Fig. 2

Phenotype of the lux-tn66 transposon strain. a Growth and luminescence of the lux-tn66 transposon strain on solid MS medium in comparison to the WT-lux strain (MG03), TopA-depleted lux strain (MG04) and the negative control—wild type strain with empty pFLUXH vector (MG01). Left panel: plate view (after 5 days of growth), right panel: luminescence intensity (after 48 h). b Luminescence of mutant reporter strains after 24 h of growth in liquid 79 medium compared to the WT-lux (MG03) and the negative control—wild-type strain with empty pFLUXH vector (MG01). c Western blot analysis of TopA protein level (left panel) and the relative transcription of the native topA gene, as well as luxC reporter gene, in the mutant lux-tn66 strain determined using RT-qPCR analysis performed on 24-h 79 medium cultures, compared to the WT-lux strain (right panel). d The growth curves of the lux-tn66 strain (79 medium, Bioscreen C, measurements every 20 min) compared to the WT-lux (MG03) and TopA-depleted lux strain (MG04), as well as to MG04 with restored TopA protein level (after induction with 0.5 µg/ml thiostrepton). e Supercoiling density of the reporter plasmids pWHM3Hyg or pWHM3Spec isolated from the transposon mutant lux-tn66 derivative (lux-tn66_RP) strain, the wild-type strain derivative (MS10) and the TopA-depleted (MS11) strain (representative image of two independent experiments). The figure shows topoisomers detected in agarose gel as well as band intensity measurements performed using ImageJ software. f The level of gyrB transcript in lux-tn66 strain determined using RT-qPCR analysis performed on 24-h 79 medium cultures, compared to the WT-lux strain (MG03)