Domestication process and the subsequent breeding may result in a genetic mutation and selection pressures that possiblycontributed to the emergence of two phenotypes of barley spikelets, namely six-row and two-row barley. The present studyused microsatellite genetic markers to investigate the population genetics of a selected set of accessions of domesticatedbarley representing different populations of Jordan, to identify a signature of selection resulted from domestication process.Genomic and statistical approaches, such as the analysis of molecular variance (AMOVA) and the so-called hierarchical“outlier” tests, were utilized to identify signatures of selection. Generally, there was a high level of polymorphisms in allthe studied populations, which ranged from 19.23 for Ramtha to 69.23 (%) for Zarga, with an average of 53.61 (%). Theaverage number of allele per locus for the entire sample set was of 9.346 and it ranged 2 occurring at [(Bmag0136),(EBmac0970_a), (EBmac0970_c) loci], to 27 at (Bmac0040) locus. The results of AMOVA showed that the main portion(~72%) of total diversity was attributed to differentiations within populations. Other portions of diversity (11.040%) wereexplained by the diversity among populations. As revealed by the hierarchical outlier tests and AMOVA results, 9microsatellites were identified to be under possible selection pressures, possibly indicates that these loci were important inthe past improvement of barley by early cultivators. In conclusion, the present study shows that barley landraces of Jordanpossess high levels of genetic diversity and allelic richness that could be utilized in barley improvement and breedingprogram.