Lefrançois Philippe, Zheng Wei, Snyder Michael
Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA.
Methods Enzymol. 2010;470:77-104. doi: 10.1016/S0076-6879(10)70004-5. Epub 2010 Mar 1.
Much of eukaryotic gene regulation is mediated by binding of transcription factors near or within their target genes. Transcription factor binding sites (TFBS) are often identified globally using chromatin immunoprecipitation (ChIP) in which specific protein-DNA interactions are isolated using an antibody against the factor of interest. Coupling ChIP with high-throughput DNA sequencing allows identification of TFBS in a direct, unbiased fashion; this technique is termed ChIP-Sequencing (ChIP-Seq). In this chapter, we describe the yeast ChIP-Seq procedure, including the protocols for ChIP, input DNA preparation, and Illumina DNA sequencing library preparation. Descriptions of Illumina sequencing and data processing and analysis are also included. The use of multiplex short-read sequencing (i.e., barcoding) enables the analysis of many ChIP samples simultaneously, which is especially valuable for organisms with small genomes such as yeast.
真核生物的基因调控大多是通过转录因子与目标基因附近或内部的结合来介导的。转录因子结合位点（TFBS）通常使用染色质免疫沉淀（ChIP）进行全基因组鉴定，其中使用针对目标因子的抗体分离特定的蛋白质 - DNA 相互作用。将 ChIP 与高通量 DNA 测序相结合，可以直接、无偏差地鉴定 TFBS；这种技术称为 ChIP 测序（ChIP-Seq）。在本章中，我们描述了酵母 ChIP-Seq 实验步骤，包括 ChIP、输入 DNA 制备和 Illumina DNA 测序文库制备的方案。还包括 Illumina 测序以及数据处理和分析的描述。多重短读测序（即条形码技术）的使用能够同时分析多个 ChIP 样本，这对于像酵母这样基因组较小的生物体尤其有价值。
