Dual Luciferase Assay for circRNA-miRNA Interaction

The dual luciferase assay kit (cat. no. KGAF040) was purchased from KeyGEN BioTECH (Nanjing, Jiangsu, China), and all the plasmids were constructed by Sangon Biotech Company (Shanghai, China). Reporter assays were performed using the dual luciferase assay system as described previously [36 (link), 37 (link)]. The luciferase reporter constructs were generated from the miRNA target expression vector pmirGLO (Promega, Madison, WI, USA). Full-length binding-site-mutant and wild-type circRNA_37492 constructs, truncated mutant and wild-type 3’UTR of fibrinogen beta chain (Fgb) gene were individually co-transfected with miR-7682-3p mimics or Control into BUMPT cells for 48 h. Luciferase activities were then measured using a Synergy™ HTX multi-mode reader (Biotek, Winooski, VT, USA) and normalized according to renilla luciferase activity.