Research Paper Volume 12, Issue 19 pp 19233—19253

CDR1as sponges miR-7-5p in HB cells. (A) Schematic illustration showing overlapping target miRNAs of CDR1as that were predicted by the Circular RNA Interactome, Circbank, and circMIR; (B) Schematic diagram for the pull-down assay using biotinylated microRNA; (C) The relative level of 12 miRNA candidates in the HepG2 and HUH6 lysates was detected by real-time PCR. Multiple miRNAs were pulled down by CDR1as, and miR-7-5p was pulled down by CDR1as in two cell lines; (D) Potential binding sites between CDR1as and miR-7-5p; (E, F) The possible binding sites of CDR1as with miR-7-5p were predicted by RegRNA2.0, (E) Graph of predicted RNA secondary structure. The yellow region indicates the RNA fold predicted structure of the motif. (F) RNA fold reliability of pair probabilities; (G) The biotinylated wild-type miR-7-5p (miR-7-5p biotin) or its mutant (NC-biotin) were transfected into HepG2 and HUH6 cells. After streptavidin capture, CDR1as levels were quantified by real-time PCR. GAPDH was used as a negative control; (H) Fluorescence in situ hybridization (FISH) showed the colocalization between CDR1as and miR-7-5p. Data are presented as the mean ± SEM of three experiments. *P < 0.05 (Student’s t-test).