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Abstract Background Myeloproliferative neoplasms (MPN) are heterogeneous diseases that classified by the presence of Philadelphia chromosome into Philadelphia chromosome negative (Ph-neg) and positive (Ph-pos) myeloproliferative neoplasms. In ph-neg group A somatic point mutation (c.1849G>T) in the JAK2 gene, part of the JAK2-STAT signal-transduction pathway, causes substitution of phenylalanine for valine (V617F) in the JAK2 protein and has been identified. This mutation was seen in PV by 65% to 97% and ET (30–57%) and primary myelofibrosis (35–95%). Highly sensitive methods have been used to determine the presence of the JAK2V617F mutation instead of direct sequencing. We aimed to assess JAK2 exon14 mutations by high-resolution melting (HRM) analysis, which allows variation screening in compare to other method for detecting mutation. Methods The mutation analysis included 45 individuals who were subjected for diagnosis of ph-neg MPN. Genomic DNA was isolated and different methods are performed. Results PCR RFLP, ARMS PCR and HRM method has a detection sensitivity comparable with conventional methods (Qiagen) to identify the mutations and sequencing. Conclusions For HRM analysis is cost-effective and beside that it is enzyme independence method also this method able to show amount of the mutant allele carried in samples and it’s helpful for treatments follow-up and determining MRD for them.