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Exploring the impact of terminators on transgene expression in Chlamydomonas reinhardtii with a synthetic biology approach

Katrin Geisler, Mark A. Scaife, View ORCID ProfilePaweł M. Mordaka, Andre Holzer, Payam Mehrshahi, Gonzalo Mendoza Ochoa, Alison G. Smith
doi: https://doi.org/10.1101/2021.08.04.455025
Katrin Geisler
1Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK
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Mark A. Scaife
1Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK
2Mara Renewables Corporation, Dartmouth, Nova Scotia, Canada
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Paweł M. Mordaka
1Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK
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  • ORCID record for Paweł M. Mordaka
Andre Holzer
1Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK
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Payam Mehrshahi
1Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK
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Gonzalo Mendoza Ochoa
1Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK
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Alison G. Smith
1Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK
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  • For correspondence: as25@cam.ac.uk
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Abstract

Chlamydomonas reinhardtii has many attractive features for use as a model organism for both fundamental studies and as a biotechnological platform. Nonetheless, despite the many molecular tools and resources that have been developed, there are challenges for its successful engineering, in particular to obtain reproducible and high levels of transgene expression. Here we describe a synthetic biology approach to screen several hundred independent transformants using standardised parts to explore different parameters that might affect transgene expression. We focused on terminators and, using a standardised workflow and quantitative outputs, tested 9 different elements representing three different size classes of native terminators to determine their ability to support high level expression of a GFP reporter gene. We found that the optimal size reflected the median size of element found in the C. reinhardtii genome. The behaviour of the terminator parts was similar with different promoters, in different host strains and with different transgenes. This approach is applicable to the systematic testing of other genetic elements, facilitating comparison to determine optimal transgene design.

Competing Interest Statement

The authors have declared no competing interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted August 04, 2021.
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Exploring the impact of terminators on transgene expression in Chlamydomonas reinhardtii with a synthetic biology approach
Katrin Geisler, Mark A. Scaife, Paweł M. Mordaka, Andre Holzer, Payam Mehrshahi, Gonzalo Mendoza Ochoa, Alison G. Smith
bioRxiv 2021.08.04.455025; doi: https://doi.org/10.1101/2021.08.04.455025
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Exploring the impact of terminators on transgene expression in Chlamydomonas reinhardtii with a synthetic biology approach
Katrin Geisler, Mark A. Scaife, Paweł M. Mordaka, Andre Holzer, Payam Mehrshahi, Gonzalo Mendoza Ochoa, Alison G. Smith
bioRxiv 2021.08.04.455025; doi: https://doi.org/10.1101/2021.08.04.455025

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