A surface-initiated enzymatic polymerization strategy for electrochemical DNA sensors.
In this work, we report a novel strategy of electrochemical DNA (E-DNA) sensor based on surface initiated enzymatic polymerization (SIEP). This DNA sensor employs a capture DNA probe labeled with thiol at its 3' terminal to be immobilized at gold electrode via gold-thiol chemistry. Oligo (ethylene glycol) -terminated thiols (SH-OEGs) are then used to prepare an oligonucleotide-incorporated nonfouling surface (ONS). After the sequence-specific recognition of target DNA, terminal deoxynucleotidyl transferase (TdT) is employed to catalyze the sequential addition of deoxynucleotides (dNTPs) at the 3'-OH group of target DNA without template. During the TdT-mediated extension reaction, by using biotinlated 2'-deoxyadenosine 5'-triphosphate (biotin-dATP), biotin labels are incorporated into the SIEP-generated long single-stranded DNA (ssDNA). Specific binding of avidin-horseradish peroxidase (Av-HRP) to the biotin label leads to enzyme turnover-based signal transduction. By using this new strategy, we demonstrated the high picomolar sensitivity and a broad detection range of six orders of magnitude.
School of Mechanical Engineering, Nanjing University of Science and Technology, Nanjing 210094, China.
Ying
Y
Wan
Hui
H
Xu
Yan
Y
Su
Xinhua
X
Zhu
Shiping
S
Song
Chunhai
C
Fan
D001704:Biopolymers / Q000737:chemistry; D015374:Biosensing Techniques / Q000295:instrumentation*; D003217:Conductometry / Q000295:instrumentation*; D004247:DNA / Q000032:analysis* / Q000235:genetics*; D004253:DNA Nucleotidylexotransferase / Q000737:chemistry*; D004800:Enzymes, Immobilized / Q000737:chemistry; D004867:Equipment Design; D019544:Equipment Failure Analysis; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013499:Surface Properties
D001704:Biopolymers; D004800:Enzymes, Immobilized; D004247:DNA; D004253:DNA Nucleotidylexotransferase