scholarly journals Analysis of BRCA1 Variants in Double-Strand Break Repair by Homologous Recombination and Single-Strand Annealing

2013 ◽  
Vol 34 (3) ◽  
pp. 439-445 ◽  
Author(s):  
William I. Towler ◽  
Jie Zhang ◽  
Derek J. R. Ransburgh ◽  
Amanda E. Toland ◽  
Chikashi Ishioka ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Strand Break ◽  
Double Strand Break ◽  
Single Strand ◽  
Double Strand Break Repair ◽  
Single Strand Annealing ◽  
Break Repair ◽  
Strand Annealing

scholarly journals FANCA Promotes DNA Double-Strand Break Repair by Catalyzing Single-Strand Annealing and Strand Exchange

2018 ◽  
Vol 71 (4) ◽  
pp. 621-628.e4 ◽  
Author(s):  
Anaid Benitez ◽  
Wenjun Liu ◽  
Anna Palovcak ◽  
Guanying Wang ◽  
Jaewon Moon ◽  
...  
Keyword(s):  
Strand Break ◽  
Double Strand Break ◽  
Single Strand ◽  
Strand Exchange ◽  
Double Strand Break Repair ◽  
Dna Double Strand Break ◽  
Single Strand Annealing ◽  
Break Repair ◽  
Strand Annealing

scholarly journals Aberrant Double-Strand Break Repair inrad51 Mutants of Saccharomyces cerevisiae

2000 ◽  
Vol 20 (24) ◽  
pp. 9162-9172 ◽  
Author(s):  
Leslie E. Kang ◽  
Lorraine S. Symington
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Strand Break ◽  
Double Strand Break ◽  
Single Strand ◽  
Double Strand Break Repair ◽  
Inverted Repeats ◽  
Independent Events ◽  
Single Strand Annealing ◽  
Break Repair ◽  
Strand Annealing

ABSTRACT A number of studies of Saccharomyces cerevisiae have revealed RAD51-independent recombination events. These include spontaneous and double-strand break-induced recombination between repeated sequences, and capture of a chromosome arm by break-induced replication. Although recombination between inverted repeats is considered to be a conservative intramolecular event, the lack of requirement for RAD51 suggests that repair can also occur by a nonconservative mechanism. We propose a model forRAD51-independent recombination by one-ended strand invasion coupled to DNA synthesis, followed by single-strand annealing. The Rad1/Rad10 endonuclease is required to trim intermediates formed during single-strand annealing and thus was expected to be required forRAD51-independent events by this model. Double-strand break repair between plasmid-borne inverted repeats was less efficient inrad1 rad51 double mutants than in rad1 andrad51 strains. In addition, repair events were delayed and frequently associated with plasmid loss. Furthermore, the repair products recovered from the rad1 rad51 strain were primarily in the crossover configuration, inconsistent with conservative models for mitotic double-strand break repair.

scholarly journals Intermolecular recombination between DNAs introduced into mouse L cells is mediated by a nonconservative pathway that leads to crossover products.

1990 ◽  
Vol 10 (1) ◽  
pp. 103-112 ◽  
Author(s):  
F L Lin ◽  
K Sperle ◽  
N Sternberg
Keyword(s):  
Mammalian Cells ◽  
Dna Recombination ◽  
Strand Break ◽  
Double Strand Break ◽  
Yeast Cells ◽  
Double Strand Break Repair ◽  
L Cells ◽  
Single Strand Annealing ◽  
Break Repair ◽  
Strand Annealing

We describe experiments designed to measure the efficiency of intermolecular recombination between mutant herpesvirus thymidine kinase (tk) genes introduced into mouse L cells. Recombinants were scored as stable transformants containing a functional tk gene. The two recombination substrates used were ptkB8, a pBR322-based plasmid containing a mutant tk gene, with a BamHI linker in an SphI restriction site that is centrally located within the gene, and mp10tk delta 3' delta 5', an mp10 vector with a tk gene deleted at both the 3' and 5' ends. The only homology shared by the two DNAs is 885 base pairs within the tk gene. To determine whether the double-strand break repair model that has been used to explain recombination in yeast cells (J. W. Szostak, T. L. Orr-Weaver, R. J. Rothstein, and F. W. Stahl, Cell 33:25-35, 1983) can account for recombination during the introduction of these DNAs into mammalian cells, we transformed cells with BamHI-linearized ptkB8 and supercoiled mp10tk delta 3' delta 5' replicative-form DNA. These two DNAs should recombine efficiently according to that model and should generate gene conversion products. In this reaction, the supercoiled DNA acts as the donor of information to repair the cleaved tk gene. Our results indicated that the efficiency of this reaction was very low (less than 10 transformants were obtained per 0.1 microgram of each DNA used in the reaction per 10(6) cells). In contrast, if BamHI-cleaved ptkB8 DNA was cotransformed into cells along with a circular DNA molecule containing a tk gene deleted only at its 3' end or only at its 5' end (mp10tk delta 3' or mp10tk delta 5'), then the efficiency of recombination could be more than 4 orders of magnitude higher than it was with circular mp10tk delta 3' delta 5' DNA. Recombination frequencies were highest when the tk delta 3' or tk delta 5' DNA used was cleaved at the tk deletion junction. Southern analyses of DNA from TK+ transformants generated with BamHI-cleaved ptkB8 and BamHI-cleaved mp10tk delta 3' DNAs indicated that recombination was almost always associated with the reassortment of markers flanking the reconstructed tk DNA. Together, these results are more consistent with the nonconservative single-strand annealing model for recombination that we proposed several years ago (F.-L. Lin, K. Sperle, and N. Sternberg, Mol. Cell. Biol. 4:1020-1034, 1984) than they are with the double-strand break repair model.

Intermolecular recombination between DNAs introduced into mouse L cells is mediated by a nonconservative pathway that leads to crossover products

1990 ◽  
Vol 10 (1) ◽  
pp. 103-112
Author(s):  
F L Lin ◽  
K Sperle ◽  
N Sternberg
Keyword(s):  
Mammalian Cells ◽  
Dna Recombination ◽  
Strand Break ◽  
Double Strand Break ◽  
Yeast Cells ◽  
Double Strand Break Repair ◽  
L Cells ◽  
Single Strand Annealing ◽  
Break Repair ◽  
Strand Annealing

We describe experiments designed to measure the efficiency of intermolecular recombination between mutant herpesvirus thymidine kinase (tk) genes introduced into mouse L cells. Recombinants were scored as stable transformants containing a functional tk gene. The two recombination substrates used were ptkB8, a pBR322-based plasmid containing a mutant tk gene, with a BamHI linker in an SphI restriction site that is centrally located within the gene, and mp10tk delta 3' delta 5', an mp10 vector with a tk gene deleted at both the 3' and 5' ends. The only homology shared by the two DNAs is 885 base pairs within the tk gene. To determine whether the double-strand break repair model that has been used to explain recombination in yeast cells (J. W. Szostak, T. L. Orr-Weaver, R. J. Rothstein, and F. W. Stahl, Cell 33:25-35, 1983) can account for recombination during the introduction of these DNAs into mammalian cells, we transformed cells with BamHI-linearized ptkB8 and supercoiled mp10tk delta 3' delta 5' replicative-form DNA. These two DNAs should recombine efficiently according to that model and should generate gene conversion products. In this reaction, the supercoiled DNA acts as the donor of information to repair the cleaved tk gene. Our results indicated that the efficiency of this reaction was very low (less than 10 transformants were obtained per 0.1 microgram of each DNA used in the reaction per 10(6) cells). In contrast, if BamHI-cleaved ptkB8 DNA was cotransformed into cells along with a circular DNA molecule containing a tk gene deleted only at its 3' end or only at its 5' end (mp10tk delta 3' or mp10tk delta 5'), then the efficiency of recombination could be more than 4 orders of magnitude higher than it was with circular mp10tk delta 3' delta 5' DNA. Recombination frequencies were highest when the tk delta 3' or tk delta 5' DNA used was cleaved at the tk deletion junction. Southern analyses of DNA from TK+ transformants generated with BamHI-cleaved ptkB8 and BamHI-cleaved mp10tk delta 3' DNAs indicated that recombination was almost always associated with the reassortment of markers flanking the reconstructed tk DNA. Together, these results are more consistent with the nonconservative single-strand annealing model for recombination that we proposed several years ago (F.-L. Lin, K. Sperle, and N. Sternberg, Mol. Cell. Biol. 4:1020-1034, 1984) than they are with the double-strand break repair model.

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