A plasmid DNA-launched SARS-CoV-2 reverse genetics system and coronavirus toolkit for COVID-19 research

Rihn, S. J. et al. (2021) A plasmid DNA-launched SARS-CoV-2 reverse genetics system and coronavirus toolkit for COVID-19 research. PLoS Biology, 19(2), e3001091. (doi: 10.1371/journal.pbio.3001091) (PMID:33630831) (PMCID:PMC7906417)

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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.

Item Type:Articles
Glasgow Author(s) Enlighten ID:Bryden, Dr Steven and Royle, Jamie and Li, Dr Kathy and Gestuveo, Mr Rommel and Fares, Dr Mazigh and Robertson, Professor David and Wickenhagen, Mr Arthur and Palmarini, Professor Massimo and Tong, Dr Lily and Mair, Mr Daniel and Davis, Dr Chris and Szemiel, Dr Agnieszka and Orton, Dr Richard and Herder, Dr Vanessa and Cowton, Dr Vanessa and Shepherd, Dr James and Gunson, Dr Rory and Da Silva Filipe, Dr Ana and Orr, Mrs Anne and Goldfarb, Daniel Max and Brennan, Dr Benjamin and Hughes, Dr Joseph and Donald, Dr Claire and Pinto, Dr Rute and McFarlane, Mr Steven and De Lorenzo, Dr Giuditta and Furnon, Dr Wilhelm and Alexander, Dr Akira and Loney, Mr Colin and Kerr, Ms Karen and Turnbull, Dr Matthew and Rihn, Dr Suzannah and Dunlop, Dr James and Nichols, Mrs Jenna and Carmichael, Dr Stephen and Wilson, Professor Sam and Cole, Ms Sarah and Nomikou, Dr Kyriaki and Thomson, Professor Emma and McElwee, Dr Marion and Brunker, Dr Kirstyn and Taggart, Ms Aislynn and Parr, Dr Yasmin and Johnson, Miss Natasha and Varjak, Dr Margus and Stewart, Mr Douglas and Bakshi, Dr Siddharth and Sugrue, Dr Elena and Stewart, Dr Meredith and Smollett, Dr Katherine and Cameron Ruiz, Natalia and Lieber, Mr Gauthier and Patel, Professor Arvind and Kohl, Professor Alain
Authors: Rihn, S. J., Merits, A., Bakshi, S., Turnbull, M. L., Wickenhagen, A., Alexander, A. J.T., Baillie, C., Brennan, B., Brown, F., Brunker, K., Bryden, S. R., Burness, K. A., Carmichael, S., Cole, S. J., Cowton, V. M., Davies, P., Davis, C., De Lorenzo, G., Donald, C. L., Dorward, M., Dunlop, J. I., Elliott, M., Fares, M., da Silva Filipe, A., Freitas, J. R., Furnon, W., Gestuveo, R. J., Geyer, A., Giesel, D., Goldfarb, D. M., Goodman, N., Gunson, R., Hastie, C. J., Herder, V., Hughes, J., Johnson, C., Johnson, N., Kohl, A., Kerr, K., Leech, H., Lello, L. S., Li, K., Lieber, G., Liu, X., Lingala, R., Loney, C., Mair, D., McElwee, M. J., McFarlane, S., Nichols, J., Nomikou, K., Orr, A., Orton, R. J., Palmarini, M., Parr, Y. A., Pinto, R. M., Raggett, S., Reid, E., Robertson, D. L., Royle, J., Cameron-Ruiz, N., Shepherd, J. G., Smollett, K., Stewart, D. G., Stewart, M., Sugrue, E., Szemiel, A. M., Taggart, A., Thomson, E. C., Tong, L., Torrie, L. S., Toth, R., Varjak, M., Wang, S., Wilkinson, S. G., Wyatt, P. G., Zusinaite, E., Alessi, D. R., Patel, A. H., Zaid, A., Wilson, S. J., and Mahalingam, S.
College/School:College of Medical Veterinary and Life Sciences > School of Infection & Immunity
College of Medical Veterinary and Life Sciences > School of Infection & Immunity > Centre for Virus Research
College of Medical Veterinary and Life Sciences > School of Biodiversity, One Health & Veterinary Medicine
Journal Name:PLoS Biology
Publisher:Public Library of Science
ISSN (Online):1545-7885
Published Online:25 February 2021
Copyright Holders:Copyright © 2021 Rihn et al.
First Published:First published in PLoS Biology 19(2): e3001091
Publisher Policy:Reproduced under a Creative Commons License

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
Innate Immunity and Host Species BarriersMassimo PalmariniMedical Research Council (MRC)MC_UU_12014/10III - Centre for Virus Research
656551Arbovirus interactions with arthropod hostsAlain KohlMedical Research Council (MRC)MC_UU_12014/8MVLS III - CENTRE FOR VIRUS RESEARCH
656491Basis of the host range and tissue tropism for hepatitis C virusArvind PatelMedical Research Council (MRC)MC_UU_12014/2MVLS III - CENTRE FOR VIRUS RESEARCH
301840Host and Viral Determinants of Interferon Resistance During HIV-1 TransmissionSam WilsonMedical Research Council (MRC)MR/P022642/1III - Centre for Virus Research
173514Inhibition of HIV-1 by Type II InterferonSuzannah RihnWellcome Trust (WELLCOTR)201366/Z/16/ZIII - Centre for Virus Research
168748Identifying and characterising antiretroviral interferon stimulated genes (ISGs)Sam WilsonMedical Research Council (MRC)MR/K024752/1III - Centre for Virus Research
Viral Genomics and BioinformaticsAndrew DavisonMedical Research Council (MRC)MC_UU_12014/12III-MRC-GU Centre for Virus Research
174030DOST-Newton PhD ScholarshipAlain KohlBritish Council (UK) (BRCOU-UK)279705176III-MRC-GU Centre for Virus Research