Novel cis-regulatory elements as synthetic promoters to drive recombinant protein expression from the Chlamydomonas reinhardtii nuclear genome

McQuillan, Josie L.; Berndt, Anthony J.; Sproles, Ashley E.; Mayfield, Stephen P.; Pandhal, Jagroop

doi:10.1016/j.nbt.2022.01.001

Title: Novel cis-regulatory elements as synthetic promoters to drive recombinant protein expression from the Chlamydomonas reinhardtii nuclear genome

Journal Article · 02 January 2022 · New Biotechnology

DOI: https://doi.org/10.1016/j.nbt.2022.01.001 · OSTI ID:1853821

^[1];

^[2]; Sproles, Ashley E. ^[2]; Mayfield, Stephen P. ^[2];

^[3]

Univ. of Sheffield (United Kingdom); University of California, San Diego
Univ. of California, San Diego, CA (United States)
Univ. of Sheffield (United Kingdom)

Eukaryotic green microalgae represent a sustainable, photosynthetic biotechnology platform for generating high-value products. The model green alga Chlamydomonas reinhardtii has already been used to generate high value bioproducts such as recombinant proteins and terpenoids. However, low, unstable, and variable nuclear transgene expression has limited the ease and speed of metabolic engineering and recombinant protein expression in this system. Here, novel genetic devices for transgene expression in C. reinhardtii have been developed by identifying cis-regulatory DNA elements capable of driving high transgene expression in C. reinhardtii promoters using de novo motif discovery informatics approaches. Thirteen putative motifs were synthesized as concatemers, linked to a common minimal basal promoter, and assayed for their activity to drive expression of a yellow fluorescent protein reporter gene. Following transformation of the vectors into C. reinhardtii by electroporation, in vivo measurements of yellow fluorescent protein expression by flow cytometry revealed that five of the DNA motifs analyzed displayed significantly higher reporter expression compared to the basal promoter control. Two of the concatemerized motifs, despite being much smaller minimal cis-regulatory elements, drove reporter expression at levels approaching that of the conventionally-used AR1 promoter. This analysis provides insight into C. reinhardtii promoter structure and gene regulation, and provides a new toolbox of cis-regulatory elements that can be used to drive transgene expression at a variety of expression levels.

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Research Organization:: Univ. of California, San Diego, CA (United States)

Sponsoring Organization:: Biotechnology and Biological Sciences Research Council (BBSRC); Engineering and Physical Sciences Research Council (EPSRC); USDOE Office of Energy Efficiency and Renewable Energy (EERE), Transportation Office. Bioenergy Technologies Office

Grant/Contract Number:: EE0008246

OSTI ID:: 1853821

Journal Information:: New Biotechnology, Journal Name: New Biotechnology Vol. 68; ISSN 1871-6784

Publisher:: ElsevierCopyright Statement

Country of Publication:: United States

Language:: English

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59 BASIC BIOLOGICAL SCIENCES
Algal biotechnology
Chlamydomonas reinhardtii
Metabolic engineering
Recombinant protein expression
Synthetic promoters
de novo motif discovery

Title: Novel cis-regulatory elements as synthetic promoters to drive recombinant protein expression from the Chlamydomonas reinhardtii nuclear genome

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