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Comparison of Tobacco Host Cell Protein Removal Methods by Blanching Intact Plants or by Heat Treatment of Extracts
Published on: August 8, 2016
Cre-mediated seed-specific transgene excision in tobacco
L Kopertekh1, K Schulze, A Frolov
1Julius Kuehn Institute, Federal Research Centre for Cultivated Plants (JKI), Institute for Biosafety of Genetically Modified Plants, Erwin-Baur-Str 27, 06484 Quedlinburg, Germany.
Plant Molecular Biology
|January 16, 2010
Summary
Marker-free transgenic plants were created using a seed-specific Cre-lox system to express valuable phenolic compounds. This efficient method allows for the production of trans-resveratrol and piceid in plant seeds.
Area of Science:
- Plant Biotechnology
- Molecular Biology
- Genetics
Background:
- Transgenic plants offer a promising platform for producing valuable compounds.
- Eliminating marker genes from transgenic events is crucial for commercial applications and regulatory approval.
- Developing efficient and precise methods for marker gene removal is an ongoing challenge in plant biotechnology.
Purpose of the Study:
- To develop a marker-free transformation strategy in tobacco plants.
- To enable seed-specific expression of pharmacologically valuable phenolic compounds.
- To demonstrate the efficiency of a Cre-lox recombination system for marker gene excision in plants.
Main Methods:
- Simultaneous cotransformation of tobacco with Cre-recombinase and lox-target constructs.
- Utilizing a seed-specific napin promoter to control Cre recombinase expression.
- Employing the Cre-lox system for site-specific excision of the selectable marker gene (bar).
- Analyzing T1 progeny for DNA excision and expression of the target gene (vstI) using genetic, molecular, and RP-HPLC methods.
Main Results:
- Successful generation of marker-free transgenic tobacco lines through Cre-lox mediated excision.
- Demonstrated efficient DNA excision in all tested transgenic lines.
- Confirmed the accumulation of trans-resveratrol and piceid in the seeds of marker-free plants.
- Showed that marker gene excision did not impede the expression of the pharmacologically valuable gene.
Conclusions:
- A developmentally controlled Cre-lox system provides an efficient strategy for generating marker-free transgenic plants.
- This approach facilitates the seed-specific production of high-value phenolic compounds like trans-resveratrol and piceid.
- The method is highly efficient and applicable for producing pharmacologically important compounds in plants.
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