Novel endogenous promoters for genetic engineering of the marine microalga Nannochloropsis gaditana CCMP526.

• Three novel promoters were characterised in N. gaditana for transgene expression. • Hsp90 and EPPSII promoters outperformed TUB , a commonly used constitutive promoter. • These promoters expand N. gaditana genetic resource for basic and applied research. Nannochloropsis is a marine microalga from the Eustigmatophyceae stramenopile lineage that has been studied extensively due to a broad range of industrial applications, mostly related to their oil and pigment production. However, tools to genetically engineer members of this group, and therefore further understand and maximise their industrial potential are still limited. In order to expand the potential industrial uses of this organism, several molecular tools, including gene promoters of different strength, are needed. A comprehensive and diverse set of well-characterized promoters is key to a number of genetic engineering and synthetic biology applications, such as the assembly of complex biological functions or entire metabolic pathways. In this study, we measured the promoter activity of three endogenous constitutive promoters from N. gaditana genes EPPSII (Nga02101); HSP90 (Nga00934); ATPase (Nga06354.1) in driving the expression of a Sh ble- mVenus fluorescent reporter fusion protein. Through a combined approach that includes flow cytometry, RT-qPCR and immunoblotting, we profiled the activity of these promoters at both the transcript and protein level. Two promoters HSP90 (Nga00934) and EPPSII (Nga02101) outperformed the widely used β-tubulin promoter, exhibiting 4.5 and 3.1-fold higher mVenus fluorescence, respectively. A third promoter ATPase (Nga06354.1) was also able to drive the expression of transgenes, albeit at lower levels. We show that the new promoters identified in this study are valuable tools, which can be used for genetic engineering and functional genetics studies in N. gaditana. [ABSTRACT FROM AUTHOR]