Molecular characterization of large deletions in the von Hippel/-Lindau WHO gene by quantitative real-time PCR - The hypothesis of an Alu-mediated mechanism underlying VHL gene rearrangements
Molecular characterization of large deletions in the von Hippel/-Lindau WHO gene by quantitative real-time PCR - The hypothesis of an Alu-mediated mechanism underlying VHL gene rearrangements
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DOI：
10.1007/bf03256463
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发表时间：
2006-01-01
影响因子：
4
通讯作者：
Murgia, Alessandra
中科院分区：
文献类型：
--
作者：
Casarin, Alberto;Martella, Maddalena;Murgia, Alessandra
简介：VHL基因突变是VHL疾病的原因。这是一种家族性常染色体显性遗传综合征，易发生良性和恶性肿瘤，包括中枢神经系统和视网膜血管母细胞瘤、嗜铬细胞瘤和透明细胞肾癌。使用基于PCR的突变扫描方法无法鉴定这些突变。定量Southern印迹分析已被传统上用于检测完全或部分缺失和更复杂的重排的gene.Methods：一种替代的定量方法，开发了使用定量Southern印迹分析和实时PCR的组合。用这种方法，我们研究了24个大的VHL基因的改变，以确定突变的确切性质，并可能表征的删除regions.Results的边界：这种结合的分子方法表明，所有的VHL改变研究是由于缺失，从该基因中的位置可以更精确地映射。其中一个已完全鉴定的样品在基因内有一个2.2kb的缺失，其5'和3'断裂点均位于重复序列内。结论：这是首次报道VHL大变异的分子分析。我们的研究结果和一个大的缺失的完整表征导致的假设，一个ESTA介导的机制可能是负责常见的VHL基因的大的变化发生。
Introduction: Mutations of the von Hippel-Lindau (VHL) gene are responsible for VHL disease. This is a familial autosomal-dominant syndrome, predisposing to the development of benign and malignant tumors, including CNS and retinal hemangioblastomas, pheochromocytomas, and clear cell renal carcinomas.At least 30% of the disease-causing mutations in the VHL gene involve large alterations. Identification of these mutations is not possible using PCR-based mutational scanning methods. Quantitative Southern blot analysis has been traditionally employed for the detection of complete or partial deletions and more complex rearrangements of the gene.Methods: An alternative quantitative method was developed using a combination of quantitative Southern blot analysis and real-time PCR. With this approach, we studied 24 large VHL gene alterations to determine the exact nature of the mutations and to possibly characterize the boundaries of the deleted regions.Results: This combined molecular approach showed that all the VHL alterations studied were due to deletions, from which the position in the gene could be more precisely mapped. One of the samples that was completely characterized was found to carry an intragenic 2.2kb deletion with both 5' and 3' breakpoints located within Alu-repeat sequences.Conclusion: This is the first report on the molecular analysis of large VHL alterations. The results of our study and the complete characterization of a large deletion lead to the hypothesis that an Alu-mediated mechanism may be responsible for the common occurrence of large alterations in the VHL gene.