Pulse SNP Discovery Using Targeted 454 FLX Sequencing

Increasing the number of genetic marker resources is necessary to generate dense genetic maps, facilitate association mapping and improve the efficiency of marker-assisted breeding programs for pulse crops. To this end, a SNP discovery pipeline has been established, identifying robust markers suitable for low cost, high volume genotyping platforms such as Illumina's Goldengate or Infinium assays. 3' targeted transcript libraries for a selected panel of cultivars and wild lines in four different species (lentil, chickpea, field pea, and common bean) were sequenced by 454 FLX, giving sufficient read length and depth for SNP calling. The sequences were assembled and potential SNPs located. Allele frequency was assessed across the panel of lines for each species, and the contigs were mapped against both annotated soybean and Medicago genomes to determine flanking sequence length. Filtering to determine the most robust SNPs was applied, identifying 3289 high-quality SNPS in chickpea, 5733 in pea, 8533 in lentil, and 14545 in bean. Final selection of SNPs was made to give an even coverage of Medicago genespace, and initial genotyping results from a 1536 lentil GoldenGate OPA showed 88% of the SNPs produced good-quality calls across cultivated lines.