假单胞菌M18调控基因ppbR的克隆及功能研究

Construction and Characterization of a ppbR gene Mutant of Pseudomonas sp. M18

摘要:

假单胞菌 (Pseudomonas sp.) M18 是促进植物生长的根际细菌, 能产生吩嗪-1-羧酸 (PCA) 和藤黄绿菌素 (Plt) 两种不同的抗生素。根据生物信息学分析, 铜绿假单胞菌PA2572基因编码蛋白可能是一个双元调控系统的应答调节子。本研究从假单胞菌M18基因组中扩增出PA2572同源基因片段ppbR, 利用体外定点插入突变和同源重组技术构建了M18 的ppbR突变株M18P。研究结果表明, 突变株M18P在泳动能力和群集运动能力上有显著的下降。突变株合成PCA 的能力比野生型有显著的下降, 在发酵液中PCA积累量仅为野生型的50%。在KMB培养基中, 突变株Plt的积累量和野生型没有显著的差异。

Abstract:

Pseudomonas sp. M18, one of plant-growth-promoting rhizobacteria, can produce secondary metabolites including phenazine-1-carboxylic acid (PCA) and pyoluteorin (Plt). PA2572 gene coding protein is a probable two-component response regulator in Pseudomonas according to homologous speculations. In order to investigate its genetic function, PA2572 homologous gene, ppbR, was amplified from M18 genome, inactivated by inserting a Gm cassette. The resulting reconstruct was introduced into the M18 genome using homologous recombination technique, so as to obtain the null mutant M18P. The results showed that the M18P has less flagellar swimming and swarming motility, and yielded fewer PCA. The production of PCA was only 50% of the wild type. However, there was no remarkable difference between mutant and wild type in producing pyoluteorin in KMB medium.