Development of endogenous promoters that drive high-level expression of introduced genes in the model diatom Phaeodactylum tricornutum

Yumi Watanabe; Takashi Kadono; Nozomu Kira; Kengo Suzuki; Osamu Iwata; Kouhei Ohnishi; Haruo Yamaguchi; Masao Adachi

doi:10.1016/j.margen.2018.06.003

Development of endogenous promoters that drive high-level expression of introduced genes in the model diatom Phaeodactylum tricornutum

Mar Genomics. 2018 Dec:42:41-48. doi: 10.1016/j.margen.2018.06.003. Epub 2018 Jun 28.

Authors

Yumi Watanabe¹, Takashi Kadono¹, Nozomu Kira¹, Kengo Suzuki², Osamu Iwata², Kouhei Ohnishi³, Haruo Yamaguchi¹, Masao Adachi⁴

Affiliations

¹ Laboratory of Aquatic Environmental Science, Faculty of Agriculture and Marine Science, Kochi University, Otsu-200, Monobe, Nankoku, Kochi 783-8502, Japan.
² Euglena Co., Ltd., 22F, Morinaga Plaza Building, Shiba-5-33-1, Minato-ku, Tokyo 108-0019, Japan.
³ Research Institute of Molecular Genetics, Kochi University, Otsu-200, Nankoku, Kochi 783-8502, Japan.
⁴ Laboratory of Aquatic Environmental Science, Faculty of Agriculture and Marine Science, Kochi University, Otsu-200, Monobe, Nankoku, Kochi 783-8502, Japan. Electronic address: madachi@kochi-u.ac.jp.

PMID: 30509379
DOI: 10.1016/j.margen.2018.06.003

Abstract

The marine diatom Phaeodactylum tricornutum is attractive for basic and applied diatom research. We isolated putative endogenous gene promoters derived from genes that are highly expressed in P. tricornutum: the fucoxanthin chlorophyll a/c-binding protein (FCP) C gene, the vacuolar ATP synthase 16-kDa proteolipid subunit (V-ATPase C) gene, the clumping factor A gene and the solute carrier family 34 member 2 gene. Five putative promoter regions were isolated, linked to an antibiotic resistance gene (Sh ble) and transformed into P. tricornutum. Using quantitative RT-PCR, the promoter activities in the transformants were analyzed and compared to that of the diatom endogenous gene promoter, the FCP A gene promoter which has been used for the transformation of P. tricornutum. Among the five isolated potential promoters, the activity of the V-ATPase C gene promoter was approximately 2.73 times higher than that of the FCP A gene promoter. The V-ATPase C gene promoter drove the expression of Sh ble mRNA transcripts under both light and dark conditions at the stationary phase. These results suggest that the V-ATPase C gene promoter is a novel tool for the genetic engineering of P. tricornutum.

Keywords: Constitutive expression; Diatoms; High-level expression; Phaeodactylum tricornutum; V-ATPase C promoter.

MeSH terms

Algal Proteins / genetics*
Algal Proteins / metabolism
Diatoms / genetics*
Gene Expression*
Promoter Regions, Genetic*
Transgenes*

Substances

Algal Proteins