Iran is one of the chief origins of subgenus Cerasus germplasm. In this study, the genetic variation of new Iranian sour cherries (which had such superior growth characteristics and fruit quality as to be considered for the introduction of new cultivars) was investigated and identified using 23 intersimple sequence repeat (ISSR) markers. Results indicated a high level of polymorphism of the genotypes based on these markers. According to these results, primers tested in this study specially ISSR-4, ISSR-6, ISSR-13, ISSR-14, ISSR-16, and ISSR-19 produced good and various levels of amplifications which can be effectively used in genetic studies of the sour cherry. The genetic similarity among genotypes showed a high diversity among the genotypes. Cluster analysis separated improved cultivars from promising Iranian genotypes, and the PCoA supported the cluster analysis results. Since the Iranian genotypes were superior to the improved cultivars and were separated from them in most groups, these genotypes can be considered as distinct genotypes for further evaluations in the framework of breeding programs and new cultivar identification in cherries. Results also confirmed that ISSR is a reliable DNA marker that can be used for exact genetic studies and in sour cherry breeding programs. 1. Introduction The sour cherry belongs to the family of Rosaceae, subfamily Prunoideae, genus Prunus, and subgenus Cerasus [1]. It is an allotetraploid species ( ) resulting from a natural hybridization between Prunus avium L. (Sweet Cherry) and Prunus fruticosa Pall. (Ground Cherry) [2]. This species is reported to have originated from the area that comprises Asia Minor, Iran, Iraq, and Syria [3] and has been used as rootstock and also in breeding programs for developing new commercial cultivars, dwarf, and resistant rootstocks [4–6]. According to the FAO database, Iran ranked third worldwide in 2011 for cherry production after Turkey and the USA with a total of 241 thousand tons produced [7]. Genetic variability is a prerequisite for any plant breeding program [8]. As an origin of the subgenus Cerasus, Iran has rich cherry germplasm resources. Using diverse Cerasus subgenus resources to broaden the genetic base of cherry cultivars and rootstocks and improving them for development of the cherry industry are important goals for cherry breeders in Iran [9, 10]. Therefore, it is necessary to characterize and preserve these genotypes and cultivars [11]. DNA markers are very useful in distinguishing between accessions and in investigations of genetic diversity or relatedness
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