Rad51 sc 8349

Antibodies were purchased from the following source: RAD51 (sc-8349), Myc (sc-40), and GAPDH (sc-25778) antibodies were from Santa Cruz, β-actin (5441) and Flag antibodies (F7425) were from Sigma–Aldrich, and V5 antibody was from Invitrogen. HCV core, NS3, NS5A antibodies were described elsewhere (Lim et al., 2011 (link)).

Cell lysates were generated by lysis in NETN buffer (50 mM Tris-HCl pH = 7.4, 250 mM NaCl, 5 mM EDTA, 0.1% NP-40, 10 mM NaF, 200 μM Na₂VO₃, 0.5 mM PMSF, and 1x protease inhibitor cocktail) on ice for 30 min. Non-soluble debris was precipitated by a 5 min 12000 rpm spin at 4 °C and discarded. Lysate was treated with 4x loading buffer (0.5 M Tris-HCl pH 6.8, 277 mM SDS, 40% glycerol, bromophenol blue, 4% 2-mercaptoethanol), ran on a 10% SDS-PAGE gel, and transferred onto a PVDF membrane for 1–2 hr at 500 mA. The following Santa Cruz (Dallas, TX, USA) antibodies were used: BRCA1 (sc-6954) and RAD51 (sc-8349 and in-house aliquot B32). The following Cell Signaling (Danvers, MA, USA) antibodies were used: ATM (2873), GAPDH (5174), Chk2 (2662), pChk2 T68 (2661), Chk1 (2345), and pChk1 S345 (2341). Other antibodies included DEK (610948 BD Bioscience, San Jose, CA, USA), DEK (16448-1-AP, Protein Tech, Rosemont, IL, USA), DEK (in-house K-877)72 (link), DNA-PKcs (ab1832, Abcam, Cambridge, MA, USA), pDNA-PKcs S2056 (ab18192), pATM S1981 (AF1655, R&D Systems, Minneapolis, MN), γH2AX (05-636, Millipore, Darmstadt, Germany), MRE11 (GTX70212, genetex, San Antonio, TX, USA), and NBS1 (NB100-143SS, Novus Biologicals, Littleton, CO, USA).

The following primary antibodies were used for immunoblotting and immunofluorescence: Cleaved form of caspase-3 (#9662S), phospho-AKT S473 (#4060P), AKT (#4691P), phospho-histone H2A.X Ser139 (#9718S) (Cell Signaling, Danvers, MA, USA), PARP (#436400) (Life Technologies, Carlsbad, CA, USA), γ-H2AX S139 (sc-517348), Rad51 (sc-8349) (Santa Cruz Biotechnology, Santa Cruz, CA, USA), Rad51 (ab-133534, Abcam, Burlingame, CA, USA), phosphor-RPA32 Ser4/8 (A300-245)(Bethyl Laboratories, Montgomery, TX, USA), beta-actin (A00730-200, Gene Script, Piscataway, NJ, USA), HRP-conjugated secondary antibodies for Western blotting were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA) and Alexa Fluor 488- or Texas Red-conjugated secondary antibodies for immunofluorescence staining were obtained from Invitrogen (Carlsbad, CA, USA).