Figure 9: Effects of depletion of XPF on Mus81−/− cell survival.

HCT116 and Mus81−/− cells were transfected with control siRNA or XPF siRNA as described in Fig. 8. (a) Cell cycle analysis by FACS. Cells were pulse-labelled with EdU for 45 min before harvesting. Cell cycle was tested with a Click-iT EdU kit by FACS. Examples of bivariate FACS were shown and percentage of cells in each cell cycle was shown in Supplementary Table 5. (b) Histogram of percentage of subG1 from (a). (c) The % of apoptotic cells was measured by TUNEL assay. Cells were transfected with control siRNA or XPF siRNA twice with 72 h interval and collected 72 h after the second transfection. TUNEL-positive cells were analysed by FACS (see Supplementary Fig. 6B for the FACS dot plots). (d) Clonogenic assays. Twenty-four hours after the second siRNA transfection, cells were seeded into six-well plates at 50, 250 or 1,000 cells per well. Two weeks later, cells were fixed and stained to count the colonies.