(A) Schematic representation of the MEP. β-estradiol-inducible Cre-recombinase is expressed under the control of a daughter cell-specific promoter, P SCW11 . LoxP sites are integrated surrounding components of the essential genes ubc9 and cdc20.

(B) The fold change in MEP cells inoculated at different initial densities is plotted over time, with 3-hr culture prior to the addition of β-estradiol. Error bars are SEM for 6 independent replicates.

(C) Viability plotted against replicative age for fluorescein-labeled versus unlabeled cells (n = 100 cells for each group; mean lifespans were 22.9 and 22.4, respectively).

(D) Total number of cells that fall into the fluorescein-positive fraction at various times during a representative High-Life experiment.

(E) The fraction of all cells that fall into the fluorescein-positive fraction at various times during a representative High-Life experiment.

(F) The fraction of progenitor cells viable is plotted against time, beginning after either birth of the cell (replicator) or initiation of the culture (High-Life), for representative experiments. A third line (false-positive adjusted) represents that fraction of viable cells in the High-Life environment after correcting for the false-positive rate observed with propidium iodide staining in the replicator device. Correction was performed by extrapolating a linear trend of false positives between cells in the replicator device stained with propidium iodide after 16 or 40 hr of culture and multiplying the observed High-Life viability by 1 minus the calculated false-positive fraction. 100 cells were considered for the replicator experiment. The High-Life experiment shows the mean of 48 replicate wells.

(G) The mean number of bud scars observed on fluorescein-labeled cells after 0, 8, and 24 hr of culture, plotted against the CF405M fluorescence intensity observed at the same time point. For bud scar counting, 20 cells were analyzed at each time point. CF405M intensity values are the mean of 48 replicate wells.