Molecular Marker Development and Germplasm Evaluation for Cultivated Peanut (Arachis hypogaea L.)

The development of molecular markers in cultivated peanut has been growing rapidly. This study focused on discovering and validating molecular markers from two peanut transcript databases: a NCBI EST dataset with 253,274 sequences and an in-house 454 RNAseq dataset with 288,701 sequences. From these two datasets, a total of 6,455 novel SSRs and 11,902 SNPs were identified. Out of the 6,455 SSRs, 380 representing various SSR types were selected for PCR validation. The amplification rate was 89.2%. Twenty-two (6.5%) SSRs were polymorphic between at least one pair of four genotypes. To validate the SNPs, Sanger sequencing of PCR products targeting 110 SNPs was conducted. Sequence comparisons of the PCR products from the four different genotypes revealed 13 true SNPs between tetraploid genotypes and 193 homoeologous SNPs within genotypes. Eight out of the 22 validated SSR markers were further selected to evaluate the genetic diversity of the U.S. peanut mini core collection and Florida peanut breeding lines. This marker set demonstrated high discrimination power by displaying an average polymorphism information content (PIC) value of 0.783, a combined probability of identity (I) of 10^-11, and a combined power of exclusion (Q) of 0.99991. The structure analysis revealed four sub-populations among the peanut accessions and lines evaluated. The results of this study enrich the peanut molecular marker tool box by providing over 6000 novel SSR markers covering the whole peanut genome and by providing the credentials for true peanut SNP marker development.