Establishment of a Real-time Fluorescent Recombinase Polymerase Amplification (RPA) for the Detection of African Swine Fever Virus |
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| Authors: | HADENG Chu-riya FAN Xiao-xu ZHAO Yong-gang WANG Shu-juan ZHANG Zhi-cheng GE Sheng-qiang LI Lin WU Xiao-dong WANG Zhi-liang |
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| Institution: | 1. National Research Center for Exotic Animal Disease, China Animal Health and Epidemiology Center, Qingdao 266032, China;
2. College of Veterinary Medicine, Inner Mongolia Agricultural University, Hohhot 010018, China |
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| Abstract: | African swine fever (ASF), which caused by African swine fever virus (ASFV), is an acute, highly contagious disease characterized by high fever. It leads to serious economic losses in pig industry. 3 assemblies of primers and probes targeting were designed to amplify ASFV B646L (p72) gene using recombinase polymerase amplification (RPA) technology. The Real-time fluorescent RPA method was established after screening of primers and probes, optimization of reaction conditions, tests of sensitivity, specificity and repeatability. The results showed that the method could detect 10 copies of DNA within 20 min at 39℃. No cross-reaction was found when testing swine fever virus, porcine circovirus type 2, porcine parvovirus, pseudorabies virus. According to the fluorescence intensity from 5.5×106 to 5.5×100 copies/μL at each time point, the coefficient of variation was 0.38% to 28.30%. In conclusion, this method could be used for the qualitative detection of ASFV pathogen, which might provide technical support for the early diagnosis of ASFV infection in China, and was also of great significance for the development of corresponding control measures. |
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| Keywords: | African swine fever virus (ASFV) Real-time fluorescence recombinase polymerase amplification (RPA) sensitivity specificity repeatability |
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