Patent ID: 8143015
Filing Date: 2012-03-27
Classification: A61K,A61P,C12N

Abstract:
1. A method of cleaving a target recognition site in double-stranded DNA in a non-human cell or an isolated human cell, wherein said target recognition site has a sequence that differs by at least one nucleotide modification from the wild-type I-CreI meganuclease recognition sequence of SEQ ID NO: 4: wherein said target recognition site has a sequence that comprises a substitution at a position corresponding to position −3 of the I-CreI recognition site of SEQ ID NO: 4, wherein the nucleotide corresponding to position −3 of the I-CreI meganuclease recognition sequence of SEQ ID NO: 4 has been altered to an A or a C, the method comprising: (a) introducing into a cell (i) a recombinant meganuclease, or (ii) a nucleic acid that encodes a recombinant meganuclease and causes expression of the recombinant meganuclease in said cell, wherein the recombinant meganuclease comprises a polypeptide having at least 85% sequence identity to residues 2-153 of the I-CreI meganuclease of SEQ ID NO: 1, and which differs from residues 2-153 of the I-CreI meganuclease of SEQ ID NO: 1 by a specificity-altering amino acid modification at a position corresponding to position 24 of SEQ ID NO: 1; and wherein (b) contacting the recombinant meganuclease with the double-stranded DNA, whereby the meganuclease cleaves the target recognition site.