Fig. 4

Loss of SUT457 leads to Exo1-dependent accumulation of telomeric ssDNA. a Analysis of telomeric ssDNA overhangs in isogenic wild-type and sut457Δ strains. Yeast colonies were restreaked on agar plates and genomic DNA was extracted at passages 1 (lanes 1 and 2) and 5 (lanes 3 and 4), fragmented with XhoI and subjected to native southern blotting (left panel) using a biotinylated probe against telomeric repeats. The southern blot was then treated with 0.4 N NaOH and the denatured DNA was re-probed to monitor equal loading (right panel). b Accumulation of telomeric ssDNA in eleven sut457Δ strains compared to their respective wild-type strains during passages 1 and 5 (P1 & P5). The ssDNA signal for each wild-type and mutant strain detected in the native southern blot was normalised to the corresponding signal in the denatured southern blot. Error bars represent standard error of the mean. * P < 0.05; ** P < 0.01; n/s, not significant (generated by nonparmetric one-tailed Mann–Whitney t-test). c Cell growth was monitored for the indicated isogenic strains at passages 1 and 5. d Analysis of telomeric ssDNA levels at passage 5 of wild-type, sut457Δ, exo1Δ and exo1Δsut457Δ isogenic strains generated through dissection of the heterozygous diploid double mutant EXO1/exo1ΔSUT457/sut457Δ. Southern blotting was performed as in (a) above. e Telomeric ssDNA levels analysed at passage 5 in the indicated isogenic wild-type and mutant strains generated from three independent tetrads (n = 3). The quantification of the ssDNA signal for each wild-type and mutant strain was performed as in (b) above. Error bars represent standard error of the mean. ** P < 0.01; n/s, not significant. The statistical analysis was performed using one-way ANOVA (Dunnett’s test)