Sequence-Based SNP Genotyping in Durum Wheat

A novel SNP discovery and genotyping technology, developed by KeyGene and called random Sequence Based Genotyping (rSBG), was tested and validated in durum and bread wheat. This technology enables sequence-based SNP discovery and genotyping in a single assay, which is cost-effective (no separate costs for SNP discovery and genotyping) and robust (no marker conversion required). Within the EU FP7 BioExploit project, the rSBG technology was adjusted to highly repetitive, polyploid genomes and optimized on the durum wheat parental lines Colosseo (CLS) and Lloyd (LLD). This protocol was subsequently used to genotype 91 CLS x LLD recombinant inbred lines (RILs) in a single GAII run. A total of 10,761 putative SNPs in 6,836 loci were identified between the parents, using stringent SNP mining rules. Out of these, 1,038 were mapped with high confidence (two-point LOD > 6) to a pre-existing framework map containing 709 markers (SSRs, DArT^®, SNPs from CRoPS^® technology). The relatively low percentage of SNPs available for mapping was mainly due to lack of sequencing depth. Nevertheless, this rSBG experiment allowed genotyping of the RILs at a density of approximately one SNP marker every 2.8 cM. It is expected that an increased sequencing output will generate a higher number of genetically informative SNPs. Interestingly, relaxation of three out of four mining rules resulted in the robust mapping of an additional 1300 SNPs. Since the SNPs are associated with unique sequences, we will explore the possibility of linking them to the reference conserved orthologous sets from the grass genomes, which could add additional value to the rSBG SNP set.

The rSBG and CRoPS^® technologies are covered by patents and patents owned by Keygene N.V. CRoPS is a registered trademark of Keygene N.V. Other brand names and/or trade names might be (registered) trademarks of their respective owners.