Mouse monoclonal anti gfp antibody

Human cell lines U2OS, HEK293T and HCT116 were purchased from ATCC. MDCI-, H2AX-, RNF8-, 53BP1- deficient MEFs and wild-type MEF were gereruously supplied by Dr. Xiaochun Yu described previously[19 (link)]. Human cell lines and MEF cells were maintained in DMEM medium with 10% fetal calf serum and cultivated at 37°C in 5% CO2 (v/v). Cells were lysed with NETN buffer (contains 0.5% NP40,50 mM Tris-HCl pH 8.0, 100 mM NaCl, 2 mM EDTA) with Roche Protease Inhibitor Cocktail. Immunoprecipitation and western blotting were performed following standard protocol as described previously. Rabbit anti-RNF138 antibody is purchased from Santa Cruse, anti-glutathione s-transferase (GST) antibody, monoclonal anti-β-actin antibodies and GAPDH antibody were purchased from Sigma. Mouse monoclonal anti-GFP antibody and Phospho-(Ser/Thr) ATM/ATR Substrate antibody were purchased from Cell Signaling Technology. RPA antibody was purchased from Abcam. γH2AX, RAD51, RNF8, MDC1, BRCA1, Rabbit anti-53BP1 antibody was kind gifts from Dr. Xiaochun Yu.

U2OS or 293 T cells were maintained in DMEM medium with 10% fetal bovine serum and cultivated at 37 °C in 5% CO₂ (v/v). U2OS cells were transfected with PX459 vector containing PARP1-sgRNA for PARP1 knockout. Transfected cells were plated at low density in 1.5 mg/ml puromycin (Invitrogen). Single colonies were propagated, and individual clones were assessed by western blotting. Loss of PARP1 in U2OS cells was validated by anti-PARP1 antibody which was purchased from Cell Signaling Technology (Cat# 9542).
Cells were lysed with NETN buffer (0.5% NP40, 50 mM Tris-HCl pH 8.0, 100 mM NaCl, 2 mM EDTA) with Roche Protease Inhibitor Cocktail. Immunoprecipitation and Western blotting were performed following standard protocol as described previously20 (link). Rabbit anti-CTCF antibody was purchased from Cell Signaling. Monoclonal anti-PAR antibody was purchased from Trevigen. Mouse monoclonal anti-GFP antibody was purchased from Cell Signaling.