FIG. 1.

Analysis of CLB2 mRNA levels through the cell cycle. (A) Comparison of CLB2 mRNA levels between wild-type and mutant strains carrying the nme1-P6 or cdc15-1 mutation. Cells were grown at 24°C, shifted to 37°C for 2 h, and RNA was isolated as described in Materials and Methods. An equal amount of RNA from each strain was subjected to Northern analysis. All blots were first probed for CLB2 mRNA and subsequently for ACT1 mRNA. The locations of the relevant transcripts are shown (wild-type, MES111-140; nme1-P6, MES111-P6; cdc15-1, TLG136). (B) Analysis of CLB2 mRNA levels in cell cycle-synchronized S. cerevisiae. Wild-type (MES111-140) and nme1-P6 mutant (MES111-P6) strains were grown to 10⁶ cells/ml at 25°C in SCD, arrested in hydroxyurea for 2 h at 25°C, and then shifted to 37°C for 1 h. Cells were then washed to remove the hydroxyurea, resuspended in fresh medium, and maintained at 37°C. Total RNA was made at 0, 15, 30, 45, 60, 75, 90, 105, and 120 min after release from hydroxyurea arrest (see Materials and Methods). RNA was harvested at the indicated times, and CLB2 and ACT1 mRNA levels were measured. Cell synchrony and release from arrest were monitored by examination of cell budding and morphology. The signals corresponding to the CLB2 and ACT1 mRNAs are shown.