[1]李国强,朱丽萍,朱丽臻,等.UPEC双组分信号系统KguS/KguR调控基因缺失株的构建及其生长特性研究[J].南京农业大学学报,2018,41(6):1113-1117.[doi:10.7685/jnau.201801046]
 LI Guoqiang,ZHU Liping,ZHU Lizhen,et al.Construction and growth characteristics analysis of the deletion mutant of the target gene regulated by two-component signaling system KguS/KguR of UPEC[J].Journal of Nanjing Agricultural University,2018,41(6):1113-1117.[doi:10.7685/jnau.201801046]
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UPEC双组分信号系统KguS/KguR调控基因缺失株的构建及其生长特性研究()
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《南京农业大学学报》[ISSN:1000-2030/CN:32-1148/S]

卷:
41卷
期数:
2018年6期
页码:
1113-1117
栏目:
出版日期:
2018-11-25

文章信息/Info

Title:
Construction and growth characteristics analysis of the deletion mutant of the target gene regulated by two-component signaling system KguS/KguR of UPEC
作者:
李国强1 朱丽萍1 朱丽臻2 陈蕾蕾3 颜世敢1
1. 齐鲁工业大学(山东省科学院)生物工程学院/山东省微生物工程重点实验室, 山东 济南 250353;
2. 山东省临沂市相公中心医院, 山东 临沂 276025;
3. 山东省农业科学院农产品研究所, 山东 济南 250100
Author(s):
LI Guoqiang1 ZHU Liping1 ZHU Lizhen2 CHEN Leilei3 YAN Shigan1
1. School of Bioengineering/Shandong Key Laboratory of Microbial Bioengineering, Qilu University of Technology(Shandong Academy of Sciences), Jinan 250353, China;
2. Xianggong Central Hospital of Linyi City, Shandong Province, Linyi 276025, China;
3. Institute of Agricultural Food, Shandong Academy of Agricultural Sciences, Jinan 250100, China
关键词:
尿道致病性大肠杆菌KguS/KguRc5032-c5037基因Red重组系统
Keywords:
uropathogenic Escherichia coliKguS/KguRc5032-c5037 genesRed recombination system
分类号:
Q784
DOI:
10.7685/jnau.201801046
文献标志码:
A
摘要:
[目的]构建尿道致病性大肠杆菌(uropathogenic Escherichia coli,UPEC)CFT073株的双组分信号系统KguS/KguR调控的c5032—c5037基因的缺失株,并研究基因缺失株在有氧和厌氧条件下的生长特性。[方法]运用Red重组系统将带有c5032—c5037基因同源臂的氯霉素抗性基因取代c5032—c5037基因,在温度敏感型质粒pCP20作用下消除氯霉素抗性基因,构建基因缺失株CFT073Δc5032—c5037,用PCR和基因测序验证基因敲除是否成功,测定有氧和厌氧条件下CFT073Δc5032—c5037在以α-酮戊二酸为唯一碳源的M9培养基中培养不同时间的菌液D600值。[结果]构建了CFT073Δc5032—c5037,PCR验证及基因测序结果均表明已成功敲除c5032—c5037基因;厌氧条件下CFT073Δc5032—c5037在以α-酮戊二酸为唯一碳源的M9培养基中生长比野生型菌株CFT073缓慢,且差异极显著(P<0.01),但在有氧条件下二者的生长无显著差异。[结论]成功构建了基因缺失株CFT073Δc5032—c5037,并试验证实KguS/KguR调控的c5032—c5037基因在厌氧条件下参与α-酮戊二酸的利用,为进一步研究KguS/KguR在UPEC中的代谢适应机制奠定了基础。
Abstract:
[Objectives] This study aims to delete c5032-c5037 genes regulated by two-component signaling system KguS/KguR of uropathogenic Escherichia coli (UPEC)strain CFT073, and study the growth characteristics of the mutant under aerobic or anaerobic conditions. [Methods] Red recombination system was used to replace c5032-c5037 genes with chloramphenicol resistance gene containing the homologous arms of c5032-c5037 genes, and then chloramphenicol resistance genes was eliminated by introducing a temperature sensitive plasmid pCP20. Finally c5032-c5037 genes deletion mutant CFT073Δc5032-c5037 was constructed. PCR and gene sequencing were used to confirm whether genes were successfully knocked out. The growth characteristics of the mutant CFT073Δc5032-c5037 were monitored in M9 minimal medium with α-ketoglutarate as the sole carbon source under aerobic or anaerobic conditions. [Results] The results of PCR and DNA sequencing of the mutant CFT073Δc5032-c5037 accorded with theoretical results. No significant growth differences were observed between the wild strain CFT073 and the mutant CFT073Δc5032-c5037 under aerobic conditions, while significant differences appeared between them under anaerobic conditions (P<0.01). [Conclusions] Genes deletion mutant CFT073Δc5032-c5037 was constructed successfully and the growth result revealed c5032-c5037 genes were closely correlated with the utilization of α-ketoglutarate under anaerobic conditions. This study could establish the foundation for further research in the function of KguS/KguR.

参考文献/References:

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备注/Memo

备注/Memo:
收稿日期:2018-01-25。
基金项目:国家重点研发计划项目(2017YFC1601404);山东省自然科学基金项目(ZR2018LC003);国家自然科学基金项目(31470243);山东省重点研发计划项目(2014GSF120006);山东省研究生教育创新计划项目(SDYY14024)
作者简介:李国强,硕士研究生。
通信作者:颜世敢,教授,研究方向为微生物学与免疫学,E-mail:yanshigan@126.com。
更新日期/Last Update: 2018-11-23