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. 1989 Nov;123(3):465–476. doi: 10.1093/genetics/123.3.465

Enhancement of Escherichia Coli Plasmid and Chromosomal Recombination by the Ref Function of Bacteriophage P1

C S Laufer 1, J B Hays 1, B E Windle 1, T S Schaefer 1, E H Lee 1, S L Hays 1, M R McClure 1
PMCID: PMC1203818  PMID: 2557261

Abstract

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 β-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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Selected References

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  1. Bachmann B. J. Pedigrees of some mutant strains of Escherichia coli K-12. Bacteriol Rev. 1972 Dec;36(4):525–557. doi: 10.1128/br.36.4.525-557.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blanar M. A., Sandler S. J., Armengod M. E., Ream L. W., Clark A. J. Molecular analysis of the recF gene of Escherichia coli. Proc Natl Acad Sci U S A. 1984 Aug;81(15):4622–4626. doi: 10.1073/pnas.81.15.4622. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Csonka L. N., Clark A. J. Construction of an Hfr strain useful for transferring recA mutations between Escherichia coli strains. J Bacteriol. 1980 Jul;143(1):529–530. doi: 10.1128/jb.143.1.529-530.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Doherty M. J., Morrison P. T., Kolodner R. Genetic recombination of bacterial plasmid DNA. Physical and genetic analysis of the products of plasmid recombination in Escherichia coli. J Mol Biol. 1983 Jul 5;167(3):539–560. doi: 10.1016/s0022-2836(83)80097-7. [DOI] [PubMed] [Google Scholar]
  5. Fishel R. A., James A. A., Kolodner R. recA-independent general genetic recombination of plasmids. Nature. 1981 Nov 12;294(5837):184–186. doi: 10.1038/294184a0. [DOI] [PubMed] [Google Scholar]
  6. Gillen J. R., Willis D. K., Clark A. J. Genetic analysis of the RecE pathway of genetic recombination in Escherichia coli K-12. J Bacteriol. 1981 Jan;145(1):521–532. doi: 10.1128/jb.145.1.521-532.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gottesman M. E., Yarmolinsky M. B. Integration-negative mutants of bacteriophage lambda. J Mol Biol. 1968 Feb 14;31(3):487–505. doi: 10.1016/0022-2836(68)90423-3. [DOI] [PubMed] [Google Scholar]
  8. Hays J. B., Boehmer S. Antagonists of DNA gyrase inhibit repair and recombination of UV-irradiated phage lambda. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4125–4129. doi: 10.1073/pnas.75.9.4125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Huskey R. J. Multiplicity reactivation as a test for recombination function. Science. 1969 Apr 18;164(3877):319–320. doi: 10.1126/science.164.3877.319. [DOI] [PubMed] [Google Scholar]
  10. James A. A., Morrison P. T., Kolodner R. Genetic recombination of bacterial plasmid DNA. Analysis of the effect of recombination-deficient mutations on plasmid recombination. J Mol Biol. 1982 Sep 25;160(3):411–430. doi: 10.1016/0022-2836(82)90305-9. [DOI] [PubMed] [Google Scholar]
  11. Kaiser K., Murray N. E. Physical characterisation of the "Rac prophage" in E. coli K12. Mol Gen Genet. 1979 Sep;175(2):159–174. doi: 10.1007/BF00425532. [DOI] [PubMed] [Google Scholar]
  12. Kobayashi I., Ikeda H. Formation of recombinant DNA of bacteriophage lambda by recA function of Escherichia coli without duplication, transcription, translation, and maturation. Mol Gen Genet. 1977 Jun 24;153(3):237–245. doi: 10.1007/BF00431589. [DOI] [PubMed] [Google Scholar]
  13. Konrad E. B. Method for the isolation of Escherichia coli mutants with enhanced recombination between chromosomal duplications. J Bacteriol. 1977 Apr;130(1):167–172. doi: 10.1128/jb.130.1.167-172.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Korba B. E., Hays J. B. Novel mutants of Escherichia coli that produce recombinogenic lesions in DNA. II. Properties of recombinogenic lambda phages grown on bacteria carrying arl mutations. J Mol Biol. 1980 May 25;139(3):473–489. doi: 10.1016/0022-2836(80)90142-4. [DOI] [PubMed] [Google Scholar]
  15. Kushner S. R., Nagaishi H., Templin A., Clark A. J. Genetic recombination in Escherichia coli: the role of exonuclease I. Proc Natl Acad Sci U S A. 1971 Apr;68(4):824–827. doi: 10.1073/pnas.68.4.824. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Low B. Restoration by the rac locus of recombinant forming ability in recB - and recC - merozygotes of Escherichia coli K-12. Mol Gen Genet. 1973 Apr 12;122(2):119–130. doi: 10.1007/BF00435185. [DOI] [PubMed] [Google Scholar]
  17. Mandel M., Higa A. Calcium-dependent bacteriophage DNA infection. J Mol Biol. 1970 Oct 14;53(1):159–162. doi: 10.1016/0022-2836(70)90051-3. [DOI] [PubMed] [Google Scholar]
  18. Signer E. R., Weil J. Recombination in bacteriophage lambda. I. Mutants deficient in general recombination. J Mol Biol. 1968 Jul 14;34(2):261–271. doi: 10.1016/0022-2836(68)90251-9. [DOI] [PubMed] [Google Scholar]
  19. Smith T. A., Hays J. B. Repair and recombination of nonreplicating UV-irradiated phage DNA in E. coli II. Stimulation of RecF-dependent recombination by excision repair of cyclobutane pyrimidine dimers and of other photoproducts. Mol Gen Genet. 1985;201(3):393–401. doi: 10.1007/BF00331329. [DOI] [PubMed] [Google Scholar]
  20. Taylor A. F., Schultz D. W., Ponticelli A. S., Smith G. R. RecBC enzyme nicking at Chi sites during DNA unwinding: location and orientation-dependence of the cutting. Cell. 1985 May;41(1):153–163. doi: 10.1016/0092-8674(85)90070-4. [DOI] [PubMed] [Google Scholar]
  21. Windle B. E., Hays J. B. A phage P1 function that stimulates homologous recombination of the Escherichia coli chromosome. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3885–3889. doi: 10.1073/pnas.83.11.3885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Windle B. E., Laufer C. S., Hays J. B. Sequence and deletion analysis of the recombination enhancement gene (ref) of bacteriophage P1: evidence for promoter-operator and attenuator-antiterminator control. J Bacteriol. 1988 Oct;170(10):4881–4889. doi: 10.1128/jb.170.10.4881-4889.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Windle B. E. Phage lambda and plasmid expression vectors with multiple cloning sites and lacZ alpha-complementation. Gene. 1986;45(1):95–99. doi: 10.1016/0378-1119(86)90136-8. [DOI] [PubMed] [Google Scholar]
  24. Zagursky R. J., Hays J. B. Expression of the phage lambda recombination genes exo and bet under lacPO control on a multi-copy plasmid. Gene. 1983 Sep;23(3):277–292. doi: 10.1016/0378-1119(83)90018-5. [DOI] [PubMed] [Google Scholar]

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