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全基因组单核苷酸多态性阵列显示基于多重置换的全基因组扩增具有高保真度。

Genome-wide single-nucleotide polymorphism arrays demonstrate high fidelity of multiple displacement-based whole-genome amplification.

作者信息

Tzvetkov Mladen V, Becker Christian, Kulle Bettina, Nürnberg Peter, Brockmöller Jürgen, Wojnowski Leszek

机构信息

Department of Clinical Pharmacology, Georg-August-University Göttingen, Göttingen, Germany.

出版信息

Electrophoresis. 2005 Feb;26(3):710-5. doi: 10.1002/elps.200410121.

Abstract

Whole-genome DNA amplification by multiple displacement (MD-WGA) is a promising tool to obtain sufficient DNA amounts from samples of limited quantity. Using Affymetrix' GeneChip Human Mapping 10K Arrays, we investigated the accuracy and allele amplification bias in DNA samples subjected to MD-WGA. We observed an excellent concordance (99.95%) between single-nucleotide polymorphisms (SNPs) called both in the nonamplified and the corresponding amplified DNA. This concordance was only 0.01% lower than the intra-assay reproducibility of the genotyping technique used. However, MD-WGA failed to amplify an estimated 7% of polymorphic loci. Due to the algorithm used to call genotypes, this was detected only for heterozygous loci. We achieved a 4.3-fold reduction of noncalled SNPs by combining the results from two independent MD-WGA reactions. This indicated that inter-reaction variations rather than specific chromosomal loci reduced the efficiency of MD-WGA. Consistently, we detected no regions of reduced amplification, with the exception of several SNPs located near chromosomal ends. Altogether, despite a substantial loss of polymorphic sites, MD-WGA appears to be the current method of choice to amplify genomic DNA for array-based SNP analyses. The number of nonamplified loci can be substantially reduced by amplifying each DNA sample in duplicate.

摘要

通过多重置换进行全基因组DNA扩增(MD-WGA)是一种很有前景的工具,可从少量样本中获取足够量的DNA。我们使用Affymetrix公司的基因芯片人类映射10K阵列,研究了经MD-WGA处理的DNA样本中的准确性和等位基因扩增偏差。我们观察到在未扩增的DNA和相应扩增后的DNA中检测到的单核苷酸多态性(SNP)之间具有极佳的一致性(99.95%)。这种一致性仅比所用基因分型技术的实验内重复性低0.01%。然而,MD-WGA未能扩增估计7%的多态性位点。由于用于判定基因型的算法,这仅在杂合位点中被检测到。通过合并两个独立的MD-WGA反应结果,我们使未检测到的SNP数量减少了4.3倍。这表明反应间的差异而非特定的染色体位点降低了MD-WGA的效率。一致地,除了位于染色体末端附近的几个SNP外,我们未检测到扩增减少的区域。总体而言,尽管多态性位点大量丢失,但MD-WGA似乎是目前用于基于阵列的SNP分析扩增基因组DNA的首选方法。通过对每个DNA样本进行重复扩增,未扩增位点的数量可大幅减少。

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