Delivery methods and compositions for nuclease-mediated genome engineering

A method of integrating one or more transgenes into a genome of an isolated population of hematopoietic stem cells or T-cells, the method comprising:

• introducing, into the population of hematopoietic stem cells or T-cells, (a) an adeno-associated type 6 (AAV6) donor vector comprising the one or more transgenes and (b) at least one non-naturally occurring nuclease, wherein the at least one nuclease is introduced in mRNA form and cleaves the genome of the population of hematopoietic stem cells or T-cells such that the one or more transgenes are integrated into the genome of the cell, wherein the donor vector is introduced into the cell prior to or after the at least one nuclease and further wherein
• (i) if the donor vector is introduced into the population of hematopoietic stem cells or T-cells before the at least one nuclease, the at least one nuclease is introduced into the cell 3 to 48 hours after the donor vector is introduced and;
• (ii) if the at least one nuclease is introduced before the donor vector, the donor vector is introduced into the cell within 4 hours after the at least one nuclease is introduced,
• such that at least 10% of the one or more transgenes are integrated into the genome of the population of hematopoietic stem cells or T-cells.