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《农业生物技术：英文版》
>Construction of Recombinant Expression Plasmids Containing H and F Protein Genes of Canine Distemper Virus Isolated from a Mink and Their Expression in Prokaryotic Cells
Construction of Recombinant Expression Plasmids Containing H and F Protein Genes of Canine Distemper Virus Isolated from a Mink and Their Expression in Prokaryotic Cells
[Objective] This study aimed to construct the recombinant expression plasmids containing H and F protein genes of Canine distemper virus isolated from a mink and to express these two genes in prokaryotic cells as well as to study the reactogenicity of the expressed products. [Method] RT-PCR amplification was used to obtain H and F protein genes; TA cloning and subcloning techniques were used to construct the cloning plasmids( pMD-18T-H and pMD-18T-F) and recombinant expression plasmids( pET28a-H and pET28a-F) ; SDS-PAGE and Western-blotting were adopted to verify whether the target proteins were successfully expressed. [Result] The recombinant expression plasmids pET28a-H and pET28a-F containing H and F protein genes of Canine distemper virus isolated from a mink were successfully constructed,and both the expressed H and F proteins with respectively relative molecular mass of 31 400 and 38 200 produced positive reaction with the CDV standard positive serum. [Conclusion] The H and F proteins expressed in prokaryotic cells were the same with the natural ones in terms of reactogenicity,which can be utilized for diagnosis of a CDV's infection or for an epidemiological investigation. Meanwhile,they also provide a basis for developing genetically engineered subunit vaccines.
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