Prokaryotic Expression of Capsid Protein and Preparation of Polyclonal Antibody of Novel Goose Astrovirus
In order to develop serological detection methods and highly effective antibodies against the novel goose astrovirus(GAstV),the ORF2 gene sequence of the novel GAstV HNZZ-4 strain was cloned and the pET-32a-Cap recombinant plasmid was constructed.Prokaryotic expression of capsid protein(Cap)was performed.Polyclonal antibody was prepared by immunizing rabbits with recombinant Cap.Western blot and immunoperoxidase single-layer assay were used to determine the reactivity of the polyclonal antibody.The results showed that the recombinant plasmid pET-32a-Cap was digested by the restriction enzymes NdeⅠand XhoⅠ,and two bands with the expected size of about 5 000 bp and 750 bp were obtained.Sequencing results of positive recombinant plasmid showed that the recombinant expression vector pET-32a-Cap was successfully constructed.Prokaryotic expression of pET-32a-Cap obtained GAstV recombinant Cap with molecular weight of about 27.5 ku.Rabbit polyclonal antibodies against GAstV Cap could react specifically with the recombinant Cap and bind to GAstV infecting LMH cells.The rabbit polyclonal antibody against novel GAstV Cap prepared in this study retains its antigenicity and has good specificity.
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