Figure 3.—

Cdc13-myc_18x telomere association is not diminished in ten1-101. (A) Chromatin immunoprecipitation of Cdc13myc_18x. Two hundred milliliters of wild-type untagged (hC160), CDC13myc₍₁₈₎ (hC1985) and ten1-101 CDC13myc_18x (hC2035) strains were grown in YPD or SD −His media to an OD₆₀₀ ∼1, and split into four 50-ml cultures. Two cultures were treated with nocodazole for 4 hr at 23° and 36° (Nz), and two cultures were diluted to an OD₆₀₀ = 0.4 and incubated at 23° or 36° for 4 hr (asynchronous cultures, A). Immunoprecipitations from cell lysates were carried out with an anti-myc antibody (12CA5). The percentage of the total telomeric DNA present in each IP was determined by dot blot analysis, hybridizing with a [³²P]d(GT/CA) probe, and exposing the blot on a Phosphorimager screen. To normalize the data, the percentage of telomere DNA recovered in each IP from Cdc13myc_18x strains was divided by the percentage of telomere DNA recovered in the IP from the untagged Cdc13 strain. The averages of four independent experiments are graphed, and the error bars represent the standard deviation. (B) Dot blot of ChIP DNA samples from strains grown either asynchronously or arrested in G2 with nocodazole probed with [³²P]d(GT/CA), as described in A. Shown are 10-fold dilutions of the immunoprecipitate and the input DNA for each strain.