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. 2022 May 12;50(22):12601–12620. doi: 10.1093/nar/gkac332

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© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com

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Figure 2. — SOS induction, DNA repair, DNA damage, and mortality rates of cells growing in LB medium. (A) Expression of the SOS induction reporter P_recA-yfp in cells having different rrn operon copy numbers. Each data point represents the mean value (± SD) from three independent experiments. (B) Duration of the lag phase of different mutant derivatives of rrnC⁺ strain having different capacities to repair DNA lesions calculated from growth curves (see Supplementary Figure S1B). Each dot represents the mean value (± SD) from three independent experiments. Asterisks show significant differences compared to the strain with 1 rrn. (C) DNA damage detected in the lag phase cells using TUNEL assay and flow cytometer. Each bar represents the mean value (± SD) from three independent experiments. Asterisks show significant differences between strains. (D) Detection of double-strand breaks. Lag phase cells carrying a Gam-GFP reporter fusion were analysed using a fluorescence microscope. Inserted photographs show Gam-GFP foci in the cells having 1 rrn operon. Each bar represents the mean value (± SD) from three independent experiments. Total of 1,329 and 1,606 cells having 7 and 1 rrn were analysed, respectively. Asterisks show significant differences compared to WT. (E) Cell death measured during growth in the LB medium in populations of 7 rrn and 1 rrn strains (upper panel). Dead cells were detected using propidium iodide (PI) staining and flow cytometer (lower panel). Each result represents the mean value (± SD) from three independent experiments. (F-I) Growth of individual cells in the ‘mother’ machine microfluidic device in LB medium. Each panel represents growth of an individual mother cell. (F) 99% of WT cells with 7 rrn operons start growing and dividing after short lag phase. (G) 70% of cells with 1 rrn operon start growing and dividing after longer lag phase. (H) 10% of cells with 1 rrn operon undergoing transient crisis, i.e. filamentation, after which it resumes growth. (I) 20% of cells with 1 rrn operon increase in size, divide a few times and then die. The presented results are representative of three independent experiments. t-test: *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001).