Xiaowen Ou

A highly sensitive and facile graphene oxide-based nucleic acid probe: Label-free detection of telomerase activity in cancer patient's urine using AIEgens

Molecular beacon (MB)-based sensing platforms that consist of a fluorogen-quencher pair play an important role in medical and biological researches. However, the synthesis of both fluorogen and quencher in the nucleic acid probes will increase the burden of organic synthesis works and induce the difficulties for precisely controlling the relative distance between fluorogen and quencher, which may lead to false-positive and false-negative results. In this work, initially we report a single labeled MB (FAM-MB, with carboxyfluorescein as fluorogen and without quencher) thus simplifies MBs with the aid of graphene oxide (GO) to detect telomerase activity. To further simplify this structure, namely label-free strategy, we design a facile, sensitive and selective platform using a label-free beacon (AIE-MB, without fluorogen and quencher), based on aggregation-induced emission fluorogen (silole-R). Upon the addition of telomerase, AIE-MB induced comb-like DNA structure leads to high aggregation of silole-R and thus exhibits strong fluorescence emission. By exploitation of this, we can detect telomerase with superior sensitivity and demonstrate their applications in bladder cancer diagnosis. Compared to single-labeled FAM-MB based telomerase activity assay, the label-free AIE-MB induced method could perform the sensitive detection with high signal-to-background ratio.

Regulation of DNA self-assembly and DNA hybridization by chiral molecules with corresponding biosensor applications.

Chirality is one of the fundamental biochemical properties in a living system, and a lot of biological and physiological processes are greatly influenced by the chirality of molecules. Inspired by this phenomenon, we study the covalent assembly of DNA on chiral molecule modified surfaces and further discuss the hybridization of DNA on chiral surfaces with nucleic acids. Take methylene blue (MB) modified DNA as a model molecule, we show that the peak current of the L-NIBC (NIBC, N-isobutyryl-L(D)-cysteine) modified gold surface (L-surface) is larger than the D-surface because of a stronger interaction between short-chain DNA and the L-surface; however, the D-surface has a higher hybridization efficiency than the L-surface. Moreover, we apply this result to actual application by choosing an electrochemical DNA (E-DNA) sensor as a potential platform. Furthermore, we further amplify the difference of hybridization efficiency using the supersandwich assay. More importantly, our findings are successfully employed to program the sensitivity and limit of detection.

Imparting biomolecules to a metal-organic framework material by controlled DNA tetrahedron encapsulation

Recently, the incorporation of biomolecules in Metal-organic frameworks (MOFs) attracts many attentions because of controlling the functions, properties and stability of trapped molecules. Although there are few reports on protein/MOFs composites and their applications, none of DNA/MOFs composite is reported, as far as we know. Here, we report a new composite material which is self-assembled from 3D DNA (guest) and pre-synthesized MOFs (host) by electrostatic interactions and hydrophilic interactions in a well-dispersed fashion. Its biophysical characterization is well analyzed by fluorescence spectroscopy, quartz crystal microbalance (QCM) and transmission electron microscopy (TEM). This new composite material keeps 3D DNA nanostructure more stable than only 3D DNA nanostructure in DI water at room temperature, and stores amounts of genetic information. It will make DNA as a guest for MOFs and MOFs become a new platform for the development of DNA nanotechnology.

A highly sensitive DNA-AIEgen-based “turn-on” fluorescence chemosensor for amplification analysis of Hg 2+ ions in real samples and living cells

Functional nucleic acids (FNAs)-based biosensors have shown great potential in heavy metal ions detection due to their low-cost and easy to operate merits. However, in most FNAs based fluorescence probes, the ingenious designs of double-labeled (fluorophore and quencher group) DNA sequence, not only bring the annoyance of organic synthesis, but also restrict its use as a robust biosensor in practical duties. In this paper, we design a simple AIEgens functional nucleic acids (AFNAs) probe which consists of only fluorogen but no quencher group. With the help of duplex-specific nuclease (DSN) enzyme based target recycling, high fluorescence signal and superior sensitivity towards Hg2+ are achieved. This robust assay allows for sensitive and selective detection of Hg2+ in real water samples and mapping of intracellular Hg2+, without double-labeling of oligonucleotide with a dye-quencher pair, nor the multiple assay steps.

Stereochemistry-Guided DNA Probe for Single Nucleotide Polymorphisms Analysis

Single nucleotide polymorphisms (SNPs) are the most abundant genetic polymorphisms and are responsible for many genetic diseases and cancers. In general, SNPs detection is performed by a single probe system (SPS), in which a single probe specifically hybridizes to one target. However, with the use of this method it is hard to improve the hybridization specificity and single mismatched discrimination factors (DF). In addition, the multiprobe system (MPS) requires complex probe designs and introduces at least one auxiliary probe except for the probe complementary to the target, resulting in a complicated detection system. Faced with these difficulties, we perform the SNP detection using a d/l-tryptophan (Trp) guided DNA probe and regulate the DF of electrochemical DNA (E-DNA) sensors by molecular chirality. We show that the DF of the d-Trp incubated E-DNA sensor (d-sensor) is larger than that of the l-sensor. More importantly, we achieve the high specificity by coupling d-Trp and l-Trp incubated E-DNA sensors, and the median DF is 7.21. Furthermore, the specificity of SNP detection can be further improved by supersandwich assay, and the median DF is enlarged to 37.23, which is comparable to that obtained with a multiprobe detection system.

Simultaneous detection of telomerase and miRNA with graphene oxide-based fluorescent aptasensor in living cells and tissue samples

Telomerase and microRNAs (miRNAs) as important biomarkers are closely related to cancers. Simultaneous detection of telomerase activity and miRNAs would be beneficial to improve the specificity and reliability. Here, we establish a telomerase and miRNA-21 (miR-21) simultaneous sensing platform with graphene oxide-based fluorescent aptasensors (GOFA) including graphene oxide (GO), template strand (TS) primer and fluorophore-labeled telomerase/miR-21 oligonucleotides. Owing to π-π stacking interaction, TS primer and telomerase/miR-21 probes would be loaded onto GO, resulting in fluorescence quenching. However, in the presence of the telomerase or miR-21, the double-stranded oligonucleotides would be away from the GO surface attribute to the hybridization between the extended TS primers and telomerase probe as well as miR-21 and miR-21 probe, leading to obvious fluorescence recovery. We found that GOFA could simultaneously detect telomerase activity and miR-21 with low background signal, high sensitivity and simplified operation. Moreover, GOFA could be used for accurately detecting telomerase activity and miRNA in living cells and cancer patient tissue sample. This sensing platform shows great potential in improving the accuracy in clinical diagnosis of cancer.

Correction to “Stereochemistry-Guided DNA Probe for Single Nucleotide Polymorphisms Analysis”

ACS Appl. Mater. Interfaces 2016, 8 (25), 15911−15916. DOI: 10.1021/acsami.6b03896 I the original paper, there was an error regarding the author affiliations. The affiliation of all the authors should have read as follows: Hubei Key Laboratory of Bioinorganic Chemistry & Materia Medica, School of Chemistry and Chemical Engineering, Huazhong University of Science and Technology, Wuhan 430074, P. R. China. The correct affiliation is shown above. Addition/Correction