This experiment was conducted to prepare,purify,and identify the bluetongue virus VP7 antigen in a recombinant baculovirus expression system,and lay the foundation for the development of immunological antibody detection methods for bluetongue disease virus.The signal peptide and transmembrane hydrophobic region sequence of VP7 sequence was analyzed by SignalP and TMHMM online software,and rare codon optimization was performed on the selected target gene fragment.The chemically synthesized nucleotide sequence was connected to the pFastBacHTA vector,and the corresponding recombinant plasmid Bacmid-VP7-DH10Bac was constructed.After transfection into sf9 insect cells,a recombinant baculovirus Bacmid-BTV-VP7 strain was obtained,then,using the rod-shaped virus insect cell expression system to express and prepare the bluetongue disease virus VP7 antigen.The results showed that after transfection of recombinant Bacmid-VP7-DH10Bac into sf9 insect cells and continuous blind transmission for 5 generations,a recombinant Bacmid-BTV-VP7 strain was successfully obtained,and the VP7 protein was successfully expressed and purified,with a size of about 43 kDa and a protein purity of about 94%.The VP7 recombinant protein did not cross with other disease antibodies.In summary,the recombinant VP7 protein can exhibit specific reactions with BTV positive serum,with good reactivity,high protein purity,and biological activity as a viral protein.It can be used as a diagnostic antigen for the development of a series of immunological antibody diagnostic detection technologies in the later stage.
维普
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