Rapid Visual CRISPR Assay: A Naked-Eye Colorimetric Detection Method for Nucleic Acids Based on CRISPR/Cas12a and a Convolutional Neural Network

文献类型： 外文期刊

第一作者： Xie, Shengsong

作者： Xie, Shengsong;Tao, Dagang;Fu, Yuhua;Xu, Bingrong;Pan, Wenya;Huang, Xin;Nie, Xiongwei;Zhao, Changzhi;Ruan, Jinxue;Zhang, Yi;Fu, Liangliang;Ma, Yunlong;Li, Xinyun;Liu, Xiaolei;Zhao, Shuhong;Xie, Shengsong;Tao, Dagang;Fu, Yuhua;Xu, Bingrong;Pan, Wenya;Huang, Xin;Nie, Xiongwei;Zhao, Changzhi;Ruan, Jinxue;Zhang, Yi;Fu, Liangliang;Ma, Yunlong;Li, Xinyun;Liu, Xiaolei;Zhao, Shuhong;Xie, Shengsong;Steinaa, Lucilla;Hemmink, Johanneke D.;Xie, Shengsong;Li, Xinyun;Liu, Xiaolei;Zhao, Shuhong;Tang, You;Han, Jianlin;Han, Jianlin;Li, Xinyun;Liu, Xiaolei;Zhao, Shuhong

作者机构：

关键词： nucleic acid detection; CRISPR-Cas12a assay; convolutional neural network; MagicEye software

期刊名称：ACS SYNTHETIC BIOLOGY （ 影响因子：5.249； 五年影响因子：5.373 ）

ISSN： 2161-5063

年卷期： 2022 年 11 卷 1 期

页码：

收录情况： SCI

摘要： Rapid diagnosis based on naked-eye colorimetric detection remains challenging, but it could build new capacities for molecular point-of-care testing (POCT). In this study, we evaluated the performance of 16 types of single-stranded DNA-fluorophore-quencher (ssDNA-FQ) reporters for use with clusters of regularly spaced short palindrome repeats (CRISPR)/Cas12a-based visual colorimetric assays. Among them, nine ssDNA-FQ reporters were found to be suitable for direct visual colorimetric detection, with especially very strong performance using ROX-labeled reporters. We optimized the reaction concentrations of these ssDNA-FQ reporters for a naked-eye read-out of assay results (no transducing component required for visualization). In particular, we developed a convolutional neural network algorithm to standardize and automate the analytical colorimetric assessment of images and integrated this into the MagicEye mobile phone software. A field-deployable assay platform named RApid VIsual CRISPR (RAVI-CRISPR) based on a ROX-labeled reporter with isothermal amplification and CRISPR/Cas12a targeting was established. We deployed RAVI-CRISPR in a single tube toward an instrument-less colorimetric POCT format that required only a portable rechargeable hand warmer for incubation. The RAVI-CRISPR was successfully used for the high-sensitivity detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and African swine fever virus (ASFV). Our study demonstrates this RAVI-CRISPR/MagicEye system to be suitable for distinguishing different pathogenic nucleic acid targets with high specificity and sensitivity as the simplest-to-date platform for rapid pen- or bed-side testing.

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