CYP2D6 genotyping by liquid chromatography-electrospray ionization mass spectrometry
CYP2D6 genotyping by liquid chromatography-electrospray ionization mass spectrometry
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DOI：
10.1007/s00216-010-4597-4
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发表时间：
2011-06-01
影响因子：
4.3
通讯作者：
Oberacher, Herbert
中科院分区：
文献类型：
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作者：
Beer, Beate;Erb, Robert;Oberacher, Herbert
遗传多态性可显著影响药物代谢酶细胞色素P450 2D6 （CYP2D6; OMIM 124030）的酶活性。因此，CYP2D6基因分型被认为是预测个体CYP2D6代谢状态的有效方法。我们引入离子对反相高效液相色谱-电喷雾电离质谱（ICEMS）作为表征CYP2D6基因单碱基变异、小缺失和插入的方法。建立了两步聚合酶链反应（PCR），同时扩增CYP2D6基因的9个多态性区域。通过高效液相色谱法对PCR扩增子进行清理、分离和变性。高性能分子质量测量提供了核苷酸组成谱，主要能够分辨37个已报道的CYP2D6等位基因。该方法应用于93例无亲缘关系的奥地利人的基因分型。为了验证，选择了一些样本和多态性位点，通过其他基因分型技术进行了重新分型。PCR-ICEMS分析被证明是一种准确、稳健、经济的CYP2D6基因分型策略。
Genetic polymorphisms can significantly affect the enzyme activity of the drug metabolizing enzyme Cytochrome P450 2D6 (CYP2D6; OMIM 124030). Accordingly, CYP2D6 genotyping is considered as a valid approach to predict the individual CYP2D6 metabolizing status. We introduce ion-pair reversed-phase high-performance liquid chromatography-electrospray ionization mass spectrometry (ICEMS) as method for the characterization of single base variants, small deletions, and insertions in the CYP2D6 gene. A two-step polymerase chain reaction (PCR) was developed for the simultaneous amplification of nine polymorphic regions within the CYP2D6 gene. Cleanup, separation, and denaturation of PCR amplicons were achieved by high-performance liquid chromatography. High-performance molecular mass measurements provided nucleotide composition profiles that principally enable the resolution of 37 reported CYP2D6 alleles. The developed assay was applied to the genotyping of 93 unrelated Austrian individuals. For validation, a selected number of samples and polymorphic sites were retyped by alternative genotyping technologies. The PCR-ICEMS assay turned out to be an accurate, robust, and cost-effective CYP2D6 genotyping strategy.