Evidence for the double-strand break repair model of bacteriophage lambda recombination.

Abstract

We have obtained evidence for the repair of double-strand gaps promoted by the Red function of bacteriophage lambda. A double-strand gap was made in one of the two regions of homology in an inverted orientation on a plasmid DNA molecule. The gapped plasmid was introduced into Escherichia coli cells expressing the red alpha (exo) and red beta (bet) genes of lambda. The gap was repaired by DNA synthesis copying an intact duplex. This gap repair was sometimes accompanied by reciprocal recombination (crossing over). The gap stimulated recombination about 100-fold. Our results are compatible with previous proposals that lambda homologous recombination involves the following early steps: (i) generation of double-stranded ends by the packaging machinery or by the replication machinery; (ii) production of a single-stranded tail with a 3'-hydroxyl end by 5'----3' degradation by lambda exonuclease (red alpha gene product); (iii) pairing of the single-stranded tail with a complementary strand from a homologous duplex with the help of beta protein (red beta gene product); (iv) priming of DNA synthesis at this 3'-hydroxyl end to copy the second DNA molecule.