Then, the sample was dialyzed against 10 mM sodium phosphate (pH 6.3) buffer overnight.
Subsequently, the sample was dialyzed against deionized water using a 1,000-Da 1,000-Daembrane (Thomas Scutofffic, Swedesboro, NJ, USA).
The sample was dialyzed with dialysis bags (D 0530, 12.4 kDa; Sigma-Aldrich Corporation, St. Louis, MO, USA) overnight against the same buffer and was filtered through a 0.45-μm polyethersulfone membrane (25 mm in diameter; Whatman, Maidstone, England) syringe filter.
After cleavage of the affinity tag with PreScission protease, the sample was dialyzed against buffer A (20 mM Tris-HCl, pH 8.0, 120 mM NaCl, 1 mM DTT) and passed again over the affinity resin to remove remaining uncleaved species.
Purified PGNpol from S. aureus SA113 Δlgt was incubated with 30 mM sodium-meta-periodate (NaIO4, Merck, Hohenbrunn, Germany) on ice in the dark for 30 min. To remove the NaIO4 the sample was dialyzed (MWCO 8 kDa, 0.1 M sodium acetate, pH 5.5) over night.
The sample was dialyzed overnight in 1 L HEPES-EDTA buffer containing 0.03% w/v saponin.
The sample was dialyzed against 10 mM Tris-HCl (pH 8.0) and 200 mM NaCl.
After incubation at 37°C for 1 hr, the sample was dialyzed against RB buffer at 4°C overnight.
The sample was dialyzed against 0.01 M 2- N-morpholino) ethanesulfonic acid (MES) buffer (pH 6.0) (Dojindo Laboratories, Kumamoto, Japan) at 4°C.
The sample was dialyzed against ChIP buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM TrisHCl, 167 mM NaCl).
