Modified oligonucleotides for mismatch discrimination

A modified oligonucleotide comprising at least one base of the base of 5-substituted pyrimidine and at least one base pyrazolo [3,4-d] pyrimidine unsubstituted or substituted 3-, wherein said at least one pyrimidine base 5 -sustituida has a formula consisting of: wherein at least one base pyrazolo [3,4-d] pyrimidine unsubstituted or substituted 3-is consisting in which each of the groups X1, X2 and X3 is a member independently selected from the group consisting of H, OH, NH2 and a protected amino group; and each of said R1 and R4 groups is a member independently selected from the group consisting of heteroalkyl (C1-C12) heteroalkenyl (C2-C12), heteroalkynyl (C2-C12), -O- (C1-C12), -O- (C2-C12 alkenyl), -O- (C2-C12 alkynyl), -S- (C1-C12), -S- (C2-C12 alkenyl), -S- (C2-C12 alkynyl), heterocyclyl (C1-C12) heterocyclyl (C2-C12) heterocyclyl (C2-C12), aryl (C1-C12), aryl (C2-C12), aryl (alkynyl C2- C12) aryl, heterocyclyl, halogen, -CN, -CONH2 and protected forms thereof,