Direct RNA Sequencing of Foot-and-mouth Disease Virus Genome Using a Flongle on MinION.

IF 16.4 1区 化学 Q1 CHEMISTRY, MULTIDISCIPLINARY
Lizhe Xu, Amy Berninger, Steven M Lakin, Vivian O'Donnell, Jim L Pierce, Steven J Pauszek, Roger W Barrette, Bonto Faburay
{"title":"Direct RNA Sequencing of Foot-and-mouth Disease Virus Genome Using a Flongle on MinION.","authors":"Lizhe Xu, Amy Berninger, Steven M Lakin, Vivian O'Donnell, Jim L Pierce, Steven J Pauszek, Roger W Barrette, Bonto Faburay","doi":"10.21769/BioProtoc.5017","DOIUrl":null,"url":null,"abstract":"<p><p><i>Foot-and-mouth disease (FMD) is a severe and extremely contagious viral disease of cloven-hoofed domestic and wild animals, which leads to serious economic losses to the livestock industry globally. FMD is caused by the FMD virus (FMDV), a positive-strand RNA virus that belongs to the genus</i> Aphthovirus, within the family Picornaviridae. Early detection and characterization of FMDV strains are key factors to control new outbreaks and prevent the spread of the disease. Here, we describe a direct RNA sequencing method using Oxford Nanopore Technology (ONT) Flongle flow cells on MinION Mk1C (or GridION) to characterize FMDV. This is a rapid, low cost, and easily deployed point of care (POC) method for a near real-time characterization of FMDV in endemic areas or outbreak investigation sites. Key features • Saves ~35 min of the original protocol time by omitting the reverse transcription step and lowers the costs of reagents and consumables. • Replaces the GridION flow cell from the original protocol with the Flongle, which saves ~90% on the flow cell cost. • Combines the NGS benchwork with a modified version of our African swine fever virus (ASFV) fast analysis pipeline to achieve FMDV characterization within minutes. Graphical overview Schematic of direct RNA sequencing of foot-and-mouth disease virus (FMDV) process, which takes ~50 min from extracted RNA to final loading, modified from the ONT SQK-RNA002 protocol (Version: DRS_9080_v2_revO_14Aug2019).</p>","PeriodicalId":1,"journal":{"name":"Accounts of Chemical Research","volume":null,"pages":null},"PeriodicalIF":16.4000,"publicationDate":"2024-06-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11211080/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Accounts of Chemical Research","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.21769/BioProtoc.5017","RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"CHEMISTRY, MULTIDISCIPLINARY","Score":null,"Total":0}
引用次数: 0

Abstract

Foot-and-mouth disease (FMD) is a severe and extremely contagious viral disease of cloven-hoofed domestic and wild animals, which leads to serious economic losses to the livestock industry globally. FMD is caused by the FMD virus (FMDV), a positive-strand RNA virus that belongs to the genus Aphthovirus, within the family Picornaviridae. Early detection and characterization of FMDV strains are key factors to control new outbreaks and prevent the spread of the disease. Here, we describe a direct RNA sequencing method using Oxford Nanopore Technology (ONT) Flongle flow cells on MinION Mk1C (or GridION) to characterize FMDV. This is a rapid, low cost, and easily deployed point of care (POC) method for a near real-time characterization of FMDV in endemic areas or outbreak investigation sites. Key features • Saves ~35 min of the original protocol time by omitting the reverse transcription step and lowers the costs of reagents and consumables. • Replaces the GridION flow cell from the original protocol with the Flongle, which saves ~90% on the flow cell cost. • Combines the NGS benchwork with a modified version of our African swine fever virus (ASFV) fast analysis pipeline to achieve FMDV characterization within minutes. Graphical overview Schematic of direct RNA sequencing of foot-and-mouth disease virus (FMDV) process, which takes ~50 min from extracted RNA to final loading, modified from the ONT SQK-RNA002 protocol (Version: DRS_9080_v2_revO_14Aug2019).

使用 MinION 上的 Flongle 对口蹄疫病毒基因组进行直接 RNA 测序。
口蹄疫(FMD)是蹄类家畜和野生动物的一种严重且传染性极强的病毒性疾病,给全球畜牧业造成了严重的经济损失。口蹄疫是由口蹄疫病毒(FMDV)引起的,FMDV 是一种正链 RNA 病毒,属于皮卡病毒科 Aphthovirus 属。早期检测和鉴定 FMDV 株系是控制新疫情和防止疾病传播的关键因素。在此,我们介绍一种直接 RNA 测序方法,该方法在 MinION Mk1C(或 GridION)上使用牛津纳米孔技术(ONT)Flongle 流式细胞对 FMDV 进行定性。这是一种快速、低成本、易于部署的医疗点 (POC) 方法,可在流行区或疫情调查点对 FMDV 进行近乎实时的表征。主要特点 - 省去反转录步骤,比原方案节省约 35 分钟时间,并降低试剂和耗材成本。- 用 Flongle 代替原方案中的 GridION 流动池,可节省约 90% 的流动池成本。- 将 NGS 工作台与非洲猪瘟病毒 (ASFV) 快速分析流水线的改进版相结合,在几分钟内完成 FMDV 鉴定。图解概述 口蹄疫病毒 (FMDV) 直接 RNA 测序过程示意图,从提取 RNA 到最终装载需要约 50 分钟,修改自 ONT SQK-RNA002 协议(版本:DRS_9080_v2_revO_14Aug2019)。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
Accounts of Chemical Research
Accounts of Chemical Research 化学-化学综合
CiteScore
31.40
自引率
1.10%
发文量
312
审稿时长
2 months
期刊介绍: Accounts of Chemical Research presents short, concise and critical articles offering easy-to-read overviews of basic research and applications in all areas of chemistry and biochemistry. These short reviews focus on research from the author’s own laboratory and are designed to teach the reader about a research project. In addition, Accounts of Chemical Research publishes commentaries that give an informed opinion on a current research problem. Special Issues online are devoted to a single topic of unusual activity and significance. Accounts of Chemical Research replaces the traditional article abstract with an article "Conspectus." These entries synopsize the research affording the reader a closer look at the content and significance of an article. Through this provision of a more detailed description of the article contents, the Conspectus enhances the article's discoverability by search engines and the exposure for the research.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信