The Affymetrix GeneChip® Human Mapping
10K Array is a Single Nucleotide Polymorphism (SNP) genotyping tool based
on allele-specific hybridization. By synthesizing probes on the array corresponding
to both of the two possible alleles at each SNP and hybridizing the target
to the array, it can be determined whether a SNP is AA, AB, or BB by analyzing
the resulting signal from the allele-specific probes.
Forty different 25mer oligonucleotides are tiled for each
SNP represented, each oligonucleotide with a slight variation in perfect
matches, mismatches, and flanking sequence around the SNP (illustration
of principle).
The assay requires 250 nanograms of genomic DNA, which
is digested with a restriction enzyme (XbaI), resulting in fragments of various
sizes (simplified
graphical overview). Adapters are ligated to all fragments regardless
of their size, but fragments in the 250- to 1,000-bp size range are preferentially
amplified by PCR using a generic primer that recognizes the adapter sequence.
The amplified DNA is then fragmented, end-labeled and hybridized to the GeneChip®
Mapping 10K Array. After scanning the array Affymetrix Software (GDAS 2.0)
is used to calculate a call for each individual genotype. With 11.555 SNP’s
represented on the array and a call rate of greater than 90% more than 10.000
genotypes should be expected per sample.
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