English: igure 7. S. boulardii Mutants Express Functional GFP. (A) Bright field and fluorescent images of ura3− S. cerevisiae laboratory haploid (S.c.) and S. boulardii Mutant 2 (M2) either untransformed (Control) or transformed (+GFP) with a URA3 plasmid containing GFP. The GFP fluorescence is detected in the FITC channel. Corresponding differential interference contrast (DIC) images are also shown. Scale bars show 10 µm. (B) Representative flow cytometry plots of forward-scattered light (FSC) versus GFP fluorescence for untransformed (Control) and transformed (+GFP) S. cerevisiae laboratory haploid (S.c. lab haploid) and S. boulardii Mutant 2 (M2) showing the percent of GFP positive cells in each population (n = 2 per strain). Transformed yeast were maintained in media lacking uracil prior to analysis. (C) Retention of URA3 plasmid and GFP expression was tested by comparing the percent of GFP positive cells of untransformed yeast (Control) relative to transformed yeast cultured in either selective media lacking uracil (URA−Glu), YPD (non selective media) for 4 hours (YPD 4 hr), or YPD for 24 hours (YPD 24 hr). Yeast strains analyzed include untransformed and transformed ura3− S. cerevisiae laboratory haploid (S.c.) and S. boulardii Mutants 1–3 (M1, M2, M3). Median fluorescent intensity (MFI) of GFP positive cells in each population is also depicted, indicating there is no visible decrease in average GFP expression per cell after incubation in YPD for 4 or 24 hours. Bars depict the mean of two samples per strain per incubation condition