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H3k36me3 ab9050

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H3K36me3 (ab9050) is a recombinant monoclonal antibody that recognizes the trimethylated form of histone H3 at lysine 36. This antibody is intended for use in various research applications to detect and study this histone modification.

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Lab products found in correlation

Cleaved caspase 3 9664, by Cell Signaling Technology (1 mentions) Radioimmunoprecipitation assay (ripa), by Thermo Fisher Scientific (1 mentions) Bca protein assay kit, by Beyotime (1 mentions) Protease inhibitor cocktail, by Beyotime (1 mentions) Phenyl methyl sulfonyl fluoride (pmsf), by Beyotime (1 mentions) Ecl kit, by Beyotime (1 mentions) Nucleoprotein extraction kit, by Sangon (1 mentions) Chip sequencing sample preparation kit, by Illumina (1 mentions) Simplechip enzymatic chromatin ip kit, by Cell Signaling Technology (1 mentions) Hiseq 2000, by Illumina (1 mentions) H3k27ac ab4729, by Abcam (1 mentions) H3k27me3 ab6002, by Abcam (1 mentions) H3k4me1 ab8895, by Abcam (1 mentions) H3k4me2 ab7766, by Abcam (1 mentions) Hiseq 3000, by Illumina (1 mentions) Anti med1 a300 793a, by Fortis Life Sciences (1 mentions) Anti h3k27ac ab4729, by Abcam (1 mentions) Anti tubulin sc 23948, by Santa Cruz Biotechnology (1 mentions) Donkey anti rabbit alexafluor 488 a21206, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

α-H3K36me3 (ab9050), Anti-histone H3K36me3 (clone ab9050, Ab9050 H3K36me3

6 protocols using h3k36me3 ab9050

1

Protein Extraction and Western Blot Analysis

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Total protein from tissues and cell lines was extracted using RIPA (Thermo Scientific, Rockford, IL, USA), and nuclear protein was prepared using the Nucleoprotein Extraction Kit (Sangon Biotech, Shanghai, China) in the presence of protease inhibitor cocktail and PMSF (Beyotime Institute of Biotechnology, Haimen, China). The concentration of various protein samples was determined using the BCA Protein assay kit (Beyotime Institute of Biotechnology) according to the manufacturer's instructions. Thirty micrograms of each lysate was electrophoresed in 10% SDS-PAGE gels and then transferred onto PVDF membranes and detected using the ECL kit (Beyotime Institute of Biotechnology). The SETD2 (ab69836), p53 (ab28) and H3K36me3 (ab9050) antibodies were purchased from Abcam (Cambridge, MA, USA). GAPDH (D16H11), Caspase-3 (#9662) and cleaved Caspase-3 (#9664) antibodies were obtained from Cell Signaling Technology Inc. (Vebery, MA, USA). H3 antibody was ordered from Beyotime Institute of Biotechnology. Anti-rabbit and anti-mouse IgG secondary antibodies conjugated with horseradish peroxidase were provided by Wuhan Boster Bio-engineering Limited Company (Wuhan, China). ImageJ Software version 1.36b was applied to analyze the expression levels of different proteins.
Xiang W., He J., Huang C., Chen L., Tao D., Wu X., Wang M., Luo G., Xiao X., Zeng F, & Jiang G. (2015). miR-106b-5p targets tumor suppressor gene SETD2 to inactive its function in clear cell renal cell carcinoma. Oncotarget, 6(6), 4066-4079.
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2

ChIP-seq analysis of histone modifications

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Isolated PBMCs were treated with formaldehyde (at a final concentration of 1%) to crosslink histone and DNA. After formaldehyde was quenched by adding glycine, cells were then pelleted and washed with cold PBS for 2 times. Chromatin was then digested with Micrococcal Nuclease. After nuclear membrane was disrupted by brief sonication, the sample was centrifuged and the supernatant was used for chromatin immunoprecipitation with Simple ChIP-enzymatic Chromatin IP Kit (Cell signaling, #9003). The ChIP antibodies, H3K4me3 (ab1012), H3K27me3 (ab6002), H3K9me3 (ab8898) and H3K36me3 (ab9050) were purchased from Abcam (Cambridge, MA). The cross link was reversed by treating the immunoprecipitated chromatin with proteinase K. DNA was then purified and quantified. The sequencing library was constructed using Illumina's Chip Sequencing sample preparation kit (#1003473) according to the manufacturer's instruction and sequenced by Illumina HiSeq2000 at Tufts University Genomic core facility.
Bam M., Yang X., Zhou J., Ginsberg J.P., Leyden Q., Nagarkatti P.S, & Nagarkatti M. (2015). Evidence for Epigenetic Regulation of Pro-inflammatory Cytokines, Interleukin-12 and Interferon Gamma, in Peripheral Blood Mononuclear Cells from PTSD patients. Journal of neuroimmune pharmacology : the official journal of the Society on NeuroImmune Pharmacology, 11(1), 168-181.
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3

ChIP-seq Analysis of Epigenetic Marks

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ChIP/ChIP-seq was carried out as described previously (Blackledge et al., 2010 (link)). Full details are provided in the Supplemental Experimental Procedures. The antibodies for ChIP-seq were H3K27me3 (pAB-069-100; Diagenode), H3K36me3 (AB9050; Abcam), anti-Suz12 (3737S; Cell Signaling Technology), and anti-Ring1B (Atsuta et al., 2001 (link)).
Cooper S., Dienstbier M., Hassan R., Schermelleh L., Sharif J., Blackledge N.P., De Marco V., Elderkin S., Koseki H., Klose R., Heger A, & Brockdorff N. (2014). Targeting Polycomb to Pericentric Heterochromatin in Embryonic Stem Cells Reveals a Role for H2AK119u1 in PRC2 Recruitment. Cell Reports, 7(5), 1456-1470.
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4

MNase-ChIP-seq and MNase-seq Protocols

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MNase-ChIP-seq and MNase-seq protocols were performed according to previous studies44 (link). In brief, LNCaP cells were exposed to 10 nM DHT or DMSO (Veh) for 4 h. Mono-nucleosomes with solubilized chromatin was achieved by MNase digestion of 2 min at 37 °C, then immunoprecipitated with antibody-conjugated magnetic beads. DNA is phenol extracted and ethanol precipitated. Libraries were prepared from isolated DNA and sent for sequencing on the Illumina HiSeq3000 at the UTHSA sequencing core. All samples were performed in biological replicates. Antibodies include: H3K4me1 (ab8895) 1:500 dilution, H3K4me2 (ab7766) 1:250 dilution, H3K27ac (ab4729) 1:500 dilution, H3K27me3 (ab6002) 1:250 dilution, H3K36me3 (ab9050) 1:250 dilution, H3K79me2 (ab8898) 1:250 dilution from Abcam (Cambridge, MA). H3K4me3 (07-473) 1:500 dilution, H3K9me3 (17-10242) 1:250 dilution from Millipore (Upstate).
Li T., Liu Q., Chen Z., Fang K., Huang F., Fu X., Wang Q, & Jin V.X. (2022). Dynamic nucleosome landscape elicits a noncanonical GATA2 pioneer model. Nature Communications, 13, 3145.
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5

Histone H3.3 Variant Regulation

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The retroviral pQCXIP plasmids expressing FLAG-tagged wild type (WT) or mutants of histone H3.3 including K4M, K9M, K27M and K36 were described previously (13 (link)). The following homemade antibodies have been described: anti-MLL4#3 (14 (link)), anti-MLL3#3 (15 (link)) and anti-UTX (16 (link)). Anti-RbBP5 (A300-109A), anti-BRD4 (A301-985A100) and anti-MED1 (A300-793A) were from Bethyl Laboratories. Anti-SET1A/B antibodies were described previously (17 (link)). Anti-H3 (ab1791), anti-H3K4me1 (ab8895), anti-H3K4me2 (ab7766), anti-H3K27ac (ab4729) and H3K36me3 (ab9050) were from Abcam. Anti-MLL1N (A700-010), anti-MLL1C (A300-374A), anti-MLL2 (ABE500), anti-Pol II (17-672), anti-H3K4me3 (07-473), anti-H3K9me2 (17-648), anti-H3K27me3 (07-449) and anti-H3K36me2 (07-369) were from Millipore. Anti-FLAG-M2 (F3165) and (Z)-4-hydroxytamoxifen (4OHT) (H7904) were from Sigma. Drosophila spike-in chromatin (53083) and antibody (61686) were from Active Motif.
Jang Y., Broun A., Wang C., Park Y.K., Zhuang L., Lee J.E., Froimchuk E., Liu C, & Ge K. (2018). H3.3K4M destabilizes enhancer H3K4 methyltransferases MLL3/MLL4 and impairs adipose tissue development. Nucleic Acids Research, 47(2), 607-620.
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6

Comprehensive Antibody Validation Protocol

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Anti-SETD2 (LS-B12660-50) was from LSBIO (Seattle, WA); anti-H3K36me1 (ab9048), anti-H3K36me2 (ab9049), and H3K36me3 (ab9050) were from Abcam (Cambridge, UK); rabbit-anti-Perilipin A/B (P1873) was from Sigma; rabbit-anti-Collagen I (600-400-103) was from Rockland (PA, USA); anti-Lbp (11836-1AP) and donkey-anti-OPN (AF808) were from R&D Systems (Minneapolis, MN); anit-Osx (sc-133871) and anti-tubulin (sc-23948) were from Santa Cruz Biotechnology (Texas, USA); donkey-anti-goat cy3 (705-165-147) was from Jackson ImmunoResearch (PA, USA); and donkey-anti-rabbit Alexa Fluor 488 (A21206) was from Molecular Probes.
Wang L., Niu N., Li L., Shao R., Ouyang H, & Zou W. (2018). H3K36 trimethylation mediated by SETD2 regulates the fate of bone marrow mesenchymal stem cells. PLoS Biology, 16(11), e2006522.
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