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Nat Protoc. 2008;3(5):849-65. doi: 10.1038/nprot.2008.49.

Array-MAPH: a methodology for the detection of locus copy-number changes in complex genomes.

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1
Department of Cytogenetics, The Cyprus Institute of Neurology & Genetics, Nicosia, Cyprus.

Abstract

High-throughput genome-wide screening methods to detect subtle genomic imbalances are extremely important for diagnostic genetics and genomics. Here, we provide a detailed protocol for a microarray-based technique, applying the principle of multiplex amplifiable probe hybridization (MAPH). Methodology and software have been developed for designing unique PCR-amplifiable sequences (400-600 bp) covering any genomic region of interest. These sequences are amplified, cloned and spotted onto arrays (targets). A mixture of the same sequences (probes) is hybridized to genomic DNA immobilized on a membrane. Bound probes are recovered and quantitatively amplified by PCR, labeled and hybridized to the array. The procedure can be completed in 4-5 working days, excluding microarray preparation. Unlike array-comparative genomic hybridization (array-CGH), test DNA of specifically reduced complexity is hybridized to an array of identical small amplifiable target sequences, resulting in increased hybridization specificity and higher potential for increasing resolution. Array-MAPH can be used for detection of small-scale copy-number changes in complex genomes, leading to genotype-phenotype correlations and the discovery of new genes.

PMID:
18451793
DOI:
10.1038/nprot.2008.49
[Indexed for MEDLINE]
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