Development and validation of SYBR Green- and probe-based reverse-transcription real-time PCR assays for detection of the S and M segments of Schmallenberg virus
| Autor: | Ahmet Kursat Azkur, Wim H.M. van der Poel, Murat Yildirim, Renate Hakze-van der Honing, Kader Yildiz, Emel Aksoy |
|---|---|
| Přispěvatelé: | KKÜ |
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
Orthobunyavirus
Cattle Diseases Sheep Diseases Biology Diamines Bunyaviridae Infections Real-Time Polymerase Chain Reaction Brief Communication Genome S segment Animals RT-rtPCR SYBR Green Benzothiazoles Organic Chemicals Detection limit Sheep General Veterinary Reverse Transcriptase Polymerase Chain Reaction M segment Schmallenberg virus Congenital malformations biology.organism_classification Virology Reverse transcriptase Hypervariable region Virology & Molecular Biology Virologie & Moleculaire Biologie Real-time polymerase chain reaction Quinolines Cattle Primer (molecular biology) |
| Zdroj: | Journal of Veterinary Diagnostic Investigation, 32(5), 710-717 J Vet Diagn Invest Journal of Veterinary Diagnostic Investigation 32 (2020) 5 |
| ISSN: | 1040-6387 |
| Popis: | WOS:000556848300001 PubMed: 32757829 Schmallenberg virus (SBV), discovered in Germany in 2011, causes congenital malformations in ruminants. Reverse-transcription real-time PCR (RT-rtPCR) assays based on various segments of SBV have been developed for molecular detection. We developed alternative RT-rtPCR assays for SBV detection to avoid earlier reported mutations and hypervariable regions of the S and M segments of the viral genome. For SYBR Green-based detection of the S segment, theR(2)value and efficiency of the developed assay were 0.99 and 99%, respectively. For probe-based S segment detection, 2 assays were developed; the first had anR(2)value of 0.99 and 102% efficiency, and the second had aR(2)value of 0.98 and 86% efficiency. The probe-based M segment assay had anR(2)value of 1.00 and 103% efficiency. Detection limits of the RT-rtPCR assays with new primer sets were 10(2)and 10(1)copies/mu L for the S and M segments, respectively. Field samples from cattle and sheep were also used for primary validation of the developed assays. Our assays should be suitable for SBV detection in ruminants and for in vitro studies of various SBV strains. General Directorate of Agricultural Research and Policies, Ministry of Agriculture and Forestry, Republic of Turkey [TAGEM15/AR-GE48] This study is a part of a project supported by the General Directorate of Agricultural Research and Policies, Ministry of Agriculture and Forestry, Republic of Turkey (grant TAGEM15/AR-GE48). |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|