Gaykalova Daria A, Kulaeva Olga I, Pestov Nikolai A, Hsieh Fu-Kai, Studitsky Vasily M
Department of Pharmacology, UMDNJ-Robert Wood Johnson Medical School, Piscataway, New Jersey, USA.
Methods Enzymol. 2012;512:293-314. doi: 10.1016/B978-0-12-391940-3.00013-5.
The vital process of transcription by RNA polymerase II (Pol II) occurs in chromatin environment in eukaryotic cells; in fact, moderately transcribed genes retain nucleosomal structure. Recent studies suggest that chromatin structure presents a strong barrier for transcribing Pol II in vitro, and that DNA-histone interactions are only partially and transiently disrupted during transcript elongation on moderately active genes. Furthermore, elongating Pol II complex is one of the major targets during gene regulation. Below, we describe a highly purified, defined experimental system that recapitulates many important properties of transcribed chromatin in vitro and allows detailed analysis of the underlying mechanisms.
RNA聚合酶II（Pol II）进行的关键转录过程发生在真核细胞的染色质环境中；事实上，中等转录水平的基因保留着核小体结构。最近的研究表明，染色质结构在体外对Pol II转录构成了强大障碍，并且在中等活性基因的转录延伸过程中，DNA-组蛋白相互作用只是部分且短暂地被破坏。此外，延伸中的Pol II复合物是基因调控过程中的主要靶点之一。下面，我们描述了一个高度纯化、明确的实验系统，该系统在体外重现了转录染色质的许多重要特性，并允许对潜在机制进行详细分析。
