ABSTRACT
In this paper, based on reciprocal chiral substrate specificity, taking achiral molecules, ethanolamine (EA) and malachite green (MG) as two model targets, biostable L- DNA aptamers and L-RNA aptamers were generated respectively by chiral inversion of existing D-aptamers. In the detection of EA with L-DNA aptamer-based sensors, the feasibility of our strategy was confirmed, while in the detection of MG with L-RNA aptamers, linear calibration curves were obtained in the range from 0.1 to 5µm with the detection limit of 0.065µm under optimized experimental conditions. The results demonstrated that the mirror-image L-aptamers have identical recognition capability as D-aptamers. Meanwhile, L-aptamers have superior biostability to resist nuclease digestion, protein binding interference and off-target effects, enabling their applications in complex practical samples, such as lake water and fish tissue extractions. Our work provides a simple, yet universal and efficient way to develop biostable aptamers.
Subject(s)
Aptamers, Nucleotide/chemistry , Drug Stability , Ethanolamine/chemistry , Rosaniline Dyes/chemistry , Stereoisomerism , Substrate SpecificityABSTRACT
Spectral reflectance data quality is important for computer color matching. There are two existing methods for evaluating the qualityspectral reflectance method and K/S method, which are too complex to apply. In this paper, 45°/0° and d/8° geometric conditions are used in the measurement of spectral reflectance of the offset ink samples printed on coated paper and silver-foiled paper while improvement on the geometric condition is made on the basis of the spectral reflectance method. Moreover, a new evaluation methodlightness and chromaticity comparative method is put forward, and comparison is made among the three methods. The results show that both 45°/0° and d/8° are feasible in the measurement of spectral reflectance of coated paper; however the former one cannot meet the requirement of spectral reflectance measurement of silver-foiled paper. In addition, as to d/8° Specular Component Included (SCI), when the silver-foiled paper is taken as the substrate, the reflectance of transparent white ink samples are smaller than that of other primary inks; and abnormal intersections appear in the curves of cyan and magenta ink respectively at the concentration of 60%, resulting in a poor spectra quality at high ink concentration; In the figure of lightness and chromaticity curves, there is significant divergence of the cyan and magenta ink curves from the referenced coated paper. In conclusion, the spectral reflectance of the transparent ink should be greater than or at least equal to other primary inks, and the maximum concentration of cyan and magenta should be limited; when the coated paper with good diffusion performance is taken as the reference, the comparative analysis is more intuitive than the two existing methods.
ABSTRACT
DNAzymes, an important type of metal ion-dependent functional nucleic acid, are widely applied in bioanalysis and biomedicine. However, the use of DNAzymes in practical applications has been impeded by the intrinsic drawbacks of natural nucleic acids, such as interferences from nuclease digestion and protein binding, as well as undesired intermolecular interactions with other nucleic acids. On the basis of reciprocal chiral substrate specificity, the enantiomer of D-DNAzyme, L-DNAzyme, could initiate catalytic cleavage activity with the same achiral metal ion as a cofactor. Meanwhile, by using the advantage of nonbiological L-DNAzyme, which is not subject to the interferences of biological matrixes, as recognition units, a facile and stable L-DNAzyme sensor was proposed for sensing metal ions in complex biological samples and live cells.
Subject(s)
Copper/analysis , DNA, Catalytic/chemistry , Lead/analysis , Copper/metabolism , DNA, Catalytic/metabolism , Enzyme Stability , HeLa Cells , Humans , Ions/analysis , Ions/metabolism , Lead/metabolism , Tumor Cells, CulturedABSTRACT
Here, we propose an efficient strategy for enzyme- and hairpin-free nucleic acid detection called an entropy beacon (abbreviated as Ebeacon). Different from previously reported DNA hybridization/displacement-based strategies, Ebeacon is driven forward by increases in the entropy of the system, instead of free energy released from new base-pair formation. Ebeacon shows high sensitivity, with a detection limit of 5 pM target DNA in buffer and 50 pM in cellular homogenate. Ebeacon also benefits from the hairpin-free amplification strategy and zero-background, excellent thermostability from 20 °C to 50 °C, as well as good resistance to complex environments. In particular, based on the huge difference between the breathing rate of a single base pair and two adjacent base pairs, Ebeacon also shows high selectivity toward base mutations, such as substitution, insertion, and deletion and, therefore, is an efficient nucleic acid detection method, comparable to most reported enzyme-free strategies.
Subject(s)
DNA/analysis , DNA/chemical synthesis , Entropy , Cell Line, Tumor , Chromatography, High Pressure Liquid , DNA/genetics , DNA Mutational Analysis , Humans , Nucleic Acid Amplification TechniquesABSTRACT
Cancer chemotherapy has been impeded by side effects and multidrug resistance (MDR) partially caused by drug efflux from cancer cells, which call for targeted drug delivery systems additionally able to circumvent MDR. Here we report multifunctional DNA nanoflowers (NFs) for targeted drug delivery to both chemosensitive and MDR cancer cells and circumvent MDR in both leukemia and breast cancer cell models. NFs are self-assembled via liquid crystallization of DNA generated by Rolling Circle Replication, during which NFs are incorporated with aptamers for specific cancer cell recognition, fluorophores for bioimaging, and Doxorubicin (Dox)-binding DNA for drug delivery. NF sizes are tunable (down to ~200 nm in diameter), and the densely packed drug-binding motifs and porous intrastructures endow NFs with high drug loading capacity (71.4%, wt/wt). The Dox-loaded NFs (NF-Dox) are stable at physiological pH, yet drug release is facilitated in acidic or basic conditions. NFs deliver Dox into target chemosensitive and MDR cancer cells, preventing drug efflux and enhancing drug retention in MDR cells. Consequently, NF-Dox induces potent cytotoxicity in both target chemosensitive cells and MDR cells, but not nontarget cells, thus concurrently circumventing MDR and reducing side effects. Overall, these NFs are promising to circumvent MDR in targeted cancer therapy.