Microplate-based platform for combined chromatin and DNA methylation immunoprecipitation assays.
Microplate-based platform for combined chromatin and DNA methylation immunoprecipitation assays.
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DOI：
10.1186/1471-2199-12-49
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发表时间：
2011-11-18
影响因子：
--
通讯作者：
Bomsztyk K
中科院分区：
文献类型：
--
作者：
Yu J;Feng Q;Ruan Y;Komers R;Kiviat N;Bomsztyk K
组成特定基因表达的过程比以前认为的要复杂得多，提出了前所未有的概念和机制挑战，需要开发新的工具。染色质结构是基因调控的核心，受DNA甲基化和组蛋白修饰的调控。染色质的免疫沉淀（ChIP）和甲基化DNA （MeDIP）代表了该领域的主要成就，使研究人员能够探测染色质修饰以及体内特定蛋白质-DNA相互作用，并估计全基因组特定位点的蛋白质密度。虽然DNA甲基化是染色质结构的一个关键组成部分，但人们经常独立于其他染色质事件和转录来研究DNA甲基化。为了允许DNA甲基化与其他基因组过程同时测量，我们开发并验证了一个简单易用的高通量微孔板平台，用于分析DNA甲基化。与传统的基于珠状细胞的MeDIP相比，微孔板的MeDIP更敏感，非特异性结合更低。我们将MeDIP方法与微孔板ChIP分析相结合，可以同时测量DNA甲基化和组蛋白标记，Matrix ChIP-MeDIP平台。我们展示了该平台的几种应用，将DNA甲基化与培养细胞、人类癌症和糖尿病肾病模型中选定基因的染色质和转录事件联系起来。Matrix ChIP-MeDIP在DNA甲基化过程中同时分析数十甚至数百种不同基因组事件的高通量能力，为探索培养细胞和整个组织中选定基因的复杂基因组机制提供了强大的平台。在这方面，Matrix ChIP-MeDIP应该有助于补充全基因组研究，其中丰富的染色质和转录数据库资源为追求健康和疾病感兴趣的基因的机制，功能和诊断信息提供了富有成效的基础。
The processes that compose expression of a given gene are far more complex than previously thought presenting unprecedented conceptual and mechanistic challenges that require development of new tools. Chromatin structure, which is regulated by DNA methylation and histone modification, is at the center of gene regulation. Immunoprecipitations of chromatin (ChIP) and methylated DNA (MeDIP) represent a major achievement in this area that allow researchers to probe chromatin modifications as well as specific protein-DNA interactions in vivo and to estimate the density of proteins at specific sites genome-wide. Although a critical component of chromatin structure, DNA methylation has often been studied independently of other chromatin events and transcription. To allow simultaneous measurements of DNA methylation with other genomic processes, we developed and validated a simple and easy-to-use high throughput microplate-based platform for analysis of DNA methylation. Compared to the traditional beads-based MeDIP the microplate MeDIP was more sensitive and had lower non-specific binding. We integrated the MeDIP method with a microplate ChIP assay which allows measurements of both DNA methylation and histone marks at the same time, Matrix ChIP-MeDIP platform. We illustrated several applications of this platform to relate DNA methylation, with chromatin and transcription events at selected genes in cultured cells, human cancer and in a model of diabetic kidney disease. The high throughput capacity of Matrix ChIP-MeDIP to profile tens and potentially hundreds of different genomic events at the same time as DNA methylation represents a powerful platform to explore complex genomic mechanism at selected genes in cultured cells and in whole tissues. In this regard, Matrix ChIP-MeDIP should be useful to complement genome-wide studies where the rich chromatin and transcription database resources provide fruitful foundation to pursue mechanistic, functional and diagnostic information at genes of interest in health and disease.
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影响因子：
64.8
作者：
Ernst, Jason;Kheradpour, Pouya;Mikkelsen, Tarjei S.;Shoresh, Noam;Ward, Lucas D.;Epstein, Charles B.;Zhang, Xiaolan;Wang, Li;Issner, Robbyn;Coyne, Michael;Ku, Manching;Durham, Timothy;Kellis, Manolis;Bernstein, Bradley E.
通讯作者：
Bernstein, Bradley E.
影响因子：
3.9
作者：
Bennett, D
通讯作者：
Bennett, D
影响因子：
3.6
作者：
Feng, Qinghua;Stern, Joshua E.;Hawes, Stephen E.;Lu, Hiep;Jiang, Mingjun;Kiviat, Nancy B.
通讯作者：
Kiviat, Nancy B.
影响因子：
12.3
作者：
Choi JK;Bae JB;Lyu J;Kim TY;Kim YJ
通讯作者：
Kim YJ
影响因子：
4.8
作者：
Aker, Mari;Bomsztyk, Karol;Emery, David W.
通讯作者：
Emery, David W.