An Infectious cDNA Clone of SARS-CoV-2

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

doi:10.1016/j.chom.2020.04.004

An Infectious cDNA Clone of SARS-CoV-2

Xuping Xie, Antonio Muruato, Kumari G. Lokugamage, Krishna Narayanan, Xianwen Zhang, Jing Zou, Jianying Liu, Craig Schindewolf, Nathen E. Bopp, Patricia V. Aguilar, Kenneth S. Plante, Scott C. Weaver, Shinji Makino, James W. LeDuc, Vineet D. Menachery, Pei Yong Shi

Research output: Contribution to journal › Article › peer-review

462 Scopus citations

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.

Original language	English (US)
Pages (from-to)	841-848.e3
Journal	Cell Host and Microbe
Volume	27
Issue number	5
DOIs	https://doi.org/10.1016/j.chom.2020.04.004
State	Published - May 13 2020

Keywords

COVID-19
SARS-CoV
SARS-CoV-2
antiviral
coronavirus
vaccine

ASJC Scopus subject areas

Parasitology
Microbiology
Virology

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10.1016/j.chom.2020.04.004

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title = "An Infectious cDNA Clone of SARS-CoV-2",

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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author = "Xuping Xie and Antonio Muruato and Lokugamage, {Kumari G.} and Krishna Narayanan and Xianwen Zhang and Jing Zou and Jianying Liu and Craig Schindewolf and Bopp, {Nathen E.} and Aguilar, {Patricia V.} and Plante, {Kenneth S.} and Weaver, {Scott C.} and Shinji Makino and LeDuc, {James W.} and Menachery, {Vineet D.} and Shi, {Pei Yong}",

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doi = "10.1016/j.chom.2020.04.004",

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T1 - An Infectious cDNA Clone of SARS-CoV-2

AU - Xie, Xuping

AU - Muruato, Antonio

AU - Lokugamage, Kumari G.

AU - Narayanan, Krishna

AU - Zhang, Xianwen

AU - Zou, Jing

AU - Liu, Jianying

AU - Schindewolf, Craig

AU - Bopp, Nathen E.

AU - Aguilar, Patricia V.

AU - Plante, Kenneth S.

AU - Weaver, Scott C.

AU - Makino, Shinji

AU - LeDuc, James W.

AU - Menachery, Vineet D.

AU - Shi, Pei Yong

PY - 2020/5/13

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N2 - 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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An Infectious cDNA Clone of SARS-CoV-2

Abstract

Keywords

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