An Infectious cDNA Clone of SARS-CoV-2
The ongoing pandemic of COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), underscores the urgency to develop experimental systems for studying this virus and identifying countermeasures. We report a reverse genetic system for SARS-CoV-2. Seven complimentary DNA (cDNA)...
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| Published in | Cell host & microbe Vol. 27; no. 5; pp. 841 - 848.e3 |
|---|---|
| Main Authors | , , , , , , , , , , , , , , , |
| Format | Journal Article |
| Language | English |
| Published |
United States
Elsevier Inc
13.05.2020
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| Subjects | |
| Online Access | Get full text |
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| Abstract | The ongoing pandemic of COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), underscores the urgency to develop experimental systems for studying this virus and identifying countermeasures. We report a reverse genetic system for SARS-CoV-2. Seven complimentary DNA (cDNA) fragments spanning the SARS-CoV-2 genome were assembled into a full-genome cDNA. RNA transcribed from the full-genome cDNA was highly infectious after electroporation into cells, producing 2.9 × 106 plaque-forming unit (PFU)/mL of virus. Compared with a clinical isolate, the infectious-clone-derived SARS-CoV-2 (icSARS-CoV-2) exhibited similar plaque morphology, viral RNA profile, and replication kinetics. Additionally, icSARS-CoV-2 retained engineered molecular markers and did not acquire other mutations. We generated a stable mNeonGreen SARS-CoV-2 (icSARS-CoV-2-mNG) by introducing this reporter gene into ORF7 of the viral genome. icSARS-CoV-2-mNG was successfully used to evaluate the antiviral activities of interferon (IFN). Collectively, the reverse genetic system and reporter virus provide key reagents to study SARS-CoV-2 and develop countermeasures.
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•A reverse genetic system has been established for SARS-CoV-2•Recombinant SARS-CoV-2 replicates as efficiently as the original clinical isolate•A stable mNeonGreen reporter SARS-CoV-2 has been developed•The mNeonGreen SARS-CoV-2 can be used to screen antiviral inhibitors
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a devastating global pandemic. Xie et al. generated an infectious cDNA clone of SARS-CoV-2 and a mNeonGreen reporter virus. Recombinant SARS-CoV-2 and reporter virus replicate as efficiently as the original clinical isolate. |
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| AbstractList | The ongoing pandemic of COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), underscores the urgency to develop experimental systems for studying this virus and identifying countermeasures. We report a reverse genetic system for SARS-CoV-2. Seven complimentary DNA (cDNA) fragments spanning the SARS-CoV-2 genome were assembled into a full-genome cDNA. RNA transcribed from the full-genome cDNA was highly infectious after electroporation into cells, producing 2.9 × 10
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plaque-forming unit (PFU)/mL of virus. Compared with a clinical isolate, the infectious-clone-derived SARS-CoV-2 (icSARS-CoV-2) exhibited similar plaque morphology, viral RNA profile, and replication kinetics. Additionally, icSARS-CoV-2 retained engineered molecular markers and did not acquire other mutations. We generated a stable mNeonGreen SARS-CoV-2 (icSARS-CoV-2-mNG) by introducing this reporter gene into ORF7 of the viral genome. icSARS-CoV-2-mNG was successfully used to evaluate the antiviral activities of interferon (IFN). Collectively, the reverse genetic system and reporter virus provide key reagents to study SARS-CoV-2 and develop countermeasures.
•
A reverse genetic system has been established for SARS-CoV-2
•
Recombinant SARS-CoV-2 replicates as efficiently as the original clinical isolate
•
A stable mNeonGreen reporter SARS-CoV-2 has been developed
•
The mNeonGreen SARS-CoV-2 can be used to screen antiviral inhibitors
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a devastating global pandemic. Xie et al. generated an infectious cDNA clone of SARS-CoV-2 and a mNeonGreen reporter virus. Recombinant SARS-CoV-2 and reporter virus replicate as efficiently as the original clinical isolate. The ongoing pandemic of COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), underscores the urgency to develop experimental systems for studying this virus and identifying countermeasures. We report a reverse genetic system for SARS-CoV-2. Seven complimentary DNA (cDNA) fragments spanning the SARS-CoV-2 genome were assembled into a full-genome cDNA. RNA transcribed from the full-genome cDNA was highly infectious after electroporation into cells, producing 2.9 × 10 plaque-forming unit (PFU)/mL of virus. Compared with a clinical isolate, the infectious-clone-derived SARS-CoV-2 (icSARS-CoV-2) exhibited similar plaque morphology, viral RNA profile, and replication kinetics. Additionally, icSARS-CoV-2 retained engineered molecular markers and did not acquire other mutations. We generated a stable mNeonGreen SARS-CoV-2 (icSARS-CoV-2-mNG) by introducing this reporter gene into ORF7 of the viral genome. icSARS-CoV-2-mNG was successfully used to evaluate the antiviral activities of interferon (IFN). Collectively, the reverse genetic system and reporter virus provide key reagents to study SARS-CoV-2 and develop countermeasures. The ongoing pandemic of COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), underscores the urgency to develop experimental systems for studying this virus and identifying countermeasures. We report a reverse genetic system for SARS-CoV-2. Seven complimentary DNA (cDNA) fragments spanning the SARS-CoV-2 genome were assembled into a full-genome cDNA. RNA transcribed from the full-genome cDNA was highly infectious after electroporation into cells, producing 2.9 × 106 plaque-forming unit (PFU)/mL of virus. Compared with a clinical isolate, the infectious-clone-derived SARS-CoV-2 (icSARS-CoV-2) exhibited similar plaque morphology, viral RNA profile, and replication kinetics. Additionally, icSARS-CoV-2 retained engineered molecular markers and did not acquire other mutations. We generated a stable mNeonGreen SARS-CoV-2 (icSARS-CoV-2-mNG) by introducing this reporter gene into ORF7 of the viral genome. icSARS-CoV-2-mNG was successfully used to evaluate the antiviral activities of interferon (IFN). Collectively, the reverse genetic system and reporter virus provide key reagents to study SARS-CoV-2 and develop countermeasures.The ongoing pandemic of COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), underscores the urgency to develop experimental systems for studying this virus and identifying countermeasures. We report a reverse genetic system for SARS-CoV-2. Seven complimentary DNA (cDNA) fragments spanning the SARS-CoV-2 genome were assembled into a full-genome cDNA. RNA transcribed from the full-genome cDNA was highly infectious after electroporation into cells, producing 2.9 × 106 plaque-forming unit (PFU)/mL of virus. Compared with a clinical isolate, the infectious-clone-derived SARS-CoV-2 (icSARS-CoV-2) exhibited similar plaque morphology, viral RNA profile, and replication kinetics. Additionally, icSARS-CoV-2 retained engineered molecular markers and did not acquire other mutations. We generated a stable mNeonGreen SARS-CoV-2 (icSARS-CoV-2-mNG) by introducing this reporter gene into ORF7 of the viral genome. icSARS-CoV-2-mNG was successfully used to evaluate the antiviral activities of interferon (IFN). Collectively, the reverse genetic system and reporter virus provide key reagents to study SARS-CoV-2 and develop countermeasures. The ongoing pandemic of COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), underscores the urgency to develop experimental systems for studying this virus and identifying countermeasures. We report a reverse genetic system for SARS-CoV-2. Seven complimentary DNA (cDNA) fragments spanning the SARS-CoV-2 genome were assembled into a full-genome cDNA. RNA transcribed from the full-genome cDNA was highly infectious after electroporation into cells, producing 2.9 × 106 plaque-forming unit (PFU)/mL of virus. Compared with a clinical isolate, the infectious-clone-derived SARS-CoV-2 (icSARS-CoV-2) exhibited similar plaque morphology, viral RNA profile, and replication kinetics. Additionally, icSARS-CoV-2 retained engineered molecular markers and did not acquire other mutations. We generated a stable mNeonGreen SARS-CoV-2 (icSARS-CoV-2-mNG) by introducing this reporter gene into ORF7 of the viral genome. icSARS-CoV-2-mNG was successfully used to evaluate the antiviral activities of interferon (IFN). Collectively, the reverse genetic system and reporter virus provide key reagents to study SARS-CoV-2 and develop countermeasures. [Display omitted] •A reverse genetic system has been established for SARS-CoV-2•Recombinant SARS-CoV-2 replicates as efficiently as the original clinical isolate•A stable mNeonGreen reporter SARS-CoV-2 has been developed•The mNeonGreen SARS-CoV-2 can be used to screen antiviral inhibitors Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a devastating global pandemic. Xie et al. generated an infectious cDNA clone of SARS-CoV-2 and a mNeonGreen reporter virus. Recombinant SARS-CoV-2 and reporter virus replicate as efficiently as the original clinical isolate. |
| Author | Bopp, Nathen E. Liu, Jianying Zou, Jing Weaver, Scott C. Lokugamage, Kumari G. Narayanan, Krishna Makino, Shinji Aguilar, Patricia V. Menachery, Vineet D. Xie, Xuping Plante, Kenneth S. Schindewolf, Craig Shi, Pei-Yong Muruato, Antonio LeDuc, James W. Zhang, Xianwen |
| Author_xml | – sequence: 1 givenname: Xuping surname: Xie fullname: Xie, Xuping email: xuxie@UTMB.edu organization: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 2 givenname: Antonio surname: Muruato fullname: Muruato, Antonio organization: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 3 givenname: Kumari G. surname: Lokugamage fullname: Lokugamage, Kumari G. organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 4 givenname: Krishna surname: Narayanan fullname: Narayanan, Krishna organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 5 givenname: Xianwen surname: Zhang fullname: Zhang, Xianwen organization: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 6 givenname: Jing surname: Zou fullname: Zou, Jing organization: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 7 givenname: Jianying surname: Liu fullname: Liu, Jianying organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 8 givenname: Craig surname: Schindewolf fullname: Schindewolf, Craig organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 9 givenname: Nathen E. surname: Bopp fullname: Bopp, Nathen E. organization: Department of Pathology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 10 givenname: Patricia V. surname: Aguilar fullname: Aguilar, Patricia V. organization: Department of Pathology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 11 givenname: Kenneth S. surname: Plante fullname: Plante, Kenneth S. organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 12 givenname: Scott C. surname: Weaver fullname: Weaver, Scott C. organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 13 givenname: Shinji surname: Makino fullname: Makino, Shinji organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 14 givenname: James W. surname: LeDuc fullname: LeDuc, James W. organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 15 givenname: Vineet D. surname: Menachery fullname: Menachery, Vineet D. email: vimenach@UTMB.edu organization: Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA – sequence: 16 givenname: Pei-Yong surname: Shi fullname: Shi, Pei-Yong email: peshi@UTMB.edu organization: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA |
| BackLink | https://www.ncbi.nlm.nih.gov/pubmed/32289263$$D View this record in MEDLINE/PubMed |
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| SubjectTerms | Animals antiviral Antiviral Agents - therapeutic use Betacoronavirus - genetics Betacoronavirus - pathogenicity Chlorocebus aethiops Clone Cells coronavirus Coronavirus Infections - drug therapy Coronavirus Infections - virology COVID-19 DNA, Complementary - genetics Genes, Reporter - genetics Genome, Viral - genetics Interferons - therapeutic use Organisms, Genetically Modified - genetics Organisms, Genetically Modified - pathogenicity Pandemics Pneumonia, Viral - drug therapy Pneumonia, Viral - virology RNA, Viral - genetics SARS-CoV SARS-CoV-2 vaccine Vero Cells - virology Virus Replication - physiology |
| Title | An Infectious cDNA Clone of SARS-CoV-2 |
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