There are three native varieties of edible yams (Dioscorea japonica Thunb.) in Taiwan, but germplasm has been collected from only a few. To assess the genetic diversity of 99 accessions collected from four geographically diverse regions, inter-simple sequence repeat (ISSR) DNA markers were analysed, and 90.2% of the polymorphic markers in these accessions were found. Genetic similarity among accessions ranged from 30.8 to 74.4%, averaging 48.4%. Cluster analyses revealed four main clusters plus one outlier (Dioscorea alata L.). The varieties oldhamii and pseudojaponica were separated into different clusters and var. japonica was grouped with both varieties. Since var. japonica was found to be genetically similar to var. oldhamii and var. pseudojaponica, we suggest that var. japonica is a hybrid or intermediate variety between the two varieties. Molecular analysis of variance results indicated that variation within counties (95.94%) was predominantly greater than variation among counties (3.63%) and among regions (0.43%). Overall, gene flow (Nm= 0.970) estimated from genetic differentiation (Gst= 0.340) suggests that gene flow among regions is relatively high. The highest genetic diversity (H= 0.191) and Shannon's index (I= 0.312), and a high gene flow (Nm= 3.926) of germplasm in the northern region suggest that this region is a possible centre of dispersal and domestication of yams in Taiwan. This study provided valuable information for germplasm collection and genetic improvement.