Anatolia (Asia Minor) is considered to be the cradle of viticulture, but wild grape accessions from this region have not been subjected to any genetic analysis. We present the first genetic characterization of wild grapes (Vitis vinifera subsp. sylvestris) from this region. Using 15 nuclear microsatellites, we genotyped 84 wild grape accessions collected from three Anatolian locations. The unweighted pair group method with arithmetic mean analysis revealed four genetic clusters that partially separated Anatolian wild grape populations. In general, accessions from geographically closer locations showed higher genetic similarities than those from more distant locations. In some cases, accessions from one population showed close genetic relationships to accessions from a different population. The genetic diversity between natural populations from both ends of the Mediterranean basin has shown higher genetic diversity in the Anatolian Peninsula than peripheral populations, suggesting that this area could be one centre of diversity of the species. The genetic relationship between wild and cultivated grapevine from Anatolian Peninsula indicated a clear separation between them. However, we cannot discarded a local genetic contribution. Overall, this study reveals useful information for management and potential utilization of Anatolian wild grape germplasm.