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一种基于CRISPR/Cas12a的快速和超灵敏测试,用于准确识别T偶类型的菌体

Chenhang Jiang^1,2, Yang Li³, Ping Yu⁴

¹Key Laboratory of Biomass Chemical Engineering of Ministry of Education, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, China.

Biotechnology letters

|September 5, 2025

概括

一种新的CRISPR/Cas12a和重组酶聚合酶放大 (RPA) 试验可提供超敏感的T-偶类菌体检测,这对于防止发酵污染和生化工业的经济损失至关重要.

科学领域:

生物技术
分子生物学
工业微生物学

背景情况:

菌体污染对工业发酵过程构成重大经济威胁.
在这些系统中,有毒的T偶类菌体 (T2,T4,T6) 是重要的生物危害.
现有的检测方法可能缺乏实时监测所需的灵敏度和速度.

研究的目的:

开发一种新型的超敏感检测系统,用于T-even型菌体.
针对TerL基因进行特定和可靠的菌体识别.
为生物化学工业提供快速有效的质量控制工具.

主要方法:

整合CRISPR/Cas12a技术与重组酶聚合酶放大 (RPA).
针对T偶菌体的TerL基因的特定原料和导向RNA的设计.
优化RPA-CRISPR测定以提高灵敏度和特异性.
使用合成标,纯化菌体 (T2,T4) 和加/废水样本进行验证.

主要成果:

优化的RPA-CRISPR试验实现了合成标的超敏感检测极限 (LOD) 1 aM.
T2菌体的检测值为1 PFU/μL,T4菌体的检测值为10 PFU/μL.
该试验具有很高的可靠性,与定量PCR (qPCR) 结果有很强的相关性.
在和现实废水样本中成功检测.

结论:

开发的RPA-CRISPR系统提供了一种快速,超敏感和可靠的检测T-even类型菌体的方法.
这项技术具有实时监测和控制工业发酵中的菌体污染的巨大潜力.
它为加强生物化学产业的质量控制和减轻经济损失提供了有希望的解决方案.

关键词:

在CRISPR/Cas12a中核酸检测质量控制重组酶聚合酶放大 T偶类型的菌体

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