Artificial Conversion of the Mating-Type of <i>Saccharomyces cerevisiae</i> without Autopolyploidization

Crossbreeding is a classical yeast hybridization procedure, where the mating of haploid cells of opposite mating-type, <i>MAT</i>a and <i>MATα</i> cells, produces a new heterozygous diploid. Here, we describe a method to generate haploid <i>MAT</i>a and <i>MATα</i> cells using mating-type conversion caused by expression of the <i>HO</i> gene, which encodes an endonuclease. Importantly, our method prevents the autopolyploidization that typically arises during artificial mating-type conversion. This facilitates isolation of the desired mating-type of yeast cells with simple and easy procedure. In the current study, we designed a suitable genetic circuit for each haploid cell and converted <i>MAT</i>α haploid cells into <i>MAT</i>a haploid cells and vice versa, demonstrating the utility of constructed artificial regulation network to prevent autopolyploidization. Via forced expression of the a1 gene in <i>MAT</i>α haploid cells or of α2 in <i>MAT</i>a haploid cells, the undesirable mating ability of yeast cells was completely suppressed. We confirmed the success in prevention of autopolyploidization by ploidy analysis. This new approach provides a reliable and versatile tool for yeast crossbreeding, so that it will be useful for scientific research and industrial applications of yeast