Kidder Benjamin L
Department of Oncology, Wayne State University School of Medicine; Karmanos Cancer Institute, Wayne State University School of Medicine;
J Vis Exp. 2018 May 25(135):57481. doi: 10.3791/57481.
Embryonic stem (ES) cell self-renewal and differentiation is governed by extrinsic signals and intrinsic networks of transcription factors, epigenetic regulators, and post-translation modifications of histones that combinatorially influence the gene expression state of nearby genes. RNA has also been shown to interact with various proteins to regulate chromatin dynamics and gene expression. Chromatin-associated RNA immunoprecipitation (CARIP) followed by next-generation sequencing (CARIP-Seq) is a novel method to survey RNAs associated with chromatin proteins, while chromatin immunoprecipitation followed by next-generation sequencing (ChIP-Seq) is a powerful genomics technique to map the location of post-translational modification of histones, transcription factors, and epigenetic modifiers on a global-scale in ES cells. Here, we describe methods to perform CARIP-Seq and ChIP-Seq, including library construction for next-generation sequencing, to generate global chromatin-associated RNA and epigenomic maps in ES cells.
胚胎干细胞（ES细胞）的自我更新和分化受外在信号以及转录因子、表观遗传调控因子和组蛋白翻译后修饰的内在网络所支配，这些因素共同影响附近基因的基因表达状态。RNA也已被证明可与多种蛋白质相互作用，以调节染色质动态和基因表达。染色质相关RNA免疫沉淀（CARIP）结合下一代测序（CARIP-Seq）是一种用于检测与染色质蛋白相关的RNA的新方法，而染色质免疫沉淀结合下一代测序（ChIP-Seq）是一种强大的基因组学技术，可在全球范围内绘制ES细胞中组蛋白、转录因子和表观遗传修饰因子的翻译后修饰位置。在这里，我们描述了进行CARIP-Seq和ChIP-Seq的方法，包括用于下一代测序的文库构建，以生成ES细胞中的全局染色质相关RNA和表观基因组图谱。
