Translation of the CRISPR/Cas9 system to human therapeutics holds high promise. Specificity remains a concern, however, especially when modifying stem cell populations. We show that existing rationally-engineered Cas9 high fidelity variants have reduced on-target activity using the therapeutically relevant ribonucleoprotein (RNP) delivery method. Therefore, we devised an unbiased bacterial screen to isolate variants that retain activity in the RNP format. Introduction of a single point mutation, R691A (HiFi Cas9), retained high on-target activity while reducing off-target editing. HiFi Cas9 induces robust AAV6-mediated gene targeting at five therapeutically-relevant loci ( HBB, IL2RG, CCR5, HEXB, TRAC) in human CD34 + hematopoietic stem and progenitor cells (HSPCs) as well as primary T-cells. We also show that the HiFi Cas9 mediates high-level correction of the sickle cell disease (SCD)-causing Glu6Val mutation in SCD patient derived HSPCs. We anticipate that HiFi Cas9 will have wide utility for both basic science and therapeutic genome editing applications.