破伤风毒素C基因的克隆及重组C蛋白的表达和免疫原性研究

Study on cloning,expression and immunogenicity of the fragment C of tetanuss toxin

对破伤风毒素C基因(TTC)进行了克隆,在大肠杆菌中构建了C基因-硫氧还蛋白融合表达系统并对其进行了重组表达,对表达后蛋白进行了纯化和rTTC的免疫原性初步研究。结果表明,该系统可高水平表达可溶性rTTC,rTTC表达量占可溶性蛋白的28.19%,纯化后纯度为96.92%;所获得的rTTC具有良好的免疫原性,可为开发新一代破伤风亚单位疫苗奠定基础。

To clone, express, purify protein and analyse the immunogenicity of the fragment C of tetanus toxin,the fragment C gene of tetanus toxin was amplified from Clostridium tetani genomic DNA by PCR. It was inserted into the hlgh-expresslon vector pET32a ,and expression of this plasmid in Escherichia coli BL21 （DE3） resulted in the production of a fusion protein. The recombinant fragment-C-thioredoxin proteins were purified significantly in one step by affinity chromatography ,and then used to immunize mice and test the antibody titers. The result of SDS-PAGE showed that a specific recombinant protein was expressed,accounting for 28.19% of the soluble protein. Western blot analysis indicated that this product had certain antigenicity,and the final purity was 96.92%. 14 days after the third immunization the anti-tetanus antibody titers of the mice of the group rTTC were detected. The recombinant protein is an immunogenicity antigen of tetanus toxin and thus constructs a basis for the development of vaccine in the future.