Establishment and application of TaqMan probe real-time fluorescence quantitative PCR method for detection of infectious laryngotracheitis in chickens
To establish a rapid diagnostic method for chicken infectious laryngotracheitis virus(ILTV),specific probes and primers were designed based on the ILTV gB gene,and a real-time fluorescent quantitative PCR(qPCR)method based on the TaqMan probe was established.The results showed that the standard curve of the ILTV qPCR method was y=-3.892 9x+44.607 with a linear correlation value(R2)of 0.998 8.The lowest detection limit of the method was 1.0× 10 copies/μL,and the coefficients of variation for both intra-and inter-batch reproducibility were less than 5.64％,and there was no cross-reactivity with other chicken-derived pathogens.The test results of the clinical samples showed a positive detection rate of 98.7％for the qPCR method compared to 92.61％for conventional PCR.The above results indicate that the method is sensitive,specific,and reproducible,and can be used for epidemic surveillance and epidemiologi-cal investigation of ILTV,providing a powerful tool for the prevention and diagnosis of ILTV infection.
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