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101.
DNA methylation at cytosine/guanine dinucleotide islands (CpGIs) is the most prominent epigenetic modification in prokaryotic and eukaryotic genomes. DNA methyltransferases (MTases) are responsible for genomic methylation, and their aberrant activities are closely associated with various diseases including cancers. However, the specific and sensitive detection of multiple DNA MTases has remained a great challenge due to the specificity of the methylase substrate and the rareness of methylation-sensitive restriction endonuclease species. Here, we demonstrate for the first time the cytosine-5 methylation-directed construction of a Au nanoparticle (AuNP)-based nanosensor for simultaneous detection of multiple DNA MTases at the single-molecule level. We used the methyl-directed endonuclease GlaI to cleave the site-specific 5-methylcytosine (5-mC). In the presence of CpG and GpC MTases (i.e., M.SssI and M.CviPI), their hairpin substrates are methylated at cytosine-5 to form the catalytic substrates for GlaI, respectively, followed by simultaneous cleavage by GlaI to yield two capture probes. These two capture probes can hybridize with the Cy5/Cy3–signal probes which are assembled on the AuNPs, respectively, to form the double-stranded DNAs (dsDNAs). Each dsDNA with a guanine ribonucleotide can act as the catalytic substrate for ribonuclease (RNase HII), inducing recycling cleavage of signal probes to liberate large numbers of Cy5 and Cy3 molecules from the AuNPs. The released Cy5 and Cy3 molecules can be simply quantified by total internal reflection fluorescence (TIRF)-based single-molecule imaging for simultaneous measurement of M.SssI and M.CviPI MTase activities. This method exhibits good specificity and high sensitivity with a detection limit of 2.01 × 10−3 U mL−1 for M.SssI MTase and 3.39 × 10−3 U mL−1 for M.CviPI MTase, and it can be further applied for discriminating different kinds of DNA MTases, screening potential inhibitors, and measuring DNA MTase activities in human serum and cell lysate samples, holding great potential in biomedical research, clinical diagnosis, drug discovery and cancer therapeutics.Cytosine-5 methylation-directed construction of Au nanoparticle-based nanosensors enables specific and sensitive detection of multiple DNA methyltransferases. 相似文献
102.
Nianqin Jie Zhikun Si Jinghe Yang Qiang Zhang Xirong Huang Duanling Yang 《Mikrochimica acta》1997,126(1-2):93-96
A fluorescence quenching method has been developed to determine cerium with rhodamine 6G. The method is based on the oxidation of rhodamine 6G by cerium(IV) in sulfuric acid solution. The linear calibration range is 4.0 × 10–7 -1.6 × 10–6mol/L. The detection limit is 1.0 × 10–7 mol/L. The relative standard deviation was 1.0% (n = 5). The method has been used to determine cerium in rare earth ores, with satisfactory results. The method offers the advantages of simplisity, sensitivity and selectivity. 相似文献
103.
Summary A sensitive LC method for the detection of amino acids and oligopeptides with pre-column fluorescence derivatization has been
developed. Glycine, glycylglycine, triglycine, glutathione, glutamic acid, and cysteine were separated on a reversed-phase
C18 column with methanol-water-triethylamine eluent, derivatization and chromatographic conditions were optimized. The six derivatives
were eluted in 20 min with good reproducibility. The relative standard derviations (n=6) at an analytical concentration of
2×10−6 M are <5%. Detection limits (signal-to-noise ratio=3) for the six derivatives are 23–68 fmol. 相似文献
104.
Lutetium(III) forms an association compound with a new synthetic reagent, 1,6-bi(1-phenyl-3-methyl-5-pyrazolone-4)hexandione (BPMPHD), and cetyltrimethylammonium bromide (CTMAB). The compound enhances the natural fluorescence of BPMPHD remarkably, upon which a new fluorescence method was developed for determining lutetium in rare earth (RE) samples. The determination range was 1.80 × 10–7–8.8 × 10–6 g/ml. The determination limit was 29 ng/ml. The composition of the ion associate was [Lu(BPMPHD)2]–CTMAB+. 相似文献
105.
Jingcheng Hao Hanqing Wang Shuo Shi Runhua Lu Taotao Wang Ganzuo Li Haiying Sun 《中国科学B辑(英文版)》1997,40(3):225-235
By the orthogonal design, the optimal formation conditions for the middle-phase microemulsions in the system dioctadecyldimethylammonium
chloride (DODMAC)/ sodium dodecyl sulfate (SDS)/n-butanol/n-hep-tane/brine were obtained as follows: WDODMAC: WSDS = 1:4-1:5,C
π-butanol (%) = 11.0-12.0, andC
NaCl (%) = 3.25
Investigations have been made on the effects of the concentrations of NaCl and n-butanol (l.0%-14.0%), the ratios ofWDODMAC: to WSDS, and the kinds of alcohols (n-propanol, n-butanol, and n-pentanol) on the formation, the phase behaviour, the ultralow interfacial
tensions, the optimal salinity (S*), and the length of salinity (δS). Some rules and data were worked out about the formation and characteristics of the middle-phase
microemulsions. The mi-crostructures of the middle-phase microemulsions were also studied by using FT-IR, ESR, and freeze
fracture electron microscopy techniques. The results from the three methods show that the microstructures of the middlephase
mi-croemulsions undergo the change from O/W to bicontinuous (B.C.) and to W/O. The distribution rule of the orga-nized molecule
assemblies in the middle-phase microemulsions is conducible to constructing the model of microemulsion systems, to recognizing
the microstructures of the middle-phase microemulsions, and to setting forth the relationship between the microstructures
and macro-properties of rnicroemulsions.
Project supported by the Niltional Natural Science Foundation of China and the Chinese Postdoctoral Science Foundation. 相似文献
106.
We isolated cucurbitacin Ⅱa from the rhizomes of Hemsleya pengxianensis, and tested the cytotoxicity of cucurbitacin Ⅱa against various cancer cell lines by the sulforhodamine B assay(SRB). At the same time, we preliminarily found that cucurbitacin Ⅱa has a certain inhibitory activity on kinase CDK1/cyclin B, and shows potent inhibitory activities against many cancer cell lines. The cucurbitacin Ⅱa was structurally characterized by specific optical rotation measurement, high-resolution mass spectroscopy and NMR spectroscopic analysis. In addition, the molecular structure of cucurbitacin Ⅱa was further determined by X-ray single-crystal crystallography. 相似文献
107.
108.
Glutathione (GSH) in single human erythrocytes is determined by capillary zone electrophoresis with end-column amperometric detection at a gold/mercury amalgam microelectrode. A capillary of 10 microm inner diameter is suitable for determination of GSH in an individual erythrocyte with a good signal-to-noise ratio. The limit of detection is 1 x 10(-7) mol/L or 26 amol and the linear dynamic range is 2 x 10(-7) to 2 X 10(-5) mol/L for the capillary. In this method, the calibration line is obtained with a capillary adsorbed before a certain amount of hemoglobin can be used for the quantification of GSH in the external standardization. The whole cell injection and the lack of necessity of a derivatization reaction lead to more accurate and precise results, which are closer to the macroscopic values of glutathione in human red blood cell (i.e., hemolysate) than those determined by indirect laser-induced fluorescence detection. 相似文献
109.
A self-aligning end-column amperometric detection system for capillary electrophoresis was constructed. In this system, the electrode and capillary were exchanged easily and the capillary/electrode alignment procedure is not required. Gold, gold/mercury amalgam, copper and carbon fiber could be used as the working electrode. The principle is in the use of two disk holders with the capillary and the electrode in the center, so that by inserting the disk holders into a groove in the working electrode port, the capillary and the electrode are automatically aligned and the distance between the capillary and the electrode is assured at 0.24 mm. The relative standard deviation obtained using five different gold/mercury amalgam microdisk electrodes for determination of cysteine was 1.5% for the migration time and 3.3% for the electrophoretic peak current. The simple and convenient system was attractive for the routine analysis by capillary electrophoresis with electrochemical detection. The system was applied to the determination of promethazine hydrochloride in human serum. 相似文献
110.
Recently, we proposed a new transformation between the angle of canonical coordinates and the bond angle to describe the bending motion in Potential Energy Surfaces (PES) of bent triatomic molecules. In this work we extend the transformation to include linear triatomic molecules. Results for the linear triatomic molecule N2O are reported. 相似文献