Abstract
We developed and validated a real-time polymerase chain reaction assay using fluorescent hybridization probes and melting curve analysis to identify the JAK2 V617F mutation, which is implicated in a substantial proportion of chronic myeloproliferative disorders (CMPDs). DNA from 161 samples was isolated from peripheral blood granulocytes and formalin-fixed bone marrow clot sections in patients with CMPDs and without myeloproliferative disorders previously genotyped for the JAK2 V617F (G-->T) mutation, which included 114 wild types (GG) and 47 mutants (GT and TT). Melting curve analysis of these samples yielded 114 wild types, 42 heterozygotes, and 5 homozygotes showing 100% concordance. Analytic sensitivity of the assay for mutant DNA was 5% for the LightTyper (Roche Applied Sciences, Indianapolis, IN) and 10% for the LightCycler (Roche Applied Sciences). Consistent with earlier reports, 78% of the non-chronic myelogenous leukemia CMPD patients and 8% of non-CMPD patients displayed this mutation. This study demonstrates that clinical genotyping of the JAK2 V617F mutation can be performed by melting analysis using both freshly isolated and formalin-fixed tissues.
References
Jan 1, 1997·Seminars in Thrombosis and Hemostasis·J J Michiels, E Juvonen
May 30, 2001·Experimental Hematology·S RöderH L Pahl
Mar 23, 2005·Lancet·E Joanna BaxterUNKNOWN Cancer Genome Project
Mar 29, 2005·Nature·Chloé JamesWilliam Vainchenker
Apr 20, 2005·Cancer Cell·Ross L LevineD Gary Gilliland
Apr 22, 2005·British Journal of Haematology·Rose C BeckJaroslaw P Maciejewski
Apr 29, 2005·The New England Journal of Medicine·Robert KralovicsRadek C Skoda
Apr 30, 2005·Blood·David P SteensmaAyalew Tefferi
May 3, 2005·The Journal of Biological Chemistry·Runxiang ZhaoZhizhuang Joe Zhao
May 26, 2005·Blood·Marcin W WlodarskiJaroslaw P Maciejewski
May 28, 2005·Blood·Amy V JonesNicholas C P Cross
Jun 7, 2005·Expert Review of Molecular Diagnostics·Gurunathan MurugesanRaymond Tubbs
Jun 23, 2005·Cell Cycle·Ayalew Tefferi, D Gary Gilliland
Jul 23, 2005·Blood·Jaroslav JelinekJean-Pierre J Issa
Aug 5, 2005·Leukemia·E AntonioliA M Vannucchi
Aug 6, 2005·Blood·Ross L LevineMichael W N Deininger
Aug 27, 2005·Molecular Interventions·Marko PesuOlli Silvennoinen
Sep 13, 2005·Journal of Clinical Oncology : Official Journal of the American Society of Clinical Oncology·Stefan FröhlingRoss L Levine
Oct 4, 2005·British Journal of Haematology·Alexandra P WolanskyjAyalew Tefferi
Oct 18, 2005·British Journal of Haematology·Ayalew TefferiD Gary Gilliland
Nov 5, 2005·Leukemia·R McClureT Lasho
Dec 24, 2005·Blood·Francesco PassamontiMario Cazzola
Jun 1, 2006·Current Hematologic Malignancy Reports·Martha Wadleigh, D Gary Gilliland
Citations
Jul 13, 2014·Molecular Diagnosis & Therapy·Weiwei LiuMing Guan
Jun 3, 2006·Blood·Hadrian SzpurkaJaroslaw P Maciejewski
Jun 26, 2007·Pathobiology : Journal of Immunopathology, Molecular and Cellular Biology·Kais HusseinHans Kreipe
Jun 26, 2008·Annals of Hematology·Inmaculada RapadoJoaquin Martinez-Lopez
May 19, 2006·American Journal of Clinical Pathology·Timothy C Greiner
Aug 24, 2010·Clinica Chimica Acta; International Journal of Clinical Chemistry·Jun QianZhen Qian
Feb 6, 2007·American Journal of Clinical Pathology·Michael LiewCarl Wittwer
May 19, 2007·American Journal of Clinical Pathology·Angela Y C TanAlexander Dobrovic
Oct 24, 2007·American Journal of Clinical Pathology·Christine FrantzImran Mirza
Sep 19, 2008·Journal of Forensic Sciences·Janice A Nicklas, Eric Buel
Feb 28, 2008·British Journal of Haematology·Graham Standen, Timothy Clench
Dec 9, 2014·Annals of Hematology·G SolerF Ferrer-Marin
Mar 21, 2015·PloS One·Soji MorishitaNorio Komatsu