Efficient approach to unique single-nucleotide polymorphism discovery.
Efficient approach to unique single-nucleotide polymorphism discovery.
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DOI：
10.1101/gr.9.5.499
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发表时间：
1999-05
期刊：
影响因子：
7
通讯作者：
P. Taillon-Miller;E. E. Piernot-E.;P. Kwok
中科院分区：
文献类型：
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作者：
P. Taillon-Miller;E. E. Piernot-E.;P. Kwok
单核苷酸多态性（SNP）是人类基因组中最常见的DNA序列变异。有人认为，一组密集的SNP标记可用于鉴定与复杂疾病性状相关的遗传因素。因为所有的高通量基因分型方法都需要SNP的精确序列知识，所以任何SNP发现方法都必须涉及DNA序列和等位基因频率的测定。此外，高通量基因分型还需要基因组DNA扩增步骤，使得有必要开发仅扩增含有SNP的DNA片段而不扩增基因组其余部分的序列标记位点（STS）。在这份报告中，我们展示了一个SNP筛选方法，产生的DNA序列和等位基因频率的信息，同时筛选出重复以最小的成本和努力的效用。我们的方法是基于使用纯合子完全性葡萄胎（CHM）作为参考。有了这个纯合参考，人们可以用合并的DNA测序方法鉴定和估计常见SNP的等位基因频率（而不是像通常那样必须对许多个体进行测序）。更重要的是，CHM参考比单个单个参考更可取，因为在GenBank中发现重复序列之前，它容易揭示通过PCR测定扩增的基因组的任何重复区域。这种方法将SNP发现的成本降低了60%，并消除了不能从基因组中唯一扩增的SNP标记的昂贵开发。
Single-nucleotide polymorphisms (SNPs) are the most frequently found DNA sequence variations in the human genome. It has been argued that a dense set of SNP markers can be used to identify genetic factors associated with complex disease traits. Because all high-throughput genotyping methods require precise sequence knowledge of the SNPs, any SNP discovery approach must involve both the determination of DNA sequence and allele frequencies. Furthermore, high-throughput genotyping also requires a genomic DNA amplification step, making it necessary to develop sequence-tagged sites (STSs) that amplify only the DNA fragment containing the SNP and nothing else from the rest of the genome. In this report, we demonstrate the utility of a SNP-screening approach that yields the DNA sequence and allele frequency information while screening out duplications with minimal cost and effort. Our approach is based on the use of a homozygous complete hydatidiform mole (CHM) as the reference. With this homozygous reference, one can identify and estimate the allele frequencies of common SNPs with a pooled DNA-sequencing approach (rather than having to sequence numerous individuals as is commonly done). More importantly, the CHM reference is preferable to a single individual reference because it reveals readily any duplicated regions of the genome amplified by the PCR assay before the duplicated sequences are found in GenBank. This approach reduces the cost of SNP discovery by 60% and eliminates the costly development of SNP markers that cannot be amplified uniquely from the genome.