Dataset Information

Exploring the Impact of Terminators on Transgene Expression in Chlamydomonas reinhardtii with a Synthetic Biology Approach.

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.

SUBMITTER: Geisler K

PROVIDER: S-EPMC8471596 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

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Exploring the Impact of Terminators on Transgene Expression in <i>Chlamydomonas reinhardtii</i> with a Synthetic Biology Approach.

Geisler Katrin K Scaife Mark A MA Mordaka Paweł M PM Holzer Andre A Tomsett Eleanor V EV Mehrshahi Payam P Mendoza Ochoa Gonzalo I GI Smith Alison G AG

Life (Basel, Switzerland) 20210914 9

<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 ex ...[more]

PMID: 34575113

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Project description:BackgroundIonizing radiation has been used for mutagenesis or material modification. The potential to use microalgae as a platform for antimicrobial production has been reported, but little work has been done to advance it beyond characterization to biotechnology. This study explored two different applications of ionizing radiation as a metabolic remodeler and a molecular modifier to enhance the antimicrobial activity of total protein and solvent extracts of Chlamydomonas reinhardtii cells.ResultsFirst, highly efficient transgenic C. reinhardtii strains expressing the plant-derived antimicrobial peptides, AtPR1 or AtTHI2.1, were developed using the radiation-inducible promoter, CrRPA70Ap. Low transgene expression was significantly improved through X-irradiation (12-50 Gy), with peak activity observed within 2 h. Protein extracts from these strains after X-irradiation showed enhanced antimicrobial activity against the prokaryotic bacterium, Pseudomonas syringae, and the eukaryotic fungus, Cryptococcus neoformans. In addition, X-irradiation (12 Gy) increased the growth and biomass of the transgenic strains. Second, C. reinhardtii cell extracts in ethanol were γ-irradiated (5-20 kGy), leading to molecular modifications and increased antimicrobial activity against the phytopathogenic bacteria, P. syringae and Burkholderia glumae, in a dose-dependent manner. These changes were associated with alterations in fatty acid composition. When both transgenic expression of antimicrobial peptides and molecular modification of bioactive substances were applied, the antimicrobial activity of C. reinhardtii cell extracts was further enhanced to some extent.ConclusionOverall, these findings suggest that ionizing radiation can significantly enhance the antimicrobial potential of C. reinhardtii through efficient transgene expression and molecular modification of bioactive substances, making it a valuable source of natural antimicrobial agents. Ionizing radiation can act not only as a metabolic remodeler of transgene expression in microalgae but also as a molecular modifier of the bioactive substances.

Dataset Information

Exploring the Impact of Terminators on Transgene Expression in Chlamydomonas reinhardtii with a Synthetic Biology Approach.

Publications

Exploring the Impact of Terminators on Transgene Expression in <i>Chlamydomonas reinhardtii</i> with a Synthetic Biology Approach.

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Dataset Information

Exploring the Impact of Terminators on Transgene Expression in Chlamydomonas reinhardtii with a Synthetic Biology Approach.

Publications

Exploring the Impact of Terminators on Transgene Expression in &lt;i&gt;Chlamydomonas reinhardtii&lt;/i&gt; with a Synthetic Biology Approach.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Exploring the Impact of Terminators on Transgene Expression in <i>Chlamydomonas reinhardtii</i> with a Synthetic Biology Approach.