Identification of the crossover site during FLP-mediated recombination in the Saccharomyces cerevisiae plasmid 2 microns circle.

M. McLeod, S. Craft, James Broach

Research output: Contribution to journalArticle

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Abstract

The FLP protein of the Saccharomyces cerevisiae plasmid 2 microns circle catalyzes site-specific recombination between two repeated segments present on the plasmid. In this paper we present results of experiments we performed to define more precisely the features of the FLP recognition target site, which we propose to designate FRT, and to determine the actual recombination crossover point in vivo. We found that essential sequences for the recombination event are limited to an 8-base-pair core sequence and two 13-base-pair repeated units immediately flanking it. This is the region identified as the FLP binding site in vitro and at which FLP protein promotes specific single-strand cleavages (B. J. Andrews, G. A. Proteau, L. G. Beatty, and P. D. Sadowski, Cell 40:795-803, 1985; J. F. Senecoff, R. C. Bruckner, and M. M. Cox, Proc. Natl. Acad. Sci. USA 82:7270-7274, 1985). Mutations within the core domain can be suppressed by the presence of the identical mutation in the chromatid with which it recombines. However, mutations outside the core are not similarly suppressed. We found that strand exchange during FLP recombination occurs most of the time within the core region, proceeding through a heteroduplex intermediate. Finally, we found that most FLP-mediated events are reciprocal exchanges and that FLP-catalyzed gene conversions occur at low frequency. The low level of gene conversion associated with FLP recombination suggests that it proceeds by a breakage-joining reaction and that the two events are concerted.

Original languageEnglish (US)
Pages (from-to)3357-3367
Number of pages11
JournalMolecular and cellular biology
Volume6
Issue number10
DOIs
StatePublished - Jan 1 1986

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Genetic Recombination
Saccharomyces cerevisiae
Plasmids
Gene Conversion
Base Pairing
Mutation
Saccharomyces cerevisiae Proteins
Chromatids
Binding Sites
Proteins

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

Cite this

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title = "Identification of the crossover site during FLP-mediated recombination in the Saccharomyces cerevisiae plasmid 2 microns circle.",
abstract = "The FLP protein of the Saccharomyces cerevisiae plasmid 2 microns circle catalyzes site-specific recombination between two repeated segments present on the plasmid. In this paper we present results of experiments we performed to define more precisely the features of the FLP recognition target site, which we propose to designate FRT, and to determine the actual recombination crossover point in vivo. We found that essential sequences for the recombination event are limited to an 8-base-pair core sequence and two 13-base-pair repeated units immediately flanking it. This is the region identified as the FLP binding site in vitro and at which FLP protein promotes specific single-strand cleavages (B. J. Andrews, G. A. Proteau, L. G. Beatty, and P. D. Sadowski, Cell 40:795-803, 1985; J. F. Senecoff, R. C. Bruckner, and M. M. Cox, Proc. Natl. Acad. Sci. USA 82:7270-7274, 1985). Mutations within the core domain can be suppressed by the presence of the identical mutation in the chromatid with which it recombines. However, mutations outside the core are not similarly suppressed. We found that strand exchange during FLP recombination occurs most of the time within the core region, proceeding through a heteroduplex intermediate. Finally, we found that most FLP-mediated events are reciprocal exchanges and that FLP-catalyzed gene conversions occur at low frequency. The low level of gene conversion associated with FLP recombination suggests that it proceeds by a breakage-joining reaction and that the two events are concerted.",
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Identification of the crossover site during FLP-mediated recombination in the Saccharomyces cerevisiae plasmid 2 microns circle. / McLeod, M.; Craft, S.; Broach, James.

In: Molecular and cellular biology, Vol. 6, No. 10, 01.01.1986, p. 3357-3367.

Research output: Contribution to journalArticle

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T1 - Identification of the crossover site during FLP-mediated recombination in the Saccharomyces cerevisiae plasmid 2 microns circle.

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AU - Craft, S.

AU - Broach, James

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N2 - The FLP protein of the Saccharomyces cerevisiae plasmid 2 microns circle catalyzes site-specific recombination between two repeated segments present on the plasmid. In this paper we present results of experiments we performed to define more precisely the features of the FLP recognition target site, which we propose to designate FRT, and to determine the actual recombination crossover point in vivo. We found that essential sequences for the recombination event are limited to an 8-base-pair core sequence and two 13-base-pair repeated units immediately flanking it. This is the region identified as the FLP binding site in vitro and at which FLP protein promotes specific single-strand cleavages (B. J. Andrews, G. A. Proteau, L. G. Beatty, and P. D. Sadowski, Cell 40:795-803, 1985; J. F. Senecoff, R. C. Bruckner, and M. M. Cox, Proc. Natl. Acad. Sci. USA 82:7270-7274, 1985). Mutations within the core domain can be suppressed by the presence of the identical mutation in the chromatid with which it recombines. However, mutations outside the core are not similarly suppressed. We found that strand exchange during FLP recombination occurs most of the time within the core region, proceeding through a heteroduplex intermediate. Finally, we found that most FLP-mediated events are reciprocal exchanges and that FLP-catalyzed gene conversions occur at low frequency. The low level of gene conversion associated with FLP recombination suggests that it proceeds by a breakage-joining reaction and that the two events are concerted.

AB - The FLP protein of the Saccharomyces cerevisiae plasmid 2 microns circle catalyzes site-specific recombination between two repeated segments present on the plasmid. In this paper we present results of experiments we performed to define more precisely the features of the FLP recognition target site, which we propose to designate FRT, and to determine the actual recombination crossover point in vivo. We found that essential sequences for the recombination event are limited to an 8-base-pair core sequence and two 13-base-pair repeated units immediately flanking it. This is the region identified as the FLP binding site in vitro and at which FLP protein promotes specific single-strand cleavages (B. J. Andrews, G. A. Proteau, L. G. Beatty, and P. D. Sadowski, Cell 40:795-803, 1985; J. F. Senecoff, R. C. Bruckner, and M. M. Cox, Proc. Natl. Acad. Sci. USA 82:7270-7274, 1985). Mutations within the core domain can be suppressed by the presence of the identical mutation in the chromatid with which it recombines. However, mutations outside the core are not similarly suppressed. We found that strand exchange during FLP recombination occurs most of the time within the core region, proceeding through a heteroduplex intermediate. Finally, we found that most FLP-mediated events are reciprocal exchanges and that FLP-catalyzed gene conversions occur at low frequency. The low level of gene conversion associated with FLP recombination suggests that it proceeds by a breakage-joining reaction and that the two events are concerted.

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