Transient cotransfection analysis of LF89 and expression plasmids for C/EBPα, β, and ε in 32Dwt18 cells.

32Dwt18 cells were transiently cotransfected with an LF gene promoter fragment spanning −89 bp (LF89) harboring a C/EBP site cloned into the promoterless luciferase reporter pGL3-Basic plasmid (10 μg), and expression plasmids for C/EBPα, β, and ε (5 μg each), individually or pairwise. A pCMVβgal expression plasmid (2 μg) was included in each transfection to normalize for transfection efficiency. The total concentration of DNA per transfection was maintained at 22 μg by the addition of salmon sperm DNA where necessary. Transfected cells were harvested 24 hours after transfection and assayed for luciferase and β-galactosidase activity. Normalized luciferase values have been represented as a fold increase of luciferase activity over LF89 alone. Mean ± SE for 3 experiments performed in duplicate have been illustrated.