title: Novel insights in genetic transformation of the probiotic yeast Saccharomyces boulardii.
abstract: Saccharomyces boulardii (S. boulardii) is a probiotic yeast related to Saccharomyces cerevisiae (S. cerevisiae) but with distinct genetic, taxonomic and metabolic properties. S. cerevisiae has been used extensively in biotechnological applications. Currently, many strains are available, and multiple genetic tools have been developed, which allow the expression of several exogenous proteins of interest with applications in the fields of medicine, biofuels, the food industry, and scientific research, among others. Although S. boulardii has been widely studied due to its probiotic properties against several gastrointestinal tract disorders, very few studies addressed the use of this yeast as a vector for expression of foreign genes of interest with biotechnological applications. Here we show that, despite the similarity of the two yeasts, not all genetic tools used in S. cerevisiae can be applied in S. boulardii. While transformation of the latter could be obtained using a commercial kit developed for the former, consequent screening of successful transformants had to be optimized. We also show that several genes frequently used in genetic manipulation of S. cerevisiae (e.g., promoters and resistance markers) are present in S. boulardii. Sequencing revealed a high rate of homology (> 96%) between the orthologs of the two yeasts. However, we also observed some of them are not eligible to be targeted for transformation of S. boulardii. This work has important applications toward the potential of this probiotic yeast as an expression system for genes of interest.
authors.affiliation: Fondazione Ri.MED; Palermo, Italy; University of Pittsburgh Center for Vaccine Research; Pittsburgh, PA USA.
authors.forename: Bruno
authors.initials: B
authors.lastname: Douradinha
authors.affiliation: Centro de Biotecnologia Molecular; Instituto de Ciências Biológicas; Universidade de Brasília; Brasília, Brazil.
authors.forename: Viviane C B
authors.initials: VC
authors.lastname: Reis
authors.affiliation: University of Pittsburgh Center for Vaccine Research; Pittsburgh, PA USA.
authors.forename: Matthew B
authors.initials: MB
authors.lastname: Rogers
authors.affiliation: Centro de Biotecnologia Molecular; Instituto de Ciências Biológicas; Universidade de Brasília; Brasília, Brazil.
authors.forename: Fernando A G
authors.initials: FA
authors.lastname: Torres
authors.affiliation: University of Pittsburgh Center for Vaccine Research; Pittsburgh, PA USA; Department of Microbiology and Molecular Genetics; School of Medicine; University of Pittsburgh; Pittsburgh, PA USA.
authors.forename: Jared D
authors.initials: JD
authors.lastname: Evans
authors.affiliation: University of Pittsburgh Center for Vaccine Research; Pittsburgh, PA USA; Department of Infectious Diseases and Microbiology; School of Medicine; University of Pittsburgh; Pittsburgh, PA USA.
authors.forename: Ernesto T A
authors.initials: ET
authors.lastname: Marques
mesh_terms: D060846:Cell Engineering; D004251:DNA Transposable Elements; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D016681:Genome, Fungal; D010957:Plasmids; D019936:Probiotics; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012440:Saccharomyces / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014170:Transformation, Genetic; D019076:Transgenes
chemical_list: D004251:DNA Transposable Elements; D005656:Fungal Proteins; D011994:Recombinant Proteins
keywords: Saccharomyces boulardii; expression of foreign sequences; promoter; resistance markers; screening