The Effect of DNA Concentration on the HRM Performance in Detecting Jak2 p.V617F Variant in Patients with Myeloproliferative Neoplasms

Abstract

Introduction: Janus kinase 2 (JAK2) is an intracellular signaling protein. JAK2 p. V617F is a common variant in normal karyotype myeloproliferative neoplasms (MPN). Highresolution melting (HRM) analysis is one of the essential methods for detecting the JAK2 p.V617F variant. In this study, we have investigated the effect of DNA concentration on detecting the JAK2 p.V617F variant using the HRM method. Methods: Genomic DNA was extracted from human blood and diluted ten times in distilled water from 1 to 0.03; afterward, HRM was conducted for each dilution (triplicate). Using SPSS v.20.0 software, the mean Tm of each dilution was calculated and compared. Results: The HRM results revealed the JAK2 wild type and variant to have Tms of 81/64°C and 80/76°C, respectively. At the endpoint of the pre-amplification, the dilutions had different emissions. The statistical analysis revealed no statistically significant differences in Tm between samples with varying DNA concentrations (P value > 0.05). Conclusion: There have been no significant differences obtained in the analysis of JAK2 p.V617F point variant in different DNA dilutions, implying that the HRM analysis has no relation to DNA concentration.