Development of a Lightcycler-based reverse transcription polymerase chain reaction for the detection of foot-and-mouth disease virus.
One step TaqMan real-time reverse transcription polymerase chain reaction (R/T RT-PCR) using a set of primers/probes was developed for the detection of foot-and-mouth disease (FMD) virus. The gene-specific probes labeled fluorogen for the internal ribosomal entry site, Leader sequence and 2B regions were used to detect FMD virus (FMDV). This assay specifically detected FMDV both in cell culture preparations and clinical samples, and was capable of distinguishing FMD from other viral diseases similar to clinical signs (swine vesicular disease, vesicular stomatitis and bovine viral diarrhea). This assay was shown to be 1000-fold more sensitive than the conventional RT-PCR method. The detection limits of this assay was 1 TCID(50)/ml of the FMDV RNA concentration. Quantification was obtained by a standard curves plotting threshold cycle values versus known infectivity titer. The assay was sensitive, specific and rapid enough to detect FMDV RNA genome in probang samples. As such, the described method is reliable and provides faster disease diagnostics than the conventional RT-PCR procedure to detect FMDV.
Oem
Jae Ku
JK
National Veterinary Research and Quarantine Service, Ministry of Agriculture and Forestry, Anyang, Korea. jku0622@nvrqs.go.kr
Kye
Soo Jeong
SJ
Lee
Kwang Nyeong
KN
Kim
Yong Joo
YJ
Park
Jee Yong
JY
Park
Jong Hyeon
JH
Joo
Yi Seok
YS
Song
Hee Jong
HJ
D000818:Animals; D005536:Foot-and-Mouth Disease / Q000175:diagnosis* / Q000821:virology; D029544:Foot-and-Mouth Disease Virus / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D019914:Taq Polymerase
D019914:Taq Polymerase