Genome engineering

The present invention provides an in vitro or ex-vivo method of altering target DNA in a stem cell wherein the stem cell is genetically modified to include in the genomic DNA of the cell a nucleic acid encoding a Cas9 enzyme that forms a co-localization complex with a guide RNA complementary to the target DNA and that cleaves the target DNA in a site specific manner comprising (a) introducing into the stem cell a first foreign nucleic acid encoding a guide RNA complementary to the target DNA and which guides the enzyme to the target DNA, wherein the guide RNA and the Cas9 enzyme are members of a co-localization complex for the target DNA, introducing into the stem cell a donor nucleic acid sequence, wherein the guide RNA is expressed, wherein the guide RNA and the Cas9 enzyme co-localize to the target DNA, the Cas9 enzyme cleaves the target DNA and the donor nucleic acid is inserted into the target DNA to produce altered DNA in the stem cell, and wherein the stem cell is not a totipotent or germline human cell.