rpoS基因插入突变对铜绿假单胞菌两个吩嗪合成基因簇的调控  

Effect of rpoS mutation on two gene clusters of phenazine in Psedomonas aeruginosa PAO1

作　　者：周金凤[1] 葛宜和[1] 刘婷[1] 程显好[1] 王磊[1] 高兴喜[1] 

机构地区：[1]鲁东大学生命科学学院,烟台264000

出　　处：《微生物学报》2010年第3期411-417,共7页Acta Microbiologica Sinica

基　　金：鲁东大学科技基金(20063302);鲁东大学实验室开放计划~~

摘　　要：【目的】为了研究铜绿假单胞菌rpoS基因对吩嗪(Phenazine)合成基因簇phz1和phz2的调控方式与机制。【方法】采用抗庆大霉素基因(gentamycin resistance cassette,aacC1)插入失活的策略构建了rpoS基因突变株PA-SG;同时利用lacZ的翻译融合表达载体pME6015,构建了phz1′-′lacZ和phz2′-′lacZ翻译融合表达载体pMEZ1和pMEZ2。采用电转化法分别将pMEZ1、pMEZ2和pME6015导入铜绿假单胞菌突变株PA-SG和野生株PAO1,用Miller法检测融合β-半乳糖苷酶活性。【结果】在KMB或PPM培养基中,pMEZ1在突变株PA-SG中的表达均增强,为野生株的4-5倍;而pMEZ2在突变株PA-SG中的表达均降低,野生株是突变株的2-3倍。【结论】由此推测,铜绿假单胞菌rpoS基因对两个不同吩嗪合成基因簇的调控作用具有特异性,在一定程度上,rpoS负调控phz1,正调控phz2。[Objective]As an opportunistic pathogen,Pseudomonas aeruginosa PAO1 can produce phenazine and its derivatives,which play a critical role in their pathogenesis. In many bacteria,RpoS,the product of rpoS gene,mediates biosynthesis of a set of secondary metabolites. [Objective] This study aims to elucidate rpoS gene’s function and regulation on two phenazine gene clusters in Pseudomonas aeruginosa PAO1. [Methods] The rpoS gene and its upstream and downstream fragments were cloned from the chromosome of Pseudomonas aeruginosa. With the insertion of gentamycin resistance cassette （aacC1）,the mutant PA-SG has been created by homologous recombination. Translational fusion plasmids phz1′-′lacZ（pMEZ1） and phz2′-′lacZ（pMEZ2） were constructed,and then were introduced into the wild type strain PAO1 and the mutant PA-SG,respectively. Activities of beta-galactosidase in them were determined with Miller method. [Results] In KMB or PPM medium,beta-galactosidase activity of phz1′-′lacZ in the mutant PA-SG is much more than that in the wild type strain. However,beta-galactosidase activity of phz2′-′lacZ in the wild type strain is 2-3 folds more than that in the mutant PA-SG. [Conclusion] With these results,it is suggested that regulation mediated by rpoS gene on two phenazine loci is specific and different.

关 键 词：铜绿假单胞菌 phz1 phz2 RPOS 吩嗪 

分 类 号：Q93[生物学—微生物学]