Location: Vegetable Crops ResearchTitle: Development and validation of quantitative polymerase chain reaction protocols targeting the ‘L’, ‘M’, and ‘S’ ribonucleic acid of Soybean vein necrosis virus Author
|Smith, D - University Of Wisconsin|
|Groves, C - University Of Wisconsin|
|Fritz, C - University Of Wisconsin|
Submitted to: American Phytopathological Society
Publication Type: Abstract Only
Publication Acceptance Date: 5/23/2013
Publication Date: 8/10/2013
Citation: Smith, D.L., Groves, C.L., Fritz, C., Willis, D.K. 2013. Development and validation of quantitative polymerase chain reaction protocols targeting the ‘L’, ‘M’, and ‘S’ ribonucleic acid of Soybean vein necrosis virus [abstract]. American Phytopathological Society. Paper No. 399-P.
Technical Abstract: Soybean vein necrosis virus (SVNV) was first reported in Wisconsin in 2012. SVNV is a new member of the Tospovirus family and is becoming more frequent in occurrence in the north central region USA. New real-time reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) protocols were developed for detection and quantification of SVNV particles in plants. Primer sets were developed to target each of the ribonucleic (RNA) segments of the tripartite genome of SVNV and optimized for use in RT-qPCR. We developed primers targeting the ‘L’ RNA and the ‘M’ RNA using GenBank accessions HQ728385 and HQ728386. While primers targeting the nucleoprotein coding region of the ‘S’ RNA were based on unique sequences of SVNV isolates obtained in Wisconsin. We established the sensitivity of SVNV detection using three methods: PCR, RT-qPCR, and nested PCR. Our findings indicate that nested PCR is required to reliably detect SVNV ‘S’ RNA present at less than 10 copies per sample. These qPCR methods will be used for future SVNV epidemiological studies.