human blood virome Metagenome

In 2018, 16 blood samples were collected from eight patients with traumatic sepsis in the Affiliated Hospital of Jiangsu University in Zhenjiang at two different times (before and seven days after therapy) for viral metagenomic investigation. All samples were collected and shipped on dry ice. 400 uL of blood should be pipetted into a 1.5 ml centrifuge tube together with 400 uL of PBS, which should be vortexed for 5 minutes at room temperature before centrifuging at 14,000 g for the same amount of time. The supernatant of each sample was transferred into a new 1.5-ml centrifuge tube. Supernatants were filtered through a 0.45-mm 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 (non-viral encapsidated) nucleic acids. Remaining total nucleic acid was then isolated using QIAamp MinElute Virus Spin Kit according to manufacturer's protocol. Thirty-seven libraries were then constructed using Nextera XT DNA Sample Preparation Kit (Illumina) and sequenced using the HiSeq Illumina platform with 250 bases paired ends with dual barcoding for each pool.