Figure 1.

Hysteresis in the cell cycle. A strain of genotype cln1 cln2 cln3 GAL-CLN3 cdc14-1 was grown to log phase in YEPGal (galactose medium, GAL-CLN3 on) at 23°C and blocked due to CLN deficiency by incubation in YEPRaff (raffinose medium, GAL- CLN3 off) at 23°C for 6 h. Galactose was then added to 3% final concentration to induce GAL- CLN3. At the indicated times after galactose addition, aliquots of the culture were removed. Some of the aliquot was immediately extracted for immunoblotting with anti-Clb2 antibody. To the rest of the aliquot glucose was added to 2% final concentration to repress GAL-CLN3, and the aliquot was incubated at 37°C for 2.5 h. The aliquot was then extracted for immunoblotting. The percentages of unbudded cells at the time of aliquot removal and after the 2.5-h incubation in glucose at 37°C were determined microscopically. Top: Clb2 immunoblot for cells in continuous galactose (G), after the raffinose block (R), and after the indicated amounts of time after galactose addition (R+G; all at 23°C). Bottom: Clb2 immunoblot for cells incubated in R+G for the indicated amount of time and then shifted to glucose at 37°C for 2.5 h. *An unidentified background band used for standardizing protein loading. The percentage of unbudded cells at the time of protein harvest is indicated below the gel lanes. The top and bottom immunoblots were from the same membrane and film exposure.