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6 cells/dish in 100-mm dishes) were treated with various concentrations of test compounds for 18 h. Collected cells were lysed with lysis buffer (50 mM Tris-HCl (pH 7.5), 20 mM EDTA, and 1% NP-40) for 5 min at 4 °C. Lysates were incubated with 10% SDS (final 1%) for 1 h at 56 °C and then treated with proteinase K solution (final 2.5 μg/μl) for 2 h at 37 °C. DNA was isolated by adding 10 M ammonium acetate (final 5 M) and ice-cold pure ethanol. After incubation overnight and centrifugation, DNA was washed twice with ice-cold 80% ethanol, dried in the air, and dissolved in TE buffer (10 mM Tris-HCl (pH 8.0) and 1 mM EDTA). Five micrograms of DNA of each tested sample was subjected to 2% agarose gel electrophoresis. The gel was stained with SYBR Gold (Molecular Probes) for 1 h at room temperature and visualized under UV transillumination.
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