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一种快速微量染色质免疫沉淀分析(微量ChIP)。

A rapid micro chromatin immunoprecipitation assay (microChIP).

作者信息

Dahl John Arne, Collas Philippe

机构信息

Department of Biochemistry, Faculty of Medicine, Institute of Basic Medical Sciences, University of Oslo, Oslo 0317, Norway.

出版信息

Nat Protoc. 2008;3(6):1032-45. doi: 10.1038/nprot.2008.68.

Abstract

Interactions of proteins with DNA mediate many critical nuclear functions. Chromatin immunoprecipitation (ChIP) is a robust technique for studying protein-DNA interactions. Current ChIP assays, however, either require large cell numbers, which prevent their application to rare cell samples or small-tissue biopsies, or involve lengthy procedures. We describe here a 1-day micro ChIP (microChIP) protocol suitable for up to eight parallel histone and/or transcription factor immunoprecipitations from a single batch of 1,000 cells. MicroChIP technique is also suitable for monitoring the association of one protein with multiple genomic sites in 100 cells. Alterations in cross-linking and chromatin preparation steps also make microChIP applicable to approximately 1-mm(3) fresh- or frozen-tissue biopsies. From cell fixation to PCR-ready DNA, the procedure takes approximately 8 h for 16 ChIPs.

摘要

蛋白质与DNA的相互作用介导了许多关键的核功能。染色质免疫沉淀(ChIP)是研究蛋白质-DNA相互作用的一项可靠技术。然而,目前的ChIP分析方法要么需要大量细胞,这使得它们无法应用于稀有细胞样本或小组织活检,要么涉及冗长的程序。我们在此描述一种1天的微量ChIP(microChIP)方案,适用于从一批1000个细胞中进行多达8个平行的组蛋白和/或转录因子免疫沉淀。MicroChIP技术也适用于监测100个细胞中一种蛋白质与多个基因组位点的关联。交联和染色质制备步骤的改变也使microChIP适用于约1立方毫米的新鲜或冷冻组织活检。从细胞固定到可用于PCR的DNA,该程序对16次ChIP而言大约需要8小时。

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