Automated contamination-free seed sampler and method of sampling

p00001Automated seed sampler free of contamination and sampling procedure

p00002Countryside

p00003The present disclosure is about systems and procedures for sampling biological materials such as seeds.

p00004Background

p00005The statements in this section simply provide background information related to the present disclosure and may not constitute the prior art.

p00006In the development and improvement of plants, genetic improvements are made in the plant, either by selective reproduction or by genetic manipulation, and when a desirable improvement is achieved, a commercial amount is developed by planting and harvesting seeds for several generations. Not all seeds express the desired traits and, therefore, these seeds need to be discarded from the population. To accelerate the population collection process, statistical samples are taken and tested to discard seeds that do not adequately express the desired trait. However, this statistical sampling necessarily allows some seeds without the desirable trait to remain in the population, and may also involuntarily exclude some seeds with the desirable trait of the desired population.

p00007US patent application with serial number 11 / 213,430 (filed on August 26, 2005); patent application

p00008US with serial number 11 / 213,431 (filed August 26, 2005); US Patent Application Serial No. 11 / 213,432 (filed August 26, 2005); US Patent Application Serial No. 11 / 213,434 (filed August 26, 2005); and the US patent application Serial No. 11 / 213,435 (filed on August 26, 2005), discloses apparatus and systems for automated seed sampling as well as procedures for sampling, testing and seed collection. EP 1 786 261 discloses a seed sample with the characteristics of the preamble of claim 1.

p00009However, at least some known automated sampling and testing systems allow various types of contamination to contaminate collected samples and distort the results. Therefore, there is a need for automated sampling of seeds in a manner substantially free of contamination.

p00010Summary

p00011This problem is solved with an apparatus according to claim 1 and a method according to claim 9. The present disclosure relates to systems and procedures for non-destructive sampling of seed material. The procedures are adapted in particular for automation, which allows greater sampling than was previously practical. With an automated non-destructive sampling allowed by at least some of the embodiments of the present disclosure, it is possible to test each seed in the population, and discard those seeds that do not express a desired trait. This greatly speeds up the process of collecting a given population of seeds, and can result in an improved final population.

p00012Various embodiments of the present disclosure facilitate the testing of most of the seeds, or all of them, in a population before being planted, so that time and resources are not wasted growing plants without the desired traits. In addition, various embodiments allow automated sampling of seeds in a contamination-free manner, thereby substantially eliminating a cross between samples.

p00013The systems and procedures of the present disclosure facilitate automated non-destructive sampling in a manner substantially free of contamination. They allow the testing and classification of large volumes of seeds, thereby facilitating the collection of seed populations with desirable traits. These and other features and advantages will be apparent in part, and partly noted below.

p00014Additional areas of applicability of the present teachings will be apparent from the description provided herein. It should be understood that the description and specific examples are intended for illustrative purposes only and are not intended to limit the scope of the present teachings.

p00015Drawings

p00016The drawings described herein are for illustrative purposes only and are not intended to limit the scope of the present teachings in any way.

p00018Figure 1 is a perspective view of a seed sampling system according to various embodiments of the present disclosure.

p00019Figure 2 is an enlarged perspective view of a seed loading station of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00020Figure 3 is an enlarged perspective view of a seed orientation system of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00021Figure 4 is a side elevational view of the seed orientation system shown in Figure 3, according to various embodiments of the present disclosure.

p00022Figure 5 is a perspective view of the seed orientation system shown in Figure 3 that includes a seed holder, according to various embodiments of the present disclosure.

p00023Figure 6 is an enlarged perspective view of the seed support shown in Figure 5, according to various embodiments of the present disclosure.

p00024Figure 7 is an enlarged side view of the seed support shown in Figure 6, according to various embodiments of the present disclosure.

p00025Figure 8 is a perspective view of a grinding station and a seed transport subsystem of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00026Figure 9 is a perspective view of a sampling station of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00027Figure 10 is an enlarged side view of the seed sampling station, shown in Figure 9, during the operation of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00028Figure 11 is a side elevational view of a liquid supply apparatus of the seed sampling system, shown in Figure 1, in a retracted position, according to various embodiments of the present disclosure.

p00029Figure 12 is a side elevational view of the liquid supply apparatus shown in Figure 11, in an extended position, according to various embodiments of the present disclosure.

p00030Figure 13 is a perspective view of a sample tray platform of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00031Figure 14 is a perspective view of a seed treatment station of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00032Figure 15 is a side elevational view of a seed conveyor of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00033Figure 16 is a perspective view of a seed tray platform of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00034Figure 17 is a side elevation view of a collection tube loading station of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00035Figure 18 is a perspective view of a preparation subsystem of the collection tube of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00036Figure 19 is a perspective view of a cleaning station of the seed sampling system shown in Figure 1, according to various embodiments of the present disclosure.

p00037The corresponding reference numbers indicate corresponding parts in all the various views of the drawings.

p00038Detailed description

p00039The following description is simply exemplary in nature and is not intended to limit the present teachings, their application, or their uses in any way. Similar reference numbers will be used throughout this report to refer to similar elements.

p00040Figure 1 illustrates an automated seed sampling system 10, according to various embodiments of the present disclosure. In general, the seed sampler system 10 includes a seed loading station 100, a seed orientation system 200, a seed transport subsystem 300, a grinding station 400, a sampling station 500, a subsystem 600 for collecting and transporting seeds, a liquid supply subsystem 700, a sample storage subsystem 800, a seed treatment station 900 and a 1000 seed storage subsystem.

p00041The seed sampler system 10 is structured and operable to isolate a seed from a seed container 104 of the seed loading station 100, orient the seed in the seed orientation station 200 and transfer the seed to the station 400 of grinding, by means of the transport subsystem 300. In addition, the seed sampler system 10 is structured and operable to remove a portion of the seed coating material at the grinding station 400, transfer the seed to the sampling station 500, by means of the transport subsystem 300 of seeds, in which the sample material is extracted from the seed in the place where the seed coating material has been removed. In addition, the seed sampler system 10 is also structured and operable to transport the extracted sample to the sample storage subsystem 800, by means of the sample transport subsystem 700, and deposit the extracted sample in a sample tray 14 located in the 800 sample deposit subsystem. In various embodiments, the sample material is collected in a disposable sample tube and is supplied to the sample tray 14 using liquid, as described further below. In addition, the seed sampler system 10 is also structured and operable to treat the exposed portion, for example, apply a protective coating to it, of the seed in the seed treatment station 900 and transport the seed to the subsystem 1000 of seed deposit, in which the seed is deposited in a seed tray 18 located on a platform of the seed storage subsystem 1000.

p00042It should be understood that the seed sampling system 10, as shown and described herein, includes various braces, beams, platforms, pedestals, etc. stationary to which various components, devices, mechanisms, systems, subsystems, assemblies and sub-assemblies described herein are attached, connected and / or mounted. Although such braces, beams, platforms, pedestals, feet, etc. are necessary. For the construction of the seed sampling system 10, it is not necessary to describe its placement, orientation and interconnections for a person skilled in the art to easily and completely understand the structure, function and operation of the seed sampling system 10. In particular, such braces, beams, platforms, pedestals, feet, etc. they are clearly illustrated in all the figures and, as such, their placement, orientation and interconnections can be understood by one skilled in the art. Therefore, for the sake of simplicity, such braces, beams, platforms, pedestals, feet, etc. will be referred to herein. simply as support structures of the system, missing an additional description of its placement, orientation and interconnections.

p00043Referring now to Figures 2 and 3, in various embodiments, the seed loading station includes the seed container 104 and a separation wheel 108. The separation wheel 108 is mounted to rotate in a vertical plane, so that a portion of the separation wheel 108 extends into an interior reservoir of the seed container 104. Another portion of the separation wheel 108 extends out of the seed container 104, so that one side 120 of the separation wheel 108 is positioned adjacent to a seed picker 124. The seed separation wheel 108 includes a plurality of separate recessed holes 128 that extend through face 120 and are coupled with communication to a vacuum system (not shown), so that a vacuum is provided in each of the recessed holes 128.

p00044To initiate the operation of the seed sampling system 10, seeds to be sampled and tested are placed in the inner tank of the seed container 104 and a vacuum is provided to at least some of the recessed holes 128, for example recessed holes 128 on face 120 of the portion of the separation wheel 108 extending into the interior of the inner container of the seed container 104. Then, the seed separation wheel 108 is rotated incrementally, by means of an indexing motor 132, so that the recessed holes 128 rotate sequentially through the inner tank of the seed container 104, outside the container 104 of seeds, and by the seed collector 124 before re-entering the inner tank of the seed container 104. As the separation wheel rotates incrementally and the recessed holes 128 pass incrementally through the inner reservoir of the seed container 104, individual seeds are collected and held in each of the recessed holes 128 by means of the vacuum provided in the respective recessed holes 128. As the separation wheel 108 rotates incrementally, the seeds are transported out of the seed container 104 to the seed collector 124 in which each seed is removed from the face 120 of the separation wheel 108. After each seed is removed from the separation wheel 108, the seed is channeled to a transfer tube 136 of the loading station. Then, the seed passes through the transfer tube 136 of the loading station, by means of gravity, vacuum or pressurized air, into a seed imaging device 204 of the seed orientation system 200 . The transfer tube 136 of the charging station is sized to have an internal diameter that will only allow the seed to pass through the transfer tube 136 of the charging station in a longitudinal orientation. That is, the seed can only pass through the transfer tube 136 of the loading station well

p00045in an orientation with the tip up or with the tip down and the internal diameter will not allow the seed to roll or turn as it passes through the transfer tube 136 of the loading station.

p00046In various embodiments, the seed picker 124 includes a brush (not shown) that physically displaces each seed from the respective recessed hole 128 as the separation wheel 108 rotates incrementally through the seed picker 124. Thereafter, the displaced seed passes through the transfer tube 136 of the loading station to the imaging device 204. Alternatively, in various other embodiments, each seed can be released from the respective recessed hole 128 when the vacuum is temporarily terminated in each individual recessed hole 128 when the individual recessed hole 128 is positioned adjacent to the seed collector 124. Thereafter, the displaced seed is transferred to the imaging device 204, by means of the transfer tube 136 of the loading station. In yet other embodiments, each seed can be blown from the respective recessed hole 128 by temporarily providing pressurized air in each individual recessed hole 128, since the individual recessed hole 128 is positioned adjacent to the seed collector 124. Thereafter, the displaced seed is transferred to the imaging device 204, by means of the transfer tube 136 of the loading station.

p00047In addition, in various embodiments the seed loading station 100 may include a bulk seed hopper 140 having a formed surface and a vibrating feeding mechanism 144. Large quantities of seeds can be placed in hopper 140 in which the seeds are channeled on the vibrating feeding mechanism 144. The vibrating feeding mechanism 144 for inserting seeds into the seed container 104 into which the seeds are separated and transferred to the imaging device 204 of the seed orientation system 200 can be controlled, as described above.

p00048Referring now to Figures 3 and 4, the seed orientation system 200 comprises the seed imaging device 204, an image forming device 208, and a seed orientation device 212 mounted on a central platform stationary 214 of the seed sampling system 10. The seed imaging device 204 includes a window 216 and an internal seed orientation area that is visible through the window 216. The orientation device 212 includes a tiltable actuator 220 operable to rotate the seed while that the seed is suspended in the area of ​​orientation of the seeds. The seed imaging device 204 is connected to one end of the transfer tube 136 of the loading station and the imaging device 208 is mounted on a support structure of the system adjacent to the imaging device, so that the imaging device 208 is positioned to see a seed suspended in the seed orientation area through the window 216.

p00049When a seed is transferred to the imaging device 204, by means of the transfer tube 136 of the loading station, the seed is suspended in the seed orientation area, adjacent to the window 216, and is seen by the Image forming device 208 through window 216. In various different embodiments, the seed is levitated in the seed orientation area using air provided through a transfer tube 224 of the orientation system connected to the bottom of the imaging device 204, in front of the tube 136 of transfer of the charging station. Or, in various embodiments, the seed can be physically maintained in the seed orientation area using any suitable mechanical means.

p00050As the seed adjacent to the window 216 is suspended, an image of the seed is captured within the imaging device 204 by means of the imaging device 208. The imaging device 208 may be any image forming device suitable for capturing images through the window 216 of the seeds suspended in the seed orientation area. For example, in various embodiments, the imaging device 208 comprises a high-speed high resolution digital camera, such as a mechanical vision camera of disruptive visual technology (DVT). The image is communicated to a computer-based system controller (not shown), in which a seed orientation is determined, that is, with the tip up or the tip down. In various embodiments, the seed imaging device 208 further locates a seed centroid and identifies the furthest point from the centroid as the tip.

p00051If it is determined that the seed is with the tip down, the seed is transported in the orientation with the tip down, by means of the transfer tube 224 of the orientation system, to one of a plurality of seed holders 304. If it is determined that the seed is with the tip up, the system controller instructs the tilting actuator 220 to rotate the seed 180 ° to place the seed in the orientation with the tip down. For example, the tilting actuator 220 can be operated pneumatically, so that air is used to rotate the seed until the orientation is detected with the tip down by means of the imaging device 208. Or, the tilting actuator can be a mechanical actuator that rotates the clamped seed by means of a mechanical clamping device suitable for placing the seed in the orientation with the tip down. Once in the orientation with the tip down, the seed is transported in the orientation with the tip down, by means of the transfer tube 224 of the orientation system, to one of the seed holders 304. The orientation of the seeds in the position with the tip down minimizes the

p00052impact on the viability of the seed when a sample of the seed is extracted, as described below. In various embodiments, the seeds are transported by means of the transfer tube 224 of the orientation system using gravity, that is, the seeds fall from the imaging device 204, through the transfer tube 224 and into one of the supports 304 of the seeds. In addition, each seed is maintained in the proper orientation, that is, with the tip down, during transport to the respective seed holder 304 by providing the transfer tube 224 of the orientation system with an internal diameter sized so that the seeds cannot turn to the position with the tip up.

p00053As used herein, the system controller may be a system based on a single computer, or a plurality of subsystems interconnected to each other in a network to coordinate the simultaneous operations of the seed sampler system 10, described herein. For example, the system controller may include a plurality of controller subsystems, for example, a controller subsystem for each station described herein. Each controller subsystem could include one or more processors or microprocessors that communicate with various sensors, devices, mechanisms, motors, tools, etc. of the seed sampling system, and are connected together in a network with a main computer system to cooperatively operate all the stations, systems and subsystems of the seed sampling system 10. Or alternatively, the system controller could comprise a single computer communicatively connected to all the various sensors, devices, mechanisms, motors, tools, etc., to cooperatively operate all stations, systems and subsystems of the system 10 seed sampler.

p00054The seed holders 304 are mounted on a motorized rotating platform 308, and are uniformly separated around a perimeter area thereof, from the seed transport subsystem 300. The transfer tube 224 of the orientation system is connected at a first end to the seed imaging device 204, so that a second end of the transfer tube 224 of the orientation system is placed at a specific distance above a perimeter portion of the turntable 308. More particularly, the second end of the transfer tube 224 of the orientation system is positioned above the turntable 308 sufficient distance to allow the seed holders 304 to pass under the second end of the transfer tube of the transfer system. orientation. However, the second end of the transfer tube 224 of the orientation system is also positioned above the turntable 308, so that there is only a small amount of free space between the second end and the supports 304. Therefore, each seed will remain in the orientation with the tip down as it passes from the transfer tube 224 of the orientation system to one of the seed holders 304.

p00055Referring now to Figures 5, 6 and 7, each seed holder 304 is structured and is used to rigidly retain a respective seed in the orientation with the tip facing down. Each seed holder 304 includes a pair of opposing fixing heads 312 slidably positioned within opposite fixing receptacles 316. Opposite fixing receptacles 316 are separated by a side channel 318 formed laterally along a center line C of the seed holder 304. Each fixing head 312 is connected to a respective fixing piston 320 by means of a respective fixing shaft 324. Each fixing piston 320 is slidably housed within a respective longitudinal internal piston cylinder 328 of the seed holder 304. A compression spring 332 is placed inside each piston cylinder 328 between a respective piston base and a lower part of the respective piston cylinder 328. Accordingly, each fixing head 312 is pushed towards the center line C of the support 304 seeds When a seed support 304 is in an inactive state, that is, when the respective seed support is not holding a seed or being manipulated to hold a seed, the opposite fixing heads 312 will be pushed by the springs 332 to a fully extended position, or deployed. When the fixing heads 312 are in the deployed position, an upper part of each respective piston 320 will extend into a respective fork access path 336 that extends laterally through the side support 304 of seeds on sides Opposites of channel 318 for seeds.

p00056Each fixing head 312 is made of a slightly soft resilient material, such as neoprene, so that a seed held between the opposite fixing heads 312 will not be damaged, as described below.

p00057As described above, the seed holders 304 are mounted on the turntable 308, and are uniformly separated around a perimeter area thereof. Before the seed orientation process described above, subsequently to it, or substantially simultaneously with it, the rotating platform 308 is rotated to place an empty seed holder 308, that is, one without the seed, under the tube 224 of Orientation system transfer. More specifically, the seed channel 318 is placed under the transfer tube 224 of the orientation system. When a seed holder 304 is placed under the transfer tube 224 of the orientation system an automated separator 340 of the fixing heads is activated to separate the fixing heads 312, so that a seed can be received between the heads 312 of fixation. The spacer 340 of the fixing heads is mounted on the support structure of the system adjacent to the seed orientation device 212 and includes a pair of tongues 344 of the fork

p00058coupled to a fork base 348. The spacer 340 of the fixing heads is operable to extend the base 348 of the fork and the tabs 344 of the fork towards the seed holder 304. For example, the spacer 340 of the fixing heads may be an operable pneumatic device for extending and retracting the fork base 348. Each tongue 344 of the fork has a chamfered distal end portion and is sized to fit the fork access tracks 336.

p00059After activating the spacer 340 of the fixing heads, the fork base 348 is extended to the seed holder 304, so that the tabs 344 are inserted in the fork access roads 336. As each tongue 344 slides in the respective fork access path 336, the chamfered distal end portions slide between the top of each respective piston 320 and an inner wall of the fork access path 336. As the tabs 344 extend additionally to the inside of each fork access path 336, the chamfer of each tongue forces the respective piston 320 outward and away from the center line C of the seed holder. In consequence, as the pistons move outwardly and away 320 from the center line C, to also move away and are separated from each fastening heads 312 and the center line C. Therefore, the fixing heads 312 are moved to a retracted position in which a seed can be placed between them.

p00060Once the fixing heads 312 have been retracted, an appropriately oriented seed can be transported through the transfer tube 224 of the orientation system and can be placed in the orientation with the tip down between the fixing heads 312 . In various embodiments, the seed sampler system 10 further includes a height-setting seed subsystem 360 to place the seed at a specific height within the respective seed holder 304. The seed placement subsystem in height includes a vertical positioner 364 mounted on the system support structure below the area of ​​the turntable 308, directly in front of the transfer tube 224 of the orientation system, and a plate actuator 368 reference mounted on the central platform 214 directly in front of the spacer 340 of the fixing heads. The vertical positioner 364 includes a spring loaded piston 372 mounted on a positioner head 376 and the actuator 368 of the reference plate includes a reference plate 380 mounted on an actuator head 384 of the reference plate. The vertical positioner 364 is operable to extend the positioning head 376 and the plunger 372 towards a lower part of the turntable 308 directly opposite the center line C of the seed holder. For example, the vertical positioner 364 may be a pneumatic device operable to extend and retract the plunger 372. Similarly, the actuator 368 of the reference plate is operable to extend the actuator head 384 and the reference plate 380 on the part upper channel 318 for seeds of seed support. For example, the actuator 368 of the reference plate may be an operable pneumatic device for extending and retracting the reference plate 380.

p00061Once the seed has been placed between the retracted fixing heads 312, the positioning head 376 is extended upwards to insert a shaft 388 of the plunger through a hole (not shown) in the lower part of the rotating platform 308 and a coaxially aligned hole (not shown) at the bottom of channel 318 for seed support seeds. Substantially simultaneously, the actuator 368 of the reference plate extends the actuator head 384 to place the reference plate 380 at a specified distance above the seed holder 304, directly above the hole at the bottom of the channel 318 for Seed support seeds. More specifically, as the positioning head 376 moves up, the shaft 388 of the plunger extends into the coaxially aligned holes and makes contact with the tip of the seed. Then, the seed is pushed up between the fixing heads 312 until the seed crown makes contact with the reference plate 380. The spring loaded structure of the plunger 372 allows the shaft 388 to retract into the plunger 372 when the seed crown makes contact with the reference plate 380, so that the seed is held in place without damaging the seed. Consequently, the crown of the seed is located at a specific height with respect to the top of the rotating platform 308.

p00062With the seed crown fastened against the reference plate 380 by means of the spring loaded piston 372, the spacer 340 of the fixing heads is operated to retract the base 348 from the fork and remove the tabs 344 from the respective tracks 336 of access. After removal of the tabs 344, the springs 332 push the fixing heads 312 towards the deployed position and firmly fix the seed between the fixing heads 312. The reference plate 380 and the shaft 388 of the plunger are subsequently retracted leaving the seed properly placed, or "loaded," on the respective seed holder 304. Then, the system controller rotates the turntable 308 to place the "loaded" seed holder 304 below the grinding station 400 and the next empty seed holder 304 below the seed orientation device 212.

p00063Referring now to Figure 8, as described above, the seed sampling system 10 includes the seed transport subsystem 300 for transporting the seeds between individual stations of the sampling system, for example, the seed loading station 100, grinding station 400, sampling station 500, etc. In general, the seed transport subsystem 300 may be any suitable transport mechanism such as, for example, a belt conveyor, a roller conveyor, and the like. However, in various embodiments the transport subsystem 300 comprises the round turntable 308 which is pivotally mounted at its center of rotation. The turntable 308 is divided

p00064practically in a plurality of sectors, with each sector containing a support 304 of seeds. The number of sectors available on the turntable 308 can be even or odd with a chosen number that depends largely on the diameter of the turntable 308, the size of the seed holders 304 and the needs of the transport application.

p00065Circular turntable 308 is pivotally mounted at its center in a shaft and bearing system 390. In various embodiments, a shaft (not shown) of the shaft and bearing system 390 may have been directly coupled to a drive motor 392. Alternatively, the shaft can be separated from the drive motor 392 and driven to rotate by means of a chain transmission, a pulley transmission or a suitable gear transmission. In various implementations, the drive motor 392 may be a high torque stepper motor.

p00066In operation, the drive motor 392 for the turntable 308 is driven forward (which can be either clockwise or counterclockwise, depending on the configuration) to rotating the turntable 308 from station to station of the sampler system 10. Therefore, the seed holders 304 are aligned with auxiliary devices, such as the loading station 100, the grinding station 400, the sampling station 500, etc. In this configuration, an auxiliary device may have been placed between the turntable 308 in stations that are in alignment with each position and, therefore, have precise access to the seeds and seed holders 304. To the extent necessary, the peripheral edges of the turntable 308 may be supported with rollers, guides, sliders, or the like, to contribute to a smooth rotation of the rotary conveyor.

p00067In addition, with reference to Figure 8, as described above, once each seed holder 304 is "loaded" with a seed, the system controller rotates the turntable 308 to place the "loaded" seed holder 304 by under the 400 grinding station. The grinding station 400 includes at least one grinding tool 404 mounted on the system support structure above the perimeter area of ​​the rotating platform 308. The grinding tool or tools 404 are used to remove a portion of the seed coating of each seed when the respective seed holder 304 is placed below the grinding station 400. Each grinding tool 404 includes an operable Z8 actuator 408 for lowering and raising at least a portion of the respective grinding tool 404 along the Z axis. Each milling tool 404 is controlled by the system controller and can be operated electrically, pneumatically or hydraulically.

p00068The grinding tool or tools 404 may be any suitable mechanism for removing a portion of the seed coating material from each seed. For example, in various embodiments, each grinding tool 404 is a rotating device that includes the actuator 408 of the Z axis and a rotating transmission 412 operatively coupled to a chuck 416. Each actuator 408 of the Z axis is operable to lower and raise the respective chuck 416 and a drill 420 of the grinding tool held in the chuck 416 along the Z axis. Drill 420 of the grinding tool can be any instrument suitable for removing seed coating material, such as a grinding drill, a buriladora drill, a reamer, or a scraping tool. For example, in various embodiments, the drill 420 of the grinding tool comprises an end grinding drill. Each actuator 408 of the Z axis is controlled by means of the system controller to lower the respective actuator 408 of the Z axis by a specific predetermined distance. The rotary transmission 412 of each rotating milling tool 404 works to rotate, or turn the respective chuck 416 and any drill bit 420 of the grinding tool held in the chuck 416.

p00069During operation, when a seed holder 304 is placed below a rotating milling tool 404, the rotating transmission 412 is activated to start rotating the chuck 416 and the drill bit 420 of the grinding tool. Then, the actuator 408 of the Z axis is ordered to lower the respective chuck 416 and the drill bit 420 of the grinding tool a predetermined distance. As the drill bit 420 of the grinding tool is lowered, it makes contact with the crown of the seed and removes the coating of the seed from at least a portion of the crown. This exposes a portion of the internal seed material that can be extracted and used to test and analyze the various traits of the respective seed, as described below.

p00070In various embodiments, the grinding station 400 comprises at least two grinding tools 404 mounted on a horizontal moving platform 424 of the grinding station that is mounted on the support structure of the system. The horizontal movement platform 424 of the grinding station is controlled by the system controller to place a selected one of the grinding tools 404 above a seed holder 304 placed below the grinding station 400. Then, the selected milling tool 404 is operated as described above to remove the seed coating of at least a portion of the respective crown of the seed. Subsequently, the system controller may place a second of the milling tools 404 above a subsequent seed holder 304 placed below the milling station 400. Then, the second one is operated

p00071Selected tool 404 for grinding as described above to remove the seed coating of at least a portion of the respective crown of the seed. In such embodiments, the grinding station 400 may further include at least one grinding set 428 of the grinding drill to clean the drill bit 416 of the inactive grinding tool 404, that is, when it is not being used. That is, although a milling tool 404 is operable to remove the seed coating of a respective seed, the drill 420 of a second inactive milling tool 404 can be cleaned by means of a cleaning assembly 428 in preparation for the next grinding operation In various embodiments, the grinding drill cleaning assemblies 428 use air pressure or vacuum pressure to remove and / or collect any seed coating residue that may accumulate on the drill bits 420 of the grinding tools 404.

p00072Referring now to Figure 9, once the seed coating of a seed has been removed, the system controller rotates the turntable 308 to place the respective seed holder 304 below the sampling station 500 . The sampling station 500 includes at least one sampling tool 504 mounted on the support structure of the system anchored to the central platform 214 above the rotating platform 308. The sampling tool or tools 504 are used to extract a portion, that is, a sample, of the exposed internal material of the seed when the respective seed holder 304 is placed below the sampling station 500. Each sampling tool 504 includes a Z8 actuator 508 operable to lower and raise at least a portion of the respective sampling tool 504 along the Z axis. Each sampling tool 504 is controlled by the system controller and can be operated electrically, pneumatically or hydraulically.

p00073The sampling tool 504 may be any suitable mechanism for extracting a sample of the exposed internal material from the seed of each seed. For example, in various embodiments, each sampling tool 504 is a rotating device that includes the actuator 508 of the Z axis and a rotating transmission 512 operatively coupled to a chuck 516. Each actuator 508 of the Z axis is operable to lower and raise the respective chuck 516 and a drill 520 of the sampling tool held in the chuck 516 along the Z axis. The drill 520 of the sampling tool may be any instrument having an external diameter smaller than the circumference of the area of ​​the exposed internal material of the seed, and suitable for extracting a sample of the exposed internal material of the seed, such as a drill bit, a buriladora drill, a reamer, or a sample extraction tube. It is important that the drill 520 of the sampling tool has a smaller diameter than the drill 420 of the grinding tool to ensure that the sample material is obtained from an area where the seed coating material has been removed, thereby substantially eliminating that any seed coating material contaminates the material collected from the sample.

p00074For example, in various embodiments, the drill 520 of the sampling tool comprises a drill bit with a blade tip having an external diameter that is smaller than an external diameter of the drill 420 of the grinding tool. Each actuator 508 of the Z axis is controlled by the system controller to lower the respective actuator 508 of the Z axis a specific predetermined distance. The rotating transmission 512 of each rotating sampling tool 454 functions to rotate or rotate the respective chuck 516 and any drill bit 520 of the sampling tool held in the chuck

p00075516.

p00076During operation, when a seed holder 304 is placed below a rotating sampling tool 504, the rotating transmission 512 is activated to start rotating the chuck 516 and the drill bit 520 of the sampling tool. Then, the actuator 508 of the Z axis is ordered to lower the respective chuck 516 and the drill bit 520 of the sampling tool a specific predetermined distance. As drill bit 520 is lowered from the rotating sampling tool, it makes contact with the exposed internal material of the seed and cuts a sample of the internal material. So, he retires,

p00077or the sample is extracted, to be tested and analyzed for various traits and / or characteristics of the respective seed, as described below.

p00078In various embodiments, the sampling station 500 comprises at least two sampling tools 504 mounted on a horizontally moving platform 524 of the sampling station that is mounted on the support structure of the system. The horizontal movement platform 524 of the sampling station is controlled by the system controller to place a selected one of the sampling tools 504 above a seed holder 304 positioned below the sampling station 500 of samples. Then, the selected sampling tool 504 is operated as described above to extract the sample of the exposed internal material from the respective seed. Subsequently, the system controller may place a second of the sampling tools 504 above a subsequent seed holder 304 placed below the sampling station 500. Then, the second selected sampling tool 504 is operated as described above to extract the exposed internal material sample from the respective seed. In such embodiments, the sampling station 500 may further include at least one set 528 for cleaning drill bits.

p00079Sampling to clean the sample drill 520 of the inactive sample tool 504, that is, when it is not being used. That is, although a sample tool 504 is operable to remove the sample from a respective seed, the sample drill bit 520 of a second inactive sample tool 504 can be cleaned by means of the cleaning kit 528 Drill bits in preparation for the next sampling operation. In various embodiments, the sample cleaning drill assemblies 528 use air pressure and / or vacuum pressure to remove and / or collect any residue from the internal seed material that can accumulate in the sample drill bits 520 of 504 sampling tools.

p00080With reference now to Figures 9 and 10, the sample collection and transport (SCT) subsystem 600 is controlled by the system controller to operate synchronously in coordination with the sampling station 500 to collect each sample as it is extracted from each seed. The SCT subsystem 600 includes a motorized rotating platform 604 driven by a drive motor (not shown) similar to the drive motor 392 (shown in Figure 8) of the rotating platform 308. The SCT subsystem further includes a plurality of pick-up tube placement devices (CTP) 608 uniformly separated around a perimeter area, and mounted thereon, from the rotating platform 604. Each CTP device 608 includes a pivot bar 612 having a hollow tube holder 616 mounted through a transverse perforation (not shown) in the pivot rod 612. The tube support 616 includes a distal end 618 structured to accept a base 620 of a collection tube 624 and a proximal end 628 adapted to receive a pneumatic tube (not shown).

p00081Each CTP device 608 further includes an actuator 632 of the controllable rotation bar by means of the system controller for rotating the rotation bar 612 to various positions around a longitudinal axis of the rotation bar 612. In various embodiments, the actuator 632 of the pivot bar is operable to pivot the tube holder 616 between a water discharge cleaning position, as illustrated in Figure 11, a collection position, as illustrated in Figure 10, and a loading and storage position, as illustrated in Figures 13 and 17. The CTP device 608 further includes a stop arm 636 connected to the pivot bar 612 and an adjustable stop 640, for example, an adjustment screw, adjustablely coupled to the stop arm 636. The stop arm 636 and the adjustable stop 640 pivot with the rotating bar 612 and function to precisely stop the rotation of the rotating bar 612, so that the support 616 of the tube is in the collection position.

p00082Simultaneously with the operation of the seed loading station 100, the grinding station 400 and the sampling station 500, the SCT subsystem 600 operates to load the collection tube 624 into the supports 616 of the tube of each CTP device 608, collect the samples in the collection tubes 624 as each sample is extracted, and deposit the collected samples in the sample trays 14. The loading of the collection tubes 624 in the supports 616 of the tube and the deposition of the sample collected in the sample trays 14, will be further described below with reference to Figure 17, and Figures 12 and 13, respectively. The collection tubes 624 can be any container or device suitable for mounting on the supports 616 of the tube and collecting the samples as described below. For example, in various embodiments, the collection tubes 624 are disposable, so that each sample is collected in a clean collection tube 624. An example of such a disposable collection tube 624 is a filtered pipette.

p00083As described above, the SCT subsystem 600 is controlled by the system controller to operate synchronously in coordination with the sampling station 500 to collect each sample as it is extracted from each seed. More specifically, before extracting the sample from the seed, the system controller rotates the platform 604 to place a CTP device 608 adjacent to the sampling station 500. In particular, a CTP device 608 is placed adjacent to the sampling station 500, so that the respective support 616 of the tube is aligned with the seed held in an adjacent seed holder 304 that has been placed below a sampling device 504, by means of the controlled rotation of the turntable 308. Prior to the placement of the CTP device 608 adjacent to the seed holder 304 placed in the sampling station 500, the SCT system 600 has loaded a collection tube 624 at the respective distal end 618 of the tube holder and the actuator respective 632 of the pivot bar has raised the collection tube 624 to a position above the collection position, for example, the water discharge cleaning position. Once the CTP device 608 is positioned adjacent to the respective seed holder 304, the actuator 632 of the pivot bar lowers the loaded pickup tube 624 until the adjustable stop 640 makes contact with a stop plate 648 mounted on the system support structure between the turntable 308 and the platform 604 adjacent to the sampling station 500. The adjustable stop 640 is preconfigured, that is, adjusted in advance, so that the rotation of the rotating bar 612 is stopped to precisely locate a tip 672 of the collection tube 624 in very close proximity to the crown, or in contact therewith of the seed held in the adjacent seed holder 304.

p00084Then, the sample drill bit 620 is lowered from a sample tool 504 to start extracting the sample, as described above. As the sample drill bit 620 is lowered, a vacuum is provided at the tip 672 of the collection tube. The vacuum is provided by means of the vacuum tube (not shown) connected to the proximal end 628 of the tube holder 616. The vacuum tube is also connected to a vacuum source (not shown), so that the vacuum is through the vacuum tube, of the support 616

p00085of the hollow tube and the collection tube 624. Consequently, as the sample material drill bit 620 is removed, the sample is aspirated into the collection tube 624, in which the sample is collected. In various embodiments, the sampling station 500 may include a positive pressure device (not shown) to assist the vacuum provided in the respective seed to collect substantially all of the sample extracted in the respective collection tube 624.

p00086Each collection tube includes a filter 676 that prevents the sample from being aspirated into the support 616 of the tube and the vacuum tube. Once the sample has been collected, the actuator 632 of the pivot bar raises the collection tube 624 to the water jet discharge cleaning position and the respective CTP device 608 is advanced to a position adjacent to the subsystem 700 of liquid supply. Accordingly, another CTP device 608 and an empty collection tube 624 are placed adjacent to a subsequent seed holder 304 and an unsampled seed that have been moved to the sampling station.

p00087Referring now to Figures 11 and 12, the liquid supply subsystem 700 includes a liquid injection device 704 mounted on a linear actuator 708 operable to extend and retract the liquid injection device 704 along a linear axis. M. More specifically, the linear actuator 708 is operable to insert and remove an injection needle 712, fixed to the liquid injection device 704, into and out of the tip 672 of the respective collection tube 624. When a collection tube 624 with a collected sample has been raised to the water discharge cleaning position and advanced to be placed adjacent to the liquid supply subsystem 700, the linear actuator 708 and the injection needle 712 are found in the retracted position, as illustrated in Figure 11. The actuator 632 of the pivot bar and the rotating platform 604 are controlled by means of the system controller, so that when the CTP device 608 is adjacent to the liquid supply subsystem 700 and the collection tube 624 is raised up to The water discharge cleaning position, a linear axis of the collection tube 624 is substantially coaxial with the linear axis M of the liquid injection device 704, as shown in Figure 11.

p00088Once the linear axis of the collection tube 624 is positioned to be coaxial with the M axis, the linear actuator 708 extends to insert the injection needle 712 into the tip 672 of the collection tube 624. The liquid injection device 704 is connected to a source (not shown) of extracting fluid supply through a fluid port 716 coupled to a dosing valve 720 of the liquid injection device 704. Therefore, once the injection needle 712 is inserted into the tip 672 of the collection tube, the fluid injection device 704 injects a dosed amount of extraction fluid into the collection tube 624. The injected extraction fluid cleans by discharge of water, or lava, the internal sides of the collection tube 624 and creates an aqueous solution with the respective sample, referred to herein as an aqueous sample. Therefore, any of the collected samples that may have accumulated on the inner walls of the collection tube 624 can be cleaned by water discharge, so that substantially all of the collected sample is suspended in the resulting aqueous solution. The extraction liquid may be any liquid suitable for supplying substantially all of the sample material collected within each respective collection tube 624, without interfering with the desired analysis, for example, a chemical and genetic analysis, of the sample material. For example, in various embodiments, the extraction liquid may comprise distilled water or any suitable solvent compatible with the desired analysis of the sample.

p00089Once the collected sample has been mixed with the extraction liquid, the linear actuator 708 retracts to remove the injection needle 712 from the tip 672 of the collection tube. Then, the system controller advances the turntable 604 to place the CTP device 608 above the sample storage subsystem 800. In addition, the system controller instructs the respective actuator 632 of the pivot bar to place the collection tube in the loading and storage position. The loading and storage position points the tube support 616 and the mounted collection tube 624 downwards to a substantially vertical orientation.

p00090Referring now to Figure 13, the sample storage subsystem 800 includes a sample tray platform 804 adapted to securely retain a plurality of sample trays 14 in fixed positions and orientations. Each sample tray 14 includes a plurality of wells 22 of samples, each of which is adapted to receive a collected aqueous sample. The platform 804 of the sample tray is mounted on an XY 808 platform. The XY 808 platform is a two-dimensional translation mechanism, which includes a first translation track 812 and a second translation track 816. The XY 808 platform also includes a first linear actuator 818 operable to bidirectionally move a first carrier device (not shown) along the length of the first translation track 812. The XY 808 platform further includes a second linear actuator 820 operable to bidirectionally move a second carrier device (not shown) along the length of the second translation track 816. The second translation track 816 is mounted on the first carrier device and the platform 804 of the sample tray is mounted on the second carrier device.

p00091The first and second linear actuators 818 and 820 are controlled by the system controller to precisely move the platform 804 of the two-dimensional sample tray. More particularly, the first and second actuators 818 and 820 move the platform 804 of the sample tray in an XY coordinate system to precisely position any selected well 22 of any tray

p0009214 samples selected at an objective location below the CTP device 608 that holds the collection tube 624 containing the collected aqueous sample. The target location is the location in the XY coordinate system that is directly below the tip 672 of the collection tube when the collection tube 624 is in the loading and deposit position above the platform 804 of the tray samples. Therefore, once the CTP device 608 is placed above the platform 804 of the sample tray and the respective collection tube 624 is placed in the loading and storage position, with the tip 672 pointed towards the location target, the system controller places a selected well 22, from a selected tray 14 of samples in the target location. Then, the aqueous sample is deposited in the selected well 22 by providing positive pressure to the proximal end 628 of the support 616 of the tube.

p00093As sample trays 14 are placed on platform 804 of the sample tray, an identification number of the trays, for example, a barcode, is recorded for each sample tray 14 and the location of each tray 14 of samples on platform 804. In addition, as each aqueous solution is deposited in a well 22, an XY location of the well, that is, the target location, can be recorded on platform 804 of the sample tray. The recorded positions of the trays and wells on the platform 804 of the sample tray can be compared with the XY locations of each deposited aqueous sample, to identify an aqueous sample in each well 22 of each sample tray 14.

p00094Once each aqueous sample has been deposited in a selected well 22, the system controller advances the turntable 604 to place a subsequent CTP device 608, which holds a collection tube 624 containing a subsequent aqueous sample, above of the 800 sample deposit subsystem. In addition, the CTP device 608 that holds the empty used collection tube 624 is advanced to a station 850 for disposal of collection tubes (shown in Figure 1) in which the used collection tube 624 can be removed or ejected of the respective tube holder 616 and be discarded. With brief reference to Figure 1, in various embodiments, the collection tube waste station 850 includes a collection tube disposal device 854 mounted on a linear actuator 858 operable to extend and retract an automated gripping device 862. When a CTP device 608 holding a used collection tube 624 is placed adjacent to the collection tube disposal device 854, the system controller orders the linear actuator 858 to extend and the gripping device 862 to grip the used tube 624 of collection. Then, the system controller orders the linear actuator 858 to retract, thereby removing the tube used 624 for collecting the respective support 616 from the tube. Then, the gripping device 862 can be ordered to release the used collection tube 624, allowing it to fall into a waste container (not shown).

p00095Referring now to Figure 14, in various embodiments, after a sample of a seed has been extracted at the sampling station 500, the system controller can advance the turntable 308 to place the respective support 304 of Seeds adjacent to a 900 seed treatment station. The seed treatment station 900 includes a treatment distributor 904 mounted on the system support structure above the perimeter area of ​​the rotating platform 308. The treatment distributor 904 includes an applicator 908 configured to apply a seed treatment, such as a sealant to the exposed portion of the respective seed, that is, the crown area of ​​the seed in which the coating of the seed has been removed. the seed and the sample has been extracted. The seed treatment can be any substance designed to improve one or more properties of the seed or to protect the seed from bacteria or other harmful elements that could damage the seed and destroy the viability of seed germination. For example, in various embodiments, the seed treatment is a sealant comprising a fungicide and / or a polymer supplied to the seed by means of the treatment distributor 904 by means of the applicator 908. The applicator 908 can be any suitable device for applying the desired seed treatment to the seeds, for example, a brush, a needle or a nozzle. In various embodiments, the applicator 908 comprises a spray nozzle and the treatment distributor 904 includes a fluid port 912 coupled to a metering valve 916. In such embodiments, the treatment distributor 904 is connected to a source (not shown) of liquid seed treatment supply through the fluid hole 912. Consequently, when a seed holder 304 is placed in the seed treatment station 900, below the treatment distributor 904, the system controller instructs the treatment distributor 904 to spray a metered amount of seed treatment on the seed respective.

p00096Referring now to Figures 15 and 16, after sampling and optional seed treatment, the system controller advances the turntable 308 until the respective seed holder 304 is positioned adjacent to a second spacer 1004 of the fixing heads of the seed deposit sub-system 1000. The separator 1004 of the fixing heads is mounted on the support structure of the system and includes a pair of tabs 1008 of the fork coupled to a base 1012 of the fork. The separator 1004 of the fixing heads is substantially identical in shape and function to the separator 340 of the fixing heads described above with reference to Figure 5. Accordingly, after activation of the separator 1004 of the fixing heads, the fork base 1012 is extended towards the seed holder 304, so that the tabs 1008 are inserted in the fork access tracks 336. As the tabs 1008 slide in the respective access roads 336 of the fork, the fixing heads 312 of the respective seed holder 304 are retracted, as described above similarly. As the fixing heads 312 are retracted, the respective seed is allowed to fall through the coaxially aligned holes in the part

p00097bottom of the channel 318 for seeds of the seed holder and the turntable 308 inside a funnel 1016 of a seed conveyor 1020.

p00098The seed conveyor 1020 comprises a first tube section 1024 coupled at a first end to the funnel 1016 and an inlet of a first Venturi device 1028 at a second end. A second tube section 1032 is connected at a first end to an outlet of the first Venturi device 1028 and at a second end to an inlet of a second Venturi device 1036. An outlet of the second Venturi device 1036 is connected to the seed distributor 1040 which is mounted on the system support structure above a platform 1044 of the seed tray. The first Venturi 1028 device is operable to induce an air flow in the first and second sections 1024 and 1032 of tube towards the distributor 1040 of seeds. At the same time, the second Venturi device is operable to induce a flow of air towards the funnel 1016. Therefore, the air flow induced by the first Venturi device 1028 will aspirate the seed into the first funnel 1016 and the first tube section 1020. Furthermore, as the seed enters the first section 1024 of the tube, it is propelled to the seed distributor 1040 by means of the air flow provided by the first Venturi device 1028. Subsequently, as the seed approaches the seed distributor 1040, the speed of the seed is reduced by means of the air flow provided by the second Venturi device 1036, so that the seed of the seed distributor 1040 is gently distributed inside a tray 18 of seeds without damaging the seed. In various embodiments, the air flow provided by the second Venturi device 1036 stops, in fact, the movement of the seed, allowing the seed to fall by gravity into a tray 18 of seeds. Various position sensors (not shown) can be provided in the first and second 1024 and 1032 tube sections to detect the presence of the seed, and provide an input to the system controller to control the operation of the 1020 seed conveyor.

p00099With particular reference to Figure 16, the seed storage subsystem 1000 further includes a platform 1044 of the seed tray adapted to securely retain a plurality of seed trays 18 in fixed positions and orientations. Each seed tray 18 includes a plurality of seed wells 26, each of which is adapted to receive a distributed seed from the seed distributor 1040. The seed distributor 1040 is mounted on the support structure of the system above the platform 1044 of the seed tray, so that the seeds can be distributed from the distributor 1040 of seeds in selected wells 26 of selected tray seeds 18 of seeds.

p00100The platform 1044 of the seed tray is mounted on an XY 1048 platform. The XY 1048 platform is a two-dimensional translation mechanism, which includes a first translation track 1052 and a second translation track 1056. The XY 1048 platform further includes a first linear actuator 1060 operable to bidirectionally move a first carrier device (not shown) along the length of the first translation track 1052. The XY platform 1048 further includes a second linear actuator 1064 operable to bidirectionally move a second carrier device (not shown) along the length of the second translation track 1056. The second translation track 1056 is mounted on the first carrier device and the platform 1044 of the seed tray is mounted on the second carrier device.

p00101The first and second linear actuators 1060 and 1064 are controlled by the system controller to precisely move the platform 1044 of the two-dimensional seed tray. More particularly, the first and second actuators 1060 and 1064 move the platform 1044 of the seed tray in an XY coordinate system to precisely position any selected well 26 of any selected seed tray 18 in an objective location below the 1040 seed distributor. The target location is the location in the XY coordinate system that is directly below a tip 1068 of the seed distributor 1040. Once the seed holder 304 is placed above the funnel 1016, the system controller places a selected well 26, of a selected seed tray in the target location. The seed in the seed holder 304 is released into the funnel 1016 and transported to the seed distributor 1040, as described above, and is gently deposited in the selected well.

p00102As the seed trays 18 are placed on the platform 1044 of the seed tray, a tray identification number, for example, a barcode, is recorded for each seed tray 18 and the location of each tray 18 of Seeds on platform 1044 of the seed tray. In addition, as each seed is deposited in a well 26, an XY location of the well, that is, the target location, can be recorded on the platform 1044 of the seed tray. Then, the positions of the tray and the wells in the platform 1044 of the sample tray can be compared with the XY locations of each seed deposited, to identify the seed specified in each well 26 of each seed tray 18.

p00103As described above, each of the seed trays 18 and the sample trays 14 includes a plurality of wells 26 and 22, respectively. In various embodiments, the number and arrangement of wells 26 in seed trays 18 correspond to the number and arrangement of wells 22 in sample trays 14. This facilitates a one-to-one correspondence between a seed and its extracted sample. However, in some embodiments, it may be desirable to provide multiple wells 22 in the sample trays 14 for each well 26 in the seed trays 18, for example, when they can be

p00104perform multiple tests with the samples, or when different samples of the same seed can be taken (for example, samples of different depths).

p00105Referring now to Figure 17, in various embodiments, the seed sampler system 10 further includes a charging station 1100 of the collection tube for mounting the collection tubes 624 on the supports 616 of the tube of each CTP device 608 . The tube loading station 110 includes a hopper 1104 having a formed surface and a vibrating feed line 1108 extending from an open bottom of the hopper 1104. Large quantities of tubes 624 can be deposited in the hopper 1104 in which the feed vibrating duct 1108 feeds the collection tubes 624 inside a feed bucket 1112. A gravity-based feed track 1116 is connected to an outlet 1118 of the feed vibrating bowl 1112 at a first end 1116A. A second end of the feed track 1116 ends in a pushing device 1120 of the collection tubes. The pushing device 1120 extends orthogonally downwardly from the second end 1116B of the feed track and includes a longitudinal lifting channel 1124 that extends along the length of the pushing device 1120. The pushing device 1120 further includes an internal pushing mechanism (not shown) in the pushing device 1120. The pushing mechanism may be any operable mechanism for pushing a collection tube 624, placed longitudinally in the lifting channel 1124, out of an upper end 1120A of the pushing device 1120. For example, the pushing mechanism may include a linear actuator that drives a shaped pusher to receive at least a portion of a collection tube 624.

p00106As the vibrating bowl 1112 vibrates, the collection tubes 624 migrate to the outlet 1118 of the vibrating bowl 1112. At outlet 1118, the collection tubes 624 fall into the first end 1116A of the feed track that is formed to cause the collection tubes 624 to fall into a slot in the tube (not shown) that extends the length of the 1116 power track. More specifically, the collection tubes 624 are caused to fall downwardly into the tube groove and hang in the tube groove by means of a flange 620A of the base 620 (shown in Figure 10) of the tube of collection. The gravity and vibration of the vibrating bowl 1112 causes the collection tubes 624 to move the length of the feed track 1116 and accumulate, in a single row, at the second end 1116B of the feed track. As the collection tubes 624 accumulate, in a single row at the second end 1116 the first collection tube 624 will be oriented longitudinally in the longitudinal elevation channel. Then, the pushing device 1120 is actuated, so that the pushing mechanism pushes the first collection tube 624 out of the upper end 1124A of the lifting channel 1124 of the pushing device.

p00107Prior to actuation of the pushing device 1120, the system controller will advance the turntable 604 to place a CTP device 608 over the second end 1116B of the feed track 1116. The system controller will also order the actuator 632 of the pivot bar to place the tube holder 616 in the loading and storage position, so that the distal end 618 of the tube holder is directly above the end 1124A upper elevation channel. Therefore, as the first collection tube is pushed, or raised, out of the upper end 1124A of the lift channel, the base 620 of the collection tube is pushed over the distal end 617 of the tube holder. The distal end 618 of the tube holder is sized so that there is a frictional fit between the base 620 of the collection tube and the distal end 618 of the tube holder. Accordingly, the collection tube 624 is lifted out of the pushing device 1120 and mounted on the respective tube holder. The next collection tube 624 will be placed on the feeding track 1116 in the lifting channel 1124 and a distal end 618 of a support of a subsequent tube placed to receive the collection tube 624.

p00108Referring now to Figure 18, in various embodiments, collection tubes 624 may comprise commercially available pipettes, referred to herein as pipettes 624 '. In such embodiments, pipettes 624 'may require that a portion of tip 672' be removed to allow proper sample extraction, water discharge cleaning of the pipette, and deposition of the aqueous sample in trays 14 shows. Therefore, in such embodiments, the seed sampler system 10 may include a subsystem 1150 for preparing the operable collection tube to cut a portion of each tip 672 'of the pipette after each pipette 624' has been mounted on a respective support 616 of the tube. The collection tube preparation subsystem 1150 includes a linear actuator 1154 operable to extend and retract a base 1158A from a cutter 1158 along a linear axis P. The linear actuator 1154 is mounted on a system support structure below the rotating platform 604, so that when a newly mounted pipette 624 'is advanced, that is, the pipette 624' has just been mounted on the respective support 616 of the tube, until the collection tube preparation subsystem 1150, the tip 672 'of the pipette is placed inside a cutting chamber 1162.

p00109The cutting chamber 1162 is formed between the base 1158A of the cutter and a cutting recess 1166 formed in a head 1158B of the cutter 1158. As illustrated in Figure 18, when the newly mounted pipette 624 'is advanced from the station 1100 of loading of the collection tube, the base 1158A of the cutter is in the retracted position and the tip 672 'is placed inside the cutting recess 1166. Subsequently, the system controller instructs the linear actuator 1154 to extend the base 1158a of the cutter. The cutter 1158 includes an instrument

p001101170 for cutting, for example, a knife blade, firmly coupled to the base 1158A, or attached thereto, of the cutter by means of an angle 1174 for fixing the cutting instrument. The cutting instrument is firmly positioned so that when the linear actuator 1154 extends the base 1158A of the cutter, the cutting instrument will cut the tip 672 'from the pipette, thereby removing a portion of the tip 672'.

p00111Referring now to Figure 19, in various embodiments, after the sampled seed has been deposited in a selected well 26 of a selected seed tray 18, the system controller advances the turntable 308 and places the support now empty. 304 of seeds in a 1200 cleaning station. The cleaning station 1200 is operable to clean and remove any seed sample and / or residual seed treatment, for example, sealant, from the respective seed holder 304 after the sampled seed has been transported to a seed tray 18 and before a new seed is oriented and placed in the seed holder 304. The cleaning station comprises a roller brush 1204 and a vacuum 1208. The vacuum 1208 is connected to a vacuum source (not shown) to provide a vacuum in the vacuum nozzle 1212 placed in close proximity to the seed support channel 318 when the respective seed support 304 is advanced to the station 1200 cleaning The vacuum provided will remove any sample material and / or residual seed treatment that may have accumulated in the seed holder 304. In addition, the roller brush 1204 is driven, for example, electrically or pneumatically, to rotate on an axis 1216 of the roller, or with it. Simultaneously by providing the vacuum in the vacuum nozzle 1212, the system controller rotates the roller brush 1204 to remove any sample material and / or residual seed treatment that may have accumulated in the seed holder 304.

p00112Applications

p00113The present disclosure provides methods for analyzing seeds that have a desired trait, marker or genotype. In one aspect of the disclosure, the analytical procedures allow the individual seeds that are present in a batch or a population of bulk seeds to be analyzed, so that chemical and / or genetic characteristics of the individual seeds can be determined.

p00114Samples prepared by the present disclosure can be used to determine a wide variety of physical, morphological, chemical and / or genetic traits. In general, such traits are determined by selecting the samples in search of one or more chemical or genetic characteristics indicative of the traits. Non-limiting examples of chemical characteristics include proteins, oils, starches, fatty acids and metabolites. Accordingly, non-limiting examples of chemical traits include protein content, starch content, oil content, determination of fatty acid profiles, determination of metabolite profiles, etc. Genetic features may include, for example, genetic markers, genetic marker alleles, genes, DNA-derived sequences, RNA-derived sequences, promoters, quantitative trait loci (QTL), 5'UTR, 3'UTR, satellite markers, transgenes, mRNA, ds mRNA, transcriptional profiles and methylation patterns.

p00115In some embodiments, the methods and devices of the present disclosure may be used in a breeding program to select plants or seeds that have a desired trait or genotype marker. The procedures of the present disclosure can be used in combination with any reproduction methodology and can be used to select a single generation or to select multiple generations. The choice of the breeding procedure depends on the mode of reproduction of the plants, on the heritability of the trait (s) that are being improved, and on the type of commercially used cultivated variety (for example, F1 hybrid cultivated variety, pure line cultivated variety , etc.). Selected non-limiting approaches to reproduce the plants of the present disclosure are defined below. In addition, it will be understood that any commercial and non-commercial cultivated variety can be used in a breeding program. Factors such as, for example, exit vigor, vegetative vigor, effort tolerance, disease resistance, branching, flowering, seed set, seed size, seed density, ability to stand upright, and threshing capacity, etc., will normally dictate the election.

p00116In various embodiments, the methods of the present disclosure are used to determine genetic characteristics in a marker assisted reproduction program. Such procedures allow enhanced breeding programs assisted by markers in which direct non-destructive seed sampling can be carried out while maintaining the identity of individuals from the seed sampler to the field. As a result, the marker-assisted reproduction program results in a "high yield" platform in which a population of seeds that have a desired trait, marker or genotype can be collected more efficiently over a period of time. shorter time, with less required resources from the field and labor. Such advantages will be described in more detail below.

p00117In other embodiments, the present disclosure provides a method for analyzing individual seeds in a population of seeds that have genetic differences. The method comprises extracting a sample comprising cells and seed DNA in the population without affecting the viability of seed germination; select the DNA extracted from the sample in search of the presence or absence of at least one genetic marker;

p00118select seeds from the population based on the results of the DNA selection; and grow plants from the selected seed.

p00119As described above, the sampling systems and procedures of the present disclosure protect the viability of seed germination, so that they are not destructive. Germination viability means that a predominant number of sampled seeds (that is, greater than 50% of all sampled seeds) remain viable after sampling. In some particular embodiments, at least about 75% of the sampled seeds, and in some embodiments at least about 85% of the sampled seeds remain viable. It should be noted that lower germination viability rates may be tolerable under certain circumstances or for certain applications, for example, as genotification costs are reduced over time, since a greater number of seeds could be sampled by Same price of the genotype.

p00120In yet other embodiments, the viability of germination is maintained for at least about six months after sampling to ensure that the sampled seed will be viable until it reaches the field to be planted. In some particular embodiments, the methods of the present disclosure further comprise treating sampled seeds to maintain germination viability. Such treatment may include, in general, any means known in the art to protect a seed from environmental conditions while it is stored or during transport. For example, in some embodiments, the sampled seeds may be treated with a polymer and / or a fungicide to protect the sampled seed while it is stored or during transport to the field before being planted.

p00121In various embodiments, the samples of the present disclosure are used in a high-performance non-destructive method to analyze individual seeds in a seed population. The procedure comprises removing a sample from the seed while maintaining the viability of germination of the seed; and select the sample in search of the presence or absence of one or more characteristics indicative of a genetic or chemical trait. The procedure may also include selecting seeds from the population based on the results of the selection; and grow plants from the selected seed.

p00122DNA can be extracted from the sample using any DNA extraction procedure known to those skilled in the art that provides sufficient DNA production, DNA quality and PCR response. A non-limiting example of suitable DNA extraction procedures is an SDS based centrifugal extraction. In addition, the extracted DNA can be amplified after extraction using any amplification procedure known to those skilled in the art. For example, a suitable amplification procedure is the preparation of GenmersPhi® DNA amplification from Amersham Biosciences.

p00123The extracted DNA is selected in search of the presence or absence of a suitable genetic marker. A wide variety of genetic markers are available and are known to those skilled in the art. The selection of DNA in search of the presence or absence of the genetic marker can be used for the selection of seeds in a breeding population. Selection can be used to select QTL, alleles, or genomic regions (haplotypes). The alleles, QTL, or haplotypes that are going to be selected can be identified using newer techniques of molecular biology with modifications of classical reproduction strategies.

p00124In other various embodiments, the seed is selected based on the presence or absence of a genetic marker that is genetically linked to a QTL. Examples of QTL that are of interest usually include, without limitation, production, bed resistance, height, maturity, disease resistance, pest resistance, resistance to nutritional deficiency, grain composition, herbicide tolerance, fatty acid content, protein or carbohydrate metabolism, higher oil content, higher nutritional content, stress tolerance, organoleptic properties, morphological characteristics, other agronomic traits, traits for industrial uses, traits for greater consumer attraction, and a combination of traits as an index of multiple traits. Alternatively, the seed can be selected based on the presence or absence of a marker that is genetically linked with a haplotype associated with a QTL. Examples of such QTL may include again, without limitation, production, bed resistance, height, maturity, disease resistance, pest resistance, resistance to nutritional deficiency, grain composition, herbicide tolerance, fatty acid content, metabolism. of proteins or carbohydrates, a higher oil content, a higher nutritional content, stress tolerance, organoleptic properties, morphological characteristics, other agronomic traits, traits for industrial uses, traits for greater consumer attraction, and a combination of traits as an index of multiple traits.

p00125The selection of a breeding population could already start at the F2 level of reproduction, if inbred homozygous progenitors are used at the initial breeding crossing. An F1 generation could also be sampled and advanced if one or more of the crossing's parents are heterozygous for the alleles or markers of interest. The grower can select an F2 population to retrieve the marker genotype of each individual in the population. The initial population sizes can be adjusted, only limited by the number of seeds available to be selected, to meet the desired probability of identifying with <

p00126> Success the desired number of individuals. See Sedcole, JR “Number of plants necessary to recover a trait”. Crop Sci. 17: 667-68 (1977). Consequently, the probability of finding the desired genotype, the initial population size, and the selected size of the resulting population for various reproduction methodologies and inbreeding levels of the sampled population can be modified.

p00127The selected seeds can be accumulated or kept separated depending on the reproduction methodology and the objective. For example, when a grower is selecting an F2 population in search of disease resistance, all individuals with the desired genotype can be harvested and planted in the breeding nursery. On the other hand, if multiple QTLs with varying effects for a trait such as grain production of a given population are selected, the grower can keep the individual identity preserved, going to the field to differentiate individuals with various combinations of the target QTL.

p00128Several procedures for preserving the unique identity of seeds can be used while the seeds are transferred from the husking laboratory to the field. The procedures include, without limitation, transfer selected individuals to seed tape, to a seat tray, or to an indexing tray, transplant with peat pots, and plant by hand from individual seed packages. Multiple selection cycles can be used depending on reproduction objectives and genetic complexity.

p00129In addition, the disclosure selection procedures can be used in a breeding program to perform the introgression of a trait in a plant. Such procedures comprise extracting a sample comprising cells with seed DNA in a population, selection of DNA extracted from each seed in search of the presence or absence of at least one genetic marker, selecting seeds from the population based on the results of the DNA selection; grow a fertile seed plant; and use the fertile plant either as a female parent or a male parent at a crossroads with another plant.

p00130Examples of genetic selection to select seeds for an integration of traits include, without limitation, the identification of high frequencies of the parent's recurrent allele, the tracking of transgenes of interest or the selection in search of the absence of unwanted transgenes, the selection of hybrid seed test, and the test of stiffness.

p00131The identification of high frequencies of pairs of recurring alleles by means of the selection procedures of the present disclosure again allows a reduced number of rows per population and a greater number of populations, or inbred lines, to be planted in a given unit from the countryside Therefore, the selection procedures of the present disclosure can also effectively reduce the resources required to complete the conversion of inbred lines.

p00132The procedures of the present disclosure also provide a quality assurance (QA) and quality control by ensuring that regulated or unwanted transgenes are identified and discarded before planting.

p00133In addition, the procedures of the present disclosure can be applied to identify hybrid seeds for transgene testing. For example, in a conversion of an inbred line on the BCnF1 platform, a grower could effectively create a batch of hybrid seeds (excluding a gamete selection) that is 50% hemicigote for the trait of interest and 50% homozygous for the absence of the trait to generate hybrid seeds for the test. Then, the grower could select all the F1 seeds produced in the crossover test and identify and select those seeds that were hemicigotas. Such a procedure is advantageous because interferences from the hybrid assays would represent commercial hybrid genetics with respect to the trait's stiffness.

p00134Other applications of the selection procedures of the present disclosure to identify and track features of interest have the same advantages identified above with respect to the required resources of field and labor. In general, transgenic conversion programs are run in multi-station locations that have a much larger land structure and management costs. As such, the impact of either reducing the needs of rows per population or of increasing the number of populations in a given field unit is significantly more drastic in cost compared to temperate applications.

p00135Furthermore, the selection procedures of the present disclosure can be used to improve the effectiveness of the double haploid program by selecting desired genotypes in the haploid platform and identifying the level of ploidy to eliminate non-haploid seeds so that they are not processed and advance to the field Again, both applications result in the reduction of field resources by population and the ability to evaluate a large number of populations in a given field unit.

p00136In various embodiments, the disclosure also provides an assay to predict the embryo's cyclicity for a particular gene of interest (GOI). The test predicts the embryo's cysticity based on the ratio of the relative numbers of copies of a GOI and an internal control gene (IC) per cell or per genome. In general, this assay uses an IC gene that has a known homogeneity, for example, homozygous in the locus (two IC copies per

p00138diploid cell), to normalize the GOI measurement. The ratio of the relative numbers of copies of the IC to the GOI predicts the number of copies of the GOI in the cell. In a homozygous cell, for any given gene (or a single genetic sequence), the number of copies of the gene is equal to the level of ploidy of the cell since the sequence is present in the same locus in all homologous chromosomes. Therefore, the cellity of a cell in any locus can be determined by means of the number of copies of the gene in the cell.

p00139In some particular embodiments, the disclosure provides an assay to predict the maize embryo's cigarity. In corn seeds, the endosperm tissue is triploid, while the embryo tissue is diploid. The endosperm that is homozygous for the IC will contain three copies of IC. The number of GOI copies of the endosperm can vary from 9 (homozygous negative) to 3 (homozygous positive); and a number of GOI copies of the endosperm of 1 or 2 are found in heterozygous seeds for the GOI (or hemicigotas for the GOI if the GOI is a transgene). The number of copies of the endosperm reflects the cigarity of the embryo: a homozygous endosperm (positive or negative) accompanies a homozygous embryo, heterozygous endosperm (either a number of GOI copies of 1 or 2) reflects a heterozygous embryo (a number of GOI copies of 1). The number of GOI copies of the endosperm (which may vary from 0 to 3 copies) can be determined from the ratio of the number of IC copies of the endosperm to the number of GOI copies of the endosperm (which may vary from 0 / 3 to 3/3, that is, from 0 to 1), which can then be used to predict the embryo's stiffness.

p00140The copy numbers of the GOI or IC can be determined by any convenient test technique for quantifying copy numbers, as is known in the art. Examples of suitable assays include, without limitation, Real Time (TaqMan®) PCR (Applied Biosystems, Foster City, California, USA) and Invader® (Third Wave Technologies, Madison, Wisconsin, USA). Preferably, such assays are developed in such a way that the amplification efficiency of both the IC and GOI sequences are the same or very similar. For example, in a Real Time TaqMan® PCR assay, the signal from a single copy GOI (it is determined that the source cell is heterozygous for the GOI) will be detected an amplification cycle subsequent to the signal of an IC of two copies, because the amount of the GOI is half that of the IC. For the same heterozygous sample, an Invader® assay would measure a GOI / IC ratio of approximately 1: 2 or 0.5. For a sample that is homozygous for both the GOI and the IC, the GOI signal would be detected at the same time as the IC signal (TaqMan®), and the Invader test would measure a GOI / IC ratio of approximately 2: 2 or 1.

p00141These guidelines are applicable to any polyploid cell, or haploid cells (such as pollen cells), since the number of copies of the GOI or IC remain proportional to the number of genome copies (or ploidy level) of the cell. Therefore, these tests of cyclicity in triploid tissues such as corn endosperm can be carried out.