactt</td><td>atatgcgtct</td><td>atttatgtag</td><td>gatgaaaggt</td><td>agtctagtac</td><td>ctcctgtgat</td><td> 10980</td>

<td>attatcccat</td><td>tccatgcggg</td><td>gtatcgtatg</td><td>cttccttcag</td><td>cactaccctt</td><td>tagctgttct</td><td> 11040</td>

<td>atatgctgcc</td><td>actcctcaat</td><td>tggattagtc</td><td>tcatccttca</td><td>atgcattcat</td><td>ttcctttgat</td><td> 11100</td>

<td>attggatcat</td><td>accctagaag</td><td>tattacgtga</td><td>ttttctgccc</td><td>cttaccctcg</td><td>ttgctactct</td><td> 11160</td>

<td>cctttttttc</td><td>gtgggaaccg</td><td>ctttagggcc</td><td>ctcagtgatg</td><td>gtgttttgta</td><td>atttatatgc</td><td> 11220</td>

<td>tcctcttgca</td><td>tttgtgtctc</td><td>tacttcttgt</td><td>fccgcctggag</td><td>ggaacttctt</td><td>catttgtatt</td><td> 11280</td>

<td>agcatggttc</td><td>acttcagtcç</td><td>ttccttccaa</td><td>ctcactcttt</td><td>ttttgctgta</td><td>aacgattctc</td><td> 11340</td>

<td>tgccgccagt</td><td>tcattgaaac</td><td>tattgaatat</td><td>atcctttaga</td><td>gattccggga</td><td>tgaataaatc</td><td> 11400</td>

<td>acctattaaa</td><td>gcagcttgac</td><td>gatctggtgg</td><td>aactaaagta</td><td>agcaattggg</td><td>taacgacgct</td><td> 11460</td>

<td>tacgagcttc</td><td>ataacatctt</td><td>cttccgttgg</td><td>agctggtggg</td><td>actaataact</td><td>gt.gtacaa.tc</td><td> 11520</td>

catttttctc atgagcattt cggtagctct ctfccttgtct ttctcgggca atcttcctat 11580 tattatagca atagatttgt atagttgctt tctattgtct aacagcttgt tattctgtag 11640 catcaaatct atggcagcct gacttgcttc ttgtgaagag agcataccat ttccaatcga 11700 agatacgctg gaatcttctg cgctagaatc aagaccatac ggcctaccgg ttgtgagaga 11760 ttccatgggc cttatgacat atcctggaaa gagtagctca tcagacttac gtttactctc 11820 tatatcaata tctacatcag gagcaatcat ttcaataaac agccgacata catcccagac 11880

<td>gctataagct</td><td>gtacgtgctt</td><td>ttaccgtcag</td><td colspan="2">attcttggct gtttcaatgt</td><td>cgtccatttt</td><td> 11540</td>

<td>ggttttcttt</td><td>taecagtatt</td><td>gttcgtttga</td><td>taatgtatte</td><td>ttgcttatta</td><td>cattataaaa</td><td> 12000</td>

<td>tctgtgcaga</td><td>tcacatgtca</td><td>aaacaacttt</td><td>ttatcacaag</td><td>atagtaccgc</td><td>aaaacgaacc</td><td> 12060</td>

<td>tgcgggccgt</td><td>ctaaaaatta</td><td>aggaaaagca</td><td>gcaaaggtgc</td><td>atttttaaaa</td><td>tatgaaatga</td><td> 12120</td>

<td>agataccgca</td><td>gtaccaatta</td><td>ttttcgcagt</td><td>acaaataatg</td><td>cgcggccggt</td><td>gcatttttcg</td><td> 12180</td>

<td>aaagaacgcg</td><td>agacaaacag</td><td>gacaattaaa</td><td>gttagttttt</td><td>cgagttagcg</td><td>tgtttgaata</td><td> 12240</td>

<td>ctgcaagata</td><td>caagataaat</td><td>agagtagttg</td><td>aaactagata</td><td>tcaattgcac</td><td>acaagatcgg</td><td> 12300</td>

<td>cgctaagcat</td><td>gccacaattt</td><td>ggtatattat</td><td>gtaaaacacc</td><td>acctaaggtg</td><td>cttgttcgtc</td><td> 12360</td>

<td>agtttgtgga</td><td>aaggtttgaa</td><td>agaccttcag</td><td>gtgagaaaat</td><td>agcattatgt</td><td>gctgctgaac</td><td> 12420</td>

<td>taacctattt</td><td>atgttggatg</td><td>attacacata</td><td>acggaacagc</td><td>aatcaagaga</td><td>gccacattca</td><td> 124 80</td>

<td>tgagctataa</td><td>tactatcata</td><td>agcaattcgc</td><td>tgagtttcga</td><td>tattgtcaat</td><td>aaatcactcc</td><td> 12540</td>

<td>agtttaaata</td><td>caagacgcaa</td><td>aaagcaacaa</td><td>ttctggaagc</td><td>ctcattaaag</td><td>aaattgattc</td><td> 12600</td>

<td>ctgcttggga</td><td>atttacaatt</td><td>attccttact</td><td>atggacaaaa</td><td>acatcaatct</td><td>gatatcactg</td><td> 12660</td>

<td>atattgtaag</td><td>tagtttgcaa</td><td>ttacagttcg</td><td>aatcatcgga</td><td>agaagcagat</td><td>aagggaaata</td><td> 12720</td>

<td>gccacagtaa</td><td>aaaaatgcta</td><td>aagcacttct</td><td>aagtgagggt</td><td>gaaagcatct</td><td>gggagatcac</td><td> 12780</td>

<td>tgagaaaata</td><td>ctaaattcgt</td><td>ttgaghatac</td><td>tfccgagatfct</td><td>acaaaaacaa</td><td>aaactttata</td><td> 12840</td>

<td>ccaattcctc</td><td>ttcctageta</td><td>ctttcatcaa</td><td>ttgtggaaga</td><td>ttcagcgata</td><td>ttaagaacgt</td><td> 12900</td>

<td>tgatccgaaa</td><td>tcatttaaat</td><td>tagtccaaaa</td><td>taagtatctg</td><td>ggagtaataa</td><td>tccagtgttt</td><td> 12960</td>

<td>agtgacagag</td><td>acaaagacaa</td><td>gcgttagtag</td><td>gcacatatac</td><td>ttctttagcg</td><td>caaggggtag</td><td> 13020</td>

<210> 44 <211> 15810 <212> DNA <213> vetor pSYlaMPElsH6a <400> 44 atcgataagc ttttcaattc aattcatcat tttttttcta ttcttttttt tgatttcggt 60 ttctttgaaa tttttttgat tcggtaatct ccgaacagaa ggaagaacga aggaaggagc 120 acagacttag attggtatat atacgcatat gtagtgttga agaaacatga aattgcccag 1B0 tattcttaac ecaactgcac agaacaaaaa cctgcaggaa acgaagataa atcatgtcga 240 aagctacata taaggaacgt gctgctacte atcctagtcc tgttgctgcc aagctattta 300 atatcatgca cgaaaagcaa acaaacttgt gtgcttcatt ggatgttcgt aocaccaagg 360 aattactgga gttagttgaa gcattaggtc ccaaaatttg tttactaaaa acacatgtgg 420 atatcttgac tgatttttcc atggagggca cagttaagcc gctaaaggca ttafcccgcca 480 agtacaattt tttactcttc gaagacagaa aatttgctga cattggtaat acagtcaaat 540 tgcagtactc tgcgggtgta tacagaatag cagaatgggc agacattacg aatgcacacg 600 gtgtggtggg cccaggtatt gttagcggtt tgaagcaggc ggcagaagaa gtaacaaagg 660 aacctagagg ccttttaatg ttagcagaat tgtcatgcaa gggctcccta tctactggag 720 aatatactaa gggtactgtt gacattgcga agagcgacaa agattttgtt atcggcttta 780 ttgctcaaag agacatgggt ggaagagatg aaggttacga ttggttgatt atgacacecg 840 grtgtgggttt agatgacaag ggagacgcat tgggtcaaca gtatagaacc gtggatgatg 900 tggtctctac aggatctgac attattattg ttggaagagg actatttgca aagggaaggg 960 atgctaaggt agagggtgaa cgttacagaa aagcaggctg ggaagcatat ttgagaagat 1020 gcggccagca aaactaaaaa actgtattat aagtaaatgc atgtatacta aactcacaaa 1080 ttagagcttc aatttaatta tatcagttat tacccgggaa tctcggtcgt aatgattttt 1140 ataatgacga aaaaaaaaaa attggaaaga aaaagcttta atgcggtagt ttatcacagt 1200 taaattgcta acgcagtcag gcaccgtgta tgaaatcfcaa caatgcgctc atcgtcatcc 1260 tcggcaccgt caccctggat gctgtaggca taggcttggt tatgcçggta ctgccgggcc 1320 tcttgcggga tat.cgt.ccat tccgacagea tcgccagtca ctatggcgtg cfcgctagcgc 1380 tatatgcgtt gatgcaattt ctatgcgcac ccgttctcgg agcactgtcc gaçcgctttg 1440 gccgccgccc agtcctgctc gcttcgctac ttggagccac tatcgactae gcgatcatgg 1500 cgaccacacc cgtcctgtgg atecttcaat atgcgcacat acgctgttat gfcfccaaggtc 1560 ccttcgttta agaacgaaag cggtcttcct tttgagggat gtttcaagtt gttcaaatct 1620 atcaaatttg caaatcccca gtctgtatct agagcgttga atcggtgatg cgatttgtta 1680 attaaattga tggtgtcacc attaccaggt ctagatatac caatggcaaa ctgagcacaa 1740 caataccagt ccggatcaac tggca-ocatc tctccogtag tctcatctaa tttttcttcc 1800 ggatgaggtt ccagatatac cgcaacacct ttattatggt ttccctgagg gaataataga 1860 atgtcccatt cgaaatcacc aattctaaac ctggg-cgaat tgtatttcgg gtttgttaac 1920

<td>tegttccagt</td><td>caggaatgtt</td><td>ccacgtgaag</td><td>ctatcttcca</td><td>gcaaagtctc</td><td>cacttcttca</td><td> 1980</td>

<td>tcaaattgtg</td><td>gagaatactc</td><td>ccaatgctct</td><td>tatctatggg</td><td>acttccgaga</td><td>aacacagtac</td><td> 2040</td>

<td>cgatacttcc</td><td>caattcgtct</td><td>tcagagctca</td><td>ttgtttgttt</td><td>gaagagacta</td><td>atcaaagaat</td><td> 2100</td>

<td>cgttttctca</td><td>aaaaaattaa</td><td>tatcttaact</td><td>gatagtttga</td><td>tcaaaggggc</td><td>aaaacgtagg</td><td> 2160</td>

<td>ggcaaacaaa</td><td>cggaaaaatc</td><td>gtttctcaaa</td><td>ttttctgatg</td><td>ccaagaactc</td><td>taaccagtct</td><td> 2220</td>

<td>tatctaaaaa</td><td>ttgccttatg</td><td>atccgtctct</td><td>ccggttacag</td><td>cctgtgtaac</td><td>tgattaatcc</td><td> 2280</td>

<td>tgcctttcta</td><td>atcaccattc</td><td>taatgtttta</td><td>attaagggat</td><td>tttgtcttca</td><td>ttaacggctt</td><td> 2340</td>

<td>tcgctcataa</td><td>aaatgttatg</td><td>acgttttgcc</td><td>cgcaggcggg</td><td>aaaccatcca</td><td>cttcacgaga</td><td> 2400</td>

<td>ctgatctcct</td><td>ctgccggaac</td><td>accgggcatc</td><td>tccaacttat</td><td>aagttggaga</td><td>aataagagaa</td><td> 2460</td>

<td>tttcagattg</td><td>agagaatgaa</td><td>aaaaaaaaac</td><td>cctgaaaaaa</td><td>aaggttgaaa</td><td>ecagttecct</td><td> 2520</td>

<td>gaaattattc</td><td>ccctacttga</td><td>ctaataagta</td><td>tataaagacg</td><td>gtaggtattg</td><td>attgtaattc</td><td> 2580</td>

<td>tgtaaatcta</td><td>tttcttaaac.</td><td>ttcttaaatt</td><td>ctacttttat</td><td>agttagtctt</td><td>ttttttagtt</td><td> 2640</td>

<td>ttaaaacacc</td><td>aagaacttag</td><td>tttcgaataa</td><td>acacacataa</td><td>acaaacacca</td><td>tgagatttçc</td><td> 2700</td>

<td>ttcaattttt</td><td>actgcagttt</td><td>tattcgcagc</td><td>atcctccgca</td><td>ttagctgctc</td><td>cagtcaacac</td><td> 2760</td>

<td>tacaacagaa</td><td>gatgaaacgg</td><td>cacaaattcc</td><td>ggctgaagct</td><td>gtcatcggtt</td><td>actcagattt</td><td> 2820</td>

<td>agaaggggat</td><td>ttcgatgttg</td><td>ctgttttgcc</td><td>attttceaac</td><td>agcacaaata</td><td>acgggttatt</td><td> 2880</td>

gtttataaat actactattg ccagcattgc tgctaaagaa gaaggggtat ctctagataa 2940 aaggtatgag gtgcgcaacg tgtccgggat gtaccatgtc acgaacgact gctecaactc 3000 aagcattgtg tatgaggcag cggacatgat catgcacacc cccgggtgcg tgccctgcgt 3060 tcgggagaac aactcttccc gctgctgggt agcgctcacc cccacgctçg cagctaggaa 3120 cgccagcgtc cccactacga caatacgacg ccacgtcgat ttgctcgttg gggcggctgc 3180 tttctgttcc gctatgtacg tgggggatct c.tgcggatct gtcttcctcg tctcccagçt 3240 gttcaccatc tcgcctcgcc ggcatgagac ggtgcaggac tgcaattgct caatctatcc 3300 cggccacata acgggtcacc gtatggcttg ggatatgatg atgaactggc accaccacca 3360 tcaccattaa agatctcgac ttggttgaac acgttgccaa ggcttaagtg aatttacttt 3420 aaagtcttgc atttaaataa attttctttt tatagcttta tgacttagtt tcaatttata 3480 tactatttta atgacatttt cgatt-cattg attgaaagct ttgtgttttt tcttgatgcg 3540 ctattgcatt gttcttgtct ttttcgccac atgtaatatc tgtagtagat acctgataca 3600 ttgtggatgc tgagtgaaat tttagttaat aatggaggcg ctcttaataa ttttggggat 3660 attggetttt ttttttaaag tttacaaatg aattttttcc gccaggataa cgattctgaa 3720 gttactctta gcgttcctat cggtacagcc atcaaatcat gcctataaat catgcctata 3780 fcttgcgtgca gtcagtatca tctacatgaa aaaaactccc gcaatttctt atagaatacg 3840 ttgaaaatta aatgtacgcg ccaagataag ataacatata tctagctaga tgcagtaata 3900 tacacagatt ccegcggacg tgggaaggaa aaaattagat aacaaaatct gagtgatatg 3960 gaaattecge tgtatagctc atatctttcc cttcaacacc agaaatgtaa aaatcttgtt 4020 acgaaggatc tttttgctaa tgtttctcgc tcaatcctca tttcttccct acgaagagtc 4 080 aaatctactt gttttctgcc ggtatcaaga tccatatctt ctagtttcac catcaaagtc 4140

<td>caatttctag</td><td>tatacagttt</td><td>atgtcccaac</td><td>gtaacagaca</td><td>atcaaaattg</td><td>gaaaggataa</td><td> 4200</td>

<td>gtatccttca</td><td>aagaatgatt</td><td>ctgcgctggc</td><td>tcctgaaccg</td><td>cctaatggga</td><td>acagagaagt</td><td> 4280</td>

<td>ecaaaacgat</td><td>gctataagaa</td><td>ccagaaataa</td><td>aacgataaaa</td><td>ccataccagg</td><td>atcctctacg</td><td> 4320</td>

<td>ccggacgcat</td><td>cgtggccggc</td><td>atcaocggcg</td><td>ccacaggtge</td><td>ggttgctggc</td><td>ccctatatcg</td><td> 4380</td>

<td>ccgacatcac</td><td>cgatggggaa</td><td>gatcgggctc</td><td>gccacttcgg</td><td>gctcatgagc</td><td>gettgttfccg</td><td> 4440</td>

<td>gcgtgggtat</td><td>ggtggcagge</td><td>cecgtggccg</td><td>ggggactgtt</td><td>gggcgccatc</td><td>tccttgcatg</td><td> 4500</td>

<td>caccattcct</td><td>tgcggcggcg</td><td>gtgctcaacg</td><td>gcctcaacct</td><td>actactggge</td><td>tgcttcçtaa</td><td> 4580</td>

<td>tgcaggagtc</td><td>gcataaggga</td><td>gagcgtcgac</td><td>cgatgccctt</td><td>gagagçcttc</td><td>aacccagtca</td><td> 4820</td>

<td>gctccttccg</td><td>gtgggcgcgg</td><td>ggcatgacta</td><td>tcgtcgccgc</td><td>acttatgact</td><td>gtcttcttta</td><td> 4880</td>

<td>tcatgcaact</td><td>cgtaggacag</td><td>gtgccggcag</td><td>cgctctgggt</td><td>cattttcggc</td><td>gaggaccgct</td><td> 4740</td>

<td>ttcgctggag</td><td>cgcgacgatg</td><td>atcggcctgt</td><td>cgcttgcggt</td><td>attcggaatc</td><td>ttgcacgccc</td><td> 4800</td>

<td>tcgctcaagc</td><td>cttcgtcact</td><td>ggtcccgcca</td><td>ccaaacgttt</td><td>cggcgagaag</td><td>caggccatta</td><td> 4860</td>

<td>tcgccggcat</td><td>ggcggccgac</td><td>gcgctgggct</td><td>aegtcttgct</td><td>ggcgttcgcg</td><td>acgcgaggct</td><td> 4920</td>

<td>ggatggcctt</td><td>ccccattatg</td><td>attcttctcg</td><td>cttccggcgg</td><td>catcgggatg</td><td>ccegegttgc</td><td> 4980</td>

<td>aggccatgct</td><td>gtccaggcag</td><td>gtagatgacg</td><td>accatcaggg</td><td>acagcttcaa</td><td>ggatcgcfccg</td><td> 5040</td>

<td>cggctcttac</td><td>cagcctaact</td><td>tcgatcactg</td><td>gaccgctgat</td><td>cgtcacggcg</td><td>atttatgcog</td><td> 5100</td>

<td>cctcggcgag</td><td>cacatggaac</td><td>gggttggcat</td><td>ggattgtagg</td><td>cgccgcccta</td><td>taccttgtct</td><td> 5160</td>

<td>gcctccccgc</td><td>gttgcgtcgc</td><td>ggtgcatgga</td><td>gccgggccac</td><td>•ctcgacctga</td><td>atggaagccg</td><td> 5220</td>

<td>gcggcacctc</td><td>gctaacggat</td><td>tcaccactcc</td><td>aagaattgga</td><td>gocaatcaat</td><td>tcttgcggag</td><td> 5280</td>

<td>aactgtgaat</td><td>gcgcaaacca</td><td>acccttggca</td><td>gaacatatcc</td><td>atcgcgtccg</td><td>ccatctccag</td><td> 5340</td>

<td>cagccgcacg</td><td colspan="2">cggcgcatct cgggcagcgt</td><td>tgggtcctgg</td><td>ccacgggtgc</td><td>gcatgatcgt</td><td> 5400</td>

<td>gctcctgtcg</td><td>ttgaggaccc</td><td>ggctaggctg</td><td>gcggggttgc</td><td>cttactggtt</td><td>agcagaatga</td><td> 5460</td>

<td>atcaccgata</td><td>cgcgagcgaa</td><td>cgtgaagcga</td><td>cfcgctgctgc</td><td>aaaacgtctg</td><td>cgacctgagc</td><td> 5520</td>

<td>aacaacatga</td><td>atggtctfccg</td><td>gtfctccgtgt</td><td>tfccgtaaagt</td><td>ctggaaacgc</td><td>ggaagtcagc</td><td> 5580</td>

gccctgcacc attatgttcc ggatctgcat cgcaggatgc tgctggctac cctgtggaac 5640 acctacatct gtattaacga agcgctggca ttgaccctga gtgatttttc tctggtcccg 5700 ccgcatccat accgccagtt gtttaccctc acaacgttcc agtaaccggg catgttcatc 5760 atcagtaacc cgtatcgtga gcatcctctc tcgtttcatc ggtatcatta cccccatgaa 5820 cagaaattcc cccttacacg gaggcatcaa gtgaccaaac aggaaaaaac cgcccttaac 5880 atggcccgct ttatcagaag ccagacatta acgcttctgg agaaactcaa cgagctggac 5940 gcggatgaac aggcagacat ctgtgaatcg cttcacgacc acgctgatga gctttaccgc 6000 agctgectcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag 6060

<td>acggtcacag</td><td>cttgtctgta</td><td>agcggtgccg</td><td>ggagcagaca</td><td>agcccgtcag </td><td>ggcgcgtcag</td><td> €120</td>

<td>cgggtgttgg</td><td>cgggtgtcgg</td><td>ggcgcagcca</td><td>tgacccagtc</td><td>acgtagcgat</td><td>agcggagtgt</td><td> 6180</td>

<td>atactggctt</td><td>aactatgcgg</td><td>catcagagca</td><td>gattgtactg</td><td>agagtgcacc</td><td>atatgcggtg</td><td> 6240</td>

<td>tgaaataccg</td><td>cacagatgcg</td><td>taaggagaaa</td><td>ataccgcatc</td><td>aggcgctctt</td><td>ccgcttcctc</td><td> 6300</td>

<td>gctcactgac</td><td>tcgctgcgct</td><td>cggtcgttcg</td><td>gctgcggcga</td><td>gcggtatcag</td><td>ctcactcaaa</td><td> 6360</td>

<td>ggcggtaata</td><td>cggttatcca</td><td>cagaatcagg</td><td>ggataacgca</td><td>ggaaagaaca</td><td>tgtgagcaaa</td><td> 6420</td>

<td>aggccagcaa</td><td>aaggccagga</td><td>accgtaaaaa</td><td>ggccgcgttg</td><td>ctggcgtttt</td><td>tccataggct</td><td> €480</td>

<td>ccgcccccct</td><td>gacgagcatc</td><td>acaaaaatcg</td><td>acgctcaagt</td><td>cagaggtggc</td><td>gaaacccgae</td><td> 6540</td>

<td>aggactataa</td><td>agataccagg</td><td>cgtttecccc</td><td>tggaagctcc</td><td>ctcgtgcgct</td><td>ctcctgttcc</td><td> 6600</td>

<td>gaccctgccg</td><td>cttaccggat</td><td>acctgtccgc</td><td>ctttctccct</td><td>tcgggaagcg</td><td>tggcgctttc</td><td> 6660</td>

<td>tcatagctea</td><td>cgctgtaggt</td><td>atctcagttc</td><td colspan="2">ggtgtaggtc gttcgctcca</td><td>agctgggctg</td><td> €720</td>

<td>tgtgcacgaa</td><td>cccccegttc</td><td>agcccgaccg</td><td>ctgcgcctta</td><td>tccggtaact</td><td>afccgtcttga</td><td> 6780</td>

<td>gtccaacccg</td><td>gtaagacacg</td><td>acttatcgcc</td><td>actggcagca</td><td>gccactggta</td><td>acaggattag</td><td> 6840</td>

<td>cagagcgagg</td><td>tatgtaggcg</td><td>gtgctacaga</td><td>gttcttgaag</td><td>fcggtggccta</td><td>actacggcta</td><td> 6900</td>

<td>cactagaagg</td><td>acagtatttg</td><td>gtatctgcgc</td><td>tctgctgaag</td><td colspan="2">ccagttacct tcggaaaaag</td><td>69G0</td>

<td>agttggtagc</td><td>tcttgatccg</td><td>gcaaacaaac</td><td>caccgctggt</td><td>agcggtggtt</td><td>tttttgtttg</td><td> 7020</td>

<td>caagcagcag</td><td>attacgcgca</td><td>gaaaaaaagg</td><td>atctcaagaa</td><td>gatcctttga</td><td>fccttttctac</td><td> 7080</td>

<td>ggggtctgac</td><td>gctcagtgga</td><td>acgaaaactc</td><td>acgttaaggg</td><td>attttggtca</td><td>tgagattatc</td><td> 7140</td>

<td>aaaaaggatc</td><td>ttcacctaga</td><td>tccttttaaa</td><td>ttaaaaatga</td><td>agttttaaat</td><td>caatctaaag</td><td> 7200</td>

<td>tatatatgag</td><td>taaacttggt</td><td>ctgacagtta</td><td>ccaatgcfcta</td><td>atcagtgagg</td><td>cacctatctc</td><td> 7260</td>

agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactae 7320 gatacgggag ggcttaccat ctggçcccag tgctgcaatg ataccgcgag acccacgctc 7380 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg 7440 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag 7500 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc 7560 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac 7620 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag 7680 aagtaagttg gccgcagtgt tatcactcat ggttatggca geactgcata attctcttac 7740 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg 7800 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaacacggg ataataccgc 7860 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact 7920 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg 7980 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa 8040 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt 8100 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg 8160 tatttagaaa aataaacaaa taggggttce gcgcacattt ccccgaaaag tgccacctga 8220 cgtctaagaa accattatta tcatgacatt aacctataaa aaataggcgt atcacgaggc B280 cctttcgtct tcaagaattc tcatgtttga cagcttatca tcgatccact tgfcatatttg 8340 gatgaahttt tgaggaattc tgaaccagtc ctaaaacgag taaataggac cggcaattct 8400 tcaagcaata aacaggaata ccaattatta aaagataact tagtcagatc gtacaataaa 8460 gctttgaaga aaaatgcgcc ttattcaatc tttgcataaa aaaatggccc aaaatctcac 8520 attggaagac atttgatgac ctcatttctt tcaatgaagg gcctaacgga gttgactaat 8580 gttgtgggaa attggaccga taagcgtgct tctgccgtgg ccaggacaaç gtatactcat 8640 cagataacag caatacctga tcactacttc gcactagttt ctcggtacta tgcatatgat 8700 ccaatatcaa aggaaatgat agcattgaag gatgagacta atccaattga ggagtggcag 8760 catatagaac agctaaaggg tagtgctgaa ggaagcatac gataccccgc atggaatggg 8820 ataatatcac aggaggtact agactacctt tcatcctaca taaatagacg catataagta 8880 cgcatttaag cataaacacg cactatgccg ttcttctcat gtatatatat atacaggcaa 8940 cacgcagata taggtgcgac gtgaacagtg agctgtatgt gcgcagctcg egttgcattt 9000 tcggaagcgc tcgttttcgg aaacgctttg aagttcctat tccgaagttc otattctcta 9060 gaaagtatag gaacttcaga gcgcttttga aaaccaaaag cgctctgaag acgcactttc 9120 aaaaaaccaa aaacgcaccg gactgtaacg agctactaaa atattgcgaa taccgcttcc 9180 acaaacattg ctcaaaagta tctctttgct atatatctct gtgctatatc cctatataac 9240 catcccatcc acctttcgct ccttgaactt gcatctaaac tcgacctcta cattttttat 9300 gtttatctct agtattacct cttagacaaa aaaattgtag taagaactat tcatagagtt 9360 aatcgaaaac aatacgaaaa tgtaaacatt tcctatacgt agtatataga gacaaaatag 9420 aagaaaccgt tcataatttt ctgaccaatg aagaatcatc aacgctat-ca ctttctgttc 9480 acaaagtatg cgcaatccac atcggtatag aatataatcg gggatgcctt tatcttgaaa 9S40 aaatgcaccc gcagcttcgc tagtaatcag taaacgcggg aagtggagtc aggctttttt 9600 tatggaagag aaaatagaca ccaaagtagc cttcttctaa ccttaacgga cctacagtgc 9660 aaaaagttat caagagactg cattatagag cgcacaaagg agaaaaaaag taatctaaga 9720 tgctttgtta gaaaaatagc gctctcggga tgcatttttg tagaacaaaa aagaagtata 9780

<td>gattcttgtt</td><td>ggtaaaatag</td><td>cgctctcgcg</td><td>ttgcatttct</td><td>gttctgtaaa ;</td><td>aatgcagctc</td><td> 9840</td>

<td>agattctttg</td><td>tttgaaaaat</td><td>tagcgctctc</td><td>gcgttgcatt</td><td>tttgttttac</td><td>aaaaatgaag</td><td> 9900</td>

<td>cacagattct</td><td>tcgttggtaa</td><td>aatagcgcüt</td><td>tcgcgttgca</td><td>tttctgttct</td><td>gtaaaaatgc</td><td> 9960</td>

<td>agcfccagatt</td><td>ctttgtttga</td><td>aaaattagcg</td><td>ctctcgcgtt</td><td>gcatttttgt</td><td>tctacaaaat</td><td> 10020</td>

<td>gaagcacaga</td><td>tgcttcgtta</td><td>acaaagatat</td><td>gctattgaag</td><td>tgcaagatgg</td><td>aaacgcagaa</td><td> 10080</td>

<td>aatgaaccgg</td><td>ggatgcgacg</td><td>tgcaagatta</td><td>cctatgcaat</td><td>agatgcaata</td><td>gtttctccag</td><td> 10140</td>

<td>gaaccgaaat</td><td>acatacattg</td><td>tcttccgtaa</td><td>agcgctagac</td><td>tatatattat</td><td>tatacaggtt</td><td> 10200</td>

<td>caaatatact</td><td>atctgtttca</td><td>gggaaaactc</td><td>ecaggttcgg</td><td>atgttcaaaa</td><td>ttcaatgatg</td><td> 10260</td>

<td>ggtaacaagt</td><td>acgatcgtaa</td><td>atctgtaaaa</td><td>cagtttgfccg</td><td>gatattaggc</td><td>tgtatctect</td><td> 10320</td>

<td>caaagcgtat</td><td>fccgaatatca</td><td>ttgagaagct</td><td>gcattttttt</td><td>tttttttttt</td><td>tttttttttt</td><td> 10380</td>

<td>tttttatata</td><td>tatttcaagg</td><td>atataccatt</td><td>gtaafcgtctg</td><td>cccctaagaa</td><td>gatcgtcgtt</td><td> 10440</td>

<td>ttgccaggtg</td><td>accacgttgg</td><td>tcaagaaatc</td><td>acagccgaag</td><td>ccattaaggt</td><td>tcttaaagct</td><td> 10500</td>

<td>atttctgatg</td><td>ttcgttccaa</td><td>tgtcaagttc</td><td>gatttcgaaa</td><td>atcatttaat</td><td>tggtggtgct</td><td> 10560</td>

<td>gctatcgatg</td><td>ctacaggtgt</td><td>cccacttcca</td><td>gatgaggcgc</td><td>tggaagcctc</td><td>caagaaggtt</td><td> 10620</td>

<td>gatgccgttt</td><td>tgttaggtgc</td><td>tgtgggtggt</td><td>cct.aaat.ggg</td><td>gtaccggtag</td><td>tgttagacct</td><td> 10680</td>

<td>gaacaaggtt</td><td>tactaaaaat</td><td>ccgtaaagaa</td><td>cttcaattgt</td><td>acgccaactt</td><td>aagaccatgt</td><td> 10740</td>

<td>aactttgcat</td><td>ccgactctct</td><td>tttagactta</td><td>tctccaatca</td><td>agccacaatt</td><td>tgctaaaggt</td><td> 10800</td>

<td>actgacttcg</td><td>ttgttgtcag</td><td>agaattagtg</td><td>ggaggtattt</td><td>actttggtaa</td><td>gagaaaggaa</td><td> 10860</td>

<td>gacgatggtg</td><td>atggtgtcgc</td><td>ttgggatagt</td><td>gaacaataca</td><td>ccgttccaga</td><td>agtçcaaaga</td><td> 10920</td>

<td>atcacaagaa</td><td>tggccgcttt</td><td>catggcccta</td><td>-caacatgagc</td><td>caccattgcc</td><td>tatttggtcc</td><td> 10980</td>

<td>ttggataaag</td><td>ctaatgtttt</td><td>ggcctctfcca</td><td>agattatgga</td><td>gaaaaactgt</td><td>ggaggaaacc</td><td> 11040</td>

<td>atcaagaacg</td><td>aattccctac</td><td>attgaaggtt</td><td>caacatcaat</td><td>tgattgattc</td><td>: tgccgccatg</td><td> 11100</td>

<td>atcctagtta</td><td>agaacccaac</td><td>ccacctaaat i</td><td>ggtattataa ·</td><td>tcaceagcaa <</td><td>catgtttggt</td><td> 11160</td>

<td>gatatcatct</td><td>ccgatgaagc</td><td>ctccgttatc ,</td><td>ccaggttcet</td><td>tgggtttgtt i</td><td>gccatctgcg</td><td> 11220</td>

<td>tccttggcct</td><td>ctttgccaga</td><td>caagaacacc</td><td>gcatttggtt</td><td>tgtacgaacc i</td><td>atgccacggt</td><td> 11280</td>

<td>tctgctccag</td><td>atttgccaaa</td><td>gaataaggtt</td><td>gaccctatcg</td><td>ccactat-ctt</td><td>gtctgctgca</td><td> 11340</td>

<td>atgatgttga</td><td>aattgtcatt</td><td>gaacttgcct</td><td>gaagaaggta</td><td>aggccattga</td><td>agatgcagtt</td><td> 11400</td>

<td>aaaaaggttt</td><td>tggatgcagg</td><td>tatcagaact</td><td>ggtgatttag</td><td>gtggttccaa</td><td>cagtaccacc</td><td> 11460</td>

<td>gaagtoggtg</td><td>atgctgtcgc</td><td>cgaagaagtt</td><td>aagaaaatcc</td><td>ttgcttaaaa</td><td>agattctctt</td><td> 11520</td>

<td>tttttatgat</td><td>atttgtacaa</td><td>aaaaaaaaaa</td><td>aaaaaaaaaa</td><td>aaaaaaaaaa</td><td>aaaaaaaaaa</td><td> 11580</td>

<td>aaaatgcagc</td><td>gtcacatcgg</td><td>ataataatga</td><td>tggcagccat</td><td>tgtagaagtg</td><td>ccttttgcat</td><td> 11640</td>

<td>ttctagtctc</td><td>tttctcggtc</td><td>tagctagttt</td><td>tactacatcg</td><td>cgaagataga</td><td>atcttagatc</td><td> 11700</td>

<td>acac.tgcctt</td><td>tgctgagctg</td><td>gatcatatga</td><td>gbaacaaaag</td><td>agtggtaagg</td><td>cctcgttaaa</td><td> 11760</td>

<td>ggacaaggac</td><td>ctgagcggaa</td><td>gtgtatcgta</td><td>aagtagacgg</td><td>agtatactag</td><td>tatagtctat</td><td> 11820</td>

<td>agtccgtgga</td><td>attctaagtg</td><td>ccagctttat</td><td>aatgtcattc</td><td>tcettactac</td><td>agacccgcct</td><td> 11680</td>

<td>gaaagtagac</td><td>acatcatcat</td><td>cagtaagctt</td><td>tgacaaaaag</td><td>cattgagtag</td><td>ctaactcttc</td><td> 11940</td>

<td>tatgcaatct</td><td>atagctgttt</td><td>tataaggcat</td><td>tcaatggaea</td><td>gattgaggtt</td><td>tttgaaacat</td><td> 12000</td>

<td>actagtgaaa</td><td>ttagccttaa</td><td>tcccttctcg</td><td>aagttaatca</td><td>* err-j» *· i-a ► «g</td><td>fcgtaaaaaat</td><td>*1 ** X^VDU</td>

<td>gcaactcgcg</td><td>ttgctctact</td><td>ttttcccgaa</td><td>tttccaaata</td><td>cgcagctggg</td><td>gtgattgctc</td><td> 12120</td>

<td>gatttcgtaa</td><td>cgaaagtttt</td><td>gtttataaaa</td><td>accgcgaaaa</td><td>ccttctgtaa</td><td>cagatagatt</td><td> 12180</td>

<td>tttacagcgc</td><td>tgatatacaa</td><td>tgacatcagc</td><td>tgtaatggaa</td><td>aataactgaa</td><td>atatgaatgg</td><td> 12240</td>

<td>cgagagactg</td><td>cttgçttgta</td><td>ttaagcaatg</td><td>tattatgcag</td><td>cacttccaac</td><td>ctatggtgta</td><td> 12300</td>

<td>cgatgaaagt</td><td>aggtgtgtaa</td><td>tcgagacgac</td><td>aagggggact</td><td>tttccagttc</td><td>etgatcatta</td><td> 12360</td>

<td>taagaaatac</td><td>aaaacgttag</td><td>catttgcatt</td><td>tgttggacat</td><td>gtactgaata</td><td>cagacgacac</td><td> 12420</td>

<td>accggtaatt</td><td>gaaaaagaac</td><td>tggattggcc</td><td>tgatoctgca</td><td>ctagtgtaca</td><td>atacaattgt</td><td> 12480</td>

<td>egatcgaatc</td><td>ataaatcacc</td><td>cagaattatc</td><td>acagtttata</td><td>tcggttgcat</td><td>ttattagtca</td><td>12S40</td>

<td>gttaaaggcç</td><td>accatcggag</td><td>agggtttaga</td><td>tattaatgta</td><td>aaaggcacgc</td><td>taaaccgcag</td><td> 12600</td>

<td>gggaaagggt</td><td>atcagaaggc</td><td>ctaaaggcgt</td><td>attütttaga</td><td>tacatggaat</td><td>ctccatttgt</td><td> 12660</td>

caatacaaag gtcactgcat tcttctctta tcttcgagat tataataaaa ttgccteaga 12720 atatcacaat aatactaaat tcattctcac g-ttttcatgt caagcatatt gggcatctgg 12700 cccaaacttc tccgccttga agaatgttat ttggtgctcc ataattcatg aatacatttc 12840 taagtttgtg gaaagagaac aggataaagg tcatatagga gat-caggagc taccgcctga 12900 agaggaccct tctcgtgaac taaacaatgt acaacatgaa gtcaatagtt taacggaaca 12960 agatgcggag gcggatgaag gattgtgggg tgaaatagat tcattatgtg aaaaatggca 13020 gtctgaagcg gagagtcaaa ctgaggcgga gataatagcc gacaggataa ttggaaatag 13080 ccagaggatg gcgaacctca aaattcgtcg tacaaagttc aaaagtgtct tgtatcatat 13140 actaaaggaa ctaatfccaat ctcagggaac cgtaaaggtt fcatcgcggta gtagtttttc 13200 acacgattcg ataaagataa gcttacatta tgaagagcag catattacag ccgtatgggt 13260 ctacttgata gtaaaatttg aagagcattg gaagcctgtt gatgtagagg tcgagtttag 13320 atgcaagttc aaggagcgaa aggtggatgg gtaggttata tagggatata gcacagagat 13380 atatagcaaa gagatacttt tgaggcaatg tttgtggaag cggtattcgc aatattttag 13440

<td>tagctcgtta</td><td>cagtccggtg</td><td>cgtttttggt</td><td>tttttgaaag</td><td colspan="2">tgcgtcttca gagcgctttt</td><td> 13500</td>

<td>ggttttcaaa</td><td>agcgctctga</td><td>agttcctata</td><td>ctttctagag</td><td>aataggaact</td><td>tcggaatagg</td><td> 13560</td>

<td>aacttcaaag</td><td>cgtttccgaa</td><td>aacgagcgct</td><td>tccgaaaatg</td><td>caacgcgagc</td><td>tgcgcacata</td><td> 13620</td>

<td>cagctcactg</td><td>ttcacgtcgc</td><td>acctatatct</td><td>gcgtgttgcc</td><td>tgtatatata</td><td>tatacatgag</td><td> 13680</td>

<td>aagaacggca</td><td>tagtgcgtgt</td><td>ttatacttaa</td><td>atgcgtactt</td><td>atatgcgtct</td><td>atttatgtag</td><td> 13740</td>

<td>gatgaaaggt</td><td>agtctagtac</td><td>ctcctgtgat</td><td>attatcccat</td><td>tccatgcggg</td><td>gtatcgtatg</td><td> 13800</td>

<td>-cttccttcag</td><td>cactaccctt</td><td>tagctgttct</td><td>atatgctgcc</td><td>actcctcaat</td><td>tggattagtc</td><td> 13860</td>

<td>tcatccttca</td><td>atgcattcat</td><td>ttcctttgat</td><td>attggatcat</td><td>accctagaag</td><td>tattacgtga</td><td> 13920</td>

<td>ttttctgccc</td><td>cttaccctcg</td><td>ttgctactct</td><td>cctttttttc</td><td>gtgggaaccg</td><td>ctttagggcc</td><td> 13980</td>

<td>etcagtgatg</td><td>gtgttttgta</td><td>atttatatgc</td><td>tcctcttgca</td><td>tttgtgtctc</td><td>tacttcttgt</td><td> 14040</td>

<td>tcgcctggag</td><td>ggaacttctt</td><td>catttgtatt</td><td>agcatggttc</td><td>acttcagtcc</td><td>ttccttccaa</td><td> 14100</td>

<td>etcactcttt</td><td>ttttgctgta</td><td>aacgattctc</td><td>tgocgccagt</td><td>tcattgaaac</td><td>tattgaatat</td><td> 14160</td>

<td>atcctttaga</td><td>gattccggga</td><td>fcgaataaatc</td><td>acctattaaa</td><td>gcagcttgac</td><td>gatctggtgg</td><td> 14220</td>

<td>aactaaagta</td><td>agcaattggg</td><td>taacgacgct</td><td>tacgagctte</td><td>ataacatctt</td><td>cttccgttgg</td><td> 14280</td>

<td>agctggtggg</td><td>actaataact</td><td>gtgtacaatc</td><td>catttttctc</td><td>atgagcattt</td><td>cggtagctct</td><td> 14340</td>

<td>cttcttgtct</td><td>ttctcgggca</td><td>atcttcctat</td><td>tattatagca</td><td>atagatttgt</td><td>atagttgctt</td><td> 14400</td>

<td>tctattgtct</td><td>aacagcttgt</td><td>tattctgtag</td><td>catcaaatct</td><td>atggcagcct</td><td>gacttgcttc</td><td> 14460</td>

<td>ttgtgaagag</td><td>agcataccat</td><td>ttccaatcga</td><td>agatacgctj</td><td>gaatcttctg</td><td>cgctagaatc</td><td> 14520</td>

<td>aagaccatac</td><td colspan="2">ggcctaccgg ttgtgagaga</td><td>ttccatgggc</td><td>cttatgacat</td><td>atcctggaaa</td><td> 14580</td>

<td>gagtagctca</td><td>tcagacttac</td><td>gtttaetctc</td><td>tatatcaata</td><td>tctacatcag</td><td>gagcaatcat</td><td> 14640</td>

<td>ttcaataaac</td><td>agccgacata</td><td>catcccagac</td><td>gctataagct</td><td>gtacgtgctt</td><td>ttaccgtcag</td><td> 14700</td>

<td>attcttggct</td><td>gtttcaatgt</td><td>cgtccatttt</td><td>ggfcttfccttt</td><td>taccagtatt</td><td>gttcgtttga</td><td> 14760</td>

<td>taatgtattc</td><td colspan="2">ttgcttatta cattataaaa</td><td>tctgtgcaga</td><td>tcacatgtca</td><td>aaacaacttt</td><td> 14820</td>

<td>ttatcacaag</td><td>atagtaccgc</td><td>aaaacgaacc</td><td>tgcgggocgt</td><td>ctaaaaatta</td><td>aggaaaagca</td><td> 14880</td>

<td>gcaaaggtgc</td><td>atttttaaaa</td><td>tatgaaatga</td><td>agataocgea</td><td>gtaccaatta</td><td>ttttcgcagt</td><td> 14940</td>

<td>acaaataatg</td><td>cgcggccggt</td><td>gcatttttcg</td><td>aaagaacgcg</td><td>agacaaacag</td><td>gacaattaaa</td><td> 15000</td>

<td>gttagttttt</td><td>cgagttagcg</td><td>tgtttgaata</td><td>ctgcaagata</td><td>caagataaat</td><td>agagtagttg</td><td> 15060</td>

<td>aaactagata</td><td>tcaattgcac</td><td>acaagatcgg</td><td>cgctaagcat</td><td>gccacaattt</td><td>ggtatattat</td><td> 15120</td>

<td>gtaaaacacc</td><td>acctaaggtg</td><td>cttgttcgtc</td><td>agtttgtgga</td><td>aaggtttgaa</td><td>agaccttcag</td><td> 15180</td>

<td>gtgagaaaat</td><td>agcatt&tgt</td><td>gctgctgaac</td><td>taacctattt</td><td>atgttggatg</td><td>attacacata</td><td> 15240</td>

<td>acggaacagc</td><td>aatcaagaga</td><td>gccacattca</td><td>tgagctataa</td><td>tactatcata</td><td>agcaattcgc</td><td> 15300</td>

<td>tgagtttcga</td><td>tattgtcaat</td><td>aaatcactcc</td><td>agtttaaata</td><td>caagacgcaa</td><td>aaagcaacaa</td><td> 15360</td>

<td>ttctggaagc</td><td>ctcattaaag</td><td>aaattgattc</td><td>ctgcttggga</td><td>atttacaatt</td><td>attccttact</td><td> 15420</td>

<td>atggacaaaa</td><td>acatcaatct</td><td>gatateactg</td><td>at.attgt.aag</td><td>tagtttgcaa</td><td>ttacagttcg</td><td> 15480</td>

<td>aatcatcgga</td><td>agaagcagat</td><td>aagggaaata</td><td>gccaeagtaa</td><td>aaaaatgcta</td><td>aagcacttct</td><td>15S40</td>

<td>aagtgagggt</td><td>gaaagcatct</td><td>gggagatcac</td><td colspan="2">tgagaaaata ctãaattcgt</td><td>ttgagtatac</td><td> 15600</td>

<td>ttcgagattt</td><td>acaaaaacaa</td><td>aaactttata</td><td colspan="2">eeaattcctc ttcctagcta</td><td>ctttcatcaa</td><td> 15660</td>

<td>ttgtggaaga</td><td>ttcagcgata</td><td>ttaagaaçgt</td><td>t ga t ccgaaa</td><td></td><td>tagtccaaaa</td><td> 15720</td>

<td>taagtatctg</td><td>ggagtaataa</td><td>tccagtgttt</td><td>agtgacagag</td><td>acaaagacaa</td><td>gcgttagtag</td><td> 15780</td>

gcacatatac ttctttagcg caaggggtag 15810

210> 45 <211> 3928 <212> DNA <213> vetor pBKS-E2sH6 <400> 45 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag €0 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaat-caa aagaatagac 120 ogagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga 180 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcceactac gtgaaccatc 240 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg 300 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa 360 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac 420 caccacaccc gecgogctta atgcgccgct acagggcgcg tcccatfccgc cattcaggct 480 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa 540

<td>agggggatgt gctgeaaggc gattaagttg ggtaacgcca gggttttcce</td><td>agtcaegacg</td><td>SOO</td>

<td>ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat</td><td>tgggtaecgg</td><td> 660</td>

<td>gccccccctc gaggtcgacg gtatcgataa gcttgcatgc ctgcagttaa</td><td>ttaactatta</td><td> 720</td>

<td>gtgatggtgg tgatggtgtc tgccctcgat cacctgocac tctgttgtag</td><td>acagcagcag</td><td> 780</td>

<td>cgggctaagc tctgatctat ccctgtcctc caagtcacaa cgctctcctc</td><td>gagtccaatt</td><td> 840</td>

<td>gcatgcggct tcgaacctgt gctccacgcc ccccacgtac atcctaacct</td><td>tgaagatggt</td><td> 900</td>

<td>gaagttgaca gtgcaggggt agtgccagag cctatatggg taatgaacca</td><td>tacacctagg</td><td> 960</td>

<td>tgtcagccag ggcccagaac cgcatctggc gtaggtggcc tcggggtgct</td><td>tccgaaaaca</td><td> 1020</td>

<td>gtcagtgggg caggtcaagg tgttgttgcc ggcccccccg atgttgcacg</td><td>gggggccccc</td><td> 1080</td>

<td>acacgtcttg gtgaacccag tgccattcat ccatgrtacag ccgaaccagt</td><td>tgcctcgcgg</td><td> 1140</td>

<td>cggccgcgtg ttgttgagaa tcagcacatc cgagtcgttc gccccccagt</td><td>tatacgtggg</td><td> 1200</td>

<td>gacaccaaac cgatcggtcg tccccaccac aacagggctc ggggtgaagc</td><td>aatacactgg</td><td> 1260</td>

<td>accgcacacc tgagacgcgg gtacaatacc acacggtcga ggcgcgtagt</td><td>gccagcagta</td><td> 1320</td>

<td>gggcçtctgg tccgagctgt taggctcagt gtaagtgagg ggaccccacc</td><td>cctgagcgaa</td><td> 1380</td>

<td>cttgtcgatg gagcgacagc tggccaagcg ctctgggcat ccagacgagt</td><td>tgaatttgtg</td><td> 1440</td>

<td>tttgtagaat agtgcggcaa agaaccctgt ttggagggag tcgttgcagt</td><td>tT» fTfirrr>aCrfc</td><td>Ί Eflft w w</td>

<td>cctgttgatg tgccaactgc cgttggtgtt tacgagctgg attttctgag</td><td>ccgacccggg</td><td> 1560</td>

<td>gctaaagagg gacacaaggc ccctggtatc ggaggctgct gcccctectg</td><td>acacgcgggt</td><td> 1620</td>

<td>atggtaccgg gccccccctc gaggtcgacg gtatcgataa gcttgatatc</td><td>gaattcctgc</td><td> 1680</td>

<td>agcccggggg atccactagt tctagagcgg ccgccaccgc ggtggagctc</td><td>cagcttttgt</td><td> 1740</td>

<td>tccctttagt gagggttaat ttcgagcttg gcgtaatcat ggtcatagct</td><td>gtttcctgtg</td><td> 1800</td>

tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa 1860 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct 1920 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga 1980 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc 2040 gtfccggctgc ggcgagcggt atcagctcac tcaaaggogg taatacggtt atocacagaa 2100 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt 2160 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa 2220 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt 2280 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg 2340 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gct-cacgctg taggtatctc 2400 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaçcccç cgttcagccc 2460 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta 2520 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct 2580 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc 2640 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa 2700 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa 2760 aaaggatctc aagaagat.cc tttgatcttt tctacggggt ctgacgctca gtggaacgaa 2620

<td>aactcacgtt</td><td>aagggatttt</td><td>ggtcatgaga</td><td>ttatcaaaaa</td><td>ggatcttcac</td><td>ctagatcctt</td><td> 2880</td>

<td>ttaaattaaa</td><td>aatgaagttt</td><td>taaatcaatc</td><td>taaagtatat</td><td>atgagtaaac</td><td>ttggtctgac</td><td> 2940</td>

<td>agttaccaat</td><td>gcttaatcag</td><td>tgaggcacct</td><td>atctcagcga</td><td>tctgtctatt</td><td>tcgttcatcc</td><td> 3000</td>

<td>atagttgcct</td><td>gactccccgt</td><td>cgtgtagata</td><td>actacgatac</td><td>gggagggctt</td><td>accatctggc</td><td> 3060</td>

<td>eccagtgctg</td><td>caatgatacc</td><td>gcgagaccca</td><td>cgctcaccgg</td><td>ctccagattt</td><td>atcagcaata</td><td> 3120</td>

<td>aaccagccag</td><td>ccggaagggc</td><td>cgagcgcaga</td><td>agtggtcctg</td><td>caactttatc</td><td>cgcctccatc</td><td> 3180</td>

<td>cagtctatta</td><td>attgttgccg</td><td>ggaagctaga</td><td>gtaagtagtt</td><td>cgccagttaa</td><td>tagtttgcgc</td><td> 3240</td>

<td>aacgttgttg</td><td>ccattgctac</td><td>aggcatcgtg</td><td>gfcgfcçacget</td><td>t· ♦· rrrr — σ — —a</td><td>tatggcttca</td><td> 3300</td>

<td>ttcagctccg</td><td>gttcecaaeg</td><td>atcaaggcga</td><td>gttacatgat</td><td>cccccatgtt</td><td>gtgcaaaaaa</td><td> 3360</td>

<td>gcggttagct</td><td>ccttcggtcc</td><td>tccgatcgtt</td><td>gtcagaagta</td><td>agttggccgc</td><td>agtgttatca</td><td> 3420</td>

<td>ctcatggtta</td><td>tggcagcact</td><td>gcataattct</td><td>cttactgtca</td><td>tgccatecgt</td><td>aagatgcttt</td><td> 3480</td>

<td>tctgtgactg</td><td>gtgagtactc</td><td>aaccaagtca</td><td>ttctgagaat</td><td>agtgtatgcg</td><td>gcgaccgagt</td><td> 3540</td>

<td>tgctcttgcc</td><td>cggcgtcaat</td><td>acgggataat</td><td>accgcgccac</td><td>atagcagaac</td><td>tttaaaagtg</td><td> 3600</td>

<td>ctcatcattg</td><td>gaaaacgttc</td><td>ttcggggcga</td><td>aaactctcaa</td><td>ggatcttacc</td><td>gctgttgaga</td><td> 3660</td>

<td>tccagttcga</td><td>tgtaacccac</td><td>tcgtgcaccc</td><td>aactgatctt</td><td>cagcatcttt</td><td>tactttcacc</td><td> 3720</td>

<td>agcgtttctg</td><td>ggtgagcaaa</td><td>aacaggaagg</td><td>caaaatgccg</td><td>caaaaaaggg</td><td>aataagggcg</td><td> 3780</td>

<td>acacggaaat</td><td>gttgaatact</td><td>catactcttc</td><td>ctttttcaat</td><td>attattgaag</td><td>catttatcag</td><td> 3840</td>

<td>ggttattgtc</td><td>tcatgagcgg</td><td>atacatattt</td><td>gaatgtattt</td><td>agaaaaataa</td><td>acaaataggg</td><td> 3900</td>

<td>gttccgcgca</td><td>catttccccg</td><td>aaaagtgc</td><td></td><td></td><td></td><td> 3928</td>

<210> 46 <211> 6104 <212> DNA <213> vetor pYIG5HCCL-22aH6 <400> 46

9Ί agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc €0 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcasttaat gtgagttagc 120 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa 180 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gaatttaata 240 cgactcacta tagggaattc gaggatcctt caatatgcge acatacgctg ttafcgttcaa 300 ggtcccttcg tttaagaacg aaagcggtct tccttttgag ggatgtttca agttgttcaa 360 atctatcaaa tttgcaaatc cccagtctgt atctagagcg ttgaatcggt gatgcgattt 420 gttaattaaa ttgatggtgt caccattacc aggtctagat ataccaatgg caaactgagc 480

<td>acaacaatac</td><td>cagtccggat</td><td>caactggcac</td><td>catctctccc</td><td>gtagtctcat</td><td>ctaatttttc</td><td> 540</td>

<td>ttccggatga</td><td>ggttccagat</td><td>ataccgcaac</td><td>acctttatta</td><td>tggtttccct</td><td>gagggaataa</td><td> 600</td>

<td>tagaatgtcc</td><td>cattcgaaat</td><td>caccaattct</td><td>aaacctgggc</td><td>gaattgtatt</td><td>tcgggtttgt</td><td> 660</td>

<td>taactcgttc</td><td>cagtcaggaa</td><td>tgttççacgt</td><td>gaagctatct</td><td>tccagcaaag</td><td>tctccacttc</td><td> 720</td>

<td>ttcatcaaat</td><td>tgtggagaat</td><td>actcccaatg</td><td>ctcttatcta</td><td>tgggacttcc</td><td>gggaaacaca</td><td> 780</td>

<td>gtaccgatac</td><td>ttcccaattc</td><td>gtcttcagag</td><td>ctcattgttt</td><td>gtttgaagag</td><td>actaatcaaa</td><td> 840</td>

<td>gaatcgtttt</td><td>ctcaaaaaaa</td><td>ttaatatctt</td><td>aactgatagt</td><td>ttgatcaaag</td><td>gggcaaaacg</td><td> 900</td>

<td>taggggcaaa</td><td>caaacggaaa</td><td>aatcgtttct</td><td>caaattttct</td><td>gatgccaaga</td><td>actctaacca</td><td> 960</td>

<td>gtcttatcta</td><td>aaaattgcct</td><td>tatgatccgt</td><td>ctctccggtt</td><td>acagcctgtg</td><td>taactgatta</td><td> 1020</td>

<td>atcctgcctt</td><td>tctaatcacc</td><td>attctaatgt</td><td>tttaattaag</td><td>ggattttgtc</td><td>ttcattaacg</td><td> 1080</td>

<td>gctttogctc</td><td>ataaaaatgt</td><td>tatgacgttt</td><td>tgcccgcagg</td><td>cgggaaacca</td><td>tccacttcac</td><td> 1140</td>

<td>gagactgatc</td><td>tcctctgccg</td><td>gaacaccggg</td><td>catctccaac</td><td>ttataagttg</td><td>gagaaataag</td><td> 1200</td>

<td>agaatttcag</td><td>attgagagaa</td><td>tgaaaaaaaa</td><td>aaaccctgaa</td><td>aaaaaaggtt</td><td>gaaaccagtt</td><td> 1260</td>

<td>ccctgaaatt</td><td>attcccctac</td><td>ttgactaata</td><td>agtatataaa</td><td>gacggtaggt</td><td>attgattgta</td><td> 1320</td>

<td>attctgtaaa</td><td>tctatttctt</td><td>aaacttctta</td><td>aattctactt</td><td>ttatagttag</td><td>tctttttttt</td><td> 1380</td>

<td>agttttaaaa</td><td>caccaagaac</td><td>ttagtttcga</td><td>ataaacacac</td><td>ataaacaaac</td><td>accatgagat</td><td> 1440</td>

<td>ttccttcaat</td><td>ttttactgca</td><td>gttttattcg</td><td>cagcatcctc</td><td>cg-cattagct</td><td>gctccagtca</td><td> 1500</td>

<td>acactacaac</td><td>agaagatgaa</td><td>acggcacaaa</td><td>ttccggctga</td><td>agctgtcatc</td><td>ggttactcag</td><td>1S60</td>

<td>atttagaagg</td><td>ggatttcgat</td><td>gttg-ctgttt</td><td>tgccattttc</td><td>caacagcaca</td><td>aataacgggt</td><td> 1620</td>

<td>tattgtttat</td><td>aaatacta-ct</td><td>attgccagca</td><td>ttgctgctaa</td><td>agaagaaggg</td><td>gtatctctag</td><td> 1680</td>

<td>ataaaaggca</td><td>taccegcgtg</td><td colspan="2">tcaggagggg cagcagcctc</td><td>cgataccagg</td><td>ggccttgtgt</td><td> 1740</td>

<td>ccctctttag</td><td>ccccgggtcg</td><td>gctcagaaaa</td><td>tccagctcgt</td><td>aaacaccaac</td><td>ggcagttggc</td><td> 1800</td>

<td>acatcaacag</td><td>gactgccctg</td><td>aactgcaacg</td><td>actccctcca</td><td>aacagggttc</td><td>tttgccgcac</td><td> 1860</td>

<td>tattctacaa acaeaaattc aactcgtctg gatgcccaga gcgcttggcc agctgtcgct</td><td> 1920</td>

<td>ccatcgacaa gttcgctcag gggtggggtc ccctcactta cactgagcct aacagctcgg</td><td> 1980</td>

<td>accagaggcc ctactgctgg cactacgcgc ctogaccgtg tggtattgta cccgcgtctc</td><td> 2040</td>

<td>aggtgtgcgg tccagtgtat tgcttcaccc cgagccctgt tgtggtgggg acgaccgatc</td><td> 2100</td>

<td>ggtttggtgt ccccacgtat aactgggggg cgaacgactc ggatgtgctg attctcaaca</td><td> 2160</td>

<td>acacgcggcc gccgcgaggc aactggttcg gcfcgtacatg gatgaatggc actgggfcfcca</td><td> 2220</td>

<td>ccaagacgtg tgggggcccc ccgtgcaaca tcgggggggc cggcaacaac accttgacct</td><td> 2280</td>

<td>gccccactga ctgttttcgg aagcaccccg aggccactta cgccagatgc ggttctgggc</td><td> 2340</td>

<td>cctggctgac acctaggtgt atggttcatt acccatatag gctctggcac tacccctgca</td><td> 2400</td>

<td>ctgtcaactt caccatcttc aaggttagga tgtacgtggg gggcgtggag cacaggttcg</td><td> 2460</td>

<td>aagccgcatg caattggact cgaggagagc gttgtgactt ggaggacagg gatagatcag</td><td> 2520</td>

<td>agcttagctc gctgctgctg tctacaacag agtggcaggt gatcgagggc agacaccatc</td><td> 2580</td>

<td>accaccatca ctaatagtta attaacgatc tcgacttggt tgaacacgtt gccaaggctt</td><td> 2640</td>

<td>aagtgaattt actttaaagt cttgcattta aataaatttt ctttttatag ctttatgact</td><td> 2700</td>

<td>tagtttcaat ttatatacta ttttaatgac attttcgatt cattgattga aagctttgtg</td><td> 2760</td>

<td>ttttttcttg atgcgctatt gcattgttct tgtctttttc gccacatgta atatctgtag</td><td> 2820</td>

<td>tagatacctg atacattgtg gatgctgagt gaaattttag ttaataatgg aggcgctctt</td><td> 2880</td>

<td>aataattttg gggatattgg cttttttttt taaagtttac aaatgaattt tttccgccag</td><td> 2940</td>

<td>gataacgatt ctgaagttac tcttagcgtt cctatcggta cagccatcaa atcatgccta</td><td> 3000</td>

<td>taaatcatgc ctatatttgc gtgcagtcag tafccatctac atgaaaaaaa ctcccgcaat</td><td> 3060</td>

<td>ttcttataga atacgttgaa aattaaatgt acgcgccaag ataagataac atatatctag</td><td> 3120</td>

<td>ctagatgcag taatatacac agattcccgc ggacgtggga aggaaaaaat tagataacaa</td><td> 3180</td>

<td>aatctgagtg atatggaaat fcccgctgtat agctcatatc tttcccttca acaccagaaa</td><td> 3240</td>

<td>tgtaaaaatc ttgttacgaa ggatcttttt gctaatgttt ctcgctcaat ectcatttct</td><td> 3300</td>

<td>tccctacgaa gagtcaaatc tacttgtttt çtgccggtat caagatccat atctt-ctagt</td><td> 3360</td>

<td>ttcaccabca aagtccaatt tct.agt.atac agtttatgtc ccaacgtaac agacaatcaa</td><td> 3420</td>

<td>aattggaaag gataagtatc cttcaaagaa tgattctgcg ctggctcctg aaccgcctaa</td><td> 3460</td>

<td>tgggaacaga gaagtccaaa acgatgctat aagaaccaga aataaaacga taaaaocata</td><td> 3540</td>

<td>ccaggatcca agcttggcac tggccgfccgt tttacaacgt cgtgactggg aaaaccctgg</td><td> 3600</td>

<td>cgttacccaa cttaat-cgcc ttgcagcaca tcocectttc gccagctggc gtaatagcga</td><td> 3660</td>

<td>agaggcccgc</td><td>accgatcgcc</td><td>cttcccaaca</td><td>gttgcgcagc</td><td>ctgaatggcg ,</td><td>aatgggaaat</td><td> 3720</td>

<td>tgtaaacgtt</td><td>aatattttgt</td><td>taaaattcgc</td><td>gttaaatttt</td><td>fcgttaaatca <</td><td>gctcattttt</td><td> 3780</td>

<td>taaccaatag</td><td>gccgaaatcg</td><td>gcaaaatccc</td><td>ttataaatca</td><td>aaagaataga</td><td>ccgagatagg</td><td> 3840</td>

<td>gttgagtgtt</td><td>gttccagttt</td><td>ggaacaagag</td><td>tccactatta</td><td>aagaacgtgg</td><td>actccaacgt</td><td> 3900</td>

<td>caaagggcga</td><td>aaaaccgtct</td><td>atcagggcga</td><td>tagcccacta</td><td>cgtgaaccat</td><td>caccctaatc</td><td> 3960</td>

<td>aagttttttg</td><td>gggtcgaggt</td><td>gccgtaaagc</td><td>actaaatcgg</td><td>aaccctaaag</td><td>ggagcccccg</td><td> 4020</td>

<td>atttagagct</td><td>tgacggggaa</td><td>agccggcgaa</td><td>cgtggcgaga</td><td>aaggaaggga</td><td>agaaagcgaa</td><td> 4080</td>

<td>aggagcgggc</td><td>gctagggcgc</td><td>tggcaagtgt</td><td>agcggtcacg</td><td>ctgcgcgtaa</td><td>ccaccacacc</td><td> 4140</td>

<td>cgccgcgctt</td><td>aatgcgccgc</td><td>tacagggcgc</td><td>gtcaggtggc</td><td>acttttcggg</td><td>gaaatgtgcg</td><td> 4200</td>

<td>cggaacccct</td><td>atttgtttat</td><td>ttttctaaat</td><td>acattcaaat</td><td>atgtat-ccgc</td><td>tcatgagaca</td><td> 4260</td>

<td>ataaccctga</td><td>taaatgcttc</td><td>aataatattg</td><td>aaaaaggaag</td><td>agtatgagta</td><td>ttcaacattt</td><td> 4320</td>

<td>ccgtgtcgcc</td><td>cttattccct</td><td>tttttgcggc</td><td>attttgcctt</td><td>cctgtttttg</td><td>ctcacccaga</td><td> 4380</td>

<td>aacgctggtg</td><td>aaagtaaaag</td><td>atgctgaaga</td><td>tcagttgggt</td><td>gcacgagtgg</td><td>gttacatcga</td><td> 4440</td>

<td>actggatctc</td><td>aacagcggta</td><td>agatccttga</td><td>gagttttcgc</td><td>cccgaagaac</td><td>gttttccaat</td><td>4S00</td>

<td>gatgagcact</td><td>tttaaagttc</td><td>tgctatgtgg</td><td>cgcggtatta</td><td>tcccgtattg</td><td>acgccgggca</td><td>4S60</td>

<td>agagcaactc</td><td>ggtcgccgca</td><td>tacactattc</td><td>tcagaatgac</td><td>ttggttgagt</td><td>actcaccagt</td><td> 4620</td>

<td>cacagaaaag</td><td>catcttacgg</td><td>atggcatgac</td><td>agtaagagaa</td><td>ttatgcagtg</td><td>ctgccataac</td><td> 4680</td>

<td>catgagtgat</td><td>aacactgcgg</td><td>ccaacttact</td><td>tctgacaacg</td><td>atcggaggac</td><td>cgaaggagct</td><td> 4740</td>

<td>aaccgctttt</td><td>ttgcacaaca</td><td>tgggggatca</td><td>tgfcaactcgc</td><td>cttgatcgtt</td><td>gggaaccgga</td><td> 4800</td>

<td>gctgaatgaa</td><td>gccataccaa</td><td colspan="2">acgacgagcg tgacaccacg</td><td>atgcctgtag</td><td>caatggcaac</td><td> 4860</td>

aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat 4920 agactggatg gaggcggata aagttgcagg accacttctg cgctcggocc ttccggctgg 4980 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc 5040 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc 5100 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg 5160 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta 5220 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg 5280 tgagttttcg ttccactgag cgtcagaccc -cgtagaaaag atcaaaggat cttcttgaga 5340 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt 5400 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag 5460 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa 5520

100

<td>ctctgtagça</td><td>cogcctacat</td><td>acctcgctct</td><td>gctaatcctg</td><td>ttaccagtgg ctgctgccag</td><td>S580</td>

<td>tggcgataag</td><td colspan="2">tcgtgtctta ccgggttgga</td><td>ctcaagacga</td><td>tagttaccgg ataaggcgca</td><td> 5640</td>

<td>gcggtcgggc</td><td>tgaacggggg</td><td>gttcgtgcac</td><td>acagcccagc</td><td>ttggagcgaa cgacctacac</td><td> 5700</td>

<td>cgaactgaga</td><td>tacctacagc</td><td>gtgagcattg</td><td>agaaagcgcc</td><td>acgcttcccg aagggagaaa</td><td> 5780</td>

<td>ggcggacagg</td><td>tatccggtaa</td><td>gcggcagggt</td><td>cggaacagga</td><td>gagcgcacga gggagcttcc</td><td> 5820</td>

<td>agggggaaac</td><td>gcctggtatc</td><td>tttatagtcc</td><td>tgtcgggttt</td><td>cgecacctct gacttgagcg</td><td> 5880</td>

<td>tcgatttttg</td><td>tgatgctcgt</td><td>caggggggcg</td><td>gagcctatgg</td><td>aaaaacgcca gcaacgcggc</td><td> 5940</td>

<td>ctttttacgg</td><td>ttcctggcct</td><td>tttgctggcc</td><td>ttttgctcac</td><td>atgttctttc ctgcgttatc</td><td> 6000</td>

<td>ccctgattct</td><td>gtggataacc</td><td>gtattaccgc</td><td>ctttgagtga</td><td>gctgataccg ctcgccgcag</td><td> 6060</td>

<td>ccgaacgacc</td><td>gaçcgcagcg</td><td>agtcagtgag</td><td>cgaggaagcg</td><td>gaag</td><td> 6104</td>

<210> 47 <211> 16301 <212> DNA <213> vetor pYYIGSE2H6 <400> 47 atçgataagc ttttcaattc aattcatcat ttttttttta ttcttttttt tgatttcggt 60 ttctttgaaa tttttttgat tcggtaafcct ccgaacagaa ggaagaacga aggaaggagc 120 acagacttag attggtatat atacgcatat gtagtgttga agaaacatga aattgcccag 180 tattcttaac ccaactgcac agaacaaaaa cctgcaggaa acgaagataa atcatgfccga 240 aagctacata taaggaacgt gctgctactc atcctagfccc tgttgctgcc aagctattta 300 atatcatgca cgaaaagcaa acaaaettgt gtgcttcatt ggatgttcgt accaceaagg 360 aattactgga gttagttgaa gcattaggtc ccaaaatttg tttactaaaa acacatgtgg 420 atatcttgac tgatttttcc atggagggca cagttaagcc gctaaaggca ttatccgcca 48 0 agtacaattt ttt.act.cttc gaagacagaa aatttgctga cattggtaat acagtcaaat 540 tgcagtactc tgcgggtgta tacagaatag cagaatgggc agacattacg aatgcacacg 600 gtgtggtggg cccaggtatt gttagcggtt tgaagcaggc ggcagaagaa gtaacaaagg 660 aacctagagg ccttttgatg ttagcagaat tgtcatgcaa gggctcccta tctactggag 720 aatatactaa gggtactgtt gacattgcga agagcgacaa agattttgtt atcggcttta 780 ttgctcaaag agacatgggt ggaagagatg aaggttacga ttggttgatt atgacacccg 840 gtgtgggttt agatgacaag ggagacgcat tgggtcaaca gtatagaacc gtggatgatg 900 tggtctctac aggatctgac attattattg ttggaagagg actatttgca aagggaaggg 960 atgctaaggt agagggtgaa cgttacagaa aagcaggctg ggaagcatat ttgagaagat 1020

101

<td>gcggccagca</td><td>aaactaaaaa</td><td>actgtattat</td><td>aagtaaatgc</td><td>atgtatacta ;</td><td>aactcacaaa</td><td> 1080</td>

<td>ttagagcttc</td><td>aatttaatta</td><td>tatcagttat</td><td>tacccggcaa</td><td>tctcggtcgt</td><td>aatgattttt</td><td> 1140</td>

<td>ataatgacga</td><td>aaaaaaaaaa</td><td>attggaaaga</td><td>aaaagcttta</td><td>atgcggtagt</td><td>ttatcacagt</td><td> 1200</td>

<td>taaattgcta</td><td>acgcagtcag</td><td>gcaccgtgta</td><td>tgaaatctaa</td><td>caatgcgctc</td><td>atcgtcatcc</td><td> 1260</td>

<td>tcggcaccgt</td><td>caccctggat</td><td>gctgtaggca</td><td>taggcttggt</td><td>tatgccggta</td><td>ctgccgggcc</td><td> 1320</td>

<td>tcttgcggga</td><td>tatcgtccat</td><td>tccgacagca</td><td>tcgccagtca</td><td>ctatggcgtg</td><td>ctgctagcgc</td><td> 1380</td>

<td>tatatgcgtt</td><td>gatgcaattt</td><td>ctatgcgcac</td><td>ccgttctcgg</td><td>agcactgtcc</td><td>gaccgctttg</td><td> 1440</td>

<td>gccgccgcec</td><td>agtcctgctc</td><td>gcttcgctac</td><td>ttggagccac</td><td>tatcgactac</td><td>gcgateatgg</td><td> 1500</td>

<td>cgaccacacc</td><td>cgtectgtgg</td><td>atccttcaat</td><td>atgcgeacat</td><td>acgctgttat</td><td>gttcaaggte</td><td> 1560</td>

<td>ccttcgttta</td><td>agaacgaaag</td><td>cggtcttcct</td><td>tttgagggat</td><td>gtttcaagtt</td><td>gttcaaatet</td><td> 1620</td>

<td>atcaaatttg</td><td>caaatcccca</td><td>gtctgtatct</td><td>agagcgttga</td><td>atcggtgatg</td><td>cgatttgtta</td><td> 1680</td>

<td>attaaattga</td><td>tggtgteacc</td><td>attaccaggt</td><td>ctagatatac</td><td>caatggcaaa</td><td>ctgagcacaa</td><td> 1740</td>

<td>caataccagt</td><td>ccggatcaac</td><td>tggcaccatc</td><td>tctcccgtag</td><td>tctcatctaa</td><td>tttttcttcc</td><td> 1800</td>

<td>ggatgaggtt</td><td>ccagatatac</td><td>cgcaacacct</td><td>ttattatggt</td><td>ttccctgagg</td><td>gaataataga</td><td> 1860</td>

<td>atgtcccatt</td><td>cgaaatcacc</td><td>aattctaaac</td><td>ctgggcgaat</td><td>tgtatttcgg</td><td>gtttgttaac</td><td> 1920</td>

<td>tcgttccagt</td><td>caggaatgtt</td><td>ccacgtgaag</td><td>ctatcttcca</td><td>gcaaagtctc</td><td>cacttcttca</td><td> 1980</td>

<td>tcaaattgtg</td><td>gagaatactc</td><td>ccaatgctct</td><td>tatctatggg</td><td>acttccggga</td><td>aacacagtac</td><td> 2040</td>

cgatacttcc caattcgtct tcagagctca ttgtttgttt gaagagacta atcaaagaat 2100 cgttttctca aaaaaattaa tatcttaact gatagtttga tcaaaggggc aaaacgtagg 2160 ggcaaacaaa cggaaaaatc gtttctcaaa ttttctgatg ccaagaactc taaccagtct 2220 tatctaaaaa ttgccttatg atccgtctct ccggttacag cctgtgtaac tgattaatcc 2280 tgcctttcta atcaccattc taatgtttta attaagggat tttgtcttca ttaacggctt 2340 tcgctcataa aaatgttatg acgttttgcc cgcaggcggg aaaccatcca cttcacgaga 2400 ctgatctcct ctgccggaac accgggcatc tccaacttat aagttggaga aataagagaa 2460 tttcagattg agagaatgaa aaaaaaaaac cctgaaaaaa aaggttgaaa ccagttccct 2520 gaaattattc ccctactfcga ctaataagta tataaagacg gtaggtattg attgtaattc 2580 tgtaaatcta tttcttaaac ttcttaaatt ctacttttat agttagtctt ttttttagtt 2640 ttaaaacacc aagaacttag tttcgaataa acacacataa acaaacacca tgagatttcc 2700 ttcaattttt actgcagttfc tattcgcagc atcetccgca ttagctgctc cagtcaacac 2760 tacaacagaa gatgaaacgg cacaaattcc ggctgaagct gtcatcggtt actcagattt 2820

102 agaaggggat ttcgatgttg ctgttttgcc attttccaac agcacaaata acgggttatt 2880 gtttataaat actactattg ccagcattgc tgctaaagaa gaaggggtat ctctagataa 2940 aaggcatacc cgcgtgtcag gaggggcagc agcctccgat accaggggcc ttgtgtccct 3000 ctttagcccc gggtcggcte agaaaatcca gctcgtaaac accaacggca gttggcacat 3060 caacaggact gccctgaact gcaacgactc cctccaaaca gggttctttg ccgcactatt 3120 ctacaaacac aaattcaact cgfcctggatg cccagagcgc ttggccagct gtcgctccat 3180

<td>cgacaagttc</td><td>gctcaggggt</td><td>ggggtcccct</td><td>cacttacact</td><td>gagcctaaca</td><td>gctcggacca</td><td> 3240</td>

<td>gaggccctac</td><td>tgctggcact</td><td>acgcgcctcg</td><td>accgtgtggt</td><td>attgtacccg</td><td>cgtctcaggt</td><td> 3300</td>

<td>gtgcggtcca</td><td>gtgtattgct</td><td>tcaccccgag</td><td>ccctgttgtg</td><td>gtggggacga</td><td>ccgatcggtt</td><td> 3360</td>

<td>tggtgtcccc</td><td>acgtataact</td><td>ggggggcgaa</td><td>cgactcggat</td><td>gtgctgattc</td><td>tcaacaacac</td><td> 3420</td>

<td>gcggccgccg</td><td>cgaggcaact</td><td>ggttcggctg</td><td>tacatggatg</td><td>aatggcactg</td><td>ggttcaccaa</td><td> 3480</td>

<td>gacgtgtggg</td><td>ggccccccgt</td><td>gcaacatcgg</td><td>gggggccggc</td><td>aacaacacct</td><td>tgacctgccc</td><td> 3540</td>

<td>cactgactgt</td><td>tttcggaagc</td><td>accccgaggc</td><td>cacttacgcc</td><td>agatgcggtt</td><td>ctgggccctg</td><td> 3600</td>

<td>gctgacacct</td><td>aggtgtatgg</td><td>ttcattaccc</td><td>atataggctc</td><td>tggcactacc</td><td>cctgcactgt</td><td> 3660</td>

<td>caacttcacc</td><td>atcttcaagg</td><td>ttaggatgta</td><td>cgtggggggc</td><td>gtggagcaca</td><td>ggttcgaagc</td><td> 3720</td>

<td>cgcatgcaat</td><td>tggactcgag</td><td>gagagcgttg</td><td>tgacttggag</td><td>gacagggata</td><td>gatcagagct</td><td> 3780</td>

<td>tagctcgctg</td><td>ctgctgtcta</td><td>caacagagtg</td><td>gcaggtgatc</td><td>gagggcagac</td><td>accatcacca</td><td> 3840</td>

<td>ccatcactaa</td><td>tagttaatta</td><td>acgatctcga</td><td>cttggttgaa</td><td>cacgttgcca</td><td>aggcttaagt</td><td> 3900</td>

<td>gaatttactt</td><td>taaagtcttg</td><td>catttaaata</td><td>aattttcttt</td><td>ttatagcttt</td><td>atgacttagt</td><td> 3960</td>

<td>ttcaatttat</td><td>atactatttt</td><td>aatgacattt</td><td>tcgattcatt</td><td>gattgaaagc</td><td>tttgtgtttt</td><td> 4020</td>

<td>ttcttgatgc</td><td>gctattgcat</td><td>tgttcttgtc</td><td>tttttcgcca</td><td>catgtaatat</td><td>ctgtagtaga</td><td> 4080</td>

<td>tacctgatac</td><td>attgtggatg</td><td>ctgagtgaaa</td><td>ttttagrttaa</td><td>taatggaggc</td><td>gctcttaata</td><td> 4140</td>

attttgggga tattggcttt tttttttaaa gtttacaaat gaattttttc cgccaggata 4200 acgattctga agttactctt agcgttccta toggtacagc catcaaafcca. tgcctataaa 4260 tcatgcctat atttgcgtgc agtcagtatc atctacatga aaaaaactcc cgcaatttct 4320

Jtatagaatac gttgaaaatt aaatgtacgc gccaagataa gataacatat atctagctag 4380 atgcagtaat atacacagat tcocgcggac gtgggaagga aaaaattaga taacaaaatc 4440 tgagtgatat ggaaattccg ctgtatagct catatctttc ccttcaacac cagaaatgta 4S00 aaaatcttgt tacgaaggat ctttttgcta atgtttctcg ctcaatcctc atttcttccc 4560 tacgaagagt caaatctact tgttttctgc cggtatcaag atccatatct tctagtttca 4620 ccatcaaagt ccaatttcta gtatacagtt tatgtcccaa cgtaacagac aatcaaaatt 4680

103

<td>ggaaaggata</td><td>agtatccttc</td><td>aaagaatgat</td><td>tctgcgctgg</td><td>ctcctgaacc <</td><td>gcctaatggg</td><td> 4740</td>

<td>aacagagaag</td><td>tccaaaacga</td><td>tgctataaga</td><td>accagaaata</td><td>aaaogataaa</td><td>accataccag</td><td> 4800</td>

<td>gatcctctac</td><td>gccggacgca</td><td>tcgtggccgg</td><td>catcaccggc</td><td>gccacaggtg</td><td>cggt-tgctgg</td><td> 4860</td>

<td>cccctatatc</td><td>gccgacatca</td><td>ccgatgggga</td><td>agatcgggct</td><td>cgccacttcg</td><td>ggctcatgag</td><td> 4920</td>

<td>cgcttgtttc</td><td>ggcgtgggta</td><td>tggtggcagg</td><td>ccccgtggcc</td><td>gggggactgt</td><td>tgggcgccat</td><td> 4980</td>

<td>ctccttgcat</td><td>gcaccattcc</td><td>ttgcggcggc</td><td>ggtgctcaac</td><td>ggcctcaacc</td><td>tactactggg</td><td> 5040</td>

<td>ctgcttccta</td><td>atgcaggagt</td><td>cgcataaggg</td><td>agagegtcga</td><td>ccgatgccct</td><td>tgagagoctt</td><td> 5100</td>

<td>caacccagtc</td><td>agctccttec</td><td>sgtssgcgcg</td><td>gggcatgact</td><td>atcgtcgccg</td><td>cacttatgac</td><td> 5160</td>

<td>tgtcttcttt</td><td>atcatgcaac</td><td>tcgtaggaca</td><td>ggtgccggca</td><td>gcgctctggg</td><td>tcattttcgg</td><td> 5220</td>

<td>cgaggaccgc</td><td>tttcgctgga</td><td>gcgcgacgat</td><td>gatcggcctg</td><td>tcgcttgcgg</td><td>tattcggaat</td><td> 5280</td>

<td>cttgcacgcc</td><td>ctcgctcaag</td><td>ccttcgtcac</td><td>tggtcccgcc</td><td>accaaacgtt</td><td>tcggcgagaa</td><td> 5340</td>

<td>gcaggccatt</td><td>atcgccggca</td><td>tggcggccga</td><td>cgcgctgggc</td><td>tacgtcttgc</td><td>tggcgttcgc</td><td> 5400</td>

<td>gacgcgaggc</td><td>tggatggcct</td><td>tccccattat</td><td>gattcttctc</td><td>gcttccggcg</td><td>gcatcgggat</td><td> 5460</td>

<td>gcccgcgttg</td><td>caggccatgc</td><td>tgtccaggca</td><td>ggtagatgac</td><td>gaccatcagg</td><td>gacagcttca</td><td> 5520</td>

<td>aggatcgctc</td><td>gcggctctta</td><td>ccagcctaac</td><td>ttcgatcact</td><td>ggaccgctga</td><td>tcgtcacggc</td><td>5S80</td>

<td>gatttatgcc</td><td>gcctcggcga</td><td>gcacatggaa</td><td>cgggttggca</td><td>tggattgtag</td><td>gcgccgccct</td><td> 5640</td>

<td>ataccttgtc</td><td>tgcctccccg</td><td>cgttgcgtcg</td><td>cggtgcatgg</td><td>agccgggoca</td><td>cctcgacctg</td><td> 5700</td>

<td>aatggaagcc</td><td>ggcggcacct</td><td>cgctaacgga</td><td>ttcaccactc</td><td>caagaattgg</td><td>agccaatcaa</td><td> 5760</td>

<td>ttcttgcgga</td><td>gaactgtgaa</td><td>tgcgcaaacc</td><td>aacccttggc</td><td>agaacatatc</td><td>: catcgcgtcc</td><td> 5820</td>

g-ccatcfccca gcagccgcac gcggcgcafec tcgggcagcg ttgggtcctg gccacgggtg 5880 cgcatgateg tgctcctgtc gttgaggacc cggctaggct ggcggggttg ccttactggt 5940 tagcagaatg aatcaccgat acgcgagcga acgtgaagcg actgctgctg caaaacgtct 6000 gcgacctgag caacaacatg aatggtcttc ggtttcegtg tttcgtaaag tctggaaacg 6060 cggaagtcag cgccctgcac cattatgttc cggatctgca tcgcaggatg ctgctggcta 6120 ccctgtggaa cacctacatc tgtattaacg aagcgctggc attgaccctg agtgattttt 6180 ctctggtccc gccgcatcca taccgccagt tgtttaccct cacaacgttc cagtaacegg 6240 gcatgtteat catcagtaac ccgtatcgtg ageatectct ctcgtttcat cggtatcatt 6300 acccccatga acagaaattc ccccttacac ggaggcatca agtgaccaaa caggaaaaaa 6360 ccgcccttaa catggcccgc tttatcagaa gccagacatt aacgcttctg gagaaactca 6420 acgagctgga cgcggatgaa caggcagaca tctgtgaatc gcttcacgac cacgctgatg 6480

104

<td>agctttaccg</td><td>cagctgcctc</td><td>gcgcgtttcg</td><td>gtgatgacgg</td><td>tgaaaacctc</td><td>tgacacatgc</td><td> 6540</td>

<td>agctcccgga</td><td>gacggtcaca</td><td>gcttgtctgt</td><td>aagcggtgcc</td><td>gggagcagac</td><td>aageeegtca</td><td> 6600</td>

<td>gggcgcgtca</td><td>gcgggtgttg</td><td>gcgggtgtcg</td><td>gggcgcagcc</td><td>atgacccagt</td><td>cacgtagcga</td><td> 6660</td>

<td>tagcggagtg</td><td>tatactggct</td><td>taactatgcg</td><td>gcatcagagc</td><td>agattgtact</td><td>gagagtgcac</td><td> 6720</td>

<td>catatgcggt</td><td>gtgaaatacc</td><td>gcacagatgc</td><td>gtaaggagaa</td><td>aataccgcat</td><td>caggcgctct</td><td> 6780</td>

<td>tccgcttcct</td><td>cgctcactga</td><td>ctcgctgcgc</td><td>tcggtcgttc</td><td>ggctgcggcg</td><td>agcggtatca</td><td> 6840</td>

<td>gctcactcaa</td><td>aggcggtaat</td><td>acggttatcc</td><td>acagaatcag</td><td>gggataacgc</td><td>aggaaagaac</td><td> 6900</td>

<td>atgtgagcaa</td><td>aaggccagca</td><td>aaaggccagg</td><td>aaccgtaaaa</td><td>aggecgcgtt</td><td>gctggcgttt</td><td> 6960</td>

<td>ttccataggc</td><td>tccgcccccc</td><td>tgacgagcat</td><td>cacaaaaatc</td><td>gacgctcaag</td><td>tcagaggtgg</td><td> 7020</td>

<td>cgaaacccga</td><td>caggactata</td><td>aagataccag</td><td>gcgtttcccc</td><td>ctggaagctc</td><td>cctcgtgcgc</td><td> 7080</td>

<td>tctcctgttc</td><td>cgaccctgcc</td><td>gcttaccgga</td><td>tacctgtccg</td><td>cctttctccc</td><td>ttcgggaagc</td><td> 7140</td>

<td>gtggcgcttt</td><td>ctcatagctc</td><td>acgctgtagg</td><td>tatctcagtt</td><td>cggtgtaggt</td><td>cgttcgctcc</td><td> 7200</td>

<td>aagctgggct</td><td>gtgtgcacga</td><td>accccccgtt</td><td>cagcccgacc</td><td>gctgcgcctt</td><td>atccggtaac</td><td> 7260</td>

<td>tatcgtcttg</td><td>agtccaaccc</td><td>ggtaagacac</td><td>gacttatcgc</td><td>cactggcagc</td><td>agccactggt</td><td> 7320</td>

<td>aacaggatta</td><td>gcagagcgag</td><td>gtatgtaggc</td><td>ggtgctacag</td><td>agttcttgaa</td><td>gtggtggcct</td><td> 7380.</td>

<td>aactacggct</td><td>acactagaag</td><td>gacagtattt</td><td>ggtatctgcg</td><td>ctctgctgaa</td><td>gccagttacc</td><td> 7440</td>

<td colspan="2">ttcggaaaaa gagttggtag</td><td>ctcttgatcc</td><td>ggcaaacaaa</td><td>ccaccgetgg</td><td>tagcggtggt</td><td> 7500</td>

ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 7S60 atcttttcta cggggtctga cgctcagtgg aàcgaaaact cacgttaagg gattttggtc 7620 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 7680 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 7740 gcacctatct cagcgatctg tetatttcgt tcatccatag ttgcctgact ccccgtcgtg 7800 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 7860 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aaggg-ccgag 7920 cgcagaagtg gtcctgcaa-c tttatccgcc tccatccagt ctattaattg ttgccgggaa 7980 getagagtaa gtagttcgoc agttaatagt ttgegcaacg ttgttgccat tgctgcaggc 8040 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctocggttc ccaacgatca 8100 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 8160 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat B220 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 8280 aagtcattct gagaatagtg tatgcggcga ocgagttgct cttgoccggc gtcaacacgg 8340

105 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 8400 gggcgaaaac tctcaaggat cttacegctg ttgagatcca gttcgatgta acccacfccgt 8460 gcacccaact gatcttcagc atcttttact tücaccagcg tttctgggtg agcaaaaaca Θ520 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 8580

<td>ctcttccttt</td><td>ttcaatatta</td><td>ttgaagcatt</td><td>tatcagggtt</td><td>attgtctcat i</td><td>gagcggatac</td><td> 8640</td>

<td>atatttgaat</td><td>gtatttagaa</td><td>aaataaacaa</td><td>ataggggttc</td><td>cgcgcacatt</td><td>tccccgaaaa</td><td> 8700</td>

<td>gtgccacctg</td><td>acgtctaaga</td><td>aaccattatt</td><td>atcatgacat</td><td>taacctataa</td><td>aaaataggcg</td><td> 8760</td>

<td>tatcacgagg</td><td>ccctttcgtc</td><td>ttcaagaatt</td><td>ctcatgtttg</td><td>acagcttatc</td><td>atcgatccac</td><td> 8820.</td>

<td>ttgtatattt</td><td>ggatgaattt</td><td>ttgaggaatt</td><td>ctgaaccagt</td><td>cctaaaacga</td><td>gtaaatagga</td><td> 8880</td>

<td>ccggcaattc</td><td>ttcaagcaat</td><td>aaacaggaat</td><td>accaattatt</td><td>aaaagataac</td><td>ttagtcagat</td><td> 8940</td>

<td>cgtacaataa</td><td>agctttgaag</td><td>aaaaatgcgc</td><td>cttattcaat</td><td>ctttgcataa</td><td>aaaaatggcc</td><td> 9000</td>

<td>caaaatctca</td><td>cattggaaga</td><td>catttgatga</td><td>cctcatttct</td><td>ttcaatgaag</td><td>ggcctaacgg</td><td> 9060</td>

<td>agttgactaa</td><td>tgttgtggga</td><td>aattggaccg</td><td>ataagcgtgc</td><td>ttctgccgtg</td><td>gccaggacaa</td><td> 9120</td>

<td>cgtatactca</td><td>tcagataaca</td><td>gcaatacctg</td><td>atcactactt</td><td>cgcactagtt</td><td>tctcggtact</td><td> 9180</td>

<td>atgcatatga</td><td>tccaatatca</td><td>aaggaaatga</td><td>tagcattgaa</td><td>ggatgagact</td><td>aatccaattg</td><td> 9240</td>

<td>aggagtggca</td><td>gcatatagaa</td><td>cagctaaagg</td><td>gtagtgctga</td><td>aggaagcata</td><td>cgataccccg</td><td> 9300</td>

<td>catggaatgg</td><td>gataatatca</td><td>caggaggtac</td><td>tagactacct</td><td>ttcatcctac</td><td>ataaatagac</td><td> 9360</td>

<td>gcatataagt</td><td>acgcatttaa</td><td>gcataaacac</td><td>gcactatgcc</td><td>gttcttctca</td><td>tgtatatata</td><td> 9420</td>

<td>tatacaggca</td><td>acacgcagat</td><td>ataggtgcga</td><td>cgtgaacagt</td><td>gagctgtatg</td><td>tgcgcagctc</td><td> 9480</td>

<td>gcgttgcatt</td><td>ttcggaagcg</td><td>ctcgttttcg</td><td>gaaacgcttt</td><td>gaagttccta</td><td>ttccgaagtt</td><td> 9540</td>

<td>cctattctct</td><td>agaaagtata</td><td>ggaacttcag</td><td>agcgcttttg</td><td>aaaaccaaaa</td><td>gcgctctgaa</td><td> 9600</td>

<td>gacgcacttt</td><td>caaaaaacca</td><td>aaaacgcacc</td><td>ggactgtaac</td><td>gagctacüaa</td><td>aatattgcga</td><td> 9660</td>

<td>ataccgcttc</td><td>cacaaacatt</td><td>gctcaaaagt</td><td>atctctttgc</td><td>tatatatctc</td><td>tgtgctatat</td><td> 9720</td>

<td>ccctatataa</td><td>ccatcccatc</td><td>cacctttcgc</td><td>tccttgaact</td><td>tgcatctaaa</td><td>ctcgacctct</td><td> 9780</td>

acatttttta tgtttatctc tagtattacc tctüagacaa aaaaattgta gtaagaacta 9840 ttcatagagt taatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag 9900 agacaaaata gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc 9960 actttctgtt cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct 10020 ttatcttgaa aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt 10080 caggcttttt ttatagaaga gaaaatagac accaaagtag ccttcttcta accttaa-cgg 10140

106

<td>acctacagtg</td><td>caaaaagtta</td><td>tcaagagact <</td><td>gcattataga i</td><td colspan="2">gcgcacaaag gagaaaaaaa</td><td> 10200</td>

<td>gtaatctaag</td><td>atgctttgtt</td><td>agaaaaatag</td><td>cgctctcggg i</td><td>atgcattttt ;</td><td>gtagaacaaa</td><td> 10260</td>

<td>aaagaagtat</td><td>agattcttgt</td><td>tggtaaaata</td><td>gcgctctcgc</td><td>gttgcatttc</td><td>tgttctgtaa</td><td> 10320</td>

<td>aaatgcagct</td><td>cagattcttt</td><td>gtttgaaaaa</td><td>ttagcgctct</td><td>cgcgttgcat</td><td>ttttgtttta</td><td> 10380</td>

<td>caaaaatgaa</td><td>gcacagattc</td><td>ttegttggta</td><td>aaatagcgct</td><td>ttcgcgttgc</td><td>atttctgttc</td><td> 10440</td>

<td>tgtaaaaatg</td><td>cagctcagat</td><td>tctttgtttg</td><td>aaaaattagc</td><td>gctctcgcgt</td><td>tgcatttttg</td><td> 10500</td>

<td>ttctacaaaa</td><td>tgaagcacag</td><td>atgcttcgtt</td><td>aacaaagata</td><td>tgctattgaa</td><td>gtgcaagatg</td><td>10S60</td>

<td>gaaacgcaga</td><td>aaatgaaccg</td><td>gggatgcgac</td><td>gtgcaagatt</td><td>acctatgcaa</td><td>tagatgcaat</td><td> 10620</td>

<td>agtttctcea</td><td>ggaaccgaaa</td><td>tacatacatt</td><td>gtcttccgta</td><td>aagcgctaga</td><td>ctatatatta</td><td> 10680</td>

<td>ttatacaggt</td><td>tcaaatatac</td><td>tatctgtttc</td><td>agggaaaact</td><td>cccaggttcg</td><td>gatgttcaaa</td><td> 10740</td>

<td>attcaatgat</td><td>gggtaacaag</td><td>tacgatcgta</td><td>aatctgtaaa</td><td>acagtttgtc</td><td>ggatattagg</td><td> 10800</td>

<td>ctgtatctcc</td><td>tcaaagcgta</td><td>ttcgaatatc</td><td>attgagaagc</td><td>tgcatttttt</td><td>tttttttttt</td><td> 10860</td>

<td>tttttttttt</td><td>ttttttatat</td><td>atatttcaag</td><td>gatataccat</td><td>tgtaatgtct</td><td>gcccctaaga</td><td> 10920</td>

<td>agatcgtcgt</td><td>tttgccaggt</td><td>gaccacgttg</td><td>gtcaagaaat</td><td>cacagccgaa</td><td>gccattaagg</td><td> 10980</td>

<td>ttcttaaagc</td><td>tatttctgat</td><td>gttcgttcca</td><td>atgtcaagtt</td><td>cgatttcgaa</td><td>aatcatttaa</td><td> 11040</td>

<td>ttggtggtgc</td><td>tgctatcgat</td><td>gctacaggtg</td><td>tcccacttcc</td><td>agatgaggcg</td><td>ctggaagcct</td><td> 11100</td>

<td>ccaagaaggt</td><td>tgatgccgtt</td><td>ttgttaggtg</td><td>ctgtgggtgg</td><td>tcctaaatgg</td><td>ggtaccggta</td><td> 11160</td>

<td>gtgttagacc</td><td>tgaacaaggt</td><td>ttactaaaaa</td><td>tccgtaaaga</td><td>acttcaattg</td><td>tacgccaact</td><td> 11220</td>

<td>taagaccatg</td><td>taa-ctttgca</td><td>tccgactctc</td><td>ttttagactt</td><td>atctecaatc</td><td>aagccacaat</td><td> 11280</td>

<td>ttgctaaagg</td><td>tactgacttc</td><td>gttgttgtca</td><td>gagaattagt</td><td>gggaggtatt</td><td>tactttggta</td><td> 11340</td>

<td>agagaaagga</td><td>agacgatggt</td><td>gatggtgtcg</td><td>cttgggatag</td><td>tgaacaatac</td><td>acegttccag</td><td> 11400</td>

<td>aagtgcaaag</td><td>aatcacaaga</td><td>afcggccgctt</td><td>tcatggccct</td><td>. acaacatgag</td><td> ccaccattgc</td><td> 11460</td>

<td>ctatttggtc</td><td>cttggataaa</td><td>gctaatgttt</td><td>tggcctcttc</td><td>: aagattatgg</td><td>’ agaaaaactg</td><td> 11520</td>

tggaggaaac catcaagaac gaattcccta cattgaaggt tcaacatcaa ttgattgatt 11580 ctgccgccat gatcctagtt aagaacccaa cccacctaaa tggtattata atcaccagca 11640 acatgtttgg tgatatcatc tccgatgaag cctccgttat cccaggttcc ttgggtttgt 11700 tgccatctgc gtccttggcc tctttgocag acaagaacac cgcattüggt ttgtacgaac 11760 catgccacgg ttctgctcca gatttgccaa agaataaggt tgaccctatc gccactatct 11820 tgtctgctgc aatgatgttg aaattgtcat tgaacttgcc tgaagaaggt aaggccattg 11880 aagatgcagt taaaaaggtt ttggatgcag gtatcagaac tggtgattta ggtggttcca 11940 acagtaccac cgaagtcggt gatgctgtcg ccgaagaagt taagaaaatc cttgettaaa 12000

107 aagattctct ttttttatga tatttgtaca aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa 1206 0 aaaaaaaaaa aaaaatgcag cgtcacatcg gataataatg atggcagcca ttgtagaagt 12120 gccttttgca tttctagtct ctttctcggt ctagctagtt ttactacatc gcgaagatag 12180 aatcttagat cacactgcct ttgctgagct ggatcatatg agtaacaaaa gagtggtaag 12240 gcctcgttaa aggacaagga cctgagcgga agtgtatcgt aaagtagacg gagtatacta 12300 gtatagtcta tagtccgtgg aattctaagt gceagettta taatgtcatt ctccttacta 12360 cagacccgcc tgaaagtaga cacatcatca tcagtaagct ttgacaaaaa gcattgagta 12420

<td>gctaactctt ctatgcaatc tatagctgtt ttataaggca ttcaatggac ;</td><td>agattgaggt</td><td> 124 80</td>

<td>ttttgaaaca tactagtgaa at.tagcct.ta atcccttctc gaagttaatc</td><td>atgcattatg</td><td> 12540</td>

<td>gfcgtaaaaaa tgcaactcgc gttgctctac tttttcccga atttccaaat</td><td>acgcagctgg</td><td> 12600</td>

<td>ggtgattgct cgatttcgta acgaaagttt tgtttataaa aaccgcgaaa</td><td>accttctgta</td><td> 12660</td>

<td>acagatagat ttttacagcg ctgatataca atgacatcag ctgtaatgga</td><td>aaataactga</td><td> 12720</td>

<td>aatatgaatg gcgagagact gcttgcttgt attaagcaat gtattatgca</td><td>gcacttccaa</td><td> 12780</td>

<td>cctatggtgt acgatgaaag taggtgtgta atcgagacga caagggggac</td><td>ttttccagtt</td><td> 12840</td>

<td>cctgatcatt ataagaaata caaaacgtta gcatttgcat ttgttggaca</td><td>tgtactgaat</td><td> 12900</td>

<td>acagacgaca caccggtaat tgaaaaagaa ctggattggc ctgatcctgc</td><td>actagtgtac</td><td> 12960</td>

<td>aatacaattg tcgatcgaat cataaatcac ccagaattat cacagtttat</td><td>atcggttgca</td><td> 13020</td>

<td>tttattagtc agttaaaggc caccatcgga gagggtttag atattaatgt</td><td>aaaaggcacg</td><td> 13080</td>

<td>ctaaaccgca ggggaaaggg tatcagaagg cctaaaggcg tattttttag</td><td>atacatggaa</td><td> 13140</td>

<td>tctccatttg tcaatacaaà ggtcactgca ttcttctctt atcttcgaga</td><td>ttataataaa</td><td> 13200</td>

<td>attgcctcag aatatcacaa taatactaaa ttcattctca cgttttcatg</td><td>tcaagcatat</td><td> 13260</td>

<td>tgggcatctg gcccaaactt ctccgccttg aagaatgtta tttggtgctc</td><td>cataattcat</td><td> 13320</td>

<td>gaatacattt ctaagtttgt ggaaagagaa caggataaag gtcatatagg</td><td>agatcaggag</td><td> 13380</td>

<td>ctaccgcctg aagaggaccc ttctcgtgaa ctaaacaatg tacaacatga</td><td>agrtcaatagt</td><td> 13440</td>

<td>ttaacggaac aagatgcgga ggcggatgaa ggattgtggg gtgaaataga</td><td>ttcattatgt</td><td> 13500</td>

<td>gaaaaatggc agtctgaagc ggagagtcaa actgaggcgg agataatagc</td><td>cgacaggata</td><td> 13560</td>

attggaaata gccagaggat ggcgaacctc aaaattcgtc gtacaaagtt caaaagtgtc 13620 ttgtatcata tactaaagga actaattcaa tctcagggaa ccgtaaaggt ttatcgcggt 13680 agtagttttt cacacgattc gataaagata agcttacatt atgaagagca gcatattaca 13740 gccgtatggg tctacttgat agtaaaattt gaagagcatt ggaagcctgt tgatgtagag 13800

108

<td>gtcgagttta</td><td>gatgcaagtt</td><td>caaggagcga</td><td>aaggtggatg <</td><td>ggtaggttat i</td><td>atagggatat</td><td> 13860</td>

<td>agcacagaga</td><td>tatatagcaa</td><td>agagatactt</td><td>ttgaggcaat <</td><td>gtttgtggaa <</td><td>gcggtattcg</td><td> 13920</td>

<td>caatatttta</td><td>gtagctcgtt</td><td>acagtccggt</td><td>gcgtttttgg</td><td>ttttttgaaa i</td><td>gtgcgtcttc</td><td> 13980</td>

<td>agagcgcttt</td><td>tggttttcaa</td><td>aagcgctctg</td><td>aagttcctat</td><td>actttctaga ;</td><td>gaataggaac</td><td> 14040</td>

<td>ttcggaatag</td><td>gaacttcaaa</td><td>gcgtttccga</td><td>aaacgagcgc</td><td>ttccgaaaat</td><td>gcaacgcgag</td><td> 14100</td>

<td>ctgcgcacat</td><td>acagctcact</td><td>gttcacgtcg</td><td>cacctatatc</td><td>tgogtgttgc</td><td>ctgtatatat</td><td> 14160</td>

<td>atatacatga</td><td>gaagaacggc</td><td>atagtgcgtg</td><td>tttatgctta</td><td>aatgcgtact</td><td>tatatgcgtc</td><td> 14220</td>

<td>tatttatgta</td><td>ggatgaaagg</td><td>tagtctagta</td><td>cctcctgtga</td><td>tattatccca</td><td>ttccatgcgg</td><td> 14280</td>

<td>ggtatcgtat</td><td>gcttccttca</td><td>gcactaccct</td><td>ttagctgttc</td><td>tatatgctgc</td><td>cactcctcaa</td><td> 14340</td>

<td>ttggattagt</td><td>ctcatccttc</td><td>aatgcattca</td><td>tttcctttga</td><td>tattggatca</td><td>taccctagaa</td><td> 14400</td>

<td>gtattacgtg</td><td>attttctgcc</td><td>ccttaccctc</td><td>gttgctactc</td><td>tccttttttt</td><td>cgtgggaacc</td><td> 14460</td>

<td>gctttagggc</td><td>cctcagtgat</td><td>ggtgttttgt</td><td>aatfctatatg</td><td>ctcctcttgc</td><td>atttgtgtct</td><td> 14520</td>

<td>ctacttcttg</td><td>ttcgcctgga</td><td>gggaacttct</td><td>tcatttgtat</td><td>tagcatggtt</td><td>caçttcagtc</td><td> 14580</td>

<td>cttcettcca</td><td>actcactctt</td><td>tttttgctgt</td><td>aaacgattct</td><td>ctgeegccag</td><td>ttcattgaaa</td><td> 14640</td>

<td>çtattgaata</td><td>tatcctttag</td><td>agattccggg</td><td>atgaataaat</td><td>cacctattaa</td><td>agcagcttga</td><td> 14700</td>

<td>cgatctggtg</td><td>gaactaaagt</td><td>aagcaattgg</td><td>gtaacgacgc</td><td>ttacgagctt</td><td>cataacatct</td><td> 14760</td>

<td>tcttccgttg</td><td>gagctggtgg</td><td>gactaataac</td><td>tgtgtacaat</td><td>ccatttttct</td><td>catgagcatt</td><td> 14820</td>

<td>tcggtagctc</td><td>tcttcttgtc</td><td>tttctcgggc</td><td>aatettccta</td><td>ttattatagc</td><td>aatagatttg</td><td> 14880</td>

<td>tatagttgct</td><td>ttctattgtc</td><td>taacagcttg</td><td>ttattctgta</td><td>gcatcaaatc</td><td>tatggcagcc</td><td> 14940</td>

<td>tgacttgctt</td><td>cttgtgaaga</td><td>gagcatacca</td><td>tttccaatcg</td><td>aagatacgct</td><td>ggaatcttct</td><td> 15000</td>

<td>gcgctagaat</td><td>caagaccata</td><td>cggcctaccg</td><td>gttgtgagag</td><td>attccatggg</td><td>ccttatgaca</td><td> 15060</td>

<td>tatcctggaa</td><td>agagtagctc</td><td>atcagactta</td><td>cgtttactct</td><td>ctatatcaat</td><td>atctacatca</td><td> 15120</td>

<td>ggagcaatca</td><td>tttcaataaa</td><td>cagccgacat</td><td>acatcccaga</td><td>cgctataagc</td><td>tgtacgtgct</td><td> 15180</td>

<td>tttaccgtca</td><td>gattctfcggc</td><td>tgtttcaatg</td><td>tcgtccattt</td><td>tggttttctt</td><td>ttaccagtat</td><td> 15240</td>

<td>tgttcgtttg</td><td>ataatgtatt</td><td>cttgcttatt</td><td>acattataaa</td><td>atctgtgcag</td><td>atcacatgtc</td><td> 15300</td>

<td>aaaacaactt</td><td>tttatcacaa</td><td>gatagtaccg</td><td>caaaacgaac</td><td>ctgcgggccg</td><td>tctaaaaatt</td><td> 15360</td>

aaggaaaagc agcaaaggtg catttttaaa atatgaaatg aagataccgc agtaccaatt 15420 attttcgcag tacaaataat gcgcggccgg tgcatttttc gaaagaacgc gagacaaaca 15480 ggacaattaa agttagtttt tcgagttagc gtgtttgaat actgcaagat acaagataaa 15S40 tagagtagtt gaaactagat atcaattgca cacaagatcg gcgctaagca tgccacaatt 15600 tggtatatta tgtaaaacac -cacctaaggt gcttgttcgt cagtttgtgg aaaggtttga 15660

109

<td>aagaccttca</td><td>ggtgagaaaa</td><td>tagcattatg</td><td>tgctgctgaa</td><td>ctaacctatt</td><td>tatgttggat</td><td>1S720</td>

<td>gattacacat</td><td>aacggaacag</td><td>caatcaagag</td><td>agccacattc</td><td>atgagctata</td><td>atactatcat</td><td>1S780</td>

<td>aagcaattcg</td><td>ctgagtttcg</td><td>atattgtcaa</td><td>taaatcactc</td><td>cagtttaaat</td><td>acaagacgca</td><td> 15840</td>

<td>aaaagcaaca</td><td>attctggaag</td><td>cctcattaaa</td><td>gaaattgatt</td><td>cctgcttggg</td><td>aatttacaat</td><td> 15900</td>

<td>tattccttac</td><td>tatggacaaa</td><td>aacatcaatc</td><td>tgatatcact</td><td>gatattgtaa</td><td>gtagtttgca</td><td> 15960</td>

<td>attacagttc</td><td>gaatcatcgg</td><td>aagaagcaga</td><td>taagggaaat</td><td>agccacagta</td><td>aaaaaatgct</td><td> 16020</td>

<td>aaagcacttc</td><td>taagtgaggg</td><td>tgaaagcatc</td><td>tgggagat-ca</td><td>ctgagaaaat</td><td>actaaattcg</td><td> 16080</td>

<td>tttgagtata</td><td>cttcgagatt</td><td>tacaaaaaca</td><td>aaaactttat</td><td>accaattcct</td><td>cttcctagct</td><td> 16140</td>

<td>actttcatca</td><td>attgtggaag</td><td>attcagcgat</td><td>attaagaacg</td><td>ttgatccgaa</td><td>atcatttaaa</td><td> 16200</td>

<td>ttagtccaaa</td><td>ataagtatct</td><td>gggagtaata</td><td>atccagtgtt</td><td>tagtgacaga</td><td>gacaaagaca</td><td> 16260</td>

<td>agcgttagta</td><td>ggcacatata</td><td>cttctttagc</td><td>gcaaggggta</td><td> 9</td><td></td><td> 16301</td>

<210> 48 <211> 4989 <212> DNA <213> vetor pYIG7 <400> 48

<td>agcgcccaat</td><td>acgcaaaccg</td><td>cctctccccg</td><td>cgçgttggcc</td><td>gattcattaa</td><td>tgcagctggc</td><td> 60</td>

<td>acgacaggtt</td><td>tcccgacfcgg</td><td>aaãgcgggcâ</td><td>gtgâgCgeaa</td><td>CgCaattàât</td><td>gtgagvtage</td><td> 120</td>

<td>tcactcatta</td><td>ggcaccccag</td><td>gctttacact</td><td>ttatgcttcc</td><td>ggctcgtatg</td><td>ttgtgtggaà</td><td> 180</td>

<td>ttgtgagcgg</td><td>ataacaattt</td><td>cacacaggaa</td><td>acagctatga</td><td>ocatgattac</td><td>gaatttaata</td><td> 240</td>

<td>cgactcacta</td><td>tagggaattc</td><td>ggatccttca</td><td>atatgcgcac</td><td>atacgctgtt</td><td>atgttcaagg</td><td> 300</td>

<td>tcocttcgtt</td><td>taagaacgaa</td><td>agcggtcttc</td><td>cttttgaggg</td><td>atgtttcaag</td><td>ttgttcaaat</td><td> 360</td>

<td>ctatcaaatt</td><td>tg-caaatccc</td><td>cagtctgtat</td><td>etagagegtt</td><td>gaatcggtga</td><td>tgcgatttgt</td><td> 420</td>

<td>taattaaatt</td><td>gatggtgtca</td><td>eeafcfcaçeag</td><td>gtctagatat</td><td>accaatggca</td><td>aactgagcac</td><td> 480</td>

<td>aacaatacca</td><td>gtccggatca</td><td>actggcacca</td><td>tctctcccgt</td><td>agtctcatct</td><td>aatttttctt</td><td>S40</td>

<td>ccggatgagg</td><td>ttccagatat</td><td>accgcaacac</td><td>ctttattatg</td><td>gtttcectga</td><td>gggaataata</td><td> €00</td>

<td>gaatgtccca</td><td>ttcgaaatca</td><td>ccaattctaa</td><td>acctgggcga</td><td>atfcgtatttc</td><td>gggtttgtta</td><td> 660</td>

<td>actcgttcca</td><td>gtcaggaatg</td><td>ttceacgtga</td><td>agctatcttc</td><td>cagcaaagtc</td><td>tccacttctt</td><td> 720</td>

<td>catcaaattg</td><td>tggagaatac</td><td>tcccaatgct</td><td>cttatctatg</td><td colspan="2">ggacttccgg gaaacacagt</td><td> 780</td>

<td>accgatactt</td><td>cccaattcgt</td><td>cttcagagct</td><td>cattgtttgt</td><td>ttgaagagac</td><td>taatcaaaga</td><td> 840</td>

<td>atcgttttct</td><td>caaaaaaatt</td><td>aatatcttaa</td><td>ctgatagttt</td><td>gatcaaaggg</td><td>gcaaaacgta</td><td> 900</td>

<td>ggggcaaaca</td><td>aacggaaaaa</td><td>tcgtttctca</td><td>aattttctga</td><td>tgccaagaac</td><td>tctaaccagt</td><td> 960</td>

110

<td>cttatcfcaaa aattgcctta tgatccgtct</td><td>ctccggttac agcctgtgta actgattaat</td><td> 1020</td>

<td>cctgcctttc taatcaccat tctaatgttt</td><td>taattaaggg attttgtctt cattaacggc</td><td> 1080</td>

<td>tttcgctcat aaaaatgtta tgacgttttg</td><td>cccgcaggcg ggaaaccatc cacttcacga</td><td> 1140</td>

<td>gactgatctc ctctgccgga acaccgggca</td><td>tctccaactt ataagttgga gaaataagag</td><td> 1200</td>

<td>aatttcagat tgagagaatg aaaaaaaaaa</td><td>accctgaaaa aaaaggttga aaccagttcc</td><td> 1260</td>

<td>ctgaaattat tcccctactt gactaataag</td><td>tatataaaga cggtaggtat tgattgtaat</td><td> 1320</td>

<td>tctgtaaatc tatttcttaa acttcttaaa</td><td>ttctactttt atagttagtc ttttttttag</td><td> 1380</td>

<td>ttttaaaaca ccaagaactt agtttcgaat</td><td>aaacacacat aaacaaacac catgaggtct</td><td> 1440</td>

<td>ttgctaatac tagtgctttg cttcctgccc</td><td>ctggctgctc tgggggtacc agatctcgac</td><td> 1500</td>

<td>ttggttgaac acgttgccaa ggcttaagtg</td><td>aatttacttt aaagtcttgc atttaaataa</td><td> 1560</td>

<td>attttctttt tatagcttta tgacttagtt</td><td>tcaatttata tactatttta atgacatttt</td><td> 1620</td>

<td>cgattcattg attgaaagct ttgtgttttt</td><td>tcttgatgcg etattgcatt gttcttgtct</td><td> 1680</td>

<td>ttttcgccac atgtaatatc tgtagtagat</td><td>acctgataca ttgtggatgc tgagtgaaat</td><td> 1740</td>

<td>tttagttaat aatggaggcg ctcttaataa</td><td>ttttggggat attggctttt ttttttaaag</td><td> 1800</td>

<td>tttacaaatg aattttttcc gccaggataa</td><td>cgattctgaa gttactctta gcgttcctat</td><td> 1860</td>

<td>cggtacagcc atcaaafccat gcctataaat</td><td>catgcctata tttgcgtgca gtcagtafcca</td><td> 1920</td>

<td>tctacatgaa aaaaactccc gcaatttctt</td><td>atagaatacg ttgaaaatta aatgtacgcg</td><td> 1980</td>

ccaagataag ataacatata tctagctaga Vgcagtaata tacacagatt cccgcggacg 2040 tgggaaggaa aaaattagat aacaaaatct gagtgatatg gaaattcogc tgtatagcfcc 2100 atatcttfccc cttcaacacc agaaatgtaa aaatcttgtt acgaaggatc tttttgctaa 2160 tgtttctcgc tcaatcctca tttcttccct acgaagagtc aaatctactt gttttctgcc 2220 ggtatcaaga tccatatctt ctagtttcac catcaaagtc caatttctag tatacagttt 2280 atgtcccaac gtaacagaca atcaaaattg gaaaggataa gtatccttca aagaatgatt 2340 ctgcgctggc tcctgaaccg cctaatggga acagagaagt ccaaaacgat gctataagaa 2400 ccagaaataa aacgataaaa ccataccagg atccaagctt ggcactggcc gtcgttttac 2460 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatccce 2520 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga togoccttcc caacagttgc 2580 gcagcctgaa tggcgaatgg gaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa 2640 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata 2700 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac 2760

111

<td>tattaaagaa</td><td>cgtggactcc</td><td>aacgtcaaag</td><td>ggcgaaaaac</td><td>cgtctatcag ggcgatggee</td><td> 2820</td>

<td>cactacgtga</td><td>accatcaccc</td><td>taatcaagtt</td><td>ttttggggtc</td><td>gaggtgccgt aaagcactaa</td><td>28B0</td>

<td>atcggaaccc</td><td>taaagggagc</td><td>ccccgafctta</td><td>gagcttgacg</td><td>gggaaagccg gcgaacgtgg</td><td> 2940</td>

<td>cgagaaagga</td><td>agggaagaaa</td><td>gcgaaaggag</td><td>cgggcgctag</td><td>ggcgctggca agtgtagpcgg</td><td> 3000</td>

<td>tcacgctgcg</td><td>cgtaaccacc</td><td>acacccgccg</td><td>cgcttaatgc</td><td>gccgctacag ggcgcgtcag</td><td> 3060</td>

<td>gtggcacttt</td><td>tcgçggaaat</td><td>gtgcgcggaa</td><td>cccctatttg</td><td>tttatttttc taaatacatt</td><td> 3120</td>

<td>caaatatgta</td><td>tccgctcatg</td><td>agacaataac</td><td>cctgataaat</td><td>gcttcaataa tattgaaaaa</td><td> 3180</td>

<td>ggaagagtat</td><td>gagtattcaa</td><td>catttccgtg</td><td>tcgcccttat</td><td>tccctttttt gcggcatttt</td><td> 3240</td>

<td>gccttcctgt</td><td>ttttgctcac</td><td>ccagaaacgc</td><td>tggtgaaagt</td><td>aaaagatgct gaagatcagt</td><td> 3300</td>

<td>tgggtgcacg</td><td>agtgggttac</td><td>atcgaactgg</td><td>atctcaacag</td><td>cggtaagatc cttgagagtt</td><td> 3360</td>

<td>ttcgccccga</td><td>agaacgtttt</td><td>ccaatgatga</td><td>gcacttttaa</td><td>agttctgcta tgfeggcgcgg</td><td> 3420</td>

<td>tattatcccg</td><td>tattgacgcc</td><td>gggcaagagc</td><td>aactcggtcg</td><td>ccgcatacac tattctcaga</td><td> 3480</td>

<td>atgacttggt</td><td>tgagtactca</td><td>ccagtcacag</td><td>aaaagcatct</td><td>tacggatggc atgacagtaa</td><td>3S40</td>

gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga 3600 caacgafccgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa 3660 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca 3720 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta 3780 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac 3840 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc 3900 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagocctcc cgtatcgtag 3960 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga 4020 taggtgcctc actgattaag cattggtaae tgtcagacca agtttactca tatatacttt 4080 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata 4140 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag 4200 aaaagatcaa aggatcttct tgagatcctt tttttctgcg egtaatcfcgc tg-cttgcaaa 4260 caaaaaaacc acegctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt 4320 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc 4380 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacataoctc gcfcctgctaa 4440 tcctgttacc agtggçtgct gccagtggcg ataagtcgtg tcttacoggg ttggactcaa 4500 gacgatagtt accggataag gcgcagcggfc cgggctgaac ggggggttcg tgcacacagc 4560 ccagcttgga gcgaacgaoc tacaccgaac tgagataect acagcgtgag cattgagaaa 4620

112 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggççttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccttcg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaag

46B0

4740

4800

4860

4920

4980

4989

<td colspan="2"><212> 5422 <212> DNA <213> vetor pY!G7El <400> 49 agcgcccaat acgcaaaccg</td><td>cctctccccg</td><td>cgcgttggcc</td><td>gattcattaa</td><td>tgcagctggc</td>

<td>acgacaggtt</td><td>tcccgactgg</td><td>aaagcgggca</td><td>gfcgagcgcaa</td><td>cgcaattaat</td><td>gtgagttagc</td>

<td>tcactcatta</td><td>ggcaccccag</td><td>gctttacact</td><td>ttatgcttcc</td><td>ggctcgtatg</td><td>ttgtgtggaa</td>

<td>ttgtgagcgg</td><td>ataacaattt</td><td>cacacaggaa</td><td>acagctatga</td><td>ccatgattac</td><td>gaatttaata</td>

<td>cgactcacta</td><td>tagggaattc</td><td>ggatccttca</td><td>atatgcgcac</td><td>atacgctgtt</td><td>atgttcaagg</td>

<td>tcccttcgtt</td><td>taagaacgaa</td><td>agcggtcttc</td><td>cttttgaggg</td><td>atgtttcaag</td><td>ttgttcaaat</td>

<td>ctatcaaatt</td><td>tgcaaatcec</td><td>cagtctgtat</td><td>etagagcgtt</td><td>gaatcggtga</td><td>tgcgatttgt</td>

<td>taattaaatt</td><td>gatggtgtca</td><td>ccattaccag</td><td>gtctagatat</td><td>accaatggca</td><td>aactgagcac</td>

<td>aacaatacca</td><td>gtccggatca</td><td>actggcacca</td><td>tctctcccgt</td><td>agtctcatct</td><td>aatttttctt</td>

<td>ccggatgagg</td><td>ttccagatat</td><td>accgcaacac</td><td>ctttattatg</td><td>gtttccctga</td><td>gggaataata</td>

<td>gaatgtccca</td><td>ttcgaaatca</td><td>ccaattctaa</td><td>acctgggcga</td><td>attgtatttc</td><td>gggtttgtta</td>

<td>actcgttcca</td><td>gtcaggaatg</td><td>ttccacgtga</td><td>agctatcttc</td><td>cagcaaagtc</td><td>tccacttctt</td>

<td>catcaaattg</td><td>tggagaatac</td><td>tcccaatgct</td><td>cttatctatg</td><td>ggacttccgg</td><td>gaaacacagt</td>

<td>accgatactt</td><td>cccaatfccgt</td><td>cttcagagct</td><td>cattgtttgt</td><td>ttgaagagac</td><td>taatcaaaga</td>

<td>atcgttttct</td><td>caaaaaaatt</td><td>aatatcttaa</td><td>etgatagttt</td><td>gatcaaaggg</td><td>gcaaaacgta</td>

<td>ggggcaaaca</td><td>aaoggaaaaa</td><td>tcgtttctca</td><td>aattttctga</td><td>tgccaagaac</td><td>tctaaccagt</td>

<td>cttatctaaa</td><td>aattgcctta</td><td>tgatccgtct</td><td>ctccggttac</td><td>agcctgtgta</td><td>actgattaat</td>

<td>cctgcctttc</td><td>taatcaccat</td><td>tctaatgttt</td><td>taattaaggg</td><td>attttgtctt</td><td>cattaacggc</td>

<td>tttcgctcat</td><td>aaaaatgtta</td><td>tgacgttttg</td><td>cccgcaggcg</td><td>ggaaaccatc</td><td>cacttcacga</td>

€0

120

180

240

300

360

420

480

540 €00

660

720

780

840

900

960

1020

1080

1140 gactgatctc ctctgccgga acaccgggca tctccaactt ataagttgga gaaataagag 1200

113

<td>aatttcagat</td><td>tgagagaatg</td><td>aaaaaaaaaa</td><td>accctgaaaa</td><td>aaaaçgttga <</td><td>aaccagfctcc</td><td> 1260</td>

<td>ctgaaattat</td><td>tcccctactt</td><td>gactaataag</td><td>tatataaaga</td><td>cggtaggtat :</td><td>tgattgtaat</td><td> 1320</td>

<td>tctgtaaatc</td><td>tatttcttaa</td><td>acttcttaaa</td><td>ttctactttt</td><td>atagttagtc</td><td>ttttttttag</td><td> 1380</td>

<td>ttttaaaaca</td><td>ccaagaactt</td><td>agtttcgaat</td><td>aaacacacat</td><td>aaacaaacac</td><td>catgaggtct</td><td> 1440</td>

<td>ttgctaatac</td><td>tagtgctttg</td><td>cttcctgccc</td><td>ctggctgctc</td><td>tggggtatga</td><td>ggtgcgcaac</td><td> 1500</td>

<td>gtgtccggga</td><td>tgtaccatgt</td><td>cacgaacgac</td><td>tgctccaact</td><td>caagcattgfc</td><td>gtatgaggca</td><td> 1560</td>

<td>gcggacatga</td><td>tcatgcacac</td><td>ccccgggtgc</td><td>gtgccctgcg</td><td>tt-cgggagaa</td><td>caactcttcc</td><td> 1620</td>

<td>cgctgctggg</td><td>tagcgctcac</td><td>ccccaegctc</td><td>gcagctagga</td><td>acgceagcgt</td><td>ccccaccacg</td><td> 1680</td>

<td>acaatacgac</td><td>gccacgtcga</td><td>tttgctcgtt</td><td>ggggcggctg</td><td>ctttctgttc</td><td>cgctatgtac</td><td> 1740</td>

<td>gtgggggacc</td><td>tctgcggatc</td><td>tgtcttcctc</td><td>gtctcccagç</td><td>tgttcaccat</td><td>ctcgcctcgc</td><td> 1800</td>

<td>cggcatgaga</td><td>cggtgcagga</td><td>ctgcaattgc</td><td>tcaatctatc</td><td>ccggccacat</td><td>aacgggtcac</td><td> 1860</td>

<td>cgtatggctt</td><td>gggatatgat</td><td>gatgaactgg</td><td>taatagaccc</td><td>ttctcacctc</td><td>ggccgataag</td><td> 1920</td>

<td>ctcagatctc</td><td>gacttggttg</td><td>aacacgttgc</td><td>caaggcttaa</td><td>gtgaatttac</td><td>tttaaagtct</td><td> 1980</td>

<td>tgcatttaaa</td><td>taaattttct</td><td>ttttatagct</td><td>ttatgactta</td><td>gtttcaattt</td><td>atatactatt</td><td> 2040</td>

<td>ttaatgacat</td><td>tttcgattca</td><td>ttgattgaaa</td><td>gctttgtgtt</td><td>ttttcttgat</td><td>gcgctattgc</td><td> 2100</td>

<td>attgttcttg</td><td>tctttttcgc</td><td>cacatgtaat</td><td>atctgtagta</td><td>gatacctgat</td><td>acattgtgga</td><td> 2160</td>

<td>tgctgagtga</td><td>aattttagtt</td><td>aataatggag</td><td>gcgctcttaa</td><td>taattttggg</td><td>gatattggct</td><td> 2220</td>

<td>ttttfctttta</td><td>aagtttacaa</td><td>atgaattttt</td><td>tccgccagga</td><td>taacgatt-ct</td><td>gaagttaetc</td><td> 2280</td>

<td>ttagcgttcc</td><td>tatcggtaca</td><td>gccatcaaat</td><td>catgcctata</td><td>aatcatgcct</td><td>atatttgcgt</td><td> 2340</td>

<td>gcagtcagta</td><td>tcatctacat</td><td>gaaaaaaact</td><td>cccgcaattt</td><td>cttatagaat</td><td>acgttgaaaa</td><td> 2400</td>

<td>ttaaatgtac</td><td>gcgccaagat</td><td>aagataacat</td><td>atatctagct</td><td>agatgcagta</td><td>. atatacacag</td><td> 2460</td>

attcccgcgg acgtgggaag gaaaaaatta gataacaaaa tctgagtgat atggaaattc 2520 cgctgtatag ctcatatctt toccttcaac aocagaaatg taaaaatctt gttacgaagg 2580 atctttttgc taatgtttct cgctcaatcc tcatttcttc ectacgaaga gtcaaatcta 2640 cttgttttct gccggtafcca agatccatat cttctagttt caccatcaaa gtccaattfcc 2700 tagtatacag tttatgtccc aacgtaacag acaatcaaaa ttggaaagga taagtatcct 2760 tcaaagaatg attctgcgct ggctcctgaa ccgcctaatg ggaacagaga agtccaaaac 2820 gatgctataa gaaccagaaa taaaacgata aaaccatacc aggatccaag cttggcactg 2880 gccgtcgttt tacaaogfccg tgactgggaa aaccctggcg ttacccaact taatcgcctt 2940 gcagcacate cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct 3000

114

<td>tcccaacagt</td><td>tgcgcagcct</td><td>gaatggcgaa</td><td>tgggaaattg</td><td colspan="2">taaacgttaa tattttgtta</td><td> 3060</td>

<td>aaattcgcgt</td><td>taaatttttg</td><td>ttaaatcagc</td><td>tcatttttta</td><td>accaataggc</td><td>cgaaatcggc</td><td> 3120</td>

<td>aaaatccctt</td><td>ataaatcaaa</td><td>agaatagacc</td><td>gagatagggt</td><td>tgagtgttgt</td><td>tccagtttgg</td><td> 3180</td>

<td>aacaagagtc</td><td>cactattaaa</td><td>gaacgtggac</td><td>tccaacgtca</td><td>aagggcgaaa</td><td>aaccgtctat</td><td> 3240</td>

<td>cagggcgatg</td><td>gcccactacg</td><td>tgaaccatca</td><td>ccctaatcaa</td><td>gttttttggg</td><td>gtcgaggtgc</td><td> 3300</td>

<td>cgtaaagcac</td><td>taaatcggaa</td><td>occtaaaggg</td><td>agcccccgat</td><td>•tt.agagct.tg</td><td>acggggaaag</td><td> 3360</td>

<td>ccggcgaacg</td><td>tggcgagaaa</td><td>ggaagggaag</td><td>aaagcgaaag</td><td>gagcgggcgc</td><td>tagggcgctg</td><td> 3420</td>

<td>gcaagtgtag</td><td>cggtcacgct</td><td>gcgcgtaacc</td><td>accacacccg</td><td>ccgcgcttaa</td><td>tgcgccgcta</td><td> 3480</td>

<td>cagggcgcgt</td><td>caggtggeac</td><td>ttttcgggga</td><td>aatgtgcgcg</td><td>gaacccctat</td><td>ttgtttattt</td><td> 3540</td>

<td>ttctaaatac</td><td>attcaaatat</td><td>gtatccgctc</td><td>atgagacaat</td><td>aaccctgata</td><td>aatgcttcaa</td><td> 3600</td>

<td>taatattgaa</td><td>aaaggaagag</td><td>tatgagtatt</td><td>caacattfccc</td><td>gtgtcgccct</td><td>tattcccttt</td><td> 3660</td>

<td>tttgcggcat</td><td>tttgccttcc</td><td>tgtttttgct</td><td>cacccagaaa</td><td>cgctggtgaa</td><td>agtaaaagat</td><td> 3720</td>

<td>gctgaagatc</td><td>agttgggtgc</td><td>acgagtgggt</td><td>tacatcgaac</td><td>tggatctcaa</td><td>cagcggtaag</td><td> 3 780</td>

<td>atccttgaga</td><td>gttttcgccc</td><td>cgaagaacgt</td><td>tttccaatga</td><td>tgagcacttt</td><td>taaagttctg</td><td> 3840</td>

<td>ctatgtggcg</td><td>cggtattatc</td><td>ccgtattgac</td><td>gccgggcaag</td><td>agcaactcgg</td><td>tcgccgcata</td><td> 3500</td>

<td>cactattete</td><td>agaatgactt</td><td>ggttgagtac</td><td>tcaccagtca</td><td>cagaaaagca</td><td>fccfcfc acncra t</td><td> 3560</td>

<td>ggcatgacag</td><td>taagagaatt</td><td>atgcagtgct</td><td>gecataacca</td><td>tgagtgataa</td><td>cactgcggcc</td><td> 4020</td>

<td>aacttacttc</td><td>tgacaacgat</td><td>cggaggaccg</td><td>aaggagctaa</td><td>ccgctttttt</td><td>gcacaacatg</td><td> 4080</td>

<td>ggggatcatg</td><td>taactcgcct</td><td>tgatcgttgg</td><td>gaaccggagc</td><td>tgaatgaagc</td><td>cataccaaac</td><td> 4140</td>

gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact 4200 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa 4260 gttgeaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct 4320 ggagccggtg agcgtgggtc fccgcggtatc attgcagcac tggggecaga tggtaagccc 4380 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga 4440 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac 4500 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag 4560 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agrttttcgtt ccactgagcg 4620 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc 4680 tgctgcttgc aaacaaaaaa accaccgcta ccageggtgg tttgtttgcc ggatcaagag 4740 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc 4800 cttctagtgt agocgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac 4860

115 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc 4920 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt 4980 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt 5040 gagcattgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc 5100 ggcagggteg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt 5160 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca 5220 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt 5260 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt 5340 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag 5400 tcagtgagcg aggaagcgga ag 5422 <210> 50 <211> 15621 <212> DNA <213> vetor pSYlYIG7Els <400> 50

<td>atcgataagc</td><td>ttttcaattc</td><td>aattcatcat</td><td>ttttttttta</td><td>ttcttttttt</td><td>tgattfccggt</td><td> 60</td>

<td>ttctttgaaa</td><td>tttttttgat</td><td>tcggtaatct</td><td>ccgaacagaa</td><td>ggaagaacga</td><td>aggaaggagc</td><td> 120</td>

<td>acagacttag</td><td>attggtatat</td><td>atacgcatat</td><td>gtagtgttga</td><td>agaaacatga</td><td>aattgcccag</td><td> 180</td>

<td>tattcttaac</td><td>ccaactgcac</td><td>agaacaaaaa</td><td>cctgcaggaa</td><td>acgaagataa</td><td>atcatgtcga</td><td> 240</td>

<td>aagctacata</td><td>taaggaacgt</td><td>gctgctactc</td><td>atcctagtcc</td><td>tgttgctgcc</td><td>aagctattta</td><td> 300</td>

<td>atatcatgca</td><td>cgaaaagcaa</td><td>acaaacttgt</td><td>gtgcttcatt</td><td>ggatgttcgt</td><td>accaccaagg</td><td> 360</td>

<td>aattactgga</td><td>gttagtfcgaa</td><td>gcattaggtc</td><td>ccaaaatttg</td><td>tttactaaaa</td><td>acacatgtgg</td><td> 420</td>

<td>atatcttgac</td><td>tgatttttcc</td><td>atggagggca</td><td>cagttaagcc</td><td>gctaaaggca</td><td>ttatccgcca</td><td> 480</td>

<td>agtacaattt</td><td>tttactcttc</td><td>gaagacagaa</td><td>aatttgctga</td><td>cattggtaat</td><td>acagbcaaat</td><td> 540</td>

<td>tgcagtactc</td><td>tgcgggtgta</td><td>tacagaatag</td><td>cagaatgggc</td><td>agacattacg</td><td>aatgcacacg</td><td> 600</td>

<td>STtgtggtggg</td><td>cccaggtatt</td><td>gttagoggtt</td><td>tgaagcaggc</td><td>ggcagaagaa</td><td>gtaacaaagg</td><td> 660</td>

<td>aacctagagg</td><td>ccttttgatg</td><td>ttagcagaat</td><td>tgtcatgcaa</td><td>gggctoccta</td><td>tctactggag</td><td> 720</td>

<td>aatatactaa</td><td>gggtactgtt</td><td>gacattgcga</td><td>agagcgacaa</td><td>agattttgtt</td><td>atcggcttta</td><td> 780</td>

<td>ttgcfccaaag</td><td>agacatgggt</td><td>ggaagagatg</td><td>aaggttacga</td><td>ttggttgatt</td><td>atgacacccg</td><td> 840</td>

<td>gtgtgggttt</td><td>agatgacaag</td><td>ggagacgcat</td><td>tgggfccaaca</td><td>gtatagaacc</td><td>gtggatgatg</td><td> 900</td>

<td>tggtctctac</td><td>aggatctgac</td><td>attattattg</td><td>ttggaagagg</td><td>actatttgca</td><td>aagggaaggg</td><td> 960</td>

atgctaaggt agagggtgaa cgttacagaa aagcaggctg ggaagcatat ttgagaagat 1020

116 gcggccagca aaactaaaaa actgtattat aagtaaatgc atctatacta aactcacaaa 1080 ttagagcttc aatttaatta tatcagttat tacccgggaa tctcggtcgt aatgattttt 1140 ataa.tgacga aaaaaaaaaa attggaaaga aaaagcttta atgcggtagt ttatcacagt 1200 taaattgcta acgcagtcag gcaccgtgta tgaaatctaa caatgcgctc atcgtcatcc 1260

<td>tcggcaccgt</td><td>caccctggat</td><td>gctgtaggca</td><td>taggcttggt</td><td>tatgccggta 1</td><td>ctgccgggcc</td><td> 1320</td>

<td>tcttgcggga</td><td>tatcgtccat</td><td>tccgacagca</td><td>tcgccagtca</td><td>ctatggcgtg</td><td>ctgctagcgc</td><td>13B0</td>

<td>tatatgcgtt</td><td>gatgcaattt</td><td>ctatgcgcac</td><td>ccgttctcgg</td><td>agcactgtcc</td><td>gaccgctttg</td><td> 1440</td>

<td>gccgccgccc</td><td>agtcctgctc</td><td>gcttcgctac</td><td>ttggagccac</td><td>tatcgactac</td><td>gcgatcatgg</td><td> 1500</td>

<td>cgaccacacc</td><td>cgtcctgtgg</td><td>atcctggtat</td><td>ggttttatcg</td><td>ttttatttct</td><td>ggttcttata</td><td> 1560</td>

<td>gcatcgtttt</td><td colspan="2">ggacttctct gttcccatta</td><td>ggcggttcag</td><td>gagccagcgc</td><td>agaatcatfcc</td><td> 1620</td>

<td>tttgaaggat</td><td>acttatcctt</td><td>tccaattttg</td><td>attgtctgtt</td><td>acgttgggac</td><td>ataaactgta</td><td> 1660</td>

<td>tactagaaat</td><td>tggactttga</td><td>tggtgaaact</td><td>agaagatatg</td><td>gatcttgata</td><td>ccggcagaaa</td><td> 1740</td>

<td>acaagtagat</td><td>ttgactcttc</td><td>gtagggaaga</td><td>aatgaggatt</td><td>gagcgagaaa</td><td>cattagcaaa</td><td> 1800</td>

<td>aagatccttc</td><td>gtaacaagat</td><td>ttttacattt</td><td>ctggtgttga</td><td>agggaaagat</td><td>atgagctata</td><td> 1860</td>

<td>cagcggaatt</td><td>tccatatcac</td><td>tcagattttg</td><td>ttatctaatt</td><td>ttttccttcc</td><td>cacgtccgcg</td><td> 1920</td>

<td>ggaatctgtg</td><td>tatattactg</td><td>catctagcta</td><td>gatatatgtt</td><td>atcttatctt</td><td>ggcgcgtaca.</td><td> 1980</td>

<td>tttaattttc</td><td>aacgtattCt</td><td>ataagaaatt</td><td>gcgggagttt</td><td>ttttcatgta</td><td>gatgatactg</td><td> 2040</td>

<td>actgcacgca</td><td>aatataggca</td><td>tgatttatag</td><td>gcatgatttg</td><td>atggctgtac</td><td>cgataggaac</td><td> 2100</td>

<td>gctaagagta</td><td>acttcagaat</td><td>cgttatcctg</td><td>gcggaaaaaa</td><td>ttcatttgta</td><td>aactttaaaa</td><td> 2160</td>

<td>aaaaaagcca</td><td>atatccccaá</td><td>aattattaag</td><td>agcgcctcca</td><td>ttattaacta</td><td>aaatttcact</td><td> 2220</td>

<td>cagcatccac</td><td>aatgtatcag</td><td>gtatetacta</td><td>cagatattac</td><td>atgtggcgaa</td><td>aaagacaaga</td><td> 2280</td>

<td>acaatgcaat</td><td>agcgcatcaa</td><td>gaaaaaacac</td><td>aaaggtttca</td><td>atcaatgaat</td><td>cgaaaatgtc</td><td> 2340</td>

<td>attaaaatag</td><td colspan="2">tatataaatt gaaactaagt</td><td>cataaagcta</td><td>taaaaagaaa</td><td>atttatttaa</td><td> 2400</td>

<td>atgcaagact</td><td>ttaaagtaaa</td><td>ttcactfcaag</td><td>ccttggcaac</td><td>gtgttcaacc</td><td>aagtcgagat</td><td> 2460</td>

<td>ctgagcttat</td><td>cggcogaggt</td><td>gagaagggtc</td><td>tattaccagt</td><td>fccatcatcat</td><td>atcccaagcc</td><td> 2520</td>

<td>atacggtgac</td><td>ccgttatgtg</td><td>gccgggatag</td><td>attgagcaat</td><td>tgcagtcctg</td><td>caccgtctca</td><td>25B0</td>

tgccggcgag gcgagatggt gaacagctgg gagacgagga agacagatcc gcagaggtcc 2640 cccacgtaca tagcggaaca gaaagcagcc gccccaacga gcaaatogac gtggcgtcgt 2700 attgtcgtgg tggggacgct ggcgttccta gctgcgagcg tgggggtgag cgctacccag 2760 cagcgggaag agttgtfcctc cagaacgpcag ggcacgcacc cgggggtgtg catgatcatg 2820

117

<td>tccgctgcct</td><td>catacacaat</td><td>gcttgagttg</td><td>gagcagtcgt</td><td>tcgtgacatg</td><td>gtacatcccg</td><td> 2880</td>

<td>gacacgttgc</td><td>gcacctcata</td><td>ccccagagca</td><td>gccaggggca</td><td>ggaagcaaag</td><td>cactagtatt</td><td> 2940</td>

<td>agcaaagacc</td><td>tcatggtgtt</td><td>tgtttatgtg</td><td>tgtttattcg</td><td>aaactaagtt</td><td>cttggtgttt</td><td> 3000</td>

<td>taaaactaaa</td><td>aaaaagacta</td><td>actataaaag</td><td>tagaatttaa</td><td>gaagtttaag</td><td>aaatagattt</td><td> 3060</td>

<td>acagaattac</td><td>aatcaatacc</td><td>tacogtcttt</td><td>atatacttat</td><td>tagtcaagta</td><td>ggggaataat</td><td> 3120</td>

<td>ttcagggaac</td><td>tggtttcaac</td><td>cttttttttc</td><td>agggtttttt</td><td>tttttcattc</td><td>tctcaatctg</td><td> 3180</td>

<td>aaattctctt</td><td>atttctccaa</td><td>cttataagtt</td><td>ggagatgccc</td><td>ggtgttccgg</td><td>cagaggagat</td><td> 3240</td>

<td>cagtctegtg</td><td>aagtggatgg</td><td>tttcccgect</td><td>gcgggcaaaa</td><td>cgtcataaca</td><td>tttttatgag</td><td> 3300</td>

<td>cgaaagccgt</td><td>taatgaagac</td><td>aaaatccctt</td><td>aattaaaaca</td><td>ttagaatggt</td><td>gattagaaag</td><td> 3360</td>

<td>gcaggattaa</td><td>tcagttacac</td><td>aggctgtaac</td><td>cggagagacg</td><td>gatcataagg</td><td>caatttttag</td><td> 3420</td>

<td>ataagactgg</td><td>ttagagttct</td><td>tggcatcaga</td><td>aaatttgaga</td><td>aacgattttt</td><td>ccgfcttgttt</td><td> 3480</td>

<td>gcccctacgt</td><td>tttgcccctt</td><td>tgatcaaact</td><td>atcagttaag</td><td>atattaattt</td><td>ttttgagaaa</td><td> 3540</td>

<td>acgattcttt</td><td>gattagtctc</td><td>ttcaaacaaa</td><td>caatgagctc</td><td>tgaagacgaa</td><td>ttgggaagta</td><td> 3600</td>

<td>tcggtactgt</td><td>gtttoccgga</td><td>agtcccatag</td><td>ataagagcat</td><td>tgggagtatt</td><td>ctccacaatt</td><td> 3660</td>

<td>tgatgaagaa</td><td>gtggagactt</td><td>tgctggaaga</td><td>tagcttcacg</td><td>tggaacattc</td><td>ctgactggaa</td><td> 3720</td>

cgagttaaca aacccgaaat acaattcgcc caggtttaga attggtgatt tcgaatggga 3780 cattctatta ttccctcagg gaaaccataa taaaggtgtt gcggtatatc tggaacctca 3640 tccggaagaa aaattagatg agactacggg agagatggtg ccagttgatc cggactggta 3900 ttgttgtgct cagtttgcca ttggtatatc tagacctggt aatggtgaca ccatcaattt 3960 aattaacaaa tcgcatcacc gattcaacgc tctagataca gactggggat ttgcaaattt 4020 gatagatttg aacaacttga aacatccctc aaaaggaaga ccgctttcgt tcttaaacga 4080 agggaccttg aacataacag cgfcatgtgcg catattgaag gatcctctac gocggacgca 4140 tcgtggccgg catcaccggc gccacaggtg cggttgctgg cocctatatc gccgacatca 4200 ccgatgggga agatcgggct cgccacttcg ggctcatgag ogcttgtttc ggcgtgggta 4260 tggtggcagg ccccgtggcc gggggactgt tgggcgocat ctccttgcat gcaccattcc 4320 ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt 4380 cgcataaggg agagogtcga cogatgccct tgagagcctt caacccagtc agctccttcc 4440 ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac 4500 tcgtaggaca ggtgocggca gcgetctggg tcattttcgg cgaggaccgc tttcgctgga 4S60 gcgcgacgat gatoggoctg tcgcttg-cgg tattcggaat cttgcacgcc ctcgctcaag 4620 ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt atcgccggca 46B0

118

<td>tggcggccga</td><td>cgcgctgggc</td><td>tacgtcttgc</td><td>tggcgttcgc</td><td>gacgcgaggc</td><td>tggatggcct</td><td> 4740</td>

<td>tccccattat</td><td>gattcttctc</td><td>gcttccggcg</td><td>gcatcgggat</td><td>gcccgcgttg</td><td>caggccatgc</td><td> 4800</td>

<td>tgtccaggca</td><td>ggtagatgac</td><td>gaccatcagg</td><td>gacagcttca</td><td>aggatcgctc</td><td>gcggctctta</td><td> 4860</td>

<td>ccagcctaac</td><td>ttcgatcact</td><td>ggaccgctga</td><td>tcgtcacggc</td><td>gatttatgcc</td><td>gcctcggcga</td><td> 4920</td>

<td>gcacatggaa</td><td>cgggttggca</td><td>tggattgtag</td><td>gcgccgccct</td><td>ataecttgtc</td><td>tgcctccccg</td><td> 4980</td>

<td>cgttgcgtcg</td><td>cggtgcatgg</td><td>agccgggcca</td><td>cctcgacctg</td><td>aatggaagcc</td><td>ggcggcacct</td><td> 5040</td>

<td>cgctaacgga</td><td>ttcaccactc</td><td>-caagaattgg</td><td>agccaatcaa</td><td>ttcttgcgga</td><td>gaactgtgaa</td><td> 5100</td>

<td>tgcgcaaacc</td><td>aacccttggc</td><td>agaacatatc</td><td>catcgcgtcc</td><td>gccatctcca</td><td>gcagccgcac</td><td> 5160</td>

<td>gcggcgcatc</td><td>tcgggcagcg</td><td>ttgggtcctg</td><td>gccacgggtg</td><td>cgcatgatcg</td><td>tgctcctgtç</td><td> 5220</td>

<td>gttgaggacc</td><td>cggctaggct</td><td>ggcggggttg</td><td>cettactggt</td><td>tagcagaatg</td><td>aatcaccgat</td><td> 5280</td>

<td>acgcgagcga</td><td>acgtgaagcg</td><td>actgctgctg</td><td>caaaacgtct</td><td>gcgacctgag</td><td>caacaacatg</td><td> 5340</td>

<td>aatggtcttc</td><td>ggtttccgtg</td><td>tttcgtaaag</td><td>tctggaaacg</td><td>cggaagt-cag</td><td>cgccctgcac</td><td> 5400</td>

<td>cattatgttc</td><td>cggatctgca</td><td>tcgcaggatg</td><td>ctgctggcta</td><td>ccctgtggaa</td><td>cacctacatc</td><td> 5460</td>

<td>tgtattaacg</td><td>aagcgctggc</td><td>attgaccctg</td><td>agtgattttt</td><td>ctctggtccc</td><td>gccgcatcca</td><td>5S20</td>

<td>taccgccagt</td><td colspan="2">tgtttaccct cacaacgttc</td><td>cagtaaccgg</td><td>gcatgttcat</td><td>catcagtaac</td><td>5S80</td>

<td>ccgtatcgtg</td><td>agcatcctct</td><td>ctcgttt-cat</td><td>cgg t at cai. t</td><td>acccccatga</td><td>acagaaattc</td><td> 5640</td>

<td>ccccttacac</td><td>ggaggcatca</td><td>agtgaccaaa</td><td>caggaaaaaa</td><td>-ccgcccttaa</td><td>catggcccgc</td><td> 5700</td>

<td>tttatcagaa</td><td>gccagac&tt</td><td>aacgctfcctg</td><td>gagaaact-cá</td><td>acgagctgga</td><td>cgcggatgaa</td><td> 5760</td>

<td>caggcagaca</td><td>tctgtgaatc</td><td>gcttcacgac</td><td>cacgctgatg</td><td>agctttaccg</td><td>cagctgcctc</td><td> 5820</td>

<td>gcgcgtttcg</td><td>gtgatgaogg</td><td>tgaaaacctc</td><td>tgacacatgc</td><td>agctcccgga</td><td>gacggtcaca</td><td> 5880</td>

<td>gcttgtctgt</td><td>aagcggtgcc</td><td>gggagcagac</td><td>aagcccgtca</td><td>gggcgcgtca</td><td>gcgggtgttg</td><td> 5940</td>

<td>gcgggtgtcg</td><td>gggcgcagcc</td><td>atgacccagt</td><td>cacgtagcga</td><td>tagcggagtg</td><td>tatactggct</td><td> 6000</td>

taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc 6060 gcacagatgc gtaaggagaa aataccgcat caggcgctct tccgctfccct cgctcactga 6120 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 6180 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 6240 aaaggccagg aaccgtaaaa aggocgcgtt gctggcgttt ttccataggc tccgcccccc 630o tgacgagcat cacaaa&atc gacgctcaag tcagaggtgg cgaaacccga caggactata €360 aagataccag gcgtttcccc ctggaagctc cct-egtgcgc tctcctgttc cgaccctgce 6420 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 6480

119

<td>acgctgtagg</td><td>tatctcagtt</td><td>cggtgtaggt</td><td>cgttcgctcc</td><td>aagctgggct </td><td>gtgtgcacga</td><td> 6540</td>

<td>accccccgtt</td><td>cagcccgacc</td><td>gctgcgcctt</td><td>atccggtaac</td><td>tatcgtettg</td><td>agtccaaccc</td><td> 6600</td>

<td>ggtaagacac</td><td>gacttatcgc</td><td>cactggcagc</td><td>agccactggt</td><td>aacaggatta</td><td>gcagagcgag</td><td> 6660</td>

<td>gtatgtaggc</td><td>ggtgctacag</td><td>agttcttgaa</td><td>gtggtggcct</td><td>aactaçggct</td><td>acactagaag</td><td> 6720</td>

<td>gacagtattt</td><td>ggtatctgcg</td><td>ctctgctgaa</td><td>gccagttacc</td><td>ttcggaaaaa</td><td>gagttggtag</td><td> 6780</td>

<td>ctcttgatcc</td><td>ggcaaacaaa</td><td>ccaccgctgg</td><td>tagcggtggt</td><td>ttttttgttt</td><td>gcaagcagca</td><td> 6840</td>

<td>gattacgcgc</td><td>agaaaaaaag</td><td>gatctcaaga</td><td>agatcctttg</td><td>atcttttcta</td><td>cggggtctga</td><td> 6900</td>

<td>cgctcagtgg</td><td>aacgaaaact</td><td>cacgttaagg</td><td>gattttggtc</td><td>atgagattat</td><td>caaaaaggat</td><td> 6960</td>

<td>cttcacctag</td><td>atccttttaa</td><td>attaaaaatg</td><td>aagttttaaa</td><td>tcaatctaaa</td><td>gtatatatga</td><td> 7020</td>

<td colspan="2">gtaaacttgg. tctgacagtt</td><td>accaatgctt</td><td>aatcagtgag</td><td>gcacetatct</td><td>cagcgatctg</td><td> 7080</td>

<td>tctatttcgt</td><td>tcatccatag</td><td>ttgcctgact</td><td>ccccgtcgtg</td><td>tagataacta</td><td>cgatacggga</td><td> 7140</td>

<td>gggcttacca</td><td>tctggcccca</td><td>gtgctgcaat</td><td>gataccgcga</td><td>gacccacgct</td><td>caccggctcc</td><td> 7200</td>

<td>agatttatca</td><td>gcaataaacc</td><td>agccagccgg</td><td>aagggccgag</td><td>cgcagaagtg</td><td>gtcctgcaac</td><td> 7260</td>

<td>tttatccgcc</td><td>tccatccagt</td><td>ctattaattg</td><td>ttgccgggaa</td><td>gctagagtaa</td><td>gtagttcg-cc</td><td> 7320</td>

<td>agttaatagt</td><td>ttgcgcaacg</td><td>ttgttgccat</td><td>tgctgcaggc</td><td>atcgtggtgt</td><td>cacgctcgtc</td><td> 7380</td>

<td>gtttggtatg</td><td>gcttcattca</td><td>gctccggttc</td><td>ccaacgatca</td><td></td><td>catgatcccc</td><td> 7**0</td>

<td>catgttgtgc</td><td>aaaaaagcgg</td><td>ttagctcctt</td><td>cggtcctccg</td><td>atcgttgtca</td><td>gaagtaagtt</td><td> 7500</td>

<td>ggccgcagtg</td><td>ttatcactca</td><td>tggttatggc</td><td>agcactgcat</td><td>aattctctta</td><td>. ctgtcatgcc</td><td>7S60</td>

atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg 7620 tatgeggcga ccgagttgçt cttgcccggc gtcaacacgg gataataccg cgccacatag 7680 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat 7740 ctta-ccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc 7600 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa 7860 aaagggaafca agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta 7920 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa 7980 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga 8040 aaccattatt atcatgacat fcaacctataa aaaataggcg tatcacgagg ccctttcgtc 8100 ttcaagaatt ctcatgtttg acagcttatc atcgatccac ttgtatattt ggatgaattt 8160 ttgaggaatt ctgaaccagt cctaaaacga gtaaatagga ccggcaattc ttcaagcaat 8220 aaacaggaat accaattatt aaaagataac ttagtcagat cgtacaataa agctttgaag 8280 aaaaatgcgc cttattcaat ctttgcataa aaaaatggcc caaaatctea cattggaaga 8340

120 catttgatga cctcatttct ttcaatgaag ggcctaacgg agttgactaa tgttgtggga 8400 aattggaccg ataagcgtgc ttctgccgtg gccaggacaa cgtatactca tcagataaca 8460 gcaatacctg atcactactt cgcactagtt tctcggtact atgcatatga tccaatatea 8520 aaggaaatga tagcattgaa ggatgagact aatccaattg aggagtggca geatatagaa 8580 cagctaaagg gtagtgctga aggaagcata cgataccccg catggaatgg gataatatea 8640 caggaggtac tagactacct ttcatcctac ataaatagac gcatataagt acgcatttaa 8700 gcataaacac gcactatgcc gttettetea tgtatatata tatacaggca acacgcagat 8760 ataggtgcga cgtgaacagt gagctgtatg tgcgcagctc gcgttgcatt ttcggaagcg 8820 ctcgttttcg gaaacgcttt gaagttccta ttccgaagtt cctattctct agaaagtata 8880 ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa gacgcacttt caaaaaacca 8940

<td>aaaacgcacc</td><td>ggactgtaac</td><td>gagctactaa</td><td>aatattgcga</td><td>ataccgcttc</td><td>cacaaacatt</td><td> 9000</td>

<td>gctcaaaagt</td><td>atctctttgc</td><td>tatatatctc</td><td>tgtgctatat</td><td>ccctatataa</td><td>ccatcccatc</td><td> 9060</td>

<td>cacctttcgc</td><td>tccttgaact</td><td>tgcatctaaa</td><td>ctcgacctct</td><td>acatttttta</td><td>tgtttatctc</td><td> 9120</td>

<td>tagtattace</td><td>tcttagacaa</td><td>aaaaattgta</td><td>gtaagaacta</td><td>ttcatagagt</td><td>taatcgaaaa</td><td> 9180</td>

<td>caatacgaaa</td><td>atgtaaacat</td><td>ttcctatacg</td><td>tagtatatag</td><td>agacaaaata</td><td>gaagaaaccg</td><td> 9240</td>

<td>ttcataattt</td><td>tctgaccaat</td><td>gaagaatcat</td><td>caacgctatc</td><td>aettfcctgfct</td><td>cacaaagt&t</td><td> 5300</td>

<td>gcgcaatcca</td><td>catcggtata</td><td>gaatataatc</td><td>ggggatgcct</td><td>ttatcttgaa</td><td>aaaatgcacc</td><td> 9360</td>

<td>cgcagcttcg</td><td>ctagtaatca</td><td>gtaaacgcgg</td><td>gaagtggagt</td><td>caggcttttt</td><td>ttatggaaga</td><td> 9420</td>

<td>gaaaatagac</td><td>accaaagtag</td><td>ccttcttcta</td><td>accttaacgg</td><td>acctacagtg</td><td>caaaaagtta</td><td> 9480</td>

<td>tcaagagact</td><td>geattataga</td><td>gçgcacaaag</td><td>gagaaaaaaa</td><td>gtaatctaag</td><td>atgctttgtt</td><td> 9540</td>

<td>agaaaaatag</td><td>cgctctcggg</td><td>atgcattttt</td><td>gtagaacaaa</td><td>aaagaagüat</td><td>agattcttgt</td><td> 9600</td>

<td>tggtaaaata</td><td>gcgctctcgc</td><td>gttgcatttc</td><td>tgttctgtaa</td><td>aaatgcagct</td><td>cagattcttt</td><td> 9660</td>

<td>gtttgaaaaa</td><td>ttagcgctct</td><td>cgcgttgcat</td><td>ttttgtttta</td><td>caaaaatgaa</td><td>gcacagattc</td><td> 9720</td>

<td>ttcgttggta</td><td>aaatagcgct</td><td>ttcgcgttgc</td><td>atttctgttc</td><td>tgtaaaaatg</td><td>cagctcagat</td><td> 9780</td>

<td>tctttgtttg</td><td>aaaaattagc</td><td>gctctcgcgt</td><td>tgcatttttg</td><td>ttctacaaaa</td><td>tgaagcacag</td><td> 9840</td>

<td>atgcttcgtt</td><td>aacaaagata</td><td>tgctattgaa</td><td>gtgcaagatg</td><td>gaaacgcaga</td><td>aaatgaaccg</td><td> 9900</td>

<td>gggatgcgac</td><td>gtgcaagatt</td><td>acctatgcaa</td><td>tagatgcaat</td><td>agtttctoca</td><td>ggaaccgaaa</td><td> 9960</td>

<td>tacatacatt</td><td>gtcttocgta</td><td>aagcgctaga</td><td>ctatatatta</td><td>ttatacaggt</td><td>tcaaatatac</td><td> 10020</td>

<td>tatctgtttc</td><td>agggaaaact</td><td>cccaggttcg</td><td>gatgttcaaa</td><td>attcaatgat</td><td>gggtaacaag</td><td> 10080</td>

<td>tacgategta</td><td>aatctgtaáa</td><td>acagtttgtc</td><td>ggatattagg</td><td>ctgtatctcc</td><td>tcaaagcgta</td><td> 10140</td>

121

<td>ttcgaatatc</td><td>attgagaagc</td><td>tgcatttttt</td><td>tttttttttt</td><td>tttttttttt ttttttatat</td><td> 10200</td>

<td>atatttcaag</td><td>gatataccat</td><td>tgtaatgtct</td><td>gcccctaaga</td><td>agatcgtcgt tttgccaggt</td><td> 10260</td>

<td>gaccacgttg</td><td>gtcaagaaat</td><td>cacagccgaa</td><td>gccattaagg</td><td>ttcttaaagc tatttctgat</td><td> 10320</td>

<td>gttcgttcca</td><td>atgtcaagtt</td><td>cgatttcgaa</td><td>aatcatttaa</td><td>ttggtggtgc tgctatcgat</td><td> 10380</td>

<td>gctacaggtg</td><td>tcccacttcc</td><td>agatgaggcg</td><td>ctggaagcct</td><td>ccaagaaggt tgatgccgtt</td><td> 10440</td>

<td>ttgttaggtg</td><td>ctgtgggtgg</td><td>tcctaaatgg</td><td>ggtaccggta</td><td>gtgttagacc tgaacaaggt</td><td> 10500</td>

<td>ttactaaaaa</td><td>tccgtaaaga</td><td>acttcaattg</td><td>tacgccaact</td><td>taagaccatg taactttgca</td><td> 10560</td>

<td>tccgactctc</td><td>ttttagactt</td><td>atctccaatc</td><td>aagccacaat</td><td>ttgctaaagg tactgacttc</td><td> 10620</td>

<td>gttgttgtca</td><td>gagaattagt</td><td>gggaggtatt</td><td>tactttggta</td><td>agagaaagga agacgatggt</td><td> 10680</td>

<td>gatggtgtcg</td><td>cttgggatag</td><td>tgaacaatao</td><td>accgttccag</td><td>aagtgcaaag aatcacaaga</td><td> 10740</td>

<td>atggccgctt</td><td>tcatggccct</td><td>acaacatgag</td><td>ccaccattgc</td><td>ctatttggtc cttggataaa</td><td> 10800</td>

<td>gctaatgttt</td><td>tggcctcttc</td><td>aagattatgg</td><td>agaaaaactg</td><td>tggaggaaac catcaagaac</td><td> 10860</td>

<td>gaattcccta</td><td>cattgaaggt</td><td>tcaacatcaa</td><td>ttgattgatt</td><td>ctgccgccat gatcctagtt</td><td> 10920</td>

<td>aagaacccaa</td><td>cccacCtaaa</td><td>tggtattata</td><td>atcaccagca</td><td>acatgtttgg tgatatcatc</td><td> 10980</td>

<td>tccgatgaag</td><td>cctccgttat</td><td>cccaggttcc</td><td>ttgggtttgt</td><td>tgccatctgc gtccttggcc</td><td> 11040</td>

<td>tctttgccag</td><td>acaagaacac</td><td>cgcatttggt</td><td>ttgtacgaac</td><td>catgccacgg ttetgctcca</td><td> 11100</td>

<td>gatttgccaa</td><td>agaataaggt</td><td>tgaccctatc</td><td>gccactatct</td><td>tgtctgctgc aatgatgttg</td><td> 11160</td>

<td>aaattgtcat</td><td>tgaacttgcc</td><td>tgaagaaggt</td><td>aaggccattg</td><td>aagatgcagt taaaaaggtt</td><td> 11220</td>

<td>ttggatgcag</td><td>gtatcagaac</td><td>tggtgattta</td><td>ggtggttcca</td><td>acagtaccac cgaagtcggt</td><td> 11280</td>

<td>gatgctgtcg</td><td>ccgaagaagt</td><td>taagaaaatc</td><td>cttgcttaaa</td><td>aagattctet ttttttatga</td><td> 11340</td>

<td>tatttgtaca</td><td>aaaaaaaaaa</td><td>aaaaaaaaaa</td><td>aaaaaaaaaa</td><td>aaaaaaaaaa aaaaatgcag</td><td> 11400</td>

<td>cgtcacatcg</td><td>gataataatg</td><td>atggcagcca</td><td colspan="2">ttgtagaagt gccttttgca tttctagtct</td><td> 11460</td>

<td>ctttctcggt</td><td>ctagctagtt</td><td>ttactacatc</td><td>gcgaagatag</td><td>< aatcttagat cacactgcct</td><td> 11520</td>

ttgctgagct ggatcatatg agtaacaaaa gagtggtaag gcctcgttaa. aggacaagga 11580 cctgagcgga agtgtategt aaagtagacg gagtatacta gtatagtcta tagtccgtgg 11640 aattctaagt gceagcttta taatgtcatt ctccttacta cagacccgoc tgaaagtaga 11700 cacatcatça tcagtaagct ttgacaaaaa gcattgagta gctaactctt ctatgcaatc 11760 tatagctgtt ttataaggca ttcaatggac agattgaggt ttttgaaaca tactagtgaa 11820 attagectta atcecttctc gaagttaatc atgcattatg gtgtaaaaaa tgcaactcgc 11880 gttgctctac tttttcccga atttccaaat acgcagctgg ggtgattgct cgatttcgta 11940 acgaaagttt tgtttataaa aaccgcgaaa accttctgta acagatagat ttttacagcg 12000

122

<td>ctgatataca</td><td>atgacatcag</td><td colspan="2">ctgtaatgga aaataactga</td><td>aatatgaatg</td><td>gcgagagact</td><td> 12060</td>

<td>gcttgcttgt</td><td>attaagcaat</td><td colspan="2">gtattatgca gcacttccaa</td><td>cctatggtgt</td><td>acgatgaaag</td><td> 12120</td>

<td>taggtgtgta</td><td>atcgagacga</td><td>caagggggac</td><td>ttttccagtt</td><td>cctgatcatt</td><td>ataagaaata</td><td> 12180</td>

<td>caaaacgtta</td><td>gcatttgcat</td><td>ttgttggaca</td><td>tgtactgaat</td><td>acagacgaca</td><td>caecggtaat</td><td> 12240</td>

<td>tgaaaaagaa</td><td>ctggattggc</td><td>ctgatcctgc</td><td>actagtgtac</td><td>aatacaattg</td><td>tcgatcgaat</td><td> 12300</td>

<td>cataaatcac</td><td>ccagaattat</td><td>cacagtttat</td><td>atcggttgca</td><td>tttattagtc</td><td>agttaaaggc</td><td> 12360</td>

<td>caccatcgga</td><td>gagggtttag</td><td>atattaatgt</td><td>aaaaggcacg</td><td>ctaaaccgca</td><td>ggggaaaggg</td><td> 12420</td>

<td>tatcagaagg</td><td>cctaaaggcg</td><td>tattttttag</td><td>atacatggaa</td><td>tctccatttg</td><td>tcaatacaaa</td><td> 12480</td>

<td>ggtcactgca</td><td>ttcttctctt</td><td>atctfccgaga</td><td>ttataataaa</td><td>attgcctcag</td><td>aatatcacaa</td><td> 12540</td>

<td>taatactaaa</td><td>ttcattctca</td><td>cgttttcatg</td><td>tcaagcatat</td><td>tgggcatctg</td><td>gcccaaactt</td><td> 12600</td>

<td>ctccgccttg</td><td>aagaatgtta</td><td>tttggtgete</td><td>cataattcat</td><td>gaatacattt</td><td>ctaagtttgt</td><td> 12660</td>

<td>ggaaagagaa</td><td>caggataaag</td><td>gtcatatagg</td><td>agatcaggag</td><td>ctaccgcctg</td><td>aagaggaccc</td><td> 12720</td>

<td>ttctcgtgaa</td><td>ctaaacaatg</td><td>tacaacatga</td><td>agtcaatagt</td><td>ttaacggaac</td><td>aagatgcgga</td><td> 12780</td>

<td>ggcggatgaa</td><td>ggattgtggg</td><td>gtgaaataga</td><td>ttcattatgt</td><td>gaaaaatggc</td><td>agtctgaagc</td><td>12B40</td>

<td>ggagagtcaa</td><td>actgaggcgg</td><td>agataatagc</td><td>cgacaggata</td><td>attggaaata</td><td>gccagaggat</td><td> 12900</td>

ggcgaacctc aaaattcgtc gtacaaagtt caaaagtgtc ttgtatcata tactaaagga 12960 actaattcaa tctcagggaa ccgtaaaggt ttatcgcggt agtagttttt cacacgattc 13020 gataaagata agcttacatt atgaagagca gcatattaca gccgtatggg tctacttgat 13080 agtaaaattt gaagagcatt ggaagcctgt tgatgtagag gtcgagttta gatgcaagtt 13140 caaggagcga aaggtggatg ggtaggttat atagggatat agcacagaga tatatagcaa 13200 agagatactt ttgaggcaat gtttgtggaa gcggtattcg caatatttta gtagctcgtt 13260 acagtccggt gcgtttttgg ttttttgaaa gtgcgtcttc agagcgcttt tggttttcaa 13320 aagcgctctg aagttcctat actttctaga gaataggaac ttcggaatag gaacttcaaa 13380 gcgtttcoga aaacgagcgc ttccgaaaat gcaacgcgag ctgcgcacat acagctcact 13440 gttcacgtcg cacctatatc tgcgtgttgc ctgtatatat atatacatga gaagaacggc 13500 atagtgcgtg tttatgctta aatgcgtact tatatgcgtc tatttatgta ggatgaaagg 13560 tagtctagta cçtcctgtga tattatocca ttccatgcgg ggtatcgtat gcttccttca 13620 gcactaccct ttagctgttc fcatatgctgc cactcctcaa ttggattagt ctcatccttc 13680 aatgcattca tttcctttga tattggatca taccctagaa gtattacgtg attttctgcc 13740 ccttaccctc gttgctaçtc tccttttttt cgtgggaacc gctttagggc cctcagtgat 13800

123 ggtgttttgt aatttatatg etcctcfctgc atttgtgtct ctacttcttg ttcgcctgga 13860 gggaacttct tcatttgtat tagcatggtt cacttcagtc cttccttcca actcactctt 13920 tttttgctgt aaacgattct ctgccgccag ttcattgaaa ctattgaata tatcctttag 13980 agattccggg atgaataaat cacctattaa agcagcttga cgatctggtg gaactaaagt 14040 aagcaattgg gtaacgacgc ttacgagctt cataacatct tcttccgttg gagctggtgg 14100 gactaa.taac tgtgtacaat ccatttttct catgagcatt tcggtagctc tcttcttgtc 14160 tttctcgggc aatcttccta ttattatagc aatagatttg tatagttgct ttctattgtc 14220 taacagcttg ttattctgta gcatcaaatc tatggcagcc tgacttgctt cttgtgaaga 14280

<td>gagcatacca tttocaatcg aagatacgct ggaatcttct gcgctagaat</td><td>caagaccata</td><td> 14340</td>

<td>cggcctaccg gttgtgagag attccatggg ccttatgaca tatcctggaa</td><td>agagtag-ctc</td><td> 14400</td>

<td>atcagactta cgtttactct ctatatcaat atctacatea agagcaatca</td><td>tttcaataaa</td><td> 14460</td>

<td>cagccgacat acatcccaga cgctataagc tgtacgtgct tttaccgtca</td><td>gattcttggc</td><td>14S20</td>

<td>tgtttcaatg tcgtecattt tggttttctt ttaccagtat tgttcgtttg</td><td>ataatgtatt</td><td>14S80</td>

<td>cttgctfcafct acattataaa atctgtgcag atcacatgtc aaaacaactt</td><td>tttatcacaa</td><td> 14640</td>

<td>gatagtaccg caaaacgaac ctgcgggccg tctaaaaatt aaggaaaagc</td><td>agcaaaggtg</td><td> 14700</td>

<td>catttttaaa atatgaaatg aagataccgc agtaccaatt attttegcag</td><td>tacaaataat</td><td> 14760</td>

<td>gcgcggccgg tgcatttttc gaaagaacgc gagacaaaca ggacaattaa</td><td>agttagtttt</td><td> 14820</td>

<td>tcgagttagc gtgtttgaat actgcaagat acaagataaa tagagtagtt</td><td>gaaactagat</td><td> 14880</td>

<td>atcaattgca cacaagatcg gcgcüaagca tgccacaatt tggtatatta</td><td>tgtaaaacac</td><td> 14940</td>

<td>cacctaaggt gcttgttcgt cagtttgtgg aaagg-tttga aagaccttca</td><td>ggtgagaaaa</td><td> 15000</td>

<td>tagcattatg tgctgctgaa ctaacctatt tatgttggat gattacacat</td><td>aacggaacag</td><td>1S060</td>

<td>caatcaagag agccacattc atgagctata atactatcat aagcaattcg</td><td>ctgagtttcg</td><td> 15120</td>

<td>atattgtcaa taaatcactc cagtttaaat acaagacgca aaaagcaaca</td><td>attctggaag</td><td> 15180</td>

<td>cctcattaaa gaaattgatt cctgcttggg aatttacaat tattccttac</td><td>tatggacaaa</td><td> 15240</td>

<td>aacatcaatc tgatatcact gatattgtaa gtagtttgca attacagttc</td><td>gaatcatcgg</td><td> 15300</td>

<td>aagaagcaga taagggaaat agccacagta aaaaaatgct aaagcacttc</td><td>taagtgaggg</td><td>1S360</td>

<td>tgaaagcatc tgggagatca ctgagaaaat actaaattcg tttgagtata</td><td>cttcgagatt</td><td> 15420</td>

<td>tacaaaaaca aaaactttat accaattcct cttcctagct actttcatca</td><td>attgtggaag</td><td> 15480</td>

<td>attcagcgat attaagaacg ttgatccgaa atcatttaaa ttagtccaaa</td><td>ataagtatct</td><td> 15540</td>

<td>gggagtaata atccagtgtt tagtgacaga gacaaagaca agcgttagta</td><td>ggcacatata</td><td> 15600</td>

cttctttagc gcaaggggta g

15621

124 <210> 51 <211> 3593 <212> DNA <213> vetor pPICZalphA <400> 51 agatctaaca tccaaagacg aaaggttgaa tgaaaccttt ttgccatccg acatccacag 60 gtccattcte acaeataagt gccaaacgca acaggagggg atacactagc agcagaccgt 120 tgcaaacgca ggacctccac tectcttctc ctcaacaccc acttttgcca tcgaaaaacc 180 agcccagtta ttgggcttga ttggagctcg ctcattccaa ttccttctat taggctacta 240 acaccatgac tttattagcc tgtctatcct ggcccccctg gegaggttca tgtttgttta 300 tttccgaatg caacaagctc cgcattacac ccgaacatca ctccagatga gggctttctg 360 agtgtggggt caaatagttt catgttcccc aaatggccca aaactgacag tttaaacgct 420 gtcttggaac ctaatatgac aaaagcgtga tctcatccaa gatgaactaa gtttggttcg 480 ttgaaatgct aacggccagt tggtcaaaaa gaaacttcca aaagtcggca taccgtttgt 540 cttgtttggt attgattgac gaatgcbcaa aaataatctc attaatgctt agcgcagtct €00 ctctatcgct tctgaacccc ggtgcacctg tgccgaaacg caaatgggga aacacccgct 660 ttttggãtga ttatgcattg tctccacatt gtatgcttcc aagattctgg tgggaatact 720 gctgatagcc taacqttcat gatcaaaatt taactgttct aacccctact tgacagcaat 780 atataaacag aaggaagctg occtgtctta aacctttttt tttatcatca ttattagctt 840 actttcataa ttgcgactgg ttccaattga caagcttttg attttaacga cttttaacga 900 caacttgaga agatcaaaaa acaactaatt attcgaaacg atgagatttc ctfccaatttt 960 tactgctgtt ttattcgcag catccfcccgc attagctgct ccagtcaaca ctacaacaga 1020 agatgaa&cg gcacaaattc cggctgaagc tgtcatcggt tactcagatt tagaagggga 1080 tttcgatgtt gctgttttgc catttt-ccaa cagcacaaat aacgggttat tgtttataaa 1140 tactactatt gccagcattg ctgctaaaga agaaggggta tctctcgaga aaagagaggc 1200 tgaagctgaa ttcacgtggc ccagccggcc gtctcggatc ggtacctcga gccgcggcgg 1260 ccgccagctt tctagaacaa aaactcatct cagaagagga tctgaatagc gccgfccgacc 1320 atcatcatca tcatcattga gtttgtagcc ttagacatga ctgttcctca gttcaagttg 1380 ggcacttacg agaagaccgg tcttgctaga ttctaatcaa gaggatgtca gaatgccatt 1440 tgcctgagag atgcaggctt catttttgat acttttttat ttgtaaccta tatagtatag 1500 gatttttttt gtcattttgt ttcttctcgt acgagcttgc tcctgatcag cctatctcgc 1560 agctgatgaa tatcttgtgg taggggtttg ggaaaatcat tcgagtttga tgtttttctt 1620

125 ggtatttccc actcctcttc agagtacaga çccccacaca ccatagcttc aaaatgtttc ggactccgcg catcgccgta ccacttcaaa tctttcttcc tctagggtgt çgttaattac gaccgcctcg tttctttttc ttcgtcgaaa tttcttgaaa tttttttfctt tagttttttt ttaataaacg gtcttcaatt tctcaagttt ttttttactt cttgttcatt agaaagaaag acaattaatc ateggcatag tatatcggca ccatggccaa gttgaocagt gccgttccgg tcgagttctg gaccgaccgg ctcgggttct gtgtggtccg ggacgacgtg accctgttca acaacaccct ggcctgggtg tgggtgcgcg aggtcgtgtc cacgaacttc cgggacgcct agccgtgggg gcgggagttc gccctgcgcg ccgaggagca ggactgacac gtccgacggc tccocctttt cctttgtcga tatcatgtaa ccccccacat ccgctctaac cgaaaaggaa atttattttt ttatagttat gttagtatta ttttttctgt acagacgcgt gtacgcatgt gttttgggac gctcgaaggc tttaatttgc cagcaaaagg ccaggaacog taaaaaggcc ccccctgacg agcatcacaa aaatcgacgc ctataaagat accaggcgtt tccccctgga ctgocgctta ccggatacct gtccgccfctt tgctcacgct gtaggtatct cagttcggtg cacgaacccc ccgttcagcc cgaccgctgc aacccggtaa gacacgactt atcgccactg gcgaggtatg taggcggtgc tacagagttc agaaggacag tatttggtat ctgcgctetg agattaagtg agaccttcgt ttgtgcggat tactcctttt ttactcttcc agattttctc acacccaagc acagcatact aaáttttccc ccgtactaaa ggtttggaaa agaaaaaaga aaggcaataa aaatttttat cacgtttctt ctctttcagt gacctccatt gatatttaag cagtttcatt tttcttgttc tattacaact catagcaatc taatctaagg ggcggtgttg tagtataata cgacaaggtg aggaactaaa tgctcaccgc gcgcgacgtc gceggagcgg cccgggactt cgtggaggac gacttcgecg tcagcgcggt ccaggaccag gtggtgccgg gcctggacga gctgtacgcc gagtggtcgg ccgggccggc catgaccgag atcggcgagc acccggccgg caactgcgtg cacttcgtgg ggcccacggg tcccaggcct cggagatceg ttagttatgt cacgcttaca ttcacgccct ggagttagac aacctgaagt ctaggtccct agaacgttat ttatatttca aatttttctt aacattatac tgaaaacctt gcttgagaag aagctggaga ccaacatgtg agcaaaaggc gcgttgctgg cgtttttaca taggctccgc tcaagtcaga ggtggcgaaa cccgacagga agctccetcg tgcgctctcc tgttccgacc ctcccttcgg gaagcgtggc gctttctcaa taggtcgttc gctccaagct gggctgtgtg gccttatccg gtaactatcg tcttgagtcc gcagcagcca -ctggtaacag gattagcaga ttgaagtggt ggcctaacfca oggctacact cbgaagocag ttaccttcgg aaaaagagtt

1680

1740

1800

1860

1920

1980

2040

2100

2160

2220

2280

2340

2400

2460

2520

2580

2640

2700

2760

2820

2880

2940

3000

3060

3120

3180

3240

3300

3360

3420

126 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag 3480 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg 3540 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag ate 3593 <210> 52 <211> 3547 <212> DMA <213> vetor pPXCZalphaD' <400> 52

<td>agatctaaca</td><td>tccaaagacg</td><td>aaaggttgaa</td><td>tgaaaccttt</td><td>ttgccatccg ,</td><td>acatccacag</td><td> €0</td>

<td>gteeattctc</td><td>acacataagt</td><td>gccaaacgca</td><td>acaggagggg</td><td>atacactagc</td><td>agcagaccgt</td><td> 120</td>

<td>tgcaaacgca</td><td>ggacctccac</td><td>tcctcttctc</td><td>ctcaacaccc</td><td>acttttgcca</td><td>tcgaaaaacc</td><td> 180</td>

<td>agcccagtta</td><td>ttgggcttga</td><td>ttggagctcg</td><td>ctcattccaa</td><td>ttccttctat</td><td>taggctacta</td><td> 240</td>

<td>acaccatgac</td><td>tttattagcc</td><td>tgtctatcet</td><td>ggcccccctg</td><td>gcgaggttea</td><td>tgtttgttta</td><td> 300</td>

<td>tttccgaatg</td><td>caacaagctc</td><td>cgcattacac</td><td>ccgaacatca</td><td>ctccagatga</td><td>gggctttctg</td><td> 360</td>

<td>agtgtggggt</td><td>caaatagttt</td><td>catgttcccc</td><td>aaatggccca</td><td>aaactgacag</td><td>tttaaacgct</td><td> 420</td>

<td>gtcttggaac</td><td>ctaatatgac</td><td>aaaagcgtga</td><td>tctcatceaa</td><td>gatgaactaa</td><td>gtttggttcg</td><td> 480</td>

<td>ttgaaatgct</td><td>aacggccagt</td><td>tggtcaaaaa</td><td>gaaacttcca</td><td>aaagtcggca</td><td>taccgtttgt</td><td> 540</td>

<td>cttgtttggt</td><td>attgafcfcgac</td><td>gaatgctcaa</td><td>aaataatctc</td><td></td><td>agcgcagtct</td><td> €00</td>

<td>ctctatcgct</td><td>tctgaacccc</td><td>ggtgcacctg</td><td>tgccgaaacg</td><td>caaatgggga</td><td>aacacccgct</td><td> €60</td>

<td>ttttggatga</td><td>ttatgcattg</td><td>tctccacatt</td><td>gtatgcttcc</td><td>aagattctgg</td><td>tgggaatact</td><td> 720</td>

<td>gctgatagcc</td><td>taacgttcat</td><td>gatcaaaatt</td><td>taactgttct</td><td>aacccctact</td><td>tgacagcaat</td><td> 780</td>

<td>atataaacag</td><td>aaggaagctg</td><td>ccctgtctta</td><td>aacctttttt</td><td>tttatcatca</td><td>ttattagctt</td><td> 840</td>

<td>actttcataa</td><td>ttgcgactgg</td><td>ttccaattga</td><td>caagcttttg</td><td>attttaacga</td><td>cttttaacga</td><td> 900</td>

<td>caacttgaga</td><td>agatcaaaaa</td><td>acaactaatt</td><td>attcgaaacg</td><td>atgagatttc</td><td>ettcaatttt</td><td> 960</td>

<td>tactgctgtt</td><td>ttattcgcag</td><td>catcctccgc</td><td>attagctgct</td><td>ccagtcaaca</td><td>ctacaacaga</td><td> 1020</td>

<td>agatgaaacg</td><td>gcacaaattc</td><td>cggctgaagc</td><td>tgtcatcggt</td><td>tactcagatt</td><td>tagaagggga</td><td> 1080</td>

<td>tttcgatgtt</td><td>gctgttttgc</td><td>cattttccaa</td><td>cagcacaaat</td><td>aacgggttat</td><td>tgtttataaa</td><td> 1140</td>

tactactatt gccagcattg ctgctaaaga agaaggggta tetetegaga aaaggggccc 1200 gaattcgcat gcggccgcca gctttctaga acaaaaactc atctcagaag aggatctgaa 1260 tagcgccgtc gaccatcatc ateateatea ttgagtttgt agccttagac atgactgttc 1320 etcagttcaa gttgggcact taegagaaga ccggtcttgc tagattetaa tcaagaggat 1380 gtcagaatgc catttgcctg agagatgcag gcttcatfctt tgatactttt ttatttgtaa 1440

127

<td>cctatatagt</td><td>ataggatttt</td><td>ttttgtcatt</td><td>ttgtttcttc</td><td>tcgtacgagc</td><td>ttgctcctga</td><td> 1500</td>

<td>tcagcctatc</td><td>tcgcagctga</td><td>tgaatatctt</td><td>gtggtagggg</td><td>tttggaaaaa</td><td>tcattcgagt</td><td> 1560</td>

<td>ttgatgtttt</td><td>tcttggtatt</td><td>tcocactcct</td><td>cttcagagta</td><td>cagaagatta</td><td>agtgagacct</td><td> 1620</td>

<td>tcgtttgtgc</td><td>ggatccccca</td><td>cacaccatag</td><td>cttcaaaatg</td><td>tttctactcc</td><td>ttttttactc</td><td> 1680</td>

<td>ttccagattt</td><td>tctcggactc</td><td>cgcgcatcgc</td><td>cgtaccactt</td><td>caaaacaccc</td><td>aagcacagca</td><td> 1740</td>

<td>tactaaattt</td><td>tccctctttc</td><td>ttcctctagg</td><td>gtgtcgttaa</td><td>ttacccgtac</td><td>taaaggtttg</td><td> 1800</td>

<td>gaaaagaaaa</td><td>aagagaccgc</td><td>ctcgtttctt</td><td>tttcttcgtc</td><td>gaaaaaggca</td><td>ataaaaattt</td><td> 1860</td>

<td>ttatcacgtt</td><td>tctttttctt</td><td>gaaatttttt</td><td>tttttagttt</td><td>ttttctcttt</td><td>cagtgacctc</td><td> 1920</td>

<td>cattgatatt</td><td>taagttaata</td><td>aacggtcttc</td><td>aatttctcaa</td><td>gtttcagttt</td><td>catttttctt</td><td> 1980</td>

<td>gttctattac</td><td>aact.tttt.tt</td><td>acttcttgtt</td><td>cattagaaag</td><td>aaagcatagc</td><td>aatctaatct</td><td> 2040</td>

<td>aaggggcggt</td><td>gttgacaatt</td><td>aatcatcggc</td><td>atagtatatc</td><td>ggcatagtat</td><td>aatacgacaa</td><td> 2100</td>

<td>ggtgaggaac</td><td>taaaccatgg</td><td>ccaagttgac</td><td>cagtgccgtt</td><td>ccggtgctca</td><td>ccgcgcgcga</td><td> 2160</td>

<td>cgtcgccgga</td><td>gcggtcgagt</td><td>tctggaccga</td><td>ccggctcggg</td><td>ttctcccggg</td><td>acttcgtgga</td><td> 2220</td>

<td>ggacgacttc</td><td>gccggtgtgg</td><td colspan="2">tccgggacga cgtgaccctg</td><td>ttcatcagcg</td><td>cggtccagga</td><td> 2280</td>

<td>ccaggtggtg</td><td>ccggacaaca</td><td>ccctggcctg</td><td>ggtgtgggtg</td><td>cgcggcctgg</td><td>acgagctgta</td><td> 2340</td>

cgccgagtgg tcggaggtcg tgtccacgaa cttccgggac gcctcegggc cggccatgac 2400 cgagatcggc gagcagccgt gggggcggga gttcgccctg cgcgacccgg ccggcaactg 2460 cgtgcacttc gtggccgagg agcaggactg acacgtccga cggcggccca cgggtcccag 2520 gcctcggaga tccgtccccc ttttcctttg tcgatatcat gtaattagtt atgtcacgct 2580 tacattcacg ccctccccce acatccgctc taaccgaaaa. ggaaggagtt agacaacctg 2640 aagtctaggt ccctatttat ttttttatag ttatgttagt attaagaacg ttatttatat 2700 ttcaaatttt tctttttttt ctgtacaga-c gcgtgtacgc atgtaacatt atactgaaaa 2760 ccttgcttga gaaggttttg ggacgctcga aggctttaat ttgcaagctg gagaccaaca 2820 tgtgagcaaa aggccagcaa aaggccagga aocgtaaaaa ggccgcgttg ctggcgtttt 2880 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc 2940 ga&acccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct 3000 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg 3060 tggcgctttc tcaatgctca cgctgrtaggt atct-cagttc ggtgtaggtc gttcgctoca 3120 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact 3180 atcgtcttga gtccaacccg gtaagacacg acttategcc actggcagca gccactggta 3240 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta 3300

128 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct 3360 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtagtt 3420 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga 3480 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca 3540 tgagatc 3547 <210> 53 <211> 3558 <212> DNA <213> vetor pPXCAalphaE' <400> 53

<td>agatctaaca tccaaagacg aaaggttgaa</td><td>tgaaaocttt ttgccatccg acatccacag</td><td> €0</td>

<td>gtccattctc acacataagt gccaaacgca</td><td>acaggagggg atacactagc agcagaccgt</td><td> 120</td>

<td>tgcaaaegca ggacctccac tcctcttctc</td><td>ctcaacaccc acttttgcca tcgaaaaacc</td><td> 180</td>

<td>agcccagtta ttgggcttga ttggagcteg</td><td>ctcatteeaâ ttcettctat taggctacta</td><td> 240</td>

<td>acaccatgac tttattagcc tgtctatcct</td><td>ggcccccctg gcgaggtfcca tgtttgttta</td><td> 300</td>

<td>tttccgaatg caacaagctc cgcattacac</td><td>ccgaacatca ctccagatga gggctttctg</td><td> 360</td>

<td>agtgtggggt caaatagttt catgttcccc</td><td>aaatggccca aaactgacag tttaaacgct</td><td> 420</td>

<td>gtcttggaac ctaatatgac aaaagcgtga</td><td>tcfccatccaa gatgaactaa gtttggttcg</td><td> 480</td>

<td>ttgaaatgct aacggccagt tggtcaaaaa</td><td>gaaacttcca aaagtcggca taccgtttgt</td><td> 540</td>

<td>cttgtttggt attgattgac gaatgctcaa</td><td>aaataatctc attaatgctt agcgcagtct</td><td> 600</td>

<td>ctctatcgct tctgaacccc ggtgcacctg</td><td>tgcogaaacg caaatgggga aacacccgct</td><td> €60</td>

<td>ttttggatga ttatgcattg tctccacatt</td><td>gtatgcttcc aagattctgg tgggaatact</td><td> 720</td>

<td>gctgatagcc taacgttcat gatcaaaatt</td><td>taactgttct a&cccctact tgacagcaat</td><td> 780</td>

<td>atataaacag aaggaagctg ccctgfccfcta</td><td>aacctttttt tttatcatca ttattagctt</td><td> 840</td>

<td>actttcataa ttgogactgg ttccaattga</td><td>caagcttttg attttaacga cttttaacga</td><td> 900</td>

<td>caacttgaga agatcaaaaa acaactaatt</td><td>attcgaaacg atgagatttc cttcaatttt</td><td> 960</td>

<td>tactgctgtt ttattcgcag catcctccgc</td><td>attagctgct ccagtcaaca ctacaacaga</td><td> 1020</td>

<td>agatgaaacg gcacaaattc cggctgaagc</td><td>fc-gfccabcggt tactcagatt tagaagggga</td><td> 1080</td>

<td>tttcgatgtt gctgttttgc cattttocaa</td><td>cagcacaaat aacgggttat tgtttataaa</td><td> 1140</td>

<td>tactactatt gccagcattg ctgctaaaga</td><td>agaaggggta tctctcgaga aaagagaggc</td><td> 1200</td>

<td>tgaagcctgc agcatatgct cgaggoogcc</td><td>agctttctag aacaaaaact catctcagaa</td><td> 1260</td>

<td>gaggatctga atagcgcogt cgaccatcat</td><td>catcatcatc attgagtfctg tagccttaga</td><td> 1320</td>

129 catgactgtt cctcagttca agttgggcac ttacgagaag accggfccttg ctagattcta 1380 atcaagagga tgtcagaatg ccatttgcct gagagatgca ggcttcattt ttgatacttt 1440 tttatttgta acctatatag tataggattt tttttgtcat tttgtttctt ctcgtacgag isoo cttgctcctg atcagcctat ctcgcagctg atgaatatct tgtggtaggg gtttgggaaa 1S60 atcattcgag tttgatgttt ttcttggtat ttcccactcc tcttcagagt acagaagatt 1620 aagtgagacc ttcgtttgtg cggatceccc acacaccata gcttcaaaat gtttctactc 1680 cttttttact cttccagatt ttctcggact ccgcgcatcg ccgtaccact tcaaaacacc 1740 caagcacagc atactaaatt ttccctcttt cttcctctag ggtgtcgtta attacccgta 1800 ctaaaggttt ggaaaagaaa aaagagaccg cctcgtttct ttttcttcgt cgaaaaaggc 1860 aataaaaatt tttatcacgt ttctttttct tgaaattttt ttttttagtt tttttctctt 1920 tcagtgacct ccattgatat ttaagttaat aaacggtctt caatttctca agtttcagtt 1980 tcatttttct tgttctatta caactttttt tacttcttgt tcattagaaa gaaagcatag 2040

<td>caatctaatc</td><td>taaggggcgg</td><td>tgttgacaat</td><td>taatcatcgg</td><td>catagtatat</td><td>cggcatagta</td><td> 2100</td>

<td>taatacgaca</td><td>aggtgaggaa</td><td>ctaaaccatg</td><td>gccaagttga</td><td>ccagtgccgt</td><td>tccggtgctc</td><td> 2160</td>

<td>accgcgcgcg</td><td>acgtcgccgg</td><td>agcggtcgag</td><td>tfcctggaccg</td><td>accggctcgg</td><td>gttctcccgg</td><td> 2220</td>

<td>gacttcgtgg</td><td>aggacgactt</td><td>cgccggtgtg</td><td>gtccgggacg</td><td>acgtgaccct</td><td>gtteateage</td><td> 2280</td>

<td>gcggtccagg</td><td>accaggtggt</td><td>gccggacaac</td><td>accctggcct</td><td>gggtgtgggt</td><td>gcgcggcctg</td><td> 2340</td>

<td>gacgagctgt</td><td>acgccgagtg</td><td>gtcggaggtc</td><td>gtgtccacga</td><td colspan="2">acttccggga -cgoctccggg</td><td> 2400</td>

<td>ccggccatga</td><td>ccgagatcgg</td><td>cgagcagccg</td><td>tgggggcggg</td><td>agttcgccct</td><td>gcgogacecg</td><td> 2460</td>

<td>gocggcaaet</td><td>gcgtgcactt</td><td>cgtggccgag</td><td>gagcaggact</td><td>gacacgtocg</td><td>acggcggccc</td><td> 2520</td>

<td colspan="2">acgggtccca ggcctcggag</td><td>atccgtcccc</td><td>cttttccttt</td><td>gtcgatatea</td><td>tgtaattagt</td><td> 2580</td>

<td>tatgtcacgc</td><td>ttacattcac</td><td>gcoctccocc</td><td>cacatccgct</td><td>ctaaccgaaa</td><td>aggaaggagt</td><td> 2640</td>

<td colspan="2">tagacaacct gaagtctagg</td><td>tccctattta</td><td>tttttttata</td><td>gttatgttag</td><td>tattaagaac</td><td> 2700</td>

<td>gttatttata</td><td>tttcaaattt</td><td>ttcttttttt</td><td>tctgtacaga</td><td colspan="2">cgcgtgtacg catgtaacat</td><td> 2760</td>

<td>tatactgaaa</td><td>accttgcttg</td><td>agaaggtttt</td><td>gggacgctcg</td><td>aaggctttaa</td><td>tttgcaagct</td><td> 2820</td>

<td>ggagaccaac</td><td>atgtgagcaa</td><td>aaggccagca</td><td>aaaggccagg</td><td>aaecgtaaaa</td><td>aggccgcgtt</td><td> 2880</td>

<td>gctggcgttt</td><td>ttccataggc</td><td>tccgcccocc</td><td>tgacgagcat</td><td>cacaaaaatc</td><td>gacgctcaag</td><td> 2940</td>

<td>tcagaggtgg</td><td>cgaaacccga</td><td>caggactata</td><td>aagataocag</td><td>gcgtttcccc</td><td>ctggaagctc</td><td> 3000</td>

<td>cctcgtgcgc</td><td>tctcctgttc</td><td>cgacectgcc</td><td>gcttaccgga</td><td>tacctgtocg</td><td>cctttetcce</td><td> 3060</td>

<td>ttcgggaagc</td><td>gtggcgcttt</td><td>ctcaatgctc</td><td>acgetgtagg</td><td>tatctcagtt</td><td>cggtgtaggt</td><td> 3120</td>

130 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 3180 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 3240 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 3300 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa 3360 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 3420 tagcggtggt ttttttgttt gcaagcagca gattacgogc agaaaaaaag gatctcaaga 3480 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 3540 gattttggtc atgagatc 3558 <210> 54 <211> 28 <212> DNA <213> iniciador ou sonda sintética <400> 54 tcgagaaaag gggcccgaat tcgcatgc 28 <210> 55 <211> 28 <212> DMA <213> iniciador ou sonda sintética <400> 55 ggccgcatgc gaattcgggc cccttttc 28 <210> 56 <211> 35 <212> DNA <213> iniciador ou sonda sintética <400> 56 tcgagaaaag agaggctgaa gcctgcagca tatgc 35 <210> 57 <211> 35 <212> DNA <213> iniciador ou sonda sintética <400> 57 ggccgcatat gctgcaggct tcagccfcctc ttttc 35 <2l0> 58 <211> 3997 <212> DNA <213> vetor pPICZalphaD' ElsH6 <400> 58 agatctaaca tccaaagacg aaaggttgaa tgaaaccttt ttgccatccg acatccacag

131 gtccattctc acacataagt gccaaacgca acaggagggg atacactagc agcagaccgt 120 tgcaaacgca ggacctccac tcctcttctc ctcaacaccc acttttgcca tcgaaaaacc 180 agcccagtta ttgggcttga ttggagcfccg ctcattccaa ttccttctat taggctacta 240 acaccatgac tttattagcc tgtctatcct ggcccccctg gcgaggttca tgtttgttta 300 tttccgaatg caacaagotc cgcattacac ccgaacatca ctccagatga gggctttctg 360 agtgtggggt caaatagttt catgttcccc aaatggccca aaactgacag tttaaacgct 420 gtcttggaac ctaatatgac aaaagcgtga tctcatccaa gatgaactaa gtttggttcg 480 ttgaaatgct aacggccagt tggtcaaaaa caaacttcca aaagtcggca taccgtttgt 540 cttgtttggt attgattgac gaatgctcaa aaataatctc attaatgctt agcgcagtct 600 ctctatcgct tctgaacccc ggtgcacctg tgccgaaacg caaatgggga aacacccget 660 ttttggatga ttatgcattg tctccacatt gtatgcttcc aagattctgg tgggaatact 720 gctgatagcc taacgttcat gatcaaaatt taactgttct aacccctacrt tgacagcaat 780 atataaacag aaggaagctg ccctgtctta aacctttttt tttatcatca ttattagctt B40 actttcataa ttgcgactgg ttccaattga caagcttttg attttaacga cttttaacga 900 caacttgaga agatcaaaaa acaactaatt attcgaaacg atgagatttc cttcaatttt 960 tactgetgtt ttattcgcag catcctccgc attagctgct ccagtcaaca ctacaacaga 1020 agatgaaacg gcacaaattc cggctgaagc tgtcatcggt tactcagatt tagaagggga 1080 tttcgatgtt gctgttttgc cattttccaa cagcacaaat aacgggttat tgtttataaa 1140 tactactatt gccagcattg ctgctaaaga agaaggggta tctctcgaga aaaggtatga 1200 ggtgegeaac gtgteeggga tgtaccatgt cacgaacgac tgctccaact caagcattgt 1260 gtatgaggca gcggacatga tcatgcacac ccccgggtgc gtgccctgcg ttcgggagaa 1320 caactcttcc cgctgctggg tagcgctcac ccccacgctc gcagctagga acgccagcgt 1380 ccccactacg acaatacgac gccacgtcga tttgctogtt ggggcggctg ctttctgttc 1440 cgctatgtac gtgggggatc tctgcggatc tgtcttcctc gtctcccagc tgttcaccat 1500 ctcgcctcgc cggcatgaga cggtgcagga ctgcaattgc tcaatctatc ccggccacat 1560 aacaggtcac cgtatggctt gggatatgat gatgaactgg caccaccacc atcaccatta 1620 aagatctaag cttgaatccc gcggccatgc gaattcgcat gcggccgcca gctttctaga 1680 acaaaaactc atctcagaag aggatctgaa tagcgccgtc gaccatcatc atcatcatca 1740 ttgagtttgt agccttagac atgactgttc ctcagttcaa gttgggeact tacgagaaga 1800 ccggtcttgc tagattctaa tcaagaggat gtcagaatgc catttgcctg agagatgcag 1860 gcttcatttt tgatactttt ttatttgtaa cctatatagt ataggatttt ttttgteatt 1920

132

<td colspan="2">ttgtttcttc tcgtacgagc</td><td>ttgctcctga</td><td>tcagcctatc</td><td>tcgcagctga</td><td>tgaatatctt</td><td> 1980</td>

<td>gtggtagggg</td><td>tttgggaaaa</td><td>tcattcgagt</td><td colspan="2">ttgatgtttt tcttggtatt</td><td>tcccactcct</td><td> 2040</td>

<td colspan="2">cttcagagta cagaagatta</td><td>agtgagacct</td><td>tcgtttgtgc</td><td>ggatccceca</td><td>cacaccatag</td><td> 2100</td>

<td>cttcaaaatg</td><td>tttctaetcc</td><td>ttttttactc</td><td>ttccagattt</td><td>tctoggactc</td><td>cgcgcatcgc</td><td> 2160</td>

<td>cgtaccactt</td><td>caaaacaccc</td><td>aagcacagca</td><td>tactaaattt</td><td>tccctctttc</td><td>ttcctctagg</td><td> 2220</td>

<td>gtgtcgttaa</td><td>ttacccgtac</td><td>taaaggtttg</td><td>gaaaagaaaa</td><td>aagagaccgc</td><td>ctcgtttctt</td><td> 2280</td>

<td>tttcttcgtc</td><td>gaaaaaggca</td><td>ataaaaattt</td><td>ttatcacgtt</td><td>tctttttctt</td><td>gaaattttfct</td><td> 2340</td>

<td>tttttagttt</td><td>ttttctcttt</td><td>cagtgacctc</td><td>cattgatatt</td><td>taagttaata</td><td>aacggtcttc</td><td> 2400</td>

<td>aatttctcaa</td><td>gtttcagttt</td><td>catttttctt</td><td>gttctattac</td><td>aacttttfctt</td><td>acttcttgtt</td><td> 2460</td>

<td>cattagaaag</td><td>aaagcatagc</td><td>aatctaatct</td><td>aaggggcggt</td><td>gttgacaatt</td><td>aatcatcggc</td><td> 2520</td>

<td>atagtatatc</td><td>ggcatagtat</td><td>aatacgacaa</td><td>ggtgaggaac</td><td>taaaccatgg</td><td>ccaagttgac</td><td> 2580</td>

<td>cagtgccgtt</td><td>ccggtgctca</td><td>ccgcgcgcga</td><td>cgtcgccgga</td><td>gcggtcgagt</td><td>tctggaccga</td><td> 2640</td>

<td>ccggctcggg</td><td>ttctcccggg</td><td>acttcgtgga</td><td>ggacgacttc</td><td>gccggtgtgg</td><td>tccgggacga</td><td> 2700</td>

<td>cgtgaccctg</td><td>ttcatcagcg</td><td>cggtccagga</td><td>ccaggtggtg</td><td>ccggacaaca</td><td>ccctggcctg</td><td> 2760</td>

<td>ggtgtgggtg</td><td>cgcggcctgg</td><td>acgagctgta</td><td>cgccgagtgg</td><td>tcggaggtcg</td><td>tgtccacgaa</td><td> 2820</td>

<td>cttccgggac</td><td>gcctccgggc</td><td>cggccatgac</td><td>cgagatcggc</td><td>gagcagccgt</td><td>crcjcrctacacrcra</td><td> 2880</td>

gttcgccctg cgcgacccgg ccggcaactg cgtg-cacttc gtggccgagg agcaggactg 2940 acacgtccga cggcggccca cgggtcccag gcctcggaga tccgtcccce ttttcctttg 3000 tcgatatcat gtaattagtt atgtcacgct tacattcacg ccctcccccc acatccgctc 3060 taaccgaaaa ggaaggagtt agacaacctg aagtctaggt coctatttat ttttttatag 3120 ttatgttagt attaagaacg ttatttatat ttcaaatttt tctttttttt ctgtacagac 3180 gcgtgtacgc atgtaacatt atactgaaaa ccttgcttga gaaggttttg ggacgctcga 3240 aggctttaat ttgcaagctg gagaccaaca tgtgagcaaa aggccagcaa aaggccagga 3300 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc 3360 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg 3420 cgtttccocc tggaagctce ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat 3480 acctgtccgc ctttctccct togggaagcg tggcgctttc tcaatgctca cgctgtaggt 3540 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc 3600 agcccgaceg ctgcgcctta tccggtaact atcgtcttga gtccaacceg gtaagacacg 3660 acttatcgcc actggcagca gecactggta acaggattag cagagcgagg tatgtaggcg 3720

133 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg 3780 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg 3840 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca 3900 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga 3960 acgaaaactc acgttaaggg attttggtca tgagatc 3997 <210> 59 <211> 4004 <212> DMA <213> vetor pPICZalphaE'ElsH6 <400> 59

<td>agat.ct.aaca</td><td>tccaaagacg</td><td>aaaggttgaa</td><td>tgaaaccttt</td><td>ttgccatccg</td><td>acatccacag</td><td> €0</td>

<td>gtccattctc</td><td>acacataagt</td><td>gccaaacgca</td><td>acaggagggg</td><td>atacactagc</td><td>agcagaccgt</td><td> 120</td>

<td>tgcaaacgca</td><td>ggacctccac</td><td>tcctcttctc</td><td>ctcaacaccc</td><td>acttttgcca</td><td>tcgaaaaacc</td><td> 180</td>

<td>agcccagtta</td><td>ttgggcttga</td><td>ttggagctcg</td><td>ctcattccaa</td><td>ttccttctat</td><td>taggctacta</td><td> 240</td>

<td>acaccatgac</td><td>tttattagcc</td><td>tgtctatcct</td><td>ggcccccctg</td><td>gcgaggttca</td><td>tgtttgttta</td><td> 300</td>

<td>tttccgaatg</td><td>caacaagctc</td><td>cgcattacac</td><td>ccgaacatca</td><td>ctccagatga</td><td>gggctttetg</td><td> 360</td>

<td>agtgtggggt</td><td>caaatagttt</td><td>catgttcccc</td><td>aaatggccca</td><td>aaactgacag</td><td>tttaaacgct</td><td> 420</td>

<td>gtcttggaac</td><td>ctaatatgac</td><td>aaaagcgtga</td><td>tctcatccaa</td><td>gatgaactaa</td><td>gtttggttcg</td><td>4B0</td>

<td>ttgaaatgct</td><td>aacggccagt</td><td>tggtcaaaaa</td><td>gaaacttcca</td><td>aaagtcggca</td><td>taccgtttgt</td><td> 540</td>

<td>cttgtttggt</td><td>attgattgac</td><td>gaatgctcaa</td><td>aaataatctc</td><td>attaatgctt</td><td>agcgcagtct</td><td> 600</td>

<td>ctctatcgct</td><td>tctgaacccc</td><td>ggtgcacctg</td><td>tgccgaaacg</td><td>caaatgggga</td><td>aacacccgct</td><td> 660</td>

<td>ttttggatga</td><td>ttatgcattg</td><td>tctccacatt</td><td>gtatgcttcc</td><td>aagattctgg</td><td>tgggaatact</td><td> 720</td>

<td>gctgatagcc</td><td>taacgttcat</td><td>gatcaaaatt</td><td>taaetgttct</td><td>aacccctact</td><td>tgacagcaat</td><td> 780</td>

<td>atataaacag</td><td>aaggaagctg</td><td>ccctgtctta</td><td>aacctttttt</td><td>tttatcatca</td><td>ttattagctt</td><td> 840</td>

<td>actttcataa</td><td>ttgcgactgg</td><td>ttccaattga</td><td>caagcttttg</td><td>attttaacga</td><td>cttttaacga</td><td> 900</td>

<td>caacttgaga</td><td>agatcaaaaa</td><td>acaactaatt</td><td>attcgaaacg</td><td>atgagatttc</td><td>cttcaatttt</td><td> 960</td>

<td>tactgctgtt</td><td>ttattcgcag</td><td>catcctccgc</td><td>attagctgct</td><td>ccagtcaaca</td><td>ctacaacaga</td><td> 1020</td>

<td>agatgaaacg</td><td>gcacaaattc</td><td>cggctgaagc</td><td>tgtcatcggt</td><td>tactcagatt</td><td>tagaagggga</td><td> 1080</td>

<td>tttcgatgtt</td><td>gctgt.ttt.gc</td><td>cattttccaa.</td><td>cagcacaaat</td><td>aacgggttat</td><td>tgtttataaa</td><td> 1140</td>

<td>tactactatt</td><td>gccagcattg</td><td>ctgctaaaga</td><td>agaaggggta</td><td>tctctcgaga</td><td>aaagagaggc</td><td> 1200</td>

<td>tgaagcctat</td><td>gaggtgcgca</td><td>acgtgtccgg</td><td>gatgtaccat</td><td>gtcacgaacg</td><td>actgctccaa</td><td> 1260</td>

<td>etcaagcatt</td><td>gtgtatgagg</td><td>cagcggacat</td><td>gatcatgcac</td><td>acccccgggt</td><td>gcgtgocctg</td><td> 1320</td>

134

<td colspan="2">cgttcgggag aacaactctt</td><td>cccgctgctg</td><td>ggtagcgctc</td><td>accoccacgc</td><td>tcgcagctag</td><td> 1380</td>

<td colspan="2">gaacgccagc gtccccacta</td><td>cgacaatacg</td><td>acgccacgtc</td><td>gatttgctcg</td><td>ttggggcggc</td><td> 1440</td>

<td>tgctttctgt</td><td>tccgctatgt</td><td>acgtggggga</td><td>tcfcctgcgga</td><td>tctgtcttcc</td><td>tcgtctccca</td><td> 1500</td>

<td>gctgttcacc</td><td>atctcgcctc</td><td>gccggcatga</td><td>gacggtgcag</td><td>gactgcaatfc</td><td>gctcaatcta</td><td>iseo</td>

<td>tcccggccac</td><td>ataacgggtc</td><td>accgtatggc</td><td>ttgggatatg</td><td>atgatgaact</td><td>ggcaccacca</td><td> 1620</td>

<td>ccatcaccat</td><td>taaagatcta</td><td>agcttgaatc</td><td>ccgcggccat</td><td>ggcatatgcg</td><td>gcegccagct</td><td> 1680</td>

<td>ttctagaaca</td><td>aaaactcatc</td><td>tcagaagagg</td><td>atctgaatag</td><td>cgccgtcgac</td><td>catcatcatc</td><td> 1740</td>

<td>atcatcattg</td><td>agtttgtagc</td><td>cttagacatg</td><td>actgttcctc</td><td>agttcaagtt</td><td>gggcacttac</td><td> 1800</td>

<td>gagaagaccg</td><td>gtcttgctag</td><td>attctaatca</td><td>agaggatgtc</td><td>agaatgccat</td><td>ttgcctgaga</td><td> 1860</td>

<td>gatgcaggct</td><td>tcatttttga</td><td>tactttttta</td><td>tttgtaacct</td><td>atatagtata</td><td>ggattttfctt</td><td> 1920</td>

<td>tgtcattttg</td><td>tttcttctcg</td><td>tacgagcttg</td><td>ctcctgatca</td><td>gcctatctcg</td><td>cagctgatga</td><td> 1980</td>

<td>atatcttgtg</td><td>gtaggggttt</td><td>gggaaaatca</td><td>ttcgagtttg</td><td>atgtttttct</td><td>tggtatttcc</td><td> 2040</td>

<td>cactcctctt</td><td>cagagtacag</td><td>aagattaagt</td><td>gagaccttcg</td><td>tttgtgcgga</td><td>tccoccacac</td><td> 2100</td>

<td>accatagctt</td><td>caaaatgttt</td><td>ctactccttt</td><td>tttactcttc</td><td>cagattttct</td><td>cggactccgc</td><td> 2160</td>

<td>gcatcgccgt</td><td>accacttcaa</td><td>aacacccaag</td><td>cacagcatac</td><td>taaattttcc</td><td>ctctttcttc</td><td> 2220</td>

<td>ctctagggtg</td><td>tcgttaatta</td><td>ççcgtactaa</td><td>agCTtttggaa</td><td>aagaaaaaag</td><td>ayci€€^CCt€</td><td>22SQ</td>

<td>gtttcttttt</td><td>cttcgtcgaa</td><td>aaaggcaata</td><td>aaaattttta</td><td>tcacgtttct</td><td>fctttcttgaa.</td><td> 2340</td>

$Ç£t.fc£ttt£, ttagt t fcttfc* tctctttcatt tCTflCCtCCSt· rr«-4—==φ a ^'y' *y« · n<Jk acw~------£*xvw

<td>ggtcttcaat</td><td>ttctcaagtt</td><td>tcagtttcat</td><td>ttttcttgtt ctattacaac</td><td>tttttttact</td><td> 2460</td>

<td>tcttgttcat</td><td>tagaaagaaa</td><td>gcatagcaat</td><td>ctaatctaag gggcggtgtt</td><td>gacaattaat</td><td> 2520</td>

<td>catcggcata</td><td>gtatatcggc</td><td>atagtataat</td><td>acgacaaggt gaggaactaa</td><td>accatggeca</td><td> 2580</td>

<td>agttgaccag</td><td>tgcogttccg</td><td>gtgctcaccg</td><td>cgcgcgacgt cgccggagcg</td><td>gtcgagttct</td><td> 2640</td>

<td>ggaccgaccg</td><td>gctcgggttc</td><td>tcccgggact</td><td>tcgtggagga cgacttcgcc</td><td>ggtgtggtcc</td><td> 2700</td>

<td>gggacgacgt</td><td>gaccctgtfcc</td><td>atcagcgcgg</td><td>tccaggacca ggtggtgccg</td><td>gacaacaccc</td><td> 2760</td>

<td>tggcctgggt</td><td>gtgggtgcgc</td><td>ggcctggacg</td><td>agctgtacgc cgagtggtcg</td><td>gaggtcgtgt</td><td> 2820</td>

<td>ccacgaactt</td><td>ccgggacgcc</td><td>tccgggccgg</td><td>ccatgaccga gatcggcgag</td><td>cagccgtggg</td><td> 2880</td>

<td>ggcgggagtt</td><td>cgccctgcgc</td><td>gacccggecg</td><td>gcaactgcgt gcacttcgtg</td><td>gccgaggagc</td><td> 2940</td>

<td>aggactgaca</td><td>cgtccgacgg</td><td>cggcccacgg</td><td>gtcccaggcc tcggagatcc</td><td>gtaocccttt</td><td> 3000</td>

<td>tcctttgtcg</td><td>atatcatgta</td><td>attagttatg</td><td>tcacgcttac attcacgccc</td><td>tocccccaea</td><td> 3060</td>

<td>tccgctctaa</td><td>ccgaaaagga</td><td>aggagttaga</td><td>caacctgaag tctaggtccc</td><td>tatttatttt</td><td> 3120</td>

<td>tttatagtta</td><td>tgrttagtatt</td><td>aagaacgtta</td><td>ttfcafcatttc aaatttttct</td><td>tttttttctg</td><td> 3180</td>

135

<td>tacagaegcg</td><td>tgtacgcatg</td><td>taacattata</td><td>ctgaaaacct</td><td>tgcttgagaa</td><td>ggttttggga</td><td> 3240</td>

<td>cgctcgaagg</td><td>ctttaatttg</td><td>caagctggag</td><td>accaacatgt</td><td>gagcaaaagg</td><td>ccagcaaaag</td><td> 3300</td>

<td>gccaggaacc</td><td>gtaaaaaggc</td><td>cgcgttgctg</td><td>gcgtttttcc</td><td>ataggetccg</td><td>cccccctgac</td><td> 3360</td>

<td>gagcatcaca</td><td>aaaatcgacg</td><td>ctcaagtcag</td><td>aggtggegaa</td><td>acccgacagg</td><td>actataaaga</td><td> 3420</td>

<td>taccaggcgt</td><td>ttccccctgg</td><td>aagctccctc</td><td>gtgcgctctc</td><td>ctgttccgac</td><td>cctgccgctt</td><td> 3480</td>

<td>accggatacc</td><td>tgtccgcctt</td><td>tctcccttcg</td><td>ggaagcgtgg</td><td>cgctttctca</td><td>atgetcacgc</td><td> 3540</td>

<td>tgtaggtatc</td><td>tcagttoggt</td><td>gtaggtcgtt</td><td>cgctccaagc</td><td>tgggctgtgt</td><td>gcacgaaccc</td><td> 3600</td>

<td>cccgttcagc</td><td>ccgaccgctg</td><td>cgccttatcc</td><td>ggtaactatc</td><td>gtcttgagtc</td><td>caacccggta</td><td> 3660</td>

<td>agacacgact</td><td>tatcgccact</td><td>ggcagcagcc</td><td>actggtaaca</td><td>ggattageag</td><td>agcgaggtat</td><td> 3720</td>

<td>gtaggcggtg</td><td>ctacagagtt</td><td>cttgaagtgg</td><td>tggcctaact</td><td>acggctacac</td><td>tagaaggaea</td><td> 3780</td>

<td>gtatttggta</td><td>tctgcgctct</td><td>gctgaagcca</td><td>gttaccttcg</td><td>gaaaaagagt</td><td>tggtagctct</td><td> 3840</td>

<td>tgatccggca</td><td>aacaaaccac</td><td>cgctggtagc</td><td>ggtggttttt</td><td>ttgtttgcaa</td><td>geageagatt</td><td> 3900</td>

<td>acgcgcagaa</td><td>aaaaaggatc</td><td>tcaagaagat</td><td>cctttgatct</td><td>tttctaeggg</td><td>gtctgacgct</td><td> 3960</td>

<td>cagtggaaeg</td><td>aaaactcacg</td><td>ttaagggatt</td><td colspan="2">ttggtcatga gatc</td><td></td><td> 4004</td>

<210> 60 <211> 4492 <212> DNA <213> vetor pPICZalphaO'E2sH6 <400> 60

<td>agatetaaca</td><td>tocaaagacg</td><td>aaaggttgaa</td><td>tgaaaccttt</td><td>ttgccatccg</td><td>acatecacag</td><td> 60</td>

<td>gtecattcte</td><td>acacataagt</td><td>gccaaacgca</td><td>acaggagggg</td><td>atacactage</td><td>agcagaccgt</td><td> 120</td>

<td>tgcaaacgca</td><td>ggacctccac</td><td>tcctcttctc</td><td>ctcaaeaccc</td><td>acttttgcca</td><td>tcgaaaaacc</td><td> 180</td>

<td>agcccagtta</td><td>ttgggcttga</td><td>ttggagctcg</td><td>ctcattccaa</td><td>ttecttetat</td><td>taggctaeta</td><td> 240</td>

<td>acaccatgac</td><td>tttattagcc</td><td>tgtcfcafccct</td><td>ggcccecetg</td><td>gegaggttca</td><td>tgtttgttta</td><td> 300</td>

<td>tttccgaatg</td><td>caacaagctc</td><td>cgcattacac</td><td>ccgaacatca</td><td>ctccagatga</td><td>gggctttctg</td><td> 360</td>

<td>agtgtggggt</td><td>caaatagttt</td><td>eatgtteccc</td><td>aaatggccca</td><td>aaaetgacag</td><td>tttaaaogct</td><td> 420</td>

<td>gtcttggaac</td><td>ctaatatgac</td><td>aaaagcgtga</td><td>tcteatecaa</td><td>gatgaactaa</td><td>gtttggttcg</td><td> 480</td>

<td>ttgaaatgct</td><td>aacggccagt</td><td>tggtcaaaaa</td><td>gaaacttcca</td><td>aaagtcggea</td><td>tacegtttgt</td><td> 540</td>

<td>cttgtttggt</td><td>attgattgac</td><td>gaatgctcaa</td><td>aaataatctc</td><td>attaatgctt</td><td>agcgcagtct</td><td> 600</td>

<td>ctctatcgct</td><td>tctgaaoccc</td><td>ggtgcacctg</td><td>tgccgaaaeg</td><td>caaatgggga</td><td>aacacccgct</td><td> 660</td>

<td>ttttggatga</td><td>ttatgcattg</td><td>tctccacatt</td><td>gtatgcttcc</td><td>aagattctgg</td><td>tgggaatact</td><td> 720</td>

<td>gctgatagcc</td><td>taacgttcat</td><td colspan="2">gateaaaatt taactgttct</td><td>aacccctact</td><td>tgaeageaat</td><td> 780</td>

136 atataaacag aaggaagctg ccctgtctta aacctttttt tttatcatca ttattagctt 840 actttcataa ttgcgactgg ttccaattga caagcttttg attttaacga cttttaacga 900 caacttgaga agatcaaaaa acaactaatt attcgaaacg atgagatttc cttcastttt 96o tactgctgtt ttattcgcag catcctocgc attagctgct ccagtcaaca ctacaacaga 1020

<td>agatgaaacg gcacaaattc cggctgaagc tgtcatcggt</td><td>tactcagatt tagaagggga</td><td> 1080</td>

<td>tttcgatgtt gctgttttgc cattttccaa cagcacaaat</td><td>aacgggttat tgtttataaa</td><td> 1140</td>

<td>tactactatt gccagcattg ctgctaaaga agaaggggta</td><td>tctctcgaga aaaggcatac</td><td> 1200</td>

<td>ccgcgtgtca ggaggggcag cagcctccga taecaggggc</td><td>cfctgtgtccc tctttagccc</td><td> 1260</td>

<td>cgggtcggct cagaaaatcc agctcgtaaa caçcaacggc</td><td>agttggcaca tcaacaggac</td><td> 1320</td>

<td>tgccctgaac tgcaacgact ccctccaaac agggttcttt</td><td>gccgcactat tctacaaaca</td><td> 1380</td>

<td>caaattcaac tcgtctggat gcccagagcg cttggccagc</td><td>tgtcgcteca tcgacaagtt</td><td> 1440</td>

<td>cgctcágggg tggggtcccc tcacttacac tgagcctaac</td><td>agctcggacc agaggcccta</td><td> 1500</td>

<td>ctgctggcac tacgcgcctc gaccgtgtgg tattgtaccc</td><td>gcgtctcagg tgtgcggtcc</td><td> 1560</td>

<td>agtgtattgc ttcaccccga gccctgttgt ggtggggacg</td><td>accgatcggt ttggtgtccc</td><td> 1620</td>

<td>cacgtataac tggggggcga acgactcgga tgtgctgatt</td><td>ctcaacaaca cgcggccgcc</td><td> 1680</td>

<td>gcgaggcaac tggttcggct gtacatggat gaatggcaet</td><td>aoâ.cc?wc£wcr</td><td> 1740</td>

<td>gggccccccg tgcaacatcg ggggggccgg caacaacacc</td><td>ttgacctgcc ccactgactg</td><td> 1800</td>

<td>ttttcggaag caccccgagg ccacctacgc cagatgcggt</td><td>tctgggccct ggctgacacc</td><td> 1860</td>

<td>taggtgtatg gttcattacc catataggct ctggcactac</td><td>ccctgcactg tcaacttcac</td><td> 1920</td>

<td>catcttcaag gttaggatgt acgtgggggg cgtggagcac</td><td>aggttcgaag ccgcatgcaa</td><td> 1980</td>

<td>ttggactcga ggagagcgtt gtgacttgga ggacagggat</td><td>agatcagagc ttagoccgct</td><td> 2040</td>

<td>gctgctgtct acaacagagt ggcaggtgat cgagggcaga</td><td>caccatcacc accatcacta</td><td> 2100</td>

<td>atagttaatt aactgcaggc atgcaagctt atcgataccg</td><td>tcgacgaatt cgcatgcggc</td><td> 2160</td>

<td>cgccagcttt ctagaacaaa aactcatctc agaagaggat</td><td>ctgaatagcg ccgtcgacca</td><td> 2220</td>

<td>tcatcatcat catcattgag tttgtagcct tagacatgac</td><td>tgttcctcag ttcaagttgg</td><td> 2280</td>

<td>gcacttacga gaagaccggt cttgctagat tctaatcaag</td><td>aggatgtcag aatgccattt</td><td> 2340</td>

<td>gcctgagaga tgcaggcttc atttttgata cttttttatt</td><td>tgtaacctat atagtatagg</td><td> 2400</td>

<td>attttttttg tcattttgtt tçttctcgta cgagcttgct</td><td>cctgatcagc -ctatetcgca</td><td> 2460</td>

gctgatgaat atcttgtggt aggggtttgg gaaaatcatt cgagtttgat gtttttcttg 2520 gtatttccca ctcctcttca gagtacagaa gattaagtga gaccttcgtt tgtgcggatc 2580

137

<td>ccccacacac</td><td colspan="2">catagcttca aaatgtttct</td><td>actccttttt</td><td>tactcttcca</td><td>gattttctcg</td><td> 2640</td>

<td>gactccgcgc</td><td colspan="2">atcgccgtac cacttcaaaa</td><td>cacccaagca</td><td>cagcatacta</td><td>aattttccct</td><td> 2700</td>

<td>ctttcttcct</td><td colspan="2">ctagggtgtc gttaattacc</td><td>cgtactaaag</td><td>gtttggaaaa</td><td>gaaaaaagag</td><td> 2760</td>

<td>accgcctcgt</td><td>ttctttttct</td><td>tcgtcgaaaa</td><td>aggcaataaa</td><td>aatttttatc</td><td>acgtttcttt</td><td> 2820</td>

<td>ttcttgaaat</td><td>tttttttttt</td><td>agtttttttc</td><td>tctttcagtg</td><td>acctçcattg</td><td>atatttaagt</td><td> 2880</td>

<td>taataaacgg</td><td>tcttcaattt</td><td>ctcaagtttc</td><td>agtttcattt</td><td>ttcttgttct</td><td>attacaactt</td><td> 2940</td>

<td>tttttacttc</td><td>ttgttcatta</td><td>gaaagaaagc</td><td>atagcaatct</td><td>aatctaaggg</td><td>gcggtgttga</td><td> 3000</td>

<td>caattaatca</td><td>tcggcatagt</td><td>atatcggcat</td><td>agtataatac</td><td>gacaaggtga</td><td>ggaactaaac</td><td> 3060</td>

<td>catggccaag</td><td>ttgaccagtg</td><td>ccgttccggt</td><td>gcVcaccgcg</td><td>cgcgaçgtcg</td><td>ccggagcggt</td><td> 3120</td>

<td>cgagttctgg</td><td>accgaccggc</td><td colspan="2">tcgggttctc -ccgggacttc</td><td>gtggaggacg</td><td>aettcgecgg</td><td> 3180</td>

<td>tgtggtccgg</td><td>gacgacgtga</td><td>ccctgttcat</td><td>cagcgcggtc</td><td>caggaccagg</td><td>tggtgccgga</td><td> 3240</td>

<td>caacaccctg</td><td>gcctgggtgt</td><td>gggtgcgcgg</td><td>cctggacgag</td><td>ctgtacgccg</td><td>agtggtcgga</td><td> 3300</td>

<td>ggtcgtgtcc</td><td>acgaacttcc</td><td>gggacgcctc</td><td>cgggccggcc</td><td>atgaccgaga</td><td>tcggcgagca</td><td> 3360</td>

<td>gccgtggggg</td><td>cgggagttcg</td><td>ccctgcgcga</td><td>cccggccggc</td><td>aactgcgtgc</td><td>acttcgtggc</td><td> 3420</td>

<td>cgaggagcag</td><td>gactgaeacg</td><td>tccgacggcg</td><td>gcccacgggt</td><td>cccaggcctc</td><td>ggagatccgt</td><td> 3480</td>

<td>cccccttttc</td><td>ctttgtcgat</td><td>atcatgtaat</td><td>tagttatgtc</td><td>acgcttacat</td><td>tcacgccctc</td><td> 3540</td>

cccccacatc cgctctaaec g&aaaggaag gagttagaca acctgaagtc taggtqccta 3600 tttatttttt tatagttatg ttagtattaa gaacgttatt tatatttcaa atttttcttt 3660 tttttctgta cagacgcgtg tacgcatgta acattatact gaaaaccttg cttgagaagg 3720 ttttgggacg ctcgaaggct ttaatttgca agctggagac caacatgtga gcaaaaggcc 3780 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc 3840 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac 3900 tataaagata ccaggcgttt ccccctggaa gctcccfccgt gcgctctcct gttccgaccc 3960 tgccgcttac cggatacctg tccgoctttc tcccttcggg aagcgtggcg ctttctcaat 4020 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc 4080 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcea 4140 acccggtaag acacgactta tcgocactgg cagcagccac tggtaacagg attagcagag 4200 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta 4260 gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg 4320 gtagctcttg atocggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc 4380 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt 4440

138 cügacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga tc 4492 <210> 61 <211> 4431 <212> DMA <213> vetox pPICZalphaE'E2sH6 <400> 61

<td>agatctaaca tccaaagacg aaaggttgaa tgaaacctfct</td><td>ttgccatccg acatccacag</td><td> 60</td>

<td>gtccattctc acacataagt gccaaacgca acaggagggg</td><td>atacactagc agcagaccgt</td><td> 120</td>

<td>tgcaaacgca ggacctccac tcctcttctc ctcaacaccc</td><td>acttttgcca tcgaaaaacc</td><td> 180</td>

<td>agcccagtta ttgggcttga ttggagctcg ctcattccaa</td><td>ttccttctat taggctacta</td><td> 240</td>

<td>acaccatgac tttattag.cc tgtctatcct ggcccccctg</td><td>gogaggttca tgtttgttta</td><td> 300</td>

<td>tttccgaatg caacaagctc cgcattacac ccgaacatca</td><td>ctccagatga gggctttctg</td><td> 360</td>

<td>agtgtggggt caaatagttt catgttcccc aaatggccca</td><td>aaactgacag tttaaacgct</td><td> 420</td>

<td>gtcttggaac ctaatafcgac aaaagcgtga tctcatccaa</td><td>gatgaact&a gtttggttcg</td><td> 480</td>

<td>ttgaaatgct aaoggccagt tggtcaaaaa gaaacttcca</td><td>aaagtcggca taccgtttgt</td><td> 540</td>

<td>cttgtttggt attgattgac gaatgctcaa aaataatctc</td><td>attaatgctt agcgcagtct</td><td> 600</td>

<td>ctctatcgct tctgaacccc ggtgcacctg tgccgaaacg</td><td>caaatgggga aacacccgct</td><td> 660</td>

<td>ttttggatga ttatgcattg tctccacatt gtafcgcttcc</td><td>aagattctgg tgggaatact</td><td> 720</td>

<td>gctgatagcc taacgttcat gatcaaaatt taactgttct</td><td>aacccctact tgacagcaat</td><td> 780</td>

<td>atataaacag aaggaagctg ccctgtctta aacctttttt</td><td>tttatcatca ttattagctt</td><td> 840</td>

<td>actttcataa ttgcgactgg ttccaattga caagcttttg</td><td>attttaacga cttttaacga</td><td> 900</td>

<td>caacttgaga agatcaaaaa acaactaatt attcgaaacg</td><td>atgagatttc cttcaatttt</td><td> 960</td>

<td>tactgctgtt ttattcgcag catcctccgc attagctgct</td><td>ccagtcaaca ctacaacaga</td><td> 1020</td>

<td>agatgaaacg gcacaaattc cggctgaagc tgtcatcggt</td><td>tactcagatt tagaagggga</td><td> 1080</td>

<td>tttcgafcgtt gctgttttgc cattttccaa cagcacaaat</td><td>aacgggttat tgtttataaa</td><td> 1140</td>

tactactatt gccagcattg ctgctaaaga agaaggggta tctctcgaga aaagagaggc 1200 tgaagcccat acccgcgtgt caggaggggc agcagcctoc gataccaggg gccttgtgtc 1260 cctctttagc cccgggtcgg ctcagaaaat ccagctcgta aacaccaacg gcagttggca 1320 catcaacagg actgccctga actgcaacga ctccctccaa acagggttct ttgccgcact 1380 attctacaaa cacaaattca actcgtctgg atgcocagag cgcttggcca gctgtcgctc 1440 catcgacaag ttcgctcagg ggtggggtcc cctcacttac actgagccta acagctcgga 1500 ccagaggccc tactgcfcggc actacgcgcc tcgaccgtgt ggtattgtac ccgcgtctca 1560

139

<td>ggtgtgcggt</td><td>ccagtgtatt</td><td>gcttcacccc</td><td>gagccctgtt</td><td>gtggtgggga ·</td><td>cgaccgatcg</td><td> 1620</td>

<td>gtttggtgtc</td><td>cccacgtata</td><td>actggggggc</td><td>gaacgactcg</td><td>gatgtgctga</td><td>ttctcaacaa.</td><td> 1680</td>

<td>cacgcggccg</td><td>ccgcgaggca</td><td>actggttcgg</td><td>ctgtacatgg</td><td>atgaatggca</td><td>ctgggttcac</td><td> 1740</td>

<td>caagacgtgt</td><td>gggggccCCC</td><td>cgtgcaacat</td><td>cgggggggcc</td><td>ggcaacaaca</td><td>ccttgacctg</td><td> 1800</td>

<td>ccccactgac</td><td>tgttttcgga</td><td>agcacoccga</td><td>ggccacctac</td><td>gccagatgcg</td><td>gttctgggcc</td><td> 1860</td>

<td>ctggctgaca</td><td>cctaggtgta</td><td>tggttcatta</td><td>cccatatagg</td><td>ctctggcact</td><td>acccctgcac</td><td> 1920</td>

<td>tgtcaacttc</td><td>accatcttca</td><td>aggttaggat</td><td>gtacgtgggg</td><td>ggcgtggagc</td><td>acaggttcga</td><td> 1980</td>

<td>agccgcatgc</td><td>aattggactc</td><td>gaggagagcg</td><td>ttgtgacttg</td><td>gaggacaggg</td><td>atagatcaga</td><td> 2040</td>

<td>gcttagcccg</td><td>ctgctgctgt</td><td>ctacaacaga</td><td>gtggcaggtg</td><td>atcgagggca</td><td>gacaccatca</td><td> 2100</td>

<td>ccaccatcac</td><td>taatagttaa</td><td>ttaactgcag</td><td>gcatgcaagc</td><td>ttatcgatac</td><td>cgtcgaccat</td><td> 2160</td>

<td>catcatcatC</td><td>atcattgagt</td><td>ttgtagcctt</td><td>agacatgact</td><td>gttcctcagt</td><td>tcaagttggg</td><td> 2220</td>

<td>cacttacgag</td><td>aagaccggtc</td><td>ttgctagatt</td><td>ctaatcaaga</td><td>ggatgtcaga</td><td>atgccatttg</td><td> 2280</td>

<td>cctgagagat</td><td>gcaggcttca</td><td>tttttgatac</td><td>ttttttattt</td><td>gtaacctata</td><td>tagtatagga</td><td> 2340</td>

<td>tttttt.tt.gt</td><td>cattttgttt</td><td>cttctcgtac</td><td>gagcttgctc</td><td>ctgatcagcc</td><td>tatctcgcag</td><td> 2400</td>

<td>ctgatgaata</td><td>tcttgtggta</td><td>ggggtttggg</td><td>aaaatcattc</td><td>gagtttgatg</td><td>tttttcttgg</td><td> 2460</td>

<td>tatttcccac</td><td>tcctcttcag</td><td>agtacagaag</td><td>attaagtgag</td><td>accttcgttt</td><td>gtgcggatcc</td><td> 2520</td>

<td>cccacacacc</td><td>atagcttcaa</td><td>aatgtttcta</td><td>ctcctttttt</td><td>actcttccag</td><td>attttctcgg</td><td> 2580</td>

<td>actccgcgca</td><td>tcgccgtacc</td><td>acttcaaaac</td><td>aoceaagcac</td><td>agcatactaa</td><td>attttccctc</td><td> 2640</td>

<td>tttcttcctc</td><td>tagggtgtcg</td><td>ttaattaccc</td><td>gtactaaagg</td><td>tttggaaaag</td><td>aaaaaagaga</td><td> 2700</td>

ccgcctcgtt tctttttctt cgtcgaaaaa ggcaataaaa atttttatca cgtttçtttt 2760 tcttgaaatt ttttttttta gtttttttct ctttcagtga cctccattga tatttaagtt 2820 aataaacggt cttcaatttc tcaagtttca gtttcatttt tcttgttcta ttacaacttt 2880 ttttacttct tgttcattag aaagaaagca tagcaatcta atctaagggg cggtgttgac 2940 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc 3000 atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc 3060 gagttctgga ccgaccggct cgggttctoc ogggacttcg tggaggacga cttcgccggt 3120 gtggtccggg acgacgtgac cctgtteatc agcgcggtcc aggaccaggt ggtgccggac 3180 aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag 3240 gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca tgaccgagat cggcgagcag 3300 ccgtgggggc gggagttcgc cctgcgcgac ccggcoggca actgcgtgca cttcgtggcc 336o

333

140

<td>gaggagcagg</td><td>actgacacgt</td><td>ccgacggcgg</td><td>cccacgggtc</td><td>ccaggcctog</td><td>gagatccgtc</td>

<td>ccecttttcc</td><td>tttgtcgata</td><td>tcatgtaatt</td><td>agttatgtca</td><td>cgcttacatt</td><td>cacgccctcc</td>

<td>ccccacatcc</td><td>gctctaaccg</td><td>aaaaggaagg</td><td>agttagacaa</td><td>ccfcgaagtct</td><td>aggtccctat</td>

<td>ttattttttt</td><td>atagttatgt</td><td>tagtattaag</td><td>aacgttattt</td><td>atatttcaaa</td><td>tttttcttfct</td>

<td>ttttctgtac</td><td>agacgcgtgt</td><td>acgcatgtaa</td><td>cattatactg</td><td>aaaaccttgc</td><td>ttgagaaggt</td>

<td>tttgggacgc</td><td>tcgaaggett</td><td>taatttgcaa</td><td>gctggagacc</td><td>aacatgtgag</td><td>caaaaggcca</td>

<td>gcaaaaggcc</td><td>aggaaccgta</td><td>aaaaggccgc</td><td>gttgctggcg</td><td>tttttccata</td><td>ggctccgccc</td>

<td>ccctgacgag</td><td>catcacaaaa</td><td>atcgacgctc</td><td>aagtcagagg</td><td>tggcgaaacc</td><td>cgacaggact</td>

<td>ataaagatac</td><td colspan="2">caggcgtttc cccctggaag</td><td>ctccctcgtg</td><td>cgctctcctg</td><td>ttccgaccct</td>

<td>gccgcttacc</td><td>ggatacctgt</td><td>ccgcctttct</td><td>cccttcggga</td><td>agcgtggcgc</td><td>tttctcaatg</td>

<td>ctcacgctgt</td><td>aggtatctca</td><td>gttcggtgta</td><td>ggtcgttcgc</td><td>tccaagctgg</td><td>gctgtgtgca</td>

<td>cgaacccccc</td><td>gttcagcccg</td><td>accgctgcgc</td><td>cttatccggt</td><td>aactatcgtc</td><td>ttgagtccaa</td>

<td>cccggtaaga</td><td>cacgacttat</td><td>cg-ccactggc</td><td>agcagccact</td><td>ggtaacagga</td><td>ttagcagagc</td>

<td>gaggtatgta</td><td colspan="2">ggcggtgcta cagagttctt</td><td colspan="2">gaagtggtgg cctaactacg</td><td>gctacactag</td>

<td>aaggacagta</td><td>tttggtatct</td><td>gcgctctgct</td><td>gaagccagtt</td><td>accttcggaa</td><td>aaagagttgg</td>

<td>tagctcttga</td><td>tccggcaaac</td><td>aaaccaccgc</td><td>tggtagcggt</td><td>ggtttttttg</td><td>tttgcaagca</td>

<td>gcagattacg</td><td>cgcagaaaaa</td><td>aaggatctca</td><td>agaagatcct</td><td>ttgatctttt</td><td>ctacggggtc</td>

<td>tgacgctcag</td><td>tggaacgaaa</td><td>actcacgtta</td><td>agggattttg</td><td>gtcatgagat</td><td>c</td>

<td colspan="2"><213> vetor pUCl8MFa</td><td></td><td></td><td></td><td></td>

<td>gcgcccaata</td><td>cgcaaaccgc</td><td>ctctccccgc</td><td>gcgttggceg</td><td>attcattaat</td><td>gcagctggca</td>

<td>cgacaggttt</td><td>cccgactgga</td><td>aagcgggcag</td><td>tgagcgcaac</td><td>gcaattaatg</td><td>tgagttagct</td>

<td>cactcattag</td><td>gcaccccagg</td><td>ctttacactt</td><td>tatgcttecg</td><td>gctcgtatgt</td><td>tgtgtggaat</td>

<td>tgtgagcgga</td><td>taacaatttc</td><td>acacaggaaa</td><td>cagctatgac</td><td>catgattacg</td><td>ccaagcttac</td>

<td>cccttcttct</td><td>ttagcagcaa</td><td>tgctggcaat</td><td>agtagtattt</td><td>ataaacaata</td><td>acccgttatt</td>

<td>tgtgctgttg</td><td>gaaaatggca</td><td>aaacagcaac</td><td>atcgaaatcc</td><td>ccttctaaat</td><td>ctgagtaacc</td>

<td>gatgacagct</td><td>tcagccggaa</td><td>tttgtgccgt</td><td>ttcafccttct</td><td>gttgtagtgt</td><td>tgactggagc</td>

<td>agctaahgcg</td><td>gaggatgctg</td><td>cgaataaaac</td><td>tgcagtaaaa</td><td>attgaaggaa</td><td>atctcatgaa</td>

<td>ttcactggcc</td><td>gtcgttttac</td><td>aacgtcgtga</td><td>ctgggaaaac</td><td>cctggcgtta</td><td>cccaacttaa</td>

3420

3480

3540

3600

3660

3720

3780

3840

3900

3960

4020

4080

4140

4200

4260

4320

4380

4431

120

180

240

300

360

420

480

540

141

<td>tcgecttgca</td><td>gcacatcccc</td><td>ctttcgccag</td><td>ctggcgtaat i</td><td>agcgaagagg .</td><td>cccgcaccga</td><td> 600</td>

<td>tcgcccttcc</td><td>caacagttgc</td><td>gcagcctgaa</td><td>tggcgaatgg -</td><td>cgcctgatgc <</td><td>ggtattttct</td><td> 660</td>

<td>ccttacgcat</td><td>ctgtgcggta</td><td>tttcacaccg</td><td>catatggtgc</td><td>actctcagta</td><td>caatctgctc</td><td> 720</td>

<td>tgatgccgca</td><td>tagttaagcc</td><td>agccccgaca</td><td>cccgccaaca</td><td>cccgctgacg</td><td>cgocctgacg</td><td> 780</td>

<td>ggcttgtctg</td><td>ctcccggcat</td><td>ccgcttacag</td><td>acaagctgtg</td><td>accgtctccg</td><td>ggagctgcat</td><td> 840</td>

<td>gtgtcagagg</td><td>ttttcaccgt</td><td>catcaccgaa</td><td>acgcgcgaga</td><td>cgaaagggcc</td><td>tcgtgatacg</td><td> 900</td>

<td>octattttta</td><td>taggttaatg</td><td>tcatgataat</td><td>aatggtttct</td><td>tagacgtcag</td><td>gtggcacttt</td><td> 960</td>

<td>tcggggaaat</td><td>gtgcgcggaa</td><td>cccctatttg</td><td>tttatttttc</td><td>taaatacatt</td><td>caaatatgta</td><td> 1020</td>

<td>tccgctcatg</td><td>agacaataaç</td><td>cctgataaat</td><td>gcttcaataa</td><td>tattgaaaaa</td><td>ggaagagtat</td><td> 1080</td>

<td>gagtattcaa</td><td>catttccgtg</td><td>tcgcccttat</td><td>tccctttttt</td><td>gcggcatttt</td><td>gccttcctgt</td><td> 1140</td>

<td>ttttgctcac</td><td>ccagaaacgc</td><td>tggtgaaagt</td><td>aaaagatgct</td><td>gaagatcagt</td><td>tgggtgcacg</td><td> 1200</td>

<td>agtgggttac</td><td>atcgaactgg</td><td>atctcaacag</td><td>cggtaagatc</td><td>cttgagagtt</td><td>ttcgccccga</td><td> 1260</td>

<td>agaacgtttt</td><td>ccaatgatga</td><td>gcacttttaa</td><td>agttctgcta</td><td>tgtggcgcgg</td><td>tattatcccg</td><td> 1320</td>

<td>tattgacgcc</td><td>gggcaagagc</td><td>aactcggtcg</td><td>ccgcatacac</td><td>tattctcaga</td><td>atgacttggt</td><td> 1380</td>

<td>tgagtactca</td><td>ccagtcacag</td><td>aaaagcatct</td><td>tacggatggc</td><td>atgacagtaa</td><td>gagaattatg</td><td> 1440</td>

<td>cagrtgctgcc</td><td>ataaccatga</td><td>gtgataacac</td><td>tgeggccaac</td><td>ttacttctga</td><td>caacgatcgg</td><td> 1500</td>

<td>aggaecgaag</td><td>gagctaaccg</td><td>cttttttgca</td><td>eaaeatgggg</td><td>gatcatgtaa</td><td>ctcgccttga</td><td>1S60</td>

<td>tcgttgggaa</td><td>ccggagctga</td><td>atgaagccat</td><td>accaaacgac</td><td>gagcgtgaca</td><td>ccacgatgcc</td><td> 1620</td>

<td>tgtagcaatg</td><td>gcaacaacgt</td><td>tgcgcaaact</td><td>attaactggc</td><td>gaactactta</td><td>ctctagcttc</td><td> 1580</td>

<td>ccggcaacaa</td><td>ttaatagact</td><td>ggatggaggc</td><td>ggataaagtt</td><td>gcaggaccac</td><td>ttctgcgctc</td><td> 1740</td>

<td>ggcccttccg</td><td>gctggctggt</td><td>ttattgctga</td><td>taaatctgga</td><td>gccggtgagc</td><td>gtgggtctcg</td><td> 1800</td>

<td>cggtatcatt</td><td>gcagcautgg</td><td>ggccagatgg</td><td>taagccctcc</td><td>cgtatcgtag</td><td>ttatctacac</td><td> 1860</td>

<td>gacggggagt</td><td>caggcaacta</td><td>tggatgaacg</td><td>aaatagacag</td><td>atcgctgaga</td><td>. taggtgcctc</td><td> 1920</td>

<td>actgattaag</td><td>cattggtaac</td><td>tgtcagacca</td><td>agtttactca</td><td>tatatacttt</td><td>. agattgattt</td><td> 1980</td>

aaaacttcat ttttaattta aaaggatcta ggtgaaga-tc -ctttttgata atctcatgac 2040 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa 2100 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc 2160 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt 2220 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg 2280 ccaccacttc aagaactctg tagcaocgcc tacatacctc gctctgctaa tcctgttacc 2340 agtggctgct gccagtggcg ataagtegtg tcttaccggg ttggactcaa gacgatagtt 2400

142 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga <210> 63 <211> 6 <2X2> PRT <213> peptídeo adaptador <400> 63

Ηχε Ηίε His Hie His His

2460

2520

2580

2640

2700

2760

2820

2880

<td><210> 64 <211> 6 <212> PRT <213> peptídeo</td><td>adaptador</td>

<td><210> 65 <211> 6 <212> PRT <213> peptídeo</td><td>adaptador</td>

<td> <213></td><td>sítio de processamento</td>

<td> <221></td><td>MISCELÂNEA</td>

<td> <223></td><td>X é qualquer amínoácid</td>

143 <400> 66

Ile Glu Gly Arg Xaa 1 5

<td> <210> <212> <212> <213></td><td>67 5 PRT sítio de processamento</td>

<td> <220> <221> <222> <22 3></td><td>MISCELÂNEA (5) . . (5) X é qualquer aminoácido</td>

Ile Asp Gly Arg Xaa. 1 5

<td> <210> <211> <212> <213></td><td>68 5 PRT sítio de processamento</td>

<td> <220> <221> <222> <22 3></td><td>MISCELÂNEA (5) . . (5) X é qualquer aminoácido</td>

Ala Glu Gly Arg Xaa 1 s

<td> <210> <211> <212> <213></td><td>69 5 PRT peptídeo adaptador</td>

Vai Ile Glu Gly Arg

<td> <210> <211> <212> <213></td><td>70 4 PRT peptídeo adaptador</td>

Ile Glu Gly Arg 1

144 <210> 71 <211> 4 <212> PRT <213> peptídeo adaptador <400> 71

Ile Asp Gly Arg 1 <210> 72 <211> 4 <212> PRT <213> peptídeo adaptador <400> 72

Ala Glu Gly Arg <210> 73 <211> 4 <212> PRT <213> HCV El <400> 73

Aen Αεη Ser ser

<td> <213></td><td>epítopo de FLAG</td>

Aatp Tyr Lye Asp Asp Asp Asp Lya

<td> <213></td><td>epítopo da proteína C</td>

Glu Asp Gin Vai Asp Pro Arg Leu Ile Asp Gly Lys 1 S 10 <210> 76 <211> 11 <212> PRT <213> epítopo de VSV

145 <400> 76

Tyr Thr Asp Ile Glu Met Asn Arg Leu Gly Lys 15 10

<td> <210> <211> <212> <213></td><td>77 9 PRT streptag</td>

Ala Trp Arg His Pro Gin Phe Gly Gly 1 5

<td> <210> <211> <212> <213></td><td>78 12 PRT epítopo de TaglOO</td>

Glu Glu Thr Ala Arg Phe Gin Pro Gly Tyr Arg Ser

<td> <210> <211> <212> <213></td><td>79 10 PRT epítopo de c-nyc</td>

Glu Gin Lys Leu Ile Ser Glu Glu Asp Leu 1 5 10

<td> <210> <211> <212> <213></td><td>80 11 PRT epítopo de HA</td>

Tyr Pro Tyr Asp Vai Pro Asp Tyr Ala Ser Leu 15 10

<td> <210> <211> <212> <213></td><td>> 81 9 PRT epitope of HA</td>

Tyr Pro Tyr Aso Go to Asp Tyr Wing 1 5

146 <210> 82 <211> 1 <212> PRT <213> HA epitope <400> 82

Cys Tyr Pro Tyr Asp Vai Pro Asp Tyr Ala Ser Leu 15 10 <210> 83 <211> 6 <212> PRT <213> thrombin dividing site <400> 83

Leu Vai Pro Arg Gly Ser

S <210> 84 <211> 4 <212> PRT <213> collagenase recognition site <220>

<221> MISCELLANEOUS <222> (2) .. (2) <22 3> is any amino acid but more often a neutral amino acid <400> 84

Pro Xaa Gly Pro

<td> <213></td><td>virus from</td><td>hepatitis C</td><td></td><td></td>

Pro Asn Ser Ser Vai Vai Tyr Glu Ala Ala Asp Ala Ile Leu hib Thr 20 25 30

Pro Gly Cys Vai Pro Cys Vai Arg Glu Gly Asn Ala Ser Arg Cys Trp 3S 40 45

Go Ala Vai Thr Pro Thr Vai Ala Thr Arg Asp Gly Lys Leu Pro Thr 50 55 eo

147

Thr- Gin Leu Arg Arg His Ile Asp Leu Leu Vai Gly Ser Ala Thr Leu 65 70 75 80

Cys Ser Ala Leu Tyr Vai Gly Asp Leu Cys Gly Ser Vai Phe Leu Vai 85 90 95

<td colspan="3"><213> vírus</td><td>! da</td><td colspan="3">hepatite C</td><td></td><td></td><td></td><td></td>

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Phe Leu Leu Ala Leu 50 55 -60

Leu Ser Cys Leu Thr Vai Pro Ala Ser Ala Tyr Gin Vai Arg Asn Ser 65 70 75 80

148

Ser Gly Leu Tyr His Vai Thr Asn Asp Cye Pro Asn Ser Ser Vai Vai 65 90 95

Tyr Glu Ala Ala Asp Ala Ile Leu His Thr Pro Gly Cys Vai Pro Cye 100 105 110

Vai Arg Glu Gly Asn Ala Ser Arg Cys Trp Vai Ala Vai Thr Pro Thr 115 120 125

Vai Ala Thr Arg Asp Gly Lys Leu Pro Thr Thr Gin Leu Arg Arg His 130 135 140

Ile Asp Leu Leu Vai Gly Ser Ala Thr Leu Cys Ser Ala Leu Tyr Vai 145 ISO 155 160

Gly Asp Leu Cys Gly Ser Vai Phe Leu Vai Gly Gin Leu Phe Thr Phe 165 170 175

Ser Pro Arg His His Trp Thr Thr Gin Asp Cys Asn Cys Ser Ile Tyr 180 185 190

Pro Gly His Ile Thr Gly His Arg Met Ala Trp Asp Met Met Met Asn 195 200 205

Trp <210> 87 <211> 192 <212> PRT <213> vírus da hepatite C <400> 87

Tyr Glu Vai Arg Asn Vai Ser Gly Met Tyr His Vai Thr Asn Asp Cys 1 5 10 15

Ser Asn Ser Ser Ile vai Tyr Glu Ala Ala Asp Met Ile Met His Thr 20 25 30

Pro Gly Cys Vai Pro Cys Vai Arg Glu Asn Asn Ser Ser Arg Cys Trp 35 40 45

Vai Ala Leu Thr Pro Thr Leu Ala Ala Arg Asn Ala Ser Vai Pro Thr 50 55 60

Thr Thr Ile Arg Arg His Vai Asp Leu Leu Vai -Gly Ala Ala Ala Phe 65 70 75 80

149

Cys Ser Ala Met Tyr Vai Gly Asp Leu Cys Gly Ser Vai Phe Leu Vai 85 50 95

Ser Gin Leu Phe Thr Xle Ser Pro Arg Arg His Glu Thr Vai Gin Asp 100 105 110

Cys Asn Cys Ser Ile Tyr Pro Gly His Ile Thr Gly His Arg Met Ala 115 120 125

Trp Asp Met Met Met Asn Trp Ser Pro Thr Thr Ala Leu Vai Val Ser 130 135 140

<210> 88 <211> 209 <212> PRT

<td><213> vírua</td><td>í da</td><td>hepatite C</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td>

Ser Gly Met Tyr His Vai Thr Asn Asp Cys Ser Asn Ser Ser Ile Vai 85 90 95

150

Tyr Glu Ala Ala 100

Asp Met Ile Met His Thr Pro Gly Cys Vai Pro Cys 105 110

Vai Arg Glu Asn 115

Asn Ser Ser Arg Cys Txp Vai Ala Leu Thr Pro Thr 120 125

Leu Ala Ala Arg 130

Asn Ala Ser Vai Pro Thr Thr Thr lie Arg Arg His 135 140

Vai Asp Leu Leu 145

Vai Gly Ala Ala Ala Phe Cys Ser Ala Met Tyr Vai 150 155 160

Gly Asp Leu Cye

Gly Ser Vai Phe Leu Vai Ser Gin Leu Phe Thr Ile 165 170 175

Ser Pro Arg Arg 180

His Glu Thr Vai Gin Asp Cys Asn Cys Ser Ile Tyr 185 190

Pro Gly His Ile 195

Thr Gly His Arg Met Ala Trp Asp Met Met Met Asn 200 205

Trp

<td>209 PRT virus</td><td>da hepatite</td><td>C</td>

5 10 15

Met Gly Tyr Ile Pro Vai Vai Gly Ala Pro Vai Gly Gly vai Ala Arg 20 25 30

Ala Leu Ala His Gly vai Arg Vai Leu Glu Asp Gly Ile Asn Tyr Ala 35 40 45

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Phe Leu Leu Ala Leu 50 55 60

Leu Ser Cye Ile Thr Thr Pro Vai Ser Ala Vai Glu Vai Lys Asn Asn 65 70 75 80

Ser Asn Ser Tyr Met Ala Thr Asn Asp Cye Ser Asn Ser Ser ile Ile BS 90 95

151

Trp Gin Leu Glu Gly Ala Vai Leu His Thr Pro Gly Cys Vai Pro Cys 100 10S 110

Glu Leu Ala Asp Asn Thr Ser Ar? Cys Trp Vai Pro Vai Thr Pro Asn 115 120 125

Met Ala He Arg Gin Pro Gly Glu Leu Thr Lys Gly Leu Arg Ala His 130 135 140

Vai Asp Vai Ile Vai Met Ser Ala Thr Leu Cys Ser Ala Leu Tyr Vai 145 150 1S5 160

Gly Asp Vai Cys Gly Ala Leu Met lie Ala Ala Gin Vai Vai Vai Vai 165 170 175

Ser Pro Gin His His His Phe Vai Gin Glu Cys Asn cys Ser lie Tyr ΙβΟ 185 190

Pro Gly His Xle Thr Gly His Arg Met Ala Trp Asp Met Met Met Asn 195 200 205

Trp <210> 90 <211> 209 <212> PRT <213> virus da hepatite C <400> 90

Met Leu Gly Lys Vai Ile Asp Thr Leu Thr Cys Gly Phe Ala Asp Leu 15 10 15

Met Gly Tyr Ile Pro Vai Vai Gly Ala Pro Vai Gly Gly Vai Ala Arg 20 25 30

Ala Leu Ala His Gly vai Az? Vai Leu Glu Asp Gly Xle Asn Tyr Ala 35 40 45

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Phe Leu Leu Ala Leu 50 55 60

Leu Ser Cys Vai Thr Ala Pro Vai Ser Ala Vai Glu Vai Lys Asn Thr €5 70 75 80

152

Ser Gin Ala Tyr Met Ala Thr Asn Asp Cys Ser Açu Asn Ser Ile Vai SS 90 55

Trp Gin Leu Glu Asp Ala Vai Leu His Vai Pro Gly Cys Vai Pro Cys 100 105 110

Glu Asn Ser Ser Gly Arg Phe His Cys Trp Ile Pro Ile Ser Pro Asn 115 120 125

Ile .Ala Vai Ser Lys Pro Gly Ala Leu Thr Lys Gly Leu Arg Ala Arg 130 135 140

Ile Asp Ala Vai Vai Met Ser Ala Thr Leu Cys Ser Ala Leu Tyr Vai 145 ISO 155 160

Gly Asp Vai Cys Gly Ala Vai Met lie Ala Ala Gin Ala Phe Ile Vai 165 170 175

Ala Pro Lys Arg His Tyr Phe Vai Gla Glu Cys Asn Cys Ser Xle Tyr 180 185 190

Pro Gly His Ile Thr Gly His Arg Met Ala Trp Asp Met Met Met Asn 195 200 205

Trp

<td> <213></td><td>virus da hepatite C</td><td></td><td></td>

Met Gly Tyr Ile Pro Leu Vai Gly Ala Pro Vai Gly Gly Vai Ala Arg 20 25 30

Ala Leu Ala His Gly Vai Arg Ala Leu Glu Asp Gly Ile Asn Phe Ala 35 40 45

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Phe Leu Leu Ala Leu 50 SS €0

Phe Ser Cys Leu Ile Kis Pro Ala Ala Ser Leu Glu Trp Arg Asn Thr €5 70 75 80

153

Ser Gly Leu Tyr Vai Leu Thr Asn Asp Cys Ser Asn Ser Ser Ile Vai 85 90 95

Tyr Glu Ala Asp Asp Vai Ile Leu His Thr Pro Gly Cys Ile Pro Cys 100 105 110

Vai Gin Asp Gly Asn Thr Ser Thr Cys Trp Thr Pro Vai Thr Pro Thr 115 120 125

Vai Ala Vai Lys Tyr Vai Gly Ala Thr Thr Ala Ser tle Arg Ser His 130 135 140

Vai Asp Leu Leu Vai Gly Ala Ala Thr Mefe Cys Ser Ala Leu Tyr Vai 145 150 155 160

Gly Asp Met Cys Gly Ala Vai phe Leu Vai Gly Gin Ala Phe Thr Phe 165 170 175

Arg Pro Arg Arg His Gin Thr Vai Gin Thr Cys Asn Cys Ser Leu Tyr 180 185 190

Pro Gly His Leu Ser Gly His Arg Met Ala Trp Asp Met Met Met Asn 195 200 205

Trp <210> 92 <211> 209 <212> PRT <213> vírus da hepatite C <400> 92

Met Ser Gly Lys Vai Ile Asp Thr Leu Thr Cys Gly Phe Ala Asp Leu 1 S 10 15

Met Gly Tyr Ile Pro Leu Vai Gly Ala Pro Vai Gly Gly Vai Ala Arg 20 2S 30

Ala Leu Ala His Gly Vai Arg Ala Vai Glu Asp Gly Ile Asn Tyr Ala 35 40 45

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Phe Leu Leu Ala Leu 50 55 60

154

Leu Ser Cys Leu Thr Vai Pro Thr Ser Ala Vai Asn Tyr Arg Asn. Ala 65 70 75 80

Ser Gly V&l Tyr His Ile Thr Asn Asp Cys Pro Asn Ser Ser Ile Vai 85 90 95

Tyr Glu Thr Glu His His Ile Leu His Leu Pro Gly Cys Leu Pro Cys 100 105 110

Vai Arg Vai Gly Asn Gin Ser Arg Cys Trp Vai Ala Leu Thr Pro Thr 115 120 125

Vai Ala Ala Pro Tyr lie Gly Ala Pro Leu Glu Ser Leu Arg Ser His 130 135 140

Vai Asp Leu Met Vai Gly Ala Ala Thr Ala Cys Ser Ala Leu Tyr ile 145 150 155 160

Gly Asp Leu Cys Gly Gly Vai Phe Leu Vai Gly Gin Met Phe Ser Phe 165 170 175

Gin Pro Arg Arg His Trp Thr Thr Gin Asp Cys Asn Cys Ser Ile Tyr 180 185 190

Ala Gly His Vai Thr Gly His Arg Met Ala Trp Asp Met Met Met Asn 195 200 205

Trp <210> 93 <211> 209 <212> PRT <213> virus da hepatite C <400> 93

Met Leu Gly Lys Vai Ile Asp Thr Leu Thr Cys Gly Phe Ala Asp Leu 1 S 10 15

Met Gly Tyr Ile Pro Leu Vai Gly Gly Pro Ile Gly Gly Vai Ala Arg 20 25 30

Ala Leu Ala His Gly Vai Arg Vai Leu Glu Asp Gly Vai Asn Tyr Ala 35 40 45

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Phe Ile Leu Ala Leu 50 55 60

155

Leu Ser Cys Leu Thr Vai Pro Ala Ser Ala Vai Pro Tyr Arg Asn Ala €5 70 75 80

Ser Gly Ile Tyr His Vai Thr Asn Asp Cys Pro Asa Ser Ser Ile Vai 85 90 95

Tyr Glu Ala Asp Asn Leu lie Leu His Ala Pro Gly Cys Vai Pro Cys 100 105 110

Vai Met Thr Gly Asn Vai Ser Arg Cys Txp Vai Gin Ile Thr Pro Thr 115 120 125

Leu Ser Ala Pro Ser Leu Gly Ala Vai Thr Ala Pro Leu Arg Arg Ala 130 135 140

Vai Asp Tyr Leu Ala Gly Gly Ala Ala Leu cys Ser Ala Leu Tyr Vai 145 150 15S 160

Gly Asp Ala Cys Gly Ala Leu Phe Leu Vai Gly Gin Met Phe Thr Tyr 165 170 175

Arg Pro Arg Gin His Ala Thr Vai Gin Asn Cys Asn Cys Ser Ile Tyr 180 185 190

Ser Gly His Vai Thr Gly His Arg Met Ala Txp Asp Met Met Met Asn 195 200 205

Txp <210> 94 <211> 209 <212> PRT <213> vírus da hepatite C <400> 94

Met Leu Gly Lys Vai Ile Asp Thr Leu Thr Cys Gly Phe Ala Asp Leu 1 S 10 15

Met Gly Tyr Ile Pno Vai Vai Gly Ala Pro Leu Gly Gly Vai Ala Ala 20 25 30

Ala Phe Ala His Gly vai Arg Ala Leu Glu Asp Gly ile Asn Tyr Ala 35 40 45

156

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Phe Leu Leu Ala Leu 50 55 60

Leu Ser Cys Leu Thr Thr Pro Ala Ser Ala Leu Thr Tyr -Gly Asn Ser 55 70 75 80

Ser Gly Leu Tyr His Leu Thr Asn Asp Cys Pro Asn Ser Ser Ile Vai 85 90 95

Leu Glu Ala Asp Ala Met Ile Leu His Leu Pro Gly Cys Leu Pro Cys 100 105 110

Vai Arg Vai Asn Asn Gin Ser Thr Cys Trp His Ala Vai Ser Pro Thr 115 120 125

Leu Ala Ile Pro Asn Ala Ser Thr Pro Ala Thr Gly Phe Arg Arg His 130 135 140

Vai Asp Leu Leu Ala Gly Ala Ala Vai Vai Cys Ser Ser Leu Tyr Ile 14S 150 155 160

Gly Asp Leu Cys Gly Ser Leu Phe Leu Ala Gly Gin Leu Phe Thr Phe 155 170 175

Gin Pro Arg Arg His Trp Thr Vai Gin Asp Cys Asn Cys Ser Ile Tyr 180 185 190

Thr Gly His Vai Thr Gly His Arg Met Ala Trp Asp Met Met Met Asn 195 200 205

Trp <210> 95 <211> 209 <212> PRT <213> vírua da hepatite C <400> 95

Met Leu Gly Lys Vai Ile Asp Thr Leu Thr Cys Gly Phe Ala Asp Leu 15 10 15

Met Gly Tyr Ile Pro Vai Vai Gly Ala Pro Leu Gly Gly Ile Ala Ala 20 25 30

Ala Leu Ala His Gly Vai Arg Ala Vai Glu Asp Gly Ile Asn Tyr Ala 35 40 45

157

Thr Gly Asa Leu Pro Gly Cys Ser Phe Ser Ile Phe Leu Leu Ala Leu 50 55 60

Leu Ser Cys Leu Thr Thr Pro Ala Ser Ala Vai His Tyr Ala Asn Lys 6S 70 75 80

Ser Gly Leu Tyr His Leu Thr Asn Asp Cys Pro Asa Ser Ser Ile Vai 85 50 95

Tyr Glu Ala Pro Ala Vai Ile Met His Leu Pro Gly Cys Vai Pro Cys 100 105 110

Vai Lys Vai Gly Asa Gla Ser Thr Cys Trp Leu Pro Ala Ser Pro Thr 115 120 125

Leu Ala Vai Pro Asn Ala Ser Thr Pro Leu Thr Arg Phe Arg Lys His 110 115 140

Vai Asp Leu Met Vai Gly Ala Ala Ala Phe Cys Ser Ala Met Tyr Vai 145 ISO 155 160

Gly Asp Ile Cys Gly Gly Leu Phe Leu Leu Gly Gin Vai Vai Thr Ile 165 170 175

Arg Pro Arg Leu His Gla Thr Vai Gin Glu Cys Asn Cys Ser Ile Tyr 180 185 190

Thr Gly Lys Ile Thr Gly His Arg Met Ala Trp Asp Ile Met Met Asn 195 200 205

Trp <210> 96 <211> 209 <212> PRT <213> virua da hepatite C <400> 96

Met Leu Gly Lys Vai Ile Asp Thr Leu Thr Cys Gly Leu Ala Asp Leu 15 10 IS

Met Gly Tyr Ile Pro Vai Leu Gly Gly Pro Leu Gly Gly Vai Ala Ala 20 25 30

158

Ala Leu Ala Hie Gly Vai Arg Ala Ile Glu Asp Gly Vai Asn Tyr Ala 35 40 45

Thr Gly Asn Leu Pro Gly Cys Ser Phe Ser Ile Leu Leu Leu Ala Leu 50 55 60

Leu Ser Cys Leu Thr Ile Pro Ala ser Ala Ile Gin Vai Lys Asn Ala 65 70 75 80

Ser Gly Ile Tyr Hie Leu Thr Asn Asp Cys Ser Asn Asn Ser Ile Vai 85 90 95

Phe Glu Ala Glu Thr Met Ile Leu His Leu Pro Gly Cys Vai Pro Cye 100 105 110

Ile Lye Ala Gly Asn Glu Ser Arg Cys Trp Leu Pro Vai Ser pro Thr 115 120 125

Leu Ala Vai Pro Asn Ser Ser Vai Pro Ile His Gly Phe Arg Arg His 130 135 140

Vai Asp Leu Leu Vai Gly Ala Ala Ala Phe Cys Ser Ala Met Tyr Ile 145 150 1S5 160

Gly Asp Leu Cys Gly Ser Ile Phe Leu Vai Gly Gin Leu Phe Thr Phe 165 170 175

Arg Pro Lys Tyr Hie Gin Vai Thr Gin Asp Cys Asn Cys Ser Ile Tyr 180 185 190

Ala Gly His Ile Thr Gly His Arg Met Ala Trp Asp Met Met Met Asn 195 200 20S

Trp <210> 97 <211> 363 <212> PRT <213> vírus da hepatite C <400> 97

Glu Thr His Vai Thr Gly Gly Asa Ala Gly Arg Thr Thr Ala Gly Pro 15 10 15

Vai Gly Leu Leu Thr Pro Gly Ala Lys Gin Asn Ile Gin Leu Ile Asn 20 25 30

159

Thr Asn

Gly Ser Trp His Ile Asn Ser Thr Ala Leu Asn Cys Asn Glu 35 40 45

Met Vai 225

Ser Leu 50

Asn Ser 65

Asp Phe

Leu Asp

Ile Vai

Ser Pro 130

Ser Trp 145

Pro Pro

Phe Thr

Asn Asn

Ala Thr 210

230 235 240

Tyr Thr

Ile Phe Lys Vai Arg Met Tyr Vai Gly Gly Vai Glu His Arg 245 250 2S5

Leu Glu Ala Ala -Cys Asn Trp Thr Arg Gly Glu Arg Cys Asp Leu Glu 260 265 270

160

Asp Arg Asp Arg Ser Glu Leu Ser Pro Leu Leu Leu Ser Thr Thr Gin 275 280 285

Trp Gin Vai Leu Pro Cys Ser Phe Thr Thr Leu Pro Ala Leu Ser Thr 290 295 300

Gly Leu ile Ηίε Leu Hie Gin Asn Ile Vai Asp Vai Gin Tyr Leu Tyr 305 310 315 320

Gly Vai Gly Ser Ser lie Ala Ser Trp Ala Ile Lys Trp Glu Tyr Vai 325 330 33S

Vai Leu Leu Phe Leu Leu Leu Ala Asp Ala Arg Vai Cys Ser Glu Phe 340 345 350

Trp Met Met Leu Leu Ile Ser Gin Ala Glu Ala 360

<td> <213></td><td>vírus da</td><td>hepatite C</td>

Vai Ser Leu Phe Ser Pro Gly Ser Ala Glu Lys Ile Gin Leu Vai Asn 20 25 30

Thr Asn Gly Ser Trp His Ile Asn Arg Thr Ala Leu Asn Cys Asn Asp 35 40 45

Ser Leu Gin Thr Gly Phe Phe Ala Ala Leu Phe Tyr Lys His Lys Phe 50 55 60

Asn Ser Ser Gly -Cys Pro Glu Arg Leu Ala ser Cys Arg Ser Ile Asp 65 70 75 80

Lys Phe Ala Gin Gly Trp Gly Pro Leu Thr Tyr Thr Glu Pro Asn Ser 85 50 55

Ser Asp Gin Arg Pro Tyr Cys Trp His Tyr Ala Pro Arg Pro Cys Gly 100 105 HO

Ile Vai Pro Ala Ser Gin Vai Cys Gly Pro Vai Tyr Cys Phe Thr Pro

161

115 120 125

Ser Pro Vai Vai Vai Gly Thr Thr Asp Arg Phe Gly Vai Pro Thr Tyr 130 135 140

Asn Trp Gly Ala Αεη Αερ Ser Asp Vai Leu Ile Leu Asn Asn Thr Arg 145 150 1S5 160 pro Pro Arg Gly Asn Trp Phe Gly Cys Thr Trp «et Asn Gly Thr Gly 165 170 175

Phe Thr Lys Thr Cys Gly Gly Pro Pro Cys Asn Ile Gly Gly Ala Gly 160 185 190

Asn Asn Thr Leu Thr Cys Pro Thr Asp Cys Phe Arg Lys His Pro Glu 195 200 205

Ala Thr Tyr Ala Arg Cys Gly Ser Gly Pro Trp Leu Thr Pro Arg Cye 210 215 220

Met Vai His Tyr Pro Tyr Arg Leu Trp His Tyr Pro Cys Thr Vai Asn 225 230 235 240

Phe Thr Ile Phe Lys Vai Arg Met Tyr Vai Gly Gly Vai Glu His Arg 245 250 2SS

Phe Glu Ala Ala Cys Asn Trp Thr Arg Gly Glu Arg Cys Asp Leu Glu 260 265 270

Asp Arg Asp Arg Ser Glu Leu Ser Pro Leu Leu Leu Ser Thr Thr Glu 275 280 285

Trp Gin Ile Leu Pro Cys Ser Phe Thr Thr Leu Pro Ala Leu Ser Thr 290 295 300

Gly Leu Ile His Leu His Gin Asn Ile Vai Asp Vai Gin Tyr Leu Tyr 305 310 315 320

Gly Vai Gly Ser Ala Vai Vai Ser Leu Vai Ile Lys Trp Glu Tyr Vai 325 330 335

Leu Leu Leu Phe Leu Leu Leu Ala Asp Ala Arg Ile Cys Ala Cys Leu 340 345 350

Trp Met Met Leu Leu Ile Ala Gin Ala Glu Ala 355 360 ·»··

Contents66

55 members in 24 offices

This recordMember, by publication datePriority claim14 more offices are in the list below

Priority claims12

Priority claims
Document	Office	Kind	Date
01870088	European Patent Office (EPO)	A	2001-04-24
01870088	European Patent Office (EPO)	A	2001-04-24
30560401	United States of America	P	2001-07-17
30560401	United States of America	P	2001-07-17
0200062	Belgium	W	2002-04-24
0200062	Belgium	W	2002-04-24
018700880	–	–	–
0200062	–	–	–
60305604	–	–	–
EP20010870088	–	–	–
US20010305604P	–	–	–
WO2002BE00062	–	–	–

Members55

Family members, oldest first
Document	Office	Kind
CA2443740A1	Canada	A1	2002-10-31
CA2443781A1	Canada	A1	2002-10-31
CA2444006A1	Canada	A1	2002-10-31
WO02085932A2	World Intellectual Property Organization (WIPO)	A2	2002-10-31
WO02086100A2	World Intellectual Property Organization (WIPO)	A2	2002-10-31
WO02086101A2	World Intellectual Property Organization (WIPO)	A2	2002-10-31
AU2002252856A1	Australia	A1	2002-11-05
WO02085932A3	World Intellectual Property Organization (WIPO)	A3	2003-03-13
US2003108561A1	United States of America	A1	2003-06-12
US2003152940A1	United States of America	A1	2003-08-14
US2003211597A1	United States of America	A1	2003-11-13
KR20030094359A	Republic of Korea	A	2003-12-11
AP2003002886A0	African Regional Intellectual Property Organization (ARIPO)	A0	2003-12-31
EP1381671A2	European Patent Office (EPO)	A2	2004-01-21
WO02086100A3	World Intellectual Property Organization (WIPO)	A3	2004-02-19
WO02086101A3	World Intellectual Property Organization (WIPO)	A3	2004-02-19
HU0303924A2	Hungary	A2	2004-03-01
HUP0303924A2	Hungary	A2	2004-03-01
EP1414942A2	European Patent Office (EPO)	A2	2004-05-06
EP1417298A2	European Patent Office (EPO)	A2	2004-05-12
IL158422A0	Israel	A0	2004-05-12
NZ529019A	New Zealand	A	2004-05-28
MXPA03009626A	Mexico	A	2004-06-30
MXPA03009632A	Mexico	A	2004-06-30
AR035867A1	Argentina	A1	2004-07-21
AR035868A1	Argentina	A1	2004-07-21
AR035869A1	Argentina	A1	2004-07-21
CZ20032853A3	Czechia	A3	2004-08-18
ZA200308277B	South Africa	B	2004-09-29
JP2004532029A	Japan	A	2004-10-21
SK13142003A3	Slovakia	A3	2004-11-03
JP2004536052A	Japan	A	2004-12-02
JP2004536582A	Japan	A	2004-12-09
BG108373A	Bulgaria	A	2004-12-30
BR0209033AThis record	Brazil	A	2005-01-11
BR0209034A	Brazil	A	2005-02-01
PL366621A1	Poland	A1	2005-02-07
ZA200308272B	South Africa	B	2005-03-30
ZA200308274B	South Africa	B	2005-03-30
RU2003130955A	Russian Federation	A	2005-04-20
CN1636050A	China	A	2005-07-06
NZ529324A	New Zealand	A	2005-08-26
HRP20030946A2	Croatia	A2	2005-10-31
RU2274643C2	Russian Federation	C2	2006-04-20
US7048930B2	United States of America	B2	2006-05-23
CO5650176A2	Colombia	A2	2006-06-30
OA13092A	African Intellectual Property Organization (OAPI)	A	2006-11-10
AU2002257392B2	Australia	B2	2007-05-10
US7238356B2	United States of America	B2	2007-07-03
US7314925B2	United States of America	B2	2008-01-01
AU2002308449B2	Australia	B2	2008-05-29
US2008124763A1	United States of America	A1	2008-05-29
JP4173741B2	Japan	B2	2008-10-29
JP4261195B2	Japan	B2	2009-04-30
KR100950104B1	Republic of Korea	B1	2010-03-30

2 legal events, as the office reported them to INPADOC

Over the term

Point at a mark for the event

Events

Legal events, newest first
	Event	Code
2011-11-16	Patent lapsed as no evidence of payment of the annual fee has been furnished to inpi [chapter 8.11 patent gazette]LapsedREFERENTE AO DESPACHO 8.6 PUBLICADO NA RPI 2103 DE 26/04/2011.B08K	B08K
2011-04-26	Application dismissed because of non-payment of annual fees [chapter 8.6 patent gazette]REFERENTE A(S) 8A E 9A ANUIDADE(S).B08F	B08F

Abstract

Term

CONSTRUCTIONS, AND METHODS 'FOR THE EXPRESSION OF RECOMBINANT HCV WRAP PROTEINS

FIELD OF. INVENTION

BACKGROUND GIVES INVENTION

SUMMARY OF THE INVENTION

PICTURE'S DESCRIPTION

EXAMPLES

EXAMPLE 1

CHHE1-F:

CHHE1-R:

EXAMPLE 2

EXAMPLE 3

EXAMPLE 4

EXAMPLE 5

EXAMPLE 6

TRANSFORMATION OF HANSENULA POLYMORPHA AND SELECTION OF

TRANSFORMANTS

EXAMPLE 7

EXAMPLE 8

EXAMPLE 9

EXAMPLE 10

TRANSFORMATION OF SACCHAROMYCES CEREVISIAE AND SELECTION OF

TRANSFORMANTS

EXAMPLE 11

TCGAGAAAAGGGGCCCGAATTCGCATGC (SEQ ID NO: 54)

NO: 56)

650: '5'-GGCCGCATATGCTGCAGGCTTCAGCCTCTCTTTTC-3' (SEQ ID

TCGAGAAAAGAGAGGCTGAAGCCTGCAGCATATGC (SEQ ID NO: 56)

CTTTTCTCTCCGACTTCGGACGTCGTATACGCCGG (SEQ ID NO: 57)

EXAMPLE 12

EXAMPLE 13

PICHIA PASTORIS TRANSFORMATION AND TRANSFORMANT SELECTION

EXAMPLE 14

CULTURE CONDITIONS FOR SACCHAROMYCES CEREVISIAE,

HANSENULA POLYMORPHA AND PI CHIA PASTORIS

EXAMPLE 15

EXAMPLE 17

PURIFICATION AND BIOCHEMICAL CHARACTERIZATION OF PROTEIN E2 FROM

HCV EXPRESSED IN HANSENULA POLYMORPHA FROM THE

EXAMPLE 18

HCV EXPRESSED IN HANSENULA POLYMORPHA FROM THE

CONSTRUCTION OF CL-H6-K-E1 CODING AND IN PROCESSING

HIT-CONTAINING PROTEIN VITRO

EXAMPLE 19

SPECIFIC REMOVAL OF LOW GLYCOSILATED FORMS OF HCV

BY HEPARINA

EXAMPLE. 20

EXAMPLE 21

ANTIGENIC EQUIVALENCE OF HC1 E1-H.6 PRODUCED BY

HANSENULA AND HCV PRODUCED 'BY MAMMALIAN CELLS

INFECTED WITH VACCINIA

EXAMPLE 22

IMMUNOGENIC EQUIVALENCE OF HC1 E1-H6 PRODUCED BY

HANSENULA AND HCV ELECTED BY VACCINIA-INFECTED MAMMALIAN CELLS

EXAMPLE 23

ANTIGENIC AND IMMUNOGENIC PROFILE OF HC1 E1-H6 PRODUCED

EXAMPLE 24

HCV E1-H6 IDENTIC ANTIGENIC REACTIVIPAPE PRODUCED

BY HANSENULA AND EL HCV PRODUCED BY MAMMALIAN CELLS

INFECTED WITH VACCINATION WITH SERUM, FROM VACCINATED CHIMPANZES '5

EXAMPLE 25

HCV EL GLYCOPERFIL BY CARBOHYDRATE ELECTROPHORESIS

AIDED BY FLUOROPHORE (FACE)

I

I

I

46B0

Priority claims12

Members55

Over the term

Events

Numbers

Titles2

Classification