Construction and identification of recombinant Mycobacterium smegmatis vaccine expressing Cysticercus cellulosae cC1 antigen
- VernacularTitle:猪囊尾蚴特异性抗原cC1重组耻垢分枝杆菌疫苗的构建与鉴定
- Author:
Xuemei WANG
;
Jiangkun LUO
;
Qian LI
;
Jiangyan LI
;
Yong CHEN
;
Zhiyong TAO
;
Hui XIA
;
Qiang FANG
- Publication Type:Journal Article
- Keywords:
Cysticercus cellulosae;
cC1;
Mycobacteria smegmatis;
Vaccine
- From:
Chinese Journal of Schistosomiasis Control
2014;(3):287-291
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct recombinant Mycobacterium smegmatis vaccine expressing Cysticercus cellulosae cC1 anti-gen. Methods The recombinant pET28a-cC1 plasmid was extracted and double digested by Xho I and BamH I restriction en-zymes,and shuttle plasmid pMV261 was extracted and double digested by Hind III and BamH I restriction enzymes. Both frag-ments were modified by Klenow fragment to form blunt end,then the large fragments of cC1 and pMV261 plasmid were purified and ligated by T4 ligase enzyme. The recombinant pMV261-cC1 plasmid was constructed and sequenced. Then the pMV261-cC1 plasmid was transformed into Mycobacterium smegmatis by the electrotransformation method. The recombinant cC1-Mycobacterium smegmatis was induced by heat and identified by the Western blotting method with the sera of cysticercosis patients. In addition, the growth states of the Mycobacterium smegmatis and the recombinant cC1-Mycobacterium smegmatis were compared and the growth curves were drawn. Results The restriction enzyme and sequencing results showed that the recombinant pMV261-cC1 plasmid was successfully constructed. After heat induction,a 40 kD band was showed by PAGE analysis of cC1-Mycobacterium smegmatis. The Western blotting results showed that the sera of cysticercosis patients could recognize the 40 kDa band,which sug-gested that cC1 protein was expressed in Mycobacterium smegmatis. Compared with the Mycobacterium smegmatis,the recombi-nant cC1-Mycobacterium smegmatis showed no significant difference in proliferation characteristics. Conclusion The recombi-nant cC1-Mycobacterium smegmatis vaccine has been successfully constructed.
