The differential sperm proteomic analysis of high- and low-quality semen from Niangya yaks, revealed 6 proteins (PSMC5, PSMD8, PSMB3, HSP90AA1, UGP2 and HSPB1), were mainly concentrated in energy production and metabolism, might play important roles in semen quality, which could serve as candidates for the selection and breeding of Niangya yaks.