动物造模,荧光定量PCR z-bio 2022-03-07 281

　　Objective To establish a convenient and sensitive fluorescent quantitative PCR method for detection of mink Aleutian virus (AMDV), so as to realize the rapid quantitative detection of AMDV in clinical samples.

　　Methods A pair of specific primers and probes were designed according to the conserved gene region of AMDV's structural protein VP2, the PCR reaction conditions were optimized, the TaqMan real-time quantitative PCR detection method was established, the standard curve was drawn, and the specificity, sensitivity and stability were analyzed. .

　　Results The established standard curve had a good linear relationship between 1.0×102 copies/μL and 1.0×108 copies/μL, and the correlation coefficient (r2) was greater than 0.994. The detection sensitivity of this method is 102 copies/μL, and there is no obvious cross-reaction with canine parvovirus, canine adenovirus, canine distemper virus and mink virus enteritis virus, and the specificity is good. The results of the repeatability test showed that the intra- and inter-group variation coefficients of the method were both less than 3%, and the repeatability was good. The results of testing 417 clinical tissue samples using this method showed that the AMDV positive rate in parts of China was 81.5%.

　　Conclusion This study established a reproducible, specific and sensitive method for AMDV real-time PCR detection, which can be used for clinical detection and epidemiological investigation of AMDV.