Abstract
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.
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Acknowledgements
The authors wish to thank Dr Catherine Stolle and her collaborators at the Molecular Genetics Laboratory, The Children’s Hospital of Philadelphia, Philadelphia, PA, USA, for kindly providing DNA samples that have been included in this study. We thank Dr Franca Anglani, Department of Medical and Surgical Sciences of the University of Padua, for fruitful discussions. The authors also wish to thank Dr Nicola Marziliano for helping with primer design. This work has been supported by the University of Padua, grant ‘ex 60%’.
The authors have no conflicts of interest that are directly relevant to the content of this article.
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Casarin, A., Martella, M., Polli, R. et al. Molecular Characterization of Large Deletions in the von Hippel-Lindau (VHL) Gene by Quantitative Real-Time PCR. Mol Diag Ther 10, 243–249 (2006). https://doi.org/10.1007/BF03256463
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DOI: https://doi.org/10.1007/BF03256463