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

493 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.",

keywords = "COVID-19, SARS-CoV, SARS-CoV-2, antiviral, coronavirus, vaccine",

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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year = "2020",

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

language = "English (US)",

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pages = "841--848.e3",

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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

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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

ASJC Scopus subject areas

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