Use of peptide nucleic acid probes for detecting DNA single-base mutations by capillary electrophoresis

作者：Basile A; Giuliani A; Pirri G; Chiari M^*

来源：Electrophoresis, 2002, 23(6): 926-929.

DOI：10.1002/1522-2683(200203)23:6<926::AID-ELPS926>3.0.CO;2-J

• capillary electrophoresis
• hybridization
• mutation detection
• nucleic acid
• peptide nucleic acid

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 singlestranded DNA. We have used as a model system the cystic fibrosis R553X and R1 162X 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.

• 出版日期2002-3