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.     

An IR study on ion-specific and solvent-specific swelling of poly(N-vinyl-2-pyrrolidone) gel

Swelling degrees of poly(N-vinyl-2-pyrrolidone) (PVP) gel were measured in aqueous salt solutions and in water/organic solvent mixtures to find marked ion- and solvent-specificities. In order to investigate any correlation of those specificities with hydration or solvation of PVP, IR spectra band of the CO group was monitored by means of ATR method both for PVP gel and the relevant solution systems. Dependence of the peak frequency on the swelling ratio suggested that hydration of PVP carbonyl group in deswollen gel systems is different from that in the corresponding solution systems. In the solution systems, PVP carbonyl band showed a high-wavenumber shift for deswollen systems, which can be well correlated with changes in water proton charge through ionic hydration and with Gutmann's acceptor number of organic solvents. In the deswollen gel systems, the CO band showed a low-wavenumber shift, suggesting a strong hydration or doubly hydrated state. This unexpected behavior was interpreted by assuming an intermolecular hydrogen bond of two carbonyl groups intermediated by water molecules.     

Athermal silica-based arrayed-waveguide grating multiplexer

A temperature dependent channel wavelength shift in a silica-based arrayed-waveguide grating multiplexer is successfully suppressed from 0.95 to 0.05 nm in the 0-85°C temperature range, which means that it can be used in practical WDM systems without the need for temperature control     

A contribution to the UV degradation of polyurethanes

The effects of the nature of the polyols used in polyurethane (PUR) synthesis on the structural transformations after ageing by UV irradiation has been studied. The investigations were performed on PUR prepared from polyester and polyether diol oligomers. The characterization of the samples before and after ageing in view of the structural changes, which influence the course of the photooxidative degradation and photocrosslinking as well as in view of changes of mechanical properties has been done. Comparative investigations were performed by spectroscopic and viscometric measurements. The results show pronounced heterogeneity of the degradative reactions, including the existence of crosslinking processes and producing thus very inhomogeneous polymeric material. The course and the intensity of photooxidative degradation of PUR differ together with changes of mechanical properties depending whether polyester or polyether polyol have been used. The processes of photooxidative degradation is less expressed in polyesterurethane then in polyurethane based on polyether, under the same conditions of experiments. Different structures of polyester diols caused the various ageing behaviour of PUR too.     

Experimental measurement on tangential momentum accommodation coefficient in a single microtube

The tangential momentum accommodation coefficient (TMAC) was investigated experimentally from the mass flow rate through a single microtube under the slip flow and the early part of the transition regime. The measurements were carried out by the constant-volume method under the mean Knudsen number smaller than 0.3, which is based on the mean pressure of the inlet and the outlet of the microtube, to apply the second-order slip boundary condition. To measure TMACs on various materials, quite large microtube was employed, which require the reduction in leakage. TMAC was obtained from the slip coefficient determined by the relation of the mass flow rate to the mean Knudsen number. The obtained mass flow rate was well explained by the theoretical equation. TMACs of deactivated-fused silica with argon, nitrogen, and oxygen were measured, showing the tangential momentum was not accommodated completely to the surface, and the values showed good agreement with previous studies. From the comparison between microtubes with different inner diameter, it is showed that TMAC is determined mainly by gas species and surface material.     

Color system for skin color extraction

This paper describes a skin color extraction method which has high speed and high accuracy. There were many systems which could achieve the requirements, but they were useless due to tough limitations such as using two cameras to recognize correct coordinates, running only a specific background, wearing several gears, learning the users detailed information in advance, making a personalized database, and so on. In contrast, our approach, which uses the YIQ color system with a background estimation method for skin color extraction, achieved a good performance more than 30 fps, with strong robustness in complicated and poorly illuminated environments. This work was presented in part at the 12th International Symposium on Artificial Life and Robotics, Oita, Japan, January 25–27, 2007     

Application of full-factorial design for optimization of the moisture determination in wheat flour using infrared scales

We studied the parameters that may affect the analysis of the moisture content in wheat flour using infrared scales. The parameters were studied by one-factor at a time methodology and then, by full-factorial design (2³). The best condition using infrared scales occurred at 120 °C, with a sample weight of 1.5 g and an exposure time to infrared radiation of 6 min. We observed the interaction effect between the three factors studied. The analysis time was 60 times lower than classical method. The optimized method for determination of the moisture content in wheat flour using infrared scales was validated to range between 9 and 17 %.     

A two-way coupled multiscale model for predicting damage-associated performance of asphaltic roadways

This paper presents a quasi-static multiscale computational model with its verification and rational applications to mechanical behavior predictions of asphaltic roadways that are subject to viscoelastic deformation and fracture damage. The multiscale model is based on continuum thermo-mechanics and is implemented using a finite element formulation. Two length scales (global and local) are two-way coupled in the model framework by linking a homogenized global scale to a heterogeneous local scale representative volume element. With the unique multiscaling and the use of the finite element technique, it is possible to take into account the effect of material heterogeneity, viscoelasticity, and anisotropic damage accumulation in the small scale on the overall performance of larger scale structures. Along with the theoretical model formulation, two example problems are shown: one to verify the model and its computational benefits through comparisons with analytical solutions and single-scale simulation results, and the other to demonstrate the applicability of the approach to model general roadway structures where material viscoelasticity and cohesive zone fracture are involved.     

Discovery of a Novel Scaffold as an Indoleamine 2,3‐Dioxygenase 1 (IDO1) Inhibitor Based on the Pyrrolopiperazinone Alkaloid,Longamide B

Indoleamine 2,3‐dioxygenase 1 (IDO1) has emerged as a key target for cancer therapy, as IDO1 plays a critical role in the capacity of tumor cells to evade the immune system. The pyrrolopiperazinone alkaloid longamide B and its derivatives were identified as novel IDO1 inhibitors based on docking studies and small library synthesis. The thioamide derivative showed higher IDO1 inhibitory activity than longamide B, and displayed an activity similar to that of a representative IDO1 inhibitor, 1‐methyl‐tryptophan. These results suggest that the pyrrolopiperazinone scaffold of longamide B could be used in the development of IDO1 inhibitors.     

Preparation of polymeric nanocapsules via nano-sized poly(methoxydiethyleneglycol methacrylate) colloidal templates

Polymeric nanocapsules are attractive devices with a number of potential applications. In the present contribution we describe a method for nanocapsule preparation which is based on the formation of nanosized templates (mesoglobules, prepared from thermo-responsive poly(methoxydiethyleneglycol methacrylate)s, PDEGMA). These mesoglobules were coated with a cross-linked shell formed by pseudo-seeded radical polymerization of either N-isopropylacrylamide or 2-hydroxyethylmethacrylate in the presence of a cross-linking agent. Dissolution and removal of templates were achieved by extensive dialysis against water at temperatures below the LCST of PDEGMA. The obtained nanocapsules were visualized by transmission electron microscopy and their dimensions were determined by dynamic light scattering. The differences in the morphology of the nanocapsules were attributed to the different structures of the cross-linked membranes.