{"title":"Single-Assay Profiling of Nucleosome Occupancy and Chromatin Accessibility.","authors":"April Cook, Jakub Mieczkowski, Michael Y Tolstorukov","doi":"10.1002/cpmb.45","DOIUrl":null,"url":null,"abstract":"<p><p>This unit describes a method for determining the accessibility of chromatinized DNA and nucleosome occupancy in the same assay. Enzymatic digestion of chromatin using micrococcal nuclease (MNase) is optimized for liberation, retrieval, and characterization of DNA fragments from chromatin. MNase digestion is performed in a titration series, and the DNA fragments are isolated and sequenced for each individual digest independently. These sequenced fragments are then collectively analyzed in a novel bioinformatics pipeline to produce a metric describing MNase accessibility of chromatin (MACC) and nucleosome occupancy. This approach allows profiling of the entire genome including regions of open and closed chromatin. Moreover, the MACC protocol can be supplemented with a histone immunoprecipitation step to estimate and compare both histone and non-histone DNA protection components. © 2017 by John Wiley & Sons, Inc.</p>","PeriodicalId":10734,"journal":{"name":"Current Protocols in Molecular Biology","volume":"120 ","pages":"21.34.1-21.34.18"},"PeriodicalIF":0.0000,"publicationDate":"2017-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmb.45","citationCount":"7","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Current Protocols in Molecular Biology","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1002/cpmb.45","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"Biochemistry, Genetics and Molecular Biology","Score":null,"Total":0}
引用次数: 7
Abstract
This unit describes a method for determining the accessibility of chromatinized DNA and nucleosome occupancy in the same assay. Enzymatic digestion of chromatin using micrococcal nuclease (MNase) is optimized for liberation, retrieval, and characterization of DNA fragments from chromatin. MNase digestion is performed in a titration series, and the DNA fragments are isolated and sequenced for each individual digest independently. These sequenced fragments are then collectively analyzed in a novel bioinformatics pipeline to produce a metric describing MNase accessibility of chromatin (MACC) and nucleosome occupancy. This approach allows profiling of the entire genome including regions of open and closed chromatin. Moreover, the MACC protocol can be supplemented with a histone immunoprecipitation step to estimate and compare both histone and non-histone DNA protection components. © 2017 by John Wiley & Sons, Inc.
核小体占用和染色质可及性的单分析分析。
本单元描述了在同一试验中确定染色化DNA和核小体占用的可及性的方法。利用微球菌核酸酶(MNase)对染色质进行酶切，用于从染色质中解放，检索和表征DNA片段。酶消化是在滴定系列中进行的，DNA片段被分离出来，并为每个单独的消化独立测序。这些测序片段然后在新的生物信息学管道中进行集体分析，以产生描述染色质(MACC)和核小体占用的MNase可及性的度量。这种方法可以分析整个基因组，包括开放和封闭染色质区域。此外，MACC方案可以补充组蛋白免疫沉淀步骤，以估计和比较组蛋白和非组蛋白DNA保护成分。©2017 by John Wiley & Sons, Inc。
本文章由计算机程序翻译，如有差异，请以英文原文为准。
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