Emys orbicularis is an Old World terrapin that has a wide distribution range from Russia to Iran, Europe, and Northwestern Africa. In this study, we compare the Southern Caspian Sea lineage (in northern Iran) of E. orbicularis with other lineages of this species in Europe using the Cyt b gene fragment. Based on our results, the Caspian Sea lineage shows a high value of PhiPT (> 0.87) in comparison with other lineages that means the lineage isolated more from other lineages. Molecular phylogenetic analyses indicate that Iranian and Caucasus lineages are distinct from other lineages. The Mantel test confirmed the pattern of isolation by distance for the lineages from Western Europe to Asia. These results indicate that the Iranian population has a long history in the Caucasian and Southern Caspian Sea region (> 6 MYA). There are many reasons to conserve this species in Iran, as high inbreeding rates, low genetic variability, accidental events on the roads, and pet trade must be considered in future conservation action plans.