A modular steroid-inducible gene expression system for use in rice
Daniela Vlad1, Basel Abu-Jamous2, Peng Wang3
1Department of Plant Sciences, University of Oxford, South Parks Rd, Oxford, OX1 3RB, UK.
BMC Plant Biology
|October 17, 2019
Summary
A new dexamethasone (DEX)-inducible gene expression system was optimized for rice, showing high sensitivity and tight regulation. Off-target effects were observed at higher concentrations but could be minimized with isopropyl β-D-1-thiogalactopyranoside (IPTG).
Area of Science:
- Plant molecular biology
- Genetics and genomics
- Biotechnology
Background:
- Chemically inducible systems are crucial for controlling gene expression in plants.
- Such systems offer spatial and temporal regulation but are underutilized in monocots like rice.
Purpose of the Study:
- To develop and evaluate a monocot-optimized dexamethasone (DEX)-inducible gene expression system (pOp6/LhGR) in rice.
- To assess the system's efficacy, sensitivity, and potential off-target effects.
Main Methods:
- Utilized Golden Gate modular cloning to construct the pOp6/LhGR system.
- Tested the system in transgenic rice callus and seedlings using the β-glucuronidase (GUS) reporter enzyme.
- Investigated dose-dependent effects of DEX and mitigation strategies using isopropyl β-D-1-thiogalactopyranoside (IPTG).
Main Results:
- Achieved tight regulation and high sensitivity to DEX, with significant GUS activity detected after 6 hours in callus and at 0.01 μM DEX in seedling roots.
- Observed severe developmental defects in transgenic plants at higher DEX concentrations.
- Identified potential non-specific gene activation due to rice genome similarity to the LhGR target sequence, which was reduced by IPTG treatment.
Conclusions:
- The pOp6/LhGR system is effective for gene expression control in rice.
- Careful optimization of DEX application and appropriate controls are necessary for specific applications.
- IPTG can be used to minimize off-target effects, enhancing the system's utility.
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