PT  - JOURNAL ARTICLE
AU  - Katrin Geisler
AU  - Mark A. Scaife
AU  - Paweł M. Mordaka
AU  - Andre Holzer
AU  - Payam Mehrshahi
AU  - Gonzalo Mendoza Ochoa
AU  - Alison G. Smith
TI  - Exploring the impact of terminators on transgene expression in <em>Chlamydomonas reinhardtii</em> with a synthetic biology approach
AID  - 10.1101/2021.08.04.455025
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.08.04.455025
4099  - http://biorxiv.org/content/early/2021/08/04/2021.08.04.455025.short
4100  - http://biorxiv.org/content/early/2021/08/04/2021.08.04.455025.full
AB  - 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 StatementThe authors have declared no competing interest.