DNA polymerase β (DNA polymerase beta (POLB)) belongs to a member of the DNA polymerase X family, mainly involved in various biological metabolic processes, such as eukaryotic DNA replication, DNA damage repair, gene recombination, and cell cycle regulation. In this study, the muscle development-related gene POLB was screened by selection signature and RNA-seq analysis and then validated for the proliferation and apoptosis of bovine primary myocytes. It was also found that overexpression of the POLB gene had a pro-apoptosis effect, but interfering with the expression of the gene had no significant effect on cells. Then, the analysis of related apoptotic genes revealed that POLB overexpression affected CASP9 gene expression.