Study Of Novel Colorimetric Methods Based On Mg²⁺-dependent DNAzyme

Colorimetric analysis has the advantages of low price,simple operation and easy signal readout,making it widely used in many fields such as life sciences,medicine,health and environmental protection.However,how to improve its detection performance is still a major challenge for researchers.DNAzyme is a kind of functional nucleic acid that displays catalytic activity.The high catalytic capability and stability of DNAzyme make it ideal signal-amplifier for colorimetric analysis.In this dissertation,we report two kinds of colorimetric methods based on Mg2+-dependent DNAzyme which are fast,accurate,sensitive and efficient.The details are as follows:1.A novel colorimetric method based on Mg2+-dependent DNAzyme to detect proteins.In this section of the dissertation,we propose a simple,sensitive and cost-effective colorimetric method for protein assay that is based on the Mg2+-dependent DNAzyme and proximity ligation conjugate.Specifically,in the presence of target protein,the DNAzyme will be assembled through two proximity probes.Then,the activated DNAzyme catalyzes cleavage of its substrates with multiple cycles,leading to the cross-linking failure between two types of DNA modified gold nanoparticles.The color of solution is still red.However,in the absence of target,the detached DNAzyme cannot cleave the substrates,resulting in a concomitant red-to-purple color change of the test solution.In this work,the disease-related platelet derived growth factor BB(PDGF-BB)has been used to demonstrate the strategy,and a low limit of detection(LOD)of 1 pg/ml as well as a wide linear range from 1 pg/ml to 10 ng/ml can be achieved.Also of note,this method also has the advantages of low price,simple operation,good stability and easy signal readout,making it realizable to use in some resource-limited area in the future.2.A novel colorimetric method based on Mg2+-dependent DNAzyme to detect small molecules.In this section of the dissertation,we propose a novel colorimetric method based on Mg2+-dependent DNAzyme to detect small molecules by applying the idea of"splitting" to aptamer,and the adenosine triphosphate(ATP)has been used to demonstrate the strategy.We split the single aptamer of ATP into two fragments and combine them with DNAzyme.In the presence of ATP,the DNAzyme will be assembled via the synchronous recognition of one ATP with two fragments of aptamer.Then,the activated DNAzyme catalyzes cleavage of its substrates with multiple cycles,leading to the cross-linking failure between two types of DNA modified gold nanoparticles.The color of solution is still red.However,in the absence of ATP,the detached aptamer cannot induce the assembly of DNAzyme to cleave linkers,resulting in the aggregation of gold nanoparticles and a concomitant red-to-purple color change of the test solution.This method may have a linear detection range from 10 pM to 1 nM with a detection limit of 10 pM for ATP.In addition,the proposed method might provide a new way to the colorimetric detection of small molecules.