An Infectious cDNA Clone of SARS-CoV-2
Open accessAbstract
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 6 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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Together with reports of preserved neutralization of SARS-CoV-2 with key B.1.1.7 and B.1.351 spike mutations and of pseudovirus with the full B.1.1.7 spike mutation set, our data indicate broad BNT162b2-elicited immune recognition.
“Together with recent reports of the preserved neutralization of SARS-CoV-2 with key S mutations found in the B.1.1.7 and the B.1.351lineage 39,40 , and neutralization of pseudovirus bearing the full set of mutations of B.1.1.7 S 41 , our data indicate broad BNT162b2-elicited immune recognition 2,13,14 .”
From BNT162b2 vaccine induces neutralizing antibodies and poly-specific T cells in humans (2021) · cites this paper as [40]
The Results describe a SARS-CoV-2 infectious clone and mNeonGreen reporter virus, while the Introduction discusses their potential use in vaccine development. Our copy does not address BNT162b2-elicited immune recognition or neutralization of B.1.1.7 or B.1.351 mutations (searched: BNT162b2, variant neutralization, and neutralizing antibodies). (LLM, conf=0.75)
Experimental — accuracy not yet measuredTwo pseudovirus variants carried either eight or ten mutations found in the S protein of the B.1.351 lineage.
“Two variants had either eight (D80A/D215G/Δ242-244/ K417N/E484K/N501Y/D614G/A701V) or ten mutations (L18F/D80A/ D215G/R246I/Δ242-244/K417N/E484K/N501Y/D614G/A701V) found in S of the B.1.351lineage (also referred to as 20C/501Y.V2) 40 .”
From BNT162b2 vaccine induces neutralizing antibodies and poly-specific T cells in humans (2021) · cites this paper as [40]
The Discussion describes an infectious SARS-CoV-2 clone and proposes using it to study future spike mutations; the Methods describe three silent marker mutations. Searches for B.1.351, pseudovirus variants, and the listed spike mutations found no passage in our copy addressing the claimed eight- or ten-mutation variants. The indexed passages provide no page numbers. (LLM, conf=0.76)
Experimental — accuracy not yet measuredN501Y is located in the viral receptor-binding domain for cell entry and increases binding to the angiotensin-converting enzyme 2 receptor.
“This mutation is located in the viral receptor-binding domain (RBD) for cell entry, increases binding to the angiotensin-converting enzyme 2 receptor”
From Neutralization of SARS-CoV-2 spike 69/70 deletion, E484K and N501Y variants by BNT162b2 vaccine-elicited sera (2021) · cites this paper as [5]
N501Y enables the virus to expand its host range to infect mice.
“and enables the virus to expand its host range to infect mice 5,6 .”
From Neutralization of SARS-CoV-2 spike 69/70 deletion, E484K and N501Y variants by BNT162b2 vaccine-elicited sera (2021) · cites this paper as [5]
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Self-citationGenetic and structural basis for SARS-CoV-2 variant neutralization by a two-antibody cocktail
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In vivo antiviral host transcriptional response to SARS-CoV-2 by viral load, sex, and age
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Self-citationCholesterol 25-hydroxylase suppresses SARS-CoV-2 replication by blocking membrane fusion
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Nucleocapsid mutations R203K/G204R increase the infectivity, fitness, and virulence of SARS-CoV-2
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Self-citationClofazimine broadly inhibits coronaviruses including SARS-CoV-2
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Self-citationEngineering SARS-CoV-2 using a reverse genetic system
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Nasal delivery of an IgM offers broad protection from SARS-CoV-2 variants
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Self-citationAn overview of current COVID-19 vaccine platforms
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Self-citationStructural basis for potent neutralization of SARS-CoV-2 and role of antibody affinity maturation
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Nucleocapsid mutations in SARS-CoV-2 augment replication and pathogenesis
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Self-citationA versatile reverse genetics platform for SARS-CoV-2 and other positive-strand RNA viruses
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Self-citationThe N501Y spike substitution enhances SARS-CoV-2 transmission
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Human NLRP1 is a sensor of pathogenic coronavirus 3CL proteases in lung epithelial cells
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Quantitative SARS-CoV-2 Serology in Children With Multisystem Inflammatory Syndrome (MIS-C)
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Self-citationSARS-CoV-2 disrupts host epigenetic regulation via histone mimicry
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Rescue of SARS-CoV-2 from a Single Bacterial Artificial Chromosome
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Systematic analysis of SARS-CoV-2 infection of an ACE2-negative human airway cell
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Targeting the m 6 A RNA modification pathway blocks SARS-CoV-2 and HCoV-OC43 replicationPreprint
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SARS-CoV-2 productively infects primary human immune system cells in vitro and in COVID-19 patients
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Self-citationPharmacological inhibition of fatty acid synthesis blocks SARS-CoV-2 replication
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SARS-CoV-2 infection triggers pro-atherogenic inflammatory responses in human coronary vessels
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An intranasal vaccine durably protects against SARS-CoV-2 variants in mice
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Self-citationNeutralization against Omicron SARS-CoV-2 from previous non-Omicron infection
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Isolation and characterization of cross-neutralizing coronavirus antibodies from COVID-19+ subjects
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SARS-CoV-2 Quantum Sensor Based on Nitrogen-Vacancy Centers in Diamond
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A selective sweep in the Spike gene has driven SARS-CoV-2 human adaptation
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Transdermal Delivery Systems for Biomolecules
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- Authors
- Xie, X., Muruato, A., Lokugamage, K. G., Narayanan, K., Zhang, X., Zou, J., Liu, J., Schindewolf, C., Bopp, N. E., Aguilar, P. V., Plante, K. S., Weaver, S. C., Makino, S., LeDuc, J. W., Menachery, V. D., & Shi, P.-Y.
- Affiliations
- The University of Texas Medical Branch at Galveston (US)
- Corresponding author
- Xie, Xuping
- Menachery, Vineet D.
- Shi, Pei-Yong
- ORCID
- Venue
- Cell Host & Microbe
- Publisher
- Elsevier BV
- Published
- 2020-05
- Volume
- 27
- Issue
- 5
- Pages
- 841-848.e3
- ISSN
- 1931-3128 (ISSN-L)
- PMC
- PMC7153529
- OpenAlex
- W3015751519
- Cited by
- 647
- References
- 30
- Citations by year
- 2026: 30 · 2025: 47 · 2024: 54 · 2023: 94 · 2022: 162 · 2021: 180 · 2020: 80
- FWCI
- 16.94
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- Locations
- Cell Host & Microbe (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Centers for Disease Control and Prevention
- National Institute on Aging
- National Institute of Allergy and Infectious Diseases
- Amon G. Carter Foundation
- Summerfield G. Roberts Foundation
- National Institutes of Health (5UC7AI094660, AI114657, AI146081, R00AG049092, R24AI120942, U19AI100625)
- University of Texas Medical Branch
- Gillson Longenbaugh Foundation
- Robert J. Kleberg, Jr. and Helen C. Kleberg Foundation
- John S. Dunn Foundation
- University of Texas System (AI127744, AI134907, AI136126, AI142759, AI145617, UL1TR001439)
- Topics
- Health Sciences › Medicine › Infectious Diseases › SARS-CoV-2 and COVID-19 Research
- Life Sciences › Agricultural and Biological Sciences › Animal Science and Zoology › Animal Virus Infections Studies
- Health Sciences › Medicine › Infectious Diseases › SARS-CoV-2 detection and testing
- Keywords
- SARS-CoV-2, infectious clone, reverse genetics, reporter virus, ORF7, interferon antiviral activity, viral replication kinetics, plaque morphology, COVID-19 pandemic
- MeSH terms
- Betacoronavirus, COVID-19, SARS-CoV-2, Animals, Antiviral Agents, Chlorocebus aethiops, Clone Cells, Interferons, Pneumonia, Viral, RNA, Viral, Vero Cells, Virus Replication, Genome, Viral, Genes, Reporter, DNA, Complementary, Coronavirus Infections, Organisms, Genetically Modified, Pandemics
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