The cecal contents bacterial DNA of each sample was extracted using HiPure Stool DNA Kit B (Magen, Shanghai, China) following the manufacturer’s instructions. The DNA extractions were quantified by ultraviolet spectroscopy and amplified using universal primers of 341F (CCTACGGGNGGCWGCAG) and 806R (GGACTACHVGGGTATCTAAT) to target the V3–V4 domain of bacterial 16S rRNA. The amplicons were normalized, pooled, and sequenced on the Illumina GAIIx platform.
The raw Illumina fastq files were quality-filtered, de-multiplexed, and analyzed using quantitative insights into microbial ecology. Sequences with more than one ambiguous nucleotide or within correct barcodes or primers were removed. The Ribosomal Database Project classifier was used to classify tags into different taxonomies against Greengenes Database (version 20101006) with confidence threshold of 0.5. The software Mothur was used to cluster tags of more than 97% identity into operational taxonomic units (OTUs), and then the abundances of OTUs were calculated.
The raw Illumina fastq files were quality-filtered, de-multiplexed, and analyzed using quantitative insights into microbial ecology. Sequences with more than one ambiguous nucleotide or within correct barcodes or primers were removed. The Ribosomal Database Project classifier was used to classify tags into different taxonomies against Greengenes Database (version 20101006) with confidence threshold of 0.5. The software Mothur was used to cluster tags of more than 97% identity into operational taxonomic units (OTUs), and then the abundances of OTUs were calculated.
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