Plant breeders today demand a more sophisticated approach than crossing plants, waiting for germination and phenotypically scoring progeny. Modern breeding programs benefit from speed and efficiency, and for many researchers this means adopting genomic technologies as early as possible. Recently, Hans Peter Koelewijn, lead scientist for cucumber genetics at Bayer Vegetable Seeds, described how Thermo Fisher Scientific AgriSeq genotyping by sequencing (GBS) technology brings increased throughput for a faster turnaround time to his work.
Vegetable genomic libraries as a tool
Crop scientists now have access to entire genomes, so they can integrate molecular information and techniques within or ahead of the traditional breeding process. Instead of crossing progeny and waiting to observe, scientists can now evaluate individuals according to genetic markers such as single nucleotide polymorphisms (SNPs), selecting plants by molecular markers to give the right combination of traits.
Molecular data on crops give breeders immediate access to the segregation of markers that correlate with desirable traits. Koelewijn believes that getting this molecular information into vegetable seed breeding helps speed up crop development. In order to maximize efficiency and reduce costs, he notes that the method used must be fast and return accurate data, with high-density coverage for markers of interest.
Optimized crop breeding
Bayer needed a solution to optimize genomic evaluations within its vegetable-breeding program. The company knew that it needed a platform that could analyze a wide variety of crops and cope with rapidly changing molecular information coming in from plant genetics research. Accordingly, Bayer researchers considered various commercial platforms, looking for a cost-effective and flexible approach that enabled higher marker density (1,000 to 10,000 SNPs) screening with accuracy and sensitivity.
AgriSeq GBS solution
As part of a European Union genomic strategy project, Koelewijn and his team investigated AgriSeq GBS solutions as a next-generation targeted genotyping approach. This method uses highly efficient multiplexed polymerase chain reaction (PCR) to target hundreds to thousands of markers simultaneously across hundreds to thousands of samples. AgriSeq GBS uniformly amplifies all the targets in a single reaction, then bar codes the amplicon libraries and pools them for simultaneous sequencing on next-generation sequencing (NGS) instruments such as the Ion Torrent. Since the reaction targets multiple variants of interest at the same time, results are fast.
Cucumber genomics at full speed
The team developed a panel of variants for SNP analysis in six types of cucumber and the various lines within them. They submitted 3,054 potential variants and from this developed 2,804 as an end library. The team developed an AgriSeq panel targeting 2,804 variants, tracking key regions across six varieties of cucumbers. For validation, the study compared analyses with wild variants.
Results from the evaluations showed an average call rate of 91.4%, with 80% of markers showing a call rate above 90%. Only 21 out of the 2,804 variants remained undetected in the samples.
During the collaborative pilot program with Thermo Fisher Scientific, Koelewijn and his colleagues found that the AgriSeq system not only supported accurate high-throughput genotyping of their targeted 3,000 markers in cucumbers, but also identified an additional 5,000 novel SNPs, further increasing the marker density and informativeness of the genotyping panel.
Koelewijn concludes that the AgriSeq GBS solution is very sensitive and low cost, and that the fast turnaround approach delivers results in around two to four days. He also thinks that the technology shows great potential for use in other crop breeding programs. Describing the cucumber as the “Arabidopsis of vegetable breeding research,” he notes that “…if it works in cucumbers, it works in other crops.”
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