Sheep represent one of the most important domestic animals in Egypt, where meat and coarse wool production are the basic breeding objectives for Egyptian sheep. Therefore, evaluation of genetic diversity of sheep breeds is necessary for implementing the most suitable breed-region conservation programs. Animals from the three major Egyptian sheep breeds (Ossimi, Rahmani and Barki) were genotyped for 14 microsatellite markers. All loci tested were highly polymorphic. Various measures of genetic variation were calculated. The total observed number of alleles per microsatellite ranged from 6 to 12 for markers MAF65 and OarHH47, respectively. Estimates of effective number of alleles were between 3.11 for TGLA53 and 6.12 for OarHH47. The mean of polymorphism information content was 0.73. Overall gene diversity for all microsatellites analyzed was 0.66, 0.69 and 0.75 for Ossimi, Rahmani and Barki, respectively. For all breeds studied, estimates of observed heterozygosity were significantly lower than the expected heterozygosity. Average observed and expected heterozygosity estimates were 0.55 and 0.67, respectively. Significant departures from Hardy-Weinberg Equilibrium (HWE) due to heterozygote deficiency were observed for all the markers analyzed. The three breeds revealed significant deviation from HWE. The overall indicator of population subdivision (FST) was calculated to be 0.071; pointing out that about 7% of genetic diversity is due to genetic variation between breeds. Estimates of total inbreeding (FIT) and within-breed inbreeding (FIS) coefficients were 0.187 and 0.118, respectively. The estimates of pair-wise genetic differentiation were 0.039, 0.051 and 0.056 for Ossimi-Rahmani, Barki-Rahmani and Ossimi-Barki pairs, respectively. Measures of genetic distance between pairs of sheep breeds ranged from 0.423 to 0.615 for Rahmani-Ossimi and Barki-Ossimi sheep breed pairs. The results obtained in this study may be useful in sustainable breeding programs of Egyptian sheep breeds.