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A plasmid DNA-launched SARS-CoV-2 reverse genetics system and coronavirus toolkit for COVID-19 research #MMPMID33630831
Rihn SJ; Merits A; Bakshi S; Turnbull ML; Wickenhagen A; Alexander AJT; Baillie C; Brennan B; Brown F; Brunker K; Bryden SR; Burness KA; Carmichael S; Cole SJ; Cowton VM; Davies P; Davis C; De Lorenzo G; Donald CL; Dorward M; Dunlop JI; Elliott M; Fares M; da Silva Filipe A; Freitas JR; Furnon W; Gestuveo RJ; Geyer A; Giesel D; Goldfarb DM; Goodman N; Gunson R; Hastie CJ; Herder V; Hughes J; Johnson C; Johnson N; Kohl A; Kerr K; Leech H; Lello LS; Li K; Lieber G; Liu X; Lingala R; Loney C; Mair D; McElwee MJ; McFarlane S; Nichols J; Nomikou K; Orr A; Orton RJ; Palmarini M; Parr YA; Pinto RM; Raggett S; Reid E; Robertson DL; Royle J; Cameron-Ruiz N; Shepherd JG; Smollett K; Stewart DG; Stewart M; Sugrue E; Szemiel AM; Taggart A; Thomson EC; Tong L; Torrie LS; Toth R; Varjak M; Wang S; Wilkinson SG; Wyatt PG; Zusinaite E; Alessi DR; Patel AH; Zaid A; Wilson SJ; Mahalingam S
PLoS Biol 2021[Feb]; 19 (2): e3001091 PMID33630831show ga
The recent emergence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the underlying cause of Coronavirus Disease 2019 (COVID-19), has led to a worldwide pandemic causing substantial morbidity, mortality, and economic devastation. In response, many laboratories have redirected attention to SARS-CoV-2, meaning there is an urgent need for tools that can be used in laboratories unaccustomed to working with coronaviruses. Here we report a range of tools for SARS-CoV-2 research. First, we describe a facile single plasmid SARS-CoV-2 reverse genetics system that is simple to genetically manipulate and can be used to rescue infectious virus through transient transfection (without in vitro transcription or additional expression plasmids). The rescue system is accompanied by our panel of SARS-CoV-2 antibodies (against nearly every viral protein), SARS-CoV-2 clinical isolates, and SARS-CoV-2 permissive cell lines, which are all openly available to the scientific community. Using these tools, we demonstrate here that the controversial ORF10 protein is expressed in infected cells. Furthermore, we show that the promising repurposed antiviral activity of apilimod is dependent on TMPRSS2 expression. Altogether, our SARS-CoV-2 toolkit, which can be directly accessed via our website at https://mrcppu-covid.bio/, constitutes a resource with considerable potential to advance COVID-19 vaccine design, drug testing, and discovery science.
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PLoS+Biol 2021 ; 19 (2): e3001091
