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.
【저자키워드】 COVID-19, SARS-CoV-2, Infectious disease, virus, CRISPR-Cas12, 【초록키워드】 viruses, public health, coronavirus, reliability, SARS-CoV, sputum, MERS-CoV, RNA, lateral flow, Protein, point-of-care, amplification, sensitivity, specificity, Coronavirus disease-19, outbreak, POCT, cleavage, reverse transcription, nasal swab, RT-RPA, change, COVID-19 patients, Concentration, Amplicon, isothermal, Middle East, acute respiratory syndrome, complex, probes, fluorescent, infectious viruses, respiratory syndrome coronavirus, applicability, gRNA, polymerase, produced, cause, anticipated, palindromic repeat, 【제목키워드】 detection,