Automating ChIP-seq Experiments to Generate Epigenetic Profiles on 10,000 HeLa Cells
Automating ChIP-seq Experiments to Generate Epigenetic Profiles on 10,000 HeLa Cells
复制标题
DOI：
10.3791/52150
复制
发表时间：
2014-12-01
影响因子：
1.2
通讯作者：
Poncelet, Dominique
中科院分区：
文献类型：
--
作者：
Berguet, Geoffrey;Hendrickx, Jan;Poncelet, Dominique
染色质免疫沉淀，然后进行下一代测序（ChIP-seq）是研究蛋白质-DNA相互作用的首选技术。ChIP-seq已用于绘制蛋白质-DNA相互作用和分配组蛋白修饰。该过程是繁琐和耗时的，并且主要限制之一是需要大量的起始材料，通常是数百万个细胞。染色质免疫沉淀分析的自动化是可能的，当程序是基于使用磁珠。染色质免疫沉淀和文库制备的成功自动化方案已经在主要专用于自动化表观遗传测定的市售机器人液体处理系统上专门设计。首先，使用针对各种组蛋白修饰的抗体验证自动化ChIP-seq测定，然后优化自动化方案以进行染色质免疫沉淀和从低细胞数开始的文库制备。这些实验的目的是为未来特定细胞类型、亚群和活检样本的表观遗传学分析提供有价值的工具。
Chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq) is a technique of choice for studying protein-DNA interactions. ChIP-seq has been used for mapping protein-DNA interactions and allocating histones modifications. The procedure is tedious and time consuming, and one of the major limitations is the requirement for high amounts of starting material, usually millions of cells. Automation of chromatin immunoprecipitation assays is possible when the procedure is based on the use of magnetic beads. Successful automated protocols of chromatin immunoprecipitation and library preparation have been specifically designed on a commercially available robotic liquid handling system dedicated mainly to automate epigenetic assays. First, validation of automated ChIP-seq assays using antibodies directed against various histone modifications was shown, followed by optimization of the automated protocols to perform chromatin immunoprecipitation and library preparation starting with low cell numbers. The goal of these experiments is to provide a valuable tool for future epigenetic analysis of specific cell types, sub-populations, and biopsy samples.