Generation of Recombinant SARS-CoV-2 Using a Bacterial Artificial Chromosome

SARS-CoV-2, the causative agent of COVID-19, has been responsible for a million deaths worldwide as of September 2020. At the time of this writing, there are no available US FDA-approved therapeutics for the treatment of SARS-CoV-2 infection. Here, we describe a detailed protocol to generate recombi...

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Bibliographic Details
Published inCurrent protocols in microbiology (Online) Vol. 59; no. 1; p. e126
Main Authors Chiem, Kevin, Ye, Chengjin, Martinez-Sobrido, Luis
Format Journal Article
LanguageEnglish
Published United States 01.12.2020
Subjects
Animals
Betacoronavirus - genetics
Betacoronavirus - growth & development
Chlorocebus aethiops
Chromosomes, Artificial, Bacterial - genetics
Containment of Biohazards
SARS-CoV-2
Transfection
Vero Cells
COVID-19
bacterial artificial chromosome
virus rescue
SARS-CoV-2
reverse genetics
infectious clone
coronavirus
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Summary:SARS-CoV-2, the causative agent of COVID-19, has been responsible for a million deaths worldwide as of September 2020. At the time of this writing, there are no available US FDA-approved therapeutics for the treatment of SARS-CoV-2 infection. Here, we describe a detailed protocol to generate recombinant (r)SARS-CoV-2 using reverse-genetics approaches based on the use of a bacterial artificial chromosome (BAC). This method will allow the production of mutant rSARS-CoV-2-which is necessary for understanding the function of viral proteins, viral pathogenesis and/or transmission, and interactions at the virus-host interface-and attenuated SARS-CoV-2 to facilitate the discovery of effective countermeasures to control the ongoing SARS-CoV-2 pandemic. © 2020 Wiley Periodicals LLC. Basic Protocol: Generation of recombinant SARS-CoV-2 using a bacterial artificial chromosome Support Protocol: Validation and characterization of rSARS-CoV-2.
ISSN:1934-8533
DOI:10.1002/cpmc.126