DNA Dendrimer-Based Directed 3D Walking Nanomachine for the Sensitive Detection and Intracellular Imaging of miRNA.

Taking advantage of the remarkable processivity and membrane penetrability, the gold nanoparticle (AuNP)-based three-dimensional (3D) DNA walking nanomachine has induced tremendous promise in molecular diagnostics and cancer therapy, whereas the executive ability of this nanomachine was eventually limited because of the disordered assembly between the walker and the track. Therefore, we developed a well-directed 3D DNA walking nanomachine by employing a DNA dendrimer as the track for intracellular imaging with high directionality and controllability. The nanomachine was constructed on a DNA dendrimer decorated with a substrate strand serving as the DNA track and a DNAzyme restrained by a locking strand as the walker. In this system, the distribution of the substrate strand and DNAzyme on the DNA dendrimer could be precisely regulated to achieve expected goals because of the specificity and predictability of the Watson-Crick base pairing, paving an explicit route for each walker to move along the track. Moreover, such a DNA dendrimer-based nanomachine owned prominent stability and anti-interference ability. By choosing microRNA-21 as a model analyte, the nanomachine was applied for the imaging of microRNA-21 in different cell lines and the monitoring of the dynamic microRNA-21 expression level in cancer cells. Therefore, we believe that this directed DNA walking nanomachine will have a variety of applications in molecular diagnostics and biological function modulation.

Oil contamination drives the transformation of soil microbial communities: Co-occurrence pattern, metabolic enzymes and culturable hydrocarbon-degrading bacteria.

The land-based oil extraction activity has led to serious pollution of the soil. While microbes may play an important role in the remediation of contaminated soils, ecological effects of oil pollution on soil microbial relationships remain poorly understood. Here, typical contaminated soils and undisturbed soils from seven oilfields of China were investigated in terms of their physicochemical characteristics, indigenous microbial assemblages, bacterial co-occurrence patterns, and metabolic enzymes. Network visualization based on k-core decomposition illustrated that oil pollution reduced correlations between co-existing bacteria. The core genera were altered to those related with oil metabolism (Pseudarthrobacter, Alcanivorax, Sphingomonas, Chromohalobacter and Nocardioides). Under oil pollution pressure, the indigenous bacteria Gammaproteobacteria was domesticated as biomarker and the enzyme expression associated with the metabolism of toxic benzene, toluene, ethylbenzene, xylene and polycyclic aromatic hydrocarbons was enhanced. Functional pathways of xenobiotics biodegradation were also stimulated under oil contamination. Finally, twelve culturable hydrocarbon-degrading microbes were isolated from these polluted soils and classified into Stenotrophomonas, Delftia, Pseudomonas and Bacillus. These results show that the soil microbial communities are transformed under oil pollution stress, and also provide useful information for future bioremediation processes.

A DNAzyme-based label-free fluorescent probe for guanosine-5'-triphosphate detection.

Guanosine-5'-triphosphate (GTP) plays a key role in many important biological processes of cells. It is not only a primer for DNA replication and one of the four essential nucleoside triphosphates for mRNA synthesis, but also an energy source for translation and other important cellular processes. It can be converted to adenine nucleoside triphosphate (ATP), and the intracellular GTP level is closely related to the specific pathological state, so it is crucial to establish a simple and accurate method for the detection of GTP. Deoxyribozymes have unique catalytic and structural properties. One of the deoxyribozymes which is named DK2 with self-phosphorylation ability can transfer a phosphate from GTP to the 5' end in the presence of manganese(ii), while lambda exonuclease (λexo) catalyzes the gradual hydrolysis of double-stranded DNA molecules phosphorylated at the 5'-end from 5' to 3', but cannot cleave the 5'-OH end. The fluorescent dye SYBR Green I (SG I) can bind to dsDNA and produce significant fluorescence, but it can only give out weak fluorescence when it is mixed with a single strand. Here, we present a novel unlabeled fluorescence assay for GTP based on the self-phosphorylation of deoxyribozyme DK2 and the specific hydrolysis of λexo. Owing to the advantages of simple operation, high sensitivity, good specificity, low cost and without fluorophore (quenching group) labeling, this method has great potential in biological applications.

Spatiotemporal control of DNAzyme activity for fluorescent imaging of telomerase RNA in living cells.

BACKGROUND: Human telomerase is a ribonucleoprotein complex that includes proteins and human telomerase RNA (hTR). Emerging evidence suggested that the expression level of hTR was high related with the development of tumor, so it is important to accurately detect the content of hTR. Optical control of DNAzyme activity shows a promising strategy for precise biosensing, biomedical imaging and modulation of biological processes. Although DNAzyme-based sensors can be controlled spatiotemporally by light, its application in the detection of hTR in living cells is still rare. Therefore, designing DNAzyme activity spatiotemporal controllable sensors for hTR detection is highly needed. RESULTS: We developed a UV light-activated DNAzyme-based nanoprobe for spatially accurate imaging of intracellular hTR. The proposed nanoprobe was named MDPH, which composed of an 8-17 DNAzyme (D) inactivated by a protector strand (P), a substrate strand (H), and MnO2 nanosheets. The MnO2 nanosheets can enhance the cellular uptake of DNA strands, so that MDPH probe can enter cells autonomously through endocytosis. Under the high concentration of GSH in cancer cells, MnO2 nanosheets can self-generate cofactors to maintain the catalytic activity of DNAzyme. When exposing UV light and in presence of target hTR, DNAzyme could cleave substrate H, resulting in the recovery of fluorescence of the system. The cells imaging results show that MDPH probe could be spatiotemporally controlled to image endogenous hTR in cancer cells. SIGNIFICANCE: With this design, telomerase RNA-specific fluorescent imaging was achieved by MDPH probe in both cancer and normal cells. Our probe made a promising new platform for spatiotemporal controllable intracellular hTR monitoring. This current method can be applied to monitor a variety of other biomarkers in living cells and perform medical diagnosis, so it may has broad applications in the field of medicine.

Accelerated DNAzyme-based fluorescent nanoprobe for highly sensitive microRNA detection in live cells.

By assembling DNAzyme on DNA nanowires through DNA hybridization, we have developed a novel accelerated DNAzyme-based fluorescent nanoprobe for fast, sensitive and selective detection of miRNA. Moreover, the strategy was successfully applied for in situ imaging of miRNA-21 in different cell lines.