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An Infectious cDNA Clone of SARS-CoV-2.

Identifieur interne : 003A22 ( Ncbi/Merge ); précédent : 003A21; suivant : 003A23

An Infectious cDNA Clone of SARS-CoV-2.

Auteurs : Xuping Xie [États-Unis] ; Antonio Muruato [États-Unis] ; Kumari G. Lokugamage [États-Unis] ; Krishna Narayanan [États-Unis] ; Xianwen Zhang [États-Unis] ; Jing Zou [États-Unis] ; Jianying Liu [États-Unis] ; Craig Schindewolf [États-Unis] ; Nathen E. Bopp [États-Unis] ; Patricia V. Aguilar [États-Unis] ; Kenneth S. Plante [États-Unis] ; Scott C. Weaver [États-Unis] ; Shinji Makino [États-Unis] ; James W. Leduc [États-Unis] ; Vineet D. Menachery [États-Unis] ; Pei-Yong Shi [États-Unis]

Source :

RBID : pubmed:32289263

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.

DOI: 10.1016/j.chom.2020.04.004
PubMed: 32289263

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pubmed:32289263

Le document en format XML

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<name sortKey="Leduc, James W" sort="Leduc, James W" uniqKey="Leduc J" first="James W" last="Leduc">James W. Leduc</name>
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<nlm:affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Galveston National Laboratory, University of Texas Medical Branch, Galveston, TX, USA.</nlm:affiliation>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Galveston National Laboratory, University of Texas Medical Branch, Galveston, TX</wicri:regionArea>
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<region type="state">Texas</region>
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<name sortKey="Menachery, Vineet D" sort="Menachery, Vineet D" uniqKey="Menachery V" first="Vineet D" last="Menachery">Vineet D. Menachery</name>
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<nlm:affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Department of Pathology and Center for Biodefense & Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: vimenach@UTMB.edu.</nlm:affiliation>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Department of Pathology and Center for Biodefense & Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX</wicri:regionArea>
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<name sortKey="Shi, Pei Yong" sort="Shi, Pei Yong" uniqKey="Shi P" first="Pei-Yong" last="Shi">Pei-Yong Shi</name>
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<nlm:affiliation>Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Sealy Institute for Vaccine Sciences, University of Texas Medical Branch, Galveston, TX, USA; Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA; Department of Pharmacology & Toxicology, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: peshi@UTMB.edu.</nlm:affiliation>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea>Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Sealy Institute for Vaccine Sciences, University of Texas Medical Branch, Galveston, TX, USA; Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA; Department of Pharmacology & Toxicology, University of Texas Medical Branch, Galveston, TX</wicri:regionArea>
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<series>
<title level="j">Cell host & microbe</title>
<idno type="eISSN">1934-6069</idno>
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<date when="2020" type="published">2020</date>
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<div type="abstract" xml:lang="en">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
<sup>6</sup>
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.</div>
</front>
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<DateRevised>
<Year>2020</Year>
<Month>04</Month>
<Day>14</Day>
</DateRevised>
<Article PubModel="Print-Electronic">
<Journal>
<ISSN IssnType="Electronic">1934-6069</ISSN>
<JournalIssue CitedMedium="Internet">
<PubDate>
<Year>2020</Year>
<Month>Apr</Month>
<Day>09</Day>
</PubDate>
</JournalIssue>
<Title>Cell host & microbe</Title>
<ISOAbbreviation>Cell Host Microbe</ISOAbbreviation>
</Journal>
<ArticleTitle>An Infectious cDNA Clone of SARS-CoV-2.</ArticleTitle>
<ELocationID EIdType="pii" ValidYN="Y">S1931-3128(20)30231-6</ELocationID>
<ELocationID EIdType="doi" ValidYN="Y">10.1016/j.chom.2020.04.004</ELocationID>
<Abstract>
<AbstractText>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
<sup>6</sup>
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.</AbstractText>
<CopyrightInformation>Copyright © 2020 Elsevier Inc. All rights reserved.</CopyrightInformation>
</Abstract>
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<Author ValidYN="Y">
<LastName>Xie</LastName>
<ForeName>Xuping</ForeName>
<Initials>X</Initials>
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<Affiliation>Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: xuxie@UTMB.edu.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Muruato</LastName>
<ForeName>Antonio</ForeName>
<Initials>A</Initials>
<AffiliationInfo>
<Affiliation>Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Lokugamage</LastName>
<ForeName>Kumari G</ForeName>
<Initials>KG</Initials>
<AffiliationInfo>
<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Narayanan</LastName>
<ForeName>Krishna</ForeName>
<Initials>K</Initials>
<AffiliationInfo>
<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Zhang</LastName>
<ForeName>Xianwen</ForeName>
<Initials>X</Initials>
<AffiliationInfo>
<Affiliation>Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Zou</LastName>
<ForeName>Jing</ForeName>
<Initials>J</Initials>
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<Affiliation>Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Liu</LastName>
<ForeName>Jianying</ForeName>
<Initials>J</Initials>
<AffiliationInfo>
<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
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<LastName>Schindewolf</LastName>
<ForeName>Craig</ForeName>
<Initials>C</Initials>
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<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
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<LastName>Bopp</LastName>
<ForeName>Nathen E</ForeName>
<Initials>NE</Initials>
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<Affiliation>Department of Pathology, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
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<LastName>Aguilar</LastName>
<ForeName>Patricia V</ForeName>
<Initials>PV</Initials>
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<Affiliation>Department of Pathology, University of Texas Medical Branch, Galveston, TX, USA; World Reference Center for Emerging Viruses and Arboviruses, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
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<LastName>Plante</LastName>
<ForeName>Kenneth S</ForeName>
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<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; World Reference Center for Emerging Viruses and Arboviruses, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
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<LastName>Weaver</LastName>
<ForeName>Scott C</ForeName>
<Initials>SC</Initials>
<AffiliationInfo>
<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; World Reference Center for Emerging Viruses and Arboviruses, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Institute for Translational Sciences, University of Texas Medical Branch, Galveston, TX, USA; Department of Pathology and Center for Biodefense & Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX, USA; Sealy Institute for Vaccine Sciences, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Makino</LastName>
<ForeName>Shinji</ForeName>
<Initials>S</Initials>
<AffiliationInfo>
<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
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<LastName>LeDuc</LastName>
<ForeName>James W</ForeName>
<Initials>JW</Initials>
<AffiliationInfo>
<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Galveston National Laboratory, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y">
<LastName>Menachery</LastName>
<ForeName>Vineet D</ForeName>
<Initials>VD</Initials>
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<Affiliation>Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Department of Pathology and Center for Biodefense & Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: vimenach@UTMB.edu.</Affiliation>
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</Author>
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<ForeName>Pei-Yong</ForeName>
<Initials>PY</Initials>
<AffiliationInfo>
<Affiliation>Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Sealy Institute for Vaccine Sciences, University of Texas Medical Branch, Galveston, TX, USA; Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA; Department of Pharmacology & Toxicology, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: peshi@UTMB.edu.</Affiliation>
</AffiliationInfo>
</Author>
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<Language>eng</Language>
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<PublicationType UI="D016428">Journal Article</PublicationType>
</PublicationTypeList>
<ArticleDate DateType="Electronic">
<Year>2020</Year>
<Month>04</Month>
<Day>09</Day>
</ArticleDate>
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<Country>United States</Country>
<MedlineTA>Cell Host Microbe</MedlineTA>
<NlmUniqueID>101302316</NlmUniqueID>
<ISSNLinking>1931-3128</ISSNLinking>
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<CitationSubset>IM</CitationSubset>
<KeywordList Owner="NOTNLM">
<Keyword MajorTopicYN="N">COVID-19</Keyword>
<Keyword MajorTopicYN="N">SARS-CoV</Keyword>
<Keyword MajorTopicYN="N">SARS-CoV-2</Keyword>
<Keyword MajorTopicYN="N">antiviral</Keyword>
<Keyword MajorTopicYN="N">coronavirus</Keyword>
<Keyword MajorTopicYN="N">vaccine</Keyword>
</KeywordList>
<CoiStatement>Declaration of Interests X.X., V.D.M, and P.-Y.S. have filed a provisional patent on the reverse genetic system of SARS-CoV-2. Other authors have no conflicts of interest to declare.</CoiStatement>
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<Month>03</Month>
<Day>22</Day>
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<Year>2020</Year>
<Month>03</Month>
<Day>30</Day>
</PubMedPubDate>
<PubMedPubDate PubStatus="accepted">
<Year>2020</Year>
<Month>04</Month>
<Day>01</Day>
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<name sortKey="Aguilar, Patricia V" sort="Aguilar, Patricia V" uniqKey="Aguilar P" first="Patricia V" last="Aguilar">Patricia V. Aguilar</name>
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<name sortKey="Leduc, James W" sort="Leduc, James W" uniqKey="Leduc J" first="James W" last="Leduc">James W. Leduc</name>
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<name sortKey="Lokugamage, Kumari G" sort="Lokugamage, Kumari G" uniqKey="Lokugamage K" first="Kumari G" last="Lokugamage">Kumari G. Lokugamage</name>
<name sortKey="Makino, Shinji" sort="Makino, Shinji" uniqKey="Makino S" first="Shinji" last="Makino">Shinji Makino</name>
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<name sortKey="Muruato, Antonio" sort="Muruato, Antonio" uniqKey="Muruato A" first="Antonio" last="Muruato">Antonio Muruato</name>
<name sortKey="Narayanan, Krishna" sort="Narayanan, Krishna" uniqKey="Narayanan K" first="Krishna" last="Narayanan">Krishna Narayanan</name>
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<name sortKey="Schindewolf, Craig" sort="Schindewolf, Craig" uniqKey="Schindewolf C" first="Craig" last="Schindewolf">Craig Schindewolf</name>
<name sortKey="Shi, Pei Yong" sort="Shi, Pei Yong" uniqKey="Shi P" first="Pei-Yong" last="Shi">Pei-Yong Shi</name>
<name sortKey="Weaver, Scott C" sort="Weaver, Scott C" uniqKey="Weaver S" first="Scott C" last="Weaver">Scott C. Weaver</name>
<name sortKey="Zhang, Xianwen" sort="Zhang, Xianwen" uniqKey="Zhang X" first="Xianwen" last="Zhang">Xianwen Zhang</name>
<name sortKey="Zou, Jing" sort="Zou, Jing" uniqKey="Zou J" first="Jing" last="Zou">Jing Zou</name>
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