Oligonucleotide probe arrays have enabled massively parallel analysis of gene expression levels from a single cDNA sample. Application of microarray technology to analyzing genomic DNA has been stymied by the sequence complexity of the entire human genome. A robust, single base–resolution direct genomic assay would extend the reach of microarray technology. We developed an array-based whole-genome genotyping assay that does not require PCR and enables effectively unlimited multiplexing. The assay achieves a high signal-to-noise ratio by combining specific hybridization of picomolar concentrations of whole genome–amplified DNA to arrayed probes with allele-specific primer extension and signal amplification. As proof of principle, we genotyped several hundred previously characterized SNPs. The conversion rate, call rate and accuracy were comparable to those of high-performance PCR-based genotyping assays.
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The International HapMap Consortium. The International HapMap Project. Nature 426, 789–796 (2003).
Johnson, G.C. et al. Haplotype tagging for the identification of common disease genes. Nat. Genet. 29, 233–237 (2001).
Daly, M.J., Rioux, J.D., Schaffner, S.F., Hudson, T.J. & Lander, E.S. High-resolution haplotype structure in the human genome. Nat. Genet. 29, 229–232 (2001).
Gabriel, S.B. et al. The structure of haplotype blocks in the human genome. Science 296, 2225–2229 (2002).
Judson, R., Salisbury, B., Schneider, J., Windemuth, A. & Stephens, J.C. How many SNPs does a genome–wide haplotype map require? Pharmacogenomics 3, 379–391 (2002).
Stephens, J.C. et al. Haplotype variation and linkage disequilibrium in 313 human genes. Science 293, 489–493 (2001).
Kwok, P.Y. & Chen, X. Detection of single nucleotide polymorphisms. Curr. Issues Mol. Biol. 5, 43–60 (2003).
Wang, D.G. et al. Large-scale identification, mapping, and genotyping of single-nucleotide polymorphisms in the human genome. Science 280, 1077–1082 (1998).
Pinkel, D. et al. High resolution analysis of DNA copy number variation using comparative genomic hybridization to microarrays. Nat. Genet. 20, 207–211 (1998).
Pollack, J.R. et al. Genome–wide analysis of DNA copy-number changes using cDNA microarrays. Nat. Genet. 23, 41–46 (1999).
Carvalho, B., Ouwerkerk, E., Meijer, G.A. & Ylstra, B. High resolution microarray comparative genomic hybridisation analysis using spotted oligonucleotides. J. Clin. Pathol. 57, 644–646 (2004).
Winzeler, E.A. et al. Direct allelic variation scanning of the yeast genome. Science 281, 1194–1197 (1998).
Borevitz, J.O. et al. Large-scale identification of single–feature polymorphisms in complex genomes. Genome Res. 13, 513–523 (2003).
Wu, D.Y., Nozari, G., Schold, M., Conner, B.J. & Wallace, R.B. Direct analysis of single nucleotide variation in human DNA and RNA using in situ dot hybridization. DNA 8, 135–142 (1989).
Storhoff, J.J. et al. Diagnostic detection systems based on gold nanoparticle probes. in Biomedical Applications of Micro- and Nanoengineering Proc. SPIE vol. 4937 (ed. Nicolau, D.V.) 1–7 (SPIE, Bellingham, Washington, 2002).
Rao, K.V. et al. Genotyping single nucleotide polymorphisms directly from genomic DNA by invasive cleavage reaction on microspheres. Nucleic Acids Res. 31, e66 (2003).
Chen, Y., Shortreed, M.R., Peelen, D., Lu, M. & Smith, L.M. Surface amplification of invasive cleavage products. J. Am. Chem. Soc. 126, 3016–3017 (2004).
Lucito, R. et al. Genetic analysis using genomic representations. Proc. Natl. Acad. Sci. USA 95, 4487–4492 (1998).
Kennedy, G.C. et al. Large-scale genotyping of complex DNA. Nat. Biotechnol. 21, 1233–1237 (2003).
Matsuzaki, H. et al. Parallel genotyping of over 10,000 SNPs using a one-primer assay on a high-density oligonucleotide array. Genome Res. 14, 414–425 (2004).
Hardenbol, P. et al. Multiplexed genotyping with sequence-tagged molecular inversion probes. Nat. Biotechnol. 21, 673–678 (2003).
Fan, J.B. et al. Highly parallel SNP genotyping. Cold Spring Harbor Symposia on Quantitative Biology LXVIII, 69–78 (CSHL, Woodbury, New York, 2003).
Dean, F.B. et al. Comprehensive human genome amplification using multiple displacement amplification. Proc. Natl. Acad. Sci. USA 99, 5261–5266 (2002).
Gunderson, K.L. et al. Decoding randomly ordered DNA arrays. Genome Res. 14, 870–877 (2004).
Wang, G. et al. DNA amplification method tolerant to sample degradation. Genome Res. 14, 2357–2366 (2004).
Bobrow, M.N., Harris, T.D., Shaughnessy, K.J. & Litt, G.J. Catalyzed reporter deposition, a novel method of signal amplification. Application to immunoassays. J. Immunol. Methods 125, 279–285 (1989).
Hacker, G.W. High performance Nanogold-silver in situ hybridisation. Eur. J. Histochem. 42, 111–120 (1998).
Shumaker, J.M., Metspalu, A. & Caskey, C.T. Mutation detection by solid phase primer extension. Hum. Mutat. 7, 346–354 (1996).
Gunderson, K.L. et al. Mutation detection by ligation to complete n-mer DNA arrays. Genome Res. 8, 1142–1153 (1998).
Pastinen, T., Kurg, A., Metspalu, A., Peltonen, L. & Syvanen, A.C. Minisequencing: a specific tool for DNA analysis and diagnostics on oligonucleotide arrays. Genome Res. 7, 606–614 (1997).
Pastinen, T. et al. A system for specific, high-throughput genotyping by allele-specific primer extension on microarrays. Genome Res. 10, 1031–1042 (2000).
Erdogan, F., Kirchner, R., Mann, W., Ropers, H.H. & Nuber, U.A. Detection of mitochondrial single nucleotide polymorphisms using a primer elongation reaction on oligonucleotide microarrays. Nucleic Acids Res. 29, E36 (2001).
Consortium, T.I.H. The International HapMap Project. Nature 426, 789–796 (2003).
Simpson, C.L. et al. MaGIC: a program to generate targeted marker sets for genome-wide association studies. Biotechniques 37, 996–999 (2004).
Di, X. et al. Dynamic model-based algorithms for screening and genotyping over 100K SNPs on oligonucleotide microarrays. Bioinformatics advance online publication, 19 January 2005 (10.1093/bioinformatics/bti275).
Pinkel, D., Straume, T. & Gray, J.W. Cytogenetic analysis using quantitative, high-sensitivity, fluorescence hybridization. Proc. Natl. Acad. Sci. USA 83, 2934–2938 (1986).
We thank our Illumina colleagues in manufacturing for providing oligos and arrays and L. Zhou for generating the nonpolymorphic control sequences used in assay development. This work was supported in part by a US National Institutes of Health National Cancer Institute grant to K.L.G.
All authors are or were employees of Illumina, Inc. and may own stock or stock options in the company.
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Gunderson, K., Steemers, F., Lee, G. et al. A genome-wide scalable SNP genotyping assay using microarray technology. Nat Genet 37, 549–554 (2005). https://doi.org/10.1038/ng1547
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