Abstract
Homology-facilitated illegitimate recombination has been described in three naturally competent bacterial species. It permits integration of small linear DNA molecules into the chromosome by homologous recombination at one end of the linear DNA substrate, and illegitimate recombination at the other end. We report that homology-facilitated illegitimate recombination also occurs in Escherichia coli during conjugation with small non-replicative plasmids, but at a low frequency of 3×10−10 per recipient cell. The fate of linear DNA in E. coli is either RecBCD-dependent degradation, or circularisation by ligation, and integration into the chromosome by single crossing-over. We also report that the observed single crossing-overs are recA-dependent, but essentially recBCD, and recFOR independent. This suggests that other, still unknown, proteins may act as mediator for the loading of RecA on DNA during single crossing-over recombination in E. coli.
Highlights
Homology-facilitated illegitimate recombination (HFIR) is a hybrid recombination reaction whereby a linear DNA molecule integrates into the bacterial chromosome during natural transformation
RecJ is a 59 to 39 single-strand-specific DNA exonuclease [5,6], which might degrade the 59 single strand extremity required for the illegitimate recombination event in A. baylyi
We report the development of a genetic screen to detect HFIR events in E. coli, following DNA conjugation with small, nonreplicative plasmids
Summary
Homology-facilitated illegitimate recombination (HFIR) is a hybrid recombination reaction whereby a linear DNA molecule integrates into the bacterial chromosome during natural transformation. Up to now, it has been reported in the three naturally competent bacterial species Streptococcus pneumoniae [1], Acinetobacter baylyi [2] and Pseudomonas stutzeri [3]. In HFIR, one end only of the linear incoming DNA molecule shares homology with the resident chromosome. Homologous recombination at this end is associated with an illegitimate recombination event within the nonhomologous region of the linear fragment (see Figure 1A). RecJ is a 59 to 39 single-strand-specific DNA exonuclease [5,6], which might degrade the 59 single strand extremity required for the illegitimate recombination event in A. baylyi
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