Electrochemical Detection of Asymmetric PCR Products by Labeling with Osmium Tetroxide

✍ Scribed by Maren Mix; Thomas Reske; Heiko Duwensee; Gerd-Uwe Flechsig

• Book ID: 102179056
• Publisher: John Wiley and Sons
• Year: 2009
• Tongue: English
• Weight: 157 KB
• Volume: 21
• Category: Article
• ISSN: 1040-0397
• DOI: 10.1002/elan.200804493

✦ Synopsis

Abstract

Single stranded DNA‐targets from asymmetric polymerase chain reaction (PCR) of a sequence of the gram positive, spore forming bacterium Clostridium acetobutylicum were detected by square‐wave voltammetry after labeling with osmium tetroxide bipyridine and hybridization with DNA capture probes immobilized on gold electrodes. The asymmetric PCR, performed with a 10‐fold excess of the forward‐primer, was used without any further purification for hybridization with protective strands and covalent labeling with osmium tetroxide bipyridine. Square‐wave voltammetric signals of 20 nmol/L targets were significantly higher at 50 °C compared with 23 °C hybridization temperature. A fully noncomplementary protective strand yielded thoroughly modified targets unable for further hybridization. Coupling this with thermal discrimination opens new opportunities for sequence specific DNA detection.