In vivo gene expression of Pseudomonas putida KT2440 in the rhizosphere of different plants.
Bottom Line: Using the IVET technology we investigated which KT2440 genes were expressed in the rhizosphere of four different plants: pine, cypress, evergreen oak and rosemary.Another 40 fusions were found to correspond to likely promoters that encode antisense RNAs of unknown function, some of which were isolated as fusions from the bacteria recovered in the rhizosphere from all of the plants, while others were specific to one or several of the plants.The results obtained in this study suggest that plant-specific signals are sensed by KT2440 in the rhizosphere and that the signals and consequent gene expression are related to the bacteria's successful establishment in this niche.
Affiliation: Bio-Iliberis Research and Development, I+D Department, 18210, Peligros, Granada, Spain.Show MeSH
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Mentions: In this study we aimed to document which genes are specifically expressed in the rhizosphere of the four plants: P. putida Δasd bearing the IVET library was inoculated as above and the plants were placed in pots with peat:sand (1:1) as substrate. We performed two independent rounds of IVET selection, with each plant tested in triplicate on each round. All plants were kept for 2 months under greenhouse conditions before recovering the viable bacterial cells on selective medium. Near 50 000 isolated after spreading serial dilutions in minimal medium agar plates with X-gal, kanamycin, DAP, lysine, methionine and threonine. Around 800 of them (1.6% of the total) formed white or very pale-blue colonies; and were unable to grow in the absence of DAP. This was indicative that the promoter that drove asd expression was expressed in the soil and not in the agar plates. For each clone the pOR1 derivative was recovered (Rainey et al., 1997) and the insert sequenced. Computational analysis of the sequences allowed us to identify a total of 79 different insertions; 39 of which were adjacent to annotated genes (Table 1), whereas the remaining 40 were antisense transcriptional clones (Table S1). From the group of 39 transcriptional fusions, six of them were isolated from the rhizosphere of all of the plants tested, while 11 were isolated from more than one type of plant, and the remaining 22 transcriptional fusions were recovered from the rhizosphere of only one of the four plants tested; specifically, three of them only in cypress, nine in evergreen oak, six in pine and four in rosemary (Fig. 2).
Affiliation: Bio-Iliberis Research and Development, I+D Department, 18210, Peligros, Granada, Spain.