Regulated luciferase expression from self-contained vectors in transfected cells. (a) Comparison of regulated luciferase expression from pGTRTL (2 μg) (white squares) and pGTLMIK (2 μg) (black squares) in transiently transfected DBA/1 embryonic fibroblasts with the temperature-sensitive large T antigen (DTF) cells (2 × 105 cells per well). Cells were grown in doxycycline (Dox)-free or Dox-containing media (concentration range of 1 pg/ml to 1 μg/ml). After 24 hours, luciferase activity was measured in cell lysates. (b) Fold induction of luciferase expression from pGTRTL (white bars) and pGTLMIK in response to Dox was determined in DTF, A431, and 293T transiently transfected cells. Cells were transfected as in (a), and fold induction was calculated by dividing the Dox-induced value by the non-induced value for each transfection. (c) Regulated expression of luciferase from pGTLMIK (1 μg) transfected into the mouse myoblast cell line C2C12 (1 × 106 cells) using the amaxa system. Transfected cells were split between the wells of a six-well plate and were either non-induced or induced with Dox (1 μg/ml) for 24 hours. In all panels, luciferase measurements were standardised for protein content and each mean value or calculated value was obtained from triplicate transfections. Vertical lines in (a) and (c) represent standard error of the mean. RLU, relative light units.