Fig. 1.

The structure of triplex histone switch. Duplex integrated safeguard based on a pair of histone genes. Histone H3 gene HHTS is regulated by a galactose promoter pGAL1, and the histone H4 gene HHFS is controlled by another galactose promoter pGAL7. Each histone switch is flanked by a pair of loxP sites. The duplex histone safeguard switches can either be on a CEN/ARS plasmid (A) or integrated into a genomic locus (B). Either the native Gal4 protein or an engineered tribrid protein called GEV turns on the transcription of the histone genes in the presence of their ligands, such that the engineered yeast survives. The third level of redundancy is use of an orthogonal control mechanism, site-specific recombination. Any pair of loxP sites (yellow diamonds) on the construct will recombine and delete one or both histone genes upon Cre, activation, leading to inviability. Both proteins can be independently controlled by small molecule(s). Hooked arrows, promoters; filled arrows, histone genes; blue lozenges, standardized vector components; HOL, HOR, sequences to Left and Right of HO gene, where the safeguard was integrated.