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The cryopreservation of sperm could affect the expression of heat-shock proteins (HSPs) which involved in the spermatogenesis and sperm quality in mammals.The present study was to investigate the relation between the 70 kDa heat-shock protein (HSP70) expression level and QinChuan bull sperm motility during the process of freezing-thawing.We assessed sperm motility, morphology, viability and concentration of semen at three different stages (fresh, after the equilibrium and frozen-thawed).We also detected the gene expressions of HSP70 at the three stages.The characteristics of the fresh semen including sperm motility, morphology, viability and concentration were evaluated.After the equilibrium and frozen-thawed, sperm quality was also assessed.Total RNA was extracted from ejaculate samples at three different stages (fresh, after the equilibrium and frozen-thawed) respectively, using the improved hot Trizol method.First-strand eDNA synthesis from RNA was catalyzed by quantseript RT Kit, and eDNA was used to produce and amplify HSP70 gene.The differential gene expressions of HSP70 at three different stages were detected according to real-time fluorescent quantitative PCR.The motility and HSP70 gene expression levels of sperm at three stages were significant difference (P < 0.05).Both the motility and the HSP70 gene expression in fresh sperm were the highest, and after frozen-thawed sperm were the lowest.The HSP70 gene expression in bull sperm gradually reduced following the process of freezing-thawing.The gene expression of HSP70 was positively related to the sperm motility.Therefore, the gene expression level of HSP70 could be considered as a potential marker to predict the quality of bull sperm.