Development of a multiplex real-time reverse transcriptase-polymerase chain reaction for equine infectious anemia virus (EIAV).
A single-tube reverse transcriptase-polymerase chain reaction (RT-PCR) using a fluorogenic real-time PCR detection method is described for the quantitation of equine infectious anemia virus (EIAV) RNA in the plasma of equids. To compensate for variations inherent in sample preparation a multiplex real-time RT-PCR system was developed that permitted the simultaneous calculation of the nucleic acid recovery rate along with the copy number of viral RNA molecules. Detection of EIAV RNA was linear from 10(9) to 10(1) molecules with intra- and inter-assay variability of less than 1% at 10(8), 10(6), 10(4) and 10(2) molecules.
Cook
R Frank
RF
Department of Veterinary Science, Gluck Equine Research Center, University of Kentucky, Lexington, KY 40546-0099, USA. rfcook@uky.edu
Cook
S J
SJ
Li
F Li
FL
Montelaro
R C
RC
Issel
C J
CJ
D000818:Animals; D015342:DNA Probes; D004859:Equine Infectious Anemia / Q000821:virology*; D005456:Fluorescent Dyes; D018628:Gene Dosage; D006736:Horses; D004860:Infectious Anemia Virus, Equine / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D012367:RNA, Viral / Q000097:blood*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019562:Viral Load
D015342:DNA Probes; D005456:Fluorescent Dyes; D012367:RNA, Viral