Research On A New Method For Detecting Copper Ion In Functional Nucleic Acid Probe

With the development of industrial society,industrial pollution is becoming more and more serious,especially in the treatment of heavy metals there are many defects.The arbitrary discharge of industrial waste water and the arbitrary use of chemical pesticides can cause serious pollution to the environment.In the natural environment,heavy metal pollution is becoming increasingly serious,especially copper ion.The harm caused by it greatly affects the ecological balance,and excessive copper ions will bring many safety risks in daily life.Therefore,the quantitative detection of copper ions has important research significance,which can greatly reduce the probability of excessive intake of copper ions.In this paper,two methods for the detection of copper ions are developed based on nucleic acid functional probe:（1）Establishment of an immune lateral chromatography method for copper ion detection based on functional nucleic acid DNAzyme.The establishment of this method is based on the basic principle of DNAzyme specific recognition of Cu²⁺combined with PCR amplification technology.After recognizing and binding Cu²⁺,DNAzyme’s cutting activity is activated,causing the substrate chain of DNAzyme to break and fall.The cleaved fragments are enriched by magnetic adsorption and used as the template for PCR amplification for exponential amplification to increase the concentration of cleaved fragments and improve the sensitivity of the experimental method.The amplified products are labeled with biotin and FITC.The C and T lines on the strip could identify and firmly capture the corresponding antigen and complex by the specific binding between streptavidin and biotin and the reaction between antigen and antibody,so as to realize the visual detection of the target.The detection limit of Cu²⁺is0.024 n M,and the linear range is 0.01 n M～1.5 n M（R²=0.99019）.（2）Establishment of a copper ion detection method based on functional nucleic acid DNAzyme and palindrome sequence assembly technology.The establishment of this method is based on the basic principle of DNAzyme’s specific recognition of Cu²⁺combined with nucleic acid hybridization.The probe is designed according to the experimental requirements.Based on the uniqueness of palindrome sequence,the long nucleic acid chain is assembled by the complementary pairing between bases.After recognizing and binding Cu²⁺,the cutting activity is activated and the substrate chain is cleaved.Since the substrate chain and enzyme chain of DNAzyme are labeled with fluorescence group FAM and quenched group Dabcyl,when the cleaved fragments of the substrate chain fall off,the fluorescence will recover in the reaction system,and the quantitative detection of the target substance is realized.The detection limit of Cu²⁺is 0.67 nM,and the linear range is 0.1 nM～100 nM（R²=0.99779）.