Direct Injection of CRISPR/Cas9-Related mRNA into Cytoplasm of Parthenogenetically Activated Porcine Oocytes Causes Frequent Mosaicism for Indel Mutations

Some reports demonstrated successful genome editing in pigs by one-step zygote microinjection of mRNA of CRISPR/Cas9-related components. Given the relatively long gestation periods and the high cost of housing, the establishment of a single blastocyst-based assay for rapid optimization of the above system is required. As a proof-of-concept, we attempted to disrupt a gene (GGTA1) encoding the α-1,3-galactosyltransferase that synthesizes the α-Gal epitope using parthenogenetically activated porcine oocytes. The lack of α-Gal epitope expression can be monitored by staining with fluorescently labeled isolectin BS-I-B₄ (IB4), which binds specifically to the α-Gal epitope. When oocytes were injected with guide RNA specific to GGTA1 together with enhanced green fluorescent protein (EGFP) and human Cas9 mRNAs, 65% (24/37) of the developing blastocysts exhibited green fluorescence, although almost all (96%, 23/24) showed a mosaic fluorescent pattern. Staining with IB4 revealed that the green fluorescent area often had a reduced binding activity to IB4. Of the 16 samples tested, six (five fluorescent and one non-fluorescent blastocysts) had indel mutations, suggesting a correlation between EGFP expression and mutation induction. Furthermore, it is suggested that zygote microinjection of mRNAs might lead to the production of piglets with cells harboring various mutation types.     

Mean molal activity coefficients of aqueous scandium chloride, nitrate, bromide and perchlorate solutions at 25.0°

The mean molal activity coefficients of aqueous scandium chloride, nitrate, bromide and perchlorate solutions were determined at 25.0° for dilute to saturated concentrations, together with the activities of water. In the dilute solutions of scandium halides, the activity coefficients were obtained from electromotive force measurements on galvanic cells, and the osmotic coefficients of all four solutions above 0.1 mol kg⁻¹ were determined from the isopiestic measurements. Least-squares equations were fitted to these coefficients, which were then used to calculate the mean molal activity coefficients and water activities. The relationships between these results and the corresponding activity data for other rare earth salts, and to the cation hydration and ionic interactions, are discussed. The results on scandium perchlorate solutions suggested that the inner-sphere hydration number of tervalent scandium ion may be seven.     

An FPGA implementation of hard‐wired sequence control system based on PLC software

Although a programmable logic controller (PLC) has been widely adopted for the sequence control of industrial machinery, its performance does not always satisfy the recent requirements in large and highly responsive systems. With the state‐of‐the‐art field programmable gate array (FPGA) technology, it is possible to implement a control program with hard‐wired logic for higher response and reduced implementation cost/space. This approach is also worthwhile for transmigration of legacy PLC software into forthcoming FPGA‐based control hardware. This study presents a systematic method to implement a hard‐wired sequence control from PLC software. PLC instructions are converted into VHDL codes, and then implemented as logic circuit with various peripheral functions. Productive PLC programs were examined with Mitsubishi Electric FX2N PLC and Altera Stratix II FPGA, and were shown to fit into a common FPGA chip. A straightforward Sequential design was estimated to be 184 times faster than PLC, while a performance‐oriented Flat design was estimated to be 44 times faster than Sequential design (i.e., 8050 times faster than PLC). A practical perfect layer winder system was actually built and successfully operated with our FPGA control board, whose logic design was implemented with our tools. © 2011 Institute of Electrical Engineers of Japan. Published by John Wiley & Sons, Inc.     

Application of step‐response test for evaluation of uncertainty of standard measuring system for lightning‐impulse high voltage

In evaluating the uncertainty of the standard measuring system for lightning‐impulse high voltages, which is composed of a standard voltage divider, a digital recorder, and calibrators, step‐response tests of the standard voltage divider may be useful. In this paper, a convolution algorithm is employed to calculate the output impulse voltage waveforms from measured step‐response waveforms. The uncertainties of peak‐value measurement due to the influence of the nominal epoch, uncertainty of the peak‐value measurement due to dispersion of the AC scale factor, and uncertainty of the virtual front‐time measurement due to long‐term stability are evaluated. Furthermore, the error of the virtual front time of the output waveforms is discussed. The front part of the step‐response waveform, t≤ T₃₀%, does not influence the error of the virtual front time. Therefore, for the standard voltage divider, the step‐response parameters, that is, the experimental response time, partial response time, settling time, and overshoot, have almost nothing to do with the error of the virtual front time. © 2012 Wiley Periodicals, Inc. Electr Eng Jpn, 180(2): 24–32, 2012; Published online in Wiley Online Library ( wileyonlinelibrary.com ). DOI 10.1002/eej.21279     

New chemical etching solution for InP and GaInAsP gratings

A new chemical etching solution of bromine-water system, which is suitable for transforming the fine resist grating mask pattern onto InP and GaInAsP surfaces, is reported. The Br2:H2O:H3PO4 (or HCl) solution does not dissolve AZ 1350 photoresist and exhibits both moderate etching rate and a pit-free etched surface.     

Colloidal platinum in hydrogen-rich water exhibits radical-scavenging activity and improves blood fluidity

The 'colloidal platinum' stabilized with polyvinylpyrrolidone (Pt/PVP-colloid) was dispersed in hydrogen-rich water (HW; hydrogen concentration, 0.82 ppm; oxidation-reduction potential, -583 mV) or regular water (RW; <0.01 ppm, +218 mV). And we evaluated the antioxidant activity of Pt/PVP-colloid in HW or RW on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and improvement of blood fluidity under 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH)-induced oxidative stress. When applied with the 0.25-0.5 ppm Pt/PVP-colloid in RW or HW, the level of DPPH radicals decreased to 77.5-59.6% or 16.1-5.6%, in contrast to the level as high as 81.3% for HW alone, respectively, as measured by an electron spin resonance method. The horse blood, which was subjected to AAPH-induced oxidative stress, was incubated for 24 hr with RW or HW, and thereafter required 13.7 sec (100%) or 5.7 sec (42.3%) for passing through the micro-channels in a rheology equipment. When treated with 0.5-1.0 ppm Pt/PVP-colloid in RW or HW, the blood passage time in the micro-channels decreased dose-dependently to 9.7-7.3 sec (71.6-53.8%) or 4.3-1.3 sec (32.8-10.3%), and the rate of micro-channels clogged with erythrocyte aggregates decreased to 23.8-21.0% or 15.8-9.8%, respectively, from 42.8% for no addition of Pt/PVP. By scanning electron microscopy, AAPH-treated erythrocytes lost intact surface morphology on the membrane together with protrusions and without hollows, being indicative of impaired transforming ability, and the rate of erythrocyte agglutination was increased to 46.2%. When treated the horse blood with HW alone significantly decreased the rate of erythrocyte agglutination to 29.6%, whereas 1.0 ppm Pt/PVP-colloid in RW or HW decreased it to 24.1% or 21.1%, respectively. Thus, DPPH-radical-scavenging and erythrocyte-protecting effects of Pt/PVP-colloid in HW were superior to those of Pt/PVP-colloid in RW or Pt/PVP-free HW. The results could be mainly attributed to the enhanced antioxidant activity of Pt/PVP in HW, which may be due to captured-hydrogen on platinum.     

Decolorization of beads-milled TiO2 nanoparticles suspension in an organic solvent

In this paper, a new method is proposed for the decolorization of a yellow-hued suspension of rutile TiO₂ nanoparticles in an organic solvent (diethylene glycol dimethylether). The presence of color has always been undesirable in a suspension of nanoparticles filler used for industrial needs, particularly for optical applications.A colorless suspension was achieved by irradiating well-dispersed TiO₂ nanoparticles in an organic solvent with UV-light (λ = 254 nm) for 5 h. TiO₂ nanoparticles of 1 and 5 wt.% were dispersed using a beads mill method. Trimethoxytrifluor(propyl) silane was used as a dispersant to achieve stability. The effect of the UV-light irradiation on the TiO₂ nanosuspension was investigated by means of a Fourier transform nuclear magnetic resonance analyzer (FT-NMR). The dispersant was partially desorbed due to the interaction of UV light and the TiO₂/dispersant complex. Thus, an enhanced transparency and the absence of color were obtained for well-dispersed TiO₂ nanoparticles in an organic solvent.     

Transverse Acoustic Response Measured by AC-cut Quartz Transducer in Superfluid Helium-3

No Heading The transverse acoustic response in the B phase of superfluid ³He has been measured using an AC-cut quartz transducer at a pressure of 17.0 bar. The measurements were performed by a CW bridge method at a frequency of 9.3 MHz utilizing a quadrature hybrid and a high-frequency lock-in amplifier. By sweeping frequency around the resonance frequency of the transducer, in-phase and quadrature responses of the resonance were obtained from the output of the lock-in amplifier. We successfully separated the quality factor and the resonance frequency. Quality factor of the transducer had a small dip just below T_c and increased as _n by cooling, while its resonance frequency decreased sharply at T_c and gradually recovered to the normal state value in the low temperature limit.PACS numbers: 67.57.Jj, 43.58.+z.     

Label-free detection of proteins in crude cell lysate with antibody arrays by a surface plasmon resonance imaging technique

We established a label-free method of measuring proteins in crude cell lysate using antibody arrays and surface plasmon resonance (SPR) imaging. The refractivity of the running buffer was adjusted with that of the lysate to overcome the bulk effect. The chemistries of the fabricated arrays were investigated to reduce nonspecific adsorption on the array surface. We found that the hydrophilicity of the poly(ethylene glycol) moiety and lower electrostatic charge on the surface provided a specific measurement of antigen-antibody interaction. We validated the system by measuring the expression of eight proteins in the mouse brain and comparing the results to those by conventional Western blotting. The detection limit of the antibody array was approximately 30 ng/mL in crude cell lysate, on the same order as that of previous SPR research. The system enabled quick, label-free, and high-throughput analysis of abundant proteins with minimal sample volume ( approximately 200 muL). It is expected that our SPR antibody array will be applicable for direct protein expression profiling of cell lysate, as well as for cell phenotyping, food analysis, discovery of new biomarkers, and immunological disease diagnostics.     

Design and fabrication of a length-extensional mode rectangular X-cut quartz resonator with zero temperature coefficient.

Rectangular X-cut quartz crystal resonators with cut angles theta > 5.0 degrees and aspect ratios Rzy (= width 2z0/length 2y0) from 0.3 to 0.5 are investigated. The resonators oscillate mode is a length-extensional mode. A semiempirical frequency equation was derived from the stress expressed in terms of the trigonometric and the hyperbolic transcendental functions with constants estimated by the regression curve fit to the stress simulated by the finite-element method (FEM). Contours on which a point satisfies a zero first order temperature coefficient condition are shown in a cut angle theta and Rzy diagram. We proved that a fabricated resonator with Rzy = 0.400 and theta = 16.0 degrees, whose design parameter is located in the area of the contour, had a zero temperature coefficient.