应用 SmartAmp 方法直接从人血样本中对细胞色素 P450 2A6 全基因缺失进行基因分型。
Direct genotyping of Cytochrome P450 2A6 whole gene deletion from human blood samples by the SmartAmp method.
机构信息
Laboratory of Drug Metabolism and Pharmacokinetics, Showa Pharmaceutical University, Machida, Tokyo 194-8543, Japan.
出版信息
Clin Chim Acta. 2011 Jun 11;412(13-14):1249-51. doi: 10.1016/j.cca.2011.03.017. Epub 2011 Mar 21.
BACKGROUND
Genetic polymorphisms of the human CYP2A6 gene are considered to be a determinant of smoking behavior and tobacco-related lung cancer risk in male Japanese smokers. We developed a SmartAmp-based genotyping method to detect whole deletion of the CYP2A6 gene directly from blood samples without DNA isolation.
METHODS
We validated the new method using CYP2A plasmids, 48 genomic DNA samples and 25 blood samples by utilizing the SmartAmp method, a unique isothermal DNA amplification process.
RESULTS
This method could discriminate the CYP2A6 gene from highly homologous CYP2A7 and CYP2A13 genes. CYP2A61 (wild-type) and CYP2A64 (whole gene deletion) were determined by the new method in perfect accordance with sequence analysis data.
CONCLUSIONS
A SmartAmp assay for genotyping the CYP2A6 gene was developed, and the reliability of the method was validated using the conventional PCR method.
背景
人类 CYP2A6 基因的遗传多态性被认为是决定日本男性吸烟者吸烟行为和与烟草相关的肺癌风险的一个因素。我们开发了一种基于 SmartAmp 的基因分型方法，可直接从血液样本中检测 CYP2A6 基因的全缺失，而无需 DNA 分离。
方法
我们使用 CYP2A 质粒、48 个基因组 DNA 样本和 25 个血液样本，利用 SmartAmp 方法（一种独特的等温 DNA 扩增过程）对新方法进行了验证。
结果
该方法能够区分高度同源的 CYP2A7 和 CYP2A13 基因。新方法可准确区分 CYP2A61（野生型）和 CYP2A64（全基因缺失），与序列分析数据完全一致。
结论
开发了一种用于 CYP2A6 基因分型的 SmartAmp 检测方法，并使用常规 PCR 方法验证了该方法的可靠性。
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