• Premise of the study: Microsatellite loci were isolated and characterized from enriched genomic libraries of two taxa of the Chilean Epipetrum group of Dioscorea to assess their levels of genetic diversity and population differentiation.• Methods and Results: Eleven microsatellite loci were identified. Six out of nine microsatellites from D. biloba amplified in D. humilis, and the two microsatellites from D. humilis amplified in both taxa. Two different sets of eight loci amplified in each of the two tested taxa, D. biloba and D. humilis. The average number of alleles was 5.75 and 5 for D. biloba and D. humilis, respectively. Higher levels of mean genetic diversity were found in D. biloba (HE = 0.639) than in D. humilis (HE = 0.414).• Conclusions: These microsatellite primers will be useful in population genetic studies and to establish conservation strategies in the endangered taxa of the Epipetrum group of Dioscorea.