Detection of JAK2 Exon 12 Mutations in JAK2 V617F-Negative Polycythemia Vera Patients by Cloning Technique

Introduction: The identification of mutations of the JAK2 gene is a useful marker in the diagnosis of polycythemia vera (PV) patients. We studied the frequency of JAK2 mutations in a group of PV patients because data are still very limited regarding this subject in Polish patients.

Methods: The JAK2 V617F mutation was examined using the amplification refractory mutation system (ARMS)-PCR method. Direct sequencing and a cloning technique were performed to determine alternations in exon 12 of the JAK2 gene.

Results: A group of 90 consecutive patients with a suspected diagnosis of polycythemia vera were investigated. In 91% of the cases, the JAK2 V617F mutation was identified. The remaining JAK2 V617F-negative patients were subjected to examination for JAK2 exon 12 by direct PCR product sequencing and the cloning technique. The following mutations were identified: H538-K539delinsL, E543-D544del and N542-E543del. These exon 12 mutants constituted 50% of PV JAK2 V617F-negative group and 4.4% (out of 90) of all PV patients (JAK2 V617F-positive and JAK2 V617F-negative).

Conclusion: Our results demonstrate the prevalence of JAK2 mutations (V617F and in exon 12) in PV cases. Moreover, the data show that direct sequencing is not an adequate technique for exon 12 mutation identification; therefore, appropriate methodology should be considered for using this molecular marker in the process of diagnosis.