Assessment of a rapid-cycle PCR assay for the identification of the recurrent c.3421C>T mutation in the ABCC6 gene in pseudoxanthoma elasticum patients
| Autor: | Thomas Brinkmann, Jens Dreier, Veronika Schulz, Alexander Grundt, Christian Götting, Christiane Szliska, Doris Hendig, Knut Kleesiek |
|---|---|
| Rok vydání: | 2004 |
| Předmět: |
Adult
Male Genotype DNA Mutational Analysis ABCC6 Biology medicine.disease_cause Polymerase Chain Reaction Pathology and Forensic Medicine Exon medicine Humans Point Mutation Pseudoxanthoma Elasticum Molecular Biology Genotyping Mutation Point mutation Reproducibility of Results DNA Cell Biology Middle Aged Pseudoxanthoma elasticum medicine.disease Molecular biology Restriction enzyme biology.protein Female Multidrug Resistance-Associated Proteins |
| Zdroj: | Laboratory Investigation. 84:122-130 |
| ISSN: | 0023-6837 |
| DOI: | 10.1038/labinvest.3700004 |
| Popis: | Pseudoxanthoma elasticum (PXE) is a heritable disorder of the connective tissue. Mutations in the ABCC6 gene could be linked to this disease and, just recently, the c.3421C>T mutation was also associated with a high risk of coronary artery disease. We have now developed new real-time PCR assays for the accurate and rapid determination of the c.3421C>T genotype. Using our new assay, we analyzed the presence of the c.3421C>T mutation in the largest collection of DNA samples from unrelated German PXE patients (n=64) and in a control cohort (n=910). For assay setup, two sets of samples with known genotype for the c.3421C>T mutation were analyzed over a period of 14 days. Results were confirmed by restriction endonuclease mapping, sequence-specific PCR and DNA sequencing. In order to ensure that no further mutations or deletions interfered with the c.3421C>T genotyping, we scanned the exon 24 of the ABCC6 gene by DHPLC and investigated the presence of the ABCC6del23–29 deletion in all patients. The assay has been set up on a group of patients with known genotype and validated on 64 PXE patients. In this group four PXE patients (6.3%) were found to be homozygous and 25 (39.0%) to be heterozygous carriers of the c.3421C>T mutation. The common ABCC6del23–29 deletion, possibly interfering with genotype determination, was searched and excluded. Furthermore, two novel mutations in the ABCC6 gene could be identified in two patients. The novel mutations c.3389C>T and c.3341G>A did not interfere with our new assay. Our new c.3421C>T genotyping assays can be used for the rapid identification of this frequent mutation in PXE patients and of the recently newly proposed cardiac risk factor in young patients with myocardial infarcts of unknown origin. |
| Databáze: | OpenAIRE |
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