Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic Repeats
Rapid and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is essential for controlling the pandemic of coronavirus disease 2019. Polymerase chain reaction (PCR)-based technique is the standard test for detection of SARS-CoV-2, which, however, requires complicated s...
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2021-02-01
|
| Series: | Biomedicines |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2227-9059/9/3/239 |
| _version_ | 1797417341231824896 |
|---|---|
| author | Jen-Hui Tsou Hongjie Liu Sanford A. Stass Feng Jiang |
| author_facet | Jen-Hui Tsou Hongjie Liu Sanford A. Stass Feng Jiang |
| author_sort | Jen-Hui Tsou |
| collection | DOAJ |
| description | Rapid and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is essential for controlling the pandemic of coronavirus disease 2019. Polymerase chain reaction (PCR)-based technique is the standard test for detection of SARS-CoV-2, which, however, requires complicated sample manipulation (e.g., RNA extraction) and is time-consuming. We previously demonstrated that clustered regularly interspaced short palindromic repeats (CRISPR) could precisely detect Human papillomavirus and somatic mutations of Epidermal growth factor receptor gene and Kirsten rat sarcoma viral oncogene homolog gene in plasma. The objective of this study was to develop CRISPR as a rapid test for sensitive detection of SARS-CoV-2. We first combined reverse transcription-isothermal recombinase polymerase amplification and CRSIPR to detect SARS-CoV-2 in genomic RNA of cells infected with the virus. The CRISPR assay with guide RNA against the M gene of SARS-CoV-2 had a sensitivity of 0.1 copies per µL for detection of the virus. We then used the CRSIPR assay to directly analyze raw SARS-CoV-2 samples. The CRISPR assay could sensitively detect SARS-CoV-2 in one hour without RNA extraction. This assay can be performed at a single temperature and with minimal equipment. The results were immediately visualized either by a UV light illuminator or paper strips. The diagnostic value of the test was confirmed in nasopharyngeal swab specimens. Altogether, we have developed a rapid CRISPR test for sensitive detection of SARS-CoV-2. |
| first_indexed | 2024-03-09T06:17:25Z |
| format | Article |
| id | doaj.art-02ad477f405b4cc08e7d253b50fd2ec4 |
| institution | Directory Open Access Journal |
| issn | 2227-9059 |
| language | English |
| last_indexed | 2024-03-09T06:17:25Z |
| publishDate | 2021-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Biomedicines |
| spelling | doaj.art-02ad477f405b4cc08e7d253b50fd2ec42023-12-03T11:50:47ZengMDPI AGBiomedicines2227-90592021-02-019323910.3390/biomedicines9030239Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic RepeatsJen-Hui Tsou0Hongjie Liu1Sanford A. Stass2Feng Jiang3Department of Pathology, University of Maryland School of Medicine, 10 South Pine Street, MSTF 7th Floor, Baltimore, MD 21201-1192, USADepartment of Epidemiology and Biostatistics, School of Public Health, University of Maryland, College Park, MD 20742-2611, USADepartment of Pathology, University of Maryland School of Medicine, 10 South Pine Street, MSTF 7th Floor, Baltimore, MD 21201-1192, USADepartment of Pathology, University of Maryland School of Medicine, 10 South Pine Street, MSTF 7th Floor, Baltimore, MD 21201-1192, USARapid and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is essential for controlling the pandemic of coronavirus disease 2019. Polymerase chain reaction (PCR)-based technique is the standard test for detection of SARS-CoV-2, which, however, requires complicated sample manipulation (e.g., RNA extraction) and is time-consuming. We previously demonstrated that clustered regularly interspaced short palindromic repeats (CRISPR) could precisely detect Human papillomavirus and somatic mutations of Epidermal growth factor receptor gene and Kirsten rat sarcoma viral oncogene homolog gene in plasma. The objective of this study was to develop CRISPR as a rapid test for sensitive detection of SARS-CoV-2. We first combined reverse transcription-isothermal recombinase polymerase amplification and CRSIPR to detect SARS-CoV-2 in genomic RNA of cells infected with the virus. The CRISPR assay with guide RNA against the M gene of SARS-CoV-2 had a sensitivity of 0.1 copies per µL for detection of the virus. We then used the CRSIPR assay to directly analyze raw SARS-CoV-2 samples. The CRISPR assay could sensitively detect SARS-CoV-2 in one hour without RNA extraction. This assay can be performed at a single temperature and with minimal equipment. The results were immediately visualized either by a UV light illuminator or paper strips. The diagnostic value of the test was confirmed in nasopharyngeal swab specimens. Altogether, we have developed a rapid CRISPR test for sensitive detection of SARS-CoV-2.https://www.mdpi.com/2227-9059/9/3/239CRISPRSARS-CoV-2COVID-19nucleic acidsdetection |
| spellingShingle | Jen-Hui Tsou Hongjie Liu Sanford A. Stass Feng Jiang Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic Repeats Biomedicines CRISPR SARS-CoV-2 COVID-19 nucleic acids detection |
| title | Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic Repeats |
| title_full | Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic Repeats |
| title_fullStr | Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic Repeats |
| title_full_unstemmed | Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic Repeats |
| title_short | Rapid and Sensitive Detection of SARS-CoV-2 Using Clustered Regularly Interspaced Short Palindromic Repeats |
| title_sort | rapid and sensitive detection of sars cov 2 using clustered regularly interspaced short palindromic repeats |
| topic | CRISPR SARS-CoV-2 COVID-19 nucleic acids detection |
| url | https://www.mdpi.com/2227-9059/9/3/239 |
| work_keys_str_mv | AT jenhuitsou rapidandsensitivedetectionofsarscov2usingclusteredregularlyinterspacedshortpalindromicrepeats AT hongjieliu rapidandsensitivedetectionofsarscov2usingclusteredregularlyinterspacedshortpalindromicrepeats AT sanfordastass rapidandsensitivedetectionofsarscov2usingclusteredregularlyinterspacedshortpalindromicrepeats AT fengjiang rapidandsensitivedetectionofsarscov2usingclusteredregularlyinterspacedshortpalindromicrepeats |