SA-β-gal and growth curves

HCT116 or MCF-7 cells were transfected with indicated siRNAs and cultured for 24 h. Subsequently, each group of cells were reseeded in 12-well plates and subjected to different treatments. For SA-β-gal staining, cells at 50 to 60% confluency were treated with Dox (50 ng/ml) for the indicated durations and then washed with ice-cold PBS and fixed for 10 min with 3% formaldehyde at room temperature. SA-β-gal staining solution (Beyotime) was prepared before use and incubated with the cells overnight at 37 °C. For growth curves, 0.75 × 10⁴ of either HCT116 or HT-29 cells from each group were plated in 96-well plates in triplicate and treated with 50 ng/ml doxorubicin. The cells were cultured for another 1 to 5 days and monitored by MTT assay.⁴⁹ The growth curves were generated accordingly.