A speedy method to detect inserted DNA fragment in cell lines transfected with retroviral vectors.

Cells transfected by retroviral vectors are brought in agene of particular interest and are very useful in avariety of experiments. It is essential to testify that theDNA fragment was successfully introduced into the cellstogether with the retroviral vectors. Polymerase chainreaction is believed to be a fast and convenient method forthis purpose when using primers flanking the cloning siteof the inserted DNA.

Induction of the differentiation of human HL-60 promyelocytic leukemia cell line by succinoyl trehalose lipids.

Four analogs of succinoyl trehalose lipid-3 (STL-3)with saturated even-number or odd-number carbonchains, and unsaturated or halogenated fatty acidswere examined for their ability to inhibit the growthand induce the differentiation of HL-60 humanpromyelocytic leukemia cells. The optimalconcentration of STL-3 at which such activities wererecognized was closed to the critical micelleconcentration of STL-3. Analog of STL-3 witheven-number or odd-number carbon chain and unsaturatedfatty acids strongly inhibited growth and induced thedifferentiation of HL-60 cells, as evaluated in termsof nitroblue tetrazilium-reducing activity and theappearance of the CD36 antigen.

Mannosylerythritol lipid increases levels of galactoceramide in and neurite outgrowth from PC12 pheochromocytoma cells.

We report here that a microbial extracellular glycolipid,mannosylerythritol lipid (MEL), induces the outgrowth ofneurites from and enhances the activity of acetylcholinesterase(AChE) in PC12 pheochromocytoma cells. Furthermore, treatment ofPC12 cells with MEL increased levels of galactosylceramide(Galbeta1-1'Cer; GalCer). Exposure of PC12 cells to exogenous GalCer caused the dose-dependent outgrowth ofneurites.

Treatment of mouse melanoma cells with phorbol 12-myristate 13-acetate counteracts mannosylerythritol lipid-induced growth arrest and apoptosis.

Mannosylerythritol lipid (MEL), an extracellularglycolipid from yeast, induces the differentiation ofHL-60 promyelocytic leukemia cells towardsgranulocytes. We show here that MEL is also a potentinhibitor of the proliferation of mouse melanoma B16cells. Flow-cytometric analysis of the cell cycle ofMEL-treated B16 cells revealed the accumulation ofcells in the sub-G(0)/G(1) phase, which is a hallmark ofcells undergoing apoptosis.

The GAF-like-domain-containing transcriptional regulator DfdR is a sensor protein for dibenzofuran and several hydrophobic aromatic compounds.

Dibenzofuran (DF) is one of the dioxin carbon skeletal compounds used as a model to study the microbial degradation of dioxins. This study analyzed the transcriptional regulation of the DF dioxygenase genes dfdA1 to dfdA4 in the DF-utilizing actinomycetes Rhodococcus sp. strain YK2 and Terrabacter sp.

Simple and rapid colorimetric detection of cofactors of aptazymes using noncrosslinking gold nanoparticle aggregation.

We developed a method for simply and rapidly detecting cofactors of aptazymes with high sensitivity using unique noncrosslinking gold nanoparticle aggregation. Applying this method to a theophylline-dependent aptazyme, 100 microM, 10 microM, and 1 microM theophylline were detected easily by the naked eye within 10 min, 20 min, and 65 min, respectively. This method is also applicable to other cleavase-aptazymes without altering the probe-DNA sequence on the gold nanoparticle.

Development of microscopic systems for high-speed dual-excitation ratiometric Ca2+ imaging.

For quantitative measurements of Ca(2+) concentration ([Ca(2+)]), ratiometric dyes are preferable, because the use of such dyes allows for correction of uneven loading or partitioning of dye within the cell as well as variations in cell thickness. Although dual-excitation ratiometric dyes for measuring [Ca(2+)], such as Fura-2, Fura-Red, and ratiometric-pericam, are widely used for a variety of applications, it has been difficult to use them for monitoring very fast Ca(2+) dynamics or Ca(2+) changes in highly motile cells. To overcome this problem, we have developed three new dual-excitation ratiometry systems.

Heterogeneity of osmium oxidation efficiency at consecutive thymines.

Consecutive thymine sequences were oxidized more efficiently with a mixture including potassium osmate, potassium hexacyanoferrate(III), and bipyridine than an isolated thymine, and, in particular, osmium oxidation at the 5'T of TT progressed much more rapidly, compared with those of other thymines.

Ultrafast pump-probe study of the primary photoreaction process in pharaonis halorhodopsin: halide ion dependence and isomerization dynamics.

Halorhodopsin is a retinal protein that acts as a light-driven chloride pump in the Haloarchaeal cell membrane. A chloride ion is bound near the retinal chromophore, and light-induced all- trans --> 13- cis isomerization triggers the unidirectional chloride ion pump. We investigated the primary ultrafast dynamics of Natronomonas pharaonis halorhodopsin that contains Cl (-), Br (-), or I (-) ( pHR-Cl (-), pHR-Br (-), or pHR-I (-)) using ultrafast pump-probe spectroscopy with approximately 30 fs time resolution.

Simple and rapid colorimetric detection of low-weight molecules using aptazymes in combination with noncrosslinking gold nanoparticle aggregation.

We developed a method for simply and rapidly detecting cofactors of aptazymes with high sensitivity using a unique noncrosslinking gold nanoparticle aggregation. Applying this method to a theophylline dependent aptazyme, 100 microM, 10 microM, and 1 microM theophylline were detected easily by the naked eye within 10 min, 20 min, and 65 min, respectively. This method is applicable to other cleavage-aptazymes without altering the probe-DNA sequence.

In vitro selection of RNA aptamer to hemin.

A new RNA aptamer-binding hemin was synthesized by the in vitro selection (SELEX) method. A pool of 103 bases single strand DNAs containing a randomized sequence of 59 bases was synthesized. The pool was incubated with hemin on hemin-immobilized affinity column.

Design of a fluorescent probe for DNA/RNA imaging.

We report new on-off fluorescent DNA probes. We used the fluorescence quenching by the exciton coupling effect of thiazole orange dyes to achieve turning off of fluorescence. Aggregation of dyes tethered to a single nucleotide in DNA dramatically changes the photophysical properties of the dyes, as demonstrated by the large spectral shifts relative to the absorption of the monomeric dyes binding to DNA.

Two-photon induced polymer nanomovement.

We present the first report of two-photon induced plastic surface deformation in solid polymer films. Exposure of azo polymer films, which absorb in the visible range (lambda(max) = 480 nm), to intense 920 nm irradiation leads to polarization dependent photofluidic polymer nanomovement caused by photoselective two-photon trans <-->cis isomerization. The deformations were induced by a gradient of light intensity; and strongly depend on the wavelength and the polarization direction of the incident laser light and the position of the focused spot with respect to the plane of the polymer film.

Band Jahn-Teller instability and formation of valence bond solid in a mixed-valent spinel oxide LiRh2O4.

We have synthesized a new spinel oxide LiRh2O4 with a mixed-valent configuration of Rh3+ and Rh4+. At room temperature, it is a paramagnetic metal, but on cooling, a metal-insulator transition occurs and a valence bond solid state is formed below 170 K. We argue that the formation of valence bond solid is promoted by a band Jahn-Teller transition at 230 K and the resultant confinement of t_{2g} holes within the xy band.

Structural determination of montanacin D by total synthesis.

The first total syntheses of acetogenin 3 and its 4 S,8 R-isomer are described. The key step involves intermolecular metathesis of an alpha,beta-unsaturated ketone carrying a tetrahydropyranyl lactone with a tetrahydrofuran derivative. Compound 3 has spectroscopic and physical data consistent with those of natural montanacin D, suggesting that the absolute configuration of the natural product is as shown in 3.

Self-directed chain reaction by small ketones with the dangling bond site on the Si(100)-(2 x 1)-H surface: acetophenone, a unique example.

Using scanning tunneling microscope (STM) at 300 K, we studied the growth of one-dimensional molecular assemblies (molecular lines) on the Si(100)-(2 x 1)-H surface through the chain reaction of small ketone (CH 3COCH 3, PhCOPh, and PhCOCH 3) molecules with dangling bond (DB) sites of the substrate. Acetone and benzophenone show the growth of molecular lines exclusively parallel to the dimer row direction. In contrast, acetophenone molecules show some molecular lines perpendicular, in addition to parallel, to the dimer row direction.

Sequence dependence of fluorescence emission and quenching of doubly thiazole orange labeled DNA: effective design of a hybridization-sensitive probe.

We have designed a doubly thiazole orange labeled nucleoside showing high fluorescence intensity for a hybrid with the target DNA and effective quenching for a single-stranded state. Knowing how much the fluorescence emission and quenching of this probe depend on the probe sequence and why there is such a sequence dependence is important for effective probe design, we synthesized more than 30 probe sequences and measured their fluorescence intensities. When the probe hybridized with the target DNA strands, there was strong emission, whereas the emission intensity was much weaker before hybridization; however, self-dimerization of probes suppressed fluorescence quenching.

Luminescence from 3,4,9,10-perylenetetracarboxylic dianhydride on Ag(111) surface excited by tunneling electrons in scanning tunneling microscopy.

The electronic excitations induced with tunneling electrons into adlayers of 3,4,9,10-perylenetetracarboxylic dianhydride (PTCDA) on Ag(111) have been investigated by in situ fluorescence spectroscopy in scanning tunneling microscopy (STM). A minute area of the surface is excited by an electron tunneling process in STM. Fluorescence spectra strongly depend on the coverage of PTCDA on Ag(111).

Terahertz beam steering and frequency tuning by using the spatial dispersion of ultrafast laser pulses.

We demonstrate a terahertz (THz) beam steering method using difference frequency generation that is based on the principle of phased array antennas. A strip-line photoconductive antenna was illuminated by two spatially dispersed beams produced from an ultrafast laser. THz radiation with a bandwidth of 65 GHz was generated from the overlapping area of the two beams, between which the frequency difference was approximately constant.

Dynamic interfacial properties of poly(ethylene glycol)-modified ferritin at the solid/liquid interface.

Poly(ethylene glycol)-modified ferritins (PEG-ferritins) with various molecular weights were synthesized by the grafting method, and their dynamic interfacial properties at the solid/liquid interface were investigated. The number of PEG grafted to ferritins was controlled by the amount of 1,1'-carbonyldiimidazole-modified PEG adding to the reaction solution. The adsorption kinetics and energy dissipation of PEG-ferritins onto bare Si substrate and amino-modified Si substrate were investigated with a quartz crystal microbalance (QCM) in 10 mM bis-Tris/HCl buffer (pH 5.

Synthesis and styrene polymerisation catalysis of eta5- and eta1-pyrrolyl-ligated cationic rare earth metal aminobenzyl complexes.

The cationic rare earth metal aminobenzyl complexes bearing mono(pyrrolyl) ligands are synthesised and structurally characterised, and the coordination mode of the pyrrolyl ligands is found to show significant influence on the polymerisation of styrene.

Synthesis and biological activities of reveromycin A and spirofungin A derivatives.

Various derivatives of reveromycin A, an inhibitor of eukaryotic cell growth, and spirofungin A, focusing on the 5S hydroxyl group and C18 hemisuccinyl group, were synthesized and their inhibitory effects on both the isoleucyl-tRNA synthetase activity and the survival of osteoclasts, and activities on the morphological reversion of src(ts)-NRK cells were examined. It was found that 2,3-dihydroreveromycin A is the promising derivative of reveromycin A based on the activity and stability.

Expression of Rab27B-binding protein Slp1 in pancreatic acinar cells and its involvement in amylase secretion.

Slp1 is a putative Rab27 effector protein and implicated in intracellular membrane transport; however, the precise tissue distribution and function of Slp1 protein remain largely unknown. In this study we investigated the tissue distribution of Slp1 in mice and found that Slp1 is abundantly expressed in the pancreas, especially in the apical region of pancreatic acinar cells. Slp1 interacted with Rab27B in vivo and both proteins were co-localized on zymogen granules.