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Rotavirus infection is a major cause of dehydrating diarrhea in infants worldwide. The non-toxic cholera toxin B subunit(CTB), known as an immunomodulatory carrier, might help to stimulate mucosal immune response when coupled to rotavirus antigens in oral immunization. Here we report for the first time the construction of a translational fusion of CTB gene 5' to the VP6 gene of a human rotavirus A(field strain T114), and expression of the CTB-VP6 fusion protein in E. coli BL21(DE3). The expressed fusion protein has a molecular weight of 56 kD, as expected, and accounts for about 15% of the total E. coli protein. Western blottings with the hyperimmune serum against rotavirus strain WA and the antibody against cholera toxin indicated that the fusion protein retains the antigenicity identical to the native CTB and VP6. The GM1-ELISA analysis proves that the renatured CTB-VP6 has strong affinity for GM1 ganglioside.

By cloning vp1 and vp2 genes of chicken anaemia virus into transfer vector pBacPAK8, recombinant transfer plasmids pBac-vp1 and pBac-vp2 were obtained. Then BmN cells were co-transfected with linearized baculovirus Bm-BacPAK6 DNA and above two recombinant plasmids respectively, recombinant viruses Bm-vp1 and Bm-vp2 were constructed and used to co-infect silkworms to express recombinant proteins. The results indicated that recombinant VP1 and VP2 could induce the corresponding antibody in chickens using immunofluorescence assay and the expression products could protect filial generation from the attack of CAV. Recombinant BmNPV expressing VP1 and VP2 is, therefore, a great hopeful production system for a subunit vaccine against CAV infection.

The complementary DNA copies of the gene 4 were prepared by reverse transcription and polymerase chain reaction (RT-PCR) from genomic RNA of simian rotavirus strain SA11. A complete VP4 gene was been inserted into a baculovirus intermediate vector pVL-1393 which was under control of the polyhedrin promoter. The outer capsid protein VP4 which was a major neutralization antigen of rotavirus, was expressed in high yield in Spodoptera frugiperda cell line. The VP4 protein expressed was recognized with a hyperimmunized serum directed against the rotavitus by immunoblot assay. Expressed VP4 protein own about 10 percent of the total proteins. The baculovirus recombinant expressed full-length VP4 protein was used to immunize animals. A high level of neutralizing antibody directed against parentalstrain SA11 virus was developed in the animals. The anti-vp4 antibody could block visible cytopathic effect (CPE) forming of SA11 virus in MA104 cells. Immunofluorescence test and Western blot also confirmed that the antibody could recognize specific antigen of rotavirus in infectious MA104 cells. The results suggest that VP4 protein expressed in a recombinant baculovirus has antigenicity and immunogenicity as well. It may be an important component in developing rotavirus recombinant and subunit vaccine.