A simple reverse genetics method to generate recombinant coronaviruses. (3rd March 2022)

• Record Type: Journal Article
• Authors: Mélade, Julien; Piorkowski, Géraldine; Touret, Franck; Fourié, Toscane; Driouich, Jean‐Sélim; Cochin, Maxime; Bouzidi, Hawa Sophia; Coutard, Bruno; Nougairède, Antoine; de Lamballerie, Xavier
• Is Part Of: EMBO reports. Volume 23:Number 5(2022)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2022-03-03
• Subjects: antivirals -- in vivo experiment -- reverse genetics -- SARS‐CoV‐2 -- serology; Molecular biology -- Periodicals; Molecular Biology -- Periodicals; Molecular biology; Periodicals; 572.8
• DOI: 10.15252/embr.202153820
• Languages: English
• ISSNs: 1469-221X

Abstract

Abstract: Engineering recombinant viruses is a pre‐eminent tool for deciphering the biology of emerging viral pathogens such as the severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2). However, the large size of coronavirus genomes renders the current reverse genetics methods challenging. Here, we describe a simple method based on "infectious subgenomic amplicons" (ISA) technology to generate recombinant infectious coronaviruses with no need for reconstruction of the complete genomic cDNA and apply this method to SARS‐CoV‐2 and also to the feline enteric coronavirus. In both cases we rescue wild‐type viruses with biological characteristics similar to original strains. Specific mutations and fluorescent red reporter genes can be readily incorporated into the SARS‐CoV‐2 genome enabling the generation of a genomic variants and fluorescent reporter strains for in vivo experiments, serological diagnosis, and antiviral assays. The swiftness and simplicity of the ISA method has the potential to facilitate the advance of coronavirus reverse genetics studies, to explore the molecular biological properties of the SARS‐CoV‐2 variants, and to accelerate the development of effective therapeutic reagents. SYNOPSIS: Engineering recombinant viruses is a pre‐eminent tool for deciphering the biology of emerging viral pathogens. This study presents a simple and user‐friendly reverse genetics system to generate recombinant infectious coronaviruses. A reverse genetics system based on "infectious subgenomic amplicons" (ISA) was used to produce recombinant viruses. Wild‐type and modified SARS‐CoV‐2 and viruses carrying a red fluorescent reporter were generated. Recombinant viruses were tested in vivo, in serology tests, and used for antiviral screening to validate the method. Abstract : Engineering recombinant viruses is a pre‐eminent tool for deciphering the biology of emerging viral pathogens. This study presents a simple and user‐friendly reverse genetic system to generate recombinant infectious coronaviruses.