(A) Representative Western Blot assessing dUTPase expression after 48 and 72 hours of siRNA treatment using dUTPase specific siRNA (sidUTPase) or a non-targeting siRNA control (siNon-t). β-Actin was used as a loading control. (B) Clonogenic survival of dUTPase depleted cells compared to siNon-t transfected or untransfected controls, treated for 48 hours with increasing concentrations of 5-FU. Data shown as average ± SEM from two independent experiments performed in triplicate. Statistical significance between untransfected and sidUTPase was determined by using a two-tailed t-test. (C) FACS analysis highlighting cell cycle alterations induced by 5-FU treatment in sidUTPase and siNon-t transfected cells. After 48 hours of siRNA transfection, cells were re-seeded and, 24 hours later, treated for 48 hours with increasing concentrations of 5-FU. DNA content was stained with PI and analyzed by FACS. Representative histograms are shown. Abbreviation: AU: arbitrary unit. (D) Quantification of the FACS experiment in Figure 1C. Data shown as average ± SEM from two independent experiments. Abbreviations: N: siNon-t, D: sidUTPase.