Strains and culture conditions

Bacterial strains used in this study included 10 strains of Xoc and 25 strains of Xoo from diverse geographic location as well as 5 strains of other phytopathogenic bacteria. These strains were cultured on peptone sucrose agar (PSA) medium at 28℃ for 2 days.


DNA extraction and PCR amplification

 Genomic DNA from the bacterial strains was extracted using BacteriaGen DNA Kit (CWBio, China) following the manufacturer’s protocol. The quantity and purity of genomic DNAwas measured using NanoDrop ND-1000 spectrophotometer (Nano-Drop Technologies, Wilmington, DE)

The 120 candidate primers were selected randomly from GMGM. For single primer reactions, 50 ng of template bacterial genomic DNA was used. For amplification of fragments directlyfrom bacterial cells, 1µl of a cell suspension (107 CFU/ml) was used per reation. The conventional PCR reaction mixture contained 1.5µl of 2.5 mM dNTPs, 2.5 µl of 10× buffer, 0.02units of Taq  polymerase, and 0.3µl of each 10 µM primer in a total volume of 25 µl. The PCR protocol included an initial denaturing step at 94°C for 3 min; followed by 35 cycles of 94°Cfor 30 s, 64°C for 30 s, and 72°C for 30 s; and a final extension at 72°C for 7 min. Products were analyzed by separation in 2% agarose gels (2% Tris-boric acid-EDTA [TBE] buffer),stained with ethidium bromide, and visualized under UV light.