Discriminating genotypes within plant collections is imperative, and DNA sequence approaches for detecting single nucleotide polymorphisms (SNPs) have proved essential in any modern analysis of germplasm. By sequencing the α-Phs and PvFRO1 genes that, respectively, encode phaseolin and an iron reductase, we prospected for SNPs in exonic and intronic regions of both genes in a sample of 31 accessions of Phaseolus vulgaris from Mesoamerican and Andean gene pools, and one accession of Phaseolus lunatus, chosen as an outgroup. Sequence alignment showed 95 SNPs in α-Phs and 83 in PvFRO1, but diversity along the nucleotide sequences was not evenly distributed in both genes. Accessions from the same gene pool showed greater similarity than those from different gene pools, and the cluster patterns obtained in this study were consistent with the hierarchical organization into two P. vulgaris gene pools. The polymorphisms detected in the α-Phs gene allowed better discrimination among the accessions within each cluster than the PvFRO1 polymorphisms. Furthermore, some variations within exons changes amino acids in both predicted protein sequences. In an unprecedented result, the phaseolin-predicted amino acid variation allowed most of the accessions to be typified.