key: cord-0869605-tijsy8ws
authors: Wang, Yuxi; Zhang, Yong; Chen, Junbo; Wang, Minjin; Zhang, Ting; Luo, Wenxin; Li, Yalun; Wu, Yangping; Zeng, Bo; Zhang, Kaixiang; Deng, Ruijie; Li, Weimin
title: Detection of SARS-CoV-2 and Its Mutated Variants via CRISPR-Cas13-Based Transcription Amplification
date: 2021-01-29
journal: Anal Chem
DOI: 10.1021/acs.analchem.0c04303
sha: bbf7e96d00c9d7aeff24b28acfaa10328bf01f0e
doc_id: 869605
cord_uid: tijsy8ws

[Image: see text] The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global health emergency, and its gene mutation and evolution further posed uncertainty of epidemic risk. Herein, we reported a light-up CRISPR-Cas13 transcription amplification method, which enables the detection of SARS-CoV-2 and its mutated variants. Sequence specificity was ensured by both the ligation process and Cas13a/crRNA recognition, allowing us to identify viral RNA mutation. Light-up RNA aptamer allows sensitive output of amplification signals via target-activated ribonuclease activity of CRISPR-Cas13a. The RNA virus assay has been designed to detect coronavirus, SARS-CoV-2, Middle East respiratory syndrome (MERS), and SARS, as well as the influenza viruses such as, H1N1, H7N9, and H9N2. It was accommodated to sense as low as 82 copies of SARS-CoV-2. Particularly, it allowed us to strictly discriminate key mutation of the SARS-CoV-2 variant, D614G, which may induce higher epidemic and pathogenetic risk. The proposed RNA virus assays are promising for point-of-care monitoring of SARS-CoV-2 and its risking variants.

. Oligonucleotide sequences S3 Table S2 . Comparations of detection performances of different assays for the detection of SARS-CoV-2 S8 Figure S1 . Gene sequences of SARS-CoV-2 (wild), SARS-CoV-2 (D614G mutated), SARS and MERS pseudovrius S9 Figure S2 . Plasmid vector for constructing SARS-CoV-2 (wild), SARS-CoV-2 (D614G mutated), SARS and MERS pseudovrius S12 Figure S3 . Working flow to prepare SARS-CoV-2 contaminated food package and frozen belt fish samples S13 Figure S4 . Detection of coronavirus species including MERS, SARS and SARS-CoV-2 by targeting E genes S14 Figure S5 . Fluorescence response towards wild and mutated SARS-CoV-2 using Pre-substrate probes targeting different detection locus S15 Figure S6 . Identify D614G mutation of SARS-CoV-2 using different crRNA sequences S16 Figure S7 . Discrimination rates for detecting D614G mutation via ligation-based recognition or the integration of ligation and Cas13/crRNA recognition using Pre-substrate probes with different recognition sequences S17 Figure S8 . Optimization of the ratio of Cas13a to crRNA S18 

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