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Key Specifications Table
| Species Reactivity | Key Applications |
|---|---|
| Vrt | WB, ChIP |
| Description | |
|---|---|
| Catalogue Number | 17-676 |
| Trade Name |
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| Description | ChIPAb+ Monomethyl-Histone H3 (Lys4) - ChIP Validated Antibody and Primer Set |
| Overview | All ChIPAb+ antibodies are individually validated for chromatin precipitation, every lot, every time. Each ChIPAb+ antibody set includes control primers (tested every lot by qPCR) to biologically validate your IP results in a locus-specific context. The qPCR protocol and primer sequences are provided, allowing researchers to validate ChIP protocols when using our antibody in their chromatin context. Each set also includes a negative control antibody to ensure specificity of the ChIP reaction. The ChIPAb+ Monomethyl-Histone H3 (Lys4) set includes the Monomethyl-Histone H3 (Lys4) antibody, normal mouse IgG, and qPCR primers which amplify a 213 bp region of human GAPDH coding region. The Monomethyl-Histone H3 (Lys4) and negative controls are supplied in a scalable "per ChIP" reaction size and can be used to functionally validate the precipitation of Monomethyl-Histone H3 (Lys4) associated chromatin. |
| Alternate Names |
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| Background Information | Histone H3 is one of the five main histone proteins involved in the structure of chromatin in eukaryotic cells. Featuring a main globular domain and a long N-terminal tail, H3 is involved with the structure of the nucleosomes of the beads on a string structure. The N-terminal tail of histone H3 protrudes from the globular nucleosome core and can undergo several different types of epigenetic modifications that influence cellular processes. These modifications include the covalent attachment of methyl or acetyl groups to lysine and arginine amino acids and the phosphorylation of serine or threonine. |
| Product Information | |
|---|---|
| Format | Purified |
| Control |
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| Presentation | Anti-Monomethyl-HisPtone H3 (Lys4) (mouse monoclonal), One vial containing 50 µg of protein G purified antibody in 50 µL PBS containing 0.05% sodium azide. Store at -20°C. Normal Mouse IgG, . Two vials containing 25 µg purified Mouse IgG in 25 µL storage buffer containing 0.1% sodium azide. Store at -20°C. ChIP Primers, human GAPDH coding region, . One vial containing 75 μL of 5 μM each primer specific for human GAPDH coding region. Store at -20°C. FOR: GGC TCC CAC CTT TCT CAT CC REV: GGC CAT CCA CAG TCT TCT GG |
| Quality Level | MQ100 |
| Applications | |
|---|---|
| Application | This ChIPAb+ Monomethyl-Histone H3 (Lys4) -ChIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers. |
| Key Applications |
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| Application Notes | Chromatin Immunoprecipitation: Sonicated chromatin prepared from HeLa cells (1 X 10E6 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 2 µg of either Normal Mouse IgG , or 2 µg Anti-Monomethyl-Histone H3 (Lys4)and the Magna ChIP™ G Kit (Cat. # 17-611). Successful immunoprecipitation of Monomethyl-Histone H3 (Lys4) associated DNA fragments was verified by qPCR using ChIP Primers GAPDH Coding Region as a positive locus, and GAPDH promoter primers (22-004) as a negative locus. (Figure 2). Data is presented as percent input of each IP sample relative to input chromatin for each amplicon and ChIP sample as indicated. Please refer to the EZ-Magna ChIP™ G (Cat. # 17-409) or EZ-ChIP™ (Cat. # 17-371) protocol for experimental details. Specificity data: HeLa acid extracts were resolved by electrophoresis, transferred to PVDF, and probed with anti-Monomethyl-Histone H3 (Lys4) (1:5000 dilution).To demonstrate specificity, antibody was preincubated with modified histone peptides: Lane 1 No peptide Lane 2 unmodified Histone H3 Lane 3 Monomethyl Histone H3 (Lys4) Lane 4 Dimethyl Histone H3 (Lys4) Lane 5 Trimethyl Histone H3 (Lys4) Lane 6 Trimethyl Histone H3 (Lys9) Lane 7 Trimethyl Histone H3 (Lys27) Lane 8 Trimethyl Histone H4 (Lys20) Proteins were visualized using donkey anti-mouse IgG conjugated to HRP and a chemiluminescence detection system. |
| Biological Information | |
|---|---|
| Immunogen | Peptide corresponding to Histone H3 containing the sequence [ART(me1-K)QTARKSTGC] on which Lys4 is acetylated. |
| Epitope | a.a. 1-12 |
| Host | Mouse |
| Specificity | This antibody detects Monomethyl-Histone H3 methylated at Lys4. |
| Isotype | IgG2bκ |
| Species Reactivity |
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| Species Reactivity Note | Wide range of cross-reactivity expected based on sequence homology. |
| Antibody Type | Monoclonal Antibody |
| Entrez Gene Number |
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| Entrez Gene Summary | Histones 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 |
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| Modifications |
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| Purification Method | Protein A Purfied |
| UniProt Number |
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| UniProt Summary | FUNCTION: 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. |
| Molecular Weight | 17 kDa |
| Product Usage Statements | |
|---|---|
| Quality Assurance | Chromatin Immunoprecipitation: Sonicated chromatin prepared from HeLa cells (1 X 10E6 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 2 µg of either Normal Mouse IgG, or 2 µg Anti-monomethyl-Histone H3 (Lys4) and the Magna ChIP™ G Kit (Cat. # 17-611). Successful immunoprecipitation of Monomethyl-Histone H3 (Lys4) associated DNA fragments was verified by qPCR using ChIP Primers GAPDH Coding Region (Figure 1). Please refer to the EZ-Magna ChIP™ G (Cat. # 17-409) or EZ-ChIP™ (Cat. # 17-371) protocol for experimental details. (Figure 1). Please refer to the EZ-Magna ChIP™ A (Cat. # 17-408) or EZ-ChIP™ (Cat. # 17-371) protocol for experimental details. |
| Usage Statement |
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| Storage and Shipping Information | |
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| Storage Conditions | Stable for 1 year at -20°C from date of receipt. Handling Recommendations: Upon first thaw, and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance. |
| Packaging Information | |
|---|---|
| Material Size | 25 assays |
| Material Package | 25 assays per set. Recommended use: 2 μg of antibody per chromatin immunoprecipitation (dependent upon biological context). |