Construction of Pseudomonas sp. M18 relA mutant and its regulation on phenazine-1-carboxylic acid biosynthesis

Abstract:

The rhizosphere bacterium Pseudomonas sp. M18 can simultaneously produce two antibiotics: phenazine-1- carboxylic acid (PCA) and pyoluteorin. ppGpp, which is synthesized by RelA, can mediate bacterial stringent response to nutritional starvation. The relA gene was PCR amplified from the M18 strain chromosomal DNA template. The relA mutant of M18 strain (M18RAG) was constructed through inserted inactivation of gentamicin resistance cassette and homologous recombination. PCA production was assayed in PPM media. It was showed that the relA mutation resulted in a significant enhancement of PCA production. PCA production of M18RAG was about 1.5 to 2 times as much as that of the wild-type strain. The negative regulation of RelA on PCA biosynthesis and its gene expression was further confirmed by the trans complementation test of relA gene and the expression analysis of phzA'-'lacZ translational fusion.