The Study Based On G4 Nucleic Acid Probe Rapid Detection Of Malachite Green

Malachite green has potential carcinogenic,teratogenic and mutagenic hazards.The discharge of industrial wastewater and the illegal addition of malachite green make it residue in aquaculture water and aquatic products,causing public concern about the quality and safety of aquatic products and environmental pollution.The traditional methods of liquid chromatography and liquid chromatography-mass spectrometry for the determination of malachite green have problems such as high cost,high operational requirements and complicated pre-treatment.New biosensors are of interest because of their convenience,speed and low cost.Studies have shown that malachite green has a good affinity with some G-quadruplexes(G4).Herein,three biosensors based on G4 nucleic acid probes were constructed for the detection of malachite green.The results show that the above sensors have the characteristics of simplicity,rapidity,low cost,high sensitivity and high specificity.The specific research contents and related results of this paper are as follows:1.Constructing G4-ThT fluorescent with G4 for rapid detection of malachite green.The functional structure of G4 that can recognizes malachite green was used to combine with the water-soluble fluorescent dye thiosphonin T(ThT)to construct a fluorescent probe G4-ThT for the rapid detection of malachite green.Based on the internal filtration effect(IFE),malachite green can quench the fluorescence of fluorescent probes.The experimental results showed that the sensing system had a good linear relationship in the range of 50-1000 μg/kg and 400-6000 μg/kg,the detection limit was 6.82 μg/kg,which has good specificity and can be applied to the detection of malachite green in temporary pond water,prawns and small yellow croakers.The recovery rate of spikes in the temporary pond water was 74.34%99.90%,in prawns was 77.00%-86.17%and in small yellow croakers was 101.14%106.11%.The DNA strands in the constructed fluorescent probe G4-ThT do not require any modification,the cost is low,and the detection of malachite green by G4-ThT fluorescent probe only requires one reaction.it can be combined with a microplate reader for highthroughput and rapid batch detection.the detection time can be controlled within 15 min.2.Using Zr-MOFs to enhance the fluorescence of G4-ThT fluorescent probes for rapid detection of malachite green.In order to further improve the detection sensitivity of G4-ThT fluorescent probes,a zirconium-metal-organic framework(ZrMOFs)compound that can be stably present in water was prepared,mixed with G4-ThT fluorescent probes,and selfassemble to form a composite probe G4-ThT@ZrMOFs for rapid and sensitive detection of malachite green.The experimental results show that in this composite probe,ZrMOFs can not only further stabilize the ThT structure,reduce molecular movement,enhance fluorescence,but also efficiently adsorb and enrich malachite green in water,the detection limit was 0.5 μg/kg,linear detection range was 0.5-100 μg/kg,and can be effectively applied to the detection of holding pond water and river water.The recovery rate of this method for the spiked pond water was 85.00%-90.36%,the river water was 79.30%-104.00%,all of which had low relative standard deviations.After combining ZrMOFs,the detection can still be completed within 15 minutes.3.Construction of an electrochemical biosensor for the sensitive detection of malachite green using DNA tetrahedra and G4 strands.The application of G4 in malachite green detection method is expanded,and the influence of matrix on the detection signal is reduced.The sulfhydryl modified DNA strands are self-assembled into DNA tetrahedra,bound to the electrode by Au-S bonds,and then the G4 nucleic acid probe was attache to the tetrahedra by base complementation.The addition of DNA tetrahedra further improves the sensitivity of electrochemical detection by constructing a reasonable probe density.Through the optimization of the enrichment time,the optimal enrichment time is selected to achieve rapid detection of malachite green.The experimental results show that the electrochemical sensor exhibits a good linear relationship from 5×10-8 to 1×10-4 mol/L.the detection limit of the electrochemical sensor was 3.65 μg/kg,and it also has good detection performance in the holding pond water...