Iran has a rich pistachio germplasm, thereby, the diversity and number of Iranian pistachio cultivars is unique in the world. In this study, genetic relationships among thirty one cultivars was assessed by using six inter simple sequence repeat ISSR primers. During the ISSR screening in this study, good amplification products were obtained from primers based on GA, CA and GAA repeats. Since, primers based on CT, GT and CAA repeats produced few large separate bands which were eliminated for the final analysis. The total of 28 bands of which 13 46/42% were polymorphic were amplified by the three primers, an average of 9.3 bands per primer. The total number of amplified fragments was between 7 to 12 and the number of polymorphic fragments ranged from three to five. The range of genetic similarity was from 0/84 to 1 and the constructed unweighted pair group method with arithmetic averages UPGMA dendrogram classified the tested genotypes into 11 main clusters. The present study showed that there is low genetic diversity among tested cultivars and the ISSR-PCR analysis produces sufficient polymorphisms for large-scale DNA fingerprinting. This study reports the first application of the ISSR technique in characterization of Iranian pistachio cultivars
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