https://scholars.lib.ntu.edu.tw/handle/123456789/508358
標題: | Simultaneous detection of single nucleotide polymorphisms and copy number variations in the CYP2D6 gene by multiplex polymerase chain reaction combined with capillary electrophoresis | 作者: | Liao H.-W. Tsai I.-L. GUAN-YUAN CHEN Kuo C.-T. Wei M.-F. TZUNG-JENG HWANG WEI J. CHEN LI-JIUAN SHEN CHING-HUA KUO |
公開日期: | 2013 | 卷: | 763 | 起(迄)頁: | 67-75 | 來源出版物: | Analytica Chimica Acta | 摘要: | CYP2D6 (cytochrome P450 2D6) is one of the most important enzymes involved in drug metabolism, and CYP2D6 gene variants may cause toxic effects of therapeutic drugs or treatment failure. In this research, a rapid and simple method for genotyping the most common mutant alleles in the Asian population (CYP2D6*1/*1, CYP2D6*1/*10, CYP2D6*10/*10, CYP2D6*1/*5, CYP2D6*5/*10, and CYP2D6*5/*5) was developed by allele-specific polymerase chain reaction (AS-PCR) combined with capillary electrophoresis (CE). We designed a second mismatch nucleotide next to the single nucleotide polymorphism (SNP) site in allele-specific primers to increase the difference in PCR amplification. Besides, we established simulation equations to predict the CYP2D6 genotypes by analyzing the DNA patterns in the CE chromatograms. The multiplex PCR combined with CE method was applied to test 50 patients, and all of the test results were compared with the DNA sequencing method, long-PCR method and real-time PCR method. The correlation of the analytical results between the proposed method and other methods were higher than 90%, and the proposed method is superior to other methods for being able to simultaneous detection of SNPs and copy number variations (CNV). Furthermore, we compared the plasma concentration of aripiprazole (a CYP2D6 substrate) and its major metabolites with the genotype of 25 patients. The results demonstrate the proposed genotyping method is effective for estimating the activity of the CYP2D6 enzyme and shows potential for application in personalized medicine. Similar approach can be applied to simultaneous detection of SNPs and CNVs of other genes. ? 2012 Elsevier B.V. |
URI: | https://scholars.lib.ntu.edu.tw/handle/123456789/508358 | ISSN: | 0003-2670 | DOI: | 10.1016/j.aca.2012.12.005 | SDG/關鍵字: | Analytical results; Aripiprazole; Copy number; Cytochrome p450; DNA patterns; DNA Sequencing; Drug metabolisms; Gene variants; Genotyping; Multiplex pcr; Mutant alleles; PCR amplification; Personalized medicines; Plasma concentration; Real-time PCR method; SIMPLE method; Simulation equations; Simultaneous detection; Single nucleotide polymorphisms; Therapeutic drugs; Toxic effect; Treatment failure; Capillary electrophoresis; Chromatographic analysis; DNA sequences; Enzyme activity; Gene therapy; Multiplexing; Nucleotides; Polymerase chain reaction; aripiprazole; cytochrome P450 2D6; genomic DNA; allele; article; base mispairing; blood sampling; capillary electrophoresis; copy number variation; correlation analysis; CYP2D6 gene; DNA determination; DNA sequence; drug blood level; enzyme activity; gene; gene activity; gene function; genotype; human; multiplex polymerase chain reaction; priority journal; sequence analysis; single nucleotide polymorphism; Cytochrome P-450 CYP2D6; DNA Copy Number Variations; Electrophoresis, Capillary; Genetic Techniques; Genetic Variation; Genotype; Humans; Multiplex Polymerase Chain Reaction; Polymorphism, Single Nucleotide |
顯示於: | 法醫學科所 |
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