omegranate is one of the oldest known fruits to the man and is one of the earliest trees cultivated in the western part of Asia and Middle East. According to historical evidences, it seems that probably Iran has been the origin of pomegranate and it has spread from this region to the other parts of the world. Due to the considerable area of the cultivated pomegranate in Iran and the distinguished position of this country as one of the greatest pomegranate producers in the world, it is very important to determine the genetic relationships among cultivars and genotypes of this fruit, in order to preserve its genetic diversity, recognition and the selection of the desired genotypes to improve it. Among the previous molecular markers used in pomegranate, microsatellite markers showed higher efficiency for separating the pomegranate genotypes, due to the co-dominant inheritance nature, high reproducibility and having high levels of polymorphism. Therefore, in the present research the investigation of the genetic diversity of pomegranate was done based on microsatellite markers on 77 Iranian and foreign genotypes of pomegranate from Japan, Turkmenistan, Russia, Italy and USA. The results showed that 10 primer pairs out of 25 microsatellite primer pairs were polymorphic and revealed 38 alleles with an average of 3.1 alleles per locus. The mean of polymorphic information content was calculated as 0.35. According to the obtained dendrogram the Iranian genotypes almost clustered in accordance to their geographical distribution. The results showed that the minimum gene diversity is among the commercial and domestic cultivars and the maximum is among the wild ones. Comparing the populations of Iranian pomegranate with the populations from other countries revealed that the average gene diversity per each locus for Iranian population is more than the other populations and the unknown genotypes show close relation to Turkmenistan and Iranian ones. Most of the Russian genotypes clustered with the domestic and commercial Iranian cultivars and some of the American genotypes grouped with the wild Iranian ones. Also close relationship was observed among two Japanese samples with the wild and ornamental genotypes of Iran. All of these indicates the close relations of foreign cultivars to Iranian ones and probably confirms the hypothesis that pomegranate spread from Iran to the other regions of the world. In order to do more advanced evolutional studies on pomegranate, the digestion of the IGS region between the conserved ribosomal RNA genes was done by 6 restriction enzymes on some of the pomegranate genotypes. The results revealed that the used restriction enzymes could not show polymorphism. Considering the related reports that indicate low genetic polymorphism in molecular researches on pomegranate, this was not unexpected and using more enzymes with the ability of 4 base pair recognition sites could give better results. Investigation of