Optimized SARS-CoV-2 Spike gene (B.1.351 - South African origin) for mammalian cell expression

pUNO1-SpikeV3 and pUNO1-SpikeV3-dfur plasmids have been specifically designed for the expression of the SARS-CoV-2 Spike (S) protein in mammalian cells with either a functional or inactivated furin (dfur) cleavage site. These plasmids encode the full-length Spike sequence from the Beta variant (B.1.351), and for optimal cellular expression, it is codon-optimized and the C‑terminal ER-retention signal has been removed [1, 2].

Beta Variant (B.1.351 lineage) Spike Expression vectors
for cell fusion and flow cytometry assays

InvivoGen also offers:

• SARS-CoV-2-Cellular Receptor Genes
• ACE2-expressing Cell Line
• SARS-CoV-2 Spike Antibodies

Available upon request:

• 293-SARS2-S-V2-dfur cells (Alpha/B.1.1.7/U.K variant)
• 293-SARS2-S-V3-dfur cells (Beta/B.1.351/S.A variant)
• 293-SARS2-S-V5-dfur cells (Gamma/P.1/BRA variant)
• 293-SARS2-S-V8-dfur cells (Delta/B.1.617.2/IND variant)

Gene Description

These plasmids encode the Spike protein from the SARS-CoV-2 Beta variant (B.1.351), first reported in South Africa in October 2020. This variant is classified as a member of Clade 20H/501Y.V2 and B.1.351 v2 lineage (Nextstrain/Pango lineage classification) [3]. It is characterized by the presence of a number of deletions and mutations within the Spike coding region, of which, several are of concern [3,4].

S1 domain: L18F, D80A, D215G, deletion L242-L244, D614G
RBD: K417N, E484K, N501Y
S2 domain: A701V

Learn more about SARS-CoV-2 Variants

The Spike protein contains a furin cleavage site that affects its cellular expression [5, in-house data]. Therefore, depending on your application InvivoGen offers:

pUNO1-SpikeV3: with a functional furin cleavage site allowing Spike/ACE2 cell fusion assays
pUNO1-SpikeV3-dfur: with an inactive furin (dfur) cleavage site allowing improved surface expression for flow cytometry detection

More details

General Plasmid Description

These plasmids feature a potent mammalian expression cassette composed of the ubiquitous human EF1α-HTLV composite promoter and the SV40 polyadenylation (pAn) signal. The codon-optimized ORF includes the native SARS-CoV-2 Spike signal sequence. The plasmids are selectable with Blasticidin in both E. coli and mammalian cells (transient and stable transfection).

Applications

Spike-mediated cell fusion assays with pUNO1-SpikeV3
Cell surface detection by flow cytometry with pUNO1-SpikeV3-dfur
Screening of SARS-CoV-2 inhibitors including small molecules, monoclonal antibodies, or convalescent plasma

References:

1. Johnson, M.C. et al. 2020. Optimized Pseudotyping Conditions for the SARS-COV-2 Spike Glycoprotein. J Virol 94.
2. Ou, X. et al. 2020. Characterization of spike glycoprotein of SARS-CoV-2 on virus entry and its immune cross-reactivity with SARS-CoV. Nat Commun 11, 1620.
3. Garcia-Beltran, W.F. et al. 2021. Multiple SARS-CoV-2 variants escape neutralization by vaccine-induced humoral immunity. Cell. doi:10.1016/j.cell.2021.03.013
4. Tegally, H. et al. 2020. Emergence and rapid spread of a new severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) lineage with multiple spike mutations in South Africa. Medrxiv. doi:10.1101/2020.12.21.20248640v1
5. Coutard, B. et al. 2020. The spike glycoprotein of the new coronavirus 2019-nCoV contains a furin-like cleavage site absent in CoV of the same clade. Antiviral Res 176, 104742.

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