Pmir report firefly luciferase transfection control vector

Prostate cells were co-transfected with CD9 3’UTR or empty 3’UTR renilla reporter vector (100ng) (SwitchGear Genomics, California, USA) and 10ng pmiR-Report firefly luciferase transfection control vector (Promega) using Lipofectamine LTX (Invitrogen, California, USA). Luciferase activity was assessed 24h post-transfection using the dual luciferase assay kit (Promega) and a Biotek Synergy 2 plate reader. Results are expressed as luminescence intensity normalized to firefly transfection control vector and then empty 3’UTR vector. For assessment of specific miRNA, CD9 3’UTR luciferase reporter vector and 50mM miRNA mimics (Bioneer Pacific, VIC, AUS) were co-transfected into HEK293FT cells.

Cells were seeded in 96 well plates and transfected with the CD9 3’UTR or empty 3’UTR renilla reporter vector (SwitchGear Genomics, Menlo Park, CA, USA) and pmiR-Report firefly luciferase transfection control vector (Promega) using Lipofectamine LTX (Invitrogen, Carlsbad, CA, USA). Renilla and firefly luciferase activity were measured as previously described, using the Dual-Glo Luciferase assay system (Promega) [19 (link)].