The authors present a method to enhance homology-directed repair (HDR) in human cells using timed delivery of Cas9-guide RNA ribonucleoprotein (RNP) complexes. By synchronizing cells at specific phases of the cell cycle, they demonstrate that HDR efficiency can be significantly increased compared to unsynchronized cells. In HEK293T cells, HDR frequencies up to 38% were achieved, with no detectable off-target editing. The approach is also effective in human primary neonatal fibroblasts and human embryonic stem cells, showing robust HDR-mediated genome editing at levels comparable to HEK293T cells. The study provides a simple and highly effective strategy for enhancing site-specific genome engineering in human cells.The authors present a method to enhance homology-directed repair (HDR) in human cells using timed delivery of Cas9-guide RNA ribonucleoprotein (RNP) complexes. By synchronizing cells at specific phases of the cell cycle, they demonstrate that HDR efficiency can be significantly increased compared to unsynchronized cells. In HEK293T cells, HDR frequencies up to 38% were achieved, with no detectable off-target editing. The approach is also effective in human primary neonatal fibroblasts and human embryonic stem cells, showing robust HDR-mediated genome editing at levels comparable to HEK293T cells. The study provides a simple and highly effective strategy for enhancing site-specific genome engineering in human cells.