Betony (stachys lavandulifolia) is one of the 34 wild species of important genus Stachys from Lamiaceae family which grows naturally in Iran. In present study, seven endemic populations of Stachys lavandulifolia (including 56 samples) from five provinces were investigated. ISSRs (Inter simple sequence repeats) were used for surveying genetic diversity and population structure of Betony. In total, 12 used primers produced 176 fragments which all of them (100%) were polymorphic. By using NTSYS-pc software and UPGMA method, the samples were divided into six groups based on Jaccard genetic similarity matrix. Principal coordinates analysis (PCOA) confirmed the accuracy of clustering results. POPGENE software was used to describe genetic variation within populations according to Shannon's index and Nei's gene diversity analysis. Genetic diversity within the populations of Aznav and Toreh was the most and the least, respectively. The average observed number of allels (na) to the effective number of allels (ne) was calculated and the number was 83%. Studies based on this average showed that the gene distribution were balanced in each population. Moreover, GeneAlex and POPGENE softwares were used to analyze genetic diversity and molecular variance among populations. Gst and Nm indexes average of the populations were accounted 0.20 and 1.94, respectively, indicating a balanced gene exchange among seven studied Betony populations. Molecular variance analysis of the populations showed that genetic diversity within and among the populations were 75% and 25%, respectively. According to the data obtained Nei's genetic similarity ranged between 0.79-0.97, the most similarity was distinguished between Toreh with Zia-Abad and Khoramdareh whereas the least one was between Toreh and Salavat-Abad. After investigating genetic structure of the populations using STRUCTURE software, they were divided into four groups. According to the obtained results, ISSR marker could be used as a valuabl
