2.4. Microbial DNA Extraction and 16S rRNA Sequencing Analysis
Total microbial genomic DNA was extracted from all fecal samples using TIANamp Stoll DNA Kit (TIANGEN Biotech Co., Ltd., Beijing, China; Catalog No.: DP328) according to the manufacturer’s protocol. The quality and concentration of total extracted DNA were identified by 1.5% agarose gel electrophoresis and Nanodrop spectrophotometer (Thermo Fisher Scientific, Inc., MA, USA). The subsequent PCR amplification of the V4 region of 16S rDNA, sequencing library construction, and high-throughput sequencing (Illumina NovaSeq/MiSeq platform) were performed at Shanghai Personalbio Technology Co., Ltd. (Shanghai, China). The raw reads were deposited in the Sequence Read Archive (SRA) database of NCBI (PRJNA903224).Microbiome bioinformatics were performed with QIIME 2 2019.4 [