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Two novel transcriptional reporter systems for monitoring Helicobacter pylori stress responses.
Plasmid ( IF 2.6 ) Pub Date : 2019-11-01 , DOI: 10.1016/j.plasmid.2019.102442
A M Belova 1 , D V Basmanov 1 , V V Babenko 1 , O V Podgorny 2 , T V Mitko 1 , K A Prusakov 1 , D V Klinov 1 , V N Lazarev 1
Affiliation  

Helicobacter pylori, a human pathogen linked to many stomach diseases, is well adapted to colonize aggressive gastric environments, and its virulence factors contribute this adaptation. Here, we report the construction of two novel H. pylori vectors, pSv2 and pSv4, carrying a reporter gene fused to the promoters of virulence factor genes for monitoring the response of single H. pylori cells to various stresses. H. pylori cryptic plasmids were modified by the introduction of the Escherichia coli origin of replication, chloramphenicol resistance cassette, and promoterless gfp gene to produce E. coli/H. pylori shuttle vectors. The promoter regions of vacA and ureA genes encoding well-characterized H. pylori virulence factors were fused to the promoterless gfp gene. Recording the GFP fluorescence signal from the genetically modified H. pylori cells immobilized in specifically designed microfluidic devices revealed the response of transcriptional reporter systems to osmotic stress, acidic stress, elevated Ni2+ concentration or iron chelation. Our observations validate the utility of the pSv2 and pSv4 vectors to monitor the regulation of virulence factor genes in diverse strains and clinical isolates of H. pylori.

中文翻译:

两个新的转录报告系统,用于监测幽门螺杆菌的应激反应。

幽门螺杆菌(Helicobacter pylori)是一种与许多胃部疾病有关的人类病原体,非常适合在侵略性胃部环境中定植,其毒力因子有助于这种适应。在这里,我们报告两个新型幽门螺杆菌载体,pSv2和pSv4的建设,携带与致病因子基因启动子融合的报告基因,用于监测单个幽门螺杆菌细胞对各种胁迫的反应。通过引入大肠杆菌的复制起点,氯霉素抗性盒和无启动子的gfp基因来修饰幽门螺杆菌隐性质粒,以产生大肠杆菌/ H。幽门螺杆菌载体。编码特征明确的幽门螺杆菌毒力因子的vacA和ureA基因的启动子区域与无启动子的gfp基因融合。从转基因的H记录GFP荧光信号。固定在专门设计的微流体装置中的幽门螺杆菌细胞揭示了转录报告系统对渗透胁迫,酸性胁迫,Ni2 +浓度升高或铁螯合的反应。我们的观察结果证实了pSv2和pSv4载体在监测幽门螺杆菌不同菌株和临床分离株中毒力因子基因的调控中的实用性。
更新日期:2019-11-01
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