Related Experiment Video
Updated: Aug 14, 2026

Recombineering Homologous Recombination Constructs in Drosophila
Published on: July 13, 2013
In vitro reconstitution of homologous recombination reactions
1Division of Biological Sciences, University of California, Davis 95616.
Abstract:
The proteins essential to homologous recombination in E. coli have been purified and their individual activities have been identified, permitting biochemical reconstitution of steps that comprise the cellular recombination process. This review focuses on the biochemical events responsible for the initiation and homologous pairing steps of genetic recombination. The properties of an in vitro recombination reaction that requires the concerted action of recA, recBCD, and SSB proteins and that is stimulated by the recombination hotspot, Chi(chi), are described. The recBCD enzyme serves as the initiator of this reaction; its DNA helicase activity produces single-stranded DNA that is used by the recA protein to promote homologous pairing and DNA strand invasion of supercoiled (recipient) DNA. The SSB protein acts to trap the single-stranded DNA produced by recBCD enzyme and to facilitate pairing by the recA protein. The chi regulatory sequence acts in cis by attenuating the nuclease, but not the helicase, activity of recBCD enzyme. This attenuation assures the preservation of ssDNA produced by the DNA helicase activity and is responsible for the simulation in vitro and, presumably, in vivo. The attenuation of nuclease activity by chi results in the loss or functional inactivation of the recD subunit.
Related Concept Videos
Fixing Double-strand Breaks
Homologous Recombination
Conservative Site-specific Recombination and Phase Variation
The recognition sites for Cre recombinase called LoxP...
DNA Isolation
Crossing Over

