The labe free proteomics technology and 4D labe free phosphorylation proteomics technology was used to systematically analyse the protein expression regulatory mechanisms of muscle tenderness. 59 differentially expressed proteins were screened by proteomic data analysis. Phosphorylated proteomic analysis showed 681 modified peptide levels were changed, of which 235 modified peptide levels corresponded to 132 proteins up-regulated and 446 modified peptide levels corresponded to 253 proteins down-regulated. Then, the two-omics analysis further predicted that the regulatory mechanism of tenderness was mainly based on glycolysis, regulating mitochondrial autophagy, apoptosis, AMPK and HIF-1 signaling pathway to regulate muscle tenderness, which was specifically manifested in the modulation of Ca