Revision 3

#3306Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IF-IC, ChIP

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

160,180

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q96KQ7

Entrez-Gene Id:

10919

Product Information

Product Usage Information

For optimal ChIP results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

Application Dilution
Western Blotting 1:1000
Immunofluorescence (Immunocytochemistry) 1:50
Chromatin IP 1:100

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

G9a/EHMT2 (C6H3) Rabbit mAb detects endogenous levels of total G9a/EHMT2 protein (both the 165 kDa G9a-L and 140 kDa G9a-S isoforms). This antibody does not cross-react with other histone methyltransferases, including GLP.

Species Reactivity:

Human, Mouse, Rat, Monkey

Species predicted to react based on 100% sequence homology

Bovine, Pig, Horse

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to the carboxy terminus of the human G9a/EHMT2 protein.

Background

G9a, also known as Euchromatic histone-lysine N-methyltransferase 2 (EHMT2), is a member of a family of histone lysine methyltransferases, each of which contains a conserved catalytic SET domain originally identified in Drosophila Su[var]3-9, Enhancer of zeste, and Trithorax proteins (1). Recombinant G9a can mono-, di- and tri-methylate histone H3 on Lys9 and Lys27 in vitro (1,2). However, in vivo G9a forms a complex with GLP, a G9a-related histone methyltransferase, and together these proteins function as the major euchromatic histone H3 Lys9 mono- and di-methyltransferases, creating transcriptionally repressive marks that facilitate gene silencing (3,4). G9a methylates itself on Lys165, a modification that regulates the association of HP1 repressor proteins with the G9a/GLP complex (5). The G9a/GLP complex also contains Wiz, a zinc finger protein that is required for G9a/GLP hetero-dimerization and complex stability (6). Wiz contains two CtBP co-repressor binding sites, which mediate the association of the G9a/GLP with the CtBP co-repressor complex (6). In addition, G9a and GLP are components of other large transcriptional co-repressor complexes, such as those involving E2F6 and CDP/cut (7-9). G9a interacts with DNMT1, and both proteins are required for methylation of DNA and histone H3 (Lys9) at replication foci, providing a functional link between histone H3 Lys9 and CpG methylation during DNA replication (10). G9a activity is critical for meiotic prophase progression, as mutant mice deficient in germ line G9a show a large loss of mature gametes (11). In addition, G9a facilitates increased global levels of di-methyl histone H3 (Lys9) during hypoxic stress and increased G9a expression is associated with hepatocelluar carcinoma (12,13).

  1. Tachibana, M. et al. (2001) J Biol Chem 276, 25309-17.
  2. Patnaik, D. et al. (2004) J Biol Chem 279, 53248-58.
  3. Tachibana, M. et al. (2002) Genes Dev 16, 1779-91.
  4. Tachibana, M. et al. (2005) Genes Dev 19, 815-26.
  5. Sampath, S.C. et al. (2007) Mol Cell 27, 596-608.
  6. Ueda, J. et al. (2006) J Biol Chem 281, 20120-8.
  7. Ogawa, H. et al. (2002) Science 296, 1132-6.
  8. Shi, Y. et al. (2003) Nature 422, 735-8.
  9. Nishio, H. and Walsh, M.J. (2004) Proc Natl Acad Sci USA 101, 11257-62.
  10. Estève, P.O. et al. (2006) Genes Dev 20, 3089-103.
  11. Tachibana, M. et al. (2007) EMBO J 26, 3346-59.
  12. Kondo, Y. et al. (2007) Hepatol Res 37, 974-83.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IF-IC: Immunofluorescence (Immunocytochemistry) ChIP: Chromatin IP

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

使用に関する制限

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Revision 3
#3306

G9a/EHMT2 (C6H3) Rabbit mAb

Western Blotting Image 1: G9a/EHMT2 (C6H3) Rabbit mAb Expand Image
Western blot analysis of extracts from HeLa and 293 cells using G9a/EHMT2 (C6H3) Rabbit mAb.
Immunofluorescence Image 1: G9a/EHMT2 (C6H3) Rabbit mAb Expand Image
Confocal immunofluorescent analysis of HeLa cells using G9a/EHMT2 (C6H3) Rabbit mAb (green). Actin filaments have been labeled with DY-555 phalloidin (red).
Chromatin Immunoprecipitation Image 1: G9a/EHMT2 (C6H3) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from K562 cells and either G9a/EHMT2 (C6H3) or 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 ZNF19 Intron 3 Primers #98428, human VRK3 intron 11 primers, and SimpleChIP® Human GAPDH Exon 1 Primers #5516. 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.