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PubMed · 12137775

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

Susumu Goto·Aki Takahashi·Keiichi Kamisango·Koichi Matsubara
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Abstract

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 30min. 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.

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AcridinesAldehyde DehydrogenaseAldehyde Dehydrogenase, MitochondrialApolipoproteins EBiotinylationDNADNA PrimersDNA ProbesGenome, HumanGenotypeHumansNucleic Acid HybridizationOligonucleotide Array Sequence AnalysisPolymerase Chain ReactionPolymorphism, Single NucleotideReproducibility of ResultsJournal Article

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Susumu Goto, Aki Takahashi, Keiichi Kamisango, Koichi Matsubara. 2002-08-01. Single-nucleotide polymorphism analysis by hybridization protection assay on solid support.. https://doi.org/10.1016/s0003-2697(02)00019-2

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