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Paper Device Combining CRISPR/Cas12a and Reverse-Transcription Loop-Mediated Isothermal Amplification for SARS-CoV-2 Detection in Wastewater
Haorui Cao; Kang Mao; Fang Ran; Pengqi Xu; Yirong Zhao; Xiangyan Zhang; Hourong Zhou; Zhugen Yang; Hua Zhang; Guibin Jiang
2022
Source PublicationEnvironmental Science & Technology
Abstract

Wastewater-based surveillance of the COVID-19 pandemic holds great promise; however, a point-of-use detection method for SARS-CoV-2 in wastewater is lacking. Here, a portable paper device based on CRISPR/Cas12a and reverse-transcription loop-mediated isothermal amplification (RT-LAMP) with excellent sensitivity and specificity was developed for SARS-CoV-2 detection in wastewater. Three primer sets of RT-LAMP and guide RNAs (gRNAs) that could lead Cas12a to recognize target genes via base pairing were used to perform the high-fidelity RT-LAMP to detect the N, E, and S genes of SARS-CoV-2. Due to the trans-cleavage activity of CRISPR/Cas12a after high-fidelity amplicon recognition, carboxyfluorescein-ssDNA-Black Hole Quencher-1 and carboxyfluorescein-ssDNA-biotin probes were adopted to realize different visualization pathways via a fluorescence or lateral flow analysis, respectively. The reactions were integrated into a paper device for simultaneously detecting the N, E, and S genes with limits of detection (LODs) of 25, 310, and 10 copies/mL, respectively. The device achieved a semiquantitative analysis from 0 to 310 copies/mL due to the different LODs of the three genes. Blind experiments demonstrated that the device was suitable for wastewater analysis with 97.7% sensitivity and 82% semiquantitative accuracy. This is the first semiquantitative endpoint detection of SARS-CoV-2 in wastewater via different LODs, demonstrating a promising point-of-use method for wastewater-based surveillance.

KeywordSars-cov-2 Crispr/cas12a Rt-lamp Paper Device Wastewater
DOI10.1021/acs.est.2c04727
URL查看原文
Language英语
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Cited Times:50[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.gyig.ac.cn/handle/42920512-1/13582
Collection环境地球化学国家重点实验室
Affiliation1.State Key Laboratory of Environmental Geochemistry, Institute of Geochemistry, Chinese Academy of Sciences, Guiyang 550081, China
2.School of Water, Energy, and Environment,Cranfield University, Cranfield MK43 0AL, UK
3.University of Chines Academy of Sciences, Beijing 100049, China
4.Precision Medicine Center, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen 518107, China
5.Guizhou Provincial People’s Hospital,Guiyang 550002, China
6.Jiangjunshan Hospital of Guizhou Province, Guiyang 550001, China
7.State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China
Recommended Citation
GB/T 7714
Haorui Cao,Kang Mao,Fang Ran,et al. Paper Device Combining CRISPR/Cas12a and Reverse-Transcription Loop-Mediated Isothermal Amplification for SARS-CoV-2 Detection in Wastewater[J]. Environmental Science & Technology,2022.
APA Haorui Cao.,Kang Mao.,Fang Ran.,Pengqi Xu.,Yirong Zhao.,...&Guibin Jiang.(2022).Paper Device Combining CRISPR/Cas12a and Reverse-Transcription Loop-Mediated Isothermal Amplification for SARS-CoV-2 Detection in Wastewater.Environmental Science & Technology.
MLA Haorui Cao,et al."Paper Device Combining CRISPR/Cas12a and Reverse-Transcription Loop-Mediated Isothermal Amplification for SARS-CoV-2 Detection in Wastewater".Environmental Science & Technology (2022).
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