幽门螺杆菌细胞空泡毒素基因毒性片段的克隆与表达  

Cloning and expressing of vacA gene related segment with vacuolating activity of Helicobacter pylori

作　　者：刘淼[1] 杨致邦[1] 林珊珊[1] 吴利先[1] 

机构地区：[1]重庆医科大学基础医学院微生物学教研室,重庆400016

出　　处：《重庆医科大学学报》2003年第6期720-723,共4页Journal of Chongqing Medical University

摘　　要：目的 :通过基因于程技术获得具有良好抗原性的重组细胞空泡毒素毒性亚单位蛋白。方法 :利用PCR从幽门螺杆菌 (Hp)空泡毒素基因中扩增毒性片段V ,克隆及序列分析后在大肠杆菌中高效表达 ,用免疫印迹法 (Westernblotting)检测其抗原性。结果 :序列分析所得片段完整 ,与标准株AF36 170 0比较有 1.5 %的bp及一个氨基酸发生变异。与文献报道的中国菌株相比有高度的同源性。SDS -PAGE显示表达产物相对分子量为 2 70 0 0 ,表达量为 30 %以上 ,Westernblotting显示该蛋白可被HP全菌抗血清所识别。结论 :基因重组Hp细胞空泡毒素毒性蛋白具有良好的抗原性 ,可用于制备Hp感染的诊断试剂与疫苗。Objective:To obtain the antigen of recombinant vacuolating segment of Helicobacter pylori vacuolating cytotoxing by gene-engineering.Methods:The objective gene was cloned into PQE-30,sequenced and expressed using PCR and recombinant techniques.After expression,the antigenicity of recombinant protein was analyzed by Western blotting.Results:DNA sequence analysis identified that the sequence was integrated.Compared with standard strain AF361700,1.5% of the cloned gene was mutated,one amino acid radicle was changed.SDS-PAGE revealed the molecular weight of objective recombinant protein was 27 000 as it was predicted;the level of expression product was over 30%.Western blotting results showed the objective protein could be recognized by anti-serum against HP.Conclusion:This recombinant protein has perfect antigenicity,so it may be used as a protective antigen in preventing HP infection.

关 键 词：幽门螺杆菌 空泡毒素 基因克隆 表达 

分 类 号：R378.2[医药卫生—病原生物学] R392.11[医药卫生—基础医学]