表达肠出血性大肠杆菌 O157:H7 EspA 抗原的减毒沙门氏菌株的构建

目的 利用平衡致死系统构建表达O157:H7 EspA 的减毒鼠伤寒沙门氏菌.方法 构建表达EspA的重组质粒,再将其转入终宿主菌减毒鼠伤寒沙门氏菌x4550 株中构建成口服活疫苗株,用IPTG进行诱导表达,经聚丙烯酰胺凝胶电泳、Westen-blot检测EspA蛋白的表达情况.并观察重组菌体外培养的稳定性.结果 利用宿主-载体平衡致死系统构建了表达O157:H7 EspA的重组减毒沙门氏菌,经Tricine-SDS-PAGE电泳出现了1条Mr约21 000的蛋白条带,Westen-blot检测能与抗EspA的单克隆抗体发生特异性反应.且在没有选择压力的条件下体外能稳定地繁殖、生长和传代.结论 表达O157:H7 EspA的重组减毒沙门氏菌构建成功,为发展口服抗EHEC O157:H7的疫苗奠定了初步基础.

Construction of attenuated Salmonella typhimurium vaccine strain expressing EHEC O157:H7 EspA

Objective To construct an attenuated Salmonella typhimurium vaccine strain expressing the gene EspA of EHEC O157:H7 with balanced lethal system. Methods The gene of EspA of EHEC O157:H7 was amplified, and then recombined with a vector (asd ). The recombinant was transfected into attenuated Salmonella typhimurium x4550. The expression of EspA was analyzed by Tricine-SDS-PAGE and Western blotting. The stability of vaccine strains was investigated by generation culture in vitro. Results The stable attenuated Salmonella typhmurium vaccine strains expressing O157:H7 EspA was constructed with balanced lethal system. T The expressed products with Mr21000 could react with the antibody of EspA. Conclusion The attenuated Salmonella typhimurium vaccine strain could probably serve as an oral live vaccine against O157:H7 infection.