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Preparation of long single-strand DNA concatemers for high-level fluorescence in situ hybridization.


ABSTRACT: Fluorescence in situ hybridization (FISH) is a powerful tool to visualize transcripts in fixed cells and tissues. Despite the recent advances in FISH detection methods, it remains challenging to achieve high-level FISH imaging with a simple workflow. Here, we introduce a method to prepare long single-strand DNA concatemers (lssDNAc) through a controllable rolling-circle amplification (CRCA). Prepared lssDNAcs are used to develop AmpFISH workflows. In addition, we present its applications in different scenarios. AmpFISH shows the following advantages: 1) enhanced FISH signal-to-noise ratio (SNR) up to 160-fold compared with single-molecule FISH; 2) simultaneous detection of FISH signals and fluorescent proteins or immunofluorescence (IF) in tissues; 3) simple workflows; and 4) cost-efficiency. In brief, AmpFISH provides convenient and versatile tools for sensitive RNA/DNA detection and to gain useful information on cellular molecules using simple workflows.

SUBMITTER: Cao D 

PROVIDER: S-EPMC8545947 | biostudies-literature | 2021 Oct

REPOSITORIES: biostudies-literature

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Preparation of long single-strand DNA concatemers for high-level fluorescence in situ hybridization.

Cao Dongjian D   Wu Sa S   Xi Caili C   Li Dong D   Zhu Kaiheng K   Zhang Zhihong Z   Gong Hui H   Luo Qingming Q   Yang Jie J  

Communications biology 20211025 1


Fluorescence in situ hybridization (FISH) is a powerful tool to visualize transcripts in fixed cells and tissues. Despite the recent advances in FISH detection methods, it remains challenging to achieve high-level FISH imaging with a simple workflow. Here, we introduce a method to prepare long single-strand DNA concatemers (lssDNAc) through a controllable rolling-circle amplification (CRCA). Prepared lssDNAcs are used to develop AmpFISH workflows. In addition, we present its applications in diff  ...[more]

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