BMI-1 is a direct target of miR-218. (A) Conservation of BMI-1 3′UTR binding site for miR-218 among different species. (B) The conserved miR-218 binding sequence or its mutated form was inserted into the C-terminal of the luciferase gene to generate pMIR-BMI-1-3′UTR or pMIR-BMI-1-mut-3’UTR, respectively. (C) miR-218 targets the wild-type but not the mutant 3′UTR of BMI-1. miR-218 was cotransfected with pMIR-REPORT (the empty vector) or reporters constructed containing either wild-type or mutant BMI-1 3′UTR into HCT116 cells. pRL-CMV was used as an internal control reporter (n = 3, mean ± SD). The luciferase activity assay was repeated three times in triplicate. *P < 0.05.