Purification of Specific Chromatin Domains from Single-Copy Gene Loci in Saccharomyces cerevisiae
Purification of Specific Chromatin Domains from Single-Copy Gene Loci in Saccharomyces cerevisiae
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DOI：
10.1007/978-1-62703-706-8_26
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发表时间：
2014-01-01
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影响因子：
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通讯作者：
Griesenbeck, Joachim
中科院分区：
文献类型：
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作者：
Hamperl, Stephan;Brown, Christopher R.;Griesenbeck, Joachim
目前可用于体内染色质分析的大多数方法都依赖于假定染色质成分或其翻译后修饰状态的先验知识。确定的天然染色体区域的分离提供了一种有吸引力的替代方案，以获得染色质的基本无偏见的分子描述。在这里，我们描述了一种将染色体上的位点特异性重组与高效串联亲和纯化方案相结合的策略，以从酿酒酵母中分离出单拷贝基因座。该方法可以对目标染色质结构域进行稳健富集，使其适合成分、结构和生化分析。该技术似乎适合于获得酵母中几乎任何基因组位点的染色质组成和特定翻译后组蛋白修饰状态的详细描述。
Most methods currently available for the analysis of chromatin in vivo rely on a priori knowledge of putative chromatin components or their posttranslational modification state. The isolation of defined native chromosomal regions provides an attractive alternative to obtain a largely unbiased molecular description of chromatin. Here, we describe a strategy combining site-specific recombination at the chromosome with an efficient tandem affinity purification protocol to isolate a single-copy gene locus from the yeast Saccharomyces cerevisiae. The method allows robust enrichment of a targeted chromatin domain, making it amenable to compositional, structural, and biochemical analyses. This technique appears to be suitable to obtain a detailed description of chromatin composition and specific posttranslational histone modification state at virtually any genomic locus in yeast.