The construction of Schistosoma japonicum vaccine BCG-Sj26GST and its identification.
The expression of foreign gene, Schistosoma Japonicum 26 ku antigen (Sj26GST), in Bacillus Calmette-Guerin (BCG), Mycobacterium (M. smegmatis) and Escherichia coli (E. coli) were studied. The cDNA fragment encoding Sj26GST was amplified by PCR using plasmid pGEX, which could express Sj26GST in E. coli as template. The Sj26GST cDNA was cloned into the down-stream of human M. tuberculosis heat shock protein (hsp) 70 promoter with correct reading frame, and then the DNA fragment containing hsp70 promoter and Sj26GST gene were subcloned together into E. coli-Mycobacteria shuttle plasmid pBCG-2000 to construct the expression shuttle plasmid pBCG-Sj26. The recombinant BCG and M. smegmatis mc(2)155, which were electroplated with pBCG-Sj26, could express Sj26GST and the recombinant Schistosoma Japonicum vaccine BCG-Sj26GST was made. The recombinant Sj26GST (rSj26GST) were soluble and could be observed on SDS-PAGE at molecular weight of 26 ku. The content of rSj26GST accounted for 15% and 10% of total bacterial protein in BCG and M. smegmatis respectively. The results of Western blot showed the combination of rSj26GST with antibody of GST.
Huangfu
Y
Y
Department of Medical Molecular Biology, Research Center of Experimental Medicine, Tongji Medical University, Wuhan 430030.
Zheng
B
B
Cheng
J
J
Liang
J
J
Feng
Z
Z
D000818:Animals; D000947:Antigens, Helminth / Q000096:biosynthesis / Q000235:genetics*; D001500:BCG Vaccine / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D012549:Schistosoma japonicum / Q000235:genetics* / Q000276:immunology; D014614:Vaccines, Synthetic / Q000235:genetics / Q000276:immunology*
D000947:Antigens, Helminth; D001500:BCG Vaccine; D018840:HSP70 Heat-Shock Proteins; D011993:Recombinant Fusion Proteins; D014614:Vaccines, Synthetic