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Focusing on the research and development and application of isothermal nucleic acid amplification technology
Development of a Real-Time Fluorescent RAA Assay for African Swine Fever Virus—Nanjing Agricultural University & Animal Health
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To establish a simple and efficient grassroots clinical detection method for African swine fever virus (ASFV), this study selected the conserved gene B646L (P72) sequence of ASFV, designed specific primers and probes, and developed a real-time fluorescent recombinase-aided isothermal amplification (RAA) assay for ASFV detection. The results showed that the developed method exhibits comparable sensitivity to the fluorescence quantitative PCR method recommended by the OIE, with a detection limit as low as 10 copies/μL. The entire detection process takes only 10 minutes. Moreover, the assay demonstrates no cross-reactivity with foot-and-mouth disease virus (FMDV), porcine parvovirus (PPV), pseudorabies virus (PRV), porcine circovirus type 2 (PCV2), porcine reproductive and respiratory syndrome virus (PRRSV), or classical swine fever virus (CSFV). The detection results for 100 clinical samples were consistent with those obtained by the OIE-recommended fluorescence quantitative PCR method. However, compared to the 2-hour detection time required for fluorescence quantitative PCR, the RAA assay significantly reduces the detection time. These results indicate that the method developed in this study features high sensitivity and good specificity, providing technical support for the early clinical diagnosis of ASFV.
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