Sheep satellite cells more than satellite cells of the rat and mouse are similar to human satellite cells. These cells are widely used in the modeling and treatment of diseases like heart insufficiency, neurological diseases, muscular dystrophy, cerebral cell transplantation for the treatment of migraines, screening, and the production of new drugs. This study was aimed to isolate and culture primary satellite cells (PSCs) obtained from sheep fetus, and perform clonal expansion of transfected PSCs. Skeletal muscle tissues of hind limbs were collected from sheep fetuses obtained from a local abattoir. After enzymatic digestion, flasks were replaced after 3 hours to isolate non-myogenic cells, such as fibroblasts. After six days, the cells were differentiated to myoblasts. Using a differentiation medium containing the horse serum, myotube cells were observed in the flask, indicating that the cultured cells were satellite cells. The mRNA expression of the PAX7 gene was used to confirm the presence of satellite cells. In addition, the results showed that satellite cells grow in a culture medium containing 5% FBS without differentiation, while 10% FBS initiates their differentiation.