Detection of infectious viruses using crispr-cas12-based assay

Chandana S. Talwar; Kwang Hyun Park; Woo Chan Ahn; Yong Sam Kim; Oh Seok Kwon; Dongeun Yong; Taejoon Kang; Euijeon Woo

doi:10.3390/bios11090301

Detection of infectious viruses using crispr-cas12-based assay

Chandana S. Talwar, Kwang Hyun Park, Woo Chan Ahn, Yong Sam Kim, Oh Seok Kwon, Dongeun Yong, Taejoon Kang, Euijeon Woo

Department of Laboratory Medicine

Research output: Contribution to journal › Article › peer-review

17 Citations (Scopus)

Abstract

The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease-19 (COVID-19), has severely influenced public health and economics. For the detection of SARS-CoV-2, clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated protein (Cas)-based assays have been emerged because of their simplicity, sensitivity, specificity, and wide applicability. Herein, we have developed a CRISPR-Cas12-based assay for the detection of SARS-CoV-2. In the assay, the target amplicons are produced by isothermal reverse transcription recombinase polymerase amplification (RT-RPA) and recognized by a CRISPR-Cas12a/guide RNA (gRNA) complex that is coupled with the collateral cleavage activity of fluorophore-tagged probes, allowing either a fluorescent measurement or naked-eye detection on a lateral flow paper strip. This assay enables the sensitive detection of SARS-CoV-2 at a low concentration of 10 copies per sample. Moreover, the reliability of the method is verified by using nasal swabs and sputum of COVID-19 patients. We also proved that the current assay can be applied to other viruses, such as Middle East respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus (SARS-CoV), with no major changes to the basic scheme of testing. It is anticipated that the CRISPR-Cas12-based assay has the potential to serve as a point-of-care testing (POCT) tool for a wide range of infectious viruses.

Original language	English
Article number	301
Journal	Biosensors
Volume	11
Issue number	9
DOIs	https://doi.org/10.3390/bios11090301
Publication status	Published - 2021 Sept

Bibliographical note

Funding Information:
Funding: This research was supported by National R&D Programs through National Research Foundation (NRF) of Korea funded by Ministry of Science and ICT (MSIT) of Korea (NRF-2021R1A2C2011328, NRF-2021M3A9G8025599, NRF-2019R1C1C1006867, NRF-2021M3H4A1A02051048, NRF-2021M3E5E3080379, and NRF-2018M3A9E2022821), Global Frontier Program through Center for BioNano Health-Guard funded by MSIT of Korea (H-GUARD_2013M3A6B2078950), Technology Development Program for Biological Hazards Management in Indoor Air through Korea Environment Industry & Technology Institute (KEITI) funded by Ministry of Environment (ME) of Korea (2021003370003), Nanomedical Devices Development Program of NNFC (CSM2105M101), and KRIBB Research Initiative Program (KGS-1052113 and 1711134081).

Publisher Copyright:
© 2021 by the authors. Licensee MDPI, Basel, Switzerland.

All Science Journal Classification (ASJC) codes

Analytical Chemistry
Biotechnology
Biomedical Engineering
Instrumentation
Engineering (miscellaneous)
Clinical Biochemistry

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This output contributes to the following UN Sustainable Development Goals (SDGs)

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title = "Detection of infectious viruses using crispr-cas12-based assay",

abstract = "The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease-19 (COVID-19), has severely influenced public health and economics. For the detection of SARS-CoV-2, clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated protein (Cas)-based assays have been emerged because of their simplicity, sensitivity, specificity, and wide applicability. Herein, we have developed a CRISPR-Cas12-based assay for the detection of SARS-CoV-2. In the assay, the target amplicons are produced by isothermal reverse transcription recombinase polymerase amplification (RT-RPA) and recognized by a CRISPR-Cas12a/guide RNA (gRNA) complex that is coupled with the collateral cleavage activity of fluorophore-tagged probes, allowing either a fluorescent measurement or naked-eye detection on a lateral flow paper strip. This assay enables the sensitive detection of SARS-CoV-2 at a low concentration of 10 copies per sample. Moreover, the reliability of the method is verified by using nasal swabs and sputum of COVID-19 patients. We also proved that the current assay can be applied to other viruses, such as Middle East respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus (SARS-CoV), with no major changes to the basic scheme of testing. It is anticipated that the CRISPR-Cas12-based assay has the potential to serve as a point-of-care testing (POCT) tool for a wide range of infectious viruses.",

author = "Talwar, {Chandana S.} and Park, {Kwang Hyun} and Ahn, {Woo Chan} and Kim, {Yong Sam} and Kwon, {Oh Seok} and Dongeun Yong and Taejoon Kang and Euijeon Woo",

note = "Funding Information: Funding: This research was supported by National R&D Programs through National Research Foundation (NRF) of Korea funded by Ministry of Science and ICT (MSIT) of Korea (NRF-2021R1A2C2011328, NRF-2021M3A9G8025599, NRF-2019R1C1C1006867, NRF-2021M3H4A1A02051048, NRF-2021M3E5E3080379, and NRF-2018M3A9E2022821), Global Frontier Program through Center for BioNano Health-Guard funded by MSIT of Korea (H-GUARD_2013M3A6B2078950), Technology Development Program for Biological Hazards Management in Indoor Air through Korea Environment Industry & Technology Institute (KEITI) funded by Ministry of Environment (ME) of Korea (2021003370003), Nanomedical Devices Development Program of NNFC (CSM2105M101), and KRIBB Research Initiative Program (KGS-1052113 and 1711134081). Publisher Copyright: {\textcopyright} 2021 by the authors. Licensee MDPI, Basel, Switzerland.",

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AU - Yong, Dongeun

AU - Kang, Taejoon

AU - Woo, Euijeon

N1 - Funding Information: Funding: This research was supported by National R&D Programs through National Research Foundation (NRF) of Korea funded by Ministry of Science and ICT (MSIT) of Korea (NRF-2021R1A2C2011328, NRF-2021M3A9G8025599, NRF-2019R1C1C1006867, NRF-2021M3H4A1A02051048, NRF-2021M3E5E3080379, and NRF-2018M3A9E2022821), Global Frontier Program through Center for BioNano Health-Guard funded by MSIT of Korea (H-GUARD_2013M3A6B2078950), Technology Development Program for Biological Hazards Management in Indoor Air through Korea Environment Industry & Technology Institute (KEITI) funded by Ministry of Environment (ME) of Korea (2021003370003), Nanomedical Devices Development Program of NNFC (CSM2105M101), and KRIBB Research Initiative Program (KGS-1052113 and 1711134081). Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

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Detection of infectious viruses using crispr-cas12-based assay

Abstract

Bibliographical note

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UN SDGs

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