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Co-operational PCR coupled with dot blot hybridization for the detection of Phaeomoniella chalmydospora on infected grapevine wood

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Co-operational PCR coupled with dot blot hybridization for the detection of Phaeomoniella chalmydospora on infected grapevine wood

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dc.contributor.author Martos, Soledad es_ES
dc.contributor.author Torres, Ester es_ES
dc.contributor.author bdessamad El Bakali, Mohamed es_ES
dc.contributor.author Raposo, Rosa es_ES
dc.contributor.author Gramaje, David es_ES
dc.contributor.author Armengol Fortí, Josep es_ES
dc.contributor.author Luque, Jordi es_ES
dc.date.accessioned 2017-05-03T12:09:09Z
dc.date.available 2017-05-03T12:09:09Z
dc.date.issued 2011-04
dc.identifier.issn 0931-1785
dc.identifier.uri http://hdl.handle.net/10251/80508
dc.description.abstract [EN] The technique consisting of the co-operational PCR coupled with dot blot hybridization and posterior colorimetric visualization was developed for the detection of Phaeomoniella chlamydospora, one of the major pathogenic fungi involved in the Petri disease of grapevine. A partial region of the fungal rDNA including the internal transcribed spacer (ITS) region was amplified through co-operational PCR for P. chlamydospora and 17 additional grapevine-associated fungi included in the genera Botryosphaeria, Cryptovalsa, Cylindrocarpon, Dematophora, Diplodia, Dothiorella, Eutypa, Fomitiporia, Lasiodiplodia, Neofusicoccum, Phaeoacremonium, Phomopsis and Stereum, by using the primer pairs NSA3/NLC2 (external pair) and NSI1/NLB4 (inner pair). A specific probe (Pch2D) targeting the ITS2 region in the rDNA was developed for the detection of P. chlamydospora. Dot blot hybridizations carried out with the PCR products showed the specificity of the probe. Results indicated that Pch2D only hybridized with DNA amplicons of P. chlamydospora isolates, thus proving the specific detection of this fungus, while the 17 remaining species tested for the Pch2D probe resulted in negative results. Sensitivity of the technique was established below 0.1 pg of genomic DNA. This technique was further validated using artificially inoculated grapevine cuttings with P. chlamydospora. The efficacy of detection was established at 80% after two independent blind assays. es_ES
dc.description.sponsorship This work was financed by the Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria (INIA) under projects RTA03-058-C2-1 and RTA2007-00023-C04 (Spain) and the European Regional Development Fund. Soledad Martos was supported by the Departament d'Educacio i Universitats de la Generalitat de Catalunya (Regional Government of Catalonia, Spain) and the European Social Fund. David Gramaje was supported by a grant from the Universidad Politecnica de Valencia (FPI-UPV). The authors thank J. Bech for his contribution to the laboratory work. en_EN
dc.language Inglés es_ES
dc.publisher Wiley es_ES
dc.relation Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria (INIA) u[RTA03-058-C2-1] [RTA2007-00023-C04] es_ES
dc.relation.ispartof Journal of Phytopathology es_ES
dc.rights Reserva de todos los derechos es_ES
dc.subject Co-operational PCR es_ES
dc.subject Detection, es_ES
dc.subject Petri disease es_ES
dc.subject Vitis vinifera es_ES
dc.subject.classification PRODUCCION VEGETAL es_ES
dc.title Co-operational PCR coupled with dot blot hybridization for the detection of Phaeomoniella chalmydospora on infected grapevine wood es_ES
dc.type Artículo es_ES
dc.identifier.doi 10.1111/j.1439-0434.2010.01758.x
dc.rights.accessRights Cerrado es_ES
dc.contributor.affiliation Universitat Politècnica de València. Escuela Técnica Superior de Ingeniería Agronómica y del Medio Natural - Escola Tècnica Superior d'Enginyeria Agronòmica i del Medi Natural es_ES
dc.description.bibliographicCitation Martos, S.; Torres, E.; Bdessamad El Bakali, M.; Raposo, R.; Gramaje, D.; Armengol Fortí, J.; Luque, J. (2011). Co-operational PCR coupled with dot blot hybridization for the detection of Phaeomoniella chalmydospora on infected grapevine wood. Journal of Phytopathology. 159(4):247-254. doi:10.1111/j.1439-0434.2010.01758.x es_ES
dc.description.accrualMethod Senia es_ES
dc.relation.publisherversion http://doi.org/10.1111/j.1439-0434.2010.01758.x es_ES
dc.description.upvformatpinicio 247 es_ES
dc.description.upvformatpfin 254 es_ES
dc.type.version info:eu-repo/semantics/publishedVersion es_ES
dc.description.volume 159 es_ES
dc.description.issue 4 es_ES
dc.relation.senia 213769 es_ES


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