Utilization of Myosin and Actin Bundles for the Transport of Molecular Cargo

The utilization of motor proteins for the movement and assembly of synthetic components is currently a goal of nanoengineering research. Application of the myosin actin motor system for nanotechnological uses has been hampered due to the low flexural rigidity of individual F‐actin filaments. Here it is demonstrated how actin bundling can be used to affect the translational behavior of myosin‐propelled filaments, transport molecules across a motor‐patterned surface, and that the movement of bundled actin can be regulated photonically. These data suggest that actin bundling may significantly improve the applicability of the myosin motor for future nanotechnological applications.     

Reaction between magnesium ammine complex compound and lithium hydride

The possibility of using ammonia as a hydrogen carrier is examined for the reaction between magnesium ammine complex MgCl₂(NH₃)₆ and lithium hydride LiH. Sample was milled at low temperature of −40 °C to avoid decomposition of MgCl₂(NH₃)₆ during the milling. The effects of milling time, milling speed (revolutions per minute), and catalysts on hydrogen storage properties were investigated by thermogravimetry, thermal desorption mass spectroscopy, and X-ray diffraction experiments. Experimental results indicated that a milled composite of Mg(NH₃)₆Cl₂ and catalyzed-LiH desorbed the ∼100% H₂ gas even at 125 °C in a closed system. The reverse reaction also proceeded by separately cooling MgCl₂ at lower temperature than 100 °C and heating LiNH₂ at 300 °C in the closed system.     

Surface wettability of silicon rubber after irradiation with a glow discharge plasma

Silicon rubber, PDMS, was irradiated by Ar, Ar/H₂ and Ar/O₂ plasmas to increase the surface wettability to water which was found to increase with the irradiation time for every application of plasma irradiation. The use of Ar/O₂ plasma was found to be particularly effective in enhancing the surface wettability. The aging behavior due to exposure to air, ethanol or water was also measured. In the case of continuous exposure to water after plasma treatment, the wettability could be maintained for more than several months with the water immersion.     

Detection of genetically modified organisms in foreign-made processed foods containing corn and potato

Investigations of the validity of labeling regarding genetically modified (GM) products were conducted using polymerase chain reaction (PCR) methods for foreign-made processed foods made from corn and potato purchased in the Tokyo area and in the USA. Several kinds of GM crops were detected in 12 of 32 samples of processed corn samples. More than two GM events for which safety reviews have been completed in Japan were simultaneously detected in 10 samples. GM events MON810 and Bt11 were most frequently detected in the samples by qualitative PCR methods. MON810 was detected in 11 of the 12 samples, and Bt11 was detected in 6 of the 12 samples. In addition, Roundup Ready soy was detected in one of the 12 samples. On the other hand, CBH351, for which the safety assessment was withdrawn in Japan, was not detected in any of the 12 samples. A trial quantitative analysis was performed on six of the GM maize qualitatively positive samples. The estimated amounts of GM maize in these samples ranged from 0.2 to 2.8%, except for one sample, which contained 24.1%. For this sample, the total amount found by event-specific quantitative analysis was 23.8%. Additionally, Roundup Ready soy was detected in one sample of 21 potato-processed foods, although GM potatoes were not detected in any sample.     

A multiplex PCR method of detecting recombinant DNAs from five lines of genetically modified maize

Seven lines of genetically modified (GM) maize have been authorized in Japan as foods and feeds imported from the USA. We improved a multiplex PCR method described in the previous report in order to distinguish the five lines of GM maize. Genomic DNA was extracted from GM maize with a silica spin column kit, which could reduce experimental time and improve safety in the laboratory and potentially in the environment. We sequenced recombinant DNA (r-DNA) introduced into GM maize, and re-designed new primer pairs to increase the specificity of PCR to distinguish five lines of GM maize by multiplex PCR. A primer pair for the maize intrinsic zein gene (Ze1) was also designed to confirm the presence of amplifiable maize DNA. The lengths of PCR products using these six primer pairs were different. The Ze1 and the r-DNAs from the five lines of GM maize were qualitatively detected in one tube. The specific PCR bands were distinguishable from each other on the basis of the expected length. The r-DNA could be detected from maize samples containing 0.5% of each of the five lines of GM maize. The sensitivity would be acceptable to secure the verification of non-GMO materials and to monitor the reliability of the labeling system.     

Preparation of polyynes by liquid-phase laser ablation using different irradiation target materials and solvents

Polyynes were prepared from pellets of a perylene derivative, 3,4,9,10-perylene tetracarboxylic dianhydride (PTCDA), and graphite by liquid-phase laser ablation, using solvents of methanol, ethanol, 1-propanol, 1-butanol, t-butyl alcohol, n-hexane, and decalin. The amounts of polyynes produced from PTCDA strongly depended on the polarity of the solvent, while those from graphite did not. Polar functional groups of acid-anhydride side chains in PTCDA are believed to influence the amounts of C₂ radicals produced by laser ablation. The long-chain polyyne C₂₈H₂ was detected in a chromatogram of a solution obtained from a graphite pellet and solvent decalin.     

Strain selection and scale-up fermentation for FR901379 acylase production by Streptomyces sp. no. 6907

Micafungin (FK463) is a widely used treatment for life-threatening, deep-seated fungal infections. It is an echinocandin-like lipopeptide derived from the chemical modification of deacylated FR901379, a type of lipopeptide antibiotic produced by Coleophoma empetri F-11899. The palmitoyl moiety of FR901379 is deacylated by FR901379 acylase produced by Streptomyces sp. no. 6907. In this study, our goal was to generate an improved strain of Streptomyces sp. no. 6907 capable of hyperproducing the FR901379-acylase enzyme. To accomplish this goal, modified strains of Streptomyces sp. no. 6907 were generated using UV-irradiation mutagenesis, and strain selection was performed using an agar-plate screening method to efficiently select an acylase-hyperproducing strain. Three marker indices were shown to correlate with elevated acylase production: decreased candidacidal activity of FR901379, decreased proteolytic activity on skim milk, and phenotypic characteristics. Cloning and subsequent sequencing of the acylase gene from the hyperproducing mutant revealed no mutations in either the acylase structural gene or the 5'-flanking region required for gene expression. The growth medium was also modified to maximize acylase production. We successfully increased acylase activity approximately 65-fold, compared with the original growth conditions (wild strain cultured in the original unmodified medium). To minimize formation of excess foam during the fermentation process, we optimized the parameters of agitation speed, as calculated from the discharge flow rate. Using our improved strain and the optimized medium and growth conditions, we have developed an improved and highly reproducible method for stable large-scale production of FR901379-acylase.     

Determination of residual avoparcin in chicken muscle by HPLC with ultraviolet and amperometric detection

An analytical method was developed for determination of residual avoparcin in chicken muscle by measuring alpha- and beta-avoparcin, major components of the pharmaceutical preparation avoparcin, using HPLC with UV and amperometric detectors. The analytical HPLC was run on a Cosmosil 5C18-AR column (4.6 mm x 25 cm) with a gradient formed from A: 2.5% acetic acid, 0.01 mol/L sodium heptane sulfonic acid-acetonitrile (88.5:11.5) (pH 4.0) and B: 2.5% acetic acid-acetonitrile (10:90), using UV and amperometric detection (AMD) with glassy-carbon electrode (+900 mV). Avoparcin was extracted from chicken muscle by homogenization with methanol-0.2 mol/L sulfuric acid (6:4) followed by centrifugation after pH adjustment to 4 with 1 mol/L sodium hydroxide. The supernatant was evaporated to dryness, and the residue was dissolved in water. The aqueous layer was adjusted to pH 4 by adding 1 mol/L sodium hydroxide. Then it was purified on a Sep-Pak tC18 plus ENV cartridge. The cartridge was washed with water, and retained substances were eluted with 50% methanol. The eluate was evaporated to dryness under reduced pressure. The residue was dissolved in water and determined by HPLC. Recoveries of avoparcin spiked in chicken muscle were 73.1-88.1% at levels of 2-10 micrograms/g. The detection limits were 0.5 microgram/g (UV) and 0.2 microgram/g (AMD).     

A histochemical method using a substrate of beta-glucuronidase for detection of genetically modified papaya

A histochemical assay for detecting genetically modified (GM) papaya (derived from Line 55-1) is described. GM papaya, currently undergoing a safety assessment in Japan, was developed using a construct that included a beta-glucuronidase (GUS) reporter gene linked to a virus coat protein (CP) gene. Histochemical assay was used to visualize the blue GUS reaction product from transgenic seed embryos. Twelve embryos per fruit were extracted from the papaya seeds using a surgical knife. The embryos were incubated with the substrate 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide (X-Gluc) in a 96-well microtiter plate for 10-15 hours at 37 degrees C. Seventy-five percent of GM papaya embryos should turn blue theoretically. The histochemical assay results were completely consistent with those from a qualitative polymerase chain reaction (PCR) method developed by this laboratory. Furthermore, the method was validated in a five-laboratory study. The method for detection of GM papaya is rapid and simple, and does not require use of specialized equipment.