FIG. 1.
Telomeric recombination in budding yeast. (A) In the standard setup, a telomerase-negative (tlc1Δ) rad52Δ (::LEU2) double mutant issued from parents heterozygous for both TLC1 and RAD52 was unviable (lower row), unlike the tlc1Δ RAD52+ mutant (upper row). In this setup, the tlc1Δ mutant used to construct the original diploid was not yet recombining at the time of mating. Subsequently, following sporulation of the diploid and selection of the desired genotypes (day 1), cells were grown in liquid YEPD medium at 29°C and diluted down to 105 cells/ml every 24 h (one cell cycle is ∼90 min) for 6 continuous days. Aliquots of the cultures were dropped onto YEPD plates to assess viability every day (from one image to the next). (B) Schematic representation of three different methodologies used to overcome death by senescence in telomerase-negative cells (TLC1 encodes the RNA subunit of telomerase) in the absence of the RAD52 recombination protein. (C) Survival from telomerase inactivation generates two types of postsenescence telomeric recombination: type I, amplifying the Y′ subtelomeric sequences, and type II, amplifying the TG1-3 telomeric sequences, best identified after XhoI digestion as schematically explained. See also Materials and Methods.