RT-RPA-Cas12a-based discrimination of SARS-CoV-2 variants of concern
RT-RPA-Cas12a-based discrimination of SARS-CoV-2 variants of concern
Abstract Timely and accurate detection of SARS-CoV-2 variants of concern (VOCs) is urgently needed for pandemic surveillance and control. However, current methods are limited by the low sensitivity, long turn-around time or high cost. Here, we report a nucleic acid testing-based method aiming to detect and discriminate SARS-CoV-2 VOCs by combining RT-RPA and CRISPR-Cas12a detecting assays (RRCd). With a detection limit of 10 copies RNA/reaction, RRCd was validated in 204 clinical samples, showing 99% positive predictive agreement and 100% negative predictive agreement, respectively. Critically, using specific crRNAs, representatives of single nucleotide polymorphisms and small deletions in SARS-CoV-2 VOCs including N501Y, T478K and ΔH69-V70 were discriminated by RRCd, demonstrating 100% accuracy in clinical samples with Ct < 33. The method completes within 65 min and could offer visible results without using any electrical devices, which may facilitate point-of-care testing of SARS-CoV-2 and its variants.
Yang Yujiao、Long Fei、Sun Jingxian、Cai Kezhu、Li Shenwei、Wang Zhiyi、Liu Jiakun、Ma Yingxin、Zhang Zilong、Zou Siwei、Tang Guiyue、Zhao Guo-Ping、Mao Guobing、Tan Wei、Li Yingying、Zhao Wei、Tian Zhen-Gan
CAS Key Laboratory of Synthetic Biology, CAS Center for Excellence in Molecular Plant Sciences, Chinese Academy of Sciences||CAS Key Laboratory of Synthetic Biology, University of Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences||School of Engineering, Southern University of Science and TechnologyShanghai International Travel Healthcare Center, Shanghai Customs District P. R. ChinaShanghai International Travel Healthcare Center, Shanghai Customs District P. R. ChinaCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesShanghai International Travel Healthcare Center, Shanghai Customs District P. R. ChinaCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences||CAS Key Laboratory of Synthetic Biology, CAS Center for Excellence in Molecular Plant Sciences, Chinese Academy of Sciences||CAS Key Laboratory of Synthetic Biology, University of Chinese Academy of Sciences||State Key Lab of Genetic Engineering & Institutes of Biomedical Sciences, Department of Microbiology and Microbial Engineering, School of Life Sciences, Fudan UniversityCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesCAS Key Laboratory of Quantitative Engineering Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of SciencesShanghai International Travel Healthcare Center, Shanghai Customs District P. R. China
医学研究方法基础医学分子生物学
SARS-CoV-2variant of concernRRCdpoint-of-care testingCRISPR
Yang Yujiao,Long Fei,Sun Jingxian,Cai Kezhu,Li Shenwei,Wang Zhiyi,Liu Jiakun,Ma Yingxin,Zhang Zilong,Zou Siwei,Tang Guiyue,Zhao Guo-Ping,Mao Guobing,Tan Wei,Li Yingying,Zhao Wei,Tian Zhen-Gan.RT-RPA-Cas12a-based discrimination of SARS-CoV-2 variants of concern[EB/OL].(2025-03-28)[2025-04-27].https://www.medrxiv.org/content/10.1101/2022.05.11.22274884.点此复制
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