Construction, identification and expression of a recombinant Efs-Em Ⅱ/3-Em14-3-3 vaccine against Echinococcus multilocularis mediated by Enterococcus faecalis
10.3760/cma.j.cn231583-20230403-00078
- VernacularTitle:粪肠球菌介导的多房棘球绦虫重组Efs-EmⅡ/3-Em14-3-3疫苗的构建、鉴定及表达
- Author:
Wengui LI
1
;
Xingkun OU
;
Ailin HE
Author Information
1. 重庆医科大学附属第一医院传染病寄生虫病研究所，重庆 400016
- Keywords:
Enterococcus faecalis;
Echinococcus multilocularis;
EmⅡ/3;
Em14-3-3;
Vaccine
- From:
Chinese Journal of Endemiology
2023;42(11):876-882
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct a recombinant Efs-EmⅡ/3-Em14-3-3 vaccine against Echinococcus multilocularis (Em) using Enterococcus faecalis (Efs) as a vector, and investigate its antigenicity. Methods:The recombinant plasmid pGEX-EmⅡ/3-Em14-3-3 was transformed into Efs ATCC47077 strain using electroporation method, and the recombinant Efs-EmⅡ/3-Em14-3-3 vaccine was constructed. The plasmid was extracted for PCR amplification and identification. The recombinant Efs-EmⅡ/3-Em14-3-3 vaccine was expressed through isopropyl-β-D-thiogalactoside (IPTG) induction, and the recombinant protein was verified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting, and the proportion of expressed proteins in the total proteins of the bacteria was analyzed by thin layer scanning technology.Results:Using the plasmid extracted from recombinant Efs bacteria as a template, the EmⅡ/3-Em14-3-3 fusion gene with a size of about 2 554 bp could be amplified by PCR. The relative molecular mass ( Mr) of the expressed EmⅡ/3-Em14-3-3 fusion protein was approximately 119 × 10 3 by SDS-PAGE; after 5 h induction by IPTG, the expression level of target protein was high, accounting for about 9% of the total bacterial protein. Western blotting showed that the expressed protein could be recognized by mouse serum infected with alveolar hydatid cyst. Conclusion:The recombinant Efs-EmⅡ/3-Em14-3-3 vaccine is successfully constructed, and the expressed fusion protein shows specific antigenicity.
