Detection of JAK2 mutations in paraffin marrow biopsies by high resolution melting analysis: identification of L611S alone and in cis with V617F in polycythemia vera.
JAK2 mutations provide an objective major criterion for diagnosis of myeloproliferative neoplasms in the 2008 World Health Organization classification, and are particularly useful for cases of polycythemia vera (PV). High resolution melting analysis (HRM) using DNA from fresh samples for mutation detection in JAK2 exons 12 and 14 has been reported. Here, we describe two new HRM techniques that use both fresh and formalin-fixed paraffin-embedded marrow samples, allowing for both prospective and retrospective analyses. We used these novel assays to screen DNA isolated from paraffin-embedded marrow biopsies from 104 patients with PV for mutations. HRM mutation-positive samples were subsequently sequenced. Ninety-seven samples (93.3%) were positive solely for the JAK2 p.V617F. One harbored a 33 bp duplication in exon 12 along with an exon 14 p.V617F mutation, another had an exon 12 p.H538_K539delinsL, and a third carried a previously unreported mutation in PV, p.L611S, alone, and in cis with p.V617F. This indicates that these assays can detect known and novel JAK2 mutations in exons 12 and 14 using either fresh or paraffin-embedded archival materials.
ARUP Laboratories and Department of Pathology, University of Utah, Salt Lake City, UT 84108, USA. roberto.nussenzveig@aruplab.com
Roberto H
RH
Nussenzveig
Tatiana
T
Burjanivova
Mohamed E
ME
Salama
Natalie W
NW
Ogilvie
Juraj
J
Marcinek
Lukas
L
Plank
Archana M
AM
Agarwal
Sherrie L
SL
Perkins
Josef T
JT
Prchal
D019943:Amino Acid Substitution; D001483:Base Sequence; D001706:Biopsy; D001853:Bone Marrow / Q000378:metabolism* / Q000473:pathology; D004252:DNA Mutational Analysis / Q000379:methods*; D005091:Exons / Q000235:genetics; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D053614:Janus Kinase 2 / Q000235:genetics*; D008867:Microtomy; D008969:Molecular Sequence Data; D009154:Mutation; D009691:Nucleic Acid Denaturation; D016612:Paraffin Embedding; D011087:Polycythemia Vera / Q000097:blood / Q000235:genetics* / Q000473:pathology; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results
C507924:JAK2 protein, human; D053614:Janus Kinase 2