Co-freezing localized CRISPR-Cas12a system enables rapid and sensitive nucleic acid analysis.

IF 10.6 1区 生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Journal of Nanobiotechnology Pub Date : 2024-10-05 DOI:10.1186/s12951-024-02831-8
Lifeng Zhang, Shihua Luo, Wenbin Li, Wanting Su, Siting Chen, Chunchen Liu, Weilun Pan, Bo Situ, Lei Zheng, Ling Li, Xiaohui Yan, Ye Zhang
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Abstract

Rapid and sensitive nucleic acid detection is vital in disease diagnosis and therapeutic assessment. Herein, we propose a co-freezing localized CRISPR-Cas12a (CL-Cas12a) strategy for sensitive nucleic acid detection. The CL-Cas12a was obtained through a 15-minute co-freezing process, allowing the Cas12a/crRNA complex and hairpin reporter confined on the AuNPs surface with high load efficiency, for rapid sensing of nucleic acid with superior performance to other localized Cas12a strategies. This CL-Cas12a based platform could quantitatively detect targets down to 98 aM in 30 min with excellent specificity. Furthermore, the CL-Cas12a successful applied to detect human papillomavirus infection and human lung cancer-associated single-nucleotide mutations. We also achieved powerful signal amplification for imaging Survivin mRNA in living cells. These findings highlight the potential of CL-Cas12a as an effective tool for nucleic acid diagnostics and disease monitoring.

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共冷冻定位 CRISPR-Cas12a 系统可实现快速、灵敏的核酸分析。
快速灵敏的核酸检测对疾病诊断和治疗评估至关重要。在此,我们提出了一种用于灵敏核酸检测的共冷冻定位CRISPR-Cas12a(CL-Cas12a)策略。CL-Cas12a通过15分钟的共冷冻过程获得,使Cas12a/crRNA复合物和发夹式报告物以高负载效率被限制在AuNPs表面,用于快速检测核酸,其性能优于其他Cas12a定位策略。这种基于 CL-Cas12a 的平台可在 30 分钟内定量检测低至 98 aM 的目标物,具有极佳的特异性。此外,CL-Cas12a 还成功应用于检测人类乳头瘤病毒感染和人类肺癌相关单核苷酸突变。我们还实现了活细胞中 Survivin mRNA 成像的强大信号放大。这些发现凸显了 CL-Cas12a 作为核酸诊断和疾病监测有效工具的潜力。
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来源期刊
Journal of Nanobiotechnology
Journal of Nanobiotechnology BIOTECHNOLOGY & APPLIED MICROBIOLOGY-NANOSCIENCE & NANOTECHNOLOGY
CiteScore
13.90
自引率
4.90%
发文量
493
审稿时长
16 weeks
期刊介绍: Journal of Nanobiotechnology is an open access peer-reviewed journal communicating scientific and technological advances in the fields of medicine and biology, with an emphasis in their interface with nanoscale sciences. The journal provides biomedical scientists and the international biotechnology business community with the latest developments in the growing field of Nanobiotechnology.
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