Figure 5

Ht1080 populations transduced by either MOV-scT6 or MOV-scT6cA were enriched via FACS. From both initial populations the fractions of cells inducible at 30 ng/ml Dox (circles) were enriched close to homogeneity. (A) Fluorescence based analysis of induction profiles of initial and enriched populations. The percentage of inducible cells (R2) at different Dox concentrations is inserted into the blots. (B) Northern blot analysis of initial (pool) and enriched (T6+ or T6cA+) populations. Total RNA was extracted from cells induced for 96 hrs with 30 ng/ml Dox and analysed after separation on 1.2% Agarose-MOPS-formaldehyde gel. Detection was performed with biotinylated probes against the M2-transactivator or GAPDH. Because of subsequent development of the blots, the M2 signals were partially visible in the blot probed with GAPDH. Densitometric analysis was done on appropriate developed blots (NIH 1.57 software), the relative values obtained for MOV-scT6 cells in the off-state (pool -Dox) was set to 1. The signal intensity of the T6cA+ could not be triggered into a linear range of signal intensity (*).