Kgaa3d5

Total protein from cells and tissues was lysed in Whole Cell Lysis Assay (Cat. KGP250, KeyGen BioTECH, China). A total of 30 µg of protein per sample was resolved by 10% SDS-PAGE and transferred to PVDF membranes. The membranes were first incubated overnight at 4°C in BSA in TBS containing 0.05% Tween 20 with primary antibodies against E-cadherin, N-cadherin, β-catenin, ZEB1 (1:1000, Cat.9782, Cell Signaling Technology, USA), and ZEB2 (1:1000, cat. 14,026–1-AP, Proteintech, USA) diluted in antibody diluent (cat. 8112 L, Cell Signaling Technology, USA), followed by incubation with secondary antibodies (Cat. KGAA3d5, KeyGen BioTECH, China) conjugated with horseradish peroxidase at room temperature for 1 h. The protein bands were detected by using an enhanced chemiluminescence plus kit (Millipore) as recommended by the manufacturer.

Total protein from tissues was lysed in a whole‐cell lysis assay (Cat. KGP250, KeyGen BioTECH, China) containing protease inhibitors (Cat. KGP603, KeyGen BioTECH, China) and phosphatase inhibitors (Cat. KGP602, KeyGen BioTECH, China). Then, 10 µg of protein per sample was resolved by 10% SDS‐PAGE and transferred onto PVDF membranes. The membranes were first incubated overnight at 4°C in BSA in TBS containing 0.05% Tween 20 with primary antibodies against β‐actin (1:5,000, Cat. M20010, Abmart, USA) and PMS2 (1:1,000, Cat. 66075‐1, Proteintech, USA), followed by incubation with secondary antibodies (Cat. KGAA3d5, KeyGen BioTECH, China) conjugated with horseradish peroxidase at room temperature for 1 hr. The protein bands were detected using an enhanced chemiluminescence plus kit (Millipore) according to the manufacturer's instructions.