Visualizing Xanthomonas citri Chromosome

The initial cultures of X. citri wt and X. citri pMAJIIc-envC were prepared by cultivating bacteria in 5.0 mL of NB medium for approximately 16 h at 30 °C and 200 rpm. The cultures were then diluted to an OD 600 nm of 0.1 using fresh NB medium for a final volume of 5.0 mL and subsequently cultivated under the same conditions until an OD 600 nm of 0.3 was reached. At this point, arabinose was added to the medium to a final concentration of 0.05%, and the cultures were maintained at 30 °C and 200 rpm. After a minimum of 2 h of induction, 5 µL drops of cell cultures were placed on agarose-covered microscope slides for direct microscope observation [21 (link)]. For chromosome visualization, X. citri wt, ΔenvC and ΔenvC pMAJIIc-envC cells were cultivated under the same conditions described above and stained with DAPI using the protocol described in Ref [22 (link)]. Bacterial visualization was conducted using an Olympus BX61 microscope equipped with a monochromatic OrcaFlash2.8 camera (Hamamatsu, Japan) and TxRed and DAPI filters. Data collection and analysis were carried out using the CellSens Version 11 software (Olympus). Statistical analyses were conducted using GraphPad Prism version 6.