Validation of the use of whole-genome amplified DNA in a population-based study

描述(由申请人提供)：摘要全基因组扩增(WGA)技术提供了从原本枯竭的天然DNA中扩增DNA的机会。这项技术有可能促进分子流行病学研究的持续生产力，这些研究限制了剩余DNA。尽管可能会出现等位基因扩增错误，但WGA在SNP基因分型中的表现通常报告了非常好的一致性和重复性。然而，从口腔刷子中获得的WGA DNA的质量尚未被彻底评估以用于突变筛查或测序，已发表的报告大多涉及从其他来源提取的有限组DNA样本，并测试基于非PCR的WGA方法的保真度。这项验证性研究的目的是确定通过直接测序从基于群体的研究中检测大量WGA DNA样本中胚系突变和多态的敏感性和特异性。我们计划对3580个口腔刷子的DNA样本进行CDKN2A/p16基因测序，这些样本已经通过GenomePlex(R)试剂盒进行了全基因组扩增，然后将测序数据与已经在原始或天然DNA中获得的结果进行了比较。我们努力证明，我们可以使用WGA随机片段-聚合酶链式反应方法，允许使用该资源对生殖系DNA进行几乎无限的未来研究，以调查与黑色素瘤风险和/或进展有关的假设。这项研究还具有更广泛的科学意义，因为它将为今后的流行病学调查提供关于口腔样本用途的概括性知识。

DESCRIPTION (provided by applicant): Abstract Whole-genome amplification (WGA) technologies offer the opportunity to expand DNA from otherwise depleted native DNAs. This technique has the potential to facilitate the continued productivity of molecular epidemiological studies that have limited remaining DNA. Although allele amplification errors can occur, the performance of WGA in SNP genotyping has generally reported very good concordance and reproducibility. However the quality of WGA DNA obtained from buccal brushes has not been thoroughly evaluated for screening of mutations or sequencing, and published reports have mostly involved limited sets of DNA samples extracted from other sources and tested for the fidelity of non- PCR based WGA methods. The objective of this validation study is to determine the sensitivity and specificity of detecting germline mutations and polymorphisms in a large set of WGA DNA samples from a population-based study by direct sequencing. We plan to sequence the CDKN2A/p16 gene in 3580 DNA samples from buccal brushes that has been whole-genome amplified with the GenomePlex(R) kit, and then compare the sequencing data with those results already obtained in the original or native DNAs. We endeavor to demonstrate that we can use the WGA random fragmentation- PCR method to allow nearly unlimited future studies of germline DNA using this resource to investigate hypotheses in relation to melanoma risk and/or progression. The study also has a broader scientific relevance in that it will provide generalizable knowledge about the utility of buccal samples for future epidemiological investigations.