Cell Signaling Technology

Product Pathways - Cell Cycle / Checkpoint

Rpb1 CTD (4H8) Mouse mAb #2629

chip-on-chip   chip-validated   chipkit   chromatin-IP   CTD   CTD-4H8   immunoprecipitation   RNA pol   rna-pol-II-CTD   rna-polymerase-II   rnapII   rpb1   sc-56767   sc-899  

No. Size Price
2629S 100 µl ( 10 western blots ) ¥3,100.00 现货查询 购买询价
2629 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Mouse,Rat,Monkey, Endogenous 250 Mouse IgG1
IP 1:50
ChIP 1:50

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, ChIP=Chromatin IP,

Homology

Species predicted to react based on 100% sequence homology: Hamster, D. melanogaster, S. cerevisiae,

Specificity / Sensitivity

Rpb1 CTD (4H8) Antibody detects endogenous levels of total Rpb1 protein (both phosphorylated and unphosphorylated forms).Rpb1 CTD (4H8) Antibody 能够检测内源性Rpb1总蛋白(包括磷酸化和非磷酸化形式)。

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide containing 10 heptapeptide repeats [Tyr1, Ser2, Pro3, Thr4, Ser5, Pro6, Ser7] in which Ser5 is phosphorylated.该单克隆抗体是由合成的针对10次重复的七肽重复序列[Tyr1, Ser2, Pro3, Thr4, Ser5, Pro6, Ser7]中丝氨酸(5位)的磷酸化肽段免疫动物生产的。

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Rpb1 CTD (4H8) Mouse mAb.Western blot 方法检测不同细胞提取物,使用的抗体为Rpb1 CTD (4H8) Mouse mAb。

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells and either 10 μl of Rpb1 CTD (4H8) Mouse mAb or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human RPL30 Exon 3 Primers #7014, SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human MyoD1 Exon 1 Primers #4490, and SimpleChIP® Human MYT-1 Exon 1 Primers #4493. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.采用来自4 x 106 Hela细胞的交叉结合染色质进行免疫共沉淀实验,细胞通过紫外线处理(100 J/m2,恢复3小时)和10 μl Rpb1 CTD (4H8) Mouse mAb或者2 μl Normal Rabbit IgG #2729,使用的试剂盒为SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003。随后通过实时PCR定量富集的DNA,使用的引物为SimpleChIP® Human RPL30 Exon 3 Primers #7014,SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human MyoD1 Exon 1 Primers #4490,和SimpleChIP® Human MYT-1 Exon 1 Primers #4493。每个样品的免疫沉淀DNA的量以相对于输入染色质总量(相当于1)的信号来表示

Background

RNA polymerase II is a complex of 12 proteins that participates in transcription, mRNA processing, and transcription-coupled DNA repair (1,2). The largest subunit, Rpb1, contains a unique heptapeptide: Tyr1, Ser2, Pro3, Thr4, Ser5, Pro6, Ser7, which is repeated up to 52 times in the carboxy-terminal domain (CTD) of Rpb1 (3). This CTD is phosphorylated by cyclin-dependent kinases (CDKs), p44/42 MAPK, DNA-PKcs, and c-Abl (4). DNA damage caused by UV-light, hydrogen peroxide, or cisplatin results in ubiquitination and proteasomal degradation of Rpb1 (1,3). The kinase inhibitor 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole prevents ubiquitination of Rpb1, suggesting that CTD phosphorylation is required for proteolysis (5). RNA 聚合酶 II是由12种蛋白组成的复合体,参与转录、mRNA处理和转录偶联DNA修复(1,2)。最大的亚基Rpb1包含一个独特的七肽:Tyr1, Ser2, Pro3, Thr4, Ser5, Pro6, Ser7,在Rpb1的羧基末端结构域(CTD)发生高达52次的重复(3)。该结构域能够被周期蛋白依赖性激酶 (CDKs),p44/42 MAPK,DNA-PKcs和c-Abl 磷酸化(4)。紫外线、过氧化氢和顺铂引起的DNA损伤能够导致Rpb1的泛素化和蛋白酶体降解(1,3)。激酶抑制剂5,6-二氯-1-β-D-呋喃核糖基苯并咪唑能够阻止Rpb1的泛素化表明CTD磷酸化为蛋白酶解所必需(5)。

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  14. Egloff, S. et al. (2010) J Biol Chem 285, 20564-9.
  15. Egloff, S. et al. (2012) Mol Cell 45, 111-22.

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Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.

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