The invention discloses a method for separating, identifying and proliferating myoblasts of small-tailed han sheep, which comprises the following steps: (1) taking and treating
skeletal muscle tissues: selecting healthy small-tailed han sheep, collecting the
skeletal muscle tissues under sterile conditions, washing and
cutting into pieces to obtain tissue blocks; (2) primary myoblast isolated culture: digesting, filtering and centrifuging the obtained tissue block, and culturing; (3) myoblast purification: purifying the cultured primary myoblasts to obtain purified myoblasts; (4) identifying myoblasts: identifying the obtained purified myoblasts; and (5) proliferation of myoblasts: after identification is completed, carrying out
cell proliferation on the purified myoblasts obtained in the step (3) by utilizing a proliferation culture medium containing
vitamin E. According to the method provided by the invention, high-purity and high-activity myoblasts can be obtained;
vitamin E is utilized, so that the proliferation capacity of myoblasts can be remarkably improved, and the
cell differentiation function of the myoblasts is not influenced.