Glycerol stocks of the E. coli cell cultures, which were prepared beforehand [30], were diluted to final concentrations of 500 ~ 1000 cells/mL in test tubes. Dilution was performed with either the rich medium LB (Luria-Bertani, Sigma) or the minimal medium M63 [30], corresponding to the agar plates used for analysing colony growth. Then, 100 μL of each diluted cell culture was plated on agar plates (1.5% agar), and the plates were incubated at 37 °C in an incubator (THS030PA, ADVANTEC). A total of nine replicates were performed for each condition, and colony growth on the agar plate was imaged by CCD (charge-coupled device) photography. The agar plates were photographed with a high-sensitivity monochrome CCD camera of a gel imager (AE-6932GXES print graph, ATTO Co., Ltd.). The brightness, contrast, saturation, hue and sharpness were set at 50, 73, 50, 50 and 0%, respectively. The OSD (on-screen display) time and exposure time were set at 10 s and 1 s, respectively. Temporal changes in colony growth were observed at 12, 18, 24, 36 and 48 h for the LB medium and 24, 36, 48, 60, 72, 84, 96 and 108 h for the M63 medium. The images were saved as TIF files and subjected to the computational analysis described below. A total of 242 plates and 638 images (photos) were analysed in the present study.

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