A real-time RT-quantative(q)PCR for the detection of bovine ephemeral fever virus.
A quantitative reverse-transcriptase real-time PCR assay, using TaqMan chemistry, for detecting bovine ephemeral virus (BEFV) is described. Available G gene sequences of viral RNA were aligned, and primers and probes were designed to recognize the virus. To quantitate the viruses, cDNA containing the real-time amplicon was prepared with a forward primer carrying the T7 promoter sequences. Run-off transcription from the T7 promoter amplicon template was used to prepare cRNA. Ten-fold dilutions of the run-off viral transcript were used as templates for the reaction in which they served as standards to quantitate unknown viral samples. By using this system it was shown that as few as 10-100 copies of a viral genome could be detected.
Stram
Yehuda
Y
Virology Division, Kimron Veterinary Institute, P.O. Box 12, Beit Dagan 50250, Israel. stramy@int.gov.il
Kuznetzova
Larisa
L
Levin
Aviad
A
Yadin
Hagai
H
Rubinstein-Giuni
Marisol
M
D000818:Animals; D002417:Cattle; D017931:DNA Primers; D004810:Ephemeral Fever / Q000175:diagnosis; D018115:Ephemeral Fever Virus, Bovine / Q000235:genetics / Q000302:isolation & purification*; D006023:Glycoproteins / Q000235:genetics; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012680:Sensitivity and Specificity; D015678:Viral Structural Proteins / Q000235:genetics
D017931:DNA Primers; D006023:Glycoproteins; D015678:Viral Structural Proteins