基因Ⅰ型日本脑炎病毒亚单位疫苗的构建与免疫原性评价OA

Given the urgency of Japanese encephalitis(JE)prevention and control in pig farms in China,as well as the uncertainty regarding the protective efficacy of currently available commercial genotype Ⅲ(GⅢ)Japanese encephalitis virus(JEV)vaccines against circulating genotype Ⅰ(GⅠ)JEV strains,there is an urgent need to develop safe,highly effective,and broad-spectrum novel JEV vaccines.The E protein gene of GⅠJEV was inserted into the multiple cloning sites of the baculovirus vector pFastBac Dual to construct a recombinant plasmid.The sequence-verified positive recombinant plasmid was transformed into DH10Bac competent cells,and recombinant bacmids were obtained through blue-white colony screening.After PCR verification,the recombinant bacmids were transfected into Sf9 cells to generate recombinant baculovirus rvBacmid-GⅠ-E.After successive passaging,expression and purification were performed,and the target protein was successfully identified by SDS-PAGE and Western-blot.Mouse immunization experiments demonstrated that vaccination with GⅠ-E significantly stimulated the production of JEV-specific IgG antibodies and markedly increased the levels of cytokines,including IL-2,IL-4,TNF-α,and IFN-γ.This study provides a novel immunization strategy and theoretical basis for the prevention of JE.