Fig. 3

Dual luciferase reporter assays verify that miR-720 regulates its targets by binding directly to their 3′-UTR. a The diagram shows that 3′-UTR of Rab35 was cloned downstream into the open reading frame of the pcDNA3.0-Vector, and also displays the detailed binding sites or the mutated binding sites of the 3′-UTR for miR-720 targeting; b The results of analysis indicate that the relative luciferase activities of the pMIR-REPORT vector containing wide-type 3′-UTR of INSIG1, KCTD15, METTL2B, NRXN3, Rab35, RASAL2, or TET2 could be decreased by upregulating miR-720; Firefly luciferase activity was normalized to Renilla luciferase activity. The relative luciferase expression level is expressed as the mean ± standard deviation (SD). Three independent experiments were performed, and representative data are shown. *p < 0.05 shows the significant difference. c Normalized luciferase activity in cells transfected with reporter vector containing mutant type 3′-UTR of KCTD15, INSIG1, or RAB35 could not be affected by overexpression of miR-720 compared with cells transfected with wild type 3′-UTR. The relative luciferase expression level is expressed as the mean ± standard deviation (SD). Three independent experiments were performed, and representative data are shown. *p < 0.05 shows the significant difference