Genetic Engineering Using Homologous Recombination¹

In the past few years, in vivo technologies have emerged that, due to their efficiency and simplicity, may one day replace standard genetic engineering techniques. Constructs can be made on plasmids or directly on the Escherichia coli chromosome from PCR products or synthetic oligonucleotides by homologous recombination. This is possible because bacteriophage-encoded recombination functions efficiently recombine sequences with homologies as short as 35 to 50 base pairs. This technology, termed recombineering, is providing new ways to modify genes and segments of the chromosome. This review describes not only recombineering and its applications, but also summarizes homologous recombination in E. coli and early uses of homologous recombination to modify the bacterial chromosome. Finally, based on the premise that phage-mediated recombination functions act at replication forks, specific molecular models are proposed.