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目的研究H-NS对副溶血弧菌(Vibrio parahaemolyticus,VP)泳动能力的调控作用。方法将VP接种于泳动实验平板上,37℃静置培养4.5 h后,通过比较不同菌株间菌苔直径的差异来判定H-NS对泳动表型的影响。提取WT和hns基因突变株(Δhns)的总RNA,采用实时定量RT-PCR方法研究H-NS对极鞭毛蛋白基因fla A的转录调控关系。将fla A启动子区DNA克隆入p HRP309质粒的β-半乳糖苷酶基因上游,构建Lac Z重组质粒,并将该重组质粒转入WT和Δhns中,通过比较二者中β-半乳糖苷酶活性的差异研究H-NS对fla A的调控关系。结果与结论动力表型结果显示,H-NS可促进VP的泳动能力;实时定量RT-PCR和Lac Z报告基因融合实验结果表明H-NS正调控fla A的转录。这表明H-NS至少可通过激活fla A的转录而促进VP的泳动能力。
Objective To investigate the regulatory effect of H-NS on the motility of Vibrio parahaemolyticus (VP). Methods VP was inoculated on the plate of swimming experiment. After standing at 37 ℃ for 4.5 h, the effect of H-NS on the motility phenotype was determined by comparing the differences in the diameters of the bacteria among the different strains. The total RNA of WT and hns gene mutant (Δhns) was extracted and the transcriptional regulation of flaA gene was detected by real-time quantitative RT-PCR. The fla A promoter region DNA was cloned into the upstream of the β-galactosidase gene of p HRP309 plasmid to construct a Lac Z recombinant plasmid and the recombinant plasmid was transformed into WT and Δhns and compared with β-galactoside The difference of enzyme activity of H-NS on fla A regulation. RESULTS AND CONCLUSION: The results of kinetic phenotype showed that H-NS could promote the migration of VP. The results of real-time quantitative RT-PCR and Lac Z reporter gene fusion show that H-NS regulates the transcription of fla A. This suggests that H-NS promotes the VP’s ability to swim at least by activating fla A transcription.