Development of a High-Resolution Melting Curve Analysis Screening Test for SRSF2 Splicing Factor Gene Mutations in Myetodyspiastic Syndromes

作者：Garza Eduardo; Fabiani Emiliano; Noguera Nelida; Panetta Paola; Piredda Maria L; Borgia Loredana; Maurillo Luca; Catalano Gianfranco; Voso Maria T; Lo Coco Francesco^*

来源：Journal of Molecular Diagnostics, 2015, 17(1): 85-89.

DOI：10.1016/j.jmoldx.2014.08.002

Somatic mutations of the spliceosome machinery have been recently identified by whole genome analysis in hematologic diseases, such as myelodysplastic syndrome, chronic Lymphocytic Leukemia, myeloproliferative neoplasms, acute myeloid leukemia, and advanced forms of mastocytosis, and also in nonhematologic conditions. SRSF2 is a member of the serine/arginine-rich family pre-mRNA splicing factors that plays a role in mRNA export from the nucleus and translation. We describe a high-resolution melting (HRM) curve analysis to screen for SRSF2 hotspot mutations in a fast, sensitive, and reliable way. Fifty bone marrow samples from patients with myelodysplastic syndrome were analyzed by the FIRM assay and by direct sequencing. FIRM screening identified four melting patterns corresponding to a negative (wild-type) group and three different mutated groups. Each mutated group was identified according to the positive control used: P95H, P95L, and P95R, respectively. An FIRM mutated pattern was identified in seven patients. Positive and negative results from FIRM were compared with direct sequencing results with a sensitivity and specificity of 100% (95% CI, 0.56-1, and 95% CI, 0.89-1, respectively). Analytical sensitivity analysis revealed a detection threshold of up to 1:9 (mutated/wild type) dilution. This rapid screening method may provide useful information for clinical decision making and be helpful to optimize laboratory resources and reduce turnaround time.

• 出版日期2015-1