The invention relates to a method for separation and purification, long-term passage cultivation,
cryopreservation and
recovery of sheep spermatogonia stem cells. The method includes the steps that various solutions are prepared, 2-3 g of tissue blocks of the seminiferous
tubule are taken out from the testis of a sheep of 3-4 mouths of age, single-
cell suspension is prepared through enzymolysis and
digestion of two steps, purified spermatogonia stem cells are separated according to the difference adherence method, the long-term passage cultivation of the sheep spermatogonia stem cells is achieved through an SSCM culture solution, and the sheep spermatogonia stem cells are cryopreserved and recovered after the long-term passage cultivation. By the technical scheme, the sheep spermatogonia stem cells with high purity can be obtained without using specific sheep spermatogonia
stem cell antibodies, a long-term passage cultivation
system of the sheep spermatogonia stem cells is built, and the method for preserving the sheep spermatogonia stem cells for a long term through a
liquid nitrogen freezing method is successfully provided. The sheep spermatogonia stem cells achieving separation, purification and passage according to the method are cultivated at least for more than 40 generations, and the longest cultivation period reaches four years if the
cryopreservation time is included.