Abstract: Neonatal pig skeletal muscle satellite cells were obtained by the two step method of collegenaseⅠand trypsin, and were subject to primary as well as secondary culture in vitro. Morphological observation and growth curve were used to evaluate the proliferative and differentiation ability of skeletal muscle satellite cells. The cells were identified with RT-PCR and cellular immunochemical stain. The results showed that the two step method with collegenaseⅠ and trypsin was suitable for collection of skeletal muscle satellite cells. The cells showed strong proliferative ability in the proliferative media and could form myotubes in differentiation media. In all, a method for culture, identification of the neonatal pig skeletal muscle satellite cell was established, which laying a foundation for the establishment of the pig skeletal muscle satellite cell line.