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A novel salt-inducible CrGPDH3 promoter of the microalga Chlamydomonas reinhardtii for transgene overexpression

Abstract

The expression of transgenes in the nucleus is an attractive alternative for the expression of recombinant proteins in the green microalga Chlamydomonas reinhardtii. For this purpose, a strong inducible promoter that allows protein accumulation without possible negative effects on cell multiplication and biomass accumulation is desirable. A previous study at our laboratory identified that the CrGPDH3 gene from C. reinhardtii was inducible under NaCl treatments. In this study, we cloned and characterized a 3012 bp sequence upstream of the start codon of the CrGPDH3 gene, including the 285 bp 5′ untranslated region. This region was identified as the full-length promoter and named PromA (− 2727 to + 285). Deletion analysis of PromA using GUSPlus as a reporter gene enabled us to identify PromC (− 653 to + 285) as the core promoter, displaying basal expression. A region named RIA1 (− 2727 to − 1672) was suggested to contain the NaCl response elements. Moreover, deletion analysis of RIA1 enabled us to identify a region of 577 bp named RIA3 (− 2727 to − 2150) that, when cloned upstream of PromC, was able to drive the expression of GUSPlus in response to 5 and 100 mM NaCl, and 100 mM KCl, similar to the native CrGPDH3 promoter. These results expand our understanding of the transcriptional mechanism of CrGPDH3 and clearly show that CrGPDH3 promoter and its chimeric forms are highly salt-inducible and can be used as inducible promoters for the overexpression of transgenes in C. reinhardtii.

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Acknowledgements

The authors would like to thank the editor and reviewers of this manuscript for their critical review and their valuable comments. The authors thank Edanz (www.edanzediting.com) for editing the English text of a draft of this manuscript.

Funding

This research was funded by Consejo Nacional de Ciencia y Tecnologia (CONACYT, Mexico) (Grant number 169217). Anayeli Guadalupe Beltran-Aguilar received funding from CONACYT (Scholarship number 426890).

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VAH-V and SP-E conceived the study. VAH-V, SP-E, and LAL-O designed the experiments. AGB-A and ICB-A performed the experiments. AGB-A, VAH-V, SP-E, and LAL-O analyzed the results. AGB-A and VAH-V wrote the manuscript.

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Correspondence to Virginia Aurora Herrera-Valencia.

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The authors declare that they have no conflict of interest.

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This article does not contain any studies with human participants or animals performed by any of the authors.

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Beltran-Aguilar, A.G., Peraza-Echeverria, S., López-Ochoa, L.A. et al. A novel salt-inducible CrGPDH3 promoter of the microalga Chlamydomonas reinhardtii for transgene overexpression. Appl Microbiol Biotechnol 103, 3487–3499 (2019). https://doi.org/10.1007/s00253-019-09733-y

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  • DOI: https://doi.org/10.1007/s00253-019-09733-y

Keywords

  • Chlamydomonas reinhardtii
  • Inducible promoter
  • Chimeric promoter
  • Recombinant protein