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A Real-Time Polymerase Chain Reaction Assay for Rapid, Sensitive, and Specific Quantification of the JAK2V617F Mutation Using a Locked Nucleic Acid-Modified.

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Presentation on theme: "A Real-Time Polymerase Chain Reaction Assay for Rapid, Sensitive, and Specific Quantification of the JAK2V617F Mutation Using a Locked Nucleic Acid-Modified."— Presentation transcript:

1 A Real-Time Polymerase Chain Reaction Assay for Rapid, Sensitive, and Specific Quantification of the JAK2V617F Mutation Using a Locked Nucleic Acid-Modified Oligonucleotide  Barbara Denys, Hakim El Housni, Friedel Nollet, Bruno Verhasselt, Jan Philippé  The Journal of Molecular Diagnostics  Volume 12, Issue 4, Pages (July 2010) DOI: /jmoldx Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

2 Figure 1 Schematic overview of the mutation-specific LNA-based qPCR reaction. At the left part of the figure, the perfect matching between the hydrolysis probe and the mutated DNA sequence allows annealing of the probe to the target DNA and consequently the Taq polymerase can hydrolyze the probe. In contrast, the single bp mismatch between the LNA oligonucleotide sequence and the mutated DNA sequence prevents specifically its annealing. At the right part of the figure, the LNA oligonucleotide anneals to wild-type target DNA, thereby preventing the detection of wild-type JAK2 sequences. The LNA oligonucleotide thus allows preferential amplification of the mutated DNA if present. Moreover, the mutated specific hydrolysis probe cannot efficiently hybridize to wild-type target because of a single bp mismatch between both. LNA indicates locked nucleic acid. The Journal of Molecular Diagnostics  , DOI: ( /jmoldx ) Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

3 Figure 2 Proportion of JAK2V617F allele burden of the 36 JAK2 mutant patients included in this study. Box-and-Whisker presentation of the results from patients with PV (n = 11), ET (n = 14), PMF (n = 8), and post-PV MF (n = 3). The central box represents 25th to 75th percentile, and the middle line the median. The black vertical line extends from the minimum to the maximum value, excluding “outside” and “far out” values. P values are represented for comparisons between all disease entities. PV indicates polycythemia vera; ET, essential thrombocythemia; PMF, primary myelofibrosis; post-PV MF, post-polycythemic myelofibrosis. The Journal of Molecular Diagnostics  , DOI: ( /jmoldx ) Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

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