Patent ID: 8609374

Claim:
A method of assembling a plurality of double-stranded DNA fragments into DNA molecules in a single in vitro recombination reaction comprising contacting in vitro the plurality of double-stranded DNA fragments with a bacterial extract from a RecA deficient bacterial strain so as to assemble the plurality of DNA fragments into DNA molecules, wherein each DNA fragment has a 3′ end and a 5′ end, wherein fragments assemble with each other when the 5′ end of one fragment has 20 bp to 100 bp that are homologous with the 3′ end of another fragment, wherein heterologous sequences, when present, of up to 1000 bp that flank the homologous region are removed prior to assembling the DNA fragments into DNA molecules, and wherein the in vitro recombination reaction is performed without the addition of any of an exogenous DNA restriction enzyme, an exogenous DNA modifying enzyme, or an exogenous DNA ligase.