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Cellular Detection of G-Quadruplexes by UNIT 4.33Optical Imaging Methods

机译:单位4.33光学成像方法G-Quadruplees的细胞检测

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摘要

G-quadruplexes (G4s) are higher-order nucleic acid structures that fold fromguanine (G)–rich DNA and RNA strands. This field of research gains tractionas a major chemical biology area since it aims at uncovering many key cellularmechanisms in which quadruplexes are involved. The wealth of knowledgeacquired over the past three decades strongly supports pivotal roles of G4 inthe regulation of gene expression at both transcriptional (DNA quadruplexes)and translational levels (RNA quadruplexes). Recent biochemical discoveriesuncovered myriad of additional G4 actions: from chromosomal stability to thefiring of replication origins, from telomere homeostasis to functional dysregulationsunderlying genetic diseases (including cancers and neurodegeneration).Here, we listed a repertoire of protocols that we have developed over thepast years to visualize quadruplexes in cells. These achievements were madepossible thanks to the discovery of a novel family of versatile quadruplexselectivefluorophores, the twice-as-smart quadruplex ligands named TASQ(for template-assembled synthetic G-quartet). The versatility of this probe allowsfor multiple imaging techniques in both fixed and live cells, includingthe use of the multiphoton microscopy, confocal microscopy, and real-timefluorescent image collection.
机译:G-四(G4S)是高阶核酸结构,其折叠fromguanine(G)丰富的DNA和RNA链。研究涨势此字段tractionas的主要化学生物学领域,因为它的目的是揭示在四链涉及许多关键cellularmechanisms。的knowledgeacquired过去三个十年的财富强烈支持在转录(四链DNA)基因表达的G4在矿井调控和翻译水平(四链RNA)的关键作用。最近的生化discoveriesuncovered无数的额外G4操作:从染色体的稳定性thefiring复制起点的,从端粒动态平衡到功能dysregulationsunderlying遗传性疾病(包括癌症和神经退行性疾病)。在这里,我们列出的协议的一个剧目,我们已经在thepast年来可视化开发四链的细胞。这些成绩的取得是madepossible得益于新家族的多功能quadruplexselectivefluorophores,命名TASQ的两次作为智能四重配体(模板组装合成G-四)的发现。这个探测器的allowsfor固定和活细胞多种成像技术的通用性,includingthe利用多光子显微镜,激光共聚焦显微镜和实时timefluorescent图像采集的。

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