Abstract
In recent years, real-time reverse transcription polymerase chain reaction (rRT-PCR) has become one of the most widely used methods for the diagnosis of infectious pathogens. The combined properties of high sensitivity, specificity, and speed, along with a low contamination risk, have made real-time PCR technology a highly attractive alternative to more conventional diagnostic methods. Numerous robust rRT-PCR systems have been developed and validated for important epizootic diseases of livestock, and in this chapter we describe an rRT-PCR protocol for the detection of bluetongue virus. The assay uses oligonucleotide primers to specifically amplify target regions of the viral genome and a dual-labeled fluorogenic (TaqMan®) probe which allows for the assay to be performed in a closed-tube format, thus minimizing the potential for cross-contamination.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Gibson UEM, Heid CA, Williams PM (1996) A novel method for real time quantitative RT PCR. Genome Res 6:995–1001
Heid CA, Stevens J, Livak KJ, Williams PM (1996) Real time quantitative PCR. Genome Res 6:986–994
Livak KJ, Flood SJA, Marmaro J, Giusti W, Deetz K (1995) Oligonucleotides with fluorescent dyes at opposite ends provide a quenched probe system useful for detecting PCR product and nucleic-acid hybridization. PCR Methods Appl 4:357–362
Hoffmann B, Beer M, Reid SM et al (2009) A review of RT-PCR technologies used in veterinary virology and disease control: sensitive and specific diagnosis of five livestock diseases notifiable to the World Organisation for Animal Health. Vet Microbiol 139:1–23
Hofmann M, Griot C, Chaignat V, Perler L, Thur B (2008) Bluetongue disease reaches Switzerland. Schweiz Arch Tierheilkd 150:49–56
Chaignat VGW, Scherrer N, Hilbe M et al (2009) Toggenburg orbivirus, a new bluetongue virus: initial detection, first observations in field and experimental infection of goats and sheep. Vet Microbiol 138:11–19
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Batten, C., Frost, L., Oura, C. (2015). Real-Time Reverse Transcriptase PCR for the Detection of Bluetongue Virus. In: Cunha, M., Inácio, J. (eds) Veterinary Infection Biology: Molecular Diagnostics and High-Throughput Strategies. Methods in Molecular Biology, vol 1247. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2004-4_8
Download citation
DOI: https://doi.org/10.1007/978-1-4939-2004-4_8
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2003-7
Online ISBN: 978-1-4939-2004-4
eBook Packages: Springer Protocols