Genotyping and mapping assay of single-nucleotide polymorphisms in CYP3A5 using DNA-binding zinc(II) complexes

It is clinically important to detect the single-nucleotide polymorphism (SNP) of CYP3A5⁎3 (6986A>G) associated with enzymatic activity for drug metabolism. The aim of this study was to establish an accurate strategy for SNP screening. Polyacrylamide gel electrophoresis (PAGE) using zinc(II) compl...

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Bibliographic Details
Published in:Clinical biochemistry 2010-02, Vol.43 (3), p.302-306
Main Authors: Kinoshita, Eiji, Kinoshita-Kikuta, Emiko, Nakashima, Hiromi, Koike, Tohru
Format: Article
Language:English
Subjects:
Base Sequence
Biological and medical sciences
Chromosome Mapping
Cyclen
CYP3A5
Cytochrome P-450 CYP3A - genetics
DNA - chemistry
DNA - metabolism
Electrophoresis
Electrophoresis, Polyacrylamide Gel - methods
General pharmacology
Genotype
Humans
Investigative techniques, diagnostic techniques (general aspects)
Isoenzymes - genetics
Medical sciences
Miscellaneous. Technology
Molecular Sequence Data
Mutation
PAGE
Pathology. Cytology. Biochemistry. Spectrometry. Miscellaneous investigative techniques
Pharmacokinetics. Pharmacogenetics. Drug-receptor interactions
Pharmacology. Drug treatments
Phos-tag
Polymorphism, Single Nucleotide
Protein Binding
Sequence Analysis, DNA
SNP
Zinc - chemistry
Zinc(II) complex
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Summary:It is clinically important to detect the single-nucleotide polymorphism (SNP) of CYP3A5⁎3 (6986A>G) associated with enzymatic activity for drug metabolism. The aim of this study was to establish an accurate strategy for SNP screening. Polyacrylamide gel electrophoresis (PAGE) using zinc(II) complexes were applied for SNP detection. Genomic analyses of 19 healthy subjects were conducted by using both Zn 2+–Phos-tag-PAGE and Zn 2+–cyclen-PAGE methodologies. Zn 2+–Phos-tag PAGE permitted identification of the following allele genotypes: the A/A homozygote ( CYP3A5⁎1/⁎1) in 3 individuals, the G/G homozygote ( CYP3A5⁎3/⁎3) in 14 individuals, and the A/G heterozygote ( CYP3A5⁎1/⁎3) in 2 individuals. Zn 2+–cyclen PAGE demonstrated not only reproducibility of the genotyping but also existence of a novel heterozygous SNP (6929G>A) in the subject with CYP3A5⁎1/⁎1. We demonstrated reliable SNP genotyping and mapping in CYP3A5 using the combination method of Zn 2+–Phos-tag PAGE and Zn 2+–cyclen PAGE.
ISSN:0009-9120
1873-2933
DOI:10.1016/j.clinbiochem.2009.09.007