| Download | - View supplementary information part 1: Rapid, high-yield production of full-length SARS-CoV-2 spike ectodomain by transient gene expression in CHO cells (PDF, 236 KiB)
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| DOI | Resolve DOI: https://doi.org/10.1016/j.jbiotec.2020.12.005 |
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| Author | Search for: Stuible, Matthew1; Search for: Gervais, Christian1; Search for: Lord-Dufour, Simon1; Search for: Perret, Sylvie1; Search for: L'Abbé, Denis1; Search for: Schrag, Joseph1; Search for: St-Laurent, Gilles1; Search for: Durocher, Yves1ORCID identifier: https://orcid.org/0000-0002-2268-4111 |
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| Affiliation | - National Research Council of Canada. Human Health Therapeutics
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| Format | Text, Article |
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| Subject | SARS-CoV-2; trimeric spike; HEK293; CHO; transient gene expression; polyethylenimine |
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| Abstract | Recombinant forms of the spike protein of SARS-CoV-2 and related viruses have proven difficult to produce with good yields in mammalian cells. Given the panoply of potential COVID-19 diagnostic tools and therapeutic candidates that require purified spike protein and its importance for ongoing SARS-CoV-2 research, we have explored new approaches for spike production and purification. Three transient gene expression methods based on PEI-mediated transfection of CHO or HEK293 cells in suspension culture in chemically-defined media were compared for rapid production of full-length SARS-CoV-2 spike ectodomain. A high-cell-density protocol using DXB11-derived CHO BRI/55E1 cells gave substantially better yields than the other methods. Different forms of the spike ectodomain were expressed, including the wild-type SARS-CoV-2 sequence and a mutated form (to favor expression of the full-length spike ectodomain stabilized in pre-fusion conformation), with and without fusion to putative trimerization domains. An efficient two-step affinity purification method was also developed. Ultimately, we have been able to produce highly homogenous preparations of full-length spike, both monomeric and trimeric, with yields of 100−150 mg/L in the harvested medium. The speed and productivity of this method support further development of CHO-based approaches for recombinant spike protein manufacturing. |
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| Publication date | 2020-12-07 |
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| Publisher | Elsevier |
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| In | |
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| Language | English |
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| Peer reviewed | Yes |
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| NRC number | NRC-HHT_53506FT |
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| Export citation | Export as RIS |
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| Report a correction | Report a correction (opens in a new tab) |
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| Record identifier | e1925bd3-b7b4-41cd-9e1b-c79e9000c46a |
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| Record created | 2021-01-06 |
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| Record modified | 2021-01-06 |
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