gut of Mouse
Fecal genomic DNA was extracted from the fecal samples with the QIAamp DNA Stool Mini Kit (Qiagen, Hilden, Germany), according to the manufacturers protocol. The bacterial genomic DNA was used as the template to amplify the V3V4 hypervariable region of the 16S rRNA gene with the forward primer (5-CCTACGGGNGGCWGCAG-3)and the reverse primer (5-GACTACHVGGGTATCTAATCC-3). Each sample was independently amplified three times. Finally, the PCR products were checked by agarose gel electrophoresis, and the PCR products from the same sample were pooled. The pooled PCR product was used as a template, and the index PCR was performed by using index primers for adding the Illumina index to the library. The amplification products were checked using gel electrophoresis and were purified using the Agencourt AMPure XP Kit (Beckman Coulter, CA, USA). The purified products were the indexed in the 16S V3V4 library. The library quality was assessed on the Qubit@2.0 Fluorometer (Thermo Scientific) and Agilent Bioanalyzer 2100 systems. Finally, the pooled library was sequenced on an Illumina Novaseq6000 Sequencer for generating 2250 bp paired-end reads.