Dnmt1

Ginsenoside 20(S)-Rg3 was obtained from Tasly Pharmaceutical Company (Tianjin, China) and dissolved at a concentration of 4 mg/ml in DMSO as a stock solution (stored at −20°C). It was then further diluted in cell culture medium to create working concentrations. The maximum final concentration of DMSO was less than 0.1% for each treatment, and was also used as a control. Antibodies including DNMT3A, DNMT3B, E-cadherin, N-cadherin, vimentin, and β-actin were from Cell Signaling Technology (Beverly, MA), FSCN1 was from Abcam (Cambridge, MA, USA), and DNMT1 was from Active Motif (Carlsbad, CA, USA).

Total protein was collected from cells by RIPA lysis buffer containing protease inhibitors (Roche, Indianapolis, IN, USA) and 1 mM PMSF on ice. Protein concentration was measured using the BCA-200 Protein Assay kit (Pierce, Rockford, IL, USA). After heat denaturation at 100°C for 5 min, proteins were separated by electrophoresis on 10% SDS–PAGE gels and then transferred onto nitrocellulose membranes (Pall Life Science, NY, USA). The membranes were blocked with 5% non-fat milk at room temperature for 1 h, and then incubated overnight at 4°C with rabbit anti-human E-cadherin(1:1000, Cell Signaling Technology, Danvers, MA, USA), N-cadherin(1:1000, Cell Signaling Technology, Danvers, MA, USA), vimentin(1:500, Cell Signaling Technology, Danvers, MA, USA), DNMT3A (1:500, Cell Signaling Technology, Danvers, MA, USA), DNMT3B(1:500, Cell Signaling Technology, Danvers, MA, USA), DNMT1(1:1000, Active Motif; Carlsbad, CA, USA), FSCN1(1:100000, Abcam, Cambridge, MA, USA), and mouse anti-human β-actin(1:1000, Cell Signaling Technology, Danvers, MA, USA). After washing with TBST, the blots were incubated with horse radish peroxidase (HRP)-conjugated goat anti-rabbit or anti-mouse IgG. Blots were visualized using ECL reagents (Pierce, Rockford, IL, USA) by a chemiluminescence imaging system (Bio-Rad, Richmond, CA, USA).