Acetylated h3k14

Proteins in the nuclear extracts (60 μg protein) were separated by SDS–PAGE gels (456–1043, Bio-Rad, Hercules, CA), transferred onto PVDF membranes (162–0174, Bio-Rad), and incubated with anti-histone antibodies: acetylated H3K9 (1/2000: ab4441, abcam, Cambridge, MA), acetylated H3K14 (1/5000: 06–911, EMD Millipore, Billerica, MA), acetylated H4K5 (1/2000: 07–327, EMD Millipore). Bound antibody was detected by horseradish peroxidase-conjugated anti-rabbit secondary antibody (1/20000: 1705046, Bio-Rad) and developed using Clarity^™ Western ECL Substrate (1705060, Bio-Rad). Signals were detected and captured by ImageQuant^™ LAS 4000 mini (GE healthcare, Pittsburgh, PA), and band intensities were quantified with ImageJ software. H3K9 acetylation intensity in individual lanes was reported relative to the normalized Mock treatment, and calculated using this formula: Relative H3K9ace intensity for each timepoint = (H3K9ace / PCNA) / averaged (Mock H3K9ace / Mock PCNA). Statistical significance was determined by unpaired t test (two-tailed) against mock at each time point.

Proteins in the nuclear extracts (60 μg protein) were separated by SDS-PAGE gels (456-1043, Bio-Rad, Hercules, CA), transferred onto PVDF membranes (162-0174, Bio-Rad), and incubated with anti-histone antibodies: acetylated H3K9 (1/2000: ab4441, abcam, Cambridge, MA), acetylated H3K14 (1/5000: 06-911, EMD Millipore, Billerica, MA), and acetylated H4K5 (1/2000: 07-327, EMD Millipore). Bound antibody was detected by horseradish peroxidase-conjugated anti-rabbit secondary antibody (1/20,000: 1705046, Bio-Rad) and developed using Clarity Western ECL Substrate (1705060, Bio-Rad). Signals were detected and captured using ImageQuant LAS 4000 mini (GE Healthcare, Pittsburgh, PA), and band intensities were quantified with ImageJ software. H3K9 acetylation intensity in individual lanes was reported relative to the normalized Mock treatment and calculated using this formula: relative H3K9ace intensity for each timepoint = (H3K9ace/PCNA)/averaged (Mock H3K9ace/Mock PCNA). Statistical significance was determined by unpaired t-test (two-tailed) against mock at each time point.