Performance of amplified DNA in an Illumina GoldenGate BeadArray assay.
Performance of amplified DNA in an Illumina GoldenGate BeadArray assay.
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DOI：
10.1158/1055-9965.epi-07-2849
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发表时间：
2008-07
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影响因子：
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通讯作者：
Goode EL
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文献类型：
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作者：
Cunningham JM;Sellers TA;Schildkraut JM;Fredericksen ZS;Vierkant RA;Kelemen LE;Gadre M;Phelan CM;Huang Y;Meyer JG;Pankratz VS;Goode EL
全基因组扩增（WGA）为流行病学研究提供了一种富集DNA的方法。我们使用1，536个单核苷酸多态性（SNP）和2，368个样本的卵巢癌研究来评估使用Illumina GoldenGate BeadArray的多重置换扩增（MDA）WGA的性能。初步筛选显示成功的基因分型的WGA样品的93.4%和99.3%的基因组样品，和93.2%的SNP的WGA样品和96.3%的SNP的基因组样品。SNP失败通过Illumina提供的可设计性等级、%GC（P ≤ 0.002）以及仅WGA、与端粒的距离和Illumina提供的SNP评分（P ≤ 0.002）来预测。端粒的距离和%GC高度相关; %GC的调整消除了端粒的距离和SNP失败之间的关联。虽然普遍较高，但每个SNP的调用率与可设计性等级、SNP评分、%GC、次要等位基因频率、与端粒的距离（P ≤ 0.01）以及仅对于WGA，Illumina提供的验证类别（P < 0.001）相关。我们发现在124个WGA：基因组重复、15个WGA重复、相同WGA制备物的88个重复等分试样和25个基因组重复中通常具有优异的一致性（>99.0%）。在存在不一致的情况下，其是跨WGA的：基因组重复，但仅限于其他重复中的少数样品，表明引入了误差。可设计性等级和SNP得分与WGA：基因组一致性相关（P < 0.001）。总之，使用MDA WGA DNA是可行的;然而，对于SNP选择和分析需要谨慎。我们建议在使用MDA WGA DNA创建GoldenGate检测时考虑生物SNP特征，特别是与端粒的距离和GC含量（推荐<50% GC），以及Illumina提供的指标。
Whole genome amplification (WGA) offers a means to enrich DNA quantities for epidemiologic studies. We used an ovarian cancer study of 1,536 single nucleotide polymorphisms (SNPs) and 2,368 samples to assess performance of multiple displacement amplification (MDA) WGA using an Illumina GoldenGate BeadArray. Initial screening revealed successful genotyping for 93.4% of WGA samples and 99.3% of genomic samples, and 93.2% of SNPs for WGA samples and 96.3% of SNPs for genomic samples. SNP failure was predicted by Illumina-provided designability rank, %GC (P ≤ 0.002), and for WGA only, distance to telomere and Illumina-provided SNP score (P ≤ 0.002). Distance to telomere and %GC were highly correlated; adjustment for %GC removed the association between distance to telomere and SNP failure. Although universally high, per-SNP call rates were related to designability rank, SNP score, %GC, minor allele frequency, distance to telomere (P ≤ 0.01), and, for WGA only, Illumina-provided validation class (P < 0.001). We found excellent concordance generally (>99.0%) among 124 WGA:genomic replicates, 15 WGA replicates, 88 replicate aliquots of the same WGA preparation, and 25 genomic replicates. Where there was discordance, it was across WGA:genomic replicates but limited to only a few samples among other replicates suggesting the introduction of error. Designability rank and SNP score correlated with WGA:genomic concordance (P < 0.001). In summary, use of MDA WGA DNA is feasible; however, caution is warranted regarding SNP selection and analysis. We recommend that biological SNP characteristics, notably distance to telomere and GC content (<50% GC recommended), as well as Illumina-provided metrics be considered in the creation of GoldenGate assays using MDA WGA DNA.