canine metagenome Genome sequencing. canine metagenome

For viral metagenomic analysis,from 2020 to 2021, 780 fecal samples of healthy dogs, 170 fecal samples of diseased dogs were collected from Yuzhu Tibetan Autonomous Prefecture of Qinghai Province, China and 840 fecal samples of healthy dogs and 180 fecal samples of diseased dogs were collected from Gulo Tibetan Autonomous Prefecture of Qinghai Province, China, for a total of 1970 samples. Samples were randomized faccording to region and level of health.The fecal samples were individually homogenized with a mortar and pestle, resuspended in 1 mL DPBS, and then frozen and thawed rapidly three times on dry-ice. The supernatants were then collected after centrifugation (10 min, 15,000 g). 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. One hundred and ninety-seven libraries were then constructed using Nextera XT DNA Sample Preparation Kit (Illumina) and sequenced using the NovaSeq 6000 Illumina platform with 250 bases paired ends with dual barcoding for each pool.

本数据集用于病毒宏基因组学分析。2020至2021年间，研究团队从中国青海省玉树藏族自治州采集健康犬粪便样本780份、患病犬粪便样本170份，另从中国青海省果洛藏族自治州采集健康犬粪便样本840份、患病犬粪便样本180份，总计1970份样本。所有样本按采集区域与健康状况完成随机化分组。粪便样本均采用研钵与研杵单独均质化，重悬于1mL Dulbecco's磷酸缓冲盐溶液（DPBS）中，随后置于干冰上快速冻融三次。经10分钟、15000×g离心后收集上清液。上清液经0.45μm孔径滤膜（密理博（Millipore））过滤，以去除真核细胞与细菌尺寸的颗粒；随后从每批样本池的上清液中取200μL，加入核酸酶混合液以降解游离（未被病毒衣壳包裹）的核酸。剩余总核酸采用QIAamp MinElute病毒离心柱试剂盒（QIAamp MinElute Virus Spin Kit），依照制造商提供的实验流程完成提取。随后采用Nextera XT DNA样本制备试剂盒（Illumina）构建197个测序文库，依托Illumina NovaSeq 6000测序平台进行250bp双端测序，每个样本池均采用双条形码标记。