Biochemical techniques for the characterization of G-quadruplex structures: EMSA, DMS footprinting, and DNA polymerase stop assay.
Biochemical techniques for the characterization of G-quadruplex structures: EMSA, DMS footprinting, and DNA polymerase stop assay.
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G-四链体结构表征的生化技术：EMSA，DMS足迹和DNA聚合酶停止测定。
DOI：
10.1007/978-1-59745-363-9_5
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发表时间：
2010
期刊：
影响因子：
--
通讯作者：
Hurley, Laurence H
中科院分区：
文献类型：
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作者：
Sun, Daekyu;Hurley, Laurence H
许多人类生长相关基因的近端启动子区域包含一个多嘌呤/多嘧啶通道，作为Sp1或其他转录因子的多重结合位点。这些小束通常含有富含鸟嘌呤的序列，由三个或更多连续的鸟嘌呤组成，由一个或多个碱基分开，对应于已知的分子内g四重体形成的一般基序。最近的研究结果提供了强有力的证据，表明特定的g -四重体结构可以由许多人类启动子区域的这些多嘌呤/多嘧啶束中的富含g的序列自然形成，这提高了这些基因的转录控制可以通过g -四重体相互作用剂调节的可能性。在本章中，我们描述了三种通用的生化方法，电泳迁移率转移测定（EMSA），二甲基硫酸盐（DMS）足迹和DNA聚合酶停止测定，这可以用于g -富g序列形成的g -四重体结构的初始表征。
The proximal promoter region of many human growth-related genes contains a polypurine/polypyrimidine tract that serves as a multiple binding site for Sp1 or other transcription factors. These tracts often contain a guanine-rich sequence consisting of four runs of three or more contiguous guanines separated by one or more bases, corresponding to a general motif known for the formation of an intramolecular G-quadruplex. Recent results provide strong evidence that specific G-quadruplex structures can be formed naturally by the G-rich sequence within these polypurine/polypyrimidine tracts of many human promoter regions, raising the possibility that the transcriptional control of these genes can be modulated by G-quadruplex-interactive agents. In this chapter, we describe three general biochemical methodologies, electrophoretic mobility shift assay (EMSA), dimethylsulfate (DMS) footprinting, and the DNA polymerase stop assay, which can be useful for initial characterization of G-quadruplex structures formed by G-rich sequences.