Research And Application Of Biosensor Based On DNA Strand Hybridization Reaction

DNA strand hybridization reaction is widely used in medical diagnostics,genetic testing,and environmental monitoring in low cost,real-time,and high sensitivity.Two methods of DNA strand hybridization between strand displacement reaction and hybridization chain reaction are often used to detect cells,DNA,proteins,and metal ions.The strand displacement reaction refers to the process of stripping the old strand by DNA strand hybridization,which has been used for various in vitro isothermal amplification strategies,such as rolling circle amplification,loop-mediated isothermal amplification,and the like.The hybridization chain reaction is a modern signal amplification mechanism.It uses the target to trigger the opening of multiple hairpin probes,so that the signal is amplified,the whole process is not used with enzymes,and the reaction conditions are mild and easy to control.Based on DNA strand hybridization reaction,three different biosensors were developed in this paper to determine HCV DNA,HPV DNA and H7N9 DNA,which are summarized as follows:?1?Based on strand displacement reaction and hybridization chain reaction,enzyme-free label-free fluorescence detection HCV DNA.After the addition of HCV DNA,HCV DNA hybridizes with HP1,triggering a chain displacement reaction between HP1 and HP2 to form a HP1-HP2 complex.After the addition of HP3 and HP4,the HP1-HP2 complex can initiate hybridization of HP3 and HP4,forming a long double-stranded structure of DNA.The added SG I binds to the long double strand of DNA and emits a strong fluorescent signal.The experimental results show that the fluorescence intensity difference value of the system and the concentration of H7N9 DNA is linear in the range of 0.1-10 nM,and the detection limit is 82 pM.?2?Target-triggering multiple-cycle amplification strategy for the visual and fluorometric determination of HPV DNA.After the addition of HPV DNA,HP1 and HP2 can be opened to undergo chain displacement reaction to form a long double-stranded structure,and Nt.BbvCI is added,resulting in a number of G-base-rich DNA strands.After the addition of ThT,G-quadruplexs can be formed to enhance fluorescence.The HPV DNA can be detected by fluorescence method.When hemin is added,the G-quadruplex linked with hemin can catalyze the oxidation of ABTS^2-for color development,so HPV DNA can be detected efficiently and sensitively by multiple methods.The linear range of fluorescence detection is 0.05-5nM,the detection limit is 32.9 pM;The linear range of UV detection is 0.4-15 nM,the detection limit is 0.284 nM;And the concentration of HPV DNA is 0.4-15 nM,the color change of the experimental system is gradually deepened,and the experimental results can be directly observed by visual colorimetry.There is also good linearity in this concentration range in human serum samples.?3?Detection of H7N9 DNA based on exonuclease ?-assisted target loop-free label-free fluorescence assay.After the addition of H7N9 DNA,the target is released,recycled,and a large amount of G-base-rich DNA is produced by the shearing property of Exo ?.When combined with the fluorescent dye ThT,G-quadruplex is formed,and the system fluorescence is enhanced.The experimental results show that the fluorescence intensity difference value of the system and the H7N9 DNA concentration is in the range of 0.05-25 nM,and the detection limit is 17 pM.