体外筛选脱氧核酶用于检测 RNA 修饰。
In Vitro Selection of Deoxyribozymes for the Detection of RNA Modifications.
机构信息
Institute of Organic Chemistry, Julius-Maximilians-University Würzburg, Würzburg, Germany.
出版信息
Methods Mol Biol. 2022;2533:167-179. doi: 10.1007/978-1-0716-2501-9_10.
Deoxyribozymes are artificially evolved DNA molecules with catalytic abilities. RNA-cleaving deoxyribozymes have been recognized as an efficient tool for detection of modifications in target RNAs and provide an alternative to traditional and modern methods for detection of ribose or nucleobase methylation. However, there are only few examples of DNA enzymes that specifically reveal the presence of a certain type of modification, including N -methyladenosine, and the knowledge about how DNA enzymes recognize modified RNAs is still extremely limited. Therefore, DNA enzymes cannot be easily engineered for the analysis of desired RNA modifications, but are instead identified by in vitro selection from random DNA libraries using synthetic modified RNA substrates. This protocol describes a general in vitro selection stagtegy to evolve new RNA-cleaving DNA enzymes that can efficiently differentiate modified RNA substrates from their unmodified counterpart.
脱氧核酶是具有催化能力的人工进化的 DNA 分子。RNA 切割脱氧核酶已被认为是检测靶 RNA 修饰的有效工具，并为检测核糖或核苷甲基化的传统和现代方法提供了替代方法。然而，能够特异性揭示特定类型修饰（包括 N-甲基腺苷）存在的 DNA 酶仅有少数几个例子，并且关于 DNA 酶如何识别修饰 RNA 的知识仍然极其有限。因此，DNA 酶不能轻易地针对所需 RNA 修饰进行工程设计，而是通过使用合成修饰的 RNA 底物从随机 DNA 文库中进行体外选择来鉴定。本协议描述了一种通用的体外选择策略，用于进化新的 RNA 切割 DNA 酶，这些酶可以有效地将修饰的 RNA 底物与未修饰的 RNA 底物区分开。
Zotero 插件