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United States Department of Agriculture

Agricultural Research Service

Title: Two-Hybrid Gene Switch: A Chemical Inducible Ecdysone Receptor (ECR) - Based Gene Regulation System

Authors
item Tavva, Venkata - UNIVERSITY OF KENTUCKY
item Dinkins, Randy
item Palli, Subba - UNIVERSITY OF KENTUCKY
item Collins, Glenn - UNIVERSITY OF KENTUCKY

Submitted to: Biennial Conference on Molecular and Cellular Biology of the Soybean
Publication Type: Abstract Only
Publication Acceptance Date: June 20, 2004
Publication Date: August 11, 2004
Citation: Tavva, V.S., Dinkins, R.D., Palli, S.R., Collins, G.B. 2004. Two-Hybrid Gene Switch: A Chemical Inducible Ecdysone Receptor (ECR) - Based Gene Regulation System. Biennial Conference on Molecular and Cellular Biology of the Soybean. page 137, abstract 85.

Interpretive Summary: Expression of transgenes is becoming a powerful tool in many biotechnology applications including food product improvement. Constitutive promoters, those that are expressed all the time in all tissues, are presently the primary means used to express transgenes in plants. Metabolic energy waste, negative pleiotropic effects and potential gene escape are some of the disadvantages associated with the use of constitutive promoters. To counter these problems, gene switches that can regulate the expression of transgenes through hormone analogs and antibiotics have been developed. However, most of the compounds that are used to induce gene expression are impractical or inappropriate for regulation of transgenes for large-scale use in green houses and in the field. We have developed a gene regulation system to control expression of multiple genes simultaneously and independently in the same plant by using chemicals for regulation of transgenes for large-scale production and that are presently commercially available and registered for field use. DNA vectors testing the utility of the system have been introduced into soybean, as well as corn and tobacco protoplasts and verified.

Technical Abstract: Chemical inducible gene regulation systems that activate or inactivate transgene expression have many potential applications in the basic understanding of gene function in plants. We have developed a two-hybrid EcR-based gene switch that regulates the expression of a reporter gene (luciferase) placed under the control of a minimal promoter and 5X GAL4 response elements. The two receptor constructs along with the reporter construct were electroporated into protoplasts isolated from cell suspension cultures of soybean (W82), as well as corn (BMS) and tobacco (Xanthi). The electroporated protoplasts were exposed to varying concentrations of methoxyfenozide and luciferase activity was measured after 24 h. Based on transient expression studies, the two-hybrid gene switch was found to be more sensitive with lower background and higher induction and expression of the reporter gene compared to the single format gene switch.

Last Modified: 9/1/2014
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