Cahill Patrick, Bakis Michele, Hurley James, Kamath Veena, Nielsen William, Weymouth Dina, Dupuis Josee, Doucette-Stamm Lynn, Smith Douglas R
Genome Therapeutics Corporation, Waltham, Massachusetts 02453, USA.
Genome Res. 2003 May;13(5):925-31. doi: 10.1101/gr.939903. Epub 2003 Apr 14.
We report the validation of a new assay for typing single nucleotide polymorphisms (SNPs) that takes advantage of the 3'-to-5' exonuclease proofreading activity of many DNA polymerases. The assay uses one or more primers labeled on the 3' nucleotide base, and can be implemented in a variety of formats including a one-step PCR reaction that allows SNP typing directly from genomic DNA samples. The detection of genotypes can be accomplished by means of fluorescence detection on assays that have been purified to remove excess primer, or by means of fluorescence polarization without any additional cleanup. We also demonstrate that the Exo-Proofreading SNP assay can be used on pooled samples to obtain allele frequency data.
我们报告了一种用于单核苷酸多态性(SNP)分型的新检测方法的验证,该方法利用了许多DNA聚合酶的3'至5'核酸外切酶校对活性。该检测方法使用一个或多个在3'核苷酸碱基上标记的引物,并且可以以多种形式实施,包括一步PCR反应,该反应允许直接从基因组DNA样本中进行SNP分型。基因型的检测可以通过对已纯化以去除过量引物的检测进行荧光检测来完成,或者通过荧光偏振而无需任何额外的纯化来完成。我们还证明了外切校对SNP检测方法可用于混合样本以获得等位基因频率数据。