Lysozyme chloride from chicken egg white

Genomic DNA was extracted from each strain as previously described^{50 (link)}. Briefly, the bacterial cells cultured in BHI were collected and incubated with 62.5 μl of lysozyme chloride from chicken egg white (2.0 mg/ml; Sigma-Aldrich Co., St. Louis, MO, USA) and 0.25 μl of lysozyme hydrochloride from chicken egg white (10 mg/ml; Fujifilm Wako Pure Chemical Industries, Osaka, Japan) for 90 min at 37 °C. For genomic DNA extraction, the samples were incubated in 600 μl of Cell Lysis Solution (Qiagen, Düsseldorf, Germany) at 80 °C for 5 min, followed by addition of 3 μl of RNase A (10 mg/ml; Qiagen) and incubation at 37 °C for 30 min. Next, 200 μl of Protein Precipitation Solution (Qiagen) was added and vortexed vigorously for 20 min, followed by centrifugation at 10,000 × g for 3 min. The supernatant was combined with 600 μl of isopropanol (Fujifilm Wako Pure Chemical Industries) and centrifuged. The precipitate was resuspended in 70% ethanol (Fujifilm Wako Pure Chemical Industries), centrifuged, combined with 100 μl of DNA Hydration Solution (Qiagen), and stored as a DNA extract.