The Olive (Olea europaea L.) is a member of the Oleaceae family, which includes approximately 30 genera and 600 species. In this study collected 88 leaf samples from 17 olive varieties from five geographical regions in Iraq. The Deoxyribonucleic acid (DNA) was extracted from 17 varieties. The DNA samples were analyzed by polymerase chain reaction (PCR) using 9 microsatellite (SSR) primers (DCA7, DCA9, DCA11, DCA16, DCA18, EMO90, GAPU71A, GAPU71B, and GAPU103). For statistical analysis, Power Marker v.3.25 was used to assess different molecular genetics parameters.  Mega software was used to generate a phylogenetic tree.
The size of the alleles ranged from 91 bp in DAC11 to 245/270 bp in DAC9. The computed mean number of allele was 44.33, and the allele frequency was 0.0892. The observed heterozygosity was 0.2577, with heterozygosity 0.258. Cluster analysis of the total 9 microsatellite markers was used to study the genetic relationships among the 88 different olive genotypes, and they were divided into two main clusters and five sub-clusters. The local cultivars and the foreign cultivar shared the same SSR alleles and they were genetically similar. There were small molecular variations (1%) among the studied geographical regions. The study aimed to investigate the genetic connections among these cultivars and offer valuable insights that can guide forthcoming efforts in cultivation expansion and breeding initiatives.