Isolation of ethanol-tolerant mutants of yeast by continuous selection

Mutants of Saccharomyces uvarum, 5D-cyc with increased tolerance to ethanol have been isolated by a continuous selection technique which allows the culture itself to determine the intensity of selection via a feedback control circuit. The output of CO2 from a continuous culture of the yeast was monitored using an infrared analyser and the signal from that analyser fed to a potentiometric controller which regulated the introduction of a concentrated ethanol solution into the culture vessel. The frequency of ethanol addition to the culture thus increased as the tolerance of the organisms improved. The use of this system permitted the selection of mutants of yeast which were viable in the presence of 12% w/v ethanol and which showed higher fermentation rates (as measured by CO2 production) than the wild-type in the presence of 10% w/v ethanol and above. The technique of continuous selection with feedback should be generally applicable to the isolation of mutants of any microorganism to improved tolerance to any inhibitory condition of either its physical or chemical environment. © 1982 Springer-Verlag.