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Propidium iodide (PI) staining cell cycle analysis
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HCC cells were planted into 6-well plate at 3 × 105 (HepG2) or 1.5 × 105 (QGY-7703) per well. Approximately 106 transfected HCC cells were fixed in 70% ethanol for at least 2 h. After washing with PBS, cells were stained in 1 mL PI staining solution containing 10 μg/mL of PI, 100 μg/mL of RNase A, and 0.1% Triton X-100 dissolved in PBS, for 30 min in the dark. Cell cycle distribution was analyzed by flow cytometry using an Accuri C6 Plus flow cytometer (BD). The cell cycle tests were performed in two independent experiments with each group in duplicate.
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