Site-Specific Recombination-Based Genetic System for Reporting Transient or Low-Level Gene Expression

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2002-07-01
Authors
Casavant, N. Carol
Phillips, Gregory
Halverson, Larry
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Beattie, Gwyn
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Microbiology
Microbiology allows you to learn about the microorganisms that affect us every day and how they interact with their surroundings. Through the program, you will be equipped with the knowledge to work in areas related to agriculture, the environment and medicine.
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Agronomy

The Department of Agronomy seeks to teach the study of the farm-field, its crops, and its science and management. It originally consisted of three sub-departments to do this: Soils, Farm-Crops, and Agricultural Engineering (which became its own department in 1907). Today, the department teaches crop sciences and breeding, soil sciences, meteorology, agroecology, and biotechnology.

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The Department of Agronomy was formed in 1902. From 1917 to 1935 it was known as the Department of Farm Crops and Soils.

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1902–present

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  • Department of Farm Crops and Soils (1917–1935)

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Abstract

We report here the construction, characterization, and application of a plasmid-based genetic system that reports the expression of a target promoter by effecting an irreversible, heritable change in a bacterial cell. This system confers strong repression of the reporter gene gfp in the absence of target promoter expression and utilizes the site-specific recombination machinery of bacteriophage P22 to trigger high-level reporter gene expression in the original cell and its progeny after target gene induction. We demonstrate the effectiveness of this genetic system by tailoring it to indicate the availability of arabinose to the biological control agent Enterobacter cloacae JL1157 in culture and in the barley rhizosphere. The presence of bioavailable arabinose triggered the production of P22 excisionase and integrase from the reporter plasmid pAraLHB in JL1157, and this led to excision of the cI repressor gene, which is flanked by att sites, and the subsequent irreversible expression of gfp in the original cell and in its progeny. In culture, nearly 100% of an E. cloacae JL1157(pAraLHB) population expressed gfpafter exposure to 6.5 to 65 μM arabinose for 3 h. We used this biosensor to demonstrate that arabinose was released from the seeds of several legumes and grass species during germination and from roots of barley seedlings grown hydroponically or in soil. When introduced into microcosms containing barley, the biosensor permitted the localization of arabinose along the roots. Arabinose was present near the root-seed junction and on the seminal roots but was not detected at the root tips. This recombination-based reporter system should be useful for monitoring bacterial exposure to transient or low levels of specific molecules directly in the environment.

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This is an article from Applied and Environmental Microbiology 68 (2002): 3588, doi: 10.1128/AEM.68.7.3588–3596.2002. Posted with permission.

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Tue Jan 01 00:00:00 UTC 2002
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