Prokaryotic expression and preparation of the polyclonal antibodies of E protein and domain Ⅲ in Zika virus
10.3969/j.issn.1002-2694.2018.00.002
- VernacularTitle:寨卡病毒E蛋白及第三结构域的原核表达和多克隆抗体制备
- Author:
Chen-Xi DING
1
;
Xu-Hui ZHU
;
Le-Le AI
;
Fu-Qiang YE
;
Wei-Long TAN
;
Dan HU
;
Jia-Feng CHEN
;
Xiao-Lu GUO
;
Xiu-Zhen PAN
;
Chang-Jun WANG
Author Information
1. 第三军医大学军事预防医学院
- Keywords:
Zika virus;
envelope protein;
prokaryotic expression;
polyclonal antibody
- From:
Chinese Journal of Zoonoses
2018;34(1):23-28
- CountryChina
- Language:Chinese
-
Abstract:
To clone,express and purify the E Protein and EDⅢ of Zika virus in E.coli and prepare two kinds of polyclonal antibodies,the virus was amplified by Vero E6 cell culture.Total RNA was extracted by RT-PCR and reverse transcribed into cDNA.The prokaryotic expression vectors pET32a/E and pET28a/EDⅢ were constructed by cDNA sequence of E and EDⅢ gene.Then,recombinant plasmids were transformed into E.coli BL21 and induced by IPTG,and purified by Ni+ column affinity chromatography.BALB/C mice were immunized with purified recombinant proteins.Antiserum was collected and titer was determined by indirect ELISA.Western blot was used to detect the specificity.Results showed that the recombinant proteins were successfully expressed and purified.The titer of the polyclonal antibodies both reached 1:409 600.Western Blot analysis showed that the polyclonal antibodies could specifically recognize the recombinant proteins.Thus,the specific polyclonal antibody were successfully prepared,laying a foundation for further study on the pathogenesis,detection methods and immune strategies of Zika virus.
