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Genetics, Vol 123, 465-476, Copyright © 1989
INVESTIGATIONS |
Enhancement of Escherichia coli Plasmid and Chromosomal Recombination by the Ref Function of Bacteriophage P1
C. S. Laufer, J. B. Hays, B. E. Windle, T. S. Schaefer, E. H. Lee, S. L. Hays and M. R. McClure
Department of Chemistry, University of Maryland Baltimore County, Catonsville, Maryland 21228 Present address: Department of Biology, Hood College, Frederick, Maryland 21701.
The Ref activity of phage P1 enhances recombination between two defective lacZ genes in the Escherichia coli chromosome (lac(-) X lac(-) recombination). Plasmid recombination, both lac(-) X lac(-) and tet(-) X tet(-), was measured by transformation of recA strains, and was also assayed by measurement of {beta}-galactosidase. The intracellular presence of recombinant plasmids was verified directly by Southern blotting. Ref stimulated recombination of plasmids in rec(+) and rec(BCD) cells by 3-6-fold, and also the low level plasmid recombination in recF cells. RecA-independent plasmid recombination, either very low level (recA cells) or high level (recB recC sbcA recA cells), was not stimulated. Ref stimulated both intramolecular and intermolecular plasmid recombination. Both normal and Ref-stimulated lac(-) X lac(-) chromosomal recombination, expected to be mostly RecBC-dependent in wild-type bacteria, were affected very little by a recF mutation. We have previously reported Ref stimulation of lac(-) X lac(-) recombination in recBC sbcB bacteria, a process known to be RecF-dependent. Chromosomal recombination processes thought to involve activated recombination substrates, e.g., Hfr conjugation, P1 transduction, were not elevated by Ref activity. We hypothesize that Ref acts by unknown mechanisms to activate plasmid and chromosomal DNA for RecA-mediated recombination, and that the structures formed are substrates for both RecF-dependent (plasmid, chromosomal) and Rec(BCD)-dependent (chromosomal) recombination pathways.
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