1 Introduction
deoxyribonucleic Acid (DNA) is one of the four biological macromolecules contained in biological cells. Its function is to encode, store and transmit genetic information
[1]. In 1982, Seeman successfully used the complementary base pairing mechanism of DNA to construct cross-shaped nanostructures, which marked the beginning of DNA as a nanomaterial
[2]. Among them, DNA hydrogel materials constructed by DNA as a key functional molecule have attracted wide attention of researchers. DNA hydrogel is a kind of intelligent hydrogel which combines the three-dimensional grid structure of hydrogel and the excellent characteristics of DNA molecules, and can obtain specific functions by accurately designing the sequence of DNA. These novel nanomaterials with excellent properties have been widely used in many fields, such as intelligent devices, nano-machinery, medicine and medical fields
[3][4,5][6⇓⇓⇓~10][11,12].
Compared with traditional hydrogels, DNA hydrogels have biological properties (specific recognition of targets and aptamers, DNA can be cut and spliced by enzymes, etc.). For example, DNA ligases can catalyze the formation of phosphodiester bonds by using restriction nucleases to cleave phosphodiester bonds between nucleic acid monomers, linking DNA strands together via the phosphodiester bond
[13⇓~15]. At present, many enzyme-assisted or enzyme-stimulus-responsive DNA hydrogels have been synthesized, and the integration of specific enzyme sites in hydrogels can directly help to achieve the modification, splicing, splicing and functionalization of DNA hydrogels
[16]. Not only enzymes can stimulate DNA hydrogels, but also some metal ions can produce the same effect, which greatly expands the stimulus-response mode of DNA hydrogels and makes them show great potential in the field of sensing
[17]. Therefore, more and more researchers began to study the structural modification of DNA hydrogels, and through the precise design of DNA chains, many intelligent hydrogel materials with excellent properties were designed
[18⇓⇓⇓⇓⇓⇓⇓⇓⇓⇓⇓⇓⇓⇓⇓⇓~35]. The appearance of DNA hydrogel fills the gap of nano-materials in the field of sensing application.In this paper, the synthesis methods, sensing mechanism and application examples of DNA hydrogel are reviewed from the perspective of sensing applications, and its development in the field of sensing is prospected.
2 Synthesis of DNA Hydrogel
The hydrogel with DNA as the structural or functional unit is called DNA hydrogel. Matsuda and Nagahara were the first to synthesize DNA hydrogels
[36]. They used DNA as a crosslinking molecule in water-soluble copolymers to prepare swellable hydrosols by taking advantage of the solubility of DNA in water. Complementary single-stranded DNA (ssDNA) can hybridize spontaneously at room temperature, so bioactive molecules and other substances can be immobilized in hydrogels at room temperature without chemical reaction during the formation of hydrogels. Since then, scholars have found that DNA has great advantages in the construction of functional hydrogels. In 2006, Luo et al. Extracted the first pure DNA hydrogels from branched DNA nanostructures
[37]. The formation of 3D network in DNA hydrogels mainly depends on three ways: the entanglement of linear DNA strands, the self-assembly of dendritic DNA and the hybridization of other materials.
2.1 Winding of linear DNA strands
DNA hydrogels can be produced by the extension of DNA. In 2006, Luo et al. Spliced and extended DNA strands using polymerase Φ29
[38]. The hydrogel can lengthen the primer DNA by cyclically replicating the circular DNA and at the same time continuously displace the synthetic DNA fragments from the template to lengthen the DNA strand, a method known as rolling circle amplification (RCA). The DNA hydrogel has different mechanical properties compared to other hydrogels: it is solid in water and becomes liquid in the absence of water. Moreover, this deformation is reversible. The work of Luo et al. opens up a new way for the synthesis of DNA hydrogels using RCA technology. Using the RCA technique, DNA hydrogels can be prepared by amplifying functional fragments into the RCA-generated DNA strands. Tian et al. amplified the i-motif structure as a repetitive sequence into the DNA strand produced by RCA, and prepared a stimulus-responsive DNA hydrogel
[39]. In 2017, N Nöll et al. Prepared DNA hydrogels from plasmid DNA, which can be prepared in large quantities by fermentation
[40]. Plasmid DNA is enzymatically digested to obtain linear DNA constructs with complementary "sticky" end genes, which are then ligated into linear DNA strands and entangled into the gel by a ligase mediated catalytic reaction.
2.2 Self-assembly of Dendritic DNA
Most DNA hydrogels are assembled from branched DNA building blocks with "sticky" end groups that can adhere to each other to form a 3D network. In 2017, Wang et al. Developed a DNA hydrogel preparation technology for hybrid chain reaction
[41]. As shown in fig. 1, the hairpin DNA strand H1 can hybridize at the 5 'end (segment d) through a loopback structure to form a hairpin dimer (H1-dimer). The I strand first binds to the toe region of H1, and then the stem and loop are opened by the DNA strand displacement process. Finally, H1 opens the stem and loop of H2 in the same way, and then H2 opens the stem and loop of another H1. An H1-dimer has two functional branches, so H1 can form a three-arm or four-arm structure, which forms a 3D network for the subsequent hybridization chain reaction, which is called the clamping hybridization chain reaction. The initiator is previously set to a desired pattern by a linear hybrid chain reaction, and then the hydrogel starts to expand from the DNA chain on which the initiator is immobilized, forming a two-dimensional DNA hydrogel pattern without external restriction. The method for constructing the DNA hydrogel is simple and can be used for constructing the DNA hydrogel with specific composition and shape.
2.3 Hybridization of DNA hydrogel
Functional groups or inorganic nanomaterials are used to compound and improve DNA hydrogels to obtain multi-functional hybrid hydrogels, which can make up for the defects of traditional DNA hydrogels in response speed, temperature performance and other aspects, and can obtain more characteristics. Shin et al. Synthesized TNA hybrid hydrogel (TA + DNA) by integrating tannic acid (TA) into DNA hydrogel
[42]. The reversible linkage of phosphodiester bonds makes TA a glue between molecules (molecular glue). The hybrid DNA hydrogel can be adhered to human tissues, and has excellent hemostatic effect in vivo due to rich polyphenol groups. Willner et al. synthesized a dynamic polymer-DNA hybrid hydrogel by incorporating a stimulus-responsive DNA structure into an acrylamide polymer
[43]. DNA chains can self-assemble into i-motif structures under acidic conditions and dissolve under alkaline conditions, which enables DNA chains to achieve the transition between sol and gel. Singh et al. Reported a DNA-carbon dot hybrid hydrogel for targeted drug delivery in vivo
[44]. Eguchi et al. Synthesized a hybrid DNA hydrogel using gold nanoparticles and polymerase chain reaction
[45]. In this hydrogel, DNA participates in protein expression and gold nanoparticles act as crosslinks. In 2014, Zhou et al synthesized an intelligent mesoporous silica nanoparticle-DNA/polymer hybrid hydrogel, which can encapsulate multiple aptamers at the same time
[46]. In 2018, Geng et al. Made a multi-functional hybrid DNA hydrogel by combining ultra-long linear DNA chains with silver nanoclusters (AgNCs)
[47]. AgNCs enable that hydrogel to have both fluorescent and antibacterial function. Compared with pure DNA hydrogels, hybrid DNA hydrogels have better performance and relatively lower cost, so DNA hybrid hydrogels are increasingly favored by material scientists.
3 Biosensing mechanism
The "swelling behavior" (absorption and retention of a certain amount of water) and the chemical and physical properties of DNA hydrogels are important properties required for sensor materials. DNA hydrogels have high water content, porosity, consistency and excellent biocompatibility, which make them widely used in the field of sensing. The most important characteristic of DNA hydrogels is their response to external stimuli. By inserting appropriate functional groups into the polymer backbone, hydrogels can be made to respond to specific chemical, physical or biological stimuli. Mesh size, the degree of crosslinking, and even the ionic charge of the backbone are all elements that can affect the stimulus response of hydrogels.
In 2012, Buenger et al. Proposed many important physicochemical mechanisms for the application of hydrogels in the field of sensing
[48]. Hydrogels can respond to a variety of environmental stimuli, such as changes in network structure, optical, mechanical, and electrical properties. These external environments can be temperature changes, light stimuli, certain biological molecules, or free antigens
[49]. Among them, the response of the hydrogel to a temperature stimulus is as follows: the temperature at which the hydrogel responds is referred to as a critical temperature (T
c), and if the polymer and the solvent are phase-separated above the critical temperature, the critical temperature is referred to as a low critical solution temperature (LCST), and if not, it is called a high critical solution temperature (UCST)
[50]. Similarly, pH-responsive hydrogels have the phenomenon that in an anionic network, if ionization occurs at a pH of the environment above the characteristic pK
a of the acidic groups, the response of the hydrogel corresponds to a network with higher hydrophilicity, a higher number of charge immobilizations, and a stronger repulsion between DNA strands. In contrast, the network has higher hydrophobicity and higher compactness.
3.1 Embedment encapsulation method
3.1.1 Enzyme-based embedment release
Enzymes are highly specific molecules that act like biocatalysts to accelerate chemical reactions and reduce activation energy
[51]. In 1912, Michaelis and Menten proposed the model of enzyme action, which divided the enzyme reaction into the formation of enzyme-substrate complex in the transitional state of reaction and the release of chemical reaction products and the reduction of enzyme
[52]. They emphasized that the rate of enzyme catalysis depends on the concentration of the enzyme and the substrate, that when the concentration of the substrate is high enough, the enzyme will be saturated, and that the rate of reaction depends only on the conversion from substrate to product. Many DNA hydrogel sensors use enzyme-substrate interactions to detect analytes. Compared with other immobilization technologies, DNA hydrogels have a large number of hydroxyl groups in the matrix structure, which can provide a good microenvironment for enzymes to maintain their natural structure, so DNA hydrogels have higher sensitivity as enzyme encapsulation materials.
3.1.2 Antigen-antibody based entrapment release
The basic principle of antigen-antibody interaction is that an antibody (or antigen) that recognizes an antigen (or antibody) on an analyte produces a signal whose intensity is proportional to the concentration of the analyte. Antibodies (or antigens) may be embedded in the DNA hydrogel matrix, they may be bound in a network, or a copolymerization reaction may occur between the antigen-antibody binding fragment and the DNA hydrogel. Ishii-Mizuno et al. Constructed a physical gel that could induce higher levels of antigen-ovalbumin-specific IgG by mixing chitosan with sDNA hydrogel
[53].
3.1.3 Ncapsulation release based on nanomaterial
By embedding nanomaterials in the hydrogel, the DNA hydrogel gradually disintegrates under the stimulation of target molecules, releasing nanomaterials as a signal source. This new detection method does not need special instruments to amplify the signal or improve the accuracy, which improves the portability of the detection and reduces the cost of the detection. In 2017, Mao et al. Proposed a portable visual detection method based on functional DNA hydrogels for different targets, using DNA hydrogels as target response units and gold nanorods (AuNRs) as multi-color signal readout circuits
[54]. The color change of AuNRs is related to the concentration of the target, and different molecules can be detected after adjusting the whole system. Lin et al. Synthesized a DNA hydrogel in 2019
[55]. As shown in fig. 2, firstly, Au @ HKUST-1 is obtained by uniformly coating gold nanoparticles on a copper-based metal organic framework with oxidase activity,Acrylate-modified adenosine aptamer (Acry-A-Apt) solution and single-stranded DNA (Acry-ssDNA) solution were mixed with Au @ HKUST-1 solution in Tris buffer and incubated twice at 50 ℃, then hemin aptamer was added to the solution and incubated three times at 50 ℃. And finally cool to room temperature to obtain that adenosine reactive DNA hydrogel. Au @ HKUST-1 was encapsulated in the crosslinked body of acrylate-modified adenosine aptamer and acrylate-modified single-stranded DNA. When adenosine was contacted with this hydrogel, the G-quadruplex structure of the hydrogel was disrupted and the preferential interaction between adenosine and adenosine aptamer resulted in complete dissolution of the DNA hydrogel. The simultaneous release of G-quadruplex/heme and Au @ HKUST-1 achieves double amplification of the signal, and the DNA hydrogel biosensor can detect adenosine down to the 1.04×10
-4nmol·L
-1.
3.2 Non-embedding encapsulation method
The three-dimensional mesh structure of the hydrogel is used for embedding signal molecules, so that the precise measurement of the target can be realized, and the portability is improved. However, most of these methods require the complete release of the embedding material to produce a measurement signal, which requires many target molecules to stimulate the hydrogel material. The introduction of optical signals gets rid of the dilemma that a large number of target molecules are needed to cleave DNA hydrogels, but such methods require large equipment to detect and analyze optical signals, which is not portable. The hydrogel can be gradually disintegrated under the action of the target, does not depend on the complete degradation of the DNA hydrogel, and can be used for detecting various target molecules in a portable mode by detecting signals generated in the disintegration process. Bai et al. developed a bioimprinted DNA hydrogel that uses volume change as an output signal
[56]. Two different methacrylamide-modified aptamers were bound to the target to produce a complex, which was inserted into the hydrogel by copolymerization. The separation of the aptamer and the protein causes the hydrogel to swell, thereby increasing the length of the hydrogel. The re-added protein binds to the aptamer, causing the length of the hydrogel to decrease. In 2005, Okumura et al. Developed a platform for diagnosing multiple targets using a microfluidic paper analyzer (μPAD) integrated with DNA
[57]. The platform can diagnose a variety of targets such as adenosine, cocaine and Pd
2+. Different DNA hydrogels are constructed according to different detection targets, and the DNA hydrogel structure is cracked by combining the targets, so that the solution with the colored dye molecules begins to flow. In 2019, Li et al. Built a platform for quantitative detection of cocaine by changing the permeability of hydrogel membranes
[58]. When cocaine is present in the solution, the internal structure of the DNA hydrogel film will be destroyed by cocaine molecules, resulting in an increase in the permeability of the film and a faster flow rate of the solvent in the capillary. The higher the concentration of cocaine molecules, the faster the flow rate.
4 Application of DNA hydrogel biosensor
When the DNA cross-linking structure in the hydrogel network is triggered by the target, the gel network structure will disintegrate and release the previously encapsulated signal substance, thus generating a signal that can be captured. In 2008, Tan et al. First demonstrated that the transformation of responsive DNA hydrogels from gels to sols is feasible
[59]. They used the released gold nanoparticles as a signal to detect adenosine. The good capture ability makes DNA hydrogels a good platform for embedding and encapsulating catalyst substances. When the catalyst is released, they can further catalyze other substances to produce larger and more obvious signals. Many scholars use DNA hydrogels to encapsulate enzymes, DNase or catalytic metal nanoparticles as catalysts, and develop various colorimetric visual sensors for various biological targets
[60⇓⇓⇓⇓~65]. For covalently bonded hydrogels, 3D scaffolds of hydrogel networks can provide abundant conjugation sites, which become efficient immobilization substrates on which functional DNA units can be attached to design biosensing systems. By introducing nano-enzyme, silver nanoclusters and photonic crystals, a DNA hydrogel biosensor system which is very sensitive to the surrounding environment can be designed
[66⇓~68].
4.1 Application in the field of environment
4.1.1 Detection of heavy metal ions
Heavy metal ions are difficult to biodegrade in nature, and can accumulate in organisms and cause chronic poisoning. Therefore, the detection of heavy metal ions in water has been a research direction that can not be ignored. The selectivity, binding, porosity, permeability, safety and self-supporting of DNA hydrogel are very suitable for heavy metal ion detection
[69]. The target molecule or ion causes a shift in the rheological properties of the DNA hydrogel
[70]. The sensing system based on this property can replace the traditional biosensors based on electrochemistry and immunosensing.
In 2020, Wang et al. Used DNA hydrogel as a valve to control the flow rate of the solution in the capillary to detect Pb
2+[71]. As shown in fig. 3, solutions of the purified 5 '-terminal acrylate-modified DNA strands a (SA) and T (ST) were mixed with an aqueous solution of acrylamide monomer, respectively, to prepare polyacrylamide-DNA conjugates P-SA and P-ST. Then, P-SA and P-ST were mixed with DNAzyme in Tris (hydroxymethyl) aminomethane-acetic acid solution and heated to prepare DNA hydrogel. The end of the capillary was placed on the surface of the heated DNA hydrogel for about 2 s to allow the DNA hydrogel to plug the capillary. When the solution containing the Pb
2+ comes into contact with the DNA hydrogel, the Pb
2+ will decompose the 3D structure of the DNA hydrogel, thus changing the blocking effect of the DNA hydrogels on the solution. When the concentration of the Pb
2+ is high, the flow rate of the solution is high, and vice versa.
Considering the volume and loading and the one-to-one correspondence between the target and the aptamer, the use of DNase can greatly improve the performance and reduce the amount of DNA hydrogel. Many metal cations can trigger DNase,Uch as Pb2+[72], Hg2+[73], Cr6+[74],Cu2+[75], Ca2+[76], Zn2+[77], etc.
Liu et al synthesized a DNA functionalized polyacrylamide hydrogel rich in thymine for the detection of Hg
2+[78,79]. The Hg
2+ can be selectively inserted into two thymine bases, and the hydrogel is made to fluoresce yellow by the use of a dye. The addition of a stain to a DNA hydrogel produces green fluorescence in the presence of a Hg
2+. Meanwhile, the acrylamide-DNA hybridization hydrogel has good capability of adsorbing the Hg
2+, so that the heavy metal ion pollution is removed while the Hg
2+ is detected. Yang et al prepared a DNA hydrogel sensor by using DNase as the cross-linking agent of DNA hydrogel
[80]. Gold nanoparticles (AuNPs) were encapsulated in the hydrogel as an indicator, and the DNAzyme cleavage substrate was activated when the Pb
2+ came into contact with the DNA hydrogel, resulting in the release of AuNPs from the hydrogel into the supernatant. The presence of Pb
2+ can be detected by colorimetric analysis of the supernatant with a detection limit of 10 nM. In order to achieve quantitative visual analysis, the volume bar graph chip is introduced. When the catalytic decomposition of H
2O
2 produces a large amount of O
2, the air pressure pushes the ink in the V-shaped chip to move, so that visual quantitative detection can be achieved, as shown in Figure 4 and Figure 5.
4.1.2 Detection of other pollutants in the environment
Antibiotics and toxins in the environment pose a great threat to human health. The abuse of antibiotics increases the probability of the emergence of drug-resistant bacteria or viruses. And antibiotics can accumulate in the environment. Therefore, there is an urgent need for a rapid and portable method to detect antibiotics. In 2016, Tan et al. Developed a DNA hydrogel based on graphene oxide (GO), which can sensitively detect oxytetracycline (OTC) in water
[81]. Adenosine and oxytetracycline binding aptamers are used as cross-linking agents for the GO sheet.When the aptamer binds to oxytetracycline, a target-aptamer complex is formed, which greatly weakens the binding force between the fluorescein imide (FAM) and GO.The increase in the distance between the FAM and GO prevents fluorescence resonance energy transfer from occurring, resulting in fluorescence. The presence of ochratoxin in food and rivers is highly toxic. Liu et al. Successfully detected ochratoxin in solution by using DNA hydrogel and gold nanoparticles to achieve visual quantitative analysis through V-chip, with a detection limit of 1.27 nM
[82]. Aflatoxin B
1 can also be detected using the volume bar graph chip with a detection limit of 1.77 nM
[83]. Microcystin-LR (MC-LR) is harmful to human health and aquatic organisms. As shown in fig. 6, Wu et al. fabricated a detection device in which Cu/Au/Pt alloy nanoparticles were embedded in a DNA hydrogel
[84]. A DNA hydrogel was obtained by hybridizing the MC-LR aptamer with two complementary DNA strands on a polyacrylamide chain, and the Cu/Au/Pt alloy nanoparticles were embedded in the DNA hydrogel. In the presence of MC-LR, the 3D network structure of the DNA hydrogel is destroyed, and the released alloy nanoparticles catalyze the reaction of H
2O
2 and tetramethylbenzidine (TMB), resulting in a color change. The detection range of the hydrogel is 4.0~10 000 ng·L
-1.
4.2 Applications in the medical field
4.2.1 Nucleic acid detection
Enzymes, fluorophores, or radioisotope labels are used to detect nucleic acid molecules, and these methods ultimately require special instruments to capture signals and improve sensitivity. Compared with these methods, the use of DNA hydrogels for the detection of nucleic acids highlights the advantages of simplicity and convenience. Liu et al. Prepared DNA-functionalized polyacrylamide hydrogel, which showed red color after mixing with target DNA, and pointed out that the content of hydrogel would greatly affect the adsorption effect of AuNPs (gold nanoparticles)
[85]. In 2014, Weng et al. Used graphene oxide and salmon sperm DNA to prepare a hydrogel electrode, and used impedance method to detect mitochondrial DNA. When the target DNA hybridized with the probe DNA immobilized on the hydrogel electrode, the impedance would be reduced. By measuring the change of impedance, the interfering sequence could be distinguished from the matching sequence cDNA
[86]. The combination of DNA hydrogel and target molecules will produce structural changes, which will lead to changes in mechanical properties. Therefore, when the hydrogel film is prepared, the change of the permeability of the hydrogel film can be used for detecting target molecules
[87,88]. Weng et al. immobilize a DNA hydrogel film at that end of a capillary and developed a DNA hydrogel sensor driven by capillary action
[86]. The sensor detects a target miRNA by hybridization between the miRNA and a DNA probe in a DNA hydrogel. As shown in fig. 7, by immersing the sensor in the sample solution, the hybridization of the DNA probe and the target miRNA increases the permeability of the hydrogel membrane, and the solution flows into the capillary by capillary action, so that the miRNA can be quantitatively detected by measuring the flow distance of the sample solution in the capillary.
4.2.2 Protein detection
In the process of clinical detection, the type and content of protein are important physiological indicators affecting diagnosis. Targeted reaction DNA hydrogels can detect ions, toxins, proteins, etc
[88,89][90]. Therefore, DNA hydrogels have great prospects in medicine and clinical diagnosis
[91,92]. Lei et al. construct an aptamer-functionalized DNA hydrogel formed by base complementary crosslink between structural motifs and linker DNA
[88]. Thrombin, as a target molecule, can bind to the linker DNA, resulting in the destruction of the DNA hydrogel structure and the release of negatively charged gold nanoparticles. The positively charged polyethyleneimine quantum dots interact with the negatively charged gold nanoparticles through electrostatic interaction and fluorescence resonance energy transfer, resulting in the weakening of the fluorescence signal. The DNA hydrogel has a good recognition ability for thrombin with a detection limit of 67 nM. Sun et al. Used the aptamer of T-2 toxin as the cross-linking agent of the hydrogel, and the DNA hydrogel wrapped the horseradish peroxidase. When the target T-2 toxin combined with the aptamer, the hydrogel collapsed and released the horseradish peroxidase, which catalyzed the reaction of H
2O
2 and KI to produce I
2, thus corroding the gold nanoparticles
[89]. As shown in fig. 8, the UV spectrum of the gold nanoparticles is blue-shifted after the gold nanoparticles are etched. The degree of blue shift was proportional to the content of T-2 toxin. The lower limit of detection was 0.87 pg·mL
-1 and the lower limit of quantification was 2.90 pg·mL
-1.
Lai et al. simultaneously formed an aptamer with a polymerizable methacrylamide group and a bifunctional aptamer-protein complex (as a small cross-linking agent), which hydrogel undergoes macroscopic volume shrinkage in the presence of the target protein,Thrombin and PDGF-ββ proteins released during blood clotting, which are biomarkers of blood vessel production and atherosclerosis, are bound to DNA hydrogels
[93]. The DNA hydrogel prepared by the method overcomes the defects of semi-quantitative analysis and low sensitivity.
4.2.3 Metabolic small molecule detection
Metabolic small molecules are important physiological indicators for clinical diagnosis. Its abnormal expression is often closely related to diseases. Therefore, portable and rapid detection of small metabolic molecules is of great significance for clinical diagnosis.
Zhong et al. Used glucose detection by loading glucose oxidase as a catalytic matrix for DNA into a DNA hydrogel, and the biosensor could be printed directly into a flexible electrochemical electrode
[94]. The addition of electrodes enables the system to have good electrochemical performance and be compatible with 3D printing. In 2016, Huang et al. Used terminal deoxynucleotidyl transferase to polymerize DNA chains, and prepared DNA hydrogels with X-DNA as an intercalation fragment. X-DNA was elongated during polymerization and formed the building blocks of hydrogels by hybridization with "sticky" end groups. The hydrogels carried out effective cascade reactions to achieve visual detection of glucose
[95]. Ma et al. Grafted two short complementary DNA strands as crosslinkers onto a linear polyacrylamide chain, and encapsulated the modified gold nanoparticles in a hydrogel as a visible detection signal
[96]. As shown in fig. 9, in the presence of the corresponding target glucose, the complex of glucose and its boric acid derivative ligand disrupts the hydrogel structure, causing the hydrogel to collapse and release the gold nanoparticles,Aggregation of gold nanoparticles in the supernatant appears as a distinct red color, which can be directly observed by the naked eye, with a detection limit of 0.44 mM in the buffer when analyzed by UV-Vis spectrophotometry.
Adenosine triphosphate (ATP) is an important multifunctional molecule with the function of extracellular signal transduction, which plays an important role in cell metabolism. Recently, DNA hydrogel-based biosensors have been used for ATP measurement in vivo because the sensitivity, resolution and accuracy of luciferase methods need to be improved.Cell viability, including cell proliferation, migration, differentiation, apoptosis, cytokine release and necrosis, was determined by using biosensors to detect cellular ATP, adenosine diphosphate (ADP) and uridine triphosphate (UTP)
[97,98]. Liao et al. Synthesized DNA-acrylamide-based hydrogels using pre-designed nucleic acid hairpin units and single-strand binders modified polyacrylamide chains, and bound CaCO
3 to nucleic acid promoter units to form DNA-acrylamide-based hydrogel microcapsules
[97]. As shown in fig. 10, in that presence of ATP or cocaine aptamer, the duplex aptamer separate and the microcapsule partially decomposes to release the load fluorescence, enabling detection of ATP.
4.2.4 Cell Detection
When examining and repairing human tissues, it is necessary to accurately locate the spatial location of different cell types to replicate the tissue structure. Hydrogels have been used as a soft matrix because of their structural similarity to soft tissue and extracellular matrix (ECM). The characteristics of complementary binding on the surface of oligonucleotide hydrogel make it have the ability to distinguish single cells and selectively recognize different kinds of cells. However, the number of aptamer binding sites on the hydrogel surface will directly affect the number of cells that can be identified in one assay. Richards et al. Reported a DNA hydrogel that can modify the cell surface
[99]. The polymer synthesized by the hybrid chain reaction of single-stranded oligonucleotides was used as a scaffold for DNA, and then the aptamer was hybridized to the surface of the hydrogel. Thereby forming a DNA hydrogel with surface modification. Each DNA scaffold in the hydrogel provides more sites for the aptamer, directly realizing the surface functionalization of the multivalent aptamer. Compared with specific aptamers, the surface of DNA hydrogels functionalized with multivalent ligands has stronger ligand-aptamer binding ability, and more than 40% of target cells can be observed to bind to DNA hydrogels after 90 min.
The specific interaction of DNA hydrogels with cells is often used for cell and biological tissue detection. In tissue engineering, ECM mimics can accurately identify and locate biological signals or cells during tissue culture. Locating a large number of cells with different functions in a single material is a formidable challenge for tissue engineering. In multicellular organisms, important cellular activities (proliferation and differentiation) are affected by ECM signaling. Therefore, if we can simulate ECM signals, we can control cell activity to identify cell types. As shown in fig. 11, Finke et al. used Polymerase chain reaction (PCR) to extend the DNA hydrogel network structure as an artificially designed ECM
[100]. The amino-functionalized primer was first used to perform PCR with two primers having different branches, and the resulting product was bound to activated glass. Then in the primer extension reaction, the primers that did not participate in the first PCR are elongated and the branched primers incorporate them into the surface binding network. The specific combination of the functional primer and the cell surface marker is used to attach the target cell to the network to achieve the effect of cell recognition. In a similar way, the types of functional primers can be adjusted to recognize different target cells.
5 Conclusion and prospect
In this paper, the synthesis methods, physical and chemical properties of DNA hydrogels are introduced in detail, and some examples of their applications in the field of sensing are listed. Compared with traditional hydrogels, DNA hydrogels have a series of unique properties required for sensing applications: structural designability, precise recognition, intelligent response and good biocompatibility. From the perspective of basic research, DNA hydrogels, as a unique biopolymer, provide a platform for the study of nanoscale molecules. From the perspective of practical sensing applications, the integration of DNA nanostructures into hydrogels through rational design can achieve unique effects. However, there are still many challenges in the application of DNA hydrogels in the field of sensing:
(1) DNA hydrogels depend on the recognition efficiency of aptamers when detecting target molecules. However, not all existing aptamers can recognize targets efficiently and quickly, which limits the sensitivity of DNA hydrogels to a certain extent. Developing more fast and effective aptamers will be a direction of our efforts.
(2) that storage modulus of the DNA hydrogel is low, and the maximum storage modulus is only a few thousand Pa, which is far low than that of other conventional polymer hydrogels. This has a certain impact on the transfer and storage of DNA hydrogel sensing devices.
(3) The high cost and the difficulty of large-scale preparation are still the main obstacles to the practical application of DNA hydrogels. Consideration should be given to the development of more efficient synthesis methods to reduce the cost of preparation or to explore the technology of extracting more DNA from biomass, or to develop new sensing systems based on trace DNA hydrogels.
(4) The implementation of the existing DNA hydrogel sensing method relies on professional technicians and large-scale equipment, which is complex to operate and has poor portability.
It is hoped that the above challenges can be overcome in the future research, and the development of portable, sensitive detection devices that do not rely on professional technicians and large-scale equipment will be the development trend of smart hydrogels in the field of sensing.
CAS Key Laboratory of Bio-inspired Materials and Interfacial Science, Chinese Academy of Sciences and National Natural Science Foundation of China(61901042)
CAS Key Laboratory of Bio-inspired Materials and Interfacial Science, Chinese Academy of Sciences and National Natural Science Foundation of China(21975019)
CAS Key Laboratory of Bio-inspired Materials and Interfacial Science, Chinese Academy of Sciences and National Natural Science Foundation of China(31870816)
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