Detection of equine arteritis virus (EAV) by polymerase chain reaction (PCR) and differentiation of EAV strains by restriction enzyme analysis of PCR products.
A polymerase chain reaction (PCR) based assay capable of detecting and differentiating seven strains of equine arteritis virus (EAV) from around the world was developed. The primers for the PCR were chosen from the ORF6 gene encoding the unglycosylated membrane protein (M). Viral RNA from cell culture fluids infected with each of the seven EAV strains and RNA from the live vaccine, Arvac, was detected by PCR using four sets of primers. The sensitivity of detection was increased from 100 to 1,000 times by performing nested PCR enabling the detection of RNA at a level of 0.5-5 PFU. Differentiation among the virus strains and the live vaccine was achieved by cutting the PCR-amplified products from three sets of primers with six restriction endonucleases. Using this procedure it was possible to distinguish among the seven EAV strains used.
Sekiguchi
K
K
Epizootic Research Station, Equine Research Institute, Japan Racing Association, Tochigi-Ken.
Sugita
S
S
Fukunaga
Y
Y
Kondo
T
T
Wada
R
R
Kamada
M
M
Yamaguchi
S
S
D001483:Base Sequence; D017931:DNA Primers; D004262:DNA Restriction Enzymes; D007240:Equartevirus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000032:analysis* / Q000235:genetics; D012680:Sensitivity and Specificity; D014763:Viral Matrix Proteins / Q000235:genetics
D017931:DNA Primers; D012367:RNA, Viral; D014763:Viral Matrix Proteins; D004262:DNA Restriction Enzymes