鹅星状病毒 Spike 蛋白与铁蛋白融合表达及纳米颗粒自组装OA

To prepare nanoparticles of goose astrovirus(GoAstV)spike protein,the Spike gene was cloned into the prokaryotic expression vector Fer-pET32a carrying the ferritin gene.After identification of double enzyme digestion and sequencing,the recombinant plasmid was transformed into competent E.coli BL21 cells.The Spike-Fer recombinant protein was induced by IPTG and purified by Ni-column affinity chromatography.The recombinant protein was analyzed and identified by Western blot,transmission electron microscopy,and nanoparticle size analyzer.The results showed that an 880 bp Spike gene fragment was successfully amplified,The constructed Spike-Fer-pET32a recombinant plasmid was digested with restriction endonucleases EcoRⅠ and XhoⅠ,and a vector band at 5 900 bp and a Spike-Fer tandem gene band at 1 425 bp were observed,which was consistent with the expected results.The Spike-Fer recombinant protein was successfully expressed in E.coli mainly in soluble form with a molecular weight of about 70 kDa.The Spike-Fer recombinant protein could specifically react with the anti-His tag monoclonal antibody.Uniform spherical nanoparticles with a diameter of about 50.7 nm formed by self-assembly of the Spike-Fer recombinant protein were observed under transmission electron microscopy.In conclusion,self-assembled spherical nanoparticles of goose astrovirus were successfully prepared,which provides a material basis for the development of a goose astrovirus nanoparticle vaccines.