Expression of Hemagglutinin Gene of Avian Influenza Virus Subtype H5N1 in Insect Cells

[Objective] The study aimed to study the expression of Hemagglutinin(HA) gene of avian influenza virus subtype H5N1 in insect cells.[Method] The HA gene of avian influenza virus subtype H5N1 was cleaved with BamHI and NotI from plasmid pcH5HA and inserted into baculovirus transfer vector pFastBac1 for constructing the recombinant transfer vector pFast-H5HA.The pFast-H5HA was transformated into E.coli DH10Bac cells with shuttle plasmid by transposition.The white colonies were screened on the agar plate with X-gal,IPTG,gentamicin,kanamycin and tetracycline and then the stable recombinant plasmid was constructed as reBacmid-H5HA through continuous passage for 4 times.[Result] The recombinant baclovirus with HA gene of avian influenza virus subtype H5N1 was obtained by transfecting sf9 cells with the recombinant plasmid reBacmid-H5HA.The results of IFA and Western-blot assay demonstrated that HA protein was expressed in the sf9 insect cells infected with recombinant baculovirus and could agglutinate chicken red blood cells,which could be inhibited by the monoclonal antibody against H5N1 HA.[Conclusion] The recombinant HA protein could get the right expression in baculovirus,which give a good base for replacements of AIV in developing detection antigen and subunit vaccine