H3K36me3 Polyclonal Antibody

$ChIP was performed with 5 \u03bcl of the antibody against H3K36me3 (bs-53041R) on sheared chromatin from 1 million HeLaS3 cells. IgG (2 \u03bcg\/IP) was used as a negative IP control. The IP\u2019d DNA was analysed by QPCR with optimized PCR primer pairs for the coding and promoter region of the active GAPDH gene, for the coding region of the inactive TSH2B gene and for the Sat2 satellite repeat. The IP\u2019d DNA was subsequently analysed with an Illumina Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer\u2019s instructions. The 36 bp tags were aligned to the human genome using the ELAND algorithm. The figure shows the results in 200 kb regions of chromosome 12 (including the GAPDH positive control), 6 and 7 and 14. These results clearly show an enrichment of the H3K36me3 at active genes$

ChIP was performed with 5 \u03bcl of the antibody against H3K36me3 (bs-53041R) on sheared chromatin from 1 million HeLaS3 cells. IgG (2 \u03bcg\/IP) was used as a negative IP control. The IP\u2019d DNA was analysed by QPCR with optimized PCR primer pairs for the coding and promoter region of the active GAPDH gene, for the coding region of the inactive TSH2B gene and for the Sat2 satellite repeat. The IP\u2019d DNA was subsequently analysed with an Illumina Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer\u2019s instructions. The 36 bp tags were aligned to the human genome using the ELAND algorithm. The figure shows the results in 200 kb regions of chromosome 12 (including the GAPDH positive control), 6 and 7 and 14. These results clearly show an enrichment of the H3K36me3 at active genes

$ChIP assays were performed using human osteosarcoma (U2OS) cells, the antibody against H3K36me3 (bs-53041R) and optimized PCR primer sets for qPCR. ChIP was performed using sheared chromatin from 1.6 million cells. A titration of the antibody consisting of 2, 5, 10 and 15 \u03bcl per ChIP experiment was analyzed. IgG (5 \u03bcg\/IP) was used as a negative IP control. Quantitative PCR was performed using primer sets for the housekeeping gene GAPDH and for myogenic differentiation gene (MYOD). The figure shows the recovery expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis.$

ChIP assays were performed using human osteosarcoma (U2OS) cells, the antibody against H3K36me3 (bs-53041R) and optimized PCR primer sets for qPCR. ChIP was performed using sheared chromatin from 1.6 million cells. A titration of the antibody consisting of 2, 5, 10 and 15 \u03bcl per ChIP experiment was analyzed. IgG (5 \u03bcg\/IP) was used as a negative IP control. Quantitative PCR was performed using primer sets for the housekeeping gene GAPDH and for myogenic differentiation gene (MYOD). The figure shows the recovery expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis.

To determine the titer, an ELISA was performed using a serial dilution of the antibody directed against H3K36me3 (bs-53041R). The antigen used was a peptide containing the histone modification of interest. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:12,700.

A Dot Blot analysis was performed to test the cross-reactivity of the antibody against H3K36me3 (bs-53041R) with peptides containing other modifications of histone H3. Other histone modifications include mono- and dimethylation of the same lysine and mono-, di- and trimethylation of lysine 9, 27 and 79. One hundred to 0.2 pmol of peptide containing the respective histone modification were spotted on a membrane. The antibody was used at a dilution of 1:100,000. The figure shows a high specificity of the antibody for the modification of interest.

$Histone extracts (15 \u03bcg) from HeLa cells were analyzed by Western blot using the antibody against H3K36me3 (bs-53041R) diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is indicated on the left; the marker (in kDa) is shown on the right.$

Histone extracts (15 \u03bcg) from HeLa cells were analyzed by Western blot using the antibody against H3K36me3 (bs-53041R) diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is indicated on the left; the marker (in kDa) is shown on the right.

Applications

WB
ELISA
dot-blot
ChIP-seq

Reactivity

Human
Mouse

Overview
Catalog #	bs-53041R
Product Name	H3K36me3 Polyclonal Antibody
Applications	WB, ELISA, dot-blot, ChIP-seq
Reactivity	Human, Mouse
Specifications
Conjugation	Unconjugated
Host	Rabbit
Source	Polyclonal antibody raised in rabbit against histone H3 containing the trimethylated lysine 36 (H3K36me3), using a KLH-conjugated synthetic peptide
Clonality	Polyclonal
Isotype	Not Determined
Concentration	Not Determined
Purification	Whole antiserum from rabbit containing 0.05% azide
Storage Buffer	Serum with 0.05% sodium azide.
Storage Condition	Store at -20°C for 12 months.
Target
Gene ID	8350
Swiss Prot	P68431
Background	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.
Application Dilution
WB	1:300-5000
ELISA	1:500-1000
dot-blot
ChIP-seq