Phenotypes of ρ0 cells, ρΔ cells, and parental MB35 cells. (a) Membrane potential of mitochondria. The membrane potential of mitochondria was monitored by staining of cells with the cationic dye MitoTracker Orange CMTMRos (MTK) at a concentration of 0.5 μM for 30 minutes. For comparison, we show here the membrane potential of mitochondria in ρΔ cells with a reduced amount of mtDNA, which were produced by incubating LpCEco cells in Tet-minus growth medium for a relatively short period (8 hours). Rather stronger staining by MitoTracker Orange is observed in many of mitochondria contained in the ρΔ cells compared to MB35 cells. In contrast, the mitochondrial membrane potential is completely vanished in ρ0 cells that were produced by incubating LpCEco cells in Tet-minus growth medium for a relatively long period (48 hours). Bars, 20 μm. (b) Development of starved MB35 cells and ρ0 cells on agar. MB35 cells and ρ0 cells were washed twice in BSS and plated on 1.5% non-nutrient agar at a density of 5 × 106 cells/cm2. This was followed by incubation for the indicated times at 22°C. Bars, 0.5 mm.