Masters Degrees (Genetics)
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Browsing Masters Degrees (Genetics) by Author "Du Plessis, Hanri"
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Item Open Access Mutational analysis of the TET2 gene in Philadelphia negative myeloproliferative neoplasms(University of the Free State, 2014-07) Du Plessis, Hanri; De Kock, A.; Viljoen, C. D.English: Myeloproliferative neoplasms (MPNs) are clonal haematopoietic stem cell disorders which are characterized by the excessive proliferation of one or more of the myeloid cell lineages. Mutations in several genes have been found to contribute to the pathogenesis of the Philadelphia (Ph) -negative MPNs. These MPNs include polycythaemia vera (PV), essential thrombocythaemia (ET) and primary myelofibrosis (PMF). The most common genes associated with Ph-negative MPNs are the JAK2 and MPL genes, which also form an integral part of the 2008 World Health Organization (WHO) diagnostic criteria for PV, ET and PMF. Mutations in the JAK2 and MPL genes are, however, not always detected in PV, ET and PMF patients. The ten-eleven-translocation (TET) 2 tumor suppressor gene has been identified to be a potential contributor to the normal development of haematopoietic cells, particularly the myeloid blood cells. While the exact role of TET2 in haematopoiesis is not yet completely understood, mutations in the TET2 gene have recently been found to occur in PV, ET and PMF patients. It has been suggested that mutations occurring in the conserved domains of TET2 could interfere with the catalytic activity of the protein, which ultimately has the potential to result in an inactive tumor suppressor gene. TET2 mutations situated outside of the conserved domains also have the potential to alter protein function. Defective expression of TET2 has previously been shown to alter haematopoietic development, resulting in the increased production of various myeloid cells. Previous studies in which mice models were used illustrated that the loss of TET2 function leads to characteristics similar to that of MPN patients. The aim of our study was to screen 25 PV, ET and PMF patients for mutations in the TET2 gene using high resolution melting (HRM) analysis and DNA sequencing analysis. In this study only two of the 24 primer sets could successfully be used to detect mutations using HRM analysis. The presence of non-specific amplification products, primer dimer, multiple melting domains and high melting background were suspected to be the causes of the unsuccessful HRM analysis. The DNA sequencing analysis of the TET2 gene was, however, successful. No novel variants were found in the current study. A total of 24 previously published TET2 variants were detected in the 25 Ph-negative MPN patients. The TET2 variants included missense single nucleotide polymorphisms (SNPs), synonymous SNPs, intronic SNPs, intronic deletions and intronic insertions. Unlike in previous studies, TET2 variants appeared to be more common in JAK2 V617F-negative MPN patients than in JAK2 V617F-positive MPN patients. It is speculated that most of the TET2 variants detected in the current study occur naturally in the TET2 gene and not exclusively in MPN patients. The prognostic value of TET2 variants in MPNs is as yet unknown. Since most of the SNPs in the current study appeared to be natural variants of the TET2 gene, it is possible that TET2 does not play a direct role in the development of MPNs. However, limited literature regarding the function of TET2 and how it contributes to abnormal haematopoiesis is available. Further investigation is therefore needed to establish the exact role of TET2 in the pathogenesis of MPNs and whether variants of this gene could predispose individuals to the development of MPNs.