Abstract
Microalgae not only serve as raw materials for biofuel but also have uses in the food, pharmaceutical, and cosmetic industries. However, regulated gene expression in microalgae has only been achieved in a few strains due to the lack of genome information and unstable transformation. This study developed a species-specific transformation system for an oleaginous microalga, Ettlia sp. YC001, using electroporation. The electroporation was optimized using three parameters (waveform, field strength, and number of pulses), and the final selection was a 5 kV cm−1 field strength using an exponential decay wave with one pulse. A new strong endogenous promoter CRT (Pcrt) was identified using transcriptome and quantitative PCR analysis of highly expressed genes during the late exponential growth phase. The activities of this promoter were characterized using a codon optimized cyan fluorescent protein (CFP) as a reporter. The expression of CFP was similar under Pcrt and under the constitutive promoter psaD (PpsaD). The developed transformation system using electroporation with the endogenous promoter is simple to prepare, is easy to operate with high repetition, and utilizes a species-specific vector for high expression. This system could be used not only in molecular studies on microalgae but also in various industrial applications of microalgae.
Highlights
Microalgae serve as raw materials for biofuel and have uses in the food, pharmaceutical, and cosmetic industries
The metabolic engineering of microalgae offers an alternative method for increasing metabolites without impairing growth by regulating only the gene expression related to the targeted metabolic p athway[5,6,7]
This study investigated the use of electroporation to establish the stable transformation of an oleaginous microalga, Ettlia sp
Summary
Microalgae serve as raw materials for biofuel and have uses in the food, pharmaceutical, and cosmetic industries. The developed transformation system using electroporation with the endogenous promoter is simple to prepare, is easy to operate with high repetition, and utilizes a species-specific vector for high expression. This system could be used in molecular studies on microalgae and in various industrial applications of microalgae. The development of a strong novel endogenous promoter based on various omics studies has been carried out for the metabolic engineering of microalgae for stable gene insertion and expression and high-yield production of recombinant proteins. The development of an endogenous promoter and an optimized expression system is essential for the metabolic engineering of Ettlia sp
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