1. Optimization of high-resolution melting analysis for simultaneous genotyping of two 11β-hydroxysteroid dehydrogenase type 1 gene polymorphisms
- Author
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Barbara Mlinar, Barbara Ostanek, Adlija Causevic, Tanja Dujic, and Janja Marc
- Subjects
Genetics ,Genotype ,Gene Expression Profiling ,General Medicine ,Biology ,Polycystic ovary ,Polymorphism, Single Nucleotide ,High Resolution Melt ,law.invention ,chemistry.chemical_compound ,chemistry ,law ,11β-hydroxysteroid dehydrogenase type 1 ,11-beta-Hydroxysteroid Dehydrogenase Type 1 ,biology.protein ,Humans ,Female ,Restriction fragment length polymorphism ,Genotyping ,Gene ,Genetics (clinical) ,DNA ,Polymerase chain reaction ,Polycystic Ovary Syndrome - Abstract
Background: Polymorphisms in HSD11B1, the gene encoding 11β-hydroxysteroid dehydrogenase type 1 enzyme, have been associated with obesity, metabolic syndrome, and type 2 diabetes. In this study, we present an optimized high-resolution melting (HRM) method for genotyping two common polymorphisms of the HSD11B1 gene: rs846910: G>A and rs45487298: insA. Methods: One hundred DNA samples from patients with polycystic ovary syndrome and healthy controls were genotyped by HRM. The results were compared with those obtained with classic polymerase chain reaction followed by restriction fragment length polymorphism analysis. Results: Various approaches were used during HRM specificity optimization. With the optimized method, genotyping accuracy of 100% was achieved. Conclusions: HRM analysis is a fast, simple, and cost-effective method compared with the alternative genotyping approaches. The work required for optimizing the method (improvement of specificity) is minor compared to the advantages.
- Published
- 2010