Resumé
A novel method eliminating DNA-mediated PCR product formation in reverse transcription PCR (RT-PCR) amplification of specific RNA sequences is described. The method exploits the higher melting temperature values of peptide nucleic acid (PNA)/DNA duplexes compared with DNA/DNA duplexes by binding a sequence-specific PNA probe to a genomic sequence immediately overlapping one of the PCR-primer attachment sites within the sequence of interest. Hybridization of the blocking probe precludes primer attachment to DNA without affecting attachment of the same primer to the reverse transcription-generated cDNA sequence. A four-step PCR cycle is used that allows the PNA probe to hybridize to the DNA strand at a higher temperature just prior to the primer annealing step.
Originalsprog | Engelsk |
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Sider (fra-til) | 609-614 |
Antal sider | 6 |
Tidsskrift | BioTechniques |
Vol/bind | 42 |
Udgave nummer | 5 |
DOI | |
Status | Udgivet - maj 2007 |
Programområde
- Programområde 2: Vandressourcer