Effect of cryopreservation protocols on the phenotypic stability of yeast

Eight cryopreservation protocols were assessed for their effects on the viability and phenotypic stability of the yeast Saccharomyces cerevisiae during a five-year study. It is found that viability and phenotypic features have remained largely unchanged when the yeast was preserved in glycerol, dimethyl sulphoxide, or sucrose at -80 degrees C or in liquid nitrogen. When sorbitol was used as a cryoprotectant, yeast cells frozen and stored at -80 degrees C manifested great decreases in viability after six months in storage and concomitantly large fluctuations in the rate of the trpl auxotrophic reversion. This phenotypic reversion was stable passage after passage. Such a degree of phenotypic fluctuations, however, was not observed for yeast cells preserved in the same sorbitol solution that went through a controlled freezing program and were subsequently stored in liquid nitrogen. These results indicate that some combinations of cryoprotective agent, freezing program, and storage temperature disturb biomaterials more profoundly during cryopreservation and imply a genetic basis of this phenotypic change.