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假单胞菌M18是一株能同时合成吩嗪-1-羧酸(PCA)和藤黄绿菌素两种抗生素的植物根际分离细菌。RelA催化合成的效应分子ppGpp能介导细菌因营养饥饿引起的应激反应。以M18菌株染色体DNA为模板,PCR扩增获得relA基因,通过庆大霉素抗性片段插入失活与同源重组技术,构建假单胞菌M18的relA突变菌株M18RAG。在PPM培养基中进行PCA发酵分析,发现突变菌株M18RAG的PCA产量显著升高,约为野生型菌株的1.5-2倍。relA基因反式互补实验以及phzA′-′lacZ翻译融合测定结果,均进一步证明了RelA对PCA生物合成及其基因表达具有抑制作用。
Pseudomonas sp. M18 is a plant rhizobacterium that can simultaneously synthesize phenazine-1-carboxylic acid (PCA) and garcinin. RelA catalyzed the synthesis of the effector molecule ppGpp can mediate stress-induced bacterial stress caused by nutrition. The relA gene was amplified by PCR using the chromosomal DNA of M18 strain as a template. The relA mutant strain M18RAG of Pseudomonas sp. M18 was constructed by insertion and inactivation of the gentamicin-resistant fragment and homologous recombination. Analysis of PCA fermentation in PPM medium showed that the PCA production of the mutant strain M18RAG was significantly increased, about 1.5-2 times that of the wild-type strain. relA gene trans-complementation assay and phzA ’-’ lacZ translational fusion assay further demonstrated that RelA has an inhibitory effect on PCA biosynthesis and its gene expression.