N1-Methyladenosine detection with CRISPR-Cas13a/C2c2
N1-Methyladenosine detection with CRISPR-Cas13a/C2c2
复制标题
使用 CRISPR-Cas13a/C2c2 检测 N1-甲基腺苷
DOI：
10.1039/c8sc03408g
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发表时间：
2019-03-14
期刊：
影响因子：
8.4
通讯作者：
Zhou, Xiang
中科院分区：
文献类型：
--
作者：
Chen, Yi;Yang, Shixi;Zhou, Xiang
近年来的研究表明，N-1-甲基腺苷（m（1）A）的广泛存在在环境胁迫、核糖体生物合成和抗生素抗性等方面起着重要作用。RNA引导的、RNA靶向的CRISPR Cas 13 a在以高灵敏度识别靶标时表现出混杂RNA酶活性的“附带效应”。受CRISPR Cas 13 a优势的启发，我们设计了一种检测m（1）A诱导的错配的系统，提供了一种快速、简单和基于荧光的m（1）A检测。对于A-ssRNA，基于Cas 13 a的分子检测平台显示出高荧光信号。对于m（1）A-ssRNA，存在约90%的荧光下降。此外，该方法还可用于RNA中m（1）A的定量分析，并可应用于AlkB对28 SrRNA的动态去甲基化分析。
Recent studies suggested that the widespread presence of N-1-methyladenosine (m(1)A) plays a very important role in environmental stress, ribosome biogenesis and antibiotic resistance. The RNA-guided, RNA-targeting CRISPR Cas13a exhibits a "collateral effect" of promiscuous RNase activity upon target recognition with high sensitivity. Inspired by the advantage of CRISPR Cas13a, we designed a system to detect m(1)A induced mismatch, providing a rapid, simple and fluorescence-based m(1)A detection. For A-ssRNA, the Cas13a-based molecular detection platform showed a high fluorescence signal. For m(1)A-ssRNA, there is an about 90% decline of fluorescence. Moreover, this approach can also be used to quantify m(1)A in RNAs and applied for the analysis of dynamic m(1)A demethylation of 28S rRNA with AlkB.