A set of eight highly polymorphic microsatellite markers was isolated and characterized from a genomic library enriched for dinucleotide repeats in the European pond turtle, Emys orbicularis . The markers were tested for polymorphism in a total of 33 turtles sampled in two natural ponds in the nature reserve of Kerkini, northern Greece. Number of alleles varied from 10 to 18, and expected heterozygosity ranged between 0.738 and 0.921. This novel set of loci will be particularly useful to assess fine‐scale population structure and for parentage analysis in E. orbicularis .