一种用于检测GSTM1和GSTT1拷贝数的多重实时聚合酶链反应方法。
A multiplex real-time PCR method for detection of GSTM1 and GSTT1 copy numbers.
作者信息
Timofeeva Maria, Jäger Birgit, Rosenberger Albert, Sauter Wiebke, Wichmann Heinz-Erich, Bickeböller Heike, Risch Angela
机构信息
German Cancer Research Center (DKFZ), Division of Epigenomics and Cancer Risk Factors (C010), Im Neuenheimer Feld 280, Heidelberg, Germany.
出版信息
Clin Biochem. 2009 Apr;42(6):500-9. doi: 10.1016/j.clinbiochem.2008.12.011. Epub 2008 Dec 30.
OBJECTIVES
Deletion polymorphisms of Glutathione-S-transferase (GST) M1 and T1 are considered risk factors for various diseases. However, most previous studies only distinguished "null" and "non-null" genotypes. Our aim was to develop a reliable, high-throughput GSTM1/T1 genotyping method able to determine allele copy numbers.
DESIGN AND METHODS
We developed a multiplex real time PCR method to distinguish between heterozygous (1/0) and homozygous (1/1) GSTM1 and GSTT1 genotypes. The principle of relative quantification was applied and an expectation-maximisation (EM) algorithm was developed to assign one of 3 possible genotypes: 1/1, 1/0 or 0/0 for each of the two genes.
RESULTS
1320 Caucasians were genotyped using the newly developed method. The observed genotype distributions did not deviate from the expected and were in Hardy-Weinberg equilibrium. GSTM1 duplication was detected in one sample.
CONCLUSION
This new semiquantitative genotyping method is a sensitive and promising tool for large-scale molecular epidemiological and clinical studies.
目的
谷胱甘肽 - S - 转移酶（GST）M1和T1的缺失多态性被认为是多种疾病的危险因素。然而，大多数先前的研究仅区分“无效”和“非无效”基因型。我们的目的是开发一种可靠的、高通量的GSTM1/T1基因分型方法，能够确定等位基因拷贝数。
设计与方法
我们开发了一种多重实时PCR方法，以区分杂合子（1/0）和纯合子（1/1）的GSTM1和GSTT1基因型。应用相对定量原理，并开发了一种期望最大化（EM）算法，为两个基因中的每一个指定三种可能的基因型之一：1/1、1/0或0/0。
结果
使用新开发的方法对1320名白种人进行了基因分型。观察到的基因型分布与预期没有偏差，处于哈迪 - 温伯格平衡。在一个样本中检测到GSTM1重复。
结论
这种新的半定量基因分型方法是大规模分子流行病学和临床研究的一种敏感且有前景的工具。
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