[Recombineering and its application].

ABSTRACT

Driven by the need of functional genomics, a homologous recombination-based, highly efficient genetic engineering system that termed "recombineering" has recently been developed. Recombineering has been defined as a genetic engineering with phage-encoded recombination function that utilizes short homologies, a convenient term to describe homologous-dependent, recombination-mediated, genetic engineering. The bacteriophage lambda Red recombination system has critical differences from standard E. coli RecA-dependent recombination pathway. The phage systems have unique advantage in that they can catalyze efficient recombination with very short regions of sequence homology (< 50 bp). Recombineering does not require construction of plasmid or phage DNA intermediates containing the appropriately pre-engineered homology segment. All that is required in vitro is the synthesis of standard oligonucleotides or construction of PCR products that provide the homology. Importantly, they function even in the absence of RecA. These approaches do not rely on the presence of suitable restriction site, and can be used to insert, delete, clone or substitute genomic DNA sequences at any desired position on a target molecule in Escherichia Coli. Recombineering also facilitates many kinds of genomic experiments difficult to be carried out. In this article, the bacteriophage lambda Red recombinase system, the progression and applications of this powerful new technique are reviewed according to the data published recently.