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通过体内外基因重组,将大肠杆菌粘附因子cs3基因定位整合到痢疾杆菌福氏2a疫苗株T32菌染色体的asd基因内,使asd基因灭活;将来内O抗原基因克隆至无抗药性表达载体pXL378,获得重组质粒pXL390,将其转化asd-的T32受体菌,构建成福氏2a和宋内双价苗苗株FS01。实验表明:重组质粒pXL390在不带任何抗菌素基因的情况下,在asd-的T32受体菌内是稳定的。FS01株遗传稳定,能表达两种痢疾菌的PLS-O抗原,无明显毒性作用。动物试验表明,以FS01株皮下免疫的小鼠对福氏2a和宋内有毒株的腹腔攻击有100%的保护。
Through in vitro and in vivo genetic recombination, the cs3 gene of Escherichia coli was localized and integrated into the asd gene of the chromosome of T32 strain of Shigella flexneri 2a vaccine strain to inactivate the asd gene. In the future, intracellular O antigen gene was cloned into non-drug resistant expression vector pXL378, the recombinant plasmid pXL390 was obtained, which was transformed into asd- T32 receptor bacterium to construct the bivalent seedling strain FS01 of Fuzhou 2a and Song. The experiment shows that the recombinant plasmid pXL390 is stable in asd- T32 receptor bacteria without any antibiotic gene. The FS01 strain is genetically stable and can express PLS-O antigen of two kinds of dysentery bacteria without obvious toxic effect. Animal experiments showed that mice subcutaneously immunized with strain FS01 had 100% protection against intraperitoneal attacks by both the Furae 2a and the toxic strains in the Song family.