PCR Amplification and Sequencing of rpoB Gene

The genomic DNAs of tested SCVs and corresponding parental strains were extracted from fresh bacterial colonies using the Biospin Bacterial Genomic DNA Extraction kit (Bioflux, Tokyo, Japan). The rpoB gene was identified by polymerase chain reaction (PCR) using the rpoB forward primer −5ʹ-TTATGCTGCACCTTCGTG-3ʹ and rpoB reverse primer −5ʹ-CAAGTGCCCATACCTCCCATC-3ʹ. An annealing temperature of 50°C was used for PCR reactions, and the extension time was set (1 min/1 kb). Positive PCR products were sent to Shanghai BGI Technology Co. (Shanghai, China) for sequencing. The sequences obtained were analyzed using the BLAST program (www.ncbi.nlm.nih.gov/BLAST).