Establishment of Real-time Fluorescence Loop-mediated Isothermal Assay for Detection of Classical Swine Fever Virus
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Abstract:
In order to make early diagnosis and prompt prevention of swine fever, a real-time fluorescent loop-mediated isothermal amplification method for rapid detection of classical swine fever virus (CSFV) was established. Using Primer Explorer version 4 to design 4 sets of primers for the 5'UTR non-coding region of CSFV, the best primers were selected to establish a real-time fluorescent loop-mediated isothermal amplification assay for CSFV, and the specificity, sensitivity and applicability were verified. The results showed that the real-time fluorescent loop-mediated isothermal amplification method established in this experiment was consistent with the real-time fluorescent polymerase chain reaction detection method for 128 clinical samples and 22 artificial interference sample by hog cholera live vaccine. This method only has amplified reaction for classical swine fever virus and the minimum detectable concentration was 10 fg/μL. There were no amplified reactions for eight pathogens such as virulence, highly pathogenic highly pathogenic porcine reproductive and respiratory syndrome virus and porcine circovirus type 2. The real-time fluorescent loop-mediated isothermal amplification method established in this experiment has strong specificity, high sensitivity, good detection effect and simple operation, so is suitable for rapid detection of clinical samples of CSFV.
