González-Giraldo Yeimy, Rodríguez-Dueñas Marisol, Forero Diego A
Department of Nutrition and Biochemistry, School of Sciences, Pontificia Universidad Javeriana, Bogotá, Colombia.
Laboratory of NeuroPsychiatric Genetics, Biomedical Sciences Research Group, School of Medicine, Universidad Antonio Nariño, Bogotá, Colombia.
Mol Biotechnol. 2016 Mar;58(3):197-201. doi: 10.1007/s12033-016-9915-4.
Insertion/Deletion polymorphisms (InDels) are a common type of genetic variation, with a growing role in population genetics and applied genomics. There is the need for the development of novel cost-effective assays for genotyping InDels of high importance. The main objective of this study was to develop high-resolution melting-based assays for genotyping two commonly studied Alu insertion polymorphisms: FXIIIB and PV92 (rs70942849 and rs3138523). Three primers (two forward and one reverse) were designed for each marker, and high-resolution melting (HRM) analyses in a qPCR platform were performed, using EvaGreen fluorescent dye. For each one of the two Alu insertion polymorphisms, HRM analyses identified distinguishable peaks for the three genotypes, allowing a robust genotyping. Results were validated using 96 DNA samples previously genotyped and the assays worked with different DNA concentrations. In this study, we developed novel cost-effective assays, using qPCR, for genotyping two Alu insertion polymorphisms (widely used as ancestry markers). Our results highlight the feasibility of using HRM analyses for genotyping InDel polymorphisms of medical and biotechnological importance.
插入/缺失多态性（InDels）是一种常见的遗传变异类型，在群体遗传学和应用基因组学中发挥着越来越重要的作用。因此，需要开发新的具有成本效益的方法来对具有高度重要性的InDels进行基因分型。本研究的主要目的是开发基于高分辨率熔解的方法，用于对两个常用的Alu插入多态性进行基因分型：FXIIIB和PV92（rs70942849和rs3138523）。针对每个标记设计了三条引物（两条正向引物和一条反向引物），并使用EvaGreen荧光染料在qPCR平台上进行高分辨率熔解（HRM）分析。对于这两个Alu插入多态性中的每一个，HRM分析都能识别出三种基因型的可区分峰，从而实现可靠的基因分型。使用96个先前已进行基因分型的DNA样本对结果进行了验证，并且该方法适用于不同浓度的DNA。在本研究中，我们使用qPCR开发了新的具有成本效益的方法，用于对两个Alu插入多态性（广泛用作祖先标记）进行基因分型。我们的结果突出了使用HRM分析对具有医学和生物技术重要性的InDel多态性进行基因分型的可行性。
