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Rapid and Sensitive Genotyping of SARS-CoV-2 Key Mutation L452R with an RPA-PfAgo Method.
Zhao, Chenjie; Yang, Lihong; Zhang, Xue; Tang, Yixin; Wang, Yue; Shao, Xiaofu; Gao, Song; Liu, Xin; Wang, Pei.
  • Zhao C; Jiangsu Key Laboratory of Marine Biological Resources and Environment, Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, School of Pharmacy, Jiangsu Ocean University, Lianyungang 222005, China.
  • Yang L; Jiangsu Key Laboratory of Marine Biological Resources and Environment, Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, School of Pharmacy, Jiangsu Ocean University, Lianyungang 222005, China.
  • Zhang X; Jiangsu Key Laboratory of Marine Biological Resources and Environment, Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, School of Pharmacy, Jiangsu Ocean University, Lianyungang 222005, China.
  • Tang Y; Jiangsu Key Laboratory of Marine Biological Resources and Environment, Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, School of Pharmacy, Jiangsu Ocean University, Lianyungang 222005, China.
  • Wang Y; Jiangsu Key Laboratory of Marine Biological Resources and Environment, Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, School of Pharmacy, Jiangsu Ocean University, Lianyungang 222005, China.
  • Shao X; Jiangsu Key Laboratory of Marine Biological Resources and Environment, Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, School of Pharmacy, Jiangsu Ocean University, Lianyungang 222005, China.
  • Gao S; Jiangsu Key Laboratory of Marine Biological Resources and Environment, Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, School of Pharmacy, Jiangsu Ocean University, Lianyungang 222005, China.
  • Liu X; Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan 430074, China.
  • Wang P; School of Food Science and Pharmaceutical Engineering, Nanjing Normal University, Nanjing 210023, China.
Anal Chem ; 2022 Dec 02.
Article in English | MEDLINE | ID: covidwho-2150966
ABSTRACT
In the two years of COVID-19 pandemic, the SARS-CoV-2 variants have caused waves of infections one after another, and the pandemic is not ending. The key mutations on the S protein enable the variants with enhanced viral infectivity, immune evasion, and/or antibody neutralization resistance, bringing difficulties to epidemic prevention and control. In support of precise epidemic control and precision medicine of the virus, a fast and simple genotyping method for the key mutations of SARS-CoV-2 variants needs to be developed. By utilizing the specific recognition and cleavage property of the nuclease Argonaute from Pyrococcus furiosus (PfAgo), we developed a recombinase polymerase amplification (RPA) and PfAgo combined method for a rapid and sensitive genotyping of SARS-CoV-2 key mutation L452R. With a delicate design of the strategy, careful screening of the RPA primers and PfAgo gDNA, and optimization of the reaction, the method achieves a high sensitivity of a single copy per reaction, which is validated with the pseudovirus. This is the highest sensitivity that can be achieved theoretically and the highest sensitivity as compared to the available SARS-CoV-2 genotyping assays. Using RPA, the procedure of the method is finished within 1.5 h and only needs a minimum laboratorial support, suggesting that the method can be easily applied locally or on-site. The RPA-PfAgo method established in this study provides a strong support to the precise epidemic control and precision medicine of SARS-CoV-2 variants and can be readily developed for the simultaneous genotyping of multiple SARS-CoV-2 mutations.

Full text: Available Collection: International databases Database: MEDLINE Type of study: Prognostic study Topics: Variants Language: English Year: 2022 Document Type: Article Affiliation country: Acs.analchem.2c03563

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Full text: Available Collection: International databases Database: MEDLINE Type of study: Prognostic study Topics: Variants Language: English Year: 2022 Document Type: Article Affiliation country: Acs.analchem.2c03563