Testing the overexpression of a bacterial PETase using a novel overexpression system in Nannochloropsis oceanica

Microalgae have become highly interesting organisms for biotechnological use due to a range of conveniences including their high photosynthetic efficiency or the possibility to be used as “green” biofactories that generate molecules of our interest, for example proteins. However, overexpression systems for protein production are rather scarce in biotechnologically relevant microalgae. In this project we therefore explored a novel overexpression system in the microalga Nannochloropsis oceanica towards the production of recombinant protein. This system relies on a fluorescence reporter gene in the microalgal genome, which can be replaced by a desired DNA fragment via homologous recombination, resulting in the loss of fluorescence. Protein overexpression is regulated by an RNA polymerase I promoter, found to be highly efficient in this microalga. To test the potential for protein production, a gene encoding a PET-degrading enzyme from Ideonella sakaiensis was cloned into an appropriate plasmid which was then delivered into the microalgae cells. After the evaluation of the loss of fluorescence of the colonies, they were tested for the presence of the trans-gene via PCR. The overexpression of the protein was tested via Western blot. This overexpression system strategy could lead to the development of environmentally friendly solutions to produce proteins using photosynthetic organisms.