Construction and characterization of enterohemorrhagic Escherichia coli O157:H7 ppk-deleted strain

VernacularTitle:肠出血型大肠埃希菌O157：H7 ppk基因缺失株的构建及其生物学特性分析
Author: Peng HAN ¹ ; Qi SUN ; Suhui ZHAO ; Qiwei ZHANG ; Chengsong WAN
Author Information

1. 南方医科大学公共卫生与热带医学学院BSL-3实验室
Keywords: enterohemorrhagic Escherichia coli O157:H7; ppk gene; Red recombination system; gene mutation
From: Journal of Southern Medical University 2014;(6):904-908
CountryChina
Language:Chinese
Abstract: Objective To construct enterohemorrhagic Escherichia coli (EHEC) O157: H7 ppk gene deletion strains and study its biological characteristics. Methods The gene fragment of kanamycin resistance was amplified using a pair of homologous arm primers whose 5' and 3' ends were homologous with ppk gene and kanamycin resistance gene, respectively. EHEC O157:H7 EDL933w competent strains were prepared and transformed via electroporation with the amplification products. The ppk gene was replaced by kanamycin resistance gene using pKD46-mediated Red recombination system. The recombinant strain was confirmed by PCR and sequencing, and its morphology, growth ability and adhesion were assessed using Gram staining, OD60 value and Giemsa staining. Results and Conclusion We established a ppk-deleted EHEC O157:H7 EDL933w strain with kanamycin resistance and compared the biological characteristics of the wild-type and mutant strains, which may facilitate further study of the regulatory mechanism of ppk gene.