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Comparative Genomic Hybridization: DNA Labeling, Hybridization and Detection

  • Richard Redon
  • Tomas Fitzgerald
  • Nigel P. Carter
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 529)

Abstract

Array-CGH involves the comparison of a test to a reference genome using a microarray composed of target sequences with known chromosomal coordinates. The test and reference DNA samples are used as templates to generate two probe DNAs labeled with distinct fluorescent dyes. The two probe DNAs are co-hybridized on a microarray in the presence of Cot-1 DNA to suppress unspecific hybridization of repeat sequences. After slide washes and drying, microarray images are acquired on a laser scanner and fluorescent intensities from every target sequence spot on the array are extracted using dedicated computer programs. Intensity ratios are calculated and normalized to enable data interpretation. Although the protocols explained in this chapter correspond primarily to the use of large-insert clone microarrays in either manual or automated fashion, necessary adaptations for hybridization on microarrays comprising shorter target DNA sequences are also briefly described.

Key words

Probe labeling random priming hybridization detection comparative genomic hybridization array-CGH 

Notes

Acknowledgments

The authors would like to thank Heike Fiegler who developed some of the methods described in this chapter. This work was supported by the Wellcome Trust.

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Richard Redon
    • 1
  • Tomas Fitzgerald
    • 1
  • Nigel P. Carter
    • 1
  1. 1.Wellcome TrustSanger InstituteUK

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