Single-nucleotide polymorphism analysis by hybridization protection assay on solid support

The clinical need for high-throughput typing methods of single-nucleotide polymorphisms (SNPs) has been increasing. Conventional methods do not perform well enough in terms of speed and accuracy to process a large number of samples, as in clinical testing. We report a new DNA microarray method that...

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Veröffentlicht in:Analytical biochemistry 2002-08, Vol.307 (1), p.25-32
Hauptverfasser: Goto, Susumu, Takahashi, Aki, Kamisango, Keiichi, Matsubara, Koichi
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Sprache:eng
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Zusammenfassung:The clinical need for high-throughput typing methods of single-nucleotide polymorphisms (SNPs) has been increasing. Conventional methods do not perform well enough in terms of speed and accuracy to process a large number of samples, as in clinical testing. We report a new DNA microarray method that uses hybridization protection assay (HPA) by acridinium-ester-labeled DNA probes. Probes were immobilized on the bottom of streptavidin-coated microtiter plates by streptavidin–biotin binding. We studied aldehyde dehydrogenase 2 (ALDH2) genotyping using two probes, discriminating A/G polymorphism. We also designed four probes to type the Alzheimer's disease-related gene ApoE, which has three genotypes (ApoE2, 3, and 4) determined by two SNP loci (C/T polymorphism). SNP analysis of the ALDH2 gene or the ApoE gene from human genome samples by solid-phase HPA was successful. Unlike other methods, the microarray by HPA does not require a washing step and can be completed within 30 min. It also has advantages in discriminating one-base mismatch in targets. These characteristics make it a good candidate for practical SNP analysis of disease-related genes or drug-metabolizing enzymes in large numbers of samples.
ISSN:0003-2697
1096-0309
DOI:10.1016/S0003-2697(02)00019-2