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CRISPR-Cas13a-based detection method for avian influenza virus

文献类型： 外文期刊

作者： 吴语涵;;战佳兴;;单朝萌;;李雁冰;;刘伊宁;;李妍;;王轶鑫;;刘哲;;文雪霞;;王秀荣

作者机构：

关键词： avian influenza virus;CRISPR-cas13a;lateral flow dipstick assay;recombinase polymerase amplification;universal detection

期刊名称： Frontiers in Microbiology

ISSN： 1664-302X

年卷期： 2023 年 14 卷

页码：

收录情况： SCIE（2023版）

摘要： Avian influenza virus (AIV) causes huge losses to the global poultry industry and poses a threat to humans and other mammals. Fast, sensitive, and portable diagnostic methods are essential for efficient avian influenza control. Here, a clustered regularly interspaced short palindromic repeats (CRISPR)-Cas13a based platform was developed to detect AIV. This novel method was developed to specifically detect H1–H16 subtypes of AIV with fluorescence and lateral flow-based readouts and exhibited no cross-reactivity with Newcastle disease virus, avian infectious bronchitis virus, or infectious bursal disease virus. The limit of detection was determined to be 69 and 690 copies/μL using fluorescence and lateral flow as readouts, respectively. The developed assay exhibited 100% consistency with quantitative real-time polymerase chain reaction in detecting clinical samples. The heating of unextracted diagnostic samples to obliterate nuclease treatment was introduced to detect viral RNA without nucleic acid extraction. Single-step optimization was used to perform reverse transcription, recombinase polymerase amplification, and CRISPR-Cas13a detection in a tube. These advances resulted in an optimized assay that could specifically detect AIV with simplified procedures and reduced contamination risk, highlighting the potential to be used in point-of-care testing.

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