Flexible Use of High-Density Oligonucleotide Arrays for Single-Nucleotide Polymorphism Discovery and Validation
Abstract
A method for identifying and validating single nucleotide polymorphisms (SNPs) with high-density oligonucleotide arrays without the need for locus-specific polymerase chain reactions (PCR) is described in this report. Genomic DNAs were divided into subsets with complexity of ∼10 Mb by restriction enzyme digestion and gel-based fragment size resolution, ligated to a common adaptor, and amplified with one primer in a single PCR reaction. As a demonstration of this approach, a total of 124 SNPs were located in 190 kb of genomic sequences distributed across the entire human genome by hybridizing to high-density variant detection arrays (VDA). A set of independent validation experiments was conducted for these SNPs employing bead-based affinity selection followed by hybridization of the affinity-selected SNP-containing fragments to the same VDA that was used to identify the SNPs. A total of 98.7% (74/75) of these SNPs were confirmed using both DNA dideoxynucleotide sequencing and the VDA methodologies. With flexible sample preparation, high-density oligonucleotide arrays can be tailored for even larger scale genome-wide SNP discovery as well as validation.
Footnotes
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↵1 Corresponding author.
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E-MAIL Shoulian_Dong{at}affymetrix.com; FAX (408) 481–0422.
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Article published on-line before print: Genome Res.,10.1101/gr. 171101.
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Article and publication are at http://www.genome.org/cgi/doi/10.1101/gr.171101.
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- Received November 30, 2000.
- Accepted May 14, 2001.
- Cold Spring Harbor Laboratory Press