Our project involves genetically modifying the yeast Saccharomyces cerevisiae to express the enzyme phytase. S. cerevisiae is classified as GRAS (generally regarded as safe), and the enzyme that is being overexpressed is not toxic.
We are using an auxotrophic marker (URA3) for transforming S. cerevisiae with an episomal plasmid containing the phytase gene. For genome integration too, an auxotrophic marker (TRP1) will be used. No antibiotic markers will be used.
However, as the plasmids we are using are E. coli-S. cerevisiae shuttle vectors, the bla gene (ampicillin antibiotic marker) is present in the vector. Integration of a linear DNA fragment containing just the expression cassette and an auxotrophic marker in the yeast genome is an approach which we plan to take once we have established proof of concept.
As the constructed yeast strain will be used for leavening breads, which are baked before consumption, we do not expect release of the modified organism in the environment.