Transient Transfection of Reporter Constructs into Differentiated 3T3-L1 adipocytes and Luciferase Assay桹n day 4 of differentiation, 3T3-L1 adipocytes were transfected with 1 礸 of DNA using Lipofectamine Plus reagents (Invitrogen). Briefly, cells were washed with phosphate-buffered saline and incubated with reduced serum medium (Opti-MEM? modification of Eagle's minimal essential medium, Invitrogen) for 45 min at 37 癈. Pre-complex DNA-Plus-Lipofectamine^TM reagent was prepared according to the manufacturer's protocol and added into each well for transfection. After overnight transfection, cells were replenished with 10% fetal bovine serum Dulbecco's modified Eagle's medium and incubated for another 36?8 h for expression. For monitoring transfection efficiency, pRL-TK vector (containing Renilla luciferase, Promega) was co-transfected. Cells co-transfected with various UTR-pGL3 constructs and pRL-TK vectors were harvested in lysis buffer, and activities of firefly and Renilla luciferase were measured by using the Dual-Luciferase assay kit (Promega) in a luminometer (Turner Designs, Sunnyvale, CA). The reporter activity was expressed as arbitrary LUC units (firefly/Renilla).