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Optimizing Promoters and Subcellular Localization for Constitutive Transgene Expression in Marchantia polymorpha
Sze Wai Tse1, Davide Annese1, Facundo Romani1
1Department of Plant Sciences, University of Cambridge, Cambridge CB2 3EA, UK.
Plant & Cell Physiology
|June 1, 2024
Summary
We benchmarked constitutive promoters in Marchantia polymorpha for synthetic biology. New promoters offer high gene expression across tissues without growth penalties, expanding genetic tools for bioproduction.
Area of Science:
- Plant molecular biology
- Synthetic biology
- Biotechnology
Background:
- Marchantia polymorpha is a key model organism for comparative studies and synthetic biology.
- A robust genetic parts toolbox is essential for predictable heterologous gene expression in Marchantia for bioproduction.
- Existing constitutive promoters in Marchantia require benchmarking to assess their utility.
Purpose of the Study:
- To systematically characterize and compare constitutive promoters for gene expression in Marchantia polymorpha.
- To identify promoters that enable high-level gene expression without negatively impacting plant growth.
- To expand the available genetic tools for researchers in the Marchantia community.
Main Methods:
- Comparison of expression levels from previously characterized and novel constitutive promoters.
- Evaluation of promoter performance across different tissues and subcellular compartments.
- Assessment of plant growth and biomass accumulation as indicators of promoter utility.
- Demonstration of promoter function by expressing a polycistronic betalain synthesis gene cassette (RUBY).
Main Results:
- The cauliflower mosaic virus 35S promoter (pro35S×2) yielded the highest protein levels but inhibited growth.
- Promoters from Marchantia ETHYLENE RESPONSE FACTOR 1 and CLASS II HOMEODOMAIN-LEUCINE ZIPPER genes provided high, tissue-wide expression without growth penalties.
- Targeting heterologous proteins to the cytosol enhanced expression levels without significant growth burden.
- Successful coordinated expression of metabolic enzymes for betalain synthesis was achieved using these promoters.
Conclusions:
- New constitutive promoters have been characterized, significantly expanding the genetic toolkit for Marchantia polymorpha.
- These promoters offer predictable and tunable gene expression, crucial for synthetic biology applications and bioproduction.
- The findings provide valuable resources for advancing research and development in the Marchantia model system.
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