viral metagenome Metagenome

For viral metagenomic analysis, 665 fecal specimens were collected from 72 species of mammals which from several zoos in Hangzhou, Zhejiang Province in 2019. Before use, swab was put into 1.5 mL centrifuge tubes 500 uL PBS and vortexed for 5 min at room temperature. The supernatants were then collected after centrifugation (5 min, 14,000g). 665 sample supernatants were pooled into 69 pools. Supernatants were filtered through a 0.45um filter (Millipore) to remove eukaryotic and bacterial cell sized particles, and 200 uL of supernatant from each pool was then subjected to a mixture of nuclease enzymes to reduce the concentration of free (nonviral encapsidated) nucleic acids. Remaining total nucleic acid was then isolated using QIAamp MinElute Virus Spin Kit according to manufacturer's protocol. Eight libraries were then constructed using Nextera XT DNA Sample Preparation Kit (Illumina) and sequenced using the MiSeq Illumina platform with 250 bases paired ends with dual barcoding for each pool.