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. 2004 Feb 6;171(1):131–140. doi: 10.1016/0042-6822(89)90519-9

Nucleotide sequence of bovine rotavirus gene 1 and expression of the gene product in Baculovirus☆

J Cohen ∗,1, A Charpilienne ∗, S Chilmonczyk ∗, MK Estes †
PMCID: PMC7131587  PMID: 2545026

Abstract

The nucleotide sequence of the gene that encodes for the structural viral protein VP1 of bovine rotavirus (RF strain) has been determined. The sequence data indicate that segment 1 contains 3302 by and is A+T rich (65.3%). The positive strand of segment 1 contains a single open reading frame that extends 1088 codons and possesses 5′- and 3′-terminal untranslated regions of 18 and 20 bp, respectively. The first AUG conforms to the Kozak consensus sequence and if utilized, would yield a protein having a calculated molecular weight of 124–847, very close to the apparent molecular weight of VP1 (M.W. 125,000). The deduced amino acid sequence presents significant similarities with RNA-dependent RNA polymerase of several RNA viruses. VP1 was also synthesized in baculovirus using two transfer vecors: pAC461 and pVL941. Following infection of Sf9 cells with a recombinant baculovirus, a full-length nonfusion protein was synthesised which shares properties with authentic VP1 made in monkey kidney cells. The level of VP1 synthesis was about 10-fold higher when the baculovirus recombinant was derived from the pVL941 transfer vector. In that case, VP1 was expressed in yields approximately equivalent to 10% of the cellular protein. The recombinant protein was immunoprecipitated by hyperimmune serum raised against purified rotavirus. It also was immunogenic; a hyperimmune serum made in guinea pigs reacted with VP1 using immunoprecipitation and Western blot. This serum did not possess neutralization activity.

Footnotes

☆

Sequence Data from this article have been deposited with the EMBL/GenBank Data Libraries under Accession No. J04346.

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