Cloning and Identification of an Unknown Gene Encoding 10.6 kDa Protein of Schistosoma japonicum
- VernacularTitle:日本血吸虫10.6 kDa膜蛋白基因的克隆及表达
- Author:
Jijia SHEN
;
Zuojun JIANG
;
Xinbing YU
;
Xuelong WANG
;
Wei WANG
- Publication Type:Journal Article
- Keywords:
Schistosoma japonicum, cDNA, gene clone, expression, vaccine
- From:
Chinese Journal of Parasitology and Parasitic Diseases
1987;0(03):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To screen a new schistosome vaccine candidate. \ Methods\ Schistosoma japonicum adult cDNA library was screened using sera from immune rabbits vaccinated with irradiated cercariae and monoclonal antibodies against membrane antigen of S.japonicum schistosomula. Three different fragments of S.japonicum cDNA genes were cloned into pGEM-T vector. The sequences of the inserts were determined using an automatic DNA sequencer and were analysed using Blast program. One of the unknown genes (B8) was selected and its ORF sequence (291 bp) was subcloned into eukaryotic expression vector. The recombinant plasmids were identified by restrictive enzymes and PCR amplification. The positive recombinant plasmids (pBK/SjB8) were transformed into host bacteria XL1-blue, and were then induced by IPTG for expression. SDS-PAGE and Western blotting analysis of total cellular protein from the bacteria were performed to detect the gene products. Results The results demonstrated that ORF of SjB8 gene was subcloned into the plasmid pBK-CMV and could express as fusion protein in XL1-blue. The results of SDS-PAGE and Western-blot also showed that the molecular weight of the fusion protein with 3 kDa ?-galactosidase was approximately 13\^6 kDa and the actual molecular weights of the SjB8 was 10\^6 kDa. The expressed fusion product of pBK/Sj-B8 could be recognized by immune serum and McAb. Conclusion A new gene of S.japonicum vaccine candidate (SjB8) was cloned into eukaryotic expression vector pBK-CMV and could express 10\^6 kDa schistosome protein. The results provide foundation for further study of the protein for its posibility as candidate vaccine.
