| Metal ions and small molecules have important influence on life activity and health in many vital processes. Mercuric ion (Hg2+) is a serious environmental pollutant with high toxicity and bioaccumulation. It accumulates in vital organs though food chain and damages nervous system, liver, kidney and other organs. Adenosine (AD) is a kind of small molecule drug for the treatment of cardiovasculardisease and plays an important role in the peripheral and central nervous system. The evidences suggest the expression level of adenosine is closely related to the survival and proliferation of cancer cells. So, the highly selective and sensitive detection of Hg2+ and adenosine is of great significance for the health as well as the diagnosis and treatment of diseases.Functional nucleic acids are widely used to construct the biosensing platforms due to their properties in selective combination of targets or output detectable signals. Aptamers, G-rich sequences and DNAzymes are common functional nucleic acids. Among them, T-T mismatched bases, as Hg2+ aptamer, combine with Hg2+ for the formation of T-Hg2+-T metallic base pairs. Adenosine aptamer can combine with adenosine. Both achieve the selective combination of the target. DNAzymes can catalyze the cleavage of the molecule beacons and G-quadruplex formed from G-rich sequences can bind with special dyes, which obtain enhanced detectable signals. In an effective biosensing platform, some signal amplification strategies should be introduced to ensure the high sensitivity. In view of the above analysis, target-triggered combination-based fluorescence signal amplification biosensing platforms are built for highly selective and sensitive detection of Hg2+ and adenosine. The main researches included in the dissertation are presented as following: The first chapter is the introduction to overview the detection significance and traditional detection methods of Hg2+ and AD as well as the role of the functional nucleic acids in biosensing platforms. The common enzyme-assisted signal amplification strategies are summarized, including endonuclease-assisted signal amplification strategies, exonuclease-assisted signal amplification strategies, polymerase-assisted signal amplification strategies and polymerase/nickase-assisted signal amplification strategies.In the second chapter, T-T mismatched bases and Hg2+ combination-triggered fluorescence signal amplification biosensing platform based on Exo Ⅲ-assisted target cycle and DNAzyme cycle is developed for detection of Hg2+. T-T mismatched bases combine with Hg2+ to ensure the highly selective combination of target. The Exo Ⅲ-assisted target cycle and DNAzyme cycle-based fluorescence signal amplification triggered by combination achieves the sensitive detection of Hg2+. The linear range of the method is from 2.0×10-10 mol·L-1 to 1.0×10-8 mol·L-1, with the detection limit of 7.2×10-11 mol·L-1. The detection platform only responses to Hg2+ among many metal ions. Addition, the detection paltform can quantify the spiked Hg2+ in tap water, river water and lake water, and even Hg2+ in waste water.In chapter three, a label-free fluorescence signal amplification biosensing platform based on target-triggered proximity combination is developed for the sensitive detection of AD. Difunctional probe, containing two split adenosine aptamers and the template of polymerase/nicking reaction, is designed for selective combination of adenosine and initiates the stranded displacement amplification. Compared with the split aptamer in two probes, the local concentration of the aptamer in the designed probe is larger and the aptamer is easier to combine with targets for efficient combination. Through the stranded displacement amplification and rolling circle amplification strategies, large amount of G-quadruplex sequences are inserted the ThT dyes for the label-free fluorescence signal. And the linear range of the method is from 5.0×10-7 mol·L-1 to 2.0×10-5 mol·L-1, with the detection limit of 8.4 ×10-8 mol·L-1, which meets the requirements for the detection of adenosine in human urine samples. This detection platform has potential applications in the practical sample detection. |
