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Bonetti et. al., PLoS Genet. 2010 May; 6(5): e1000966.<br />
YLL2730 UCC5913 yku70Δ::URA3 mre11Δ::NATMX bar1Δ::HPHMX<br />
YLL2731 UCC5913 rif2Δ::NATMX exo1Δ::URA3 bar1Δ::HPHMX<br />
YLL2733 UCC5913 rap1Δ::KANMX4 [CEN-HIS3-rap1Δ670-807]<br />
exo1Δ::URA3 bar1Δ::HPHMX<br />
YLL2736 UCC5913 rap1Δ::KANMX4 [CEN-HIS3-rap1Δ670-807]<br />
mre11Δ::NATMX bar1Δ::HPHMX<br />
K699 MATa ade2-1 can1-100 his3-11,15 leu2-3,112 trp1-1 ura3-<br />
1 rad5-535<br />
DMP5108/<br />
19A<br />
DMP5108/<br />
20A<br />
K699 cdc13-1<br />
K699 cdc13-1 rif2Δ::KANMX4<br />
Plasmids are in<strong>di</strong>cated by brackets. All strains are from this study except the strains<br />
UCC5913: Diede SJ, Gottschling DE (2001) Exonuclease activity is required for<br />
sequence ad<strong>di</strong>tion and Cdc13p loa<strong>di</strong>ng at a de novo telomere. Curr Biol 11: 1336-1340.<br />
RMY169: Michelson RJ, Rosenstein S, Weinert T (2005) A telomeric repeat sequence<br />
adjacent to a DNA double-stranded break produces an anticheckpoint. Genes Dev 19:<br />
2546-2559.<br />
Resection assay<br />
Visualization of the single-stranded overhangs at native telomeres was done<br />
as described [31]. The same DNA samples were separated on a 0.8% agarose<br />
gel, denatured and hybri<strong>di</strong>zed with the end-labeled C-rich oligonucleotide for<br />
loa<strong>di</strong>ng control. To monitor resection at the HO-derived telomeres, RsaI- and<br />
EcoRV-<strong>di</strong>gested genomic DNA was subjected to denaturing polyacrilammide<br />
gel electrophoresis and then hybri<strong>di</strong>zed with the single-stranded riboprobes A<br />
or B, which anneal to the 5′ C-strand or the 3′ G-strand, respectively, to a site<br />
located 212 nt from the HO cutting site. Resection of the C-rich strand in<br />
Figure 3E and Figure 4C was monitored by hybri<strong>di</strong>zing RsaI-<strong>di</strong>gested genomic<br />
DNA with riboprobe A. To monitor resection of the 5′-strand at the HOinduced<br />
DSB, EcoRV-<strong>di</strong>gested genomic DNA was hybri<strong>di</strong>zed with a singlestranded<br />
riboprobe, which anneal to the 5′-strand to a site located 215 nt from<br />
the HO cutting site. For quantitative analysis of C-strand and G-strand signals,<br />
the ratios between the intensities of ssDNA and loa<strong>di</strong>ng control bands were<br />
calculated by using the NIH image program.<br />
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