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An Improved Restriction Enzyme Accessibility Assay for Analyzing Changes in Chromatin Structure in Samples of Limited Cell Number

机译：一种改进的限制酶可及性分析用于分析有限细胞数样品中染色质结构的变化

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Studies investigating mechanisms that control gene regulation frequently examine the accessibility of specific DNA sequences to nuclease cleavage. In general, sequences that are sensitive to nuclease cleavage are considered to be in an \u22open\u22 chromatin conformation that is associated with regulatory factor binding, while sequences resistant to nuclease cleavage are considered to be in a \u22closed\u22 conformation commonly associated with chromatin that is neither poised for transcription nor being actively transcribed. Changes in nuclease accessibility at specific genomic sequences reflect changes in the local chromatin structure that can occur as a result of signaling cues in the extracellular environment. These changes in chromatin structure usually precede or are coincident with changes in gene expression patterns and are therefore a useful marker of regulatory events controlling transcription. We describe a method to perform restriction enzyme accessibility assays (REAA) that utilizes ligation-mediated polymerase chain reaction (LM-PCR) technology and that permits assessment of samples from any source containing as few as 1,000 cells. Use of this modified REAA protocol will enhance analysis of chromatin structural changes at specific DNA sequences of interest by making it possible to analyze samples where unrestricted amounts of sample are not readily available.
机译：研究控制基因调控的机制的研究经常检查特定DNA序列对核酸酶切割的可及性。通常，对核酸酶切割敏感的序列被认为与调控因子结合相关的染色质构象，而对核酸酶切割抵抗的序列被认为通常与相关的核酸闭合构象。既不准备转录也不活跃转录的染色质。特定基因组序列上核酸酶可及性的变化反映了局部染色质结构的变化，这种变化可能是由于细胞外环境中信号提示的结果而发生的。染色质结构的这些改变通常在基因表达模式改变之前或与之同时发生，因此是控制转录的调控事件的有用标记。我们描述了一种利用连接介导的聚合酶链反应（LM-PCR）技术进行限制酶可及性分析（REAA）的方法，该方法可以评估来自任何包含1,000个细胞的来源的样品。使用这种改良的REAA方案将可以分析不受限数量样品的样品，从而增强了对特定目标DNA序列染色质结构变化的分析。

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