Propofol is one of the most widely clinically used intravenous anesthetic, and it induces apoptosis in human and murine leukemia cell lines. Yet, whether propofol causes DNA damage and affects the mRNA expression of repair-associated genes in cancer cells remains undetermined. In the present study, we investigated the effects of propofol on DNA damage and associated mRNA gene expression in RAW264.7 cells. Comet assay and DNA gel electrophoresis were used to evaluate DNA damage in RAW264.7 cells and propofol-inhibited cell growth in vitro. The results revealed a longer DNA tail and DNA fragmentation. Real-time PCR assay was used to examine mRNA gene expression of DNA damage and DNA repair-associated genes. Following exposure to propofol for 48 h, a decrease in the mRNA expression of DNA-PK, BRCA1, MGMT and p53 was noted in the RAW264.7 cells. Results from the western blotting indicated that p53, MGMT, 14-3-3-σ, BRCA1 and MDC1 proteins were decreased while p-p53 and p-H2A.X(S140) were increased in the RAW264.7 cells following exposure to propofol. In conclusion, exposure to propofol caused DNA damage and inhibited mRNA expression and protein levels of repair-associated genes in RAW264.7 cells.