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Updated: Jun 14, 2025

Robust 3D DNA FISH Using Directly Labeled Probes

Robust 3D DNA FISH Using Directly Labeled Probes

Published on: August 15, 2013

在单个细胞中可视化表观遗传修饰及其空间近距离,使用三个DNA编码的放大FISH成像策略:BEA-FISH,PPDA-FISH和细胞交谈

Feng Chen¹, Xinyin Li¹, Min Bai¹

¹Institute of Analytical Chemistry and Instrument for Life Science, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'An, P. R. China.

Nature protocols

|September 4, 2024

概括

这项研究引入了三种新的DNA编码放大FISH成像策略,以检测低丰度表观遗传修饰及其在细胞中的空间近距离. 这些方法克服了当前技术的局限性,使分子相互作用的精确可视化和分析成为可能.

科学领域:

分子生物学分子生物学
表观遗传学在表观遗传学中,表观遗传学是指表观遗传学.
细胞生物学细胞生物学

背景情况:

表观遗传修饰和分子接近对于细胞功能和疾病至关重要.
如FISH和免疫光等现有的成像技术在检测低丰度修饰及其空间关系方面存在局限性.

研究的目的:

开发先进的现场成像策略,以检测低丰度表观遗传修饰及其空间近距离.
克服当前细胞成像方法在可视化微妙分子相互作用方面的局限性.

主要方法:

开发了三种基于FISH的DNA编码放大成像策略:BEA-FISH,PPDA-FISH和细胞交谈.
利用DNA编码放大来将分子特征转换为DNA条形码,用于滚动圆的放大和FISH分析.
关键反应包括目标标记,DNA编码和固定细胞中的滚动圆放大成像.

主要成果:

在现场计数低丰度修饰物 (例如,5-基甲基细胞因子,5-甲基细胞因子).
启用对对近距离差异化可视化两个修饰和蛋白质周围的一对多近距离分析.
在单细胞中成功可视化了核酸和基因素修饰的组合模式.

结论:

开发的成像策略为研究表观遗传修饰和分子近距离提供了更高的灵敏度和特异性.
这些方法为探索在生理和病理背景下复杂的分子相互作用提供了新的工具.
该协议可适应各种应用,可以在2-4天内完成.

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3D Multicolor DNA FISH Tool to Study Nuclear Architecture in Human Primary Cells

3D Multicolor DNA FISH Tool to Study Nuclear Architecture in Human Primary Cells

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Combined Immunofluorescence and DNA FISH on 3D-preserved Interphase Nuclei to Study Changes in 3D Nuclear Organization

Combined Immunofluorescence and DNA FISH on 3D-preserved Interphase Nuclei to Study Changes in 3D Nuclear Organization

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相关概念视频

FISH - Fluorescent In-situ Hybridization

FISH - Fluorescent In-situ Hybridization

Fluorescence in situ hybridization, or FISH, was developed in the early 1980s and has quickly become one of the most widely used techniques in cytogenetics. Labeled probes are used to bind complementary DNA or RNA sequences on a chromosome or in a region within a cell. Earlier, the probes could only be obtained by cloning or reverse transcription of a DNA template. Currently, the probe oligonucleotides can be synthesized synthetically. Additionally, with the advancement of optical techniques,...