Multimodal Protein Expression Analysis

For qRT-PCR, total RNA was isolated with TRIzol reagent (Invitrogen), and cDNA was synthesized using the First Strand cDNA Synthesis kit (Yeasen). For the quantitative real-time PCR (qRT-PCR), the SYBR Green Supermix kit (Yeasen) was used on the StepOnePlus™ Real-Time PCR System (Applied Biosystems). The primer sequences for qRT-PCR were as follows:
Western blotting procedures were described in our previous study (18 (link)). The primary antibodies used were MISP (1:1,000, cat.no. 26338-1-AP, Proteintech); ZO-1 (1:1,000, cat. no. 8193P; Cell Signaling Technology); ZEB1 (1:1,000, cat. no. 70512S; Cell Signaling Technology), N-cadherin (1:1,000, cat. no. 13116T; Cell Signaling Technology); vimentin (1:1,000, cat. no. 3932S; Cell Signaling Technology); E-cadherin (1:1,000, cat.no. 20874-1-AP, Proteintech.), Slug (1:1,000, cat. no. 9585T; Cell Signaling Technology, Inc.) and β-actin (1:2,000, cat.no.GB12001, Servicebio.), GAPDH (1:2,000, cat.no. GB12002, Servicebio.); HRP-linked secondary antibodies (1:5,000; cat. no. 7074P2 and 7076S; Cell Signaling Technology, Inc.).