Figure 4. Tm_2–8 − 0.53 × miTm_x–y value-dependent different silencing activities of miRNAs.

The indicated concentration of synthetic miRNA, psiCHECK-SM construct (10 ng), and pGL3-control (1 µg) were simultaneously transfected into HeLa cells, and Renilla luciferase activity / firefly luciferase activity was determined 24 after transfection. The results of luciferase reporter assays using (a) miR-302a-3p/372/373-3p/520c-3p family miRNAs with same seed sequences but different structures, or (b) miR-376a-2-3p/499-5p/628-3p with same composition of nucleotides in the seed regions. The value of Tm_2–8 − 0.53 × miTm_1–5 of each miRNA was represented in parenthesis. The structures of psiCHECK-SM and psiCHECK-CM reporters are same as those shown in Figure 1a and 1b. The black bar indicates the result of psiCHECK-SM. The white bar, the result of psiCHECK-CM. The ordinate represents relative luciferase activity (%) and the abscissa represents miRNA concentration. Yellow indicates the common seed sequence in miR-302a-3p/372/373-3p/520c-3p family miRNAs, blue indicates the seed region with same nucleotide compositions in miR-376a-2-3p/499-5p/628-3p miRNAs.