Download Supplementary Figure 1 (A) Flow cytometry analysis of DNA content

Supplementary Figure 1 (A) Flow cytometry analysis of DNA content in exponentially
growing wild type cells exposed to HU, MMS or CPT as described in Fig. 3. Cells are
collected before induction of Gal-HO. (B) Viability of cells exposed to Zeocin and HU as
described in Fig. 2C. 100 cells were washed and plated on YPD. Viable colonies were
counted after 48h at 30°C.
Supplementary Figure 2 (A) PCR analysis of DSB repair at the MAT locus after induction of
HO in wild type cells (PP723) exposed or not to HU, MMS and CPT. Cell growth and arrest
was performed as described in figure 3. *: ARG5,6 DNA. (B) Position of PCR primers is
indicated with arrows. (C) Quantitation of the intensity of the DNA repair band in untreated
(Ctrl) cells or in cells exposed to HU, MMS and CPT. Band intensity was normalized to
ARG5,6.
Supplementary Figure 3 Analysis of DSB resection at the MAT locus by alkaline gel
electrophoresis in wild type cells exposed to HU, MMS and CPT. (A) Wild type cells (PP648)
were exposed to genotoxic drugs as indicated in Fig. 3 and were harvested at the indicated
times after induction of the HO endonuclease. Genomic DNA was digested with StyI and was
analyzed by alkaline gel electrophoresis. Resection generates ssDNA fragments of increasing
length (fragments 1, 2 and 3) due to the progressive loss of StyI restriction sites. C: HO cut
fragment. *: StyI fragments detected by the probe. (B) Southern blot analysis of HO-induced
resection in untreated cells or in cells exposed to HU, MMS or CPT. (C) Quantitation of the
intensity of resection bands 1, 2 and 3 normalized to a control fragment on chromosome III.
Supplementary Figure 4 (A) Schematic representation of the MAT locus. The progressive
loss of StyI restriction fragments upon induction of the HO endonuclease with galactose and
resection of 5’-ends the break is shown. See figure 4C for the corresponding Sothern blot. (B)
Quantitation of unequal sister-chromatid exchange (uSCE) in wild type (PP915) and rad51
(PP985) cells exposed for 45 minutes to 0.02% MMS. The experiment was performed as
described previously (Fasullo et al., 2001).