(A) Equal numbers of early logarithmic phase cells from WT
and ubc1Δ strains with or without an endogenous
Hcm1-TAP tag were lysed followed by TAP Western analysis.
(B) Assessment of Hcm1 protein stability over 3 hours in WT
and ubc1Δ strains in the presence of cycloheximide,
added at time (0), in 2% glucose. Equal cell numbers were removed at the
indicated time points with 30 μg protein loaded.
(C) Fluorescent microscopy of genomically integrated
GFP-tagged Hcm1 expressed from its endogenous promoter in isogenic WT
and ubc1Δ strains. Both strains were arrested in G1
followed by release, with cells collected at the indicated
timepoints.
(D) Flow cytometry analysis of cells collected at the
timepoints in (A), highlighting the relative proportion of replicated
DNA (2n) upon release from G1.