Electrochemical detection of E. coli 16S rDNA sequence using air-plasma-activated fullerene-impregnated screen printed electrodes.
A new DNA modified electrode for the electrochemical detection of 16S rDNA extracted from Escherichia coli (JCM1649) is proposed. The electrodes were fabricated by screen printing a fullerene-impregnated carbon ink onto a poly(methylmethacrylate) substrate and immobilizing a probe DNA on the surface after activating the electrode with air plasma. The results indicated a dramatic improvement in the surface coverage of the immobilized probe DNA, and of the reduction peak of the redox indicator (Co(phen)(3)(3+)) due to the incorporation of fullerene. By immobilizing the probe onto the fullerene-impregnated screen-printed electrodes, the PCR product of the 16S rDNA extracted from E. coli was directly detected without any pretreatment. A well defined signal difference was observed between the perfectly matching oligonucleotide and the mismatching one, and it was possible to detect the target at the modified electrode. This method enabled us to clearly detect the two base mismatches in the ca. 1500-bases long 16S rDNA sequence.
Shiraishi
Haruki
H
Department of Applied Chemistry, Ritsumeikan University, Noji Higashi 1-1-1, Kusatsu, Shiga, Japan. h-s@se.ritsumei.ac.jp <h-s@se.ritsumei.ac.jp>
Itoh
Tomohito
T
Hayashi
Hiroki
H
Takagi
Kazuyoshi
K
Sakane
Masayasu
M
Mori
Tatsuhiko
T
Wang
Joseph
J
D000388:Air; D020137:Base Pair Mismatch; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics*; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics*; D004563:Electrochemistry / Q000295:instrumentation / Q000379:methods*; D004566:Electrodes; D004926:Escherichia coli / Q000235:genetics*; D037741:Fullerenes / Q000737:chemistry; D006358:Hot Temperature; D017403:In Situ Hybridization / Q000295:instrumentation / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000737:chemistry / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods*; D013499:Surface Properties
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D037741:Fullerenes; D012336:RNA, Ribosomal, 16S