Gut microbial metagenomic DNA extraction and microbial diversity analysis
Experiment 1: Twelve 6 week old mice were fed a basal diet for 2 weeks for acclimatization, then randomly to Control (Con: standard chow diet, 10% fat, 0.1% choline) with sterile carboxymethyl cellulose (CMC) slurry around the right maxillary second molar (M2) or Pg group (Pg: same diet) with periodontitis induced by ligating the right maxillary M2 and applying CMC slurry containing Pg. At week 8, the mice were intraperitoneally administered pentobarbital sodium at a dose of 50 mg/kg for preliminary anesthesia, and then subjected to cervical dislocation to confirm euthanasia. The plasma, cecal contents, small intestine, large intestine, and liver tissues of the mice were harvested for subsequent analysis.Experiment 2: Fifteen 6 week old mice were fed a basal diet for 2 weeks for acclimatization, then randomly divided into three groups: Con (standard chow diet and sterile CMC slurry), Choline (standard chow diet supplemented with 1% choline and sterile CMC slurry), or Choline + Pg (standard chow diet supplemented with 1% choline and CMC slurry). Applications were performed five times per week for 6 weeks. All mice were euthanized humanely at week 8 for tissue sample collection.