A sensitive fluorescence biosensor based on ligation-transcription and CRISPR/Cas13a-assisted cascade amplification strategies to detect the H1N1 virus

文献类型： 外文期刊

第一作者： Xue, Lulu

作者： Xue, Lulu;Xu, Mengyao;Xu, Yao;Li, Zehong;Bu, Shengjun;Wei, Jiaqi;Zhou, Hongyu;Hao, Zhuo;Wan, Jiayu;Bu, Shengjun

作者机构：

关键词： SplintR ligase; T7 RNA polymerase; CRISPR/Cas13a

期刊名称：ANALYTICAL AND BIOANALYTICAL CHEMISTRY （ 影响因子：4.3； 五年影响因子：4.0 ）

ISSN： 1618-2642

年卷期： 2024 年 416 卷 13 期

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收录情况： SCI

摘要： We propose a sensitive H1N1 virus fluorescence biosensor based on ligation-transcription and CRISPR/Cas13a-assisted cascade amplification strategies. Products are generated via the hybridization of single-stranded DNA (ssDNA) probes containing T7 promoter and crRNA templates to a target RNA sequence using SplintR ligase. This generates large crRNA quantities in the presence of T7 RNA polymerase. At such crRNA quantities, ternary Cas13a, crRNA, and activator complexes are successfully constructed and activate Cas13a to enhance fluorescence signal outputs. The biosensor sensitively and specifically monitored H1N1 viral RNA levels down to 3.23 pM and showed good linearity when H1N1 RNA concentrations were 100 pM-1 mu M. Biosensor specificity was also excellent. Importantly, our biosensor may be used to detect other viral RNAs by altering the sequences of the two probe junctions, with potential applications for the clinical diagnosis of viruses and other biomedical studies.

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