Reaserch And Application Of Electrochemical DNA Sensor Based On Graphene

Due to the high sensitivity, good selectivity and low cost for the detection of specific single-stranded DNA (ssDNA) sequences with very small volume samples, electrochemical DNA sensor can be used in many fields including medical diagnosis, biological engineering, environmental protection and so on. Graphene (GR), which is a two-dimensional and single layer of carbon atom sheet, has many important applications in electrochemical sensors and biosensors due to its specific structure and properties such as wide potential windows, high stability, large surface area, excellent electrocatalytic activities, high conductivity and biocompatible microenviroment. This thesis mainly includes the following parts:(1) A TiO2nanorod and GR composite was prepared, which was cast on the surface of carbon ionic liquid electrode (CILE) with chitosan as film forming material. The modified electrodes were characterized by scanning electron microscopy, cyclic voltammetry and electrochemical impedance spectroscopy. The ssDNA probe sequences were electrostatic adsorbed on the electrode surface to get the DNA sensor, which could be hybridized with the target ssDNA sequence. Differential pulse voltammetric responses of the electrochemical indicator methylene blue (MB) on different DNA modified electrode were recorded to monitor the hybridization reaction. The polymerase chain reaction (PCR) products of MON89788gene sequences from soybean samples were successfully detected by the proposed method.(2) An electrochemical DNA biosensor was fabricated by using electrochemical reduced graphene oxide modified CILE as the working electrode and MB as the hybridization indicator. The probe ssDNA sequences were immobilized on the modified electrode surface through electrostatic adsorption. After hybridization with target ssDNA sequence, differential pulse voltammetric responses of MB on different DNA modified electrode were recorded for monitoring the hybridization reaction. The PCR products of MON810gene sequences from transgenic maize were successfully detected by the proposed method.(3) A partially reduced graphene oxide modified CILE was fabricated by the controllable electrochemical reduction method, which was used as the platform for the immobilization of probe ssDNA quence. NH2modified ssDNA probe sequences were immobilized on the electrode surface via the covalent bond between the unreduced oxygen groups on the modified electrode surface and the amine group of ssDNA at the5’end. The prepared ssDNA modified electrode was furthur hybridized with the target ssDNA sequence with MB adopted as the electrochemical hybridization indicator. The fabricated DNA sensor was applied to detect the A2704-12gene sequence and the PCR products of A2704-12gene sequences from transgenic soybean were successfully detected by the proposed method.(4) A carboxyl functionalized graphene oxide was synthetized and cast on the surface of electropolymerized ploy-L-lysine modified glassy carbon electrode to give the modified electrode. The NH2modified probe ssDNA sequences were immobilized on the modified electrode surface by the formation of amide bonds to get the DNA sensor, which was stable and furthur hybridized with the target ssDNA sequence. Differential pulse voltammetry was used to monitor the hybridization events with MB as electrochemical indicator. And the current responses of MB on different DNA modified electrode was recorded for the detection of tlh gene sequence related to the vibrio parahaemolyticus. The PCR products of tlh gene from oyster samples were detected with satisfactory resultsIn summary, four kinds of electrochemical DNA sensors were constructed in this thesis, which were applied to detect specific target ssDNA sequences with high sensitivity, good selectivity, and the ability to distinguish the complementary, mismatched and non-complementary ssDNA sequence.