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Phenotype MicroArrays used in the study of two-component systems regulating carbon and nitrogen metabolism in Pseudomonas

  • Regulation of carbon and nitrogen utilization by CbrAB and NtrBC two-component systems in Pseudomonas aeruginosa. Li W, Lu CD. Department of Biology, Georgia State University, Atlanta, GA 30303, USA. In this study, PM1, PM2, and PM3 were used to characterize carbon and nitrogen metabolism of a cbrAB deletion mutant and it's supressors (ntrB alleles). Combining PM studies with activity profile data for key enzymes, the authors were able to observe clear metabolic phenotypes and their partial reversal by suppressors. "This study supports the notion that CbrAB and NtrBC form a network to control the C/N balance in P. aeruginosa."

  • Dual Involvement of CbrAB and NtrBC in the Regulation of Histidine Utilization in Pseudomonas fluorescens SBW25. Zhang XX, Rainey PB. Institute of Molecular Biosciences and New Zealand Institute for Advanced Study, Massey University, Auckland, New Zealand. Effects on transcription and metabolism were reported for cbrB, ntrC, and cbrB ntrC double mutants. The authors found metabolic deficiencies in the double mutant that were not present in either single mutant indicating redundancy for these functions. This data is used to "show that NtrC is involved in nitrogen metabolism and that CbrB is primarily a regulator of carbon-source utilization; however, CbrB is also involved in the utilization of nitrogen substrates-and in some cases utilisation of substrates as both carbobn and nitrogen sources".



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    Brief description of PM technology

    Phenotype MicroArray™ technology tests cellular phenotypes. Included in the tests are assays of basic cellular nutritional pathways for C, N, P, and S metabolism, osmotic and pH and sensitivity, and sensitivity to chemical agents.

    The most common application is to assess the phenotypic effects of mutations. A change in genotype of a cell should lead to one or more changes in phenotype. PMs allow testing of knock-out or knock-in mutants to help discern the biological changes that occur consequent to genetic changes.

    Another common application is phenotypic characterization of a collection of related strains. For example, it is possible to determine the phenotypic relatedness of a collection of isolates of a given species. PM analysis has been successfully implemented for a wide variety of model microbial cells including Escherichia coli,

    Salmonella enterica, Pseudomonas aeruginosa, Pseudomonas putida, Sinorhizobium meliloti, Saccharomyces cerevisiae, Bacillus subtilis, Bacillus cereus, Shewanella oneidensis, Proteus mirabilis, and many other species. In the last year, Biolog has successfully developed universal protocols that allow us to test most species of interest. Contact us for details.

    Phenotype MicroArray™ technology uses Biolog's OmniLog® instrument, which is also used for microbial identification. The OmniLog® automatically incubates, reads and interprets the Biolog Phenotype MicroArray MicroPlates™. It continuously processes samples but allows

    the user complete access at any time during a sample run. Samples can be loaded when ready and removed when complete without disturbing other samples still in-process. Inside the Reader there are 25 trays. Each tray holds 2 MicroPlates, giving the Reader a total capacity to incubate and read 50 MicroPlates. Before the user inoculates the appropriate MicroPlates, they log the MicroPlate information into the OmniLog® software.

    By simply following the software's instructions, the user then opens the door of the OmniLog® Reader and places each MicroPlate in the appropriate tray slot indicated by the software. Once all the MicroPlates are loaded and the door of the Reader is closed, the OmniLog® software takes over the responsibility for incubating, reading, saving, and printing the results. The OmniLog® PM System utilizes Windows-based software with an elegantly simple user interface. The status of all the samples can be observed by simply looking at the Read menu screen of the OmniLog® PM System software. All of the information for a specific MicroPlate is contained on a single line of this screen. Once the system has completed a specific MicroPlate it indicates this status with a check mark icon. A clock icon is used for those samples that are still incubating

    because a result has not yet been determined. The OmniLog® PM System's software is extremely flexible and can be integrated into virtually any laboratory's workflow. New MicroPlates can be entered into the Reader and MicroPlates that have already completed can be removed from the Reader at almost any time. This allows the user to either perform experiments one at a time or batch them.

    PM Technology and the OmniLog® instrument are ideal for a core facility or for purchase under a shared equipment grant.

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