Development Of Highly Sensitive DNA And Aptamer-based Electrochemical Platforms

In this thesis, according to different target analyte to build three new type ofelectrochemical biosensing platform based on DNA and aptamer. They have beenwidely recognized as highly promising tools for specific sequences of DNA fragmentsand adenosine triphosphate (ATP). The main contents are listed below:1.Herein, a novel enzyme-free and label-free ultrasensitive electrochemical DNAbiosensing platform for the detection of target DNA was firstly proposed, in whichthree auxiliary DNA probes were ingeniously designed to construct the dendritic DNAconcatamer and used as hexaammineruthenium (III) chloride (RuHex) carrier forsignal amplification. With the developed dendritic DNA concatamer-based signalamplification strategy, the DNA biosensor could achieve an ultrasensitiveelectrochemical detection of DNA with a superior detection limit as low as5aM,respectively, and also demonstrate a high selectivity for DNA detection. The currentlyproposed dendritic DNA concatamer opens a promising direction to constructultrasensitive DNA biosensing platform for biomolecular detection in bioanalysis andclinical biomedicine,which offers the distinct advantage of simplicity owing to noneed of any kind of enzyme, chemical modification or labeling.2.In this work, a simple, immobilization-free, isothermal and ultrasensitiveelectrochemical DNA biosensing platform has been developed based on anexonuclease III (Exo III)-aided autocatalytic target recycling strategy. Aferrocene-labeled hairpin probe (HP1) is ingeniously designed, which contains aprotruding DNA fragment at3’termini as the recognition unit for target DNA. Also,the DNA fragment that could be used as secondary target was introduced but it wascaged in the stem region of HP1. In the presence of target DNA, its recognition withthe protruding fragment of HP1triggers the Exo III cleavage process, accompanyingwith the target recycling and autonomous generation of secondary target DNA. Thisaccordingly results into the autonomous accumulation of ferrocene-labeled monucleotide, inducing a distinct increase in the electrochemical signal owing to itselevated diffusivity toward ITO electrode surface. The current developed autocatalyticstrategy provides an ultrasensitive electrochemical detection of DNA down to the0.1pM level, respectively, with a high selectivity. It should be used as a generalautocatalytic strategy toward the detection of a wide spectrum of analytes and may beassociated with more analytical techniques. Thus, it holds great potential for thedevelopment of ultrasensitive biosensing platform for the applications in bioanalysis,disease diagnostics and clinical biomedicine.3.This chapter describes a novel electrochemical aptasensor biosensor for simple,rapid, and specific detection of ATP based on a new three-way DNA junction-drivenToe-hold strand displacement mode assay. This biosensor was constructed byself-assembly of a5’thiolated capture probe and short partial complementary probe onthe gold electrode,firstly.ATP detection was realized by outputting a remarkable redoxcurrent of the3’ ferrocene (Fc) tail labeled aptamer probe. When the target ATP wasinputed into the system, the aptamer as the toehold domain-carrying strand could befull complementary to the5’thiolated capture probe,resulting in forming a stableduplex complex. As a result, the Fc probe was proximate to the electrode surface,andthe Faradaic current was observed.This aptamer biosensor was proved to have a lowdetection limit (5nM) and a wide dynamic range (from5nM to10μM) due to thestable hybridization mode. In addition, the sensing system could discriminate the ATPanaloguesfromATP,with high sensitivity and wonderful stability.