A gradient PCR-based screen for use in site-directed mutagenesis

Site-directed mutagenesis is widely used to study protein and nucleic acid structure and function. Despite recent advancements in the efficiency of procedures for site-directed mutagenesis, the fraction of site-directed mutants by most procedures rarely exceeds 50% on a routine basis and is never 100%. Hence it is typically necessary to sequence two or three clones each time a site-directed mutant is constructed. We describe a simple and robust gradient-PCR-based screen for distinguishing site-directed mutants from the starting, unmutated plasmid. The procedure can use either purified plasmid DNA or colony PCR, starting from a single colony. The screen utilizes the primer used for mutagenesis and a common outside primer that can be used for all other mutants constructed with the same template. Over 30 site-specific mutants in a variety of templates were successfully screened and all of the mutations detected were subsequently confirmed by DNA sequencing. A single base pair mismatch could be detected in an oligonucleotide of 36 bases. Detection efficiency was relatively independent of starting template concentration and the nature of the outside primer used. (C) 2003 Elsevier Science (USA). All rights reserved.

Biaxial strain induced electrical inhomogenities and phase separation in the ferromagnetic metallic phase in thin films of La0.7Ca0.3MnO3: A scanning tunneling potentiometry/spectroscopy study

We have investigated the effect of biaxial strain on local electrical/electronic properties in thin films of La0.7Ca0.3MnO3 with varying degrees of biaxial strain in them. The local electrical properties were investigated as a function of temperature by scanning tunneling spectroscopy (STS) and scanning tunneling potentiometry (STP), along with the bulk probe like conductance fluctuations.The results indicate a positive correlation between the lattice mismatch biaxial strain and the local electrical/electronic inhomogenities observed in the strained sample. This is plausible since the crystal structure of the manganites interfere rather strongly with the magnetic/electronic degrees of freedom. Thus even a small imbalance (biaxial strain) can induce significant changes in the electrical properties of the system.

A scanning tunneling microscopy and potentiometry study of epitaxial thin films of La0.7Ca0.3MnO3

To investigate the role of grain boundaries and other growth related microstructure in manganite films, a scanning tunneling microscope is used to simultaneously probe surface topography and local potential distribution under current flow at nanometer level in films of epitaxial thin films of La0.7Ca0.3MnO3 deposited on single crystal SrTiO3 and NdGaO3 substrate by laser ablation. We have studied two types of films strained and strain relaxed. Thin (50nm) films (strained due to lattice mismatch between substrate and the film) show step growth (unit cell steps) and have very smooth surfaces. Relatively thicker films (strain relaxed, thickness 200nm) do not have these step growths and show rather smooth well connected grains. Charge transport in these films is not uniform on the nanometer level and is accompanied by potential jumps at the internal surfaces. In particular scattering from grain boundaries results in large variations in the local potential resulting in fields as high as 104-105V/cm located near the grain boundaries. We discuss the role of local strain and strain inhomogeneties in determining the current transport in these films and their resistance and magnetoresistivity. In this paper we attempt to correlate between bulk electronic properties with microscopic electronic conduction using scanning tunneling microscopy and scanning tunneling potentiometry.

Soft classification based Sub-pixel allocation model

Land cover (LC) refers to what is actually present on the ground and provide insights into the underlying solution for improving the conditions of many issues, from water pollution to sustainable economic development. One of the greatest challenges of modeling LC changes using remotely sensed (RS) data is of scale-resolution mismatch: that the spatial resolution of detail is less than what is required, and that this sub-pixel level heterogeneity is important but not readily knowable. However, many pixels consist of a mixture of multiple classes. The solution to mixed pixel problem typically centers on soft classification techniques that are used to estimate the proportion of a certain class within each pixel. However, the spatial distribution of these class components within the pixel remains unknown. This study investigates Orthogonal Subspace Projection - an unmixing technique and uses pixel-swapping algorithm for predicting the spatial distribution of LC at sub-pixel resolution. Both the algorithms are applied on many simulated and actual satellite images for validation. The accuracy on the simulated images is ~100%, while IRS LISS-III and MODIS data show accuracy of 76.6% and 73.02% respectively. This demonstrates the relevance of these techniques for applications such as urban-nonurban, forest-nonforest classification studies etc.

Improved utilisation of an HF transformer in DC–AC application

Power conversion using high frequency (HF) link converters is popular because of compact size and light weight of highfrequency transformer. This study focuses on improved utilisation of HF transformer in DC–AC applications. In practical application, the operating condition of the power converter deviates significantly from the designed considerations. These deviating factors are commutation requirements (dead-time, overlap), mismatch in device drops and presence of the fundamental frequency in load current. As a result, the HF transformer handles some amount of low-frequency components (including DC) other than desired HF components. This causes the operating point in B-H curve to shift away from its normal or idealised position and hence results poor utilisation of the HF transformer and unwanted losses. This study investigates the nature of the problem with experimental determination of approximate lumped parameter modelling and saturation behaviour (B-H curve) of the HF transformer. A simple closed-loop control algorithm with online tuning of the controller parameters is proposed to improve the utilisation of the isolation transformer. The simulation and experimental results are presented.

Relation between Local Structure and Glass Forming Ability of Liquid Alloys

An attempt has been made to describe the glass forming ability (GFA) of liquid alloys, using the concepts of the short range order (SRO) and middle range order (MRO) characterizing the liquid structure.A new approach to obtain good GFA of liquid alloys is based on the following four main factors: (1) formation of new SRO and competitive correlation with two or more kinds of SROs for crystallization, (2) stabilization of dense random packing by interaction between different types of SRO, (3) formation of stable cluster (SC) or middle range order (MRO) by harmonious coupling of SROs, and (4) difference between SRO characterizing the liquid structure and the near-neighbor environment in the corresponding equilibrium crystalline phases. The atomic volume mismatch estimated from the cube of the atomic radius was found to be a close relation with the minimum solute concentration for glass formation. This empirical guideline enables us to provide the optimum solute concentration for good GFA in some ternary alloys. Model structures, denoted by Bernal type and the Chemical Order type, were again tested in the novel description for the glass structure as a function of solute concentration. We illustrated the related energetics of the completion between crystal embryo and different types of SRO. Recent systematic measurements also provide that thermal diffusivity of alloys in the liquid state may be a good indicator of their GFA.

Nonlinear dielectric behavior in three-component ferroelectric superlattices

Three-component ferroelectric superlattices consisting of alternating layers of SrTiO3, BaTiO3, and CaTiO3 (SBC) with variable interlayer thickness were fabricated on Pt(111)/TiO2/SiO2/Si (100) substrates by pulsed laser deposition. The presence of satellite reflections in x-ray-diffraction analysis and a periodic concentration of Sr, Ba, and Ca throughout the film in depth profile of secondary ion mass spectrometry analysis confirm the fabrication of superlattice structures. The Pr (remnant polarization) and Ps (saturation polarization) of SBC superlattice with 16.4-nm individual layer thickness (SBC16.4) were found to be around 4.96 and 34 μC/cm2, respectively. The dependence of polarization on individual layer thickness and lattice strain were studied in order to investigate the size dependence of the dielectric properties. The dielectric constant of these superlattices was found to be much higher than the individual component layers present in the superlattice configuration. The relatively higher tunability ( ∼ 55%) obtained around 300 K indicates that the superlattice is a potential electrically tunable material for microwave applications at room temperature. The enhanced dielectric properties were thus discussed in terms of the interfacial strain driven polar region due to high lattice mismatch and electrostatic coupling due to polarization mismatch between individual layers.

The influence of Zr layer thickness on contact deformation and fracture in a ZrN-Zr multilayer coating

In order to understand the influence of ductile metal interlayer on the overall deformation behavior of metal/nitride multilayer, different configurations of metal and nitride layers were deposited and tested under indentation loading. To provide insight into the trends in deformation with multilayer spacings, an FEM model with elastic-perfect plastic metal layers alternate with an elastic nitride on top of an elastic-plastic substrate. The strong strain mismatch between the metal and nitride layers significantly alters the stress field under contact loading leading to micro-cracking in the nitride, large tensile stresses immediately below the contact, and a transition from columnar sliding in thin metal films to a more uniform bending and microcracking in thicker coatings.

Fabrication of Transparent BaNaB9O15 Glass-Microcrystal Composites

Transparent glasses in the BaO-Na2O-B2O3 (BNBO) system were fabricated via the conventional melt-quenching technique. The amorphous and the glassy nature of the as-quenched samples were confirmed by x-ray powder diffraction (XRD) and differential thermal analysis (DTA), respectively. Cyclic heat treatment of the as-quenched glasses yielded transparent glass-microcrystal composites. The volume fraction of the crystallites and their sizes could be easily controlled by this process. Heat-treated samples were highly transparent owing to the minimum mismatch between the refractive indices of the crystallites and the glass residual matrix. BNBO samples that were heat treated at 540A degrees C for 4 h for 10 cycles were found to be 60% to 70% transparent in the 500 nm to 900 nm wavelength range.

Distinct mechanisms of DNA repair in mycobacteria and their implications in attenuation of the pathogen growth

About a third of the human population is estimated to be infected with Mycobacterium tuberculosis. Emergence of drug resistant strains and the protracted treatment strategies have compelled the scientific community to identify newer drug targets, and to develop newer vaccines. In the host macrophages, the bacterium survives within an environment rich in reactive nitrogen and oxygen species capable of damaging its genome. Therefore, for its successful persistence in the host, the pathogen must need robust DNA repair mechanisms. Analysis of M. tuberculosis genome sequence revealed that it lacks mismatch repair pathway suggesting a greater role for other DNA repair pathways such as the nucleotide excision repair, and base excision repair pathways. In this article, we summarize the outcome of research involving these two repair pathways in mycobacteria focusing primarily on our own efforts. Our findings, using Mycobacterium smegmatis model, suggest that deficiency of various DNA repair functions in single or in combinations severely compromises their DNA repair capacity and attenuates their growth under conditions typically encountered in macrophages. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Stochastically driven instability in rotating shear flows

The origin of hydrodynamic turbulence in rotating shear flows is investigated, with particular emphasis on the flows whose angular velocity decreases but whose specific angular momentum increases with the increasing radial coordinate. Such flows are Rayleigh stable, but must be turbulent in order to explain the observed data. Such a mismatch between the linear theory and the observations/experiments is more severe when any hydromagnetic/magnetohydrodynamic instability and then the corresponding turbulence therein is ruled out. This work explores the effect of stochastic noise on such hydrodynamic flows. We essentially concentrate on a small section of such a flow, which is nothing but a plane shear flow supplemented by the Coriolis effect. This also mimics a small section of an astrophysical accretion disc. It is found that such stochastically driven flows exhibit large temporal and spatial correlations of perturbation velocities and hence large energy dissipations of perturbation, which presumably generate the instability. A range of angular velocity (Omega) profiles of the background flow, starting from that of a constant specific angular momentum (lambda = Omega r(2); r being the radial coordinate) to a constant circular velocity (v(phi) = Omega r), is explored. However, all the background angular velocities exhibit identical growth and roughness exponents of their perturbations, revealing a unique universality class for the stochastically forced hydrodynamics of rotating shear flows. This work, to the best of our knowledge, is the first attempt to understand the origin of instability and turbulence in three-dimensional Rayleigh stable rotating shear flows by introducing additive noise to the underlying linearized governing equations. This has important implications to resolve the turbulence problem in astrophysical hydrodynamic flows such as accretion discs.

Supervised matching of comments with news article segments

Comments constitute an important part of Web 2.0. In this paper, we consider comments on news articles. To simplify the task of relating the comment content to the article content the comments are about, we propose the idea of showing comments alongside article segments and explore automatic mapping of comments to article segments. This task is challenging because of the vocabulary mismatch between the articles and the comments. We present supervised and unsupervised techniques for aligning comments to segments the of article the comments are about. More specifically, we provide a novel formulation of supervised alignment problem using the framework of structured classification. Our experimental results show that structured classification model performs better than unsupervised matching and binary classification model.

Effect of stress orientation on microstructural evolution during creep of near-lamellar Ti-47Al-2Cr-2Nb

Microstructural evolution was studied in a near-lamellar two phase (alpha(2) + gamma) Ti-47Al-2Cr-2Nb alloy under high temperature creep and exposure conditions. The aim of this study was to probe the role of stress orientation, with respect to lamellar plates, on microstructural changes during primary creep. Creep testing was complemented with SEM and TEM based microstructural characterization. It was observed that retention of excess alpha(2) resulted in an unstable microstructure. Under stress and temperature, excess alpha(2) was lost and Cr-rich precipitates formed. Depending on stress orientation, the sequence of precipitates formed was different. alpha(2) loss was accompanied by formation of the non-equilibrium C14 Laves phase when lamellar plates were oriented parallel to the stress axis. In contrast, alpha(2) loss did not result in formation of the C14 phase in perpendicular samples. It was concluded that C14 formed preferentially in certain test orientations because of its effectiveness in relieving residual stresses in alpha(2) that arose from lattice misfit and modulus mismatch. (c) 2012 Elsevier B.V. All rights reserved.

Seismic site classification and correlation between standard penetration test N value and shear wave velocity for Lucknow city in Indo-Gangetic basin

Subsurface lithology and seismic site classification of Lucknow urban center located in the central part of the Indo-Gangetic Basin (IGB) are presented based on detailed shallow subsurface investigations and borehole analysis. These are done by carrying out 47 seismic surface wave tests using multichannel analysis of surface waves (MASW) and 23 boreholes drilled up to 30 m with standard penetration test (SPT) N values. Subsurface lithology profiles drawn from the drilled boreholes show low- to medium-compressibility clay and silty to poorly graded sand available till depth of 30 m. In addition, deeper boreholes (depth >150 m) were collected from the Lucknow Jal Nigam (Water Corporation), Government of Uttar Pradesh to understand deeper subsoil stratification. Deeper boreholes in this paper refer to those with depth over 150 m. These reports show the presence of clay mix with sand and Kankar at some locations till a depth of 150 m, followed by layers of sand, clay, and Kankar up to 400 m. Based on the available details, shallow and deeper cross-sections through Lucknow are presented. Shear wave velocity (SWV) and N-SPT values were measured for the study area using MASW and SPT testing. Measured SWV and N-SPT values for the same locations were found to be comparable. These values were used to estimate 30 m average values of N-SPT (N-30) and SWV (V-s(30)) for seismic site classification of the study area as per the National Earthquake Hazards Reduction Program (NEHRP) soil classification system. Based on the NEHRP classification, the entire study area is classified into site class C and D based on V-s(30) and site class D and E based on N-30. The issue of larger amplification during future seismic events is highlighted for a major part of the study area which comes under site class D and E. Also, the mismatch of site classes based on N-30 and V-s(30) raises the question of the suitability of the NEHRP classification system for the study region. Further, 17 sets of SPT and SWV data are used to develop a correlation between N-SPT and SWV. This represents a first attempt of seismic site classification and correlation between N-SPT and SWV in the Indo-Gangetic Basin.

Circulating power test setup for a PWM rectifier motor drive

Power converters burn-in test consumes large amount of energy, which increases the cost of testing, and certification, in medium and high power application. A simple test configuration to test a PWM rectifier induction motor drive, using a Doubly Fed Induction Machine (DFIM) to circulate power back to the grid for burn-in test is presented. The test configuration makes use of only one power electronic converter, which is the converter to be tested. The test method ensures soft synchronization of DFIM and Squirrel Cage Induction Machine (SCIM). A simple volt per hertz control of the drive is sufficient for conducting the test. To synchronize the DFIM with SCIM, the rotor terminal voltage of DFIM is measured and used as an indication of speed mismatch between DFIM and SCIM. The synchronization is done when the DFIM rotor voltage is at its minimum. Analysis of the DFIM characteristics confirms that such a test can be effectively performed with smooth start up and loading of the test setup. After synchronization is obtained, the speed command to SCIM is changed in order to load the setup in motoring or regenerative mode of operation. The experimental results are presented that validates the proposed test method.