H3K27me3S28p | Histone H3 (trimethylated Lys27, p Ser28)
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Chromatin Immunoprecipitation: using anti-H3K27me3S28p antibodies. Chromatin from one million formaldehyde cross-linked HeLa cells was used with 2 μg of H3K27me3S28p and 20 μl of magnetic IgG beads per immunoprecipitation. A no antibody (No Ab) control was also used. Immunoprecipitated DNA was quantified using quantitative PCR and normalized to the input chromatin.
Dot Blot: using anti-H3K27me3S28p antibodies. Lane 1: S28p/K27 unmodified. Lane 2: S28p N-Term. Lane 3: S28p C-term. Lane 4: K27Me3. Lane 5: S28p/K27Me3. Load: 1, 10, and 100 picomoles of peptide. Primary antibody at 1 μg/ml for 45 min at 4°C. Secondary antibody: Dylight®488 rabbit secondary antibody at 1:10 000 for 45 min at RT.
Western Blot: using anti-H3K27me3S28p antibodies. Lane 1: HeLa histone extracts. Lane 2: NIH-3T3 histone extracts. Lane 3: C. elegans embryo cell lysate. Load: 30 μg per lane. Primary antibody used at 1 μg/ml overnight at 4°C. Secondary antibody: IRDye800™ rabbit secondary antibody at 1:10 000 for 45 min at RT.