Southern Blot Localization of blaKPC-2 Gene

The plasmid location of the bla_KPC–2 gene was determined by Southern blot experiments according to the previous study (Wang et al., 2020 (link)). Briefly, whole chromosomal DNA was digested with S1-nuclease (TaKaRa, Japan). The digested fragments were electrophoresed on a CHEF-mapper XA pulsed-field gel electrophoresis (PFGE) system (Bio-Rad, United States) for 18 h at 14°C. The DNA fragments were transferred to a positively charged nylon membrane (Millipore, United States) and then hybridized with a digoxigenin-labeled bla_KPC–2-specific probe. The fragments was detected by an NBT/BCIP color detection kit (Roche, Germany). The Salmonella enterica serotype Braenderup H9812 was used as the size marker.