metagenome of cecal contents: adult male rat
Total genomic DNA was extracted from fecal samples using the E.Z.N.A. tissue DNA Kit (Omega Bio-tek, Norcross, GA, U.S.) according to manufacturers instructions. Concentration and purity of extracted DNA was determined with TBS-380 and NanoDrop2000, respectively. DNA extract quality was checked on 1% agarose gel. DNA extract was fragmented to an average size of about 300 bp using Covaris M220 (Gene Company Limited, China) for paired-end library construction. Paired-end library was constructed using TruSeqTM DNA Sample Prep Kit (Illumina, San Diego, CA, USA). Adapters containing the full complement of sequencing primer hybridization sites were ligated to the blunt-end of fragments. Paired-end sequencing was performed on Illumina HiSeq4000 platform (Illumina Inc., San Diego, CA, USA) at Majorbio Bio-Pharm Technology Co., Ltd. (Shanghai, China) using HiSeq 3000/4000 PE Cluster Kit and HiSeq 3000/4000 SBS Kit according to the manufacturers instructions (www.illumina.com). The readings were assembled and predicted according to the published method (5). The KEGG annotation was conducted using BLASTP (Version 2.2.28+) against the Kyoto Encyclopedia of Genes and Genomes database (6) (http://www.genome.jp/keeg/) with an e-value cutoff of 1e-5.