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Characterization of double-strand break-induced recombination: homology requirements and single-stranded DNA formation.
Mol Cell Biol. 1992 Feb; 12(2):563-75.MC

Abstract

In the yeast Saccharomyces cerevisiae, a double-strand chromosome break created by the HO endonuclease is frequently repaired in mitotically growing cells by recombination between flanking homologous regions, producing a deletion. We showed that single-stranded regions were formed on both sides of the double-strand break prior to the formation of the product. The kinetics of the single-stranded DNA were monitored in strains with the recombination-deficient mutations rad52 and rad50 as well as in the wild-type strain. In rad50 mutants, single-stranded DNA was generated at a slower rate than in the wild type, whereas rad52 mutants generated single-stranded DNA at a faster rate. Product formation was largely blocked in the rad52 mutant. In the rad50 rad52 double mutant, the effects were superimposed in that the exonucleolytic activity was slowed but product formation was blocked. rad50 appears to act before or at the same stage as rad52. We constructed strains containing two ura3 segments on one side of the HO cut site and one ura3 region on the other side to characterize how flanking repeats find each other. Deletions formed preterentially between the homologous regions closest to the double-strand break. By varying the size of the middle ura3 segment, we determined that recombination initiated by a double-strand break requires a minimum homologous length between 63 and 89 bp. In these competition experiments, the frequency of recombination was dependent on the length of homology in an approximately linear manner.

Authors+Show Affiliations

Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, Massachusetts 02254-9110.No affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

1732731

Citation

Sugawara, N, and J E. Haber. "Characterization of Double-strand Break-induced Recombination: Homology Requirements and Single-stranded DNA Formation." Molecular and Cellular Biology, vol. 12, no. 2, 1992, pp. 563-75.
Sugawara N, Haber JE. Characterization of double-strand break-induced recombination: homology requirements and single-stranded DNA formation. Mol Cell Biol. 1992;12(2):563-75.
Sugawara, N., & Haber, J. E. (1992). Characterization of double-strand break-induced recombination: homology requirements and single-stranded DNA formation. Molecular and Cellular Biology, 12(2), 563-75.
Sugawara N, Haber JE. Characterization of Double-strand Break-induced Recombination: Homology Requirements and Single-stranded DNA Formation. Mol Cell Biol. 1992;12(2):563-75. PubMed PMID: 1732731.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Characterization of double-strand break-induced recombination: homology requirements and single-stranded DNA formation. AU - Sugawara,N, AU - Haber,J E, PY - 1992/2/1/pubmed PY - 1992/2/1/medline PY - 1992/2/1/entrez SP - 563 EP - 75 JF - Molecular and cellular biology JO - Mol Cell Biol VL - 12 IS - 2 N2 - In the yeast Saccharomyces cerevisiae, a double-strand chromosome break created by the HO endonuclease is frequently repaired in mitotically growing cells by recombination between flanking homologous regions, producing a deletion. We showed that single-stranded regions were formed on both sides of the double-strand break prior to the formation of the product. The kinetics of the single-stranded DNA were monitored in strains with the recombination-deficient mutations rad52 and rad50 as well as in the wild-type strain. In rad50 mutants, single-stranded DNA was generated at a slower rate than in the wild type, whereas rad52 mutants generated single-stranded DNA at a faster rate. Product formation was largely blocked in the rad52 mutant. In the rad50 rad52 double mutant, the effects were superimposed in that the exonucleolytic activity was slowed but product formation was blocked. rad50 appears to act before or at the same stage as rad52. We constructed strains containing two ura3 segments on one side of the HO cut site and one ura3 region on the other side to characterize how flanking repeats find each other. Deletions formed preterentially between the homologous regions closest to the double-strand break. By varying the size of the middle ura3 segment, we determined that recombination initiated by a double-strand break requires a minimum homologous length between 63 and 89 bp. In these competition experiments, the frequency of recombination was dependent on the length of homology in an approximately linear manner. SN - 0270-7306 UR - https://www.unboundmedicine.com/medline/citation/1732731/Characterization_of_double_strand_break_induced_recombination:_homology_requirements_and_single_stranded_DNA_formation_ L2 - https://journals.asm.org/doi/10.1128/mcb.12.2.563-575.1992?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.org&rfr_dat=cr_pub=pubmed DB - PRIME DP - Unbound Medicine ER -