Microbial community characterization of fermented stool
Aliquots (1 ml) of fermented stool samples were centrifuged (20,784 g, 15 min), and the supernatant discarded. Automated DNA extraction of the pelleted material was performed using a QIAcube Connect instrument with the QIAamp PowerFecal Pro DNA kit (Qiagen, Germantown, MD) per manufacturer instructions. Recovered genomic DNA was used as a template for amplification of the V4 variable region of microbial 16 S ribosomal RNA (rRNA) genes using primers 515 F (GTGCCAGCMGCCGCGGTAA) and 806 R (GGACTACHVGGGTWTCTAAT) (custom primers from Integrated DNA Technologies, Coralville, IA). Amplicons were generated using a two-stage PCR amplification protocol