CRISPR/Cas12a-based biosensing platform for the on-site detection of single-base mutants in gene-edited rice

文献类型： 外文期刊

第一作者： Wang, Mengyu

作者： Wang, Mengyu;Liu, Xiaojing;Yang, Jiangtao;Wang, Zhixing;Wang, Xujing;Wang, Haoqian

作者机构：

关键词： single-base mutant detection; fast visual detection; on-site detection; CRISPR; Cas12a; recombinase polymerase amplification

期刊名称：FRONTIERS IN PLANT SCIENCE （ 影响因子：6.627； 五年影响因子：7.255 ）

ISSN： 1664-462X

年卷期： 2022 年 13 卷

页码：

收录情况： SCI

摘要： At present, with the accelerated development of the global biotechnology industry, novel transgenic technologies represented by gene editing are developing rapidly. A large number of gene-edited products featuring one or a few base indels have been commercialized. These have led to great challenges in the use of traditional nucleic acid detection technology and in safety regulation for genetically modified organisms (GMOs). In this study, we developed a portable clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins 12a-based (CRISPR/Cas12a-based) biosensing platform named Cas12aFVD (fast visual detection) that can be coupled with recombinase polymerase amplification (RPA) for on-site detection of mutants in gene-edited rice in one tube. The detection procedure can be accomplished in 40 min with a visible result, which can be observed by the naked eye under blue light (470-490 nm). By accurate recognition of targets based on Cas12a/CRISPR RNA (crRNA), Cas12aFVD exhibits excellent performance for the detection of two- and three-base deletions, one-base substitution, and one-base insertion mutants with a limit of detection (LOD) of 12 copies/mu l showing great potential for mutant detection, especially single-base mutants. The Cas12aFVD biosensing platform is independent of laboratory conditions, making it a promising and pioneering platform for the detection of gene-edited products.

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