DNA Replication Antibody Sampler Kit

• 产品编号：CST-8341S      品牌：Cell Signaling Technology       原厂货号：8341S
• 产品分类：抗体 > 一抗 > 复合抗体试剂盒
• 应用分类：

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描述：

单克隆抗体是由合成的人源的针对MCM2蛋白氨基末端肽段或MCM3蛋白氨基末端肽段、MCM7蛋白羧基末端肽段、RPA70蛋白苏氨酸（164位）肽段免疫动物生产的。 单克隆抗体由重组的蛋白A-PCNA融合蛋白或全长重组人p58引发酶免疫动物生产的。多克隆抗体是由合成的人源的针对CDT1蛋白氨基末端序列免疫动物，采用蛋白A和多肽亲和层析技术纯化生产的。DNA Replication Antibody Sampler Kit能够快捷经济地评价调节DNA复制的多种靶蛋白。该试剂盒内含足够4次western blot实验的一抗和二抗。哺乳动物细胞中的DNA复制起始是一个受几种蛋白质复合物调控的高度协调过程。复制起始物(ORCs)，复制开始的地方，分散在整个基因组。经由前复制和复制因子的有序结合来启动复制叉形成可以调节它们的活性，复制叉是DNA合成的活性结构。起始识别复合物被认为在整个细胞周期结合于染色质(1,2)。前复制复合物(Pre-RC)形式在有丝分裂后期/ G1早期，始于CDT1和CDC6结合到起始物。连同CDT1和CDC6能够促进微小染色体维持蛋白复合体(MCM)的装载。此过程被称为“染色质发牌”。许可允许的染色质DNA每次细胞周期只复制一次，有助于确保不会发生遗传变异和肿瘤细胞生长(3中已论述)。经典的MCM蛋白复合体(MCM2-7)是一个六个相关磷酸蛋白家族，可作为真核细胞DNA复制解旋酶部分地发挥功能(3,4)。MCM2，MCM3，MCM4和MCM6亚基的磷酸化和泛素化调节MCM复合体的活性并启动DNA合成(5-7)。MCM蛋白在DNA复制过程中被清除，引起染色质无照，抑制Pre-RC改革。除了DNA聚合酶，DNA复制起始需要一对引发酶亚基。 DNA引物酶的活性催化前导链和后随链上RNA /DNA引物(引发剂DNA)的从头合成，而聚合酶活性延伸引发剂DNA(8)。48和58 kDa的引物酶亚基在小RNA引物的合成中合作。P48是活性的催化亚单位(9)，而p58 偶联p48至聚合酶允许引物转移至活性位点。P58亚基可能也在引物长度的调控中发挥了作用(10,11)。一旦开始复制，增殖细胞核抗原(PCNA) 作为DNA聚合酶Δ和ξ的辅助因子，拴住这些聚合酶到复制过程中的模板DNA。PCNA与DNA聚合酶的相互作用能增加前导链的持续合成。PCNA，DNA滑动压板蛋白家组的一员，是一个同源三聚体环，环绕沿着作为复制叉进展的DNA双螺旋结构(12)。PCNA和调节DNA合成的多种蛋白质参与DNA复制，DNA修复，细胞周期调控并调节DNA合成。PCNA以ATP-依赖的方式的被装载到DNA上面，通过多蛋白压板装载器-复制因子C(RFC)(13)。RFC转而通过与DNA单链结合蛋白复合物复制蛋白A(RPA)相互作用来协同DNA。经典的RPA复合物是异源三聚体且由RPA1 (RPA70)， RPA2 (RPA32)和RPA3 (RPA14)亚基组成。RPA能够识别和稳定修复过程中的单链DNA以及DNA重组，并在复制过程中起重要作用(14-17)。

原厂资料：

Specificity / Sensitivity

All antibodies recognize endogenous levels of the respective target protein.

Source / Purification

Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near: the amino terminus of human MCM2 protein, the amino terminus of human MCM3 protein, the carboxy terminus of human MCM7 protein, or residues surrounding Thr164 of human RPA70 protein. Monoclonal antibodies are produced by immunizing animals with: a recombinant Protein A-PCNA fusion protein or full-length human recombinant p58 primase. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human CDT1 protein. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.

Description

The DNA Replication Antibody Sampler Kit provides a fast and economical means of evaluating multiple targets regulating DNA replication. The kit contains enough primary antibodies to perform four western blots with each antibody.

Background

The initiation of DNA replication in mammalian cells is a highly coordinated process that is regulated by several protein complexes. Origins of replication (ORCs), at which replication is initiated, are dispersed throughout the genome. Their activities are regulated via the sequential binding of pre-replication and replication factors that initiate formation of replication forks, the active structures at which DNA is synthesized. The origin recognition complex is thought to be bound to chromatin throughout the cell cycle (1,2). The pre-replication complex (Pre-RC) forms in late mitosis/early G1 phase beginning with the binding of CDT1 and CDC6 to the origin. Together CDT1 and CDC6 promote the loading of the heterohexameric minichromosome maintenance (MCM) complex. This process is referred to as chromatin licensing. Licensing of the chromatin permits the DNA to replicate only once per cell cycle, helping to ensure that genetic alterations and malignant cell growth do not occur (reviewed in 3). The canonical MCM complex proteins (MCM2-7) are a family of six related phospho-proteins that function, in part, as the eukaryotic replicative DNA helicase (3,4). Phosphorylation and ubiquitination of the MCM2, MCM3, MCM4, and MCM6 subunits appears to regulate MCM complex activity and the initiation of DNA synthesis (5-7). MCM proteins are removed during DNA replication, causing chromatin to become unlicensed, inhibiting Pre-RC reformation. In addition to DNA polymerase, initiation of DNA replication requires a pair of primase subunits. DNA Primase activity catalyzes de novo synthesis of an RNA/DNA primer (initiator DNA) on the leading and lagging strands, while polymerase activity extends the initiator DNA (8). The 48 and 58 kDa primase subunits cooperate in the synthesis of small RNA primers. p48 is the catalytically active subunit (9), while p58 couples p48 to the polymerase to allow the transfer of primers to the active site. The p58 subunit may also play a role in regulation of primer length (10,11). Once replication is initiated, Proliferating Cell Nuclear Antigen (PCNA) serves as an accessory factor for DNA polymerases delta and epsilon, acting to tether these polymerases to template DNA during replication. Interactions of PCNA with DNA polymerases increase the processivity of leading strand synthesis. PCNA, a member of DNA sliding clamp family, is a homotrimeric ring complex that encircles and slides along the DNA double helix as the replication fork progresses (12). Multiple proteins involved in DNA replication, DNA repair, and cell cycle control bind to PCNA and regulate DNA synthesis. PCNA is loaded onto the DNA in an ATP-dependent manner by a multiprotein clamp loader, Replication Factor C (RFC) (13). RFC, in turn, associates with DNA via interactions with the single-stranded DNA binding protein complex, Replication Protein A (RPA). The canonical RPA complex is heterotrimeric and composed of RPA1 (RPA70), RPA2 (RPA32), and RPA3 (RPA14) subunits. RPA recognizes and stabilizes single stranded DNA in repair processes and DNA recombination, and plays a role in replication (14-17). 

注意事项：

Storage: Antibodies are supplied in 10 mM sodium HEPES
(pH 7.5), 150 mM NaCl, 100 μg/ml BSA, 50% glycerol, and less
than 0.02% sodium azide. Store at –20°C. Do not aliquot the
antibodies