Accuracy of genotyping for single nucleotide polymorphisms by a microarray-based single nucleotide polymorphism typing method involving hybridization of short allele-specific oligonucleotides.
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.
Iwasaki
Hironori
H
Department of Molecular Biology, Institute of Gerontology, Nippon Medical School, Kawasaki, Japan.
Ezura
Yoichi
Y
Ishida
Ryota
R
Kajita
Mitsuko
M
Kodaira
Mina
M
Knight
Jim
J
Daniel
Steve
S
Shi
Michael
M
Emi
Mitsuru
M
D000483:Alleles; D005838:Genotype; D006801:Humans; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide; D015203:Reproducibility of Results