| Nucleic acid is the carrier of inheritance information, which plays a significantrole in the body of organisms and also has a wide application in vitro. Aptamer isalways a strand of DNA or RNA molecular, which is obtained though systematicevolution of ligands by exponential enrichment (SELEX). As a rule, aptamer couldbond its targets including amino acid, protein, even a whole cell with high affinityand specifity. Therefore, it has capacious application prospects in bioanalytical assay,environment monitoring, basic medicine, new drug synthesis or selection. This paperaims to aptamer technology as the foundation and builts biosensors throughcombining smallmolecular probes self-assembly and exponential polymeraseamplification reaction. Through our methods, trace detection of lead and silver ionswere carried out and analysis efforts were fine. The detailed points of the thesis aredescribed as follows:Thrombin aptamer (TBA) and pyrene probe (one positive charge) were selectedas sensing elements in the first section. The nucleic acids could induce pyreneaggregating due to polyanion properties of its phosphoric acid framework. Strongpyrene excimer emission was tested because of pyrene aggregation. Meanwhile, TBAis a G-rich oligonucleotide. A appropriate G-rich oligonucleotide folded into stablequadruplex structure when mixed with lead in an aqueous solution, which can bemarkedly resistant to the digestion of single-stranded nucleic acid specific nucleaseS1. Subsequently, pyrene probe was added into the solution. A G-quadruplex that wasnot digested by nuclease S1could induce the aggregation of pyrene probes.Consequently, the excimer emission was checked out and the fluorescence intensitywas propotional with Pb2+added. A simple and sensitive method for sensing Pb2+wasestablished.In the second part, a cytosine-rich oligonucleotide oligo-C and a cationic perylene derivative constitute a biosensor for testing Ag+. In the presence of Ag+, oneAg+ion could interact specifically with two cytosine nucleic acid bases by covalentbond, which made oligo-C fold into a stable hairpin structure by gradually cooling toroom temperature. Afterwards, perylene probes were added into the mixture. Theelectrostatic interactions were weakened between oligo-C and perylene probes due tooligo-C rebounding into hairpin structure. As a result, the free monomer of peryleneprobes was released into the solution and a fluorescence intensity increase wasobserved. This method obtained a limit of detection of5nM.In the third segment, a lead reliant DNAzyme called GR-5was utilized for thisstudy. The GR-5DNAzyme can catalyze cleavage of an RNA base embedded DNAsubstrate in the presence of Pb2+. Two short oilgonucleotides were produced.However, the5′-part of the cleaved substrate contains a2′,3′-cyclic phosphate at its3’-part, which prevents its direct use as a primer for the subsequent stranddisplacement amplification reaction. Exonuclease III could successfully remove the2′,3′-cyclic phosphate, and the3′-hydroxyl was exposed during the experiment. Andthen, it can be used as a primer for strand displacement amplification reaction. Thenucleic acid exponential amplification reaction was used as amplification manner,which was effective, time-saving and highly special. Finally, molecular beacon couldincorporate with the ampification products through bases complementing andincreased fluorescence intensity was checked out. This method shows well seletive,high sensitivity and low background, which provides a novel idea for building upbiosensors. |
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