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Accuracy of Genotyping for Single Nucleotide Polymorphisms by a Microarray-Based Single Nucleotide Polymorphism Typing Method Involving Hybridization of Short Allele-Specific Oligonucleotides

2002; University of Oxford; Volume: 9; Issue: 2 Linguagem: Inglês

10.1093/dnares/9.2.59

1756-1663

Hironori Iwasaki, Yoichi Ezura, Ryota Ishida, Mitsuko Kajita, M. Kodaira, Jim Knight, Steve Daniel, Michael Shi, Mitsuru Emi,

Molecular Biology Techniques and Applications

Advances in technologies for identifying genetic polymorphisms rapidly and accurately will dramatically accelerate the discovery of disease-related genes. Among a variety of newly described methods for rapid typing of single-nucleotide polymorphisms (SNPs), gene detection using DNA microarrays is gradually achieving widespread use. This method involves the use of short (11- to 13-mer) allele-specific oligonucleotides. This method allows simultaneous analysis of many SNPs in DNAs from a large number of individuals, in a single experiment. In this work, we evaluated the accuracy of a new microarray-based short allele-specific oligonucleotide (ASO) hybridization method. There is a 96-well formatted array on a single plate, in which up to 256 spots are included in each well. Fluorescent probes for our experiments were produced by multiplex PCR amplification often target SNP-containing regions. We genotyped 192 individuals across a panel of ten single base variations, which included an insertion/deletion polymorphism. For comparison, we genotyped the same individuals for the same SNPs by the method of single-base extension with fluorescence detection. The typing accuracies of the microarray-based PCR-ASO and single-base extension methods were calculated as 99.9% and 99.1%, respectively, on the basis of genotyping results determined by direct sequencing. We conclude that the microarray-based hybridization method using short ASO probes represents a potential breakthrough technology for typing large numbers of SNPs rapidly and efficiently.