U-2-OS cells were transfected with siRNA for 24 hours and treated with UCN-01 for 4 hours or irradiated with 20 Gy of ionizing radiation. Cells (1 × 106) were molded into 1% low-melting agarose plugs (InCert Agarose, Lonza), followed by treatment with proteinase K buffer [10 mM tris-Cl (pH 7.5), 50 mM EDTA, 1% N-laurylsarcosyl, proteinase K (2 mg/ml)] at 50°C for 48 hours. Plugs were then subjected to PFGE (1% agarose, CHEF-DR II system; Bio-Rad Laboratories; 120° angle, 60- to 240-s switch time, and 4 V/cm) for 20 hours. DNA was visualized by ethidium bromide staining. Quantification was performed using ImageJ software.

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