中文摘要：

目的：构建奇异变形菌（PMI）聚磷酸盐激酶1（polyphosphate kinase 1，PPK1）基因缺失株，为研究 ppk1基因在PM I致尿路感染中的作用及其机制奠定基础。方法通过融合PCR将目的基因上游及下游同源臂连接成一个片段，并克隆入自杀质粒pCVD442，将重组质粒转化入大肠埃希菌SM10λpir中，再与PMI受体菌进行接合试验，通过氨苄西林、蔗糖筛选得到 p pk1基因缺失株（△ pk1），进行PCR和DNA测序鉴定；体外比较野生株和缺失株在低营养条件下的生存能力。结果利用融合PCR将目的基因上、下游片段连接成一个重组片段，并连接至pCVD442自杀质粒；重组自杀质粒进入PMI后出现了同源重组事件，同源片段发生替换，ppk1基因被敲除，通过PCR及测序分析证实基因组目的基因已缺失；经体外测定MOPS培养基中的生长曲线，发现敲除株与野生株相比，在低营养环境中生存能力明显降低。结论自杀质粒同源重组方法可用于PM I基因敲除株的构建，是研究PMI基因功能的重要手段，ppk1基因与PMI在低营养环境中的生存能力有关。

英文摘要：

OBJECTIVE To construct the Proteus mirabilis strain with polyphosphate kinase 1 （PPK1） gene dele‐tion so as to lay foundation for the study of the p pk1 gene in pathogenesis of urinary tract infections induced by P . mirabilis .METHODS The upstream and downstream flanking DNA fragments of target gene were fused by PCR and then cloned into the suicide vector pCVD442 .The recombinant plasmid was electransformed into SM10λpir of E .coli strain and introduced into P .mirabilis by conjugation .The ppk1 gene deletion strains were screened by ampicillin and sucrose and identified by PCR and DNA sequencing ;then the in vitro comparative study was con‐ducted for the survival abilities of the wild strains and the ppk1 gene deletion strains .RESULTS The upstream and downstream fragments of the target gene were fused to a recombinant fragment and connected to the suicide vector pCVD442 .After the recombinant suicide plasmid was electransformed into the PMI gene ,the homologous recom‐bination events occurred ,the homologous fragments were replaced ,the p pk1 gene was deleted ,and the PCR and DNA sequencing proved the deletion of the target gene of genome .The in vitro determination of the growing curve of MOPS in culture medium revealed that as compared with the wild strain ,the survival ability of the ppk1 gene deletion strains under the poor nutrition condition was remarkably reduced .CONCLUSION The suicide plasmid ho‐mologous recombination can be applied in the construction of the ppk1 gene deletion P .mirabilis strain and plays an important role in the study of the gene function of the P .mirabilis ,and the p pk1 gene is associated with sur‐vival ability of the P .mirabilis strain under the poor nutrition condition .