Construction of Recombinant Porcine rotavirus VP7 Expression Vector and Immunogenicity Analysis
WANG Hong-Tao1, WANG Pi-Wu2,*
1 College of Life Science, Tonghua Normal University, Tonghua 134000, China; 2 College of Agronomy, Jilin Agricultural University, Changchun 130118, China
Abstract：Porcine rotavirus (PoRV) is a member of the Reovirus family, belonging to the genus Rotavirus. PoRV is a widely prevalent zoonotic disease in the world. At present, developing a safe and effective vaccine is one of the important means to control rotavirus infection. The aim of this study was to express Porcine rotavirus VP7 yeast protein and to identify its immunological properties. First, the primers were designed according to the full sequence of VP7 published in GenBank (NC_011503.2), and established recombinant yeast engineering strain with fusion expression vector psep-2-VP7 from the G-2 proton containing VP7 to amplification of the purpose of the fragment, isopropyl-β-D-sulfur semi-lactose glycoside (isopropyl β-D-thiogalactoside, IPTG) induces the expression of glutathione-S-transferase (GST)-VP7 fusion protein, after which the optimized fermentation process conditions of the fusion protein GST-VP7 were isolated and purified and immunoimmune to mice (Mus musculus) for immunogenic analysis. The results showed that the size of the cloned VP7 gene was consistent with the expected glycoprotein gene, and the molecular weight of the recombinant protein was about 61 kD expressed in millet merozoa. After the fermentation conditions were optimized, the protein concentration was up to 0.99 mg/mL. The experimental results showed that after 2 weeks of VP7 protein immunization, the average geometric titer of IgG antibody in the nasal drip (IN) and intramuscular injection groups (IM) reached 495 and 680, respectively, and the average geometric titer of SIgA IN the nose drop group reached 43.5, which could produce high level of γ-interferon (IFN-γ), indicating that VP7 protein immunity could effectively induce the body to produce cellular immunity. The eukaryotic expression of VP7 protein in this study provided conditions for the follow-up study of this protein. The study on the immunogenicity of this protein has laid a foundation for the study on diagnostic antigens of rotavirus and related vaccines.
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