Figure 2A–B. Direct inhibitory effect of miR-25 and miR-92a on Ins1. (A) Dual-Luciferase ® reporter assay quantitation of the effects of anti- or pre-miR-25 or miR-92a interaction with the wild-type and mutated 3′ UTR of Ins1. For the mutated construct, the target sequence 5′-TGCAAT-3′ for the miRNA seed region was mutated to 5′-GCGCCG-3′. The plasmid constructs together with anti or pre miR-25 or miR-92a were co-transfected independently into INS-1 cells. Relative luminescence for luciferase gene activity in treated samples were obtained 48 h post-transfection by normalizing the values against control plasmids transfected with corresponding anti- or pre-scrambled miRNAs. The luminescence for luciferase activity of the controls is considered to be 100%. All transfections of miRNAs were done at 30 nM concentration. Data are presented as mean ± SEM (n = 3) against control. Statistically significant differences are tested using t-test at *p < 0.05 significance. (B) Fold change in the expression of preproinsulin, miR-25 and miR-92a in pancreatic islets and INS-1 cells subjected to increasing glucose concentration (G10, 15, and 20 mM). Data presented as mean ± SEM (n = 3) against control at 5 mM glucose concentration. Statistically significant differences are tested using t-test at p < 0.05 significance. *p < 0.05, **p < 0.01.