通过高通量测序进行病毒检测。

Viral detection by high-throughput sequencing.

作者信息

Motooka Daisuke, Nakamura Shota, Hagiwara Katsuro, Nakaya Takaaki

机构信息

Department of Infection Metagenomics, Research Institute for Microbial Disease, Osaka University, Osaka, Japan.

出版信息

Methods Mol Biol. 2015;1236:125-34. doi: 10.1007/978-1-4939-1743-3_11.

DOI:10.1007/978-1-4939-1743-3_11

PMID:25287501

Abstract

We applied a high-throughput sequencing platform, Ion PGM, for viral detection in fecal samples from adult cows collected in Hokkaido, Japan. Random RT-PCR was performed to amplify RNA extracted from 0.25 ml of fecal specimens (N = 8), and more than 5 μg of cDNA was synthesized. Unbiased high-throughput sequencing using the 318 v2 semiconductor chip of these eight samples yielded 57-580 K (average: 270 K, after data analysis) reads in a single run. As a result, viral genome sequences were detected in each specimen. In addition to bacteriophage, mammal- and insect-derived viruses, partial genome sequences of plant, algal, and protozoal viruses were detected. Thus, this metagenomic analysis of fecal specimens could be useful to comprehensively understand viral populations of the intestine and food sources in animals.

摘要

我们应用了一种高通量测序平台——Ion PGM，对从日本北海道采集的成年奶牛粪便样本进行病毒检测。进行随机逆转录聚合酶链反应（Random RT-PCR）以扩增从0.25毫升粪便标本（N = 8）中提取的RNA，并合成了超过5微克的互补脱氧核糖核酸（cDNA）。使用这八个样本的318 v2半导体芯片进行无偏差高通量测序，单次运行产生了57 - 580 K（数据分析后平均为270 K）条读数。结果，在每个标本中都检测到了病毒基因组序列。除了噬菌体、哺乳动物和昆虫来源的病毒外，还检测到了植物、藻类和原生动物病毒的部分基因组序列。因此，这种粪便标本的宏基因组分析可能有助于全面了解动物肠道和食物来源中的病毒群体。