SimpleChIP? Mouse Oct-4 Promoter Primers

• 产品编号：CST-4653S      品牌：CST       原厂货号：4653S
• 产品分类：PCR/RT-PCR/qRT-PCR > 其它 > 引物 > 其他引物与寡核苷酸
• 应用分类：

描述：

小鼠Oct-4启动子引物含有一个正向反向PCR引物混合物，特异针对人类Oct-4启动子区域。这些引物可能被用于扩增被染色质免疫沉淀分离的DNA。引物已经被优化用于SYBR®绿色定量实时PCR并由SimpleChIP®酶染色质IP试剂盒#9002 和#9003以及Cell Signaling Technology公司ChIP验证的抗体测试过。Oct-4基因在干细胞中高度活跃，启动子被Oct-4, Sox2, 和Nanog蛋白结合。染色质免疫沉淀分析(ChIP)是一种通用强效用于探测细胞染色质自然状态蛋白DNA相互作用的技术[1,2]。这种分析可以被用于确定与基因组特异区域相关的多种蛋白或确定被特异蛋白结合的基因组内各种区域[3-6]。ChIP可被用于检测各种蛋白被招募到基因启动子的特异顺序或测量整个基因座特殊组蛋白修饰相对量[3,4]。除组蛋白外，ChIP分析可用于分析转录因子和共因子，DNA复制因子，DNA修复蛋白，当做ChIP实验时，细胞首先由甲醛（可逆蛋白DNA 交联剂）固定，保持DNA蛋白相互作用在细胞内[1,2]。细胞裂解，收获染色质超声或酶解成片段。染色质片段由特异蛋白或组蛋白修饰免疫沉淀。任何序列与蛋白或组蛋白相关DNA将随着部分交联染色质复合物被共沉淀，相对量DNA序列由免疫筛选富集。在免疫沉淀后，蛋白DNA交联被逆转，DNA被纯化。标准PCR或实时定量PCR常通过蛋白特异沉淀用于测量特殊DNA序列的富集量。另外，ChIP实验能与基因组铺板芯片技术，高通量测序激素，或者克隆技术联用，所有这些可对蛋白质DNA相互作用和组蛋白修饰进行基因组范围分析[5-8]。SimpleChIP®引物已被优化用于实时定量PCR扩增ChIP分离DNA，并提供重要阳性和阴性对照用于确认ChIP实验成功。

1. Orlando, V. (2000) Trends Biochem Sci 25, 99-104.
2. Kuo, M.H. and Allis, C.D. (1999) Methods 19, 425-33.
3. Agalioti, T. et al. (2000) Cell 103, 667-78.
4. Soutoglou, E. and Talianidis, I. (2002) Science 295, 1901-4.
5. Mikkelsen, T.S. et al. (2007) Nature 448, 553-60.
6. Lee, T.I. et al. (2006) Cell 125, 301-13.
7. Weinmann, A.S. and Farnham, P.J. (2002) Methods 26, 37-47.
8. Wells, J. and Farnham, P.J. (2002) Methods 26, 48-56.

原厂资料：

Description

SimpleChIP® Mouse Oct-4 Promoter Primers contain a mix of forward and reverse PCR primers that are specific to a region of the mouse Oct-4 promoter. These primers can be used to amplify DNA that has been isolated using chromatin immunoprecipitation (ChIP). Primers have been optimized for use in SYBR® Green quantitative real-time PCR and have been tested in conjunction with SimpleChIP® Enzymatic Chromatin IP Kits #9002 and #9003 and ChIP-validated antibodies from Cell Signaling Technology®. The Oct-4 gene is highly active in stem cells and the promoter is bound by Oct-4, Sox2, and Nanog proteins.

Background

The chromatin immunoprecipitation (ChIP) assay is a powerful and versatile technique used for probing protein-DNA interactions within the natural chromatin context of the cell (1,2). This assay can be used to either identify multiple proteins associated with a specific region of the genome or to identify the many regions of the genome bound by a particular protein (3-6). ChIP can be used to determine the specific order of recruitment of various proteins to a gene promoter or to "measure" the relative amount of a particular histone modification across an entire gene locus (3,4). In addition to histone proteins, the ChIP assay can be used to analyze binding of transcription factors and co-factors, DNA replication factors, and DNA repair proteins. When performing the ChIP assay, cells are first fixed with formaldehyde, a reversible protein-DNA cross-linking agent that "preserves" the protein-DNA interactions occurring in the cell (1,2). Cells are lysed and chromatin is harvested and fragmented using either sonication or enzymatic digestion. Fragmented chromatin is then immunoprecipitated with antibodies specific to a particular protein or histone modification. Any DNA sequences that are associated with the protein or histone modification of interest will co-precipitate as part of the cross-linked chromatin complex and the relative amount of that DNA sequence will be enriched by the immunoselection process. After immunoprecipitation, the protein-DNA cross-links are reversed and the DNA is purified. Standard PCR or quantitative real-time PCR are often used to measure the amount of enrichment of a particular DNA sequence by a protein-specific immunoprecipitation (1,2). Alternatively, the ChIP assay can be combined with genomic tiling micro-array (ChIP on chip) techniques, high throughput sequencing (ChIP-Seq), or cloning strategies, all of which allow for genome-wide analysis of protein-DNA interactions and histone modifications (5-8). SimpleChIP® primers have been optimized for amplification of ChIP-isolated DNA using real-time quantitative PCR and provide important positive and negative controls that can be used to confirm a successful ChIP experiment.

1. Orlando, V. (2000) Trends Biochem Sci 25, 99-104.
2. Kuo, M.H. and Allis, C.D. (1999) Methods 19, 425-33.
3. Agalioti, T. et al. (2000) Cell 103, 667-78.
4. Soutoglou, E. and Talianidis, I. (2002) Science 295, 1901-4.
5. Mikkelsen, T.S. et al. (2007) Nature 448, 553-60.
6. Lee, T.I. et al. (2006) Cell 125, 301-13.
7. Weinmann, A.S. and Farnham, P.J. (2002) Methods 26, 37-47.
8. Wells, J. and Farnham, P.J. (2002) Methods 26, 48-56.

注意事项：

Storage: Supplied in nuclease-free water at a concentration of
5 μM (each primer is at a final concentration of 5 μM). Store at
-20°C.