Fig. 2.

Schematic outline of the experimental design for the crossbreeding of sake yeast strains. a Outline of the basic method used for the crossbreeding of sake yeast strains. Plasmids pLhyS-2 K-Pa1 and pHhyS-3 K-2α were used for isolation of a- and α-type derivative cells, respectively, and plasmids pLY-hygro and pHY-kan were used for the hybridization of derivatives. Unnecessary plasmids were removed from yeast cells by cultivating cells in the absence of selection pressure. b Improved method for continuous crossbreeding without the requirement for removal of plasmid pLhyS-2 K-Pa1. c A second improved method for continuous crossbreeding without the requirement for removal of plasmid pHhyS-3 K-2α. Plasmid pAUR112 was used for the hybridization of obtained derivatives in (b) and (c)