Lipopolysaccharide and O-polysaccharide Extraction

LPS extraction was performed as described previously (Sun et al., 2018 (link)). Briefly, the cells were collected by centrifugation and resuspended in ddH₂O. Then, an equal volume of 90% phenol was added, and the mixture was shaken vigorously at 68°C for 30 min. After centrifuging at 7,000 g for 20 min at 4°C, the supernatant was collected. Phenol was removed from the supernatant using a dialysis bag in ddH₂O for 2 days. Then, DNase (5 μg/mL; Solarbio, Beijing, China), RNase (1 μg/mL; Solarbio) and proteinase K (20 μg/mL; Solarbio) were sequentially added to the dialyzed sample. After incubating at the optimal temperature, the solution was placed in a boiling water bath for 10 min and then centrifuged at 7,000 g for 10 min to obtain LPS. To obtain OPS, glacial acetic acid was added to the LPS solution with a final concentration of 1% and incubated in a boiling water bath for 90 min. The pH was then adjusted to 7.0 with NaOH. Finally, the mixture was centrifuged at 40,000 g for 5 h, and the supernatant was collected.