The importance of gene dosage studies:: mutational analysis of the parkin gene in early-onset parkinsonism
The importance of gene dosage studies:: mutational analysis of the parkin gene in early-onset parkinsonism
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DOI：
10.1093/hmg/10.16.1649
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发表时间：
2001-08-01
影响因子：
3.5
通讯作者：
Klein, C
中科院分区：
文献类型：
--
作者：
Hedrich, K;Kann, M;Klein, C
早发性帕金森综合征（EOP）可能与帕金基因的不同突变有关，包括外显子缺失和重复。为了测试基因剂量改变，我们开发了一种新的定量双重PCR方法，该方法使用LightCycler（Roche Diagnostics）上的荧光共振能量转移技术。在21例EOP患者中，通过常规突变筛查（单链构象多态性和序列分析）检测到3个突变（外显子3的单碱基对替换和外显子9的小缺失），而在7例患者中发现基因剂量改变。我们鉴定了外显子2、3、5和7的杂合和复合杂合缺失。后者也被发现在纯合子状态。此外，还观察到外显子4的两个杂合重复。值得注意的是，两名患者携带了两种以上的帕金突变。这是第一项通过实时动力学定量系统筛选Parkin所有12个外显子的研究，并清楚地表明Parkin基因的突变分析应包括基因剂量研究。此外，我们的定量PCR方法很容易适用于任何其他基因的缺失或重复的整个外显子进行筛选。
Early-onset parkinsonism (EOP) may be associated with different mutations in the Parkin gene, including exon deletions and duplications. To test for gene dosage alterations, we developed a new method of quantitative duplex PCR using the fluorescence resonance energy transfer technique on the LightCycler (Roche Diagnostics). In 21 patients with EOP, three mutations (a single base pair substitution in exon 3 and small deletions in exon 9) were detected by conventional mutational screening (single-strand conformation polymorphism and sequence analysis), while alterations of gene dosage were found in seven patients. We identified heterozygous and compound heterozygous deletions of exons 2, 3, 5 and 7. The latter was also found in the homozygous state. In addition, two heterozygous duplications of exon 4 were observed. Remarkably, two patients; carried more than two Parkin mutations. This is the first study systematically screening all 12 exons of Parkin by real-time, kinetic quantification and clearly shows that mutational analysis of the Parkin gene should include gene dosage studies. Furthermore, our method of quantitative PCR is easily applicable to any other gene to be screened for deletions or duplications of whole exons.