Rapid single nucleotide polymorphism analysis by primer extension and capillary electrophoresis using polyvinyl pyrrolidone matrix
| Autor: | Maria Sasvari-Szekely, Zsolt Ronai, Csaba Barta, András Guttman |
|---|---|
| Rok vydání: | 2001 |
| Předmět: |
Clinical Biochemistry
Mutant Biology Biochemistry Molecular biology Primer extension Analytical Chemistry law.invention chemistry.chemical_compound Capillary electrophoresis chemistry law Primer dimer Multiplex ligation-dependent probe amplification Primer (molecular biology) DNA Polymerase chain reaction |
| Zdroj: | ELECTROPHORESIS. 22:779-782 |
| ISSN: | 1522-2683 0173-0835 |
| Popis: | Rapid molecular diagnosis of 21-hydroxylase deficiency by detecting the most common mutation in the 21-hydroxylase gene is presented using primer extension and capillary electrophoresis with a polyvinyl pyrrolidone matrix. DNA samples were subjected to polymerase chain reaction (PCR) in order to amplify a 422 bp fragment of the CYP21 gene containing the single nucleotide polymorphism (SNP) site. This product served as a template in the primer extension reaction using a fluorescently labeled primer in close proximity to the SNP. ddGTP was used to block the extension if the mutation was present and the other three dNTPs to enable elongation of the primer. Fast analysis of the resulting fragments was performed by capillary electrophoresis using 10% polyvinylpyrrolidone as sieving and wall coating matrix. The Cy5-labeled primer and the two possible primer extension products (mutant and wild type) were completely separated in 90 s. |
| Databáze: | OpenAIRE |
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