The exponential growth of the human population is limiting the wildlife habitat all around the word. In recent years habitat loss and fragmentation is one of the main reasons that threats the wild life species. The Grévy’s zebra (Equus grevyi) is the most endangered member of Zebras. Their historical range was previously from north Ethiopia to southwest Somalia and to northern Kenya. Currently they are distributed only in fragmented habitats in central and eastern part of Ethiopia and in the north of Kenya. They are listed as endangered in the IUCN red list, as their population has declined 68% in 27 years. There are very few studies on genetic structure of this species, and investigating the genetic connection between different populations is needed. Molecular markers are one of the best tools to understand the level of fragmentation, population bottlenecks or potential inbreeding. In this study, the population structure of Ethiopian zebra population from Alledeghi Wildlife Reserve (WR) and Sarite area was studied using non-invasively obtained fecal samples collected during 2001-2011. This study analyzes genetic variation at 10 microsatellite loci and a 350-bp fragment of the mitochondrial DNA control region. The results showed that the genetic diversity is very low between the populations (π=0.00116 for Alledeghi WR and π=0 for Sarite population). The population of Alledeghi WR is probably isolated from the population of Sarite, as they don’t share any haplotypes. As the population of Alledeghi WR is separated from the ones from Sarite and Kenya, applying more conservational programs in this area is needed to protect the genetic diversity of the Grévy’s zebras in this area.