Duffy Kelly J, Littrell Jack, Locke Adam, Sherman Stephanie L, Olivier Michael
Human and Molecular Genetics Center, Medical College of Wisconsin, Milwaukee, WI, USA.
Nucleic Acids Res. 2008 Dec;36(22):e145. doi: 10.1093/nar/gkn736. Epub 2008 Oct 21.
Individuals with trisomy 21 display complex phenotypes with differing degrees of severity. Numerous reliable methods have been established to diagnose the initial trisomy in these patients, but the identification and characterization of the genetic basis of the phenotypic variation in individuals with trisomy remains challenging. To date, methods that can accurately determine genotypes in trisomic DNA samples are expensive, require specialized equipment and complicated analyses. Here we report proof-of-concept results for an Invader(R) assay-based genotyping procedure that can determine SNP genotypes in trisomic genomic DNA samples in a simple and cost-effective manner. The procedure requires only two experimental steps: a real-time measurement of the fluorescent Invader(R) signal and analysis with a specifically designed clustering algorithm. The approach was tested using genomic DNA samples from 23 individuals with trisomy 21, and results were compared to genotypes previously determined with pyrosequencing. Additional assays for 15 SNPs were tested in a set of 21 DNA samples to assess assay performance. Our method successfully identified the correct SNP genotypes for the trisomic genomic DNA samples tested, and thus provides an alternative to determine SNP genotypes in trisomic DNA samples for subsequent association studies in patients with Down syndrome and other trisomies.
21三体综合征患者表现出严重程度不同的复杂表型。已经建立了许多可靠的方法来诊断这些患者最初的三体情况，但是确定三体患者表型变异的遗传基础并对其进行特征描述仍然具有挑战性。迄今为止，能够准确确定三体DNA样本中基因型的方法成本高昂，需要专门的设备和复杂的分析。在此，我们报告了基于侵入法（Invader®）的基因分型程序的概念验证结果，该程序能够以简单且经济高效的方式确定三体基因组DNA样本中的单核苷酸多态性（SNP）基因型。该程序仅需两个实验步骤：实时测量荧光侵入信号，并使用专门设计的聚类算法进行分析。使用来自23名21三体综合征患者的基因组DNA样本对该方法进行了测试，并将结果与先前通过焦磷酸测序确定的基因型进行了比较。在一组由21个DNA样本组成的样本集中对另外15个SNP进行了检测，以评估检测性能。我们的方法成功地为所测试的三体基因组DNA样本鉴定出了正确的SNP基因型，因此为在唐氏综合征和其他三体患者中进行后续关联研究确定三体DNA样本中的SNP基因型提供了一种替代方法。
