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An Ecdysone Receptor-based Singular Gene Switch for Deliberate Expression of Transgene with Robustness, Reversibility, and Negligible Leakiness
Published on: May 7, 2018
Development of a methoxyfenozide-responsive gene switch for applications in plants
Venkata S Tavva1, Randy D Dinkins, Subba R Palli
1Plant and Soil Sciences Department, 1405 Veterans Road, University of Kentucky, Lexington, KY 40546-0312, USA.
The Plant Journal : for Cell and Molecular Biology
|January 18, 2006
Summary
Scientists developed a novel two-hybrid gene switch using the ecdysone receptor (EcR) for enhanced transgene regulation in plants. This improved system achieves nanomolar sensitivity, significantly advancing gene switch technology for agricultural and research applications.
Area of Science:
- Molecular Biology
- Plant Biotechnology
- Gene Regulation
Background:
- Ecdysone receptor (EcR)-based gene switches are used for conditional transgene expression.
- Existing plant EcR gene switches are monopartate and require high ligand concentrations (micromolar), limiting their practical application.
- There is a need for more sensitive and efficient gene switch systems in plants.
Purpose of the Study:
- To develop a highly sensitive two-hybrid gene switch system based on the Choristoneura fumiferana ecdysone receptor (CfEcR).
- To improve the sensitivity of EcR-based gene switches from micromolar to nanomolar concentrations of the ligand methoxyfenozide.
- To evaluate the efficacy of the novel gene switch in various plant species and protoplasts.
Main Methods:
- Constructed two-hybrid receptor systems by fusing CfEcR DEF domains to GAL4 DNA-binding domain and retinoid X receptor (RXR) EF domains to VP16 activation domain.
- Tested constructs using a luciferase reporter gene under the control of GAL4 response elements and a minimal promoter.
- Evaluated gene switch performance in tobacco, corn, and soybean protoplasts, as well as in transgenic Arabidopsis and tobacco plants.
Main Results:
- The two-hybrid CfEcR gene switch system successfully regulated transgene expression in plants.
- The sensitivity of the gene switch was significantly enhanced, responding to nanomolar concentrations of methoxyfenozide.
- The CfEcR + Locusta migratoria RXR (LmRXR) two-hybrid switch demonstrated 25 to 625 times greater sensitivity compared to monopartate systems, varying by plant species.
Conclusions:
- The developed two-hybrid CfEcR gene switch offers a substantial improvement in sensitivity for transgene regulation in plants.
- Methoxyfenozide, a registered pesticide, serves as an effective and safe ligand for this enhanced gene switch system.
- This technology holds promise for advanced applications in plant biotechnology, research, and agriculture requiring precise gene control.
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