A novel method for hybridization of Saccharomyces species without genetic markers

Protoplasts of Saccharomyces cerevisiae were inactivated by treatment with different concentrations of antifungal compounds for various periods. Of the 14 compounds tested, N-ethylmaleimide proved to be the most efficient. The inactivation effect was fully reproducible. The inactivated protoplasts could be reactivated and still function as fusion partners. They were fused with untreated protoplasts by polyethylene glycol treatment and produced viable hybrid cells. Nuclear and extrachromosomal genetic analysis and chromosome separation of the fusion products from fusion experiments involving inactivated and non-inactivated protoplasts revealed that N-ethylmaleimide did not affect either of the genomes and hence it was perfectly suited for the hybridization of any type yeast cells without genetic markers.