We have improved a marker system to examine the genetic diversity in Xanthomonads species by using whole genome sequence analysis and repetitive regions in the bacterial genome. Specifically, we amplified microsatellite regions using PCR, and the resulting fragments were digested with specific enzymes. These fragment profiles served as molecular markers. Consequently, a more precise marker for bacterial identification was developed by combining the microsatellite and RFLP methods to differentiate Xanthomonads species. In Addition, we analysed the ancestral ordering of various Xanthomonas species\' genomes available in NCBI using Progressive Mauve Alignment. The data revealed unique collinear regions characteristic of Xanthomonas species. These regions were also associated with cut genomic fragments used as markers, enabling the discrimination of Xanthomonas species infecting tomato and pepper. We propose that these findings contribute to understanding the genetic diversity of Xanthomonas and rapidly diagnosis.