Fig. 3

Time course characterization of the pFREE plasmid-curing system. Curing of pZ-plasmids expressing GFP with either pSC101 (pZS-GFP, green), ColE1 (pZE-GFP, red) or p15A (pZA-GFP, blue) replicon. The solid lines indicate induced cultures with rhamnose (Rham) and anhydrotetracycline (aTc), whereas the dashed lines refer to non-induced (Ø). Plating was performed at induction time (0) and 3, 7, 11 and 24 h after induction. Between 100 and 150 colony forming units (CFUs) were counted from each replicate and the ratio between fluorescent and non-fluorescent cells were determined. The percentage of plasmid-carrying cells is depicted. Of the non-fluorescent and tested cells, all had lost the pFREE plasmid after 24 h. Data points represent mean value of three biological replicates with error-bars showing standard deviation. Representative LB agar plates for pZE-GFP with equal number of cells plated with cultures induced with rhamnose and aTc (top) and non-induced (Ø) (bottom) of the pFREE system after 24 h