Numerous studies have demonstrated that triptolide induces cell cycle arrest and apoptosis in human cancer cell lines. However, triptolide-induced DNA damage and inhibition of DNA repair gene expression in human skin cancer cells has not previously been reported. We sought the effects of triptolide on DNA damage and associated gene expression in A375.S2 human malignant melanoma cells in vitro. Comet assay, DAPI staining and DNA gel electrophoresis were used for examining DNA damage and results indicated that triptolide induced a longer DNA migration smear based on single cell electrophoresis and DNA condensation and damage occurred based on the examination of DAPI straining and DNA gel electrophoresis. The real-time PCR technique was used to examine DNA damage and repair gene expression (mRNA) and results indicated that triptolide led to a decrease in the ataxia telangiectasia mutated (ATM), ataxia-telangiectasia and Rad3-related (ATR), breast cancer 1, early onset (BRCA-1), p53, DNA-dependent serine/threonine protein kinase (DNA-PK) and O6-methylguanine-DNA methyltransferase (MGMT) mRNA expression. Thus, these observations indicated that triptolide induced DNA damage and inhibited DNA damage and repair-associated gene expression (mRNA) that may be factors for triptolide-mediated inhibition of cell growth in vitro in A375.S2 cells.