Isothermal amplification is an efficient way to amplify DNA with high accuracy; however, the real-time monitoring for quantification analysis mostly relied on expensive and precisely designed probes. In the present study, a graphene oxide (GO)-based nanoprobe was used to real-time monitor the isothermal amplification process. The interaction between GO and different DNA structures was systematically investigated, including single-stranded DNA (ssDNA), double-stranded DNA (dsDNA), DNA 3-helix, and long rolling circle amplification (RCA) and hybridization chain reaction (HCR) products, which existed in one-, two-, and three-dimensional structures. It was found that the high rigid structures exhibited much lower affinity with GO than soft ssDNA, and generally the rigidity was dependent on the length of targets and the hybridization position with probe DNA. On the basis of these results, we successfully monitored HCR amplification process, RCA process, and the enzyme restriction of RCA products with GO nanoprobe; other applications including the detection of the assembly/disassembly of DNA 3-helix structures were also performed. Compared to the widely used end-point detection methods, the GO-based sensing platform is simple, sensitive, cost-effective, and especially in a real-time monitoring mode. We believe such studies can provide comprehensive understandings and evocation on design of GO-based biosensors for broad application in various fields.Keywords: interaction of graphene oxide and DNA; isothermal amplification; length and position dependent; real-time detection; various DNA structures;

DNA nanostructures have attracted wide interest in biomedical applications, especially as nanocarriers for drug delivery. Therefore, it is important to ensure the structural integrity of DNA nanostructures under ambient temperature storage. In this study, we examined lyophilization-based preservation of DNA nanostructures by investigating the structural integrity of different DNA nanostructures reconstituted from lyophilization. We demonstrated that lyophilization under appropriate ionic strength is amenable to the storage of DNA nanostructures. Compared with that stored in liquid solution, DNA nanostructure carriers reconstituted from lyophilization showed significantly better structural integrity after an accelerated aging test equivalent to 100-day room-temperature storage.Keywords: DNA nanostructures; ionic strength; lyophilization; storage; structure friendly;

Herein, we have developed a simple and facile method to synthesize yolk–shell nanostructured Fe3O4@C nanoparticles (NPs) as a multifunctional biosensing platform for the label-free colorimetric detection of H2O2 and glucose. It was demonstrated that Fe3O4@C yolk–shell nanostructures (YSNs) retained the magnetic properties that can be used for separation and concentration. Also importantly, the Fe3O4@C YSNs exhibited an intrinsic peroxidase-like activity that could quickly catalyze the enzyme substrate in the presence of H2O2 and produce a blue color. Compared to other similar ferric oxide-based NPs with different structures, Fe3O4@C YSNs exhibited greatly enhanced catalytic activities due to their unique structural features. Moreover, steady-state kinetics indicated the catalytic behaviors in agreement with the classic Michaelis–Menten models. Taking advantage of the high catalytic activity, Fe3O4@C YSNs were employed as novel peroxidase mimetics for label-free, rapid, sensitive, and specific colorimetric sensing of H2O2 and glucose, suggesting that Fe3O4@C YSNs have the potential for construction of portable sensors in the application of point-of-care (POC) diagnosis and on-site tests.

The nanocarrier systems have been widely used for improving solubility, stability and therapeutic activity of drugs due to their high drug-loading efficiency, good target specificity and long circulation time. To achieve precisely controlled loading and tumor-selective release of drugs, extensive researches have been focused on developing nanocarriers with low toxicity, excellent biocomapatibility and biodegradability. As a type of nano-biomaterials with various functions and applications, self-assembled DNA nanostructures explored new ways to develop drug carriers in smart drug delivery based on their well-defined structures, good biocompatibility and stability, high cell membrane permeability and controlled drug releasing property. In this review, we summarized the developing course and the recent advances of DNA nanostructures for drug delivery, including the application of both static and dynamic DNA nanostructures. The application of dynamic DNA nanostructures for controllable drug release showed great potential in smart drug delivery.In this review, the developing course and the recent advances of DNA nanostructures for drug delivery were summerized, including the application of both static and dynamic DNA nanostructures. We also predicted the application prospect of DNA nanostructures.Download high-res image (125KB)Download full-size image

In this work, we have developed multiple-armed DNA tetrahedral nanostructures (TDNs) for dual-modality in vivo imaging using near-infrared (NIR) fluorescence and single-photon emission computed tomography (SPECT). We found that the presence of arm strands in TDNs remarkably enhanced their in vitro stability, allowing them to stay intact for at least 12 h in serum. By using NIR fluorescence imaging, we evaluated in mice the pharmacokinetics of TDNs, which exhibited distinctly different in vivo biodistribution patterns compared with those of double-stranded (ds)DNA. We also noticed that TDNs had twofold longer circulation time in the blood system than that of dsDNA. With the use of multiple-armed TDNs, we could precisely anchor an exact number of functional groups including tumor-targeting folic acid (FA), NIR emitter Dylight 755, and radioactive isotope 99mTc on prescribed positions of TDNs, which showed the capability of targeted imaging ability in cancer cells. Furthermore, we realized noninvasive tumor-targeting imaging in tumor-bearing mice by using both NIR and SPECT modalities.Keywords: dual-modality imaging; near-infrared fluorescence; single-photon emission computed tomography; tetrahedral DNA nanostructures; tumor targeting

Microarrays of biomolecules hold great promise in the fields of genomics, proteomics, and clinical assays on account of their remarkably parallel and high-throughput assay capability. However, the fluorescence detection used in most conventional DNA microarrays is still limited by sensitivity. In this study, we have demonstrated a novel universal and highly sensitive platform for fluorescent detection of sequence specific DNA at the femtomolar level by combining dextran-coated microarrays with hybridization chain reaction (HCR) signal amplification. Three-dimensional dextran matrix was covalently coated on glass surface as the scaffold to immobilize DNA recognition probes to increase the surface binding capacity and accessibility. DNA nanowire tentacles were formed on the matrix surface for efficient signal amplification by capturing multiple fluorescent molecules in a highly ordered way. By quantifying microscopic fluorescent signals, the synergetic effects of dextran and HCR greatly improved sensitivity of DNA microarrays, with a detection limit of 10 fM (1×105 molecules). This detection assay could recognize one-base mismatch with fluorescence signals dropped down to ~20%. This cost-effective microarray platform also worked well with samples in serum and thus shows great potential for clinical diagnosis.Dextran-HCR microarrays for DNA detection.

DNA nanostructures have attracted great attention due to their precisely controllable geometry and great potential in various areas including bottom-up self-assembly. However, construction of higher-order DNA nanoarchitectures with individual DNA nanostructures is often hampered with the purity and quantity of these “bricks”. Here, we introduced size exclusion chromatography (SEC) to prepare highly purified tetrahedral DNA nanocages in large scale and demonstrated that precise quantification of DNA nanocages was the key to the formation of higher-order DNA nanoarchitectures. We successfully purified a series of DNA nanocages with different sizes, including seven DNA tetrahedra with different edge lengths (7, 10, 13, 17, 20, 26, 30 bp) and one trigonal bipyramid with a 20-bp edge. These highly purified and aggregation-free DNA nanocages could be self-assembled into higher-order DNA nanoarchitectures with extraordinarily high yields (98% for dimer and 95% for trimer). As a comparison, unpurified DNA nanocages resulted in low yield of 14% for dimer and 12% for trimer, respectively. AFM images cleraly presented the characteristic structure of monomer, dimer and trimer, impling the purified DNA nanocages well-formed the designed nanoarchitectures. Therefore, we have demonstrated that highly purified DNA nanocages are excellent “bricks” for DNA nanotechnology and show great potential in various applications of DNA nanomaterials.Keywords: DNA tetrahedron; highly ordered DNA nanoarchitectures; purification; quantification; size exclusion chromatography;

In this work, we investigated the interactions between graphene oxide (GO) and conjugated polyelectrolytes (CPEs) with different backbone and side chain structures. By studying the mechanism of fluorescence quenching of CPEs by GO, we find that the charge and the molecular structure of CPEs play important roles for GO–CPEs interactions. Among them, electrostatic interaction, π–π interaction, and cation−π bonding are dominant driving forces. By using a cationic P2, we have developed a sensitive homogeneous sensor for DNA and RNA detection with a detection limit of 50 pM DNA and RNA, which increased the sensitivity by 40-fold as compared to GO-free CPE-based sensors. This GO-assisted CPE sensing strategy is also generic and shows a high potential for biosensor designs based on aptamers, proteins, peptides, and other biological probes.

We herein report a power-free microfluidic chip for fluorescent DNA detection with high single-nucleotide polymorphism discrimination, using a DNA intercalator and graphene oxide.

We designed a single-fluorophore-tagged hairpin-structured nano-beacon probe by using a superquencher, graphene oxide (GO), based on which a new method for the analysis of DNA phosphorylation detection was developed.

In this work, we report the design of a novel graphene-based molecular beacon (MB) that could sensitively and selectively detect specific DNA sequences. The ability of water-soluble graphene oxide (GO) to differentiated hairpin and dsDNA offered a new approach to detect DNA. We found that the background fluorescence of MB was significantly suppressed in the presence of GO, which increased the signal-to-background ratio, hence the sensitivity. Moreover, the single-mismatch differentiation ability of hairpin DNA was maintained, leading to high selectivity of this new method.

The single stranded DNA (ssDNA) with G-rich sequence can fold into G-quadruplex via intramolecular hydrogen-bonding interaction in the presence of ligand. This structure conversion can be specifically detected by a fluorescence method based on different interaction between SYBR Green I (SG) and various DNA structures. SG is proved to intercalate into G-quadruplex and results in high fluorescence intensity, which can be further amplified by 6-fold through fluorescence resonance energy transfer (FRET) from a water-soluble cationic conjugated polymer (CCP) to SG due to the high affinity of positively charged CCP to negatively charged rigid G-quadruplex, whereas it is not performed for ssDNA in the absence of K+. As a result, the ssDNA/SG/CCP complex can be used to detect potassium ions with improved selectivity in a label-free and cost-effective manner.Keywords: cationic conjugated polymer; fluorescence resonance energy transfer; potassium ion; SYBR Green I

Based on gold nanoparticles (AuNPs) and engineered DNA aptamers, we designed a novel bioassay strategy for the detection of adenosine as a small target molecule. In this design, an aptamer is engineered to consist of two pieces of random-coil like ssDNA which are respectively attached to AuNPs through their 5′-thiol-modified end. They can reassemble into the intact aptamer tertiary structure and induce nanoparticle aggregation in the presence of the specific target. Results have demonstrated that gold nanoparticles can effectively differentiate these two different DNA structures via their characteristic surface plasmon resonance-based color change. With this method, adenosine can be selectively detected in the low micromolar range, which means that the strategy reported here can be applicable to the detection of several other small target molecules.

We designed a single-fluorophore-tagged hairpin-structured nano-beacon probe by using a superquencher, graphene oxide (GO), based on which a new method for the analysis of DNA phosphorylation detection was developed.

We herein report a power-free microfluidic chip for fluorescent DNA detection with high single-nucleotide polymorphism discrimination, using a DNA intercalator and graphene oxide.