ChIP assay was carried out as described previously [21] (link). Chromatin extracts were immunoprecipitated using 10 μg anti-KDM4C (A300-885 A; Bethyl Laboratories); 4 μg anti-trimethylated H3K9 (61,013; Active Motif), 4 μg anti-dimethylated H3K9 (39,683; Active Motif), and 2 μg anti-total H3 (39,763; Active Motif); Anti-GST (sc-459; Santa Cruz Biotechnology) antibodies were used as mock ChIP controls. Fold enrichments were calculated by determining the ratios of the amount of immunoprecipitated DNA to that of the input sample and were normalized to the level observed at a control region, which was defined as 1.0. The ChIP primers are available on request (Supplemental Table Primers).
Anti trimethylated h3k9
Manufactured by Active Motif
Sourced in United States
Anti-trimethylated H3K9 is a laboratory product that detects the trimethylation of histone H3 at lysine 9. This modification is associated with heterochromatin formation and gene silencing. The product can be used in various applications, such as chromatin immunoprecipitation (ChIP) and western blotting, to study epigenetic mechanisms.
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2 protocols using anti trimethylated h3k9
1
Chromatin Immunoprecipitation Assay Protocol
2
Investigating Histone Methylation at NOTCH1 Promoter
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In order to explore whether GASC1 could affect the level of methylation modification of histone H3K9me2/me3 in NOTCH1 promoter region, we performed ChIP assay in ALDH+ KYSE150 cells with GASC1 knockdown or CA treatment. ChIP assay was carried out as described previously [9 (link)]. Antibodies were used as anti-GASC1 (Bethyl Laboratories, USA), anti-trimethylated H3K9, anti-dimethylated H3K9, and anti-total H3 (Active Motif, USA). Anti-GST antibody (Santa Cruz Biotechnology, USA) was used as mock ChIP control. The ChIP-qPCR primers are as follows: nonpromoter control: forward: 5′-TTCTTGAGTTTGGCATGAAAGA-3′; reverse: 5′-TCTTAATCCAGCATTGGCAGT-3′; NOTCH1 promoter: forward: 5′-CCCAATGGGCAAGAAGTCTA-3′; reverse: 5′-CACAATGTGGTGGTGGGATA-3′.
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