Spot assay for assessing the MMS, CPT, HU and X-ray sensitivities of yeast cells producing S. cerevisiae Rad51 mutants. Cells were spotted in tenfold serial dilutions on plates, in the absence or presence of these DNA-damaging factors. The plates were incubated at 30 C for one day (CPT and X-ray), two days (no treatment), three days (MMS) or seven days (HU). Reproducibility of the spot assay was confirmed by three independent experiments (Supplementary Fig. 2b). Bottom right, bar graphs and dot plots showing the relative growth of yeast cells with newly introduced Rad51 genes compared with wild-type Rad51 after X-ray irradiation. The quantification was performed using the third spot (1:100 dilution). Data are mean s.d. (n = 3 independent replicates). P values obtained by one-sided Welch’s t-test were 1.9 10 −5 (+ empty vector), 2.4 10 −4 (+ rad51 K122A ), 0.22 (+ rad51 K128A ) and 3.9 10 −4 (+ rad51 K122A/K128A ). Asterisks indicate significance at P < 0.05.
