the interior intestinal 16S microbiome sequencing of control group in 60-days trial
The total genomic DNA of intestinal specimen was extracted by HiPure Stool DNA extraction kit (Magen, China) in accordance of manufacturers protocol. Then, the extracted DNA was used as DNA template for the PCR amplification of the V3-V4 hypervariable regions of prokaryotic 16S rDNA gene in the following PCR cycles: 94 C for 2 min, followed by 30 cycles at 98 C for 10 s, 62-66 C for 30 s, and 68 C for 30 s and a final extension at 68 C for 5 min. The primer pairs used were 341 F and 806R with sequences 5 CCTACGGGNGGCWGCAG 3 and 5 GGACTACHVGGGTATCTAAT 3.  Purified amplicons were then pooled in equimolar and paired-end sequenced (250 bp) on Illumina Novaseq 6000 platform.