Construction of the recombinant plasmids encoding nonstructural proteins of SFTS virus and immunogenicity study
10.3760/cma.j.issn.1003-9279.2017.03.013
- VernacularTitle: SFTS病毒非结构蛋白重组质粒的构建及其体液免疫原性研究
- Author:
Yujie AI
1
;
Yaping HAN
;
Ke JIN
;
Ming YUE
;
Yiqing ZHOU
;
Jun LI
Author Information
1. First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China
- Publication Type:Journal Article
- Keywords:
Severe fever with thrombocytopenia syndrome(SFTS) virus;
Nonstructural protein;
Recombinant plasmid;
Humoral immune response
- From:
Chinese Journal of Experimental and Clinical Virology
2017;31(3):240-244
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To study the construction and humoral immunogenicity of the recombinant plasmids encoding nonstructural proteins(NSs) of severe fever with thrombocytopenia syndrome(SFTS) virus(SFTSV).
Methods:Recombinant plasmids encoding NSs gene and optimized NSs gene of SFTSV were constructed by polymerase chain reaction(PCR) and cloned into eukaryotic expression vector pJW4303, which were named pJW4303-NSs and pJW4303-NSs-opt, respectively. Then, the plasmid pJW4303-NSs was tagged with Flag, named pJW4303-NSs-Flag. Meanwhile, Nhe I restrict site and tissue plasminogen activator(tPA) signal sequence were inserted to construct bi-optimized recombinant plasmid named pJW4303-tPA-NSs-opt. All plasmids were identified by sequencing. The transient expression of NSs was confirmed by Western blotting in human embryonic kidney 293T cells. The NSs-specific IgG antibodies in BALB/c mice which were immunized by intramuscular injection with electroporation were examined by enzyme-linked immunosorbent assay(ELISA).
Results:The recombinant plasmids pJW4303-NSs, pJW4303-NSs-opt, pJW4303-tPA-NSs-opt and pJW4303-NSs-Flag were successfully constructed. The expression of NSs was confirmed in lysates and supernatants of 293T cells. The NSs-specific IgG responses of all three recombinant plasmids were detected by ELISA in BALB/c mice. It was found that optimized recombinant plasmid pJW4303-NSs-opt elicited higher levels of the NSs-specific IgG than that of pJW4303-NSs in week 6, 8 which induced stronger immune response.
Conclusions:The recombinant plasmids encoding SFTSV NSs possess the satisfied immunogenicity. In addition, the plasmid pJW4303-NSs-opt could induce the strongest humoral immune response.
