Figure 3.

RARRES3 could suppress EMT process while involving the up-regulation of β-catenin. A. Knock-down efficiency of candidate shRNA target sequences was determed by western-blotting. Both the 3 different shRNA vectors as well as shRNA control (scramble) vetor were transfected in CRC cell line HT-29. shRNA-RARRES3-2 was chosen as the best which was abbreviated for shRNA-RARRES3 in the following funcational exmperiment. B. The endogenous expression of RARRES3 in a panel of CRC cell lines was detected using immunoblotting. β-Tubulin was loading control. 80μg total protein was loaded per lane, which was separated by 10% SDS-PAGE followed by visualization with Western Breeze kit (Invitrogen, USA). C. Immunoblotting assay for HT-29 and SW-620 after knock-down and over-expression of RARRES3 respectively using retroviral shRNA and over-expression vector transfection for 96 hours.