DNA double-strand breaks (DSBs) are the principal cytotoxic lesions caused by many exogenous and endogenous factors. In response to DSBs, cells have evolved complex and highly conserved systems, which mainly consist of homologous recombination repair (HR) and non-homologous end joining (NHEJ) pathways, to effectively repair the lethal lesions. The two pathways play crucial roles in preserving the genomic integrity. Here, we provide an overview of detailed process, concerned molecules and regulatory factors in these pathways. Accumulated knowledge of DSBs repair may offer opportunities to develop more effective treatments for cancer.