Electrochemical DNA Sensors Based On Polythionine/Prussian Blue-Gold Nanocomposite/Gold Nanoparticles

Electrochemical DNA sensor has good specificity, good stability, high sensitivity, with simple preparation process,versatile, and many other advantages.There is a wide range of applications in genetic engineering, gene diagnosis,gene therapy, environmental monitoring and other fields. In the field of DNA sensors, thionine (Th) and Prussian Blue-Gold Nanocomposite (PB-Au) are good mediators, which are able to improve the conductivity of the electrode, thus widespread attention. Gold nanoparticles (AuNPs) have a large surface area, high reactivity and good biocompatibility.And it is easy to conbine biologically active substances (eg, DNA, proteins) via Au-NH2bond, fixing the biological macromolecules to the electrode surface. In this thesis, according to PB-Au’s good performance of the transmission of electron, stability and conductivity of polythionine film.we use cyclic voltammetry and electrodeposition technique to prepare current type DNA sensor based on PTh/PB-Au/AuN-Ps,and it is used in the detection of DNA and DNA damage.(1) The electric polymerization of thionine on glassy carbon electrode surfaceWe studied the potential of the scan range, buffer type, acidity, concentration of thionine concentration in the polymerization solution and a buffer solution for electrical thionine’s process by cyclic voltammetry. The results showed that when the CV’s potential in ABS was-0.45～1.1Vand in PBS was-0.45～1.05V, the GCE/PTh was obtained. When the anode potential was less than1.0V, the GCE/Th was obtained. When ABS was pH7.0,PBS was pH6.0and the concentration of thionine was0.005mol/L, the electrochemical redox peaks current is the maximum and RCt is the minimum.(2) Electrochemical DNA sensors Based on PTh/PB-Au/AuNPsThionine was electropolymerized on the glassy carbon electrode surface, and then PB-Au and AuNPs were electrodeposited on modified electrode surface. DNA probes antibodies were conjugated with AuNPs through Au-NH2bond,so receive GCE/PTh/PB-Au/AuNPs/DNA. ssDNAimmobilization and hybridization were characterized by Electrochemi-cal impedance spectroscopy(EIS) and cyclic voltammetry(CV). By differential pulse voltam-metry curves measured the slope of the DNA work curve is-13.442A-ml/μg, the linear range of1.0×1014-1.0×10-7μg/mL, the detection limit is1.0×10-14μg/mL, the detection limit is6.17×10-15μg/mL, the linear correlation coefficient r=0.9936. The sensor has good specificity and stability.(3) heavy metal ions on DNA damage We used prepared GCE/PTh/PB-Au/AuNPs/DNA to research Pb2+, Cr2+, Cd2+and Hg2+metal ions on DNA damage.EIS and DPV characterization results show that, the prepared GCE/PTh/PB-Au/AuNPs/dsDNA can detect the extent of damage to the DNA with high sensitivity.Some experiment conditions were investigated, such as the temperature of the detect solution and the pH of the detect solution.To optimize the experiment conditions, by differential pulse voltammetry curves measured the slope of the DNA work curve is larger, the linear range of1.0×10-13～1.0×10-7mol/L, the detection limit is lower. The DNA sensors showed good consistency. The prepared sensitive can be used to study other genetic damage.