Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S5) antibody - ChIP Grade

• Supplier: Abcam
• Catalog: ab5131
• Prices: $403.00
• Sizes: 50 µg
• Host: Rabbit
• Clonality: Polyclonal
• Isotype: IgG
• Applications: IP IHC-P ELISA IHC IHC-F ChIPseq WB ChIP ICC/IF ICC/IF
• Species Reactivities: Mouse, Rat, Human, Yeast, Xenopus, Arabidopsis thaliana, C. elegans, Drosophila, S. pombe, Zebrafish, Broad species
• Antigen: Synthetic peptide of Saccharomyces cerevisiae RNA polymerase II CTD repeat YSPTSPS, phosphorylated at S5
• Blocking Peptide: Human RNA polymerase II CTD repeat YSPTSPS (phospho S5) peptide
• Description: Rabbit Polyclonal
• Gene: CELE_F36A4.7
• Conjugate: Unconjugated

Product images

IHC - Wholemount of Caenorhabditis elegans larvae labelling RNA polymerase II CTD repeat YSPTSPS (phospho S5) with ab5131. The sample was incubated with primary antibody (1/500 in PBS + 3% BSA + 0.1% Triton X-100) for 12 hours at 4°C. ab150077, a goat anti-rabbit Alexa 488 - (1/1000), was used as the secondary antibody.See Abreview

All lanes : Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S5) antibody - ChIP Grade (ab5131) at 1/5000 dilutionLane 1 : Xenopus laevis whole tissue lysate treated with DMSO for 24 hoursLane 2 : Xenopus laevis whole tissue lysate treated with CDK inhibitor for 24 hoursSecondaryHRP-conjugated goat anti-rabbit IgG polyclonal at 1/10000 dilutiondeveloped using the ECL techniquePerformed under reducing conditions.

All lanes : Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S5) antibody - ChIP Grade (ab5131) at 1 µg/mlLane 1 : HeLa (Human epithelial carcinoma cell line) Nuclear LysateLane 2 : S.cerevisiae (Y190) Whole Cell Lysate Lane 3 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate with S. cerevisiae RNA polymerase II CTD repeat YSPTSPS peptide (ab12795) at 1 µg/mlLane 4 : S.cerevisiae (Y190) Whole Cell Lysate with S. cerevisiae RNA polymerase II CTD repeat YSPTSPS peptide (ab12795) at 1 µg/mlLane 5 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate with Human RNA polymerase II CTD repeat YSPTSPS (phospho S5) peptide (ab18488) at 1 µg/mlLane 6 : S.cerevisiae (Y190) Whole Cell Lysate with Human RNA polymerase II CTD repeat YSPTSPS (phospho S5) peptide (ab18488) at 1 µg/mlLane 7 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate with S. cerevisiae RNA polymerase II CTD repeat YSPTSPS (phospho S2) peptide (ab12793) at 1 µg/mlLane 8 : S.cerevisiae (Y190) Whole Ce

Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 mins. The  ChIP was performed with 25 µg of chromatin, 2 µg of  ab5131 (blue), and 20 µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified on the inactive AFM and F8 promoters, the GAPDH promoter (active) and over the y-Actin gene (active). Schematic diagram of the y-Actin gene is shown on the top of the figure. Black boxes represent exons and thin lines represent introns. PCR products are depicted as bars under the gene.

Lanes 1, 3 & 4 : Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S5) antibody - ChIP Grade (ab5131) at 1/500 dilutionLane 2 : Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S5) antibody - ChIP Grade (ab5131) at 1/2000 dilutionLane 1 : HeLa nuclear extract Lane 2 : HeLa nuclear extract Lane 3 : HeLa nuclear extract with Human RNA polymerase II CTD repeat YSPTSPS (phospho S5) peptide (ab18488) at 1 µg/mlLane 4 : HeLa nuclear extract with S. cerevisiae RNA polymerase II CTD repeat YSPTSPS peptide (ab12795) at 1 µg/mlLysates/proteins at 20 µg per lane.SecondaryGoat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/5000 dilutionPerformed under reducing conditions.

ICC/IF image of ab5131 stained human HeLa cells. The cells were methanol fixed (5 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab5131, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue).

(Image courtesy of Human Protein Atlas) ab5131 staining RNA Polymerase in Human urinary bladder tissue. Paraffin embedded human skin tissue was incubated with ab5131 for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6. ab5131 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines. Further results for this antibody can be found at www.proteinatlas.org

HeLa cells were fixed with 4% formaldehyde in PEM buffer. The coverslip was incubated in blocking buffer of 5% powdered milk in TBS-T plus 0.02% sodium azide for 1 hour at room temperature. Blocking buffer was removed and primary antibody was added at a dilution of 1/160 and incubated overnight at 4 degrees celsius. The coverslips were then washed 4-5 times with blocking buffer for 5 minutes. Secondary antibody, goat anti-rabbit Alexa 594, was added at a dilution of 1/1000 and incubated at room temperature for one hour. From this point on coverslips were covered with foil to protect them from light. They were washed 5 times with TBS-T and then one time with PEM, for 5 minutes each wash. The coverslips were fixed 10-30 minutes in 4% formaldehyde in PEM buffer, then washed 3 times with PEM buffer for 5 minutes. 0.1M ammonium chloride in PEM buffer was added for 10 minutes to quench auto-florescence, and then slips were washed 2 times for 5 minutes in PEM followed by 3 washes for 5 minute

Product References

Integrated transcriptome analysis of mouse spermatogenesis. - Integrated transcriptome analysis of mouse spermatogenesis.

Margolin G, Khil PP, Kim J, Bellani MA, Camerini-Otero RD. BMC Genomics. 2014 Jan 18;15:39.

Targeted chromatin profiling reveals novel enhancers in Ig H and Ig L chain Loci. - Targeted chromatin profiling reveals novel enhancers in Ig H and Ig L chain Loci.

Predeus AV, Gopalakrishnan S, Huang Y, Tang J, Feeney AJ, Oltz EM, Artyomov MN. J Immunol. 2014 Feb 1;192(3):1064-70.

The transcript elongation factor SPT4/SPT5 is involved in auxin-related gene - The transcript elongation factor SPT4/SPT5 is involved in auxin-related gene

Durr J, Lolas IB, Sorensen BB, Schubert V, Houben A, Melzer M, Deutzmann R, Grasser M, Grasser KD. Nucleic Acids Res. 2014 Apr;42(7):4332-47.

Polycomb binding precedes early-life stress responsive DNA methylation at the Avp - Polycomb binding precedes early-life stress responsive DNA methylation at the Avp

Murgatroyd C, Spengler D. PLoS One. 2014 Mar 5;9(3):e90277.

A dual role for the histone methyltransferase PR-SET7/SETD8 and histone H4 lysine - A dual role for the histone methyltransferase PR-SET7/SETD8 and histone H4 lysine

Kapoor-Vazirani P, Vertino PM. J Biol Chem. 2014 Mar 14;289(11):7425-37.

Nonproteolytic roles of 19S ATPases in transcription of CIITApIV genes. - Nonproteolytic roles of 19S ATPases in transcription of CIITApIV genes.

Maganti N, Moody TD, Truax AD, Thakkar M, Spring AM, Germann MW, Greer SF. PLoS One. 2014 Mar 13;9(3):e91200.

Super-resolution microscopy reveals decondensed chromatin structure at - Super-resolution microscopy reveals decondensed chromatin structure at

Wang Y, Maharana S, Wang MD, Shivashankar GV. Sci Rep. 2014 Mar 26;4:4477.

Simultaneous mapping of transcript ends at single-nucleotide resolution and - Simultaneous mapping of transcript ends at single-nucleotide resolution and

Park D, Morris AR, Battenhouse A, Iyer VR. Nucleic Acids Res. 2014 Apr;42(6):3736-49.

Loss of Caenorhabditis elegans BRCA1 promotes genome stability during replication - Loss of Caenorhabditis elegans BRCA1 promotes genome stability during replication

Wolters S, Ermolaeva MA, Bickel JS, Fingerhut JM, Khanikar J, Chan RC, Schumacher B. Genetics. 2014 Apr;196(4):985-99.

The U4/U6 recycling factor SART3 has histone chaperone activity and associates - The U4/U6 recycling factor SART3 has histone chaperone activity and associates

Long L, Thelen JP, Furgason M, Haj-Yahya M, Brik A, Cheng D, Peng J, Yao T. J Biol Chem. 2014 Mar 28;289(13):8916-30.