Journal: PLoS ONE

Article Title: MutS Homologue hMSH5: Recombinational DSB Repair and Non-Synonymous Polymorphic Variants

doi: 10.1371/journal.pone.0073284

Figure Lengend Snippet: ( A ) DSB-triggered protein loadings at the proximal and distal regions. The regions, surrounding the site of I- Sce I, used for ChIP analysis were schematically illustrated. Numbers represent the distance from the site of I- Sce I in base pairs. The levels of I- Sce I expression, at different time points post-transfection, were analyzed by Western blotting with a α-HA antibody. Representative image of ChIP analysis of locus −303/−57 was shown, in which GAPDH was used as a positive control. PCR analysis (primer set: F13/IN2R1) of an unrelated region on 6p21.3 was included as an additional ChIP control. Arrows were used to mark the positions of the PCR products. ( B ) DSB-induced hMRE11, hRad51, hMSH5, hMSH4, and c-Abl loadings were analyzed at the proximal and distal loci. Error bars represent standard deviations from the means of triplicate measurements.

Article Snippet: Antibodies used in the experiments included α-hMRE11 (NB100–142, Novus Biologicals Inc., Littleton, CO), α-c-Abl, α-hRad51 (14B4) (NB100–148, Novus Biologicals Inc), α-hMSH5 , and α-hMSH4 .

Techniques: Expressing, Transfection, Western Blot, Positive Control, Control

Journal: PLoS ONE

Article Title: MutS Homologue hMSH5: Recombinational DSB Repair and Non-Synonymous Polymorphic Variants

doi: 10.1371/journal.pone.0073284

Figure Lengend Snippet: ( A ) ChIP analysis was performed in conjunction with RNAi-mediated gene silencing to determine the interdependency of DSB-triggered protein loadings at the proximal region. Controls without RNAi treatment were from Fig. 2B – the data is presented again on this graph for the purpose of comparison. ( B ) Knockdown efficiencies of shRNA encoding construct targeting hMRE11, hRad51, hMSH5, or hMSH4. Due to the difficulty in detecting endogenous hMSH4 in 293T cells by Western blotting, the hMSH4 knockdown efficiency was determined by the use of 293T/f45 cells. ( C ) ChIP analysis of the effects of RNAi on DSB-induced protein loadings at a distal region. Levels of protein loading in the absence of RNAi treatment were from Fig. 2B and included for the purpose of comparison. Error bars represent standard deviations from the means of triplicate measurements.

Article Snippet: Antibodies used in the experiments included α-hMRE11 (NB100–142, Novus Biologicals Inc., Littleton, CO), α-c-Abl, α-hRad51 (14B4) (NB100–148, Novus Biologicals Inc), α-hMSH5 , and α-hMSH4 .

Techniques: Comparison, Knockdown, shRNA, Construct, Western Blot

Journal: PLoS ONE

Article Title: MutS Homologue hMSH5: Recombinational DSB Repair and Non-Synonymous Polymorphic Variants

doi: 10.1371/journal.pone.0073284

Figure Lengend Snippet: ( A ) ChIP analysis of the effects of hMSH5 Y742F on DSB-triggered protein loading at both the proximal and distal regions was carried out with 293T reporter cells expressing hMSH5 or hMSH5 Y742F . Briefly, cells were transfected with pcDNA6/Flag-hMSH5 or Flag-hMSH5 Y742F and selected with 10 µg/ml blasticidin. ( B ) Expression of hMSH5 and hMSH5 Y742F in selected clones was validated by Western blot analysis of approximately equal numbers of hMSH5 and hMSH5 Y742F cells. ( C ) The effects of c-Abl kinase inhibition on DSB-induced protein loading at the proximal and distal regions. 293T reporter cells were pretreated with 4 µM imatinib for 48 hrs prior to the induction of DSB by I- Sce I transfection. ChIP analysis was performed to evaluate DSB-induced hRad51, hMSH5, and hMSH4 chromatin association. ( D ) ChIP analysis of GAPDH promoter performed with α-RNAPII or the mouse IgG in the presence or absence of imatinib treatment. Error bars represent standard deviations from the means of triplicate measurements. Asterisks indicate p<0.05 by Student’s t -test.

Article Snippet: Antibodies used in the experiments included α-hMRE11 (NB100–142, Novus Biologicals Inc., Littleton, CO), α-c-Abl, α-hRad51 (14B4) (NB100–148, Novus Biologicals Inc), α-hMSH5 , and α-hMSH4 .

Techniques: Expressing, Transfection, Clone Assay, Western Blot, Inhibition

Journal: Journal of Biological Chemistry

Article Title: Cellular Redistribution of Rad51 in Response to DNA Damage

doi: 10.1074/jbc.m109.024646

Figure Lengend Snippet: FIGURE 1. DNA damage induces an increase in nuclear levels of Rad51 in Brca2-proficient and Brca2-deficient cells. HeLa (A), HCT116 (B), and Capan-1 (C) cells grown at 37 °C were harvested at the indicated times following exposure to 2 Gy of IR and fractionated as described under “Experimental Procedures” to yield cytoplasmic (Cyto), nucleoplasmic (Nuc), and chromatin (Chrom) samples. D and E, HCT116 and Capan-1 cells, respectively, were treated with cyclo- heximide (CHX; 20 M) 1 h prior to exposure to 2 Gy of IR. A portion of each fraction (30 g of total protein) was loaded onto 4–12% SDS-polyacrylamide gels, andWesternblotsweredevelopedusingamouseanti-Rad51monoclonalantibody.Blotswerealsodevelopedusingthefollowingmarkersasloadingcontrols: glyceraldehyde-3-phosphate dehydrogenase (GAPDH; cytoplasmic), Sam68 (nucleoplasmic), and fibrillarin (chromatin). F, changes in levels of nuclear Rad51 as a function of time after IR treatment in A–E were quantified as described under “Experimental Procedures.” The data shown are representative of the results of at least three separate experiments, and the S.D. observed with quantification was 20%.

Article Snippet: Antibodies—The primary antibodies used were mouse monoclonal antibodies against Rad51 (clone 3C10, Upstate), Rad51B (ab3637, Abcam), Rad51C (NB 100-177, Novus), Rad51D (sc-53432, Santa Cruz Biotechnology), Xrcc2 (ab20253, Abcam), and Xrcc3 (NB 100-180D1, Novus).

Techniques:

Journal: Journal of Biological Chemistry

Article Title: Cellular Redistribution of Rad51 in Response to DNA Damage

doi: 10.1074/jbc.m109.024646

Figure Lengend Snippet: FIGURE 2. Levels of Capan-1 nuclear Rad51 increase in an IR dose-de- pendent manner. Capan-1 cells exposed to 5 or 8 Gy of IR were grown at 37 °C for 2 h. Cells were harvested and fractionated as described under “Experimental Procedures,” and portions of the cytoplasmic (Cyto) and nucle- oplasmic (Nuc) fractions (25 g of total protein) were loaded onto 4–12% SDS-polyacrylamide gels. Western blots were developed using a mouse anti- Rad51 monoclonal antibody, and levels of cytoplasmic and nuclear Rad51 were quantified as described under “Experimental Procedures” (supplemen- tal Table 1). The blot shown is representative of four separate experiments. GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: Antibodies—The primary antibodies used were mouse monoclonal antibodies against Rad51 (clone 3C10, Upstate), Rad51B (ab3637, Abcam), Rad51C (NB 100-177, Novus), Rad51D (sc-53432, Santa Cruz Biotechnology), Xrcc2 (ab20253, Abcam), and Xrcc3 (NB 100-180D1, Novus).

Techniques: Western Blot

Journal: Journal of Biological Chemistry

Article Title: Cellular Redistribution of Rad51 in Response to DNA Damage

doi: 10.1074/jbc.m109.024646

Figure Lengend Snippet: FIGURE 4. Rad51C depletion decreases the steady-state level of nuclear Rad51 and diminishes its DNA damage-inducednucleartransportinBrca2-proficientandBrca2-deficientcells.HeLa(A)andCapan-1(B) cells were transfected with a nonspecific () or Rad51C-specific () siRNA pool (SMARTpool), grown for 42 h at 37 °C, exposed to 6 Gy of IR, and harvested 2 h later. Cytoplasmic (Cyto) and nuclear (Nuc) fractions were analyzed by Western blotting using an anti-Rad51 monoclonal antibody. Changes in levels of cytoplasmic and nuclear Rad51 were quantified as described under “Experimental Procedures.” GAPDH, glyceraldehyde-3- phosphate dehydrogenase.

Article Snippet: Antibodies—The primary antibodies used were mouse monoclonal antibodies against Rad51 (clone 3C10, Upstate), Rad51B (ab3637, Abcam), Rad51C (NB 100-177, Novus), Rad51D (sc-53432, Santa Cruz Biotechnology), Xrcc2 (ab20253, Abcam), and Xrcc3 (NB 100-180D1, Novus).

Techniques: Transfection, Western Blot

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Expression of Rad51 in HIV-1 infected primary human microglial cells. (A) Quantitation of the changes in Rad51 levels was performed by densitometric analysis of the scanned X-ray films. The results were normalized relative to expression of Grb2 and are presented as a histogram. (B) p24 levels in the supernatant of the infected microglial cells during the course of HIV-1 infection as measured by ELISA and is shown over time in days post-infection.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Expressing, Infection, Quantitation Assay, Enzyme-linked Immunosorbent Assay

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Effect of Rad51 or siRNA-Rad51 or p65 on HIV-1 LTR activity. (A) Schematic representation of the structural organization of the HIV-1 LTR enhancer region and basal/core promoter and the positions of the deletion mutants that were used in this study. (B) Primary human microglial cells were transfected with 100 ng of the HIV-1 LTR deletion mutants (−167/+66, −94/+66, −80/+66, −40/+66) fused to the luciferase reporter gene either alone or in combination with 300 ng of plasmids expressing Rad51 and/or p65 subunit of NFκB. The numbers represent the fold change in the HIV-1 promoter activity mediated by Rad51 and/or p65/NFκB. (C) The effect of silencing of p65 subunit of NFκB gene expression on HIV-1 LTR activity was assayed when by transfection the HIV-1 LTR plasmid (−167/+66) alone or in combination with 100 nM of p65 siRNA (sip65), non-targeting siRNA (ntsiRNA) or plasmid-expressing Rad51 with or without p65 siRNA. (E) Effect of silencing of Rad51 expression was also assayed by transfection of the HIV-1 LTR with 100 nM of Rad51 siRNA (siRad51), non-targeting siRNA (ntsiRNA) or plasmid-expressing p65/NFκB with or without Rad51 siRNA. The amounts of DNA in each transfection mixture were normalized with pcDNA. Luciferase activity was determined 48 hours after transfection. (D and E) The levels of expression of p65/NFκB and Rad51 were analyzed by Western blots for the lysates from the experiments shown in (C and E), respectively. Grb2 served as a loading control.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Activity Assay, Transfection, Luciferase, Expressing, Gene Expression, Plasmid Preparation, Western Blot, Control

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Colocalization of Rad51 and p65/NFκB. Microglia were transfected with plasmid expressing p65/NFκB together with plasmids expressing either GFP-Rad51 or GFP. Cells transfected with GFP-Rad51 show bright green staining that is cytoplasmic, predominantly perinuclear and punctuate staining in the nucleus (D). Green staining in GFP-transfected control cells was uniformly distributed in nuclei and cytoplasm (A). By immunofluorescence we detected p65/NFκB protein in both nuclei and cytoplasm of infected cells (B and E rhodamine). Superimposition of both fluorochromes shows colocalization of both proteins in the perinuclear region and in the nuclei of cells (G). DAPI was used for labeling nuclear DNA (C and F).

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Transfection, Plasmid Preparation, Expressing, Staining, Control, Immunofluorescence, Infection, Labeling

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Physical interaction between Rad51 and p65/NFκB. (A) Interaction between p65/NFκB and Rad51 was evaluated by immunoprecipitation (IP)/western blot analysis. Total cell lysate prepared from U-87MG cells was immunoprecipitated using anti-Rad51 antibody (lane 2). Nonimmune mouse serum (NMS) was used for the negative control for the IP reaction (lane 1). After IP, protein complexes were washed, separated by SDS-PAGE followed by Western blot analysis using anti-p65 antibody. Fifty micrograms of the total cell lysate protein extract was included on the gel as a positive control (lane 3). (B) Protein extracts from U-87MG cells were incubated with GST-Rad51 or its deletion mutants (1 µM) in a GST pull-down assay for identification of the region of Rad51 that binds to p65/NFκB. GST was used as a negative control for the binding reaction (lane 1). NFκB/p65 was assessed by Western blot. Fifty micrograms of total cell protein extract was used as a positive control for protein expression (lane 7). (C) The integrity of GST and the GST-Rad51 fusion proteins used in the assay was demonstrated directly by SDS-PAGE followed by Coomassie blue staining. (D) Schematic representation of the organization of Rad51, the deletion mutants and their relative binding to p65/NFκB.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Immunoprecipitation, Western Blot, Negative Control, SDS Page, Positive Control, Incubation, Pull Down Assay, Binding Assay, Expressing, Staining

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Effect of Rad51 on the interaction of p65/NFκB with κB DNA sequence element of HIV-1 LTR measured by gel shift in nuclear extracts from U87 MG cells. (A) Approximately 100,000 cpm of synthetic [γ32P]-labeled double-stranded DNA oligonucleotide probe corresponding to the HIV-1 LTR κB site was incubated with 10 µg of nuclear extracts prepared from U87 MG cells transfected with 100 nmol siRNA for Rad51 (lane 3) or 200 nmol siRad51 (lane 4), 200 nmol non-target siRNA (lane 5), pcDNA6A-RAD51 (1–339) (lane 6), with p65/NFκB (lanes 11–14) and control pcDNA (lane 2). Labeled probe was also incubated with nuclear extracts prepared from pcDNA-transfected U87 MG cells in the presence of a specific DNA competitor (cold probe, lane 7), non-specific competitor (an unrelated dsDNA) (lanes 8 and 12), anti-p65 antibody (lanes 9 and 13) and normal mouse serum (NMS) (lanes 10 and 14). (B) Western blots were performed to determine the levels of expression of Rad51 with Lamin A as a loading control (lower part). (C) To identify the region of Rad51 involved in activation of the HIV-1 LTR, primary human microglial cells were transfected with 100 ng of LTR-luciferase plasmid (−167/+66) either alone or in combination with 300 ng of plasmids expressing Rad51: full length (1–339), Rad51 (1–160), Rad51 (1–80), Rad51 (1–40) and Rad51 (80–339). The amount of DNA in each transfection mixture was normalized with pCDNA6A. Luciferase activity was determined 48 hours after transfection. (D) Expression of Rad51 and deletion mutants was verified by western blot analysis of cell lysates prepared from the transfected primary human microglial cells using antibody against MYC-tagged Rad51.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Sequencing, Gel Shift, Labeling, Incubation, Transfection, Control, Western Blot, Expressing, Activation Assay, Luciferase, Plasmid Preparation, Activity Assay

Journal: Molecular and Cellular Biology

Article Title: Minichromosome Maintenance Proteins Interact with Checkpoint and Recombination Proteins To Promote S-Phase Genome Stability

doi: 10.1128/mcb.01717-07

Figure Lengend Snippet: FIG. 6. siRNA against MCM4 or MCM7 in HeLa cells generates DNA damage and cell inviability. HeLa cells were transfected with buffer only (mock) or with siRNA against MCM4 or MCM7, and the cells were analyzed at 0 and 48 h after transfection. (A) Proteins were analyzed by 10% SDS-PAGE, followed by immunoblotting (IB) with antibodies to detect MCM4 or MCM7, and subsequently immunoblotted with anti-PCNA antibody as a loading control. (B) DNA content was assessed by flow cytometry. HeLa cells mock treated or treated with MCM4 siRNA or MCM7 siRNA for 48 h were stained with (C) DAPI to visualize the DNA, and (D) phospho-H2AX and DAPI to detect DNA damage. (E) The DNA damage correlates with loss of MCM localization and corresponds to sites of new DNA synthesis. Stretched chromatin fibers prepared from HeLa cells 48 h after siRNA treatment were immunostained for MCM7 and phospho-H2AX and counterstained with DAPI (left); chromatin fibers prepared from cells pulse-labeled with BrdU for 1 h were immunostained for BrdU and Rad51 and counterstained with DAPI. Scale bar, 10 m (approximately 20 kb). (F) MCM siRNA results in cell death by apoptosis. Cells mock treated or treated with siRNA against MCM4 or MCM7 were immunostained for cleaved caspase 3 and counterstained with DAPI. Scale bars, 10 m.

Article Snippet: HeLa cells and chromatin fibers were immunostained overnight at 4°C in a humid chamber with antibodies against phospho-H2AX (Millipore), MCM7 (Santa Cruz), and Rad51 (Santa Cruz), each used at 1:100 dilution; BrdU (Becton Dickinson) used at 1:50 dilution; or cleaved caspase 3 (Cell Signaling) used at 1:500 dilution.

Techniques: Transfection, SDS Page, Western Blot, Control, Cytometry, Staining, DNA Synthesis, Labeling

Journal: Molecular and Cellular Biology

Article Title: Minichromosome Maintenance Proteins Interact with Checkpoint and Recombination Proteins To Promote S-Phase Genome Stability

doi: 10.1128/mcb.01717-07

Figure Lengend Snippet: FIG. 7. MCM proteins interact with Rad51 in HeLa cells. (A) Whole-cell lysates prepared from asynchronous cultures of HeLa cells (asynch), cells blocked in 2 mM thymidine for 18 h and then released into S phase for 3 h (S phase), and cells blocked in early S phase with 2.5 mM HU for 18 h (HU) were immunoprecipitated with anti-Rad51 antibodies or with an irrelevant, control antibody (anti-myc). The anti-myc immuno- precipitation (IP) was performed on cells blocked in HU. Rad51 and associated proteins were analyzed by 10% SDS-PAGE, followed by immunoblotting to detect Rad51 and Mcm7. (B) Coimmunoprecipitation of MCM7 with Rad51 from HeLa cells is not dependent on DNA. Anti-Rad51 antibodies immunoprecipitate Rad51 and MCM7 from both untreated HeLa cell lysates and lysates that were treated with ethidium bromide (EtBr). AS, asynchronous cells; S, cells released from thymidine block into S phase; HU, cells arrested in hydroxyurea. C, control IP (anti-myc).

Article Snippet: HeLa cells and chromatin fibers were immunostained overnight at 4°C in a humid chamber with antibodies against phospho-H2AX (Millipore), MCM7 (Santa Cruz), and Rad51 (Santa Cruz), each used at 1:100 dilution; BrdU (Becton Dickinson) used at 1:50 dilution; or cleaved caspase 3 (Cell Signaling) used at 1:500 dilution.

Techniques: Immunoprecipitation, Control, SDS Page, Western Blot, Blocking Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: DNA damage during the G0/G1 phase triggers RNA-templated, Cockayne syndrome B-dependent homologous recombination

doi: 10.1073/pnas.1507105112

Figure Lengend Snippet: CSB recruited at transcriptionally active damage sites is independent of HR factors. (A) CSB is recruited to I-SCEI induced DSBs. TA-cherry and HA-CSB with/without I-SCEI were cotransfected into U2OS TRE cells. After 24 h, cells were fixed and stained with HA-antibody. The recruitment of CSB at TA-cherry with/without I-SCEI sites is shown. (B) siRAD51, siRAD51C, or siRAD52 transfected U2OS TRE cells were expressed with FLAG-CSB and TA-KR. The recruitment of CSB at sites of TA-KR is shown after light irradiation for 10 min followed by 30 min of incubation. (C) The fold increase of CSB intensity after knockdown of the indicated protein. (D) Knockdown effects of RAD51, RAD52, and RAD51C by siRNAs. Western blot of siCtrl, siRAD52, or siRAD51C transfected U2OS TRE cells with the indicated antibody. (E) The recruitment of CSB at sites of TA-KR in U2OS TRE cells with light irradiation for 10 min followed with 30 min incubation. Cells were pretreated for 15 min with RNaseH (15 units). (F) RAD51C interacts with endogenous CSB independently of DNA after DNA damage. Flp-in 293 cells stably expressing GFP-RAD51C were treated with 5 Gy IR with 1 h postirradiation incubation. Then cell lysates with/without 10 μg/mL EtBr were applied for IP by anti-GFP. Detection of endogenous CSB is shown. (G) Myc-CSB FL or UbΔ and TA-KR cotransfected U2OS TRE cells were irradiated with light for 10 min followed by 0.5–24 h incubation. Cells were immunostained by anti-Myc. Data are represented as mean ± SD, and the P values were determined by using Student’s two-tailed t test.

Article Snippet: Olympus FV2000 confocal microscopy was used for imaging studies. table ft1 table-wrap mode="anchored" t5 Table S1. caption a7 siRNA Catalog no. Company siCSB SR301453 Origene siRAD51 E-003530–00 Thermo Scientific Dharmacon siRAD52 E-011760–00 Thermo Scientific Dharmacon siRAD51C E-010534–00 Thermo Scientific Dharmacon siXPA 4390771-n264189 Ambion siXPC 4390771-n270792 Ambion siXPG 4392420-s4802 Ambion siCSA 4392420-s3103 Ambion Open in a separate window siRNAs used in the study table ft1 table-wrap mode="anchored" t5 Table S2. caption a7 Antibody Species Clone, catalog no. Company ɣH2AX, ser139 Mouse monoclonal JBW301, 05–636 EMD Millipore GFP Mouse monoclonal 11814460001 Roche HA Mouse monoclonal 12CA5, 11666606001 Roche c-Myc Rabbit polyclonal A-14, sc-789 Santa Cruz Biotechnology FLAG Mouse monoclonal M2, IB13026 Eastman Kodak CSB Rabbit polyclonal H-300, sc-25370 Santa Cruz Biotechnology RAD51 Rabbit polyclonal H-300, sc-9034 Santa Cruz Biotechnology RAD52 Rabbit polyclonal F-7, sc-365341 Santa Cruz Biotechnology RAD51C Rabbit polyclonal 2H11, sc-56214 Santa Cruz Biotechnology AcK9H3 Rabbit polyclonal ab4441 abcam actin Mouse monoclonal 9E10, MMS-150P Covance 4H8 Mouse monoclonal ab5408 Abcam CSB Rabbit polyclonal sc-25370 Santa Cruz Biotechnology 53BP1 Rabbit polyclonal NB100-304 Novus Biologicals Open in a separate window Antibodies used in the study

Techniques: Staining, Transfection, Irradiation, Incubation, Knockdown, Western Blot, Stable Transfection, Expressing, Two Tailed Test

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: DNA damage during the G0/G1 phase triggers RNA-templated, Cockayne syndrome B-dependent homologous recombination

doi: 10.1073/pnas.1507105112

Figure Lengend Snippet: siRNAs used in the study

Article Snippet: Olympus FV2000 confocal microscopy was used for imaging studies. table ft1 table-wrap mode="anchored" t5 Table S1. caption a7 siRNA Catalog no. Company siCSB SR301453 Origene siRAD51 E-003530–00 Thermo Scientific Dharmacon siRAD52 E-011760–00 Thermo Scientific Dharmacon siRAD51C E-010534–00 Thermo Scientific Dharmacon siXPA 4390771-n264189 Ambion siXPC 4390771-n270792 Ambion siXPG 4392420-s4802 Ambion siCSA 4392420-s3103 Ambion Open in a separate window siRNAs used in the study table ft1 table-wrap mode="anchored" t5 Table S2. caption a7 Antibody Species Clone, catalog no. Company ɣH2AX, ser139 Mouse monoclonal JBW301, 05–636 EMD Millipore GFP Mouse monoclonal 11814460001 Roche HA Mouse monoclonal 12CA5, 11666606001 Roche c-Myc Rabbit polyclonal A-14, sc-789 Santa Cruz Biotechnology FLAG Mouse monoclonal M2, IB13026 Eastman Kodak CSB Rabbit polyclonal H-300, sc-25370 Santa Cruz Biotechnology RAD51 Rabbit polyclonal H-300, sc-9034 Santa Cruz Biotechnology RAD52 Rabbit polyclonal F-7, sc-365341 Santa Cruz Biotechnology RAD51C Rabbit polyclonal 2H11, sc-56214 Santa Cruz Biotechnology AcK9H3 Rabbit polyclonal ab4441 abcam actin Mouse monoclonal 9E10, MMS-150P Covance 4H8 Mouse monoclonal ab5408 Abcam CSB Rabbit polyclonal sc-25370 Santa Cruz Biotechnology 53BP1 Rabbit polyclonal NB100-304 Novus Biologicals Open in a separate window Antibodies used in the study

Techniques: