Fig. 3.

miR-122 directly binds to the 3′-UTR of PEG10 transcript. a Schematic illustration of the luciferase reporter vector pmiR-GLO-PEG10-3′-UTR. Nine putative miR-122 binding sites were identified by bioinformatic analysis (top) at positions 64, 102, 564, 934, 1310, 1735, 2310, 2403 and 3420. Four deletion fragments and the full-length 3′-UTR of PEG10 were cloned downstream of the luciferase gene at the NheI and XhoI sites. An expanded view of the seed region of miR-122 in the PEG10-3′-UTR is shown. b Identification of the miR-122 target region in the 3′-UTR of PEG10 transcript. 293T cells were co-transfected with miR-122S and the negative control (Vector) along with pmiR-GLO-PEG10-3′-UTR F1, pmiR-GLO-PEG10-3′-UTR F2, pmiR-GLO-PEG10-3′-UTR F3, pmiR-GLO-PEG10-3′-UTR F4, pmiR-GLO-PEG10-3′-UTR F5, or pmiR-GLO-miR122 PTS; luciferase activity was determined at 48 h post-transfection. Renilla luciferase served as the internal control. Data represent the mean of three independent experiments, and error bars represent SD. c Schematic illustration of putative miR-122 binding site in the 3′-UTR of PEG10 transcript. For pmiR-GLO-PEG10 3′-UTR MTS, seven nucleotides, ACACTCC, were replaced with AGTGAGG. MTS mutation. d Identification of the miR-122 target sequence in the 3′-UTR of PEG10 transcript. 293T cells were co-transfected with either miR-122S or empty vector along with pmiR-GLO-PEG10-3′-UTR TS, pmiR-GLO-PEG10-3′-UTR MTS, or pmiR-GLO-miR122 PTS constructs