To test the applicability of the All-in-One Cas9D10A nickase vector system and screening approaches in other cell lines, we targeted the DDR genes MDC1, 53BP1 and RIF1 in human aneuploid osteosarcoma (U2OS) cells and haploid chronic myelogenous leukaemia derived (HAP1) cells. The gene discussed is RIF1; the disease is osteosarcoma.