Xu Chong-bo, Zeng Jin, Xu Chong-li, Wang Yu-jiong
College of Bioengineering, Dalian University, Dalian 116622, China.
Wei Sheng Wu Xue Bao. 2005 Apr;45(2):205-8.
Betal-toxin and beta2-toxin genes from chromosomal DNA of Clostridium perfringens type C were amplified by PCR, PCR products were cleaved with restriction endonucleases and recovered. The recombinant plasmid pETXB1-2 containing beta1-beta2 fusion genes was constructed by recombinant technique and then transformed into Escherichia coli BL21 (DE3). The beta1-beta2 fusion proteins were expressed in recombinant strain BL21 (DE3) (pETXB1-2), and the expression level of the beta1-beta2 fusion proteins was about 15.36% of total cellular protein by SDS-PAGE and thin-layer gel scanning analysis. More importantly, immunization in a mouse model with crude preparation containing the fusion protein inclusion bodies or inactivated recombinant strain induced protection against at least 1 MLD of the toxin from Clostridium perfringens type C. Hence the fusion proteins possess a good immunogenicity. The constructed recombinant strain BL21 (DE3) (pETXB1-2) can be used as a candidate of vaccine strain.
用PCR扩增产气荚膜梭菌C型染色体DNA中的β1毒素和β2毒素基因,PCR产物用限制性内切酶切割并回收。通过重组技术构建含有β1-β2融合基因的重组质粒pETXB1-2,然后转化到大肠杆菌BL21(DE3)中。β1-β2融合蛋白在重组菌株BL21(DE3)(pETXB1-2)中表达,经SDS-PAGE和薄层凝胶扫描分析,β1-β2融合蛋白的表达量约占细胞总蛋白的15.36%。更重要的是,用含有融合蛋白包涵体的粗制品或灭活的重组菌株对小鼠模型进行免疫,可诱导产生针对至少1个产气荚膜梭菌C型毒素最小致死量的保护作用。因此,融合蛋白具有良好的免疫原性。构建的重组菌株BL21(DE3)(pETXB1-2)可作为疫苗菌株的候选菌株。