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@article{1374933,
author = {Dudová, Zdenka and Bartošík, Martin and Fojta, Miroslav},
article_location = {Amsterdam},
article_number = {MAR 20 2017},
doi = {http://dx.doi.org/10.1016/j.electacta.2017.02.104},
keywords = {DNA methyltransferase; DNA methylation; magnetic beads; restriction endonuclease; carbon electrode},
language = {eng},
issn = {0013-4686},
journal = {Electrochimica Acta},
title = {Magnetic bead-based electrochemical assay for determination of DNA methyltransferase activity},
url = {http://dx.doi.org/10.1016/j.electacta.2017.02.104},
volume = {231},
year = {2017}
}
TY - JOUR
ID - 1374933
AU - Dudová, Zdenka - Bartošík, Martin - Fojta, Miroslav
PY - 2017
TI - Magnetic bead-based electrochemical assay for determination of DNA methyltransferase activity
JF - Electrochimica Acta
VL - 231
IS - MAR 20 2017
SP - 575-581
EP - 575-581
PB - Elsevier Science
SN - 00134686
KW - DNA methyltransferase
KW - DNA methylation
KW - magnetic beads
KW - restriction endonuclease
KW - carbon electrode
UR - http://dx.doi.org/10.1016/j.electacta.2017.02.104
L2 - http://dx.doi.org/10.1016/j.electacta.2017.02.104
N2 - DNA methylation, an epigenetic mechanism playing important role in many cellular processes, is carried out via action of DNA methyltransferase (MTase) enzymes. Increasing evidence shows that altered activity of MTases may cause aberrant DNA methylation, which in turn may lead to malignant transformation, and ultimately to cancer. Assessing MTase activity is thus considered of high importance in early cancer diagnostics. Electrochemistry represents an interesting alternative to current methods of MTase analysis, as it is relatively inexpensive, fast and simple. We describe here a development of electrochemical assay in which DNA probe, containing restriction sites CCGG for HapII endonuclease and bearing biotin label at its distal end, is immobilized at magnetic beads and incubated in MTase solution. Methylation of DNA then blocks restriction by HapII and the terminal biotin moiety is preserved to be available for attachment of alkaline phosphatase producing an electroactive indicator, yielding electrochemical signal from the methylated DNA. On the other hand, without MTase the DNA is not methylated and thus the biotinylated DNA end is cleaved-off, leading to the signal diminution. The assay is simple and quick, requiring less than 3 h to completely assess MTase activity. (C) 2017 Elsevier Ltd. All rights reserved.
ER -
DUDOVÁ, Zdenka, Martin BARTOŠÍK and Miroslav FOJTA. Magnetic bead-based electrochemical assay for determination of DNA methyltransferase activity. \textit{Electrochimica Acta}. Amsterdam: Elsevier Science, 2017, vol.~231, MAR 20 2017, p.~575-581. ISSN~0013-4686. Available from: https://dx.doi.org/10.1016/j.electacta.2017.02.104.
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