染色质成像及核体成像新技术。

IF 7.9 Q1 Medicine Wiley Interdisciplinary Reviews-Systems Biology and Medicine Pub Date : 2019-05-01 Epub Date: 2018-11-19 DOI:10.1002/wsbm.1442
Nicole A Szydlowski, Jane S Go, Ying S Hu
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引用次数: 6

摘要

先进荧光成像和标记技术的协同发展使染色质结构和动态在纳米级和活细胞的直接可视化成为可能。超分辨率成像包含了一类不断发展的技术,突破了荧光显微镜的衍射极限。结构照明显微镜提供了两倍的分辨率改进,可以很容易地实现使用传统的荧光团实时多色成像。单分子定位显微镜以较慢的采集速度为代价,将空间分辨率提高了大约10倍。受激发射损耗显微镜产生大约五倍的分辨率提高与成像速度成正比的扫描面积。同时,先进的标记策略已经发展到“点亮”全球和序列特异性DNA区域。DNA结合染料已被用于在单分子定位显微镜中实现高标记密度,并在相关光学和电子显微镜中增强对比度。新一代Oligopaint利用生物信息学分析来优化荧光原位杂交探针的设计。通过顺序和组合标记,DNA结构域的体积和长度以及不同拓扑相关结构域的空间组织的直接表征已被报道。在活细胞中,基因座特异性标记可以通过在目标基因旁边插入人工基因座来实现,例如阻遏因子-操作符阵列系统,或者利用基因组编辑工具,包括锌精细蛋白、转录激活因子样效应物和聚集规律间隔的短回文重复系统。结合单分子跟踪,这些标记技术可以直接可视化染色质内和染色质间的相互作用。本文分类如下:实验室方法与技术>成像。
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Chromatin imaging and new technologies for imaging the nucleome.

Synergistic developments in advanced fluorescent imaging and labeling techniques enable direct visualization of the chromatin structure and dynamics at the nanoscale level and in live cells. Super-resolution imaging encompasses a class of constantly evolving techniques that break the diffraction limit of fluorescence microscopy. Structured illumination microscopy provides a twofold resolution improvement and can readily achieve live multicolor imaging using conventional fluorophores. Single-molecule localization microscopy increases the spatial resolution by approximately 10-fold at the expense of slower acquisition speed. Stimulated emission-depletion microscopy generates a roughly fivefold resolution improvement with an imaging speed proportional to the scanning area. In parallel, advanced labeling strategies have been developed to "light up" global and sequence-specific DNA regions. DNA binding dyes have been exploited to achieve high labeling densities in single-molecule localization microscopy and enhance contrast in correlated light and electron microscopy. New-generation Oligopaint utilizes bioinformatics analyses to optimize the design of fluorescence in situ hybridization probes. Through sequential and combinatorial labeling, direct characterization of the DNA domain volume and length as well as the spatial organization of distinct topologically associated domains has been reported. In live cells, locus-specific labeling has been achieved by either inserting artificial loci next to the gene of interest, such as the repressor-operator array systems, or utilizing genome editing tools, including zinc finer proteins, transcription activator-like effectors, and the clustered regularly interspaced short palindromic repeats systems. Combined with single-molecule tracking, these labeling techniques enable direct visualization of intra- and inter-chromatin interactions. This article is categorized under: Laboratory Methods and Technologies > Imaging.

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来源期刊
CiteScore
18.40
自引率
0.00%
发文量
0
审稿时长
>12 weeks
期刊介绍: Journal Name:Wiley Interdisciplinary Reviews-Systems Biology and Medicine Focus: Strong interdisciplinary focus Serves as an encyclopedic reference for systems biology research Conceptual Framework: Systems biology asserts the study of organisms as hierarchical systems or networks Individual biological components interact in complex ways within these systems Article Coverage: Discusses biology, methods, and models Spans systems from a few molecules to whole species Topical Coverage: Developmental Biology Physiology Biological Mechanisms Models of Systems, Properties, and Processes Laboratory Methods and Technologies Translational, Genomic, and Systems Medicine
期刊最新文献
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