RT-RPA Assisted CRISPR/Cas12a Based One-Pot Rapid and Visual Detection of the Pan-Dengue Virus.

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Tác giả: Pooja Bhardwaj, Preeti Dhangur, Alagarasu Kalichamy, Rajeev Singh

Ngôn ngữ: eng

Ký hiệu phân loại: 920.71 Men

Thông tin xuất bản: United States : Journal of medical virology , 2025

Mô tả vật lý:

Bộ sưu tập: NCBI

ID: 56604

Globally ≤ 4 billion of the population are at potential risk of contracting dengue virus (DENV) infection. Seasonal outbreaks of dengue are frequently reported causing a high healthcare burden. Undiagnosed DENV can lead to severe morbidity and mortality. Early diagnosis of DENV relies on molecular methods, which are impractical in resource-constrained settings (RCSs). Dengue can be caused by any of the four distinct DENV serotypes. Therefore, a simple method for rapid diagnosis of Pan-DENV serotypes is of utmost importance at RCSs. A fluorescence detection platform for Pan-DENV using RT-RPA and CRISPR/Cas12a was developed targeting nonstructural 1 (NS1) gene for DENV-1, 2, and 3, and envelope (E) gene for DENV-2. Further, crRNA specific to DENV serotypes were designed to facilitate CRISPR/Cas12a detection. Analytical sensitivity was determined using synthetic RNA and DENV serotypes genome. Clinical validation of the assay was performed using RNA extracted from AES/AFI clinical samples. The developed CRISPR/Cas12a-based detection platform can detect all four serotypes of DENV viz 1-4 in a single pot using fluorescence detection. This assay showed the limit of detection ≥ 781 zg reaction
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