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Absolute quantification of cassava brown streak virus mRNA by real-time qPCR

dc.contributor.authorShirima, R.R.
dc.contributor.authorMaeda, D.G
dc.contributor.authorKanju, E.
dc.contributor.authorCeasar, G.
dc.contributor.authorTibazarwa, F.I.
dc.contributor.authorLegg, J.P.
dc.date.accessioned2019-12-04T11:08:03Z
dc.date.available2019-12-04T11:08:03Z
dc.date.issued2017
dc.identifier.citationShirima, R.R., Maeda, D.G., Kanju, E., Ceasar, G., Tibazarwa, F.I. & Legg, J. (2017). Absolute quantification of cassava brown streak virus mRNA by real-time qPCR. Journal of Virological Methods, 245, 1-13.
dc.identifier.issn0166-0934
dc.identifier.urihttps://hdl.handle.net/20.500.12478/1707
dc.descriptionArticle purchased
dc.description.abstractCassava brown streak disease (CBSD) is the most important virus disease of cassava and a major food security threat in Africa. Yearly economic losses of up to $100 million USD have been attributed to CBSD. The lack of information on plant-virus interactions has restricted progress in breeding for CBSD resistance. Virus quantification is becoming a major tool for the quick and reliable assessment of plant host resistance. Therefore, a protocol for specific absolute quantification of Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) was developed. CBSV and UCBSV coat protein (CP) specific standard templates: CBSV (pFer2, 826 bp) and UCBSV (pUF1-R1-1, 732) respectively were generated and maintained in a TA cloning vector. These were used to construct standard curves using a TaqMan qPCR assay. Standard curves with acceptable amplification efficiencies (90–105%) and coefficients of determination (R2) greater than 0.99 were obtained. Infected cassava plants were sampled from a screenhouse and the field and used to validate this assay. Results obtained by testing several screenhouse and field samples revealed consistent absolute quantification assays for different CBSV and UCBSV isolates. This study presents the first protocol for absolute quantification of CBSVs and is expected to accelerate screening for CBSD resistance and hence breeding for CBSD resistance. The use of the method presented here should improve the clarity of virus quantification data as the results obtained are not influenced by varietal, host, seasonal or environmental conditions. Screening efficiency will also be greatly improved as there is no need for the use of reference genes consequently allowing for a larger number of samples to be analyzed. This will increase experimental precision in a timely and cost effective manner.
dc.description.sponsorshipBill & Melinda Gates Foundation
dc.format.extent5-13
dc.language.isoen
dc.subjectCassava
dc.subjectVirus
dc.subjectFood Sceurity
dc.subjectDna
dc.subjectCassava Brown Streak Disease
dc.subjectVirus Titre
dc.titleAbsolute quantification of cassava brown streak virus mRNA by real-time qPCR
dc.typeJournal Article
dc.description.versionPeer Review
cg.contributor.affiliationInternational Institute of Tropical Agriculture
cg.contributor.affiliationUniversity of Dar es Salaam
cg.contributor.affiliationTanzania Commission for Science and Technology
cg.coverage.regionAfrica
cg.coverage.regionCentral Africa
cg.coverage.regionEast Africa
cg.coverage.countryBurundi
cg.coverage.countryCongo, Dr
cg.coverage.countryTanzania
cg.isijournalISI Journal
cg.authorship.typesCGIAR and developing country institute
cg.iitasubjectCassava
cg.iitasubjectPlant Diseases
cg.journalJournal of Virological Methods
cg.howpublishedFormally Published
cg.accessibilitystatusOpen Access
local.dspaceid82965
cg.targetaudienceScientists
cg.identifier.doihttp://doi.org/10.1016/j.jviromet.2017.03.003


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