Highly sensitive amperometric detection of genomic DNA in animal tissues.
A simple and highly sensitive method for the detection of genomic DNA in tissue samples is described. It is based on amperometric detection of target DNA by forming an analyte/polymeric activator bilayer on a gold electrode. The biotinylated target DNA is hybridized to oligonucleotide capture probes immobilized on the gold electrode, forming the first layer. A subsequent binding of glucose oxidase-avidin conjugate to the target DNA and the introduction of a second layer of a redox polymer to the electrode, via layer-by-layer electrostatic self-assembly, allow for electrochemical detection of the catalytic oxidation current of glucose in a PBS solution. Less than 2.0 fg of rat genomic DNA, for both regulated and house-keeping genes, can be easily detected in 2.5 microl droplets. The proposed procedure shows very high specificity for genomic DNA in a RT-PCR mixture.
Xie
Hong
H
Institute of Bioengineering and Nanotechnology, 51 Science Park Road, Singapore 117586, Republic of Singapore.
Yu
Yuan Hong
YH
Mao
Pei-Lin
PL
Gao
Zhiqiang
Z
D000818:Animals; D019921:Biotinylation; D004247:DNA / Q000032:analysis* / Q000235:genetics; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics; D004563:Electrochemistry / Q000379:methods*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics; D008099:Liver / Q000378:metabolism*; D009693:Nucleic Acid Hybridization; D015345:Oligonucleotide Probes / Q000235:genetics; D010084:Oxidation-Reduction; D011108:Polymers / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
D018076:DNA, Complementary; D015345:Oligonucleotide Probes; D011108:Polymers; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D004247:DNA; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases