Miniaturized electrochemical detector as a tool for detection of DNA amplified by PCR.
This paper reports on the analysis of specific sequence of Phage Lambda DNA amplified by PCR. Agarose gel electrophoresis, gel electrophoresis on chip and stationary electrochemical instrument were employed for detection of amplicons obtained after 2, 4, 6, 8, 10, 15, 20, 25, 30 and 35 cycles. In the case of agarose gel electrophoresis the lowest detectable amount of DNA was obtained after 15 PCR cycles. Gel electrophoresis on chip offers higher sensitivity because the lowest detectable amount of amplicons by this technique was obtained after eight PCR cycles. Further we employed square wave voltammetry and various working electrodes (hanging mercury drop electrode, screen-printed carbon electrode and carbon-nanotube-based screen-printed electrodes) to detect amplicons. Amplicons obtained even after two cycles were detectable at all electrodes. To improve the selectivity of electrochemical detection carbon nanoelectrodes were off-line coupled with gel electrophoresis. Into the agarose gel the electrodes were placed. Further the amplicons were loaded into agarose gel wells. DNA migrating to the detection place was electrochemically analyzed. Amplicons obtained after two cycles were detectable by this hyphenated technique.
Huska
Dalibor
D
Department of Chemistry and Biochemistry, Faculty of Agronomy, Mendel University of Agriculture and Forestry, Brno, Czech Republic.
Hubalek
Jaromir
J
Adam
Vojtech
V
Kizek
Rene
R
D004247:DNA / Q000032:analysis*; D004587:Electrophoresis, Agar Gel / Q000295:instrumentation* / Q000379:methods; D046209:Electrophoresis, Microchip / Q000295:instrumentation* / Q000379:methods; D037742:Nanotubes, Carbon / Q000737:chemistry; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods
D037742:Nanotubes, Carbon; D004247:DNA