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dc.contributor.author
Gonzalez, Mariana Selena
dc.contributor.author
de Brasi, Carlos Daniel
dc.contributor.author
Bianchini, Michele
dc.contributor.author
Gargallo, Patricia Martha
dc.contributor.author
Stanganelli, Carmen Graciela
dc.contributor.author
Zalcberg, Ilana
dc.contributor.author
Larripa, Irene Beatriz
dc.date.available
2015-05-19T18:34:48Z
dc.date.issued
2013-12-27
dc.identifier.citation
Gonzalez, Mariana Selena; de Brasi, Carlos Daniel; Bianchini, Michele; Gargallo, Patricia Martha; Stanganelli, Carmen Graciela; Zalcberg, Ilana; Larripa, Irene Beatriz; Improved Diagnosis of the Transition to JAK2V617F Homozygosity: The Key Feature for Predicting the Evolution of Myeloproliferative Neoplasms; Public Library Science; Plos One; 9; 1; 27-12-2013; 1-8;
dc.identifier.issn
1932-6203
dc.identifier.uri
http://hdl.handle.net/11336/507
dc.description.abstract
Most cases of BCR-ABL1-negative myeloproliferative neoplasms (MPNs), essential thrombocythemia, polycythemia vera and primary myelofibrosis are associated with JAK2V617F mutations. The outcomes of these cases are critically influenced by the transition from JAK2V617F heterozygosity to homozygosity. Therefore, a technique providing an unbiased assessment of the critical allele burden, 50% JAK2V617F, is highly desirable. In this study, we present an approach to assess the JAK2V617F burden from genomic DNA (gDNA) and complementary DNA (cDNA) using one-plus-one template references for allele-specific quantitative-real-time-PCR (qPCR). Plasmidic gDNA and cDNA constructs encompassing one PCR template for JAK2V617F spaced from one template for JAK2Wild Type were constructed by multiple fusion PCR amplifications. Repeated assessments of the 50% JAK2V617F burden within the dynamic range of serial dilutions of gDNA and cDNA constructs resulted in 52.5364.2% and 51.4664.21%, respectively. The mutation-positive cutoff was estimated to be 3.65% (mean +2 standard deviation) using 20 samples from a healthy population. This qPCR approach was compared with the qualitative ARMS-PCR technique and with two standard methods based on qPCR, and highly significant correlations were obtained in all cases. qPCR assays were performed on paired gDNA/cDNA samples from 20 MPN patients, and the JAK2V617F expression showed a significant correlation with the allele burden. Our data demonstrate that the qPCR method using one-plus-one template references provides an improved assessment of the clinically relevant transition of JAK2V617F from heterozygosity to homozygosity.
dc.format
application/pdf
dc.language.iso
eng
dc.publisher
Public Library Science
dc.rights
info:eu-repo/semantics/openAccess
dc.rights.uri
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.subject
Jak2 V617f
dc.subject
Qpcr
dc.subject
Gdna
dc.subject
Cdna
dc.subject.classification
Ciencias Médicas y de la Salud
dc.subject.classification
Biotecnología de la Salud
dc.subject.classification
Tecnologías Que Involucran la Identificación de Adn, Proteínas y Enzimas, y Cómo Influyen En El Conjunto de Enfermedades y Mantenimiento del Bienestar
dc.title
Improved Diagnosis of the Transition to JAK2V617F Homozygosity: The Key Feature for Predicting the Evolution of Myeloproliferative Neoplasms
dc.type
info:eu-repo/semantics/publishedVersion
dc.type
info:ar-repo/semantics/artículo
dc.type
info:eu-repo/semantics/article
dc.date.updated
2016-03-30 10:35:44.97925-03
dc.journal.volume
9
dc.journal.number
1
dc.journal.pagination
1-8
dc.journal.pais
Estados Unidos
dc.journal.ciudad
San Francisco
dc.description.fil
Fil: Gonzalez, Mariana Selena. Consejo Nacional de Invest.cientif.y Tecnicas. Instituto de Medicina Experimental;
dc.description.fil
Fil: de Brasi, Carlos Daniel. Consejo Nacional de Invest.cientif.y Tecnicas. Instituto de Medicina Experimental;
dc.description.fil
Fil: Bianchini, Michele. DTO. DE GENETICA;
dc.description.fil
Fil: Gargallo, Patricia Martha. INST. DE INVEST. HEMATOLOGICAS;
dc.description.fil
Fil: Stanganelli, Carmen Graciela. Academia Nacional de Medicina de Buenos Aires. Instituto de Invest. Hematologicas "mariano R. Castex";
dc.description.fil
Fil: Zalcberg, Ilana. Molecular Biology, Laboratory, Instituto Nacional do Caˆncer, Rio de Janeiro, Brazil;
dc.description.fil
Fil: Larripa, Irene Beatriz. Consejo Nacional de Invest.cientif.y Tecnicas. Instituto de Medicina Experimental;
dc.journal.title
Plos One
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/doi/http://dx.doi.org/10.1371/journal.pone.0086401
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/url/http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0086401
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