bluuebunny/crossref_metadata_2025 huggingface dataset:
Pulsed wire discharge apparatus for mass production of copper nanopowders
Authors: Suematsu H. , Nishimura S. , Murai K. , Hayashi Y. , Suzuki T. , Nakayama T. , Jiang W. , Yamazaki A. , Seki K. , Niihara K.
Published: June 2007

A pulsed wire discharge (PWD) apparatus for the mass production of nanopowders has been developed. The apparatus has a continuous wire feeder, which is operated in synchronization with a discharging circuit. The apparatus is designed for operation at a maximum repetition rate of 1.4 Hz at a stored energy of 160 J. In the present study, Cu nanopowder was synthesized using the PWD apparatus and the performance of the apparatus was examined. Cu nanopowder of 2.0 g quantity was prepared in N2 gas at 100 kPa for 90 s. The particle size distribution of the Cu nanopowder was analyzed by transmission electron microscopy and the mean surface diameter was determined to be 65 nm. The ratio of the production mass of the powder to input energy was 362 g/kW h.
Investigation of eddy-viscosity models modified using discrete filters: A simplified “regularized variational multiscale model” and an “enhanced field model”
Authors: Jeanmart H. , Winckelmans G.
Published: June 2007

Subgrid-scale (SGS) models for large-eddy simulation (LES) having the formalism of an effective eddy-viscosity model, but that operates on a modified velocity field, are further evaluated and new ones are proposed. The modified field is obtained using regular filtering of the LES field carried out in physical space. This is actually done by using a discrete and compact operator (only using nearest neighbors values), eventually iterated; this ensures that the proper filtering behavior is preserved, even for near wall points. The first model investigated here is inspired by the variational multiscale approach originally proposed by T. J. Hughes et al. [Phys. Fluids 13, 505 (2001)]. Here, the modelling is simplified, leading to a SGS viscosity effect operating on the “small-scale LES field” that is obtained by subtracting the LES field from its filtered counterpart. Such a model (here called RVMM for short) was already proposed and partially evaluated {e.g., see G. S. Winckelmans and H. Jeanmart [Direct and Large-Scale Eddy Simulation IV (Kluwer, Dordrecht, 2001)] and H. Jeanmart and G. S. Winckelmans (CTR Proceedings of the Summer Program, 2002), the “model M2” of A. W. Vreman [Phys. Fluids 15, L61 (2003)], the “high-pass filtered Smagorinsky model” of S. Stolz et al. [Direct and Large-Eddy Simulation V (Kluwer, Dordrecht, 2004) and S. Stolz et al., Phys. Fluids 17, 065103 (2005)]}. The other model investigated here is an “enhanced field model” (EFM). The SGS viscosity model then operates on a LES field that is artificially enhanced at the small scales; that obtained by adding to the LES field the small-scale field. The two model families are presented in a unified way; they have a behavior that combines viscous and hyperviscous effects, while remaining simple and practical. They however do not naturally have the proper y3 near wall behavior for the SGS dissipation; hence, they need some near wall damping. To ensure the proper near-wall behavior, we use here the dynamic procedure (self-consistent for each model). The performance of both models is compared to that of other models (also dynamic): the Smagorinsky model, hyperviscosity models, and a hybrid model combining explicitly a Smagorinsky term and a hyperviscosity term. The cases here investigated are LES of decaying isotropic turbulence starting at Reλ=90 and LES of turbulent channel flow at Reτ=395. A good behavior of the RVMM and EFM, as compared to the others, is observed in all cases. They constitute an easily implemented and better alternative than the dynamic Smagorinsky model.
Optical emission spectroscopy characterization of oxygen plasma during degradation of Escherichia coli
Authors: Vujošević D. , Mozetič M. , Cvelbar U. , Krstulović N. , Milošević S.
Published: June 2007

Optical emission spectroscopy was applied for plasma characterization during sterilization of substrates contaminated with bacteria. The amount of 1010∕ml cells of Escherichia coli was carefully applied to glass substrates and exposed to oxygen plasma glow discharge at different pressures between 30 and 200Pa. Plasma was created in a glass discharge tube by an inductively coupled rf generator at the frequency of 27.12MHz and output power of about 250W. The electron temperature and plasma density were estimated with a double Langmuir probe. They were between 3 and 5eV and 2 and 35×1015m−3. Density of neutral oxygen atoms was measured with a catalytic probe, and was between 2 and 6×1021m−3. Optical emission spectroscopy was performed with a low resolution spectrometer. The emission from carbon monoxide and nitrogen molecules was used to monitor the evolution of bacteria degradation. Both signals expressed a well defined maximum corresponding to peak erosion of bacteria by plasma radicals. As the sterilization was accomplished, both CO and N2 lines fell below the detection limit of the spectrometer. The bacteria degradation was also monitored by scanning electron microscope (SEM) and culturing. The SEM images corresponded well with the evolution of CO and N2 lines so the optical emission spectroscopy found a reliable tool for monitoring the sterilization process.
Distribution of blocking temperatures in nano-oxide layers of specular spin valves
Authors: Ventura J. , Araujo J. P. , Sousa J. B. , Veloso A. , Freitas P. P.
Published: June 2007

Specular spin valves show enhanced giant magnetoresistive (GMR) ratio when compared to other, simpler, spin valve structures. The enhancement of GMR results from specular reflection in nano-oxide layers (NOLs) formed by the partial oxidation of the pinned and free layer. These oxides forming the NOL order antiferromagnetically (AFM) below a temperature T∼175 K. Here, we study the effects of the pinned layer magnetization and its domain structure on the AFM ordering of the NOL by performing field cooling measurements with different cooling fields (H0). We observe enhanced (reduced) exchange field and magnetoresistive ratio for H0>0(<0), i.e., parallel (antiparallel) to the pinned magnetization. These measurements allowed us to confirm the existence of a wide distribution of blocking temperatures (TB) in the NOL of specular spin valves, having a maximum at T≈175 K, and extending to NOL regions with TB as low as 15 K.
Be ion irradiation induced p- to n-type conversion in HgCdTe
Authors: Manchanda Rachna , Sharma R. K. , Malik A. , Pal R. , Dhaul Anuradha , Dutt M. B. , Basu P. K. , Thakur O. P.
Published: June 2007

Be ion implantation/irradiation in HgCdTe has been explored from the point of view of fabricating n region on p-type structures. Be implantation was carried out in vacancy doped p-type bulk HgCdTe crystals at 200KeV ion energy and a fluence of 1×1014ions∕cm2. Implanted samples were annealed at 150°C under nitrogen ambient for different durations. Be appears to be settling as an isoelectronic impurity in HgCdTe after the implant annealing process. A n-type region with reasonably good electron mobility is formed as a consequence of the redistribution of irradiation induced defects.
Differentiation of functionals with variables coupled by conservation constraints: Analysis through a fluid-dynamical model
Authors: Gál Tamás
Published: June 2007

A guide for obtaining the corresponding constrained functional derivatives for conservation constraints coupling the functional variables is given, with analyzing an application in liquid film dynamics. The use of constrained derivatives makes the proper account for constraints possible in time-dependent, nonequilibrium physical theories with physical equations not emerging as Euler-Lagrange equations, which is especially relevant with respect to the dynamics of complex liquids.
Diffusion of spheres in isotropic and nematic networks of rods: Electrostatic interactions and hydrodynamic screening
Authors: Kang Kyongok , Wilk A. , Patkowski A. , Dhont Jan K. G.
Published: June 2007

Translational diffusion of a small charged tracer sphere in isotropic and nematic suspensions of long and thin charged rods is investigated as a function of ionic strength and rod concentration. A theory for the diffusive properties of a small sphere is developed, where both (screened) hydrodynamic interactions and charge interactions between the tracer sphere and the rod network are analyzed. Hydrodynamic interactions are formulated in terms of the hydrodynamic screening length. As yet, there are no independent theoretical predictions for the hydrodynamic screening length for rod networks. Experimental tracer-diffusion data are presented for various ionic strengths as a function of the rod concentration, both in the isotropic and nematic states. Orientational order parameters are measured for the same ionic strengths as a function of the rod concentration. The hydrodynamic screening length is determined from these experimental data and scaling relations obtained from the above mentioned theory. For the isotropic networks, a master curve is found for the hydrodynamic screening length as a function of the rod concentration. For the nematic networks the screening length turns out to be a very sensitive function of the orientational order parameter.
κ
-Minkowski representations on Hilbert spaces
Authors: Agostini Alessandra
Published: June 2007

The algebra of functions on κ-Minkowski noncommutative space-time is studied as algebra of operators on Hilbert spaces. The representations of this algebra are constructed and classified. This new approach leads to a natural construction of integration in κ-Minkowski space-time in terms of the usual trace of operators.
Minimum entropy production principle from a dynamical fluctuation law
Authors: Maes Christian , Netočný Karel
Published: June 2007

The minimum entropy production principle provides an approximative variational characterization of close-to-equilibrium stationary states, both for macroscopic systems and for stochastic models. Analyzing the fluctuations of the empirical distribution of occupation times for a class of Markov processes, we identify the entropy production as the large deviation rate function, up to leading order when expanding around a detailed balance dynamics. In that way, the minimum entropy production principle is recognized as a consequence of the structure of dynamical fluctuations, and its approximate character gets an explanation. We also discuss the subtlety emerging when applying the principle to systems whose degrees of freedom change sign under kinematical time reversal.
Another proof of Gell-Mann and Low’s theorem
Authors: Molinari Luca Guido
Published: June 2007

The theorem by Gell-Mann and Low is a cornerstone in quantum field theory and zero-temperature many-body theory. The standard proof is based on Dyson’s time-ordered expansion of the propagator; a proof based on exact identities for the time propagator is here given.
Lagrangian systems with higher order constraints
Authors: Cendra H. , Grillo S. D.
Published: June 2007

A class of mechanical systems subject to higher order constraints (i.e., constraints involving higher order derivatives of the position of the system) are studied. We call them higher order constrained systems (HOCSs). They include simplified models of elastic rolling bodies, and also the so-called generalized nonholonomic systems (GNHSs), whose constraints only involve the velocities of the system (i.e., first order derivatives in the position of the system). One of the features of this kind of systems is that D’Alembert’s principle (or its nonlinear higher order generalization, the Chetaev’s principle) is not necessarily satisfied. We present here, as another interesting example of HOCS, systems subjected to friction forces, showing that those forces can be encoded in a second order kinematic constraint. The main aim of the paper is to show that every HOCS is equivalent to a GNHS with linear constraints, in a canonical way. That is to say, systems with higher order constraints can be described in terms of one with linear constraints in velocities. We illustrate this fact with a system with friction and with Rocard’s model [Dynamique Générale des Vibrations (1949), Chap. XV, p. 246 and L’instabilité en Mécanique; Automobiles, Avions, Ponts Suspendus (1954)] of a pneumatic tire. As a by-product, we introduce some applications on higher order tangent bundles, which we expect to be useful for the study of intrinsic aspects of the geometry of such bundles.
Optical extinction monitor using cw cavity enhanced detection
Authors: Kebabian Paul L. , Robinson Wade A. , Freedman Andrew
Published: June 2007

We present details of an apparatus capable of measuring optical extinction (i.e., scattering and/or absorption) with high precision and sensitivity. The apparatus employs one variant of cavity enhanced detection, specifically cavity attenuated phase shift spectroscopy, using a near-confocal arrangement of two high reflectivity (R∼0.9999) mirrors in tandem with an enclosed cell 26cm in length, a light emitting diode (LED), and a vacuum photodiode detector. The square wave modulated light from the LED passes through the absorption cell and is detected as a distorted wave form which is characterized by a phase shift with respect to the initial modulation. The amount of that phase shift is a function of fixed instrument properties—cell length, mirror reflectivity, and modulation frequency—and of the presence of a scatterer or absorber (air, particles, trace gases, etc.) within the cell. The specific implementation reported here employs a blue LED; the wavelength and spectral bandpass of the measurement are defined by the use of an interference filter centered at 440nm with a 20nm wide bandpass. The monitor is enclosed within a standard 19in. rack-mounted instrumentation box, weighs 10kg, and uses 70W of electrical power including a vacuum pump. Measurements of the phase shift induced by Rayleigh scattering from several gases (which range in extinction coefficient from 0.4–32Mm−1) exhibit a highly linear dependence (r2=0.99997) when plotted as the cotangent of the phase shift versus the expected extinction. Using heterodyne demodulation techniques, we demonstrate a detection limit of 0.04Mm−1 (4×10−10cm−1) (2σ) in 10s integration time and a base line drift of less than ±0.1Mm−1 over a 24h period. Detection limits decrease as the square root of integration time out to ∼150s.
PbTe nanocomposites synthesized from PbTe nanocrystals
Authors: Martin J. , Nolas G. S. , Zhang W. , Chen L.
Published: June 2007

Dense lead telluride (PbTe) nanocomposites were prepared from PbTe nanocrystals synthesized employing an aqueous solution-phase reaction. This approach reproducibly synthesizes 100–150nm nanocrystals with a high yield of over 2g per batch. Densification using spark plasma sintering dimensionally integrated nanoscale grains within a bulk matrix, resulting in a uniform dispersion of nonconglomerated nanocrystals. Transport properties of PbTe nanocomposites were evaluated through temperature dependent resistivity, Hall, Seebeck coefficient, and thermal conductivity measurements. These nanocomposites show an enhancement in the thermoelectric properties compared to bulk polycrystalline PbTe with similar carrier concentrations. Our results also indicate a strong sensitivity to stoichiometry, surface oxygen adsorption, and porosity.
Chemical sputtering of carbon films by argon ions and molecular oxygen at cryogenic temperatures
Authors: Hopf C. , Schlüter M. , Jacob W.
Published: June 2007

The experiments demonstrate the existence of a synergistic interaction of molecular oxygen and energetic ions with amorphous carbon leading to enhanced erosion; although the samples, amorphous hydrogenated carbon films, are not gasified by O2 at room temperature, additional ion bombardment at the same temperature leads to erosion rates that drastically exceed those of physical sputtering. Investigation of the temperature dependence from 400 down to 113K shows that the erosion rate increases with decreasing temperature.
Molecular and kinetic characterisation of sugarcane pyrophosphate: fructose-6-phosphate 1-phosphotransferase and its possible role in the sucrose accumulation phenotype
Authors: Groenewald Jan-Hendrik , Botha Frederik Coenraad
Published: June 2007

The amount of pyrophosphate: fructose-6-phosphate 1-phosphotransferase (PFP) activity in sugarcane internodal tissue is inversely correlated with sucrose content. To help elucidate this apparent role of PFP in sucrose accumulation in sugarcane we have determined its molecular and kinetic properties. Sugarcane PFP was purified 285-fold to a final specific activity of 4.23 µmol min–1 mg–1 protein. It contained two polypeptides of 63.2 and 58.0 kDa respectively, at near equal amounts that cross-reacted with potato PFP-α and –β antiserum. In gel filtration analyses the native enzyme eluted in three peaks of 129, 245 and 511 kDa, corresponding to dimeric, tetrameric and octameric forms, respectively and fructose 2,6-bisphosphate (Fru 2,6-P2) influenced this aggregation state. Both the glycolytic (forward) and gluconeogenic (reverse) reactions had relative broad pH optima between pH 6.7 and 8.0. The Fru 2,6-P2 saturation curves were hyperbolic with approximate Ka values of 69 and 82 nm for the forward and reverse reactions, respectively. The enzyme showed hyperbolic saturation curves for all its substrates with Km values comparable with that of other plant PFP, i.e. 150, 37, 39 and 460 µM for fructose 6-phosphate, inorganic pyrophosphate, fructose 1,6-bisphosphate and inorganic phosphate, respectively. Sugarcane PFP’s molecular and kinetic characteristics differed slightly from that of other plant PFP in that: (i) Fru 2,6-P2 directly induced the octameric state from the dimeric state; (ii) Fru 2,6-P2 shifted the pH optimum for the forward reaction to a slightly more basic pH; and (iii) Fru 2,6-P2 increased the Vmax for the forward and reverse reactions by similar amounts.
Transporters in starch synthesis
Authors: Martin Thomas , Ludewig Frank
Published: June 2007

Starch is synthesised and stored in plastids. In autotrophic tissues, the carbon skeletons and energy required for starch synthesis are directly available from photosynthesis. However, plastids of heterotrophic tissues require the metabolites for starch synthesis to be imported. Depending on plant species and tissue type, import is facilitated by several different plastid inner envelope metabolite transporters. Commonly, glucose-6-phosphate/phosphate translocators and adenylate translocators are used, but in the cereal endosperm, the role is carried out by ADP glucose transporters (Brittle1, BT1). This review predominantly focuses on transporters of the plastid inner envelope membrane. Their roles are discussed within an overview of starch synthesis. We also examine additional functions of these transporters according to our current knowledge.
Starch breakdown: recent discoveries suggest distinct pathways and novel mechanisms
Authors: Zeeman Samuel C. , Delatte Thierry , Messerli Gaëlle , Umhang Martin , Stettler Michaela , Mettler Tabea , Streb Sebastian , Reinhold Heike , Kötting Oliver
Published: June 2007

The aim of this article is to provide an overview of current models of starch breakdown in leaves. We summarise the results of our recent work focusing on Arabidopsis, relating them to other work in the field. Early biochemical studies of starch containing tissues identified numerous enzymes capable of participating in starch degradation. In the non-living endosperms of germinated cereal seeds, starch breakdown proceeds by the combined actions of α-amylase, limit dextrinase (debranching enzyme), β-amylase and α-glucosidase. The activities of these enzymes and the regulation of some of the respective genes on germination have been extensively studied. In living plant cells, additional enzymes are present, such as α-glucan phosphorylase and disproportionating enzyme, and the major pathway of starch breakdown appears to differ from that in the cereal endosperm in some important aspects. For example, reverse-genetic studies of Arabidopsis show that α-amylase and limit-dextrinase play minor roles and are dispensable for starch breakdown in leaves. Current data also casts doubt on the involvement of α-glucosidase. In contrast, several lines of evidence point towards a major role for β-amylase in leaves, which functions together with disproportionating enzyme and isoamylase (debranching enzyme) to produce maltose and glucose. Furthermore, the characterisation of Arabidopsis mutants with elevated leaf starch has contributed to the discovery of previously unknown proteins and metabolic steps in the pathway. In particular, it is now apparent that glucan phosphorylation is required for normal rates of starch mobilisation to occur, although a detailed understanding of this step is still lacking. We use this review to give a background to some of the classical genetic mutants that have contributed to our current knowledge.
Aspartate-444 Is Essential for Productive Substrate Interactions in a Neuronal Glutamate Transporter
Authors: Teichman Shlomit , Kanner Baruch I.
Published: May 2007

In the central nervous system, electrogenic sodium- and potassium-coupled glutamate transporters terminate the synaptic actions of this neurotransmitter. In contrast to acidic amino acids, dicarboxylic acids are not recognized by glutamate transporters, but the related bacterial DctA transporters are capable of transporting succinate and other dicarboxylic acids. Transmembrane domain 8 contains several residues that differ between these two types of transporters. One of these, aspartate-444 of the neuronal glutamate transporter EAAC1, is conserved in glutamate transporters, but a serine residue occupies this position in DctA transporters. When aspartate-444 is mutated to serine, cysteine, alanine, or even to glutamate, uptake of d-[3H]-aspartate as well as the inwardly rectifying steady-state currents induced by acidic amino acids is impaired. Even though succinate was not capable of inducing any steady-state transport currents, the dicarboxylic acid inhibited the sodium-dependent transient currents by the mutants with a neutral substitution at position 444. In the neutral substitution mutants inhibition of the transients was also observed with acidic amino acids. In the D444E mutant, acidic amino acids were potent inhibitors of the transient currents, whereas the apparent affinity for succinate was lower by at least three orders of magnitude. Even though L-aspartate could bind to D444E with a high apparent affinity, this binding resulted in inhibition rather than stimulation of the uncoupled anion conductance. Thus, a carboxylic acid–containing side chain at position 444 prevents the interaction of glutamate transporters with succinate, and the presence of aspartate itself at this position is crucial for productive substrate binding compatible with substrate translocation.
Tree Gibbs Sampler: identifying conserved motifs without aligning orthologous sequences
Authors: Cai Xiaohui , Hu Haiyan , Li Xiaoman Shawn
Published: June 2007

Summary: Tree Gibbs Sampler is a software for identifying motifs by simultaneously using the motif overrepresentation property and the motif evolutionary conservation property. It identifies motifs without depending on pre-aligned orthologous sequences, which makes it useful for the extraction of regulatory elements in multiple genomes of both closely related and distant species.

Availability: The Tree Gibbs Sampler software is freely downloadable at https://compbio.iupui.edu/xiaomanli/LiSoftware/retrieve.php?ID=tgs

Contact:  shawnli@iupui.edu
Logistic regression for disease classification using microarray data: model selection in a largepand smallncase
Authors: Liao J.G. , Chin Khew-Voon
Published: June 2007

Motivation: Logistic regression is a standard method for building prediction models for a binary outcome and has been extended for disease classification with microarray data by many authors. A feature (gene) selection step, however, must be added to penalized logistic modeling due to a large number of genes and a small number of subjects. Model selection for this two-step approach requires new statistical tools because prediction error estimation ignoring the feature selection step can be severely downward biased. Generic methods such as cross-validation and non-parametric bootstrap can be very ineffective due to the big variability in the prediction error estimate.

Results: We propose a parametric bootstrap model for more accurate estimation of the prediction error that is tailored to the microarray data by borrowing from the extensive research in identifying differentially expressed genes, especially the local false discovery rate. The proposed method provides guidance on the two critical issues in model selection: the number of genes to include in the model and the optimal shrinkage for the penalized logistic regression. We show that selecting more than 20 genes usually helps little in further reducing the prediction error. Application to Golub's leukemia data and our own cervical cancer data leads to highly accurate prediction models.

Availability: R library GeneLogit at http://geocities.com/jg_liao

Contact:  jl544@drexel.edu
Environmental forcing and Southern Ocean marine predator populations: effects of climate change and variability
Authors: Trathan P.N , Forcada J , Murphy E.J
Published: May 2007

The Southern Ocean is a major component within the global ocean and climate system and potentially the location where the most rapid climate change is most likely to happen, particularly in the high-latitude polar regions. In these regions, even small temperature changes can potentially lead to major environmental perturbations. Climate change is likely to be regional and may be expressed in various ways, including alterations to climate and weather patterns across a variety of time-scales that include changes to the long interdecadal background signals such as the development of the El Niño–Southern Oscillation (ENSO). Oscillating climate signals such as ENSO potentially provide a unique opportunity to explore how biological communities respond to change. This approach is based on the premise that biological responses to shorter-term sub-decadal climate variability signals are potentially the best predictor of biological responses over longer time-scales. Around the Southern Ocean, marine predator populations show periodicity in breeding performance and productivity, with relationships with the environment driven by physical forcing from the ENSO region in the Pacific. Wherever examined, these relationships are congruent with mid-trophic-level processes that are also correlated with environmental variability. The short-term changes to ecosystem structure and function observed during ENSO events herald potential long-term changes that may ensue following regional climate change. For example, in the South Atlantic, failure of Antarctic krill recruitment will inevitably foreshadow recruitment failures in a range of higher trophic-level marine predators. Where predator species are not able to accommodate by switching to other prey species, population-level changes will follow. The Southern Ocean, though oceanographically interconnected, is not a single ecosystem and different areas are dominated by different food webs. Where species occupy different positions in different regional food webs, there is the potential to make predictions about future change scenarios.
Infrequently transcribed long genes depend on the Set2/Rpd3S pathway for accurate transcription
Authors: Li Bing , Gogol Madelaine , Carey Mike , Pattenden Samantha G. , Seidel Chris , Workman Jerry L.
Published: June 2007

The presence of Set2-mediated methylation of H3K36 (K36me) correlates with transcription frequency throughout the yeast genome. K36me targets the Rpd3S complex to deacetylate transcribed regions and suppress cryptic transcription initiation at certain genes. Here, using a genome-wide approach, we report that the Set2–Rpd3S pathway is generally required for controlling acetylation at coding regions. When using acetylation as a functional readout for this pathway, we discovered that longer genes and, surprisingly, genes transcribed at lower frequency exhibit a stronger dependency. Moreover, a systematic screen using high-resolution tiling microarrays allowed us to identify a group of genes that rely on Set2–Rpd3S to suppress spurious transcripts. Interestingly, most of these genes are within the group that depend on the same pathway to maintain a hypoacetylated state at coding regions. These data highlight the importance of using the functional readout of histone codes to define the roles of specific pathways.
A novel view of the transcriptome revealed from gene trapping in mouse embryonic stem cells
Authors: Roma Guglielmo , Cobellis Gilda , Claudiani Pamela , Maione Francesco , Cruz Pedro , Tripoli Gaetano , Sardiello Marco , Peluso Ivana , Stupka Elia
Published: June 2007

Embryonic stem (ES) cells are pluripotent cell lines with the capacity of self-renewal and the ability to differentiate into specific cell types. We performed the first genome-wide analysis of the mouse ES cell transcriptome using ∼250,000 gene trap sequence tags deposited in public databases. We unveiled >8000 novel transcripts, mostly non-coding, and >1000 novel alternative and often tissue-specific exons of known genes. Experimental verification of the expression of these genes and exons by RT-PCR yielded a 70% validation rate. A novel non-coding transcript within the set studied showed a highly specific pattern of expression by in situ hybridization. Our analysis also shows that the genome presents gene trapping hotspots, which correspond to 383 known and 87 novel genes. These “hypertrapped” genes show minimal overlap with previously published expression profiles of ES cells; however, we prove by real-time PCR that they are highly expressed in this cell type, thus potentially contributing to the phenotype of ES cells. Although gene trapping was initially devised as an insertional mutagenesis technique, our study demonstrates its impact on the discovery of a substantial and unprecedented portion of the transcriptome.
Animal Cap Isolation from Xenopus laevis: Figure 1.
Authors: Sive Hazel L. , Grainger Robert M. , Harland Richard M.
Published: June 2007

INTRODUCTION

This protocol presents a method for isolating Xenopus laevis animal cap cells, which are cells situated around the animal (pigmented) pole of a blastula or very early gastrula-stage embryo. This tissue is fated to become cement gland/neurectoderm on the dorsal side of the embryo and epidermis on the ventral side. Animal caps are composed of pluripotent cells that can be induced to form endodermal, mesodermal, or ectodermal cell types, and can therefore serve as a useful substrate to assess the activity of various inducing factors. Caps from embryos that have been injected with expression constructs can also be removed and analyzed to assess the activity of various genes. In addition, caps can be used in conjugation (induction) assays.
Xenopus laevis Keller Explants: Figure 1.
Authors: Sive Hazel L. , Grainger Robert M. , Harland Richard M.
Published: June 2007

INTRODUCTION

The basic Keller explant is a rectangle of dorsal mesendoderm and ectoderm from an early-gastrula-stage Xenopus laevis embryo. It is ~60° to 90° wide, extending from the bottle cells to the animal pole. This protocol describes how to dissect, assemble, and cultivate Keller explants. The purpose of Keller explants was initially to allow observation of gastrulation movements, particularly convergent extension, in culture. This is difficult to do when explants curl up, but in Keller sandwiches, the explants are cultured flat, either as a single sheet (open-face explant) or more frequently as two sheets sandwiched together with their inner surfaces apposed (closed sandwich). Explants are cultured beneath a coverslip fragment or a glass bridge resting on silicone vacuum grease until the desired stage, usually during or after neurulation. Instead of involuting beneath the ectoderm, mesoderm elongates in a plane with adjacent ectoderm. Explants are made at the onset of gastrulation before significant vertical juxtaposition of ectoderm and mesoderm has occurred.
Dissection of Tightly Adhering Xenopus laevis Tissues by Trypsin Treatment
Authors: Sive Hazel L. , Grainger Robert M. , Harland Richard M.
Published: June 2007

INTRODUCTION

In older Xenopus laevis embryos (late gastrula and beyond), tissues begin to stick to one another and cannot be peeled apart. For assays requiring isolated tissues, it is necessary to separate such tissues enzymatically, which can be readily accomplished by mild trypsin treatment. Embryos are treated singly, or in small numbers, to avoid possible toxic effects of excessive trypsin digestion. This protocol describes a method for the separation of neural tissue from a neural-plate-stage embryo, although the technique can be adapted to many different tissue types. Enzymatically dissected tissue can be used in quantitative gene expression assays or in specification or induction assays.
Chromatin Immunoprecipitation (ChIP) on Unfixed Chromatin from Cells and Tissues to Analyze Histone Modifications
Authors: Wagschal Alexandre , Delaval Katia , Pannetier Maëlle , Arnaud Philippe , Feil Robert
Published: June 2007

INTRODUCTION

In cells and tissues, the histone proteins that constitute the nucleosomes can present multiple post-translational modifications, such as lysine acetylation, lysine and arginine methylation, serine phosphorylation, and lysine ubiquitination. On their own, or in combination, these covalent modifications on the core histones are thought to play essential roles in chromatin organization and gene expression in eukaryotes. Importantly, patterns of histone modifications may be somatically conserved and can, thereby, maintain locus-specific repression/activity in defined lineages, or throughout development. Indirect immunofluorescence studies on cultured cells have been pivotal in unraveling the roles of histone modifications. However, to address in detail what happens at specific sites in vivo, chromatin immunoprecipitation (ChIP) is the method of choice. Here, we describe how ChIP can be performed on non-fixed chromatin from animal cells or tissues (fresh or frozen) to analyze histone modifications at specific chromosomal sites. These protocols are suitable only for analyzing histones and their modifications. For other applications, chromatin immunoprecipitation should be performed on cross-linked chromatin.
A Behavioral Stress Assay in Drosophila Larvae: Figure 1.
Authors: Daubert Elizabeth A. , Condron Barry G.
Published: June 2007

INTRODUCTION

This protocol describes a simple behavioral assay designed to test the response of Drosophila larvae to a homeostatic insult: bright light. Treating normally photophobic larvae with bright light before placing them near a target food source reveals a greater latency to reach the target when compared to controls not receiving light treatment. This effect is reversible given a recovery period after light treatment, and it may reveal a method by which to measure behavioral plasticity and stress responses in fruit fly larvae.
TheArabidopsisATNRT2.7 Nitrate Transporter Controls Nitrate Content in Seeds
Authors: Chopin Franck , Orsel Mathilde , Dorbe Marie-France , Chardon Fabien , Truong Hoai-Nam , Miller Anthony J. , Krapp Anne , Daniel-Vedele Françoise
Published: June 2007

In higher plants, nitrate is taken up by root cells where Arabidopsis thaliana NITRATE TRANSPORTER2.1 (ATNRT2.1) chiefly acts as the high-affinity nitrate uptake system. Nitrate taken up by the roots can then be translocated from the root to the leaves and the seeds. In this work, the function of the ATNRT2.7 gene, one of the seven members of the NRT2 family in Arabidopsis, was investigated. High expression of the gene was detected in reproductive organs and peaked in dry seeds. β-Glucuronidase or green fluorescent protein reporter gene expression driven by the ATNRT2.7 promoter confirmed this organ specificity. We assessed the capacity of ATNRT2.7 to transport nitrate in Xenopus laevis oocytes or when it is expressed ectopically in mutant plants deficient in nitrate transport. We measured the impact of an ATNRT2.7 mutation and found no difference from the wild type during vegetative development. By contrast, seed nitrate content was affected by overexpression of ATNRT2.7 or a mutation in the gene. Finally, we showed that this nitrate transporter protein was localized to the vacuolar membrane. Our results demonstrate that ATNRT2.7 plays a specific role in nitrate accumulation in the seed.
Glucocorticoid receptor gene polymorphisms and susceptibility to rheumatoid arthritis
Authors: Donn Rachelle , Payne Debbie , Ray David
Published: May 2007

Background  A defect in hypothalamic–pituitary–adrenal (HPA) axis function has been suggested to contribute to susceptibility to rheumatoid arthritis (RA).

Objective  To investigate polymorphisms of the glucocorticoid receptor (GR) gene and determine any associations with RA.

Methods  Three GR polymorphisms that tag 95% of all haplotypes across the GR gene were genotyped. These are an intron B Bcl1 polymorphism, a ttg insertion/deletion within intron F (rs2307674) and the single nucleotide polymorphism (SNP) lying in the 3′ untranslated region of exon 9b (rs6198). The dye terminator‐based SNaPshot method or size resolution by capillary electrophoresis was performed. The study population comprised 198 UK Caucasian RA cases and 393 ethnically matched controls.

Results  No significant single point or haplotypic associations were found for GR polymorphisms with RA susceptibility. Furthermore, no evidence for GR polymorphisms with aspects of RA severity was seen.

Conclusion  In this study of the most comprehensive coverage of GR polymorphisms with RA, no significant contributing role for GR polymorphisms with RA was found.
Congenital hypothyroidism with goitre caused by new mutations in the thyroglobulin gene
Authors: Caputo Mariela , Rivolta Carina M. , Esperante Sebastían A. , Gruñeiro‐Papendieck Laura , Chiesa Ana , Pellizas Claudia G. , González‐Sarmiento Rogelio , Targovnik Héctor M.
Published: May 2007

Context  Thyroid dyshormonogenesis is associated with mutations in the thyroglobulin (TG) gene and characterized by normal organification of iodide and low serum TG. These mutations give rise to congenital goitrous hypothyroidism, transmitted in an autosomal recessive mode.

Objectives  The aim of this study was to identify new mutations in the TG gene in an attempt to increase the understanding of the molecular basis of this disorder. Three unrelated patients with marked impairment of TG synthesis were studied.

Methods  The promoter and the complete coding regions of the TG gene, along with the flanking intronic regions, were analysed by direct DNA sequencing.

Results  Four different inactivating TG mutations, three novel mutations (c.548G>A, p.C164Y; c.759–760insA, p.L234fsX237; c.6701C>A, p.A2215D) and one previously identified mutation (c.886C>T, p.R277X) were identified. Multiple sequence alignment study revealed that the wild‐type cysteine residue at position 164 is strictly conserved in the TG of all the species analysed, whereas the wild‐type alanine residue at position 2215 is well conserved in the TG and acetylcholinesterase (AChE) of all the species analysed except in rabbit AChE, in which it is substituted by glutamic acid.

Conclusions  We report three patients with congenital hypothyroidism with goitre caused by two compound heterozygous mutations, p.C164Y/p.L234fsX237 and p.R277X/p.A2215D, and one homozygous mutation, p.R277X, in the TG gene. To our knowledge this is the first report of the presence of a nucleotide insertion mutation in the TG gene.
Uterine leiomyosarcomas are characterized by high p16, p53 and MIB1 expression in comparison with usual leiomyomas, leiomyoma variants and smooth muscle tumours of uncertain malignant potential
Authors: O'Neill C J , McBride H A , Connolly L E , McCluggage W G
Published: June 2007

Aims:  It has been suggested that p16 is overexpressed in uterine leiomyosarcomas in comparison with leiomyomas. In this study, p16 immunohistochemical expression was assessed in a variety of uterine smooth muscle tumours, including usual leiomyomas, leiomyoma variants, smooth muscle tumours of uncertain malignant potential (STUMPs) and leiomyosarcomas. The aim was to ascertain whether there are differences in p16 expression between these groups and whether p16 is of potential value in the assessment of problematic uterine smooth muscle neoplasms. p16 expression was also compared with that of p53 and MIB1.

Methods and results:  Cases of usual leiomyoma (n = 10), leiomyoma variants (n = 27), STUMP (n = 4) and leiomyosarcoma (n = 22) were subject to p16, p53 and MIB1 immunohistochemistry. For p16, cases were evaluated with respect to both staining distribution and intensity. There was a statistically significant difference in p16 distribution (P < 0.001) and intensity (P = 0.001) between leiomyosarcomas and the other groups. There was no difference in p16 expression between usual leiomyomas, leiomyoma variants and STUMPs. There were also statistically significant differences in p53 (P = 0.014) and MIB1 (P < 0.001) immunoreactivity between leiomyosarcomas and the other groups.

Conclusions:  p16 is overexpressed in uterine leiomyosarcomas compared with leiomyomas, benign leiomyoma variants and STUMPs, suggesting that p16 may be implicated in the pathogenesis of malignant uterine smooth muscle neoplasms. p16, in combination with p53 and MIB1, may be of value as an adjunct to morphological examination in the assessment of problematic uterine smooth muscle tumours, although further large‐scale studies with follow‐up are necessary to confirm this.
Reactive astrocytes and activated microglial cells express EAAT1, but not EAAT2, reflecting a neuroprotective potential following ischaemia
Authors: Beschorner R , Simon P , Schauer N , Mittelbronn M , Schluesener H J , Trautmann K , Dietz K , Meyermann R
Published: June 2007

Aims: Glutamate receptor antagonists have failed clinical stroke trials and it has been proposed that the action of N‐methyld‐aspartate receptors is necessary for neuronal survival. Thus, excitatory amino acid transporters (EAATs) might be a promising therapeutic target. The aim of this study was to investigate glial expression of EAATs following ischaemia.

Methods and results: Expression of EAAT1 (GLAST) and EAAT2 (Glt‐1) in 24 cases of ischaemia was examined by immunohistochemistry. Cortical expression of both EAATs in the lesion decreased within 24 h (P < 0.01, each). Whereas EAAT1+ white matter cells increased 18‐fold (P < 0.05) within 24 h in the lesion and remained elevated for months in adjacent (469‐fold,P < 0.01) and remote areas (20‐fold,P < 0.05), EAAT2+ white matter cells were equivalent in ischaemia and controls. In the first week after stroke mainly activated (ramified and amoeboid) microglia expressed EAAT1, whereas monocytic cells in perivascular spaces and foamy macrophages lacked EAAT1. After more than 1 week, predominantly reactive astrocytes expressed EAAT1.

Conclusions: Microglial EAAT1 expression is restricted to the early/intermediate stage of activation and blood‐derived (perivascular) monocytes do not contribute to EAAT1+ cells following ischaemia. Whether a pharmacological increase in glial EAAT expression may compensate for loss of cortical EAAT expression and reduce neuronal damage following stroke requires investigation by further functional studies.
The wave forms of temporomandibular joint sound clicking and crepitation
Authors: WIDMALM S.E. , WILLIAMS W.J. , ADAMS B.S.
Published: May 2007

summary
The aim of the present study was to determine the sound wave forms which correspond to auscultatory findings of temporomandibular joint (TMJ) clicking and crepitation. Such knowledge is important when selecting parts of digital recordings for spectral analysis. Electronic digital recordings were made with a sampling rate of 44 100 Hz from 60 subjects, including 51 patients referred for suspected rheumatological disease and nine healthy subjects. Accelerometers with the bandwidth 20–3600 Hz were used for all subjects and complementary recordings were made from a subgroup of nine subjects using a measurement microphone with the bandwidth 20–20 000 Hz. The clicking sounds could be classified into different types according to differences in temporal period duration (T) as measured on the analogue display. One type of clicking, found in 51% of the patients, had a T of 2–20 ms. Another type, found in 70% of the subjects, had a T of less than 1 ms, often as low 0.2 ms. This type of clicking was not seen at all in the analogue display if the sampling rate was below 3 000 Hz. The character of the two types of clicking differed: the short duration sounds had a very high pitch, while the pitch of the longer duration sound was lower. Crepitation was found in 63% of the subjects and was observed to be composed of a series of short duration sounds, occurring with brief (less than 10 ms) intervals. It is concluded that the accelerometer (or microphone) bandwidth should cover the entire audible range (20–20 000 Hz), and that sampling rates must be much higher than 3000 Hz, and preferably greater than 10 000 Hz, before the true significance of electronically recorded joint sounds/vibrations can be determined.
Effect of occlusal pressure on the histopathological changes in denture supporting tissues
Authors: HARA T. , SATO T. , & K. NAKASHIMA , MINAGI S.
Published: May 2007

summary
Histopathological changes in the denture supporting tissues in relation to the occlusal pressure exerted through an experimental denture base were studied. An experimental denture, which prescribed the occlusal pressure by the amount of subsidence of the denture base (100, 50 and 13 μm), was con‐ structed for the palate of the molar region of rat. Histopathological changes caused by the occlusal pressure were observed, and the intensity of the pressure was also estimated. In the 100 μm subsidence group, compression of epithelium and lamina propria mucosae, inflammatory change and bone resorption were caused by the occlusal pressure. In the 50 μm subsidence group, compression of epithelium and lamina propria mucosae and bone resorption were observed, while in the 13 μm subsidence group, no evidence of inflammatory change and bone resorption were observed. The epithelium and lamina propria mucosae recovered histopathologically depending on the gradual decrease of the occlusal pressure result ing from bone resorption, whereas no quantitative recovery of the resorbed bone tissue was observed in the 100 and 50 μm subsidence groups. From the results observed in this study, a high correlation was observed between the histopathological changes under a denture base and the intensity of the occlusal pressure, and the possibility of the existence of the threshold for bone resorption induced by occlusal pressure was suggested.