[show abstract][hide abstract] ABSTRACT: DNA representing RNA segment 3 of bluetongue virus (BTV) serotype 17, corresponding to the gene that codes for a group-specific antigen VP3, has been inserted into a baculovirus transfer vector in lieu of the 5' coding region of the polyhedrin gene of Autographa californica nuclear polyhedrosis virus (AcNPV). After cotransfection of Spodoptera frugiperda cells with wild-type AcNPV DNA in the presence of the derived recombinant transfer vector DNA, polyhedrin-negative recombinant baculoviruses were recovered. When S. frugiperda cells were infected with one of these recombinant viruses, a protein that was similar in size and antigenic properties to the BTV VP3 protein was synthesized. Antibodies raised in mice or rabbits to the baculovirus-expressed VP3 protein immunoprecipitated the VP3 protein of BTV-17 as well as that of BTV-10. The expressed antigen reacted with antisera representing four U.S.A. BTV serotypes in an indirect ELISA test.
Journal of General Virology 07/1987; 68 ( Pt 6):1627-35. · 3.13 Impact Factor
[show abstract][hide abstract] ABSTRACT: The complete sequence of the large RNA segment 3 (L3) of bluetongue virus serotype 10 (BTV-10) has been determined from DNA copies of the viral RNA cloned in the E. coli plasmid pBR322. The L3 viral RNA is 2772 nucleotides long with a single open reading frame of 2706 nucleotides. The L3 predicted primary gene product (VP3) is 103 342 daltons and has a net charge at neutral pH of -5. The sequence of the L3 RNA species differs by 126 point mutations from that of BTV-17 (i.e., 95.5% homology; see M. Purdy, J. Petre and P. Roy, J. Virol. 51, 754-759, 1984). The predicted L3 primary gene products of the two viruses differ by 9 amino acids. These differences correspond to 9 point mutations and represent 0.15% of the sites where nucleotide substitution could cause an amino acid change. By contrast, another 114 point mutations in the genome correspond to 6.5% of the available sites where nucleotide substitutions could be silent (i.e., where a nucleotide substitution may not cause an amino acid change). Three point mutations are in the 3' non-coding region of the RNA species. The quantitative differences between the coding and silent mutations are interpreted as representing the result of gene product conservation.
Virus Research 10/1985; 3(2):181-90. · 2.75 Impact Factor
[show abstract][hide abstract] ABSTRACT: The complete sequence of the RNA which encodes the major outer-shell-neutralizing antigen (VP2) of bluetongue virus serotype 10 was determined from overlapping cDNA clones inserted into pBR322. The segment L2 RNA was 2,926 base pairs long (1.87 X 10(6) daltons) and had, in one strand, an open reading frame capable of coding for a protein that had a calculated size of 111,122 daltons (956 amino acids) and a +11.5 net charge. The coding strands of both the L2 gene and the group-specific L3 gene of bluetongue virus serotype 17 (M. Purdy, J. Petre, and P. Roy, J. Virol. 51:754-759, 1984) had common sequences of some six nucleotides at their 5' termini (namely, GUUAAA...) and eight nucleotides (namely, ...ACACUUAC) at their 3' termini. Both had short 5' noncoding regions with AUG codons at residues 20 to 22 (L2) and 18 to 20 (L3). The sequences flanking these AUG codons were similar (A/GCCAUGG). The 3' noncoding regions were longer (36 nucleotides for L2, 49 nucleotides for L3). The predicted amino acid sequence of the L2, compared with the similarly sized L3 gene product, was rich in cysteine residues and charged amino acids.
Journal of Virology 10/1985; 55(3):826-30. · 5.08 Impact Factor