title: Viral metagenome in feces from rhesus in China
design: In 2015, a fecal sample was collected from rhesus.The sample was resuspended in 10 volumes of phosphate-buffered saline (PBS) and vigorously vortexed for 5 min. Five hundred microliters was collected after centrifugation (10 min, 15,000  g) and filtered through a 0.45-m filter (Millipore) to remove eukaryotic and bacterial cell-sized particles. The filtrate enriched in viral particles was treated with a mixture of DNases (Turbo DNase from Ambion, Baseline-ZERO from Epicentre, and benzonase from Novagen) and RNase (Fermentas) to digest unprotected nucleic acid at 37C for 90 min. Remaining total nucleic acid was then isolated using QIAamp MinElute Virus Spin Kit (Qiagen) according to manufacturer's protocol. Viral cDNA synthesis was performed using SuperScript III reverse transcriptase (Invitrogen) according to the manufactures instructions. The second strand of cDNA synthesis was performed by incubation of reverse transcribed (RT) products with Klenow Fragment DNA polymerase (New England Biolabs). Library was 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.