High throughput detection of drug-metabolizing enzyme polymorphisms by allele-specific fluorogenic 5' nuclease chain reaction assay

M. Hiratsuka, Y. Agatsuma, F. Omori, K. Narahara, T. Inoue, Y. Kishikawa, M. Mizugaki

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

We have developed an allele-specific fluorogenic 5' nuclease chain reaction assay for detecting polymorphisms in the following human drug-metabolizing enzyme genes: CYP2C9 (CYP2C9*2 and *3), CYP2C19 (CYP2C19*2 and *3), CYP2D6 (CYP2D6*4, *10, *14, *18, and *21(C8)), N-acetyltransferase 2 (NAT2*5B, *6A, and *7B), thiopurine methyltransferase(TPMT*3C), and aldehyde dehydrogenase2 (ALDH2*2). This method is a marriage of two emerging technologies, the use of allele-specific amplification primers for target DNA and hybridization of the TaqMan probe. The TaqMan probe is labeled with both a fluorescent reporter dye and a quencher dye. Genotypes are separated according to the different threshold cycles of the wild-type and mutant primers. All assays are performed using a single thermocycling protocol. This genotyping method is rapid and highly sensitive and yields a high throughput. It could be applied toward automated large-scale genotyping.

Original languageEnglish
Pages (from-to)1131-1135
Number of pages5
JournalBiological and Pharmaceutical Bulletin
Volume23
Issue number10
DOIs
Publication statusPublished - 2000 Jan 1

Keywords

  • Allele-specific amplification
  • Genotyping
  • High-throughput
  • Polymorphism

ASJC Scopus subject areas

  • Pharmacology
  • Pharmaceutical Science

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