Osmotic
stress resistant yeast Zygosaccharomyces rouxii - isolation of
auxotrophic mutants and efficient transformation by electroporation.
Lenka Pribylova,
Hana Sychrova
Dept. Membrane Transport, Inst. Physiology CzAcadSci, Videnska 1083, Prague 4,
142 20, Czech Republic (pribyll@biomed.cas.cz)
The yeast Zygosaccharomyces rouxii is known for it's high tolerance to osmotic stress. This typical feature is supposedly due to some sets of specific genes, enabling this yeast to grow in media with high concentrations of salts and/or sugars, i.e. under conditions restrictive to Saccharomyces cerevisiae. By this time, there are only few studies of this yeast carried out on molecular level. One of the reasons is the high resistance of Z. rouxii cells to routine and quick transformation procedures. So far, the only one technique for its transformation was the laborious and time-consuming spheroplasting method, and only one type of auxotrophic mutants, leu2 (derived from the wt strain ATCC 42981), has been successfully transformed. There existed only few vectors suitable for Z. rouxii (i.e. containing S. cerevisiae ARS/CEN or part of the Z. rouxii pSR1 plasmid and ScLEU2 as a marker). Here we report the isolation of first Z. rouxii ura3 mutants, protocols for efficient transformation of the leu2 and ura3 mutants by electroporation and the construction and use of different vectors (ARS/CEN vs. pSR1, ZrLEU2, ScLEU2, ScURA3 markers). Protocols for transformation by electroporation slightly differ for different Z. rouxii strains. This work was supported by grants GA CR 204/01/0272 and AVOZ 5011922.