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Whole Cell Bacterial Biosensor for Glutamine and Applications to Plants and Microbes

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Title: Whole Cell Bacterial Biosensor for Glutamine and Applications to Plants and Microbes
Author: Tessaro, Michael
Department: Department of Plant Agriculture
Program: Plant Agriculture
Advisor: Raizada, Manish
Abstract: Glutamine (Gln) is a critical intermediate in nitrogen metabolism in all organisms. Here, a whole cell biosensor (GlnLux) for Gln was constructed by transforming a bacterial Gln auxotroph with a constitutive lux reporter. The biosensor was optimized for sensitivity, linearity, efficiency, specificity and robustness to permit detection of Gln in vitro and in vivo. The optimized GlnLux biosensor achieved nanomolar sensitivity with Gln standards. Extracts from only 1 mg of maize (Zea mays L.) leaf tissue were sufficient for Gln detection by GlnLux. Measurements of Gln in leaf extracts by GlnLux correlated with quantification by high performance liquid chromatography (Spearman r = 0.95). GlnLux permitted indirect in planta imaging of Gln using a CCD camera, enabling identification of plants that had been fertilized with nitrogen. Imaging using GlnLux also resolved predicted spatial differences in leaf Gln concentration. In a second application, it was demonstrated that GlnLux embedded into agar permits non-destructive screening of co-inoculated bacterial colonies for biological nitrogen fixation (BNF). GlnLux agar was able to distinguish a Bradyrhizobium japonicum wild type strain (nif+) from a mutant strain defective in nitrogenase (nif-) following ≥8 h of co-incubation. The technology was used to screen a bacterial endophyte diversity library cultured from Zea mays (L.) seeds for biological nitrogen fixation.
URI: http://hdl.handle.net/10214/3311
Date: 2012-01


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http://creativecommons.org/licenses/by/2.5/ca/ Except where otherwise noted, this item's license is described as http://creativecommons.org/licenses/by/2.5/ca/