Direct and accurate miRNA detection based on CRISPR/Cas13a-triggered exonuclease-iii-assisted colorimetric assay

IF 2.5 4区化学 Q3 CHEMISTRY, ANALYTICAL

Journal of Analytical Science and Technology Pub Date : 2024-04-16 DOI:10.1186/s40543-024-00434-4

Yunxiao Li, Qiuxia Wang, Yali Wang

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引用次数: 0

Abstract

The abnormally expressed microRNAs (miRNAs) serve as crucial indicators for disease diagnosis and are strongly associated with the progression of various diseases. The quantification of miRNAs is highly significant for the clinical diagnosis and treatment of various types of cancers. This study utilizes the CRISPR/Cas13a system, which combines CRISPR RNA (crRNA) and CRISPR-associated Cas13a, to recognize the miRNA directly and specifically, thus activating the trans-cleavage activity of Cas13a. By integrating the CRISPR-Cas13a system with an exonuclease-iii (Exo-iii)-assisted chain cleavage of silver ions (Ag+)-aptamer and an Ag+ -based color reaction, a method for the detection of miRNA that is specific and sensitive is developed. This approach demonstrated exceptional sensitivity in detecting miRNA, with a low detection limit of 5.12 fM. Moreover, the suggested method was effectively utilized to precisely measure the amount of miRNA in intricate biological samples, demonstrating its practical applicability. Furthermore, due to its elevated sensitivity and simple probe design and fluorophore labeling, the suggested colorimetric technique holds great potential for use in clinical diagnostics.

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基于 CRISPR/Cas13a 触发的外切酶-iii 辅助比色法的直接准确 miRNA 检测

表达异常的微RNA（miRNA）是疾病诊断的重要指标，与各种疾病的进展密切相关。miRNAs的定量分析对各类癌症的临床诊断和治疗意义重大。本研究利用CRISPR/Cas13a系统，将CRISPR RNA（crRNA）和CRISPR相关的Cas13a结合起来，直接特异性地识别miRNA，从而激活Cas13a的反式裂解活性。通过将 CRISPR-Cas13a 系统与外切核酸酶-iii（Exo-iii）辅助的链式裂解银离子（Ag+）-aptamer 和基于 Ag+ 的颜色反应相结合，开发出了一种特异性和灵敏度高的 miRNA 检测方法。该方法检测 miRNA 的灵敏度极高，检测限低至 5.12 fM。此外，该方法还可用于精确测量复杂生物样本中的 miRNA 含量，证明了它的实用性。此外，由于其灵敏度高、探针设计和荧光团标记简单，所建议的比色技术在临床诊断中具有巨大的应用潜力。

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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来源期刊

Journal of Analytical Science and Technology Environmental Science-General Environmental Science

CiteScore

4.00

自引率

4.20%

发文量

审稿时长

13 weeks

期刊介绍： The Journal of Analytical Science and Technology (JAST) is a fully open access peer-reviewed scientific journal published under the brand SpringerOpen. JAST was launched by Korea Basic Science Institute in 2010. JAST publishes original research and review articles on all aspects of analytical principles, techniques, methods, procedures, and equipment. JAST’s vision is to be an internationally influential and widely read analytical science journal. Our mission is to inform and stimulate researchers to make significant professional achievements in science. We aim to provide scientists, researchers, and students worldwide with unlimited access to the latest advances of the analytical sciences.