Cost-Effective KASPar Assays for SNP Genotyping and Diversity Analysis in the Reference Set of Groundnut

Competitive allele-specific PCR (KASPar) assays have emerged as a cost-effective marker assays especially for molecular breeding applications in a range of crop species. SNP genotyping, because of tetraploid nature of the genome, is very challenging in cultivated groundnut germplasm lines. Therefore, 96 SNPs were used to develop KASPar assays and referred as GKAMs (Groundnut KASPar Assay Markers). All 96 GKAMs were screened on a set of 94 genotypes that includes parental lines of 28 mapping populations. As a result, 90 GKAMs were validated and 73 GKAMs showed polymorphism across 94 genotypes and 71 GKAMs were polymorphic between parental lines of 28 mapping populations. With an objective to assess effectiveness of KASPar assays in elucidating genetic relationships, all 90 validated GKAMs were screened on the reference set (280 diverse genotypes). The polymorphism information content value ranged from 0.01 to 0.50 with a mean of 0.24. Cluster analysis of allelic data grouped accessions according to their genome type, subspecies and botanical variety. The cultivated genotypes were sub-divided into two subspecies fastigiata and hypogaea in general; however, some overlaps too were found indicating their frequent inter-crossing during the course of evolution. The subspecies fastigiata comprising of three botanical varieties fastigiata, peruviana and vulgaris formed one group while the subspecies hypogaea containing botanical variety hirsuta and hypogaea formed another group. Furthermore, the wild species, having diploid genomes AA, BB, EE, EX and PP were grouped into a single cluster. This study demonstrates the value of GKAMs for genetics research and breeding applications in groundnut.