Applied Microbiology and Biotechnology, Vol.48, No.2, 204-207, 1997
A Counter Selectable Marker for Genetic-Transformation of the Yeast Schwanniomyces-Alluvius
We report here a counter-selectable marker system for genetic transformation of the yeast Schwanniomyces alluvius, based on the complementation of uracil auxotrophs defective in either orotidine-5’-phosphate decarboxylase (URA3) or orotidine-5’-pyrophosphatase (URA5). Uracil auxotrophs of S.alluvius were obtained by ethyl methanesulphonate mutagenesis and complemented using the ura3 gene from S.cerevisiae. A transformation frequency of approximately 10(4)/mu g DNA was obtained, which is tenfold higher than results described in earlier reports. Transformants were analysed by Southern blot hybridisation and were found to de mitotically stable. The extrachromosomal nature of the transforming DNA was confirmed by Southern hybridisation and plasmid rescue. The rescued plasmid DNA had a restriction pattern identical to that of the parent plasmid.