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目的利用Red同源重组技术构建肠出血性大肠杆菌(EHEC)O157∶H7外膜蛋白A(ompA)基因敲除菌株,研究敲除菌株的黏附能力变化,阐明OmpA在细菌黏附以及致病中的作用。方法利用Red同源重组技术对EHEC O157∶H7 ompA基因进行敲除,同时构建EHEC O157:H7 ompA回复突变菌株,最后通过细胞试验对EHEC O157∶H7野生型菌株、敲除菌株及回复突变菌株的黏附能力进行比较。结果成功构建了EHEC O157:H7 ompA基因敲除菌株及其回复突变菌株。细胞试验结果表明,与野生型菌株相比,敲除菌株的黏附能力明显下降,而将ompA基因进行回复突变后其黏附能力又得到回复。结论 OmpA在EHEC O157∶H7黏附HeLa细胞过程中发挥重要作用,ompA基因敲除菌株的获得为进一步研究其功能提供了帮助。
Objective To construct the ompA gene knockout strain of Enterohemorrhagic Escherichia coli (EHEC) O157: H7 using Red homologous recombination technology and to study the change of adhesion ability of the knockout strains, and to elucidate the role of OmpA in bacterial adhesion and pathogenicity effect. Methods EHEC O157: H7 ompA gene was knocked down by Red homologous recombination technique and the EHEC O157: H7 ompA gene was cloned. The recombinant EHEC O157: H7 ompA strain was obtained by the cell assay. Adhesion ability to compare. Results The EHEC O157: H7 ompA gene knockout strain and its backcountry mutant were successfully constructed. The cell test results showed that compared with the wild-type strain, the adhesion ability of the knock-out strain was significantly decreased, and the adhesive ability of the ompA gene was recovered after the mutation of the ompA gene. Conclusion OmpA plays an important role in the adhesion of HeLa cells to EHEC O157: H7. Obtaining the ompA gene knock-out strain can help to further study its function.