Construction and eukaryotic expression of recombinant plasmid encoding fusion protein of goat complement C3d and foot-and-mouth disease virus VP1.
- Author:
Jieyu LING
1
;
Zhao LIU
;
Tiezhu TONG
;
Huiying FAN
;
Dekun ZHANG
;
Huanchun CHEN
;
Aizhen GUO
Author Information
1. State Key Laboratory of Agricultural Microbiology, Wuhan 430070, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Capsid Proteins;
biosynthesis;
genetics;
Cloning, Molecular;
Complement C3d;
biosynthesis;
genetics;
immunology;
Female;
Foot-and-Mouth Disease Virus;
genetics;
Goats;
HeLa Cells;
Humans;
Immunologic Factors;
biosynthesis;
genetics;
immunology;
Plasmids;
genetics;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
immunology;
Transfection
- From:
Chinese Journal of Biotechnology
2008;24(2):209-213
- CountryChina
- Language:Chinese
-
Abstract:
We constructed a recombinant plasmid encoding VP1 gene of O type foot-and-mouth disease virus fused to a molecular adjuvant, goat complement C3d gene. The goat C3d gene was cloned and three copies were tandem-linked with the linker (G4S)2 sequence. VP1 gene of O type foot-and-mouth disease virus was linked to three tandem repeats of C3d through the linker sequence and cloned into pUC19 to obtain the recombinant plasmid pUC19-VP1-C3d3. The VP1-C3d3 fusion gene was then subcloned into the eukaryotic vector pcDNA3.1(+) that had been modified to contain the tissue plasminogen activator (tPA) leader sequence to obtain pcDNA3.1-tPA-VP1-C3d3. HeLa cells were transfected with pcDNA3.1-tPA-VP1-C3d3 by Lipofectamine 2000. Indirect immunofluorescent assay and Western blot assay showed that VP1-C3d3 fusion gene was successfully expressed in HeLa cells. The fusion protein with the expected size 133 kD could be secreted outside the cells. This study laid a good foundation to further research on the novel vaccine against foot-and-mouth disease virus by using goat C3d as a molecular adjuvant to enhance the immunogenicity of VP1.
