Enzymatic Incorporation of Biotin into DNA for DNA Hybridization Analysis and for Sensitive Detection of PCR-Amplified DNA

0506770 - BFÚ 2020 RIV CZ eng C - Conference Paper (international conference)
Špaček, Jan - Zenka, Martin - Haroniková, Lucia - Havran, Luděk - Fojta, Miroslav
Enzymatic Incorporation of Biotin into DNA for DNA Hybridization Analysis and for Sensitive Detection of PCR-Amplified DNA.
XXXIV. Moderní elektrochemické metody sborník přednášek. Ústí nad Labem: Best servis, 2014 - (Navrátil, T.; Fojta, M.; Pecková, K.), Č. 2014 (2014), s. 193-196. ISBN 978-80-905221-2-1.
[Moderní Elektrochemické Metody /34./. Jetřichovice (CZ), 19.05.2014-23.05.2014]
R&D Projects: GA ČR GAP206/12/2378; GA ČR GAP206/11/1638
Institutional support: RVO:68081707
Keywords : transferase * sensors * DNA hybridization * Electrochemical DNA sensors
OECD category: Electrochemistry (dry cells, batteries, fuel cells, corrosion metals, electrolysis)

We present two enzymatical electrochemical assays for DNA analysis. For hybridization analysis we used probes with biotin-14-dC introduced to 3' OH end by terminal transferase. For detection of PCR products we used Deep Vent polymerase to incorporate biotin-14-dCduring PCR. In both cases streptavidin-alkaline phosphatase conjugate was subsequently attached to the incorporated biotins and was used to catalyze dephosphorylation of 1-naphthyl phosphate to 1-naphthol, the electrochemical signal of which was utilized for detection. Compared to the former method, biotin incorporation during PCR offers lower molar detection limits, whereas application of the biotin-tailed probe can provide us with more selective detection.
Permanent Link: http://hdl.handle.net/11104/0297943