Amersham ecl prime select

Proteins were extracted using RIPA buffer with 1% Protease inhibitor cocktail (Nacalai Tesque). After determination of the protein concentration using Protein Quantification Assay (Takara Bio Inc.), all samples were denatured in Laemmli sample buffer for 5 min at 100 °C. The denatured samples were separated by SDS-polyacrylamide gel electrophoresis and transferred onto a polyvinylidene difluoride membrane (EMD Millipore). After blocking with 5% milk or 5% BSA, the membranes were incubated with the primary antibodies as follows: p16^INK4a (1:1000, IBL, 11104), p21^Cip1/Waf1 (1:1000, Cell Signaling Technology, 2947), Lamin B1 (1:5000, Abcam, ab16048), RB (1:500, Santa Cruz, sc-102), phospho-RB (Ser780) (1:1000, Cell Signaling Technology, 9307), p53 (1:1000, Santa Cruz, sc-6243), phospho-p53 (Ser15) (1:1000, Cell Signaling Technology, 9284) and α-Tubulin (1:2000, Sigma-Aldrich, T9026). The membranes were then incubated with the secondary antibodies as follows: anti-mouse IgG (1:2000, Cell Signaling Technology, 7076), anti-rabbit IgG (1:2000, Cell Signaling Technology, 7074) and visualized with Amersham ECL prime/select (GE Healthcare), followed by detection with chemiluminescence using LAS-3000mini imaging system (Fujifilm) and by analysis of data using Multi Guage V3.1 (Fujifilm). Uncropped and unprocessed scans of the blots are included in the Source data file.

Proteins were extracted using with RIPA buffer with 1% Protease inhibitor cocktail (Nacalai Tesque). After determination of the protein concentration using Protein Quantification Assay (Takara Bio Inc.), all samples were denatured in Laemmli sample buffer for 5 min at 100 °C. The denatured samples were separated by SDS-polyacrylamide gel electrophoresis and transferred onto a polyvinylidene difluoride membrane (EMD Millipore). After blocking with 5% milk or 5% BSA, the membranes were incubated with the primary antibodies as follows: β-actin (1:1000, Sigma, cat#: A5316), BRD2 (1:1000, Abcam, cat#: ab139690), BRD3 (1:1000, Santa cruz, cat#: sc-81202), BRD4 (1:1000, Cell signaling, cat#: 13440), BRD4 (1:1000, abcam, cat#: 128874). Cleaved caspase3 (1:1000, Cell signaling, cat#: 9664), LaminB1 (1:1000, Abcam, cat#: ab16048), p21 (1:1000, Cell signaling, cat#: 2947 or Abcam, cat#: ab107099), XRCC4 (1:1000, Santa cruz, cat#: sc-2711087). The membranes were then incubated with the secondary antibodies (1:1000, Cell signaling) and visualized with Amersham ECL prime/select (GE Healthcare), followed by detection with chemiluminescence using LAS-3000mini imaging system (Fujifilm) and by anlaysis of data using Multi Guage V3.1 (Fujifilm). Uncropped and unprocessed scans of the blots are included in the Source Data file.