Innovative Technology

Innovation

Focusing on the research and development and application of isothermal nucleic acid amplification technology

Development of a Recombinase-Aided Nucleic Acid Detection Method for Middle East Respiratory Syndrome Coronavirus – Ningbo National Travel & Ningbo Inspection and Quarantine

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Abstract: Objective To establish a rapid, sensitive, and specific reverse-transcription recombinase-aided assay (RT-RAA) for the detection of Middle East Respiratory Syndrome Coronavirus (MERS-CoV). Methods Specific RAA primers and probes were designed and synthesized based on the conserved sequences of the MERS-CoV genome. After optimizing reaction conditions, RT-RAA amplification was performed using a 10-fold serial dilution of pseudovirus particles generated from third-generation lentiviral vectors as the template. The specificity, sensitivity, and performance in clinical samples were evaluated. Results The established RT-RAA method demonstrated a detection sensitivity of 10 copies/mL for MERS-CoV pseudovirus particle standards, which is higher than that of real-time fluorescent RT-PCR (10 copies/mL). The RT-RAA assay showed clear amplification when detecting MERS-CoV pseudovirus particles, while no amplification was observed for four other respiratory viruses, including influenza A H1N1, influenza B, respiratory syncytial virus, and rhinovirus. The method successfully detected both clinical samples from China’s first imported case of MERS and positive inactivated samples provided by the UK OCMD proficiency testing program, with results consistent with the actual outcomes. Moreover, the assay’s detection time (20 minutes) was significantly shorter than that of real-time fluorescent RT-PCR (90 minutes). Conclusion The developed MERS-CoV RT-RAA assay is sensitive, specific, and rapid, making it suitable for rapid on-site detection of MERS-CoV infections and epidemiological investigations.

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Methodological Study on Rapid Detection of Human Rhinovirus by Isothermal Amplification Mediated by Recombinase—Chen Shudan

Development and Application of an RT_RAA Rapid Detection Method for Middle East Respiratory Syndrome Coronavirus—Ningbo National Travel & Ningbo Inspection and Quarantine

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Methodological Study on Rapid Detection of Human Rhinovirus by Isothermal Amplification Mediated by Recombinase—Chen Shudan

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Development and Application of an RT_RAA Rapid Detection Method for Middle East Respiratory Syndrome Coronavirus—Ningbo National Travel & Ningbo Inspection and Quarantine