Construction And Application Of Sensors Based On DNA Hydrogel

Deoxyribonucleic acid(DNA)is not only the genetic biomolecule in living systems,but also a promising building-block for constructing materials.Various functional DNA structures promote the construction of smart responsive materials.DNA has attracted particular research interest due to their unique advantages such as precise programming,specific molecular recognition,excellent biocompatibility,easy synthesis and modification,biological functions and so on.Taking advantage of the base pairing principle,DNA has been utilized for constructing artificial structures from the nanoscale to the bulk regime.DNA hydrogels are typical macroscale materials by DNA assembly,showing great application in the fields of sensing,medical diagnosis and treatment,tissue engineering,and so on.Given that a large amount of hydrogel and cargoes encapsulated in hydrogels are needed during the detection process,it is urgent to fabricate simple,sensitive,cost-effective,and portable sensors based on DNA hydrogel for point-of-care testing(POCT).In addition,using biotechnology to prepare large amounts of DNA is an efficient way to construct DNA hydrogels,which paves the way for constructing simple and cost-effective sensors.In this paper,we developed visual sensors for detecting lead ions(Pb2+)and uracil-DNA glycosylase(UDG)using the pore size-based separation performance of DNA hydrogel by combing DNA hydrogels with capillary tubes.In addition,we constructed a fluorescence imaging method for aflatoxin B1(AFB1)detection based on DNA hydrogel microspheres which prepared by rolling circle amplification(RCA)technology.The main work includes three parts are listed as follows:(1)We developed a DNA-based hydrogel capillary sensor for sensitive detection of Pb2+,taking advantage of the target-responsive hydrogel and capillary action of the capillary tube.First,we prepared the polyacrylamide-DNA hydrogel,and the Pb2+-dependent DNAzyme played the roles of response unit and crosslinker in the hydrogel.Based on the denaturation and renaturation characters of DNA,the hydrogel transformed into solution after being heated and flowed into the tube,and then plugged the end of the capillary tube.After the hydrogel end horizontally being immersed in the reaction solution,the target Pb2+ would activate the DNAzyme,and the crosslinker substrate could be cleaved,leading to that the mesh size of the hydrogel increases and the solution could flow into the tube in shorter time.Only by recording the distance and incubated time,the concentration of Pb2+ could be quantitatively detected by the naked eye.The tiny hydrogel without extra complicated encapsulating process was used to construct the sensor.This method has the advantages of cost-effective,simple,portable,universal,and highly sensitive,which shows good prospects in food safety,environmental monitoring,molecular diagnostics,and so on.(2)The mesh size of the hydrogel could be affected by the concentration of the crosslinker,which would influence the membrane filtration of the hydrogel.Based on this,we constructed a DNA hydrogel-based UDG sensor using a capillary tube.Prior to the hydrogel construction,UDG first reacted with the DNA substrate in the liquid system.Compared with the untreated system,the DNA substrate treated by UDG was cleaved,resulting in the occurrence of some voids and increase in mesh size of the hydrogel.Taking advantage of the membrane filtration technique which is closely related to the mesh size of the hydrogel,quantitative detection of UDG was achieved by reading the distance of the solution in the capillary tube.This method solves the problem of low activity of the enzyme for the substrate of the hydrogel and realizes the visual detection of biological macromolecules based on DNA hydrogel.This method exhibits significant advantages of simple,highly sensitive,portable,and universal,which holds great potential for POCT,clinical diagnosis,and drug screening.(3)Based on DNA hydrogel microspheres prepared by isothermal RCA technology,we proposed a sensor for the detection of AFB1 by using the fluorescence imaging method.The fabrication process of the DNA hydrogel microspheres by RCA technology is very simple,and the microstructure is in the shape of a bird nest with large surface and many reaction sites.The DNA hydrogel microspheres were used as the reaction carriers,and immobilized the fluorescent-modified aptamers through DNA hybridization.In the presence of AFB1,the aptamer could specifically bind to AFB1 and fall off from the surface of the microsphere.With the concentration of AFB1 increases,the fluorescence intensity of the microspheres gradually decreases.By measuring the fluorescence intensity of the microspheres,the concentration of AFB1 could be quantitatively detected.This method may be generally applicable for sensing various targets by incorporation of the appropriate target-related nucleotide sequence in the hydrogel.This method only needs a small amount of sample to mix with the microsphere solution,and the operation is extremely simple and time-effective.On the basis of this,this sensitive sensor could be applied for the detection of small amounts of samples,and has great potential for food safety,environmental monitoring,and medical research.