使用基于复合酶辅助放大试验辅助CRISPR/Cas12a系统的新方法检测Clostridium perfringens
Xingxing Xiao1, Qingxun Zhang2, Sihong Wu1
1Wenzhou Key Laboratory of Sanitary Microbiology, Key Laboratory of Laboratory Medicine, Ministry of Education, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China.
Transboundary and emerging diseases
|April 30, 2025
概括
对于Clostridium perfringens (C. perfringens),新的快速检测方法提供了快速而敏感的结果. 这些无仪器测定,使用复合酶辅助放大 (RAA) 和CRISPR/Cas12a,可以在不到一个小时内检测出C. perfringens.
科学领域:
- 微生物学 微生物学
- 分子生物学分子生物学
- 生物技术是生物技术.
背景情况:
- 杆菌 (Clostridium perfringens) 是人类和动物健康的重要病原体.
- 对C. perfringens的快速和敏感的诊断工具的需求对于有效的疾病控制和管理至关重要.
- 现有的检测方法可能缺乏及时干预所需的速度或灵敏度.
研究的目的:
- 开发新的,快速和灵敏的无仪器检测方法来检测Clostridium perfringens.
- 使用复合酶辅助放大 (RAA) 与CRISPR/Cas12a系统相结合,以提高检测能力.
- 为了验证这些新方法的性能与既定技术和各种样本类型相比.
主要方法:
- 开发了两种不同的C. perfringens检测试验:RAA-CRISPR/Cas12a-FL (基于光) 和RAA-CRISPR/Cas12a-LFS (侧流带).
- 使用C. perfringens基因组DNA.评估了这两种方法的检测极限 (LOD).
- 对非标细菌的评估特异性,并将结果与实时聚合酶连锁反应 (qPCR) 进行比较,使用临床和尖样本.
主要成果:
- 在RAA-CRISPR/Cas12a-FL试验中,每个反应的LOD为2个副本,而RAA-CRISPR/Cas12a-LFS的LOD为20个副本.
- 这两种方法都与qPCR结果100%一致,并且没有与其他细菌物种的交叉反应.
- 两种测试的整个检测过程在1小时内完成.
结论:
- 开发的RAA-CRISPR/Cas12a-FL和RAA-CRISPR/Cas12a-LFS测定方法提供了快速,敏感和特定的C. perfringens的检测.
- 这些无仪器方法在临床环境中对C. perfringens感染的早期诊断具有重大潜力.
- 这些测试为人类和兽医应用提供了一个有前途的新诊断方案.
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