Abstract: Alhagi graecorum Boiss. (Camel thorn) (Family Fabaceae) is a wild plant which grows in salty soils and dry environment. It is native to Mediterranean and central Asia. Several studies have revealed the use of Alhagi plants in treating a wide spectrum of diseases. In this study, eight ISSR primers were used to assess genetic diversity of five populations of A. graecorum (El-Dakhla Oasis; Botanical garden of botany department, Ain Shams University; Wadi El-Rayan; Qarun Lake and Siwa Oasis) using individual- and bulked samples based approaches. Twenty five individuals were sampled, amplified separately and amplified as bulked DNA to identify polymorphism among the studied populations. In total, 101 and 97 bands were scored with 83.3% and 78.4% polymorphism, respectively. Average polymorphism information content (PIC) for amplified DNA of individual samples was 0.875 and bulked-samples was recorded 0.69. Analysis of molecular variance (AMOVA) showed that 63% of the total variation was observed among populations while variation within populations accounted for the remaining 37%. The highest genetic similarity index was (0.99) among the studied Alhagi genotypes was recorded between the two genotypes 6 and 7 both individuals were belonging to Ain Shams University population, while among inter populations was (0.853%) between Qarun Lake and Siwa Oasis populations. The UPGMA cluster analysis divided the 25 individuals and bulked samples into two main groups. Each group was further subdivided into many sub-clusters, PCoA was also carried out and in agreement with the UPGMA. The obtained results demonstrate that the ISSR markers were efficient for evaluation of the genetic relationships between intra- and inter-population due to their efficiency in revealing polymorphism.