The CMV promoter is down regulated in mouse embryonic CCE cells and during differentiation into mesoderm. A-D. CCE cells were transfected with pEGFP-N1 and subjected to G418 selection 48 hours later. Cells expressing GFP are obvious 2 days after transfection (A, B) while few GFP positive cells remain 7 days post transfection (C, D). E-F. CCE cells were transfected with pEGFP-N1 and differentiation was initiated 2 days post transfection. No GFP+ cells were observed on day 5 of differentiation to the Flk1 stage (pool of containing both Flk1+ and Flk1- cells) when transient transfectants were used. G-I. CCE cells were transfected with pEGFP-N1 and placed under G418 selection 2 days post transfection. Undifferentiated cells (G, I) and cells differentiated to the Flk1 stage (H, I) were analysed by FACS for GFP expression 14–19 days post transfection. The data suggest that less than 50% of the undifferentiated cells express GFP, while this percentage declines during differentiation to the Flk1 stage. Similar levels of GFP expression were obtained during differentiation of cells transfected with pLK444-GFP.