Determination of single nucleotide polymorphisms in N-acetyltransferase2 gene using an electrochemical DNA chip and an automated DNA detection system.
An electrochemical DNA chip using an electrochemically active intercalator and DNA probe immobilized on a gold electrode has been developed for genetic analysis. In this study, N-acetyltransferase2 (NAT2) gene polymorphisms (C481T G590A G857A) were determined by the electrochemical DNA chip and the automated DNA detection system that performs hybridization reaction, washing, detection, and data analysis. Human genomic DNAs were extracted from blood and DNA fragments containing the three polymorphisms were amplified by the polymerase chain reaction (PCR) method. Double-stranded PCR products were treated with T7 exonuclease and single-stranded target DNAs were obtained. A sample containing the single-stranded target DNAs was injected into a cassette including the electrochemical DNA chip and set in an automated system. The turnaround time for genotyping with this system was 90 min. A total of 38 samples were automatically genotyped by an SNP determination algorithm. The results of genotype were completely consistent with those determined by the polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis. Consequently, this method requires no labeling step and has the advantage of realizing a compact and automatic system, and so the system is expected to contribute to personalized medicine based on genotype.
Nakamura
Naoko
N
Corporate Research & Development Center, Toshiba Corporation, Kawasaki.
Ito
Keiko
K
Hongo
Sadato
S
Hashimoto
Kouji
K
Furutsuka
Miyuki
M
Kubota
Ryuuji
R
Fukuda
Tsuyoshi
T
Ohno
Masako
M
Azuma
Junichi
J
Gemma
Nobuhiro
N
D001191:Arylamine N-Acetyltransferase / Q000235:genetics*; D004247:DNA / Q000097:blood; D015342:DNA Probes; D004563:Electrochemistry / Q000379:methods*; D005838:Genotype; D006801:Humans; D008839:Microelectrodes; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010597:Pharmacogenetics; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*
D015342:DNA Probes; D004247:DNA; D001191:Arylamine N-Acetyltransferase; C478900:NAT2 protein, human