Escapee analysis of a genetic biocontainment of yeast with a protein stability switch

We have developed a genetic biocontainment system based on conditional stability of essential proteins. We used a yeast-adapted destabilizing domain degron, which can be stabilized by estradiol addition (ERdd). Leveraging the yeast GFP collection and lab automation platforms, we ERdd-tagged 775 essential genes and screened for strains with estradiol dependent growth. Three genes, SPC110, DIS3 and RRP46, were found to be particularly suitable targets. Escapees strains capable of growing without estradiol derived from the ERdd-tagged containment strains RRP46-ERdd, DIS3-ERdd and RRP46-ERdd/DIS3-ERdd were subjected to whole genome sequencing to elucidate their escape mechanism.