LuxAB Gene-marked Pseudomonas Fluorescens CP1108and Its Biological Effect On Application

Phosphate-solubilizing bacterium CP1108isolated from cotton rhizosphere was sorted to Pseudomonas fluorescens, through biochemical and16S rDNA analysis. By the method of tri-parent conjugation, the luxAB gene was introduced into CP1108. The labeled strain CP1108L had a high luminescence activity and Str、Km and Tet resistance and its transfer frequency was up to （4.65±0.01）×10^-5. After being inoculated in plates for20generations, CP1108L still had the luminescence activity and resistant to three antibiotics. By the method of alkaline lysis and electrophoresis plasmid was extracted from strain CP1108L. The results showed that the marker gene luxAB was successfully transferred into strain CP1108, and founed that the physiological and biochemical character of CP1108L had not influenced by the transferred of luxAB gene. Also the plasmid transferred had no serious effect on the growth and the survival of CP1108L in soil.We studied the survival of CP1108L in soil, and found that CP1108L successfully survived in both sterilized and non-sterilized soil, and colonizing level of CP1108L in sterile soil was higher than that of non-sterile soil, and colonized in soil with the numbers between103¹04cfu g^-142days later. Rhizoboxes were used to trace the colonization dynamics of the CP1108L around cotton root. The results showed that colonizing density of CP1108L in cotton rhizoplane was higher than that of cotton rhizosphere. The strain CP1108L colonized mainly in0⁶cm root segment in vertical direction, and decreased gradually along the root. In rhizosphere soil, the colonization density reached to maximum2.31×10⁵cfu g^-1root-soil and3.18×10⁵cfu g^-1 root, after cotton grew for14d. What is more, the colonizing density of CP1108L in0²cm root segment reached to9.08×10⁴cfu g^-1 root-soil and1.44×10⁵cfu g^-1 root respectively.A binding assay for P. fluorescens CP1108L cells adsorbed to cotton roots was developed. It consisted of incubation exposing germinating seedlings to bacterial cells in cell suspension. Stable adsorption began within the very first minutes of interaction and proceeded intensively during the first8minutes, and gradually arrived to the maximum value （1.02¹.48）×10⁸cell g^-1root. The isothermal adsorption tests showed that the adsorption curve of bacterial cells accorded perfectly with Langmuir isothermal adsorption equation. The maximum saturated adsorption value q0was calculated as4.05×10⁶cell g^-1、1.45×10⁸cell g^-1and1.19×10⁸cell g^-1. Adsorption of the P. fluorescens CP1108(10⁸cells g^-1) was about100times higher than that of E. coli DH5α (10⁶cells g^-1), and binding saturation was observed when cotton seedlings were exposed to bacterial cells up to108II cells mL^-1. The research showed that the PGPR strains have high affinity to root of cotton seedlings. Thus PGPR strains could be used as microbial inoculants and introduced into the potting experiment.Pot experiment results showed that strain CP1108L had a good advance on the plant height, root length and dry weight of cotton. Our result also showed that luxAB gene did not affect strain CP1108. The strain CP1108L could be released into soil, and used as microbial inoculants.