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Microarrays pp 247-265 | Cite as

Detecting Ligated Fragments on Oligonucleotide Microarrays

Optimizing Chip Design, Array Multiplex Ligation-Dependent Probe Amplification Modification, and Hybridization Parameters
  • Ian R. Berry
  • Carol A. Delaney
  • Graham R. Taylor
Part of the Methods in Molecular Biology™ book series (MIMB, volume 381)

Abstract

Copy-number polymorphisms at specific genomic loci have been implicated in numerous human and animal disease phenotypes. Multiplex ligation-dependent probe amplification (MLPA) is a molecular genetic technique allowing targeted quantification of genomic copy-number changes (deletions and duplications), with potential for multiplexing up to 50 loci in one assay, and resolution down to the single nucleotide level. Modification of the MLPA technique to include Cy-labeled amplification primers permits parallel product detection by capillary electrophoresis and microarray hybridization. Detection and quantification of products by sequencespecific hybridization rather than size-specific capillary electrophoresis increases the potential for probe multiplexing possible in one assay and also allows for more flexible and efficient MLPA probe design. Protocols for the printing of synthetic oligonucleotide probe-sets for the detection of MLPA products, MLPA-probe amplification using array-compatible primers, and parallel product detection by quantitative capillary electrophoresis and microarray hybridization have been optimized.

Key Words

Copy number Cy-labeled PCR MLPA oligonucleotide array printing quantitative capillary electrophoresis 

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Copyright information

© Humana Press Inc., Totowa, NJ 2007

Authors and Affiliations

  • Ian R. Berry
    • 1
  • Carol A. Delaney
    • 1
  • Graham R. Taylor
    • 1
  1. 1.Regional DNA Lab, Regional Genetics ServiceSt. James’s University HospitalLeedsUK

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