Studies On DNA Biosensors And Molecular Machines Based On Nucleic Aptamers

This thesis studied on DNA biosensors, molecular machines and logic gates based on nucleic aptamers owing to its specific binding to the targets, which can be used to the detect of bioactive small moleculars—Adenosine triphosphate(ATP) and Adenosine monophosphate(AMP), proteins—thrombin and Adenosine Deaminase. This present work mainly studied on chemiluminescence DNA biosensors based on dual-amplification of thrombin and thiocyanuric acid-gold nanoparticle network, Logic-Based Dual-Functional DNA Tweezers with Protein and Small Molecules as Mechanical Activators and Eleetrogenerated chemilumineseence logic gate with Adenosine monophosphate AMP and AD as activators.The whole thesis can be divided into five chapters:Chapter 1 This chapter introduced the design principle and classification of DNA biosensors, and also introduced the present research of the DNA biosensor and the recent advances in the study of molecular machines. Chapter 2 The study on chemiluminescence DNA biosensors based on dual-amplification of thrombin and thiocyanuric acid-gold nanoparticle network.This thesis studied the dual-amplification efforts of thrombin and thiocyanuric acid-gold nanoparticle network, first, the thrombin labeled on the PbS NPs through the specific interaction with its aptamers; second, TCA–AuNP network introducing more AuNPs capped for each sandwich format. The CL detection of DNA was taken out with the use of Luminol-Au³⁺ system. On the optimum of Luminol-Au³⁺ system, the target DNA can be detected in the range of 2.0×10^-16M to 3.5×10^-14M, with a limit of detection (LOD) as low as 1.0×10^-16 M.Chapter 3 The study on Logic-Based Dual-Functional DNA Tweezers with Protein and Small Molecules as Mechanical Activators. The tweezers contained two symmetrical components, and six DNA strands (A to F) were hybridized to form the closed state. In strands C and D, there is a 8-base recognition region that is complementary to its aptamer. Strand E and F are labelled at the 5' and 3' ends with fluorophore and its quencher as functional components. In the closed state the fluorophore and quencher are close to each other and fluorophore can be quenched efficiently. In the strands C and D, there is a recognition region that is complementary to its input, driving the tweezers to open state and releaseing two inert double-stranded waste products. So the two strands E and F are far away from each other and the quench efficiency is low, so the CL signal increased in intensity relative to that for the closed state. When the strands C and D were added again, they complementary with strands A and B, driving the tweezer to closed state. In this process, the target moleculars were served as the fuel to achieve the cycling operation of the tweezer. The nucleic aptamers changed the traditional recognation of nucliec acid, which was used to storage and carry the genetic information. Due to their diversified structures, aptamers can achieve the similar fuction as the ligands of antibody and molecular probes. The aptamers have many potential applications, such as the detection of small moleculars without immunization and the targets which are unstable or toxic.Chapter 4 The study on Eleetrogenerated Chemilumineseence logic gate with AMP and AD as activators. Nucleic acid aptamers are important matrrials for the construction of logic gate, because their divesity and complexity. They can bind specifically with their ligands. The nucleic aptamers labelled with fluorophore and quencher were immoblized on the electrodes. When the targets were added in, the distance between fluorophore and quencher was changed, so the output signals of the logic gate also changed, The logic gate designed in this thesis was based on specific recognization of AMP to its aptamer and the deamation of AD. When AMP was added, it binded with its aptamer spcifically; the addition of AD catalytics the deamation of AMP, which changed the structure of AMP, therefor, the AMP would not bind with its aptamer. Finally, the output signal of the logic gate was detected by Eleetrogenerated chemilumineseence analysis. A further improvment was taken out by the introduction of the quencher. By this way, the backgroud noise was decreased and the sensitivity was improved efficiently. Chapter 5 was the conclusion of the whole work.