Development of a novel real-time RT-PCR assay with LUX primer for the detection of swine transmissible gastroenteritis virus.
Real-time RT-PCR assay, based on light upon extension (LUX) fluorogenic primer and LightCycle technology, was developed for rapid detection of transmissible gastroenteritis virus (TGEV). Viral RNA from different TGEV isolates and clinical specimens was detected. To evaluate the sensitivity of the assay, a gel-based RT-PCR method targeted at the same 101 bp sequence was also developed. Serial 10-fold dilutions of TGEV RNA were detected by the two methods. Although the real time method used only 2 microl RNA for each reaction, a 10-fold increase of sensitivity over that of the gel-based method, which used 10 microl RNA was demonstrated. The study indicates that the LUX assay reported below is rapid, reliable and sensitive and it has the potential for use as an alternative molecular method for TGEV diagnosis.
Chen
Ru
R
Animal Quarantine Laboratory, Guangdong Entry-Exit Inspection and Quarantine Bureau, No. 66 Huacheng Dadao, Zhujiangxincheng, Guangzhou 510623, PR China.
Huang
Weiming
W
Lin
Zhixiong
Z
Zhou
Zhongfang
Z
Yu
Haiqiong
H
Zhu
Daozhong
D
D000818:Animals; D002460:Cell Line; D017931:DNA Primers / Q000737:chemistry; D005453:Fluorescence; D005761:Gastroenteritis, Transmissible, of Swine / Q000175:diagnosis / Q000821:virology; D007421:Intestine, Small / Q000821:virology; D012367:RNA, Viral / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013552:Swine / Q000821:virology; D013997:Time Factors; D005760:Transmissible gastroenteritis virus / Q000235:genetics / Q000302:isolation & purification*
D017931:DNA Primers; D012367:RNA, Viral