Rotavirus as an Expression Platform of the SARS-CoV-2 Spike Protein.

BioRxiv : the Preprint Server for Biology
Asha Ann Philip, John Thomas Patton

Abstract

Rotavirus, a segmented double-stranded RNA virus, is a major cause of acute gastroenteritis in young children. The introduction of live oral rotavirus vaccines has reduced the incidence of rotavirus disease in many countries. To explore the possibility of establishing a combined rotavirus-SARS-CoV-2 vaccine, we generated recombinant (r)SA11 rotaviruses with modified segment 7 RNAs that contained coding sequences for NSP3 and FLAG-tagged portions of the SARS-CoV-2 spike (S) protein. A 2A translational element was used to drive separate expression of NSP3 and the S product. rSA11 viruses were recovered that encoded the S-protein S1 fragment, N-terminal domain (NTD), receptor-binding domain (RBD), extended receptor-binding domain (ExRBD), and S2 core (CR) domain (rSA11/NSP3-fS1, -fNTD, -fRBD, -fExRBD, and -fCR, respectively). Generation of rSA11/fS1 required a foreign-sequence insertion of 2.2-kbp, the largest such insertion yet made into the rotavirus genome. Based on isopycnic centrifugation, rSA11 containing S sequences were denser than wildtype virus, confirming the capacity of the rotavirus to accommodate larger genomes. Immunoblotting showed that rSA11/-fNTD, -fRBD, -fExRBD, and -fCR viruses expressed S products of expected ...Continue Reading

Datasets Mentioned

BETA
MN908947.3
MT655946

Methods Mentioned

BETA
transfection
immunoprecipitation assay
glycosylation
pulldown
electrophoresis
Electrophoretic profile

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