An Integrated Platform for Genome-wide Mapping of Chromatin States Using High-throughput ChIP-sequencing in Tumor Tissues.
An Integrated Platform for Genome-wide Mapping of Chromatin States Using High-throughput ChIP-sequencing in Tumor Tissues.
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DOI：
10.3791/56972
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发表时间：
2018-04
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通讯作者：
C. Terranova;M. Tang;Elias Orouji;M. Maitituoheti;Ayush T. Raman;S. Amin;Zhiyi Liu;Kunal Rai
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作者：
C. Terranova;M. Tang;Elias Orouji;M. Maitituoheti;Ayush T. Raman;S. Amin;Zhiyi Liu;Kunal Rai
组蛋白修饰构成表观基因组的主要组成部分，在确定相关位点的转录状态中发挥重要的调节作用。此外，特定修饰的存在已被用来确定非编码功能元件（例如增强子）的位置和身份。近年来，染色质免疫沉淀和新一代测序 (ChIP-seq) 已成为确定单个组蛋白修饰的全基因组谱的强大工具。然而，越来越清楚的是，染色质修饰的组合模式（称为染色质状态）决定了相关基因组位点的身份和性质。因此，需要由强大的高通量 (HT) 方法组成的工作流程，用于分析大量组蛋白修饰标记，以及能够处理无数 ChIP-Seq 分析数据集的计算分析管道，以全面确定大量样品中的表观基因组状态。这里介绍的 HT-ChIP-Seq 工作流程由两个模块组成：1) 一个实验方案，用于分析 96 孔格式的少量肿瘤样本和细胞系中的几种组蛋白修饰； 2) 计算数据分析管道，结合现有工具来计算单个标记占用率和组合染色质状态模式。这两个模块共同促进了快速高效地处理数百个 ChIP-Seq 样本。这里介绍的工作流程用于从黑色素瘤肿瘤和细胞系的 6 个组蛋白标记谱中得出染色质状态模式。总的来说，我们提出了一个全面的 ChIP-seq 工作流程，可应用于数十种人类肿瘤样本和癌细胞系，以确定各种恶性肿瘤中的表观基因组畸变。
Histone modifications constitute a major component of the epigenome and play important regulatory roles in determining the transcriptional status of associated loci. In addition, the presence of specific modifications has been used to determine the position and identity non-coding functional elements such as enhancers. In recent years, chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq) has become a powerful tool in determining the genome-wide profiles of individual histone modifications. However, it has become increasingly clear that the combinatorial patterns of chromatin modifications, referred to as Chromatin States, determine the identity and nature of the associated genomic locus. Therefore, workflows consisting of robust high-throughput (HT) methodologies for profiling a number of histone modification marks, as well as computational analyses pipelines capable of handling myriads of ChIP-Seq profiling datasets, are needed for comprehensive determination of epigenomic states in large number of samples. The HT-ChIP-Seq workflow presented here consists of two modules: 1) an experimental protocol for profiling several histone modifications from small amounts of tumor samples and cell lines in a 96-well format; and 2) a computational data analysis pipeline that combines existing tools to compute both individual mark occupancy and combinatorial chromatin state patterns. Together, these two modules facilitate easy processing of hundreds of ChIP-Seq samples in a fast and efficient manner. The workflow presented here is used to derive chromatin state patterns from 6 histone mark profiles in melanoma tumors and cell lines. Overall, we present a comprehensive ChIP-seq workflow that can be applied to dozens of human tumor samples and cancer cell lines to determine epigenomic aberrations in various malignancies.