04-790-S | Anti-dimethyl-Histone H3 (Lys4) Antibody, clone AW30, Trial Size

04-790-S
10 µL  
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      Overview

      Replacement Information

      Key Spec Table

      Species ReactivityKey ApplicationsHostFormatAntibody Type
      H, ChWB, ChIP-seq, ChIP, DB, PIARbCulture SupernatantMonoclonal Antibody
      Description
      Catalogue Number04-790-S
      Replaces05-790
      Brand Family Upstate
      Trade Name
      • Upstate
      DescriptionAnti-dimethyl-Histone H3 (Lys4) Antibody, clone AW30, Trial Size
      Alternate Names
      • H3K4me2
      • Histone H3 (di methyl K4)
      Background InformationHistones are highly conserved proteins that serve as the structural scaffold for the organization of nuclear DNA into chromatin. The four core histones, H2A, H2B, H3, and H4, assemble into an octamer (2 molecules of each). Subsequently, 146 base pairs of DNA are wrapped around the octamer, forming a nucleosome, the basic subunit of chromatin. Histones are modified post-translationally by the actions of enzymes in both the nucleus and cytoplasm. These modifications regulate DNA transcription, repair, recombination, and replication. The most commonly studied modifications are acetylation, phosphorylation, methylation, and ubiquitination. These modifications occur predominantly on the N-terminal and C-terminal tails that extend beyond the nucleosome core particle. Di- and trimethylation of histone H3 at Lys4 correlates with transcriptional activity of many genes. Dimethylation of Histone H3 at Lys4 occurs at both active and inactive euchromatic regions but not in silent heterochromatic sites, whereas trimethylation at Lys4 is present exclusively at active genes.
      References
      Product Information
      FormatCulture Supernatant
      Control
      • HeLa acid extracts
      PresentationCultured supernantant in 0.05% sodium azide
      Applications
      ApplicationAnti-dimethyl-Histone H3 (Lys4) Antibody, clone AW30 is a rabbit monoclonal antibody for detection of dimethyl-Histone H3 (Lys4) also known as H3K4me2, Histone H3 (di methyl K4) & has been validated in WB, ChIP, ChIP-seq, DB, PIA, Mplex.
      Key Applications
      • Western Blotting
      • ChIP-seq
      • Chromatin Immunoprecipitation (ChIP)
      • Dot Blot
      • Peptide Inhibition Assay
      Application NotesWestern Blot Analysis:
      A 1:1000-1:2000 dilution of this lot detected dimethyl histone H3 (Lys4) in HeLa acid extracts.

      Western Blot Analysis:
      A 1:2,000 to 1:10,000 dilution of a previous lot detected methylated histone H3 in acid extracted proteins from HeLa cells. The antibody did not detect unmethylated recombinant Histone H3.

      ChIP-seq Analysis:
      Representative lot data. Chromatin immunoprecipitation was performed using the Magna ChIP™ HiSens kit (cat# 17-10460), Anti-dimethyl-Histone H3 (Lys4) antibody (2 µl cat# 04-790-S), 20 µL Protein A/G beads, and 1e6 crosslinked HeLa cell chromatin followed by DNA purification using magnetic beads. Libraries were prepared from Input and ChIP DNA samples using standard protocols with Illumina barcoded adapters, and analyzed on Illumina HiSeq instrument. An excess of sixteen million reads from FastQ files were mapped using Bowtie (http://bowtie-bio.sourceforge.net/manual.shtml) following TagDust (http://genome.gsc.riken.jp/osc/english/dataresource/) tag removal. Peaks were identified using MACS (http://luelab.dfci.harvard.edu/MACS/), with peaks and reads visualized as a custom track in UCSC Genome Browser (http://genome.ucsc.edu) from BigWig and BED files. The highest 25% of peaks identified in the 04-790 and 05-1338 datasets showed 92 and 90% overlap with peaks identified in the ENCODE H3K4me2 BROAD Histone track for HeLa S3.

      Dot Blot Analysis:
      Absurance Histone H3 Antibody Specificity Array (Cat. No. 16-667) and Absurance Histone H2A, H2B, H4 Antibody Specificity Array (Cat. No. 16-665), which contain histone peptides with various modifications were probed with Cat. No 04-790, Anti-dimethyl Histone H3 (Lys4), clone AW30 at 1:1000 dilution. Proteins were visualized using a Donkey anti-Rabbit IgG conjugated to HRP and a chemiluminescence detection system.

      Peptide Inhibition Analysis:
      2 μM of a histone H3 peptide containing dimethyl-lysine 4 abolished detection of histone H3 by a previous lot of antibody in immunoblot analysis of HeLa acid extracts. No signal reduction was observed with histone H3 peptides containing either monomethyl or trimethyl-lysine 4 modifications
      Biological Information
      ImmunogenKLH-conjugated, synthetic peptide containing the sequence …Tme2KQT… in which me2K corresponds to dimethyl-lysine at residue 4 of human histone H3.
      Cloneclone AW30
      HostRabbit
      SpecificityHistone H3 dimethylated on lysine 4
      IsotypeIgG
      Species Reactivity
      • Human
      • Chicken
      Species Reactivity NoteHuman and chicken. The immunizing sequence is conserved from Tetrahymena
      to human, so broad species cross-reactivity is expected.
      Antibody TypeMonoclonal Antibody
      Entrez Gene Number
      Entrez Gene SummaryHistones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures. This gene is intronless and encodes a member of the histone H3 family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element. This gene is located separately from the other H3 genes that are in the histone gene cluster on chromosome 6p22-p21.3.
      Gene Symbol
      • HIST3H3
      • H3FT
      • MGC126886
      • H3t
      • MGC126888
      • H3T
      • H3/g
      • H3.4
      • H3/t
      UniProt Number
      UniProt SummaryFUNCTION: SwissProt: Q16695 # Core component of nucleosome. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling.
      SIZE: 136 amino acids; 15508 Da
      SUBUNIT: The nucleosome is a histone octamer containing two molecules each of H2A, H2B, H3 and H4 assembled in one H3-H4 heterotetramer and two H2A-H2B heterodimers. The octamer wraps approximately 147 bp of DNA.
      SUBCELLULAR LOCATION: Nucleus.
      TISSUE SPECIFICITY: Expressed in testicular cells.
      DEVELOPMENTAL STAGE: Expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
      PTM: Acetylation is generally linked to gene activation. Acetylation on Lys-10 impairs methylation at Arg-9. Acetylation on Lys-19 and Lys-24 favors methylation at Arg-18 (By similarity). & Citrullination at Arg-9 and/or Arg-18 by PADI4 impairs methylation and represses transcription (By similarity). & Asymmetric dimethylation at Arg-18 by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 by PRMT5 is linked to gene repression (By similarity). & Methylation at Lys-5, Lys-37 and Lys-80 are linked to gene activation. Methylation at Lys-5 facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 and Lys-28 are linked to gene repression. Methylation at Lys-10 is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 and acetylation of H3 and H4. Methylation at Lys-5 and Lys-80 require preliminary monoubiquitination of H2B at 'Lys-120'. Methylation at Lys-10 and Lys-28 are enriched in inactive X chromosome chromatin (By similarity). & Phosphorylated at Thr-4 by GSG2/haspin during prophase and dephosphorylated during anaphase. At centromeres, specifically phosphorylated at Thr-12 from prophase to early anaphase. Phosphorylated at Ser-11 during the whole mitosis. Phosphorylation at Ser-11, which is linked to gene activation, prevents methylation at Lys-10 but facilitates acetylation of H3 and H4. Phosphorylated at Ser-29 by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation (By similarity). & Phosphorylation at 'Ser-11' is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at 'Ser-11' is important during interphase because it enables the transcription of genes following external stimulation, like stress or growth factors. Phosphorylation at 'Ser-11' is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylation at 'Ser-11' by AURKB/Aurora-B mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. & Ubiquitinated (By similarity).
      SIMILARITY: SwissProt: Q16695 ## Belongs to the histone H3 family.
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Quality Assuranceroutinely evaluated by immunoblot on acid-extracted proteins from HeLa cells; the antibody did not detect unmethylated recombinant Histone H3 (Catalog #14-494)
      Usage Statement
      • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
      Storage and Shipping Information
      Storage ConditionsStable for 1 year at -20°C from date of receipt.
      For maximum recovery of product, centrifuge the vial prior to removing the cap.
      Packaging Information
      Material Size10 µL
      Transport Information
      Supplemental Information
      Specifications