DNA Microarray-Based Mutation Discovery and Genotyping

  • David GreshamEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 772)


DNA microarrays provide an efficient means of identifying single-nucleotide polymorphisms (SNPs) in DNA samples and characterizing their frequencies in individual and mixed samples. We have studied the parameters that determine the sensitivity of DNA probes to SNPs and found that the melting temperature (Tm) of the probe is the primary determinant of probe sensitivity. An isothermal-melting temperature DNA microarray design, in which the Tm of all probes is tightly distributed, can be implemented by varying the length of DNA probes within a single DNA microarray. I describe guidelines for designing isothermal-melting temperature DNA microarrays and protocols for labeling and hybridizing DNA samples to DNA microarrays for SNP discovery, genotyping, and quantitative determination of allele frequencies in mixed samples.

Key words

DNA microarray Single-nucleotide polymorphisms Isothermal melting temperature SNPscanner Bulk segregant mapping 



I thank the labs of David Botstein, Leonid Kruglyak, Maitreya Dunham, and Justin Borevitz where many of these methods were developed. I also thank Bo Curry, Leonardo Brizuela, and Ben Gordon at Agilent Technologies for participation in the initial study of microarray design.


  1. 1.
    Maskos U, Southern, EM (1993) A novel method for the parallel analysis of multiple mutations in multiple samples. Nucleic Acids Res 21:2269–2270Google Scholar
  2. 2.
    Gresham D, Dunham MJ, Botstein D (2008) Comparing whole genomes using DNA microarrays. Nat Rev Genet 9:291–302Google Scholar
  3. 3.
    DeRisi JL, Iyer VR, Brown PO (1997) Exploring the metabolic and genetic control of gene expression on a genomic scale. Science 278:680686Google Scholar
  4. 4.
    Lieb JD, Liu X, Botstein D et al (2001) Promoter-specific binding of Rap1 revealed by genome-wide maps of protein-DNA association. Nat Genet 28:327–334Google Scholar
  5. 5.
    Pollack JR, Perou CM, Alizadeh AA, et al (1999) Genome-wide analysis of DNA copy-number changes using cDNA microarrays. Nat Genet 23:41–46Google Scholar
  6. 6.
    Pinkel D, Segraves R, Sudar D, et al (1998) High resolution analysis of DNA copy number variation using comparative genomic hybridization to microarrays. Nat Genet 20:207–211Google Scholar
  7. 7.
    Gresham D, Ruderfer DM, Pratt SC, et al (2006) Genome-wide detection of polymorphisms at nucleotide resolution with a single DNA microarray. Science 311:1932–1936Google Scholar
  8. 8.
    Ronald J, Akey JM, Whittle J, et al (2005) Simultaneous genotyping, gene-expression measurement, and detection of allele-specific expression with oligonucleotide arrays. Genome Res 15:284–291Google Scholar
  9. 9.
    Gresham D, Curry B, Ward A, et al (2010) Optimized detection of sequence variation in heterozygous genomes using DNA microarrays with isothermal-melting probes. Proc Natl Acad Sci USA 107:1482–1487Google Scholar
  10. 10.
    Ehrenreich IM, Torabi N, Jia Y, et al (2010) Dissection of genetically complex traits with extremely large pools of yeast segregants. Nature 464:1039–1042Google Scholar
  11. 11.
    Winzeler EA, Richards DR, Conway AR, et al (1998) Direct allelic variation scanning of the yeast genome. Science 281:1194–1197.Google Scholar
  12. 12.
    Borevitz JO, Liang D, Plouffe D, et al (2003) Large-scale identification of single-feature polymorphisms in complex genomes. Genome Res 13:513–523Google Scholar
  13. 13.
    Cutler DJ, Zwick ME, Carrasquillo MM, et al (2001) High-throughput variation detection and genotyping using microarrays. Genome Research 11:1913–1925Google Scholar
  14. 14.
    Liu WM, Di X, Yang G, et al (2003) Algorithms for large-scale genotyping microarrays, Bioinformatics 19:23972403Google Scholar
  15. 15.
    SantaLucia J (1998) A unified view of polymer, dumbbell, and oligonucleotide DNA nearest-neighbor thermodynamics. Proc Natl Acad Sci USA 95:1460–1465Google Scholar
  16. 16.
    SantaLucia J, Allawi HT, Seneviratne PA (1996) Improved nearest-neighbor parameters for predicting DNA duplex stability. Biochemistry 35:3555–3562Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Department of Biology and Center for Genomics and Systems BiologyNew York UniversityNew YorkUSA

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