Product Pathways - Chromatin Regulation / Epigenetics
Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb #13969
|13969S||100 µl ( 10 western blots )||￥3,900.00 现货查询||购买询价|
|13969T||20 µl ( 2 western blots )||￥1,500.00 现货查询||购买询价|
|13969||carrier free & custom formulation / quantity||email request|
Species cross-reactivity is determined by western blot.
Applications Key: W=Western Blotting, IP=Immunoprecipitation, F=Flow Cytometry, IF-IC=Immunofluorescence (Immunocytochemistry), ChIP=Chromatin IP, ChIP-seq=Chromatin IP-seq,
Species predicted to react based on 100% sequence homology: Bovine,
Specificity / Sensitivity
Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb detects endogenous levels of histone H3 when tri-methylated on Lys9. This antibody shows some cross-reactivity with histone H3 that is di-methylated on Lys9, but does not cross-react with non-methylated or mono-methylated histone H3 Lys9. This antibody does not detect tri-methyl histone H3 Lys9 when the adjacent Ser10 residue is phosphorylated during mitosis. In addition, this antibody does not cross-react with methylated histone H3 Lys4, Lys27, Lys36, or Lys79.
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of histone H3 in which Lys9 is tri-methylated.
Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells and either 10 μl of Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human AFM Intron 1 Primers #5098, SimpleChIP® Human MYT-1 Exon 1 Primers #4493, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
Confocal immunofluorescent analysis of interphase (left) or mitotic (right) HeLa cells, untreated (upper) or λ phosphatase-treated (lower), using Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb (green) and β-Actin (8H10D10) Mouse mAb #3700 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye). As shown, this antibody does not detect tri-methyl histone H3 Lys9 in mitotic cells when the adjacent Ser10 residue is phosphorylated.
Western blot analysis of extracts from various cell lines using Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb.
Antibody specificity was determined by western blotting. HeLa and NIH/3T3 cell lysates were probed with Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb (Panel A) or Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb pre-adsorbed with 1.5 μM of various competitor peptides (panels B-M). As shown, only the tri-methyl histone H3 (Lys9) peptide (panel E) competed away binding of the antibody.
Flow cytometric analysis of HeLa cells using Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb (blue) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 Hela cells and either 10 µl of HP1β (D2F2) XP® Rabbit mAb #8676 or 10 μl of Tri-Methyl-Histone H3 (Lys9) (D4W1U) Rabbit mAb, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. DNA Libraries were prepared from 5ng enriched ChIP DNA for HP1β ChIP-seq and 50ng enriched ChIP DNA for H3K9me3 ChIP-seq using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina®, and sequenced on the Illumina NextSeq. HP1β and H3K9me3 are known to associate with each other on chromatin. The figure shows binding of both HP1β and H3K9me3 across ZNF genes on chromosome 19, known target genes of both HP1β and H3K9me3. For additional ChIP-seq tracks, please download the product data sheet.
The nucleosome, made up of four core histone proteins (H2A, H2B, H3, and H4), is the primary building block of chromatin. Originally thought to function as a static scaffold for DNA packaging, histones have now been shown to be dynamic proteins, undergoing multiple types of post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (1). Histone methylation is a major determinant for the formation of active and inactive regions of the genome and is crucial for the proper programming of the genome during development (2,3). Arginine methylation of histones H3 (Arg2, 17, 26) and H4 (Arg3) promotes transcriptional activation and is mediated by a family of protein arginine methyltransferases (PRMTs), including the co-activators PRMT1 and CARM1 (PRMT4) (4). In contrast, a more diverse set of histone lysine methyltransferases has been identified, all but one of which contain a conserved catalytic SET domain originally identified in the Drosophila Su(var)3-9, Enhancer of zeste, and Trithorax proteins. Lysine methylation occurs primarily on histones H3 (Lys4, 9, 27, 36, 79) and H4 (Lys20) and has been implicated in both transcriptional activation and silencing (4). Methylation of these lysine residues coordinates the recruitment of chromatin modifying enzymes containing methyl-lysine binding modules such as chromodomains (HP1, PRC1), PHD fingers (BPTF, ING2), tudor domains (53BP1), and WD-40 domains (WDR5) (5-8). The discovery of histone demethylases such as PADI4, LSD1, JMJD1, JMJD2, and JHDM1 has shown that methylation is a reversible epigenetic marker (9).
- Peterson, C.L. and Laniel, M.A. (2004) Curr Biol 14, R546-51.
- Kubicek, S. et al. (2006) Ernst Schering Res Found Workshop , 1-27.
- Lin, W. and Dent, S.Y. (2006) Curr Opin Genet Dev 16, 137-42.
- Lee, D.Y. et al. (2005) Endocr Rev 26, 147-70.
- Daniel, J.A. et al. (2005) Cell Cycle 4, 919-26.
- Shi, X. et al. (2006) Nature 442, 96-9.
- Wysocka, J. et al. (2006) Nature 442, 86-90.
- Wysocka, J. et al. (2005) Cell 121, 859-72.
- Trojer, P. and Reinberg, D. (2006) Cell 125, 213-7.
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
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For Research Use Only. Not For Use In Diagnostic Procedures.
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Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SimpleChIP is a registered trademark of Cell Signaling Technology, Inc.
Tween is a registered trademark of ICI Americas, Inc.
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NEBNext is a registered trademark of New England Biolabs, Inc.
Ultra is a trademark of New England Biolabs, Inc.
Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.
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