Development and primary application of the SYBR Green I RT-qPCR assay for the detection of the transmissible gastroenteritis virus (TGEV) S gene

This study established a SYBR Green I real-time quantitative PCR detection assay (RT-qPCR) for the detection of the porcine transmissible gastroenteritis virus (TGEV) using specific primers designed to amplify the highly conserved porcine TGEV S gene sequence. The threshold cycle (Ct) and the log plasmid copy numbers had a good linear relationship with an efficiency of 1.05 (R2 = 0.999). The advantages of utilizing this approach for the rapid detection of TGEV include excellent sensitivity, reproducibility, and low cost. Ninety-six porcine fecal samples were tested in this study, and 7 samples more than PCR assay were detected by this assay.

[1]  C. Sánchez,et al.  Two Amino Acid Changes at the N-Terminus of Transmissible Gastroenteritis Coronavirus Spike Protein Result in the Loss of Enteric Tropism , 1997, Virology.

[2]  R. Vemulapalli,et al.  A real-time TaqMan® RT-PCR assay with an internal amplification control for rapid detection of transmissible gastroenteritis virus in swine fecal samples , 2009, Journal of Virological Methods.

[3]  Joaquín Dopazo,et al.  Genetic evolution and tropism of transmissible gastroenteritis coronaviruses , 1992, Virology.

[4]  C. Sánchez,et al.  Complete Genome Sequence of Transmissible Gastroenteritis Coronavirus PUR46-MAD Clone and Evolution of the Purdue Virus Cluster , 2004, Virus Genes.

[5]  F. Gebauer,et al.  Mechanisms of transmissible gastroenteritis coronavirus neutralization , 1990, Virology.

[6]  D. Barlič-Maganja,et al.  Real-time RT-PCR assay for rapid and specific detection of classical swine fever virus: comparison of SYBR Green and TaqMan MGB detection methods using novel MGB probes. , 2008, Journal of virological methods.

[7]  Yuriy Fofanov,et al.  Development and Characterization of a Highly Specific and Sensitive SYBR Green Reverse Transcriptase PCR Assay for Detection of the 2009 Pandemic H1N1 Influenza Virus on the Basis of Sequence Signatures , 2010, Journal of Clinical Microbiology.

[8]  Thomas A. Kuczmarski,et al.  Limitations of TaqMan PCR for Detecting Divergent Viral Pathogens Illustrated by Hepatitis A, B, C, and E Viruses and Human Immunodeficiency Virus , 2003, Journal of Clinical Microbiology.

[9]  Feng Li,et al.  Development of TaqMan Fluorescence Quantitative RT-PCR Assay for Detection of Transmissible Gastroenteritis Virus of Swine , 2007 .

[10]  Y. Fang,et al.  Simultaneous detection and genotyping of porcine reproductive and respiratory syndrome virus (PRRSV) by real-time RT-PCR and amplicon melting curve analysis using SYBR Green. , 2008, Research in veterinary science.

[11]  Jiangping Gu,et al.  Characterization of Streptococcus suis serotype 2 blood infections using RT-qPCR to quantify glutamate dehydrogenase copy numbers. , 2010, Journal of microbiological methods.

[12]  Yuelong Shu,et al.  A SYBR Green I real-time RT-PCR assay for detection and differentiation of influenza A(H1N1) virus in swine populations. , 2009, Journal of virological methods.

[13]  L. Enjuanes,et al.  Transmissible gastroenteritis coronavirus, but not the related porcine respiratory coronavirus, has a sialic acid (N-glycolylneuraminic acid) binding activity , 1996, Journal of virology.