Abstract：To establish two-dimensional gel electrophoresis methods (2-DE) for porcine skeletal muscle proteome, the experimental conditions of 2-DE was optimized，and various extraction methods，different loading quantities and pH IPG strips，different adhesive concentration and gel staining were studied. The results showed that the process of grinding in liquid nitrogen and following by ultrasonication was more appropriate than that of grinding in liquid nitrogen, which could obtained better 2-DE map for low nucleic acid contamination and more protein spots. Compared with different pH IPG strips, 18 cm pH 4~7 IPG strips were better than pH 3~10 NL. For silver staining of 18 cm pH 4~7 IPG strip, the suitable loading quantity was 150 μg. Furthermore, the reproducibility of three gels in same sample was more 70%. Meanwhile, the common problems occurring in 2-DE experiments were analyzed. The results indicated that the 2-DE maps of porcine skeletal muscle were obtained with high resolution and reproducibility, and can be used for the further proteome analysis.
