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. 2019 Mar 26;9:70. doi: 10.3389/fcimb.2019.00070

Figure 2.

Figure 2

Depletion of DNA gyrase perturbs nucleoid morphology. (A) Effect of gyrase depletion on nucleoid organization. Morphology of 4′,6-diamidino-2-phenylindole (DAPI)-stained nucleoid of both control and gyr(-) was observed by confocal laser scanning microscopy using 100x oil objective under ultraviolet illumination. Each image is a representative of multiple fields from three different experiments. (B) Quantitative estimation of bacterial nucleoid decompaction. Status of DAPI-stained nucleoids of control and gyr(-) was analyzed by estimating median fluorescence intensity (MFI) of the dye using flow cytometry. Values were obtained from 10,000 events using FlowJo software (FlowJo LLC). FQ-treated Mtb was used as a reference control. Error bars represent SD from at least 3 replicates and statistical significance is determined by paired Student's t-test: **P < 0.01. (C) Scanning electron microscopy (SEM) analysis of bacteria. Shown are the SEM images of control and gyr(-) after 4 days of treatment with 20 ng/ml ATc. Each image is a representative of multiple fields from three different experiments. (D) Effect of gyrase depletion on bacterial cell length. Cell length of control and gyr(-) was determined under light microscopy using 100x oil objective. Shown are the mean cell lengths of ~300 cells from three different experiments. Statistical significance is determined by paired Student's t-test: ****P < 0.0001.