Patent ID: 8586362

Claim:
A method of repairing a DNA double-strand break in vitro, comprising the steps of: (a) providing a duplex DNA molecule wherein the molecule has a double-strand break and wherein the molecule is not super-coiled; (b) providing a single-stranded DNA targeting fragment, wherein 3′ end of the targeting fragment is homologous to at least 15 nucleotides of known sequence in the DNA molecule of step (a) and wherein sequence of the double-stand break is within the targeting fragment; (c) adding RecA protein or RecA protein homologue to the targeting fragment; (d) adding single-stranded DNA binding protein (SSB) or SSB homologue; (e) adding the DNA molecule of step (a) to the mixture resulting from step (d) and incubating until a strand invasion of the targeting fragment into the DNA molecule of step (a) has occurred; and (f) adding DNA polymerase and dNTPs, wherein the proteins of steps (c) and (d) and the polymerase of step (f) are the only strand-repair proteins added to the system, and incubating the resulting mixture until the invading strand has extended, wherein the strand comprises sequences from the targeting fragment and the DNA molecule of step (a).