CRISPR-Cas12-based field-deployable system for rapid detection of synthetic DNA sequence of the monkeypox virus genome
CRISPR-Cas12-based field-deployable system for rapid detection of synthetic DNA sequence of the monkeypox virus genome
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DOI：
10.1002/jmv.28385
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发表时间：
2023-01-01
影响因子：
12.7
通讯作者：
Qin, Peiwu
中科院分区：
文献类型：
--
作者：
Chen, Qun;Gul, Ijaz;Qin, Peiwu
猴痘病毒（MPXV）的全球爆发凸显了对快速且具有成本效益的MPXV检测工具的需求，以有效地监测和控制猴痘疾病。在此，我们展示了一种用于MPXV裸眼检测的便携式CRISPR-Cas系统。该系统利用了CRISPR-Cas 12的高选择性和重组酶聚合酶扩增的等温核酸扩增潜力。它可以检测到当前流行的MPXV分支和原始分支。我们使用微量滴定板读数器达到22.4aM（13.5copies/mu l）的检测限（LoD），而在两步测定中系统的视觉LoD为75 aM（45 copies/mu l），在一锅系统中进一步降低至25 aM（15 copies/mu l）。我们比较了我们的结果与定量聚合酶链反应，并获得满意的一致性。对于临床应用，我们展示了一种灵敏和精确的视觉检测方法，其具有阿托摩尔灵敏度和35分钟的样品响应时间。
The global outbreak of the monkeypox virus (MPXV) highlights the need for rapid and cost-effective MPXV detection tools to effectively monitor and control the monkeypox disease. Herein, we demonstrated a portable CRISPR-Cas-based system for naked-eye detection of MPXV. The system harnesses the high selectivity of CRISPR-Cas12 and the isothermal nucleic acid amplification potential of recombinase polymerase amplification. It can detect both the current circulating MPXV clade and the original clades. We reached a limit of detection (LoD) of 22.4aM (13.5copies/mu l) using a microtiter plate reader, while the visual LoD of the system is 75aM (45copies/mu l) in a two-step assay, which is further reduced to 25aM (15copies/mu l) in a one-pot system. We compared our results with quantitative polymerase chain reaction and obtained satisfactory consistency. For clinical application, we demonstrated a sensitive and precise visual detection method with attomolar sensitivity and a sample-to-answer time of 35min.