Successful application of FTA Classic Card technology and use of bacteriophage phi29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomes

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dc.contributor.authorOwor, Betty E
dc.contributor.authorShepherd, Dionne N
dc.contributor.authorTaylor, Nigel J
dc.contributor.authorEdema, Richard
dc.contributor.authorMonjane, Aderito L
dc.contributor.authorThomson, Jennifer A
dc.contributor.authorMartin, Darren P
dc.contributor.authorVarsani, Arvind
dc.date.accessioned2016-07-26T08:02:06Z
dc.date.available2016-07-26T08:02:06Z
dc.date.issued2007
dc.date.updated2016-07-15T15:24:26Z
dc.description.abstractLeaf samples from 155 maize streak virus (MSV)-infected maize plants were collected from 155 farmers’ fields in 23 districts in Uganda in May/June 2005 by leaf-pressing infected samples onto FTA® Classic Cards. Viral DNA was successfully extracted from cards stored at room temperature for 9 months. The diversity of 127 MSV isolates was analysed by PCR-generated RFLPs. Six representative isolates having different RFLP patterns and causing either severe, moderate or mild disease symptoms, were chosen for amplification from FTA cards by bacteriophage 29 DNA polymerase using the TempliPhiTM system. Full-length genomes were inserted into a cloning vector using a unique restriction enzyme site, and sequenced. The 1.3-kb PCR product amplified directly from FTA-eluted DNA and used for RFLP analysis was also cloned and sequenced. Comparison of cloned whole genome sequences with those of the original PCR products indicated that the correct virus genome had been cloned and that no errors were introduced by the 29 polymerase. This is the first successful large-scale application of FTA card technology to the field, and illustrates the ease with which large numbers of infected samples can be collected and stored for downstream molecular applications such as diversity analysis and cloning of potentially new virus genomes.en_ZA
dc.identifierhttp://dx.doi.org/10.1016/j.jviromet.2006.11.004
dc.identifier.apacitationOwor, B. E., Shepherd, D. N., Taylor, N. J., Edema, R., Monjane, A. L., Thomson, J. A., ... Varsani, A. (2007). Successful application of FTA Classic Card technology and use of bacteriophage phi29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomes. <i>Journal of Virological Methods</i>, http://hdl.handle.net/11427/20739en_ZA
dc.identifier.chicagocitationOwor, Betty E, Dionne N Shepherd, Nigel J Taylor, Richard Edema, Aderito L Monjane, Jennifer A Thomson, Darren P Martin, and Arvind Varsani "Successful application of FTA Classic Card technology and use of bacteriophage phi29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomes." <i>Journal of Virological Methods</i> (2007) http://hdl.handle.net/11427/20739en_ZA
dc.identifier.citationOwor, B. E., Shepherd, D. N., Taylor, N. J., Edema, R., Monjane, A. L., Thomson, J. A., ... & Varsani, A. (2007). Successful application of FTA® Classic Card technology and use of bacteriophage ϕ29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomes. Journal of virological methods, 140(1), 100-105.en_ZA
dc.identifier.issn0166-0934en_ZA
dc.identifier.ris TY - Journal Article AU - Owor, Betty E AU - Shepherd, Dionne N AU - Taylor, Nigel J AU - Edema, Richard AU - Monjane, Aderito L AU - Thomson, Jennifer A AU - Martin, Darren P AU - Varsani, Arvind AB - Leaf samples from 155 maize streak virus (MSV)-infected maize plants were collected from 155 farmers’ fields in 23 districts in Uganda in May/June 2005 by leaf-pressing infected samples onto FTA® Classic Cards. Viral DNA was successfully extracted from cards stored at room temperature for 9 months. The diversity of 127 MSV isolates was analysed by PCR-generated RFLPs. Six representative isolates having different RFLP patterns and causing either severe, moderate or mild disease symptoms, were chosen for amplification from FTA cards by bacteriophage 29 DNA polymerase using the TempliPhiTM system. Full-length genomes were inserted into a cloning vector using a unique restriction enzyme site, and sequenced. The 1.3-kb PCR product amplified directly from FTA-eluted DNA and used for RFLP analysis was also cloned and sequenced. Comparison of cloned whole genome sequences with those of the original PCR products indicated that the correct virus genome had been cloned and that no errors were introduced by the 29 polymerase. This is the first successful large-scale application of FTA card technology to the field, and illustrates the ease with which large numbers of infected samples can be collected and stored for downstream molecular applications such as diversity analysis and cloning of potentially new virus genomes. DA - 2007 DB - OpenUCT DP - University of Cape Town J1 - Journal of Virological Methods LK - https://open.uct.ac.za PB - University of Cape Town PY - 2007 SM - 0166-0934 T1 - Successful application of FTA Classic Card technology and use of bacteriophage phi29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomes TI - Successful application of FTA Classic Card technology and use of bacteriophage phi29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomes UR - http://hdl.handle.net/11427/20739 ER - en_ZA
dc.identifier.urihttp://hdl.handle.net/11427/20739
dc.identifier.vancouvercitationOwor BE, Shepherd DN, Taylor NJ, Edema R, Monjane AL, Thomson JA, et al. Successful application of FTA Classic Card technology and use of bacteriophage phi29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomes. Journal of Virological Methods. 2007; http://hdl.handle.net/11427/20739.en_ZA
dc.languageengen_ZA
dc.publisherElsevieren_ZA
dc.publisher.institutionUniversity of Cape Town
dc.rightsCreative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0)*
dc.rights.urihttps://creativecommons.org/licenses/by-nc-nd/4.0/en_ZA
dc.sourceJournal of Virological Methodsen_ZA
dc.source.urihttp://www.sciencedirect.com/science/journal/01660934
dc.subject.otherMaize streak virus (MSV)
dc.subject.otherGeminivirus
dc.subject.otherFTA cards
dc.subject.otherRolling circle amplification
dc.subject.otherPhi29 DNA polymerase
dc.titleSuccessful application of FTA Classic Card technology and use of bacteriophage phi29 DNA polymerase for large-scale field sampling and cloning of complete maize streak virus genomesen_ZA
dc.typeJournal Articleen_ZA
uct.type.filetypeText
uct.type.filetypeImage
uct.type.publicationResearchen_ZA
uct.type.resourceArticleen_ZA
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