利用 CRISPR-Cas13 快速、简便、灵敏、低成本地现场检测环境 DNA 和 RNA

IF 6.3 2区 环境科学与生态学 Q1 ECOLOGY Methods in Ecology and Evolution Pub Date : 2024-07-09 DOI:10.1111/2041-210X.14369
Jiwei Yang, Shoma Matsushita, Fei Xia, Susumu Yoshizawa, Wataru Iwasaki
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引用次数: 0

摘要

对物种分布进行环境 DNA(eDNA)监测已成为生态学、保护生物学和渔业领域确定各种生物(包括重要、受威胁和入侵物种)存在和分布的重要工具。然而,eDNA 检测的灵敏度仍有待提高,需要耗时的实时聚合酶链反应(qPCR)步骤和昂贵的机器。在本研究中,我们报告了一种基于 CRISPR-Cas13 的快速、简便、灵敏、低成本和现场检测 eDNA 的方法。这种检测鲤鱼(Cyprinus carpio)和青鳉(Oryzias latipes)核酸的方法采用两步法,首先进行重组酶聚合酶扩增(RPA),然后使用 Cas13 核酸酶进行裂解,从而有效识别线粒体 DNA 或 RNA。我们的研究结果表明,在检测微量 eDNA 方面,基于 Cas13 的方法比基于 qPCR 的方法灵敏度更高。当与环境 RNA(eRNA)的反转录相结合时,我们的方法将检测灵敏度提高了约一个数量级。使用快速核酸提取液和横向流动条,基于 Cas13 的检测方法可实现现场检测 eDNA。基于 Cas13 的 eDNA 和 eRNA 检测只需极少的培训,并可在 1 小时内完成,无需离心和 qPCR 仪器。基于 Cas13 的方法具有便携性、准确性以及设计引物和 CRISPR RNA(crRNA)的相对简便性,这突出表明它有可能拓宽 eDNA 和 eRNA 在生物多样性保护等各个领域的应用。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

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Rapid, easy, sensitive, low-cost and on-site detection of environmental DNA and RNA using CRISPR-Cas13

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来源期刊
CiteScore
11.60
自引率
3.00%
发文量
236
审稿时长
4-8 weeks
期刊介绍: A British Ecological Society journal, Methods in Ecology and Evolution (MEE) promotes the development of new methods in ecology and evolution, and facilitates their dissemination and uptake by the research community. MEE brings together papers from previously disparate sub-disciplines to provide a single forum for tracking methodological developments in all areas. MEE publishes methodological papers in any area of ecology and evolution, including: -Phylogenetic analysis -Statistical methods -Conservation & management -Theoretical methods -Practical methods, including lab and field -This list is not exhaustive, and we welcome enquiries about possible submissions. Methods are defined in the widest terms and may be analytical, practical or conceptual. A primary aim of the journal is to maximise the uptake of techniques by the community. We recognise that a major stumbling block in the uptake and application of new methods is the accessibility of methods. For example, users may need computer code, example applications or demonstrations of methods.
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