Detection of Infectious Viruses Using CRISPR-Cas12-Based Assay

Biosensors

Taejoon Kang¹  Oh Seok Kwon²  Yong-Sam Kim²  Dongeun Yong³  Chandana S. Talwar⁴  Euijeon Woo⁴  Kwang-Hyun Park⁴  Woo-Chan Ahn⁴ 

[1] Bionanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 34141, Korea;Department of Biomolecular Science, University of Science and Technology (UST), 217 Gajeong-ro, Yuseong-gu, Daejeon 34113, Korea;Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, 50-1 Yonsei-ro, Seodaemun-gu, Seoul 03722, Korea;Disease Target Structure Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 34141, Korea;

关键词: SARS-CoV-2;    COVID-19;    CRISPR-Cas12;    virus;    infectious disease;   

DOI  :  10.3390/bios11090301

来源: DOAJ

【 摘 要 】

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.

【 授权许可】

Unknown