Boedi Stefan, Reyes-Dominguez Yazmid, Strauss Joseph
Department of Applied Genetics and Cell Biology, AIT-Austrian Institute of Technology GmbH Health and Environment Department, Bioresources, University and Research Center, UFT Campus Tulln, Tulln/Donau, Austria.
Methods Mol Biol. 2012;944:221-36. doi: 10.1007/978-1-62703-122-6_16.
Chromatin immunoprecipitation (ChIP) is used to map the interaction between proteins and DNA at a specific genomic locus in the living cell. The protein-DNA complexes are stabilized already in vivo by reversible crosslinking and the DNA is sheared by sonication or enzymatic digestion into fragments suitable for the subsequent immunoprecipitation step. Antibodies recognizing chromatin-linked proteins, transcription factors, artificial tags, or specific protein modifications are then used to pull down DNA-protein complexes containing the target. After reversal of crosslinks and DNA purification locus-specific quantitative PCR is used to determine the amount of DNA that was associated with the target at a given time point and experimental condition. DNA quantification can be carried out for several genomic regions by multiple qPCRs or at a genome-wide scale by massive parallel sequencing (ChIP-Seq).
染色质免疫沉淀法（ChIP）用于在活细胞中特定基因组位点绘制蛋白质与DNA之间的相互作用。蛋白质-DNA复合物已在体内通过可逆交联得以稳定，DNA通过超声处理或酶切消化成适合后续免疫沉淀步骤的片段。然后使用识别染色质相关蛋白、转录因子、人工标签或特定蛋白质修饰的抗体来下拉包含靶标的DNA-蛋白质复合物。在交联逆转和DNA纯化后，使用位点特异性定量PCR来确定在给定时间点和实验条件下与靶标相关的DNA量。可以通过多重qPCR对几个基因组区域进行DNA定量，或者通过大规模平行测序（ChIP-Seq）在全基因组范围内进行DNA定量。
