Development of Molecular Beacon method to detect of JAK2 V617F mutation

Ali Nazemi--- Molecular Genetics part, Department of Biology, Islamic Azad University, Tonekabon branch, Mazandaran, Iran assistant professor

Abstract


40

V617F mutation of JAK2 gene is a point mutation of Somatic cells leading to permanent phosphorylation and protein kinase activity of the JAK2 protein. This mutation is a valuable marker in the diagnosis of myeloid neoplasms. Development of new techniques can be effective and highly sensitive for the detection of the mutation. The aim of this study is designing and executing a molecular beacon probe-based real-time PCR detect to V617F mutation. Test results were reviewed from a set of wild type and cloned JAK2 exon 12 mutated allele genes into plasmid vector and through amplification by Real-Time PCR system. The sensitivity and specificity of Molecular Beacon probes design were tested. The results showed that the Real-Time PCR system has %100 specificity and % 0.001 sensitivity. Based on the results obtained from the molecular beacon probe-based real-time PCR, it was proved that the system is able to detect normal and mutant alleles of JAK2 V617F position with high accuracy and a short time in a closed tube system


Keywords


Molecular Beacon Probe; JAK2V617F Mutation; Neoplasms

Full Text:

PDF

32

References


Cankovic M, Whiteley L, Hawley RC. Clinical performance of JAK2 V617F mutation detection assays in a molecular diagnostics laboratory. Am J Clin Pathol, 2009; 132:713-721.

Zhao AH, Gao R. Development of a highly sensitive method for detection of JAK2V617F. Journal of Hematology & Oncology.2011, 4:40.

Huijsmans CJ, Poodt J, Savelkoal PH. Sensitive detection and quantification of the JAK2V617F allele by Real-Time PCR. Journal of Molecular Diagnostics.2011; 13, 5.

Frantz C, Sekora DM, Henley DC, et al. Comparative evaluation of three JAK2V617F mutation detection methods.Am J Clin Pathol.2007;128:865-874.

Chen Q, Lu P, Jones AV, et al. Amplification refractory mutation system, a highly sensitive and simple polymerase chain reaction assay, for the detection of JAK2 V617F mutation in chronic myeloproliferative disorders. J Mol Diagn. 2007; 9:272-276.

Murugesan G, Aboudola S, Szpurka H, et al. Identification of the JAK2 V617F mutation in chronic myeloproliferative disorders using FRET probes and melting curve analysis. Am J Clin Pathol. 2006; 125:625-633.

Vannuchi AM, Antonioli E, Guglielmelli P, et al. Prospective identification of high-risk polycythemia vera patients based on JAK2(V617F) allele burden. Leukemia. 2007; 21:1952-1959.

Tefferi A, Vardiman JW. Classification and diagnosis of myeloproliferative neoplasms: the 2008 World Health Organization criteria and point-of-care diagnostic algorithms. Leukemia. 2008; 22:14-22.

Jeffery M. Klco, Ravi Vij, Friederike H.K, Anjum Hassan, John L.F. Molecular Pathology of Myeloproliferative Neoplasms. Am J Clin Pathol 2010;133:602-615.

Kolpashchikov DM. An Elegant Biosensor Molecular Beacon Probe: Challenges and Recent Solutions. Scientifica ,2012, p17.

Zapparoli G.V, Jorissen R.N, Hewitt C.A, McBean M, Westerman D.A, Dobrovic A,. Quantitative threefold allele-specific PCR(QuanTAS-PCR) for highly sensitive JAK2 V617Fmutant allele detection. BMC Cancer 2013, 13:206.

Zhao A.H, Gao R, Zhao Z.J. Development of a Highly Sensitive Method for Detection of JAK2V617F. Journal of Hematology & Oncology 2011, 4:40.

Wu ZH, Yuan H, Zhang X, Liu W, Xu J, Zhang W, Guan M. Development and Inter-Laboratory Validation of Unlabeled Probe Melting Curve Analysis for Detection of JAK2 V617F Mutation in Polycythemia Vera. PLoS ONE 2011,6(10).

Wu Z, Zhang Y, Zhang X, Xu X, Kang ZH, Li SH, Zhang CH, Su B, Guan M. A Multiplex Snapback Primer System for the Enrichment and Detection of JAK2 V617F and MPL W515L/K Mutations in Philadelphia-Negative Myeloproliferative Neoplasms. BioMed Research International.Volume 2014.




DOI: http://dx.doi.org/10.22037/jps.v9i2.19323

Article Metrics

Metrics Loading ...

Metrics powered by PLOS ALM

Refbacks

  • There are currently no refbacks.


"Journal of Paramdedical Sciences", is a publication of "School of Paramedical Sciences, Shahid Beheshti University of Medical Sciences" and "Iranian Society of Medical Proteomics".

"Journal of Paramdedical Sciences" is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

 

EISSN: 2008-4978

PISSN: 2008-496X