[Construction and eukaryotic expression of recombinant plasmid encoding fusion protein of goat complement C3d and foot-and-mouth disease virus VP1].
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.
Ling
Jieyu
J
State Key Laboratory of Agricultural Microbiology, Wuhan 430070, China.
Liu
Zhao
Z
Tong
Tiezhu
T
Fan
Huiying
H
Zhang
Dekun
D
Chen
Huanchun
H
Guo
Aizhen
A
D000818:Animals; D036022:Capsid Proteins / Q000096:biosynthesis / Q000235:genetics*; D003001:Cloning, Molecular; D015933:Complement C3d / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D005260:Female; D029544:Foot-and-Mouth Disease Virus / Q000235:genetics; D006041:Goats; D006367:HeLa Cells; D006801:Humans; D007155:Immunologic Factors / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D014162:Transfection
D036022:Capsid Proteins; D007155:Immunologic Factors; D011993:Recombinant Fusion Proteins; C442501:VP1 protein, Foot-and-mouth disease virus; D015933:Complement C3d