Cell cycle assay

At 48 hours after transfection, U87 and LN229 cells were harvested and washed using ice-cold PBS solution. Subsequently the cells were fixed with 70% ethanol overnight at 4 °C before being re-suspended using propidium iodide (PI)/RNase A solution (5 μg/mL PI and 100 mg/mL RNase A) and incubated for 15 min at room temperature in the dark. Then the flow cytometer (Millipore Guava) was utilized to analyze cell cycle. All the assays were conducted three times independently.