%0 Journal Article %T Establishment and application of the triplex PCR method for simultaneous detection of Cyprinid herpesvirus 2 and Cyprinid herpesvirus 3 %A PENG,Junhui %A AN,Wei %A ZHANG,Minghui %J 渔业科学进展en %@ 2095-9869 %V 42 %N 6 %D 2021 %P 158-164 %K Cyprinid herpesvirus; Triplex PCR; Carassius auratus; Cyprinus carpio %X Cyprinid herpesvirus 2 (CyHV-2) and Cyprinid herpesvirus 3 (CyHV-3) mainly infect crucian carp (goldfish), common carp, koi, and their hybrids. They are highly infectious and pathogenic viruses and can seriously damage the culture of Cyprinid fishes. In order to establish a rapid, and efficient method for simultaneously detecting the two viruses, three pairs of specific primers were designed based on the conserved sequences of the genes encoding DNA polymerase (CyHV-2, 3), helicase (CyHV-2), and thymidine kinase (CyHV-3). By optimizing the reaction conditions and system for the triplex PCR, a detection method for identifying different types of carp herpesviruses was established and validated using the Carassius auratus and Cyprinus carpio tissue samples preserved in the laboratory. The results showed that the triplex PCR method amplified only three specific bands from the herpes virus positive carp samples, showing good specificity. Using the cloned plasmid as a template with 10-fold dilution, the detection limit of the method was 2.85×102 copies/μL, showing high sensitivity. A total of 122 samples of crucian carp and 60 samples of common carp were analyzed by this method and the detection results were consistent with those of the national standard (GB/T 36194-2018) and industry standard (SC/T 7212.1-2011). In conclusion, the newly established triplex PCR method is not only highly accurate and sensitive, but can also detect two types of carp herpesvirus simultaneously, thereby effectively improve the detection efficiency. %R %U http://journal.yykxjz.cn/yykxjzen/ch/reader/view_abstract.aspx %1 JIS Version 3.0.0