A multiplex reverse transcription-nested polymerase chain reaction for detection and differentiation of wild-type and vaccine strains of canine distemper virus

Si, Wei; Zhou, Shun; Wang, Zhao; Cui, Shang-jin

RT-nPCR was used to detect the cells infected with CDV vaccine strain, wild-type strain, mixed CDV vaccine and wild-type strains, CPV, CAV, CCV, RV, NDV, and uninfected cells to test its specificity. Extracted RNA from serially diluted (10 4 , 10 3 , 10 2 , 10 1 , 10 0 , 10 -1 , 10 -2 , 10 -3 TCID 50 ) CDV cell cultures (10 6.5 TCID 50 /mL) were assayed by RT-nPCR to determine its sensitivity. RT-nPCR was performed to identify cells infected with CDV vaccine strain, wild-type strain, mixed CDV vaccine and wild-type strains, field samples from Heilongjiang and Jilin province, CPV, CAV, CCV, RV, NDV, and uninfected cells three times to validate the repeatability of the test. 30 samples in Heilongjiang and Jilin provinces, and Fifty-one field samples from dogs, raccoons, foxes, and minks in Shandong provinces were assayed by RT-nPCR; the background of the 51 field samples is listed in table 1. All the positive field samples wild-type strain were confirmed by Rapid test which BioNote, Inc. produced.