Exploring the Impact of Terminators on Transgene Expression in <i>Chlamydomonas reinhardtii</i> with a Synthetic Biology Approach.
Scaife, Mark A
Mordaka, Paweł M
Tomsett, Eleanor V
Smith, Alison G
MetadataShow full item record
Geisler, K., Scaife, M. A., Mordaka, P. M., Holzer, A., Tomsett, E. V., Mehrshahi, P., Mendoza Ochoa, G. I., & et al. (2021). Exploring the Impact of Terminators on Transgene Expression in <i>Chlamydomonas reinhardtii</i> with a Synthetic Biology Approach.. https://doi.org/10.3390/life11090964
<i>Chlamydomonas reinhardtii</i> 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 <i>C. reinhardtii</i> 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.
Algae, Terminator, Heterologous protein expression, Modular Assembly, 3′ Utr, Standardized Workflow, Quantification Of Transgene Expression
Biotechnology and Biological Sciences Research Council (BB/M018180/1, BB/I007660/1, BB/L002957/1, BB/L014130/1, BB/R01860X/1)
Bill and Melinda Gates Foundation (OPP1144)
Seventh Framework Programme (311956)
External DOI: https://doi.org/10.3390/life11090964
This record's URL: https://www.repository.cam.ac.uk/handle/1810/330081
Attribution 4.0 International
Licence URL: https://creativecommons.org/licenses/by/4.0/