Vector for chromosomal integration of the phoC gene in plant growth-promoting bacteria

Abstract

This work describes the subcloning of the gene encoding the PhoC acid phosphatase from Morganella morganii (phoC gene) in a vector that permits stable chromosomal integration of this gene in plant growth-promoting bacteria (PGPB). A plasmid was constructed using the suicide delivery vector pJMT6 (a pUT/mini Tn5 derivative vector) and the plasmid pLR1, the latter harboring the phoC gene. The recombinant construction pLF17, which contains a non-antibiotic resistance selection marker, was transformed and expressed in Escherichia coli CC118λpir. A transformant clone, E. coli CC118λpir F17 was selected and further characterized, showing phoC gene expression through an histochemical assay and zymograms developed to detect phosphatase activity. With this technique, it was possible to detect, in the whole cell extract, the 25-kDa polypeptidic component responsible for acid phosphatase activity. Acid phosphatase activity was quantified in the whole cell and in the supernatant of the culture as being higher in the transformant E. coli CC118λpir F17 than in E. coli CC118λpir without plasmids along the cultivation time.