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  • Title: Use of peptide nucleic acid probes for detecting DNA single-base mutations by capillary electrophoresis.
    Author: Basile A, Giuliani A, Pirri G, Chiari M.
    Journal: Electrophoresis; 2002 Mar; 23(6):926-9. PubMed ID: 11920878.
    Abstract:
    Peptide nucleic acid (PNA) oligomers can be used as probes in pre-gel hybridization experiments, as an alternative to Southern hybridization. In this technique, the PNA probe is hybridized to a cyanine-5 labeled DNA sample denatured at low ionic strength, and the mixture is directly injected for size separation into a capillary electrophoresis (CE) system equipped with laser-induced fluorescence (LIF) detector. The neutral backbone of PNA allows hybridization to occur at low ionic strength and assures an efficient CE separation of the PNA/DNA hybrids from both double-stranded and single-stranded DNA. We have used as a model system the cystic fibrosis R553X and R1162X single-base mutations and we have assessed the influence of various factors, such as temperature and denaturants concentration on DNA/PNA hybrid stability in order to achieve the high specificity required for a single base pair discrimination.
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