Fig 1: RNR-deg is a superior alternative to HU for inducing cell cycle arrest. (A) Schematic of RNR-deg. AID-Rnr1 is degraded by OsTIR1 in the presence of NAA, in an rnr3Δ background, halting dNTP production required for DNA replication. (B) Survival spot assay of WT vs. RNR-deg (OsTIR1, AID-rnr1, rnr3Δ) strains. OsTIR1 expression requires Gal, and degradation requires NAA. Spots contained 5 × 104, 5 × 103, 500, or 50 cells. (C) Flow cytometry profiles of RNR-deg cells that were synchronized in G1 by αF and released into YPG/Gal with the indicated treatment (200 mM HU or 250 µM NAA). (D) RNR-deg treatment reverses faster than HU treatment after washout. Asynchronous RNR-deg cells were treated with αF, released into media containing the indicated treatment for 4 h, and washed extensively by pelleting (1,000×g), then resuspending cells in water two times, followed by a final pelleting/resuspension in YPG/Gal. Cytometry profiles were acquired at the indicated times after washout. Green bar highlights cells in G2. The cell cycle speed of RNR-deg + NAA cells after wash is faster than postwash HU-treated cells, comparable to the speed of untreated cells. (E) Cytometry profiles of red-shifted fluorescence intensity from DHE-stained RNR-deg cells in the absence or presence of 250 µM NAA; the untreated profile from Fig. 2B is used for comparison.