Dual-luciferase assay
HEK293T cells were spread to 96 well plates at the concentration of
1×104 cells per well. After 24 hours, HEK293T cells
were co-transfected with dual-luciferase reporter vector
(pmirGLO-Wt-circ or pmirGLO-Mt-circ, pmirGLO-Wt-3’UTR or
pmirGLO-Mt-3’UTR) and miR-204-5p mimics or negative control (miR-NC)
using the Lipo-fectomine 3000 transfection reagent (Invitrogen, USA),
respectively. After 48h of incubation, firefly and Renilla luciferase
activities were measured using a dual-luciferase reporter assay system
(Promega, USA) according to the manufacturer’s instructions.