Anti ku80 c48e7

Total protein lysate was extracted from BMMCs using RIPA buffer (Intron Biotechnology, Korea) containing a cocktail of 1% phosphatase- and 1% protease inhibitors (Thermo Fischer Scientific, USA). Sodium dodecyl sulphate-polyacrylamide gel electrophoresis and electroblotting were performed using anti-p53 (9282, Cell Signaling Technology, US and DO-7 sc-47,698 Santa Cruz Biotechnology, US), anti-MDM2 (MABE281, Merck Millipore, US), and anti-p21^WAF-1 (OP64, Merck Millipore, US) antibodies. Equal protein loading was assessed using anti-alpha tubulin (T5168, Sigma Aldrich, US) and anti-Ku80 (C48E7, Cell Signaling Technology, US) antibodies. All antibodies were diluted in 1% (w/v) non-fat milk powder in TBS–Tween. Proteins were visualized using enhanced chemiluminescence (ECL RevelBlot Plus or Intense, Ozyme, France) depending on the intensity of the signal. Protein lysate isolated from Saos-2 p53-null cell line engineered to over-express specific p53 isoforms cDNA were used as reference for p53 isoforms identification.

Sub‐confluent cells were trypsinized and washed once in PBS. Total proteins were extracted by solubilizing the cell pellets in urea buffer (8 M urea, 0.1 M NaH₂PO₄, 10 mM Tris pH 8, 0.1% mercaptoethanol). Protein concentration was quantified by the Bio‐Rad protein assay (Bio‐Rad Laboratories, Hercules, CA). Proteins (20 μg) were separated by electrophoresis on NuPage 4%‐20% Bis‐Tris gels (Life Technologies Corporation, Carlsbad, CA) and transferred onto Immobilon‐P membranes (Millipore Sigma, Burlington, MA) according to the manufacturers' directions. Western blot analyses of proteins were carried out by using anti‐Ku80 (C48E7) (Cell Signaling Technology Inc., Danvers, MA), anti‐BAF180 (ABE70) (Millipore Sigma), anti‐BRG1/SMARCA4 (SC‐17796) (Santa Cruz Biotechnology, Santa Cruz, CA), anti‐BAF57/SMARCE1 (A300‐810A) (Bethyl Laboratories, Montgomery, TX), anti‐SNF5/ SMARCB1 (612111) (BD Biosciences, San Jose, CA), anti‐EZH2 (ab3748) (Abcam, Cambridge, MA), anti‐SUZ12 (ab126577) (Abcam), and anti‐EED (ab4469) (Abcam). The primary antibodies were detected using fluorescently labeled anti‐mouse and anti‐rabbit secondary antibodies at a dilution of 1:10 000 and obtained from Li‐Cor and were visualized by scanning the blots on a Li‐Cor Odyssey (Thermo Fisher Scientific).