| In this paper, electrochemical methods were used to study the interaction between [Cu(dmp)(H2O)Cl2], 2-aminophenoxazin-3-one and 4,4'-diaminoazobenzene with salmon sperm DNA to optimize conditions. These sensors were prepared by immobilizing single-stranded DNA probes on different electrodes and using electroactive indicators to measure the hybridization events between the DNA probes and their complementary DNA fragments. The electrochemical DNA biosensor developed might have potential application in designing of novel anti-AIDS drugs and diagnosis disease.In the first chapter, the action modes of small molecules compounds with deoxyribonucleic acid and research methods were summaried, the principle and classification of DNA biosensor was introduced. Research status of DNA biosensor and research progress of aptamer sensor were reviewed. This article described mainly the research status of DNA electrochemical biosensor.In the second chapter, 2,9-dimethyl-1,10-phenanthroline copper complex was synthesized by o-nitroaniline as starting material and characterized by elemental analysis and IR spectra. An electrochemical DNA biosensor was developed based on the recognition of target DNA by hybridization detection. The study was carried out using glassy carbon electrode (GCE) modified with lable-free 21-mer single stranded oligonucleotides related to hepatitis B virus sequence via covalent immobilization and [Cu(dmp)(H2O)Cl2] (dmp = 2,9-dimethyl-1,10- phenan- throline) as an electrochemical indicator, whose sizes are comparable to those of the small groove of native double-duplex DNA. The method, which is simple and low cost, allows the accumulation of copper complex within the DNA layer. Electochemical detection was performed by cyclic voltammetry and differential pulse voltammetry (DPV) over the potential range where the [Cu(dmp)(H2O)Cl2] was active. Numerous factors affecting the probe immobilization, target hybridization, and indicator binding reactions were optimized to maximize the sensitivity and speed the assay time. With this approach, a sequence of the hepatitis B virus could be quantified over the ranges from 8.82×10-8 to 8.82×10-7 mol·L-1 with a linear correlation of r = 0.9937 and a detection limit of 7.0×10-8 mol·L-1. The [Cu(dmp)(H2O)Cl2] signal observed from probe sequence before and after hybridization with four bases mismatch containing sequence is lower than that observed after hybridization with complementary sequence.In the third chapter, A surface-based method for the study of the interaction of DNA with redox-active 2-aminophenoxazin-3-one was described. The study was carried out using glassy carbon electrode (GCE) modified with 21-mer single stranded oligonucleotides related to hepatitis B virus sequence via covalent immobilization and 2-aminophenoxazin-3-one as an electrochemical indicator. Electochemical detection was performed by cyclic voltammetry and differential pulse voltammetry over the potential range where the AP was redox active. Numerous factors affecting the probe immobilization, target hybridization, and indicator binding reactions were optimized to maximize the sensitivity and speed the assay time. With this approach, a sequence of the hepatitis B virus from human blood could be quantified over the ranges from 3.53×10-7 to 1.08×10-6 mol·L-1 with a linear correlation of r = 0.9963 and a detection limit of 1.00×10-7 mol·L-1 (S/N = 3). The AP signal observed from probe sequence before the hybridization is higher than that observed after hybridization with complementary sequence. With the increase of the number of guanine bases, the cathodic currents were increased. AP preferred to interact with guanine bases as that MB did.In the fourth chapter, A novel electrochemical DNA biosensor based on 4,4'-diaminoazobenzene (4,4'-DAAB) and multi walled carbon nanotubes(MWNT)-modified glassy carbon electrode (GCE) for short DNA sequences related to the hepatitis B virus (HBV) hybridization detection was presented. Differential pulse voltammetry was used to investigate hybridization event. The decrease in the peak current of 4,4'-DAAB was observed upon hybridization of probe with the target. This electrochemical approach is sequence specific as indicated by the control experiments in which no peak current change was observed if a noncomplementary DNA sequence was used. Numerous factors affecting the target hybridization are optimized to maximize the sensitivity. Under optimal conditions, this sensor has a good calibration range between 7.94×10-8 and 1.58×10-6 mol·L-1, with HBV DNA sequence detection limit of 1.14×10-8 mol·L-1. In the last chapter, it was summarized for the whole thesis. |
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