The baculovirus expression vector system: A commercial manufacturing platform for viral vaccines and gene therapy vectors.

PubMed

Felberbaum, Rachael S

2015-05-01

The baculovirus expression vector system (BEVS) platform has become an established manufacturing platform for the production of viral vaccines and gene therapy vectors. Nine BEVS-derived products have been approved - four for human use (Cervarix(®), Provenge(®), Glybera(®) and Flublok(®)) and five for veterinary use (Porcilis(®) Pesti, BAYOVAC CSF E2(®), Circumvent(®) PCV, Ingelvac CircoFLEX(®) and Porcilis(®) PCV). The BEVS platform offers many advantages, including manufacturing speed, flexible product design, inherent safety and scalability. This combination of features and product approvals has previously attracted interest from academic researchers, and more recently from industry leaders, to utilize BEVS to develop next generation vaccines, vectors for gene therapy, and other biopharmaceutical complex proteins. In this review, we explore the BEVS platform, detailing how it works, platform features and limitations and important considerations for manufacturing and regulatory approval. To underscore the growth in opportunities for BEVS-derived products, we discuss the latest product developments in the gene therapy and influenza vaccine fields that follow in the wake of the recent product approvals of Glybera(®) and Flublok(®), respectively. We anticipate that the utility of the platform will expand even further as new BEVS-derived products attain licensure. Finally, we touch on some of the areas where new BEVS-derived products are likely to emerge. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Improved Production Efficiency of Virus-Like Particles by the Baculovirus Expression Vector System

PubMed Central

Bárcena, Juan; Nuñez, Maria del Carmen; Martínez-Alonso, Diego; Dudognon, Benoit; Guijarro, Eva; Escribano, José M.

2015-01-01

Vaccines based on virus-like particles (VLPs) have proven effective in humans and animals. In this regard, the baculovirus expression vector system (BEVS) is one of the technologies of choice to generate such highly immunogenic vaccines. The extended use of these vaccines for human and animal populations is constrained because of high production costs, therefore a significant improvement in productivity is crucial to ensure their commercial viability. Here we describe the use of the previously described baculovirus expression cassette, called TB, to model the production of two VLP-forming vaccine antigens in insect cells. Capsid proteins from porcine circovirus type 2 (PCV2 Cap) and from the calicivirus that causes rabbit hemorrhagic disease (RHDV VP60) were expressed in insect cells using baculoviruses genetically engineered with the TB expression cassette. Productivity was compared to that obtained using standard counterpart vectors expressing the same proteins under the control of the polyhedrin promoter. Our results demonstrate that the use of the TB expression cassette increased the production yields of these vaccine antigens by around 300% with respect to the standard vectors. The recombinant proteins produced by TB-modified vectors were fully functional, forming VLPs identical in size and shape to those generated by the standard baculoviruses, as determined by electron microscopy analysis. The use of the TB expression cassette implies a simple modification of the baculovirus vectors that significantly improves the cost efficiency of VLP-based vaccine production, thereby facilitating the commercial viability and broad application of these vaccines for human and animal health. PMID:26458221

Improved Production Efficiency of Virus-Like Particles by the Baculovirus Expression Vector System.

PubMed

López-Vidal, Javier; Gómez-Sebastián, Silvia; Bárcena, Juan; Nuñez, Maria del Carmen; Martínez-Alonso, Diego; Dudognon, Benoit; Guijarro, Eva; Escribano, José M

2015-01-01

Vaccines based on virus-like particles (VLPs) have proven effective in humans and animals. In this regard, the baculovirus expression vector system (BEVS) is one of the technologies of choice to generate such highly immunogenic vaccines. The extended use of these vaccines for human and animal populations is constrained because of high production costs, therefore a significant improvement in productivity is crucial to ensure their commercial viability. Here we describe the use of the previously described baculovirus expression cassette, called TB, to model the production of two VLP-forming vaccine antigens in insect cells. Capsid proteins from porcine circovirus type 2 (PCV2 Cap) and from the calicivirus that causes rabbit hemorrhagic disease (RHDV VP60) were expressed in insect cells using baculoviruses genetically engineered with the TB expression cassette. Productivity was compared to that obtained using standard counterpart vectors expressing the same proteins under the control of the polyhedrin promoter. Our results demonstrate that the use of the TB expression cassette increased the production yields of these vaccine antigens by around 300% with respect to the standard vectors. The recombinant proteins produced by TB-modified vectors were fully functional, forming VLPs identical in size and shape to those generated by the standard baculoviruses, as determined by electron microscopy analysis. The use of the TB expression cassette implies a simple modification of the baculovirus vectors that significantly improves the cost efficiency of VLP-based vaccine production, thereby facilitating the commercial viability and broad application of these vaccines for human and animal health.

Re-engineering adenovirus vector systems to enable high-throughput analyses of gene function.

PubMed

Stanton, Richard J; McSharry, Brian P; Armstrong, Melanie; Tomasec, Peter; Wilkinson, Gavin W G

2008-12-01

With the enhanced capacity of bioinformatics to interrogate extensive banks of sequence data, more efficient technologies are needed to test gene function predictions. Replication-deficient recombinant adenovirus (Ad) vectors are widely used in expression analysis since they provide for extremely efficient expression of transgenes in a wide range of cell types. To facilitate rapid, high-throughput generation of recombinant viruses, we have re-engineered an adenovirus vector (designated AdZ) to allow single-step, directional gene insertion using recombineering technology. Recombineering allows for direct insertion into the Ad vector of PCR products, synthesized sequences, or oligonucleotides encoding shRNAs without requirement for a transfer vector Vectors were optimized for high-throughput applications by making them "self-excising" through incorporating the I-SceI homing endonuclease into the vector removing the need to linearize vectors prior to transfection into packaging cells. AdZ vectors allow genes to be expressed in their native form or with strep, V5, or GFP tags. Insertion of tetracycline operators downstream of the human cytomegalovirus major immediate early (HCMV MIE) promoter permits silencing of transgenes in helper cells expressing the tet repressor thus making the vector compatible with the cloning of toxic gene products. The AdZ vector system is robust, straightforward, and suited to both sporadic and high-throughput applications.

On the use of finite difference matrix-vector products in Newton-Krylov solvers for implicit climate dynamics with spectral elements

DOE PAGES

Woodward, Carol S.; Gardner, David J.; Evans, Katherine J.

2015-01-01

Efficient solutions of global climate models require effectively handling disparate length and time scales. Implicit solution approaches allow time integration of the physical system with a step size governed by accuracy of the processes of interest rather than by stability of the fastest time scales present. Implicit approaches, however, require the solution of nonlinear systems within each time step. Usually, a Newton's method is applied to solve these systems. Each iteration of the Newton's method, in turn, requires the solution of a linear model of the nonlinear system. This model employs the Jacobian of the problem-defining nonlinear residual, but thismore » Jacobian can be costly to form. If a Krylov linear solver is used for the solution of the linear system, the action of the Jacobian matrix on a given vector is required. In the case of spectral element methods, the Jacobian is not calculated but only implemented through matrix-vector products. The matrix-vector multiply can also be approximated by a finite difference approximation which may introduce inaccuracy in the overall nonlinear solver. In this paper, we review the advantages and disadvantages of finite difference approximations of these matrix-vector products for climate dynamics within the spectral element shallow water dynamical core of the Community Atmosphere Model.« less

Design and construction of functional AAV vectors.

PubMed

Gray, John T; Zolotukhin, Serge

2011-01-01

Using the basic principles of molecular biology and laboratory techniques presented in this chapter, researchers should be able to create a wide variety of AAV vectors for both clinical and basic research applications. Basic vector design concepts are covered for both protein coding gene expression and small non-coding RNA gene expression cassettes. AAV plasmid vector backbones (available via AddGene) are described, along with critical sequence details for a variety of modular expression components that can be inserted as needed for specific applications. Protocols are provided for assembling the various DNA components into AAV vector plasmids in Escherichia coli, as well as for transferring these vector sequences into baculovirus genomes for large-scale production of AAV in the insect cell production system.

A vector-product information retrieval system adapted to heterogeneous, distributed computing environments

NASA Technical Reports Server (NTRS)

Rorvig, Mark E.

1991-01-01

Vector-product information retrieval (IR) systems produce retrieval results superior to all other searching methods but presently have no commercial implementations beyond the personal computer environment. The NASA Electronic Library Systems (NELS) provides a ranked list of the most likely relevant objects in collections in response to a natural language query. Additionally, the system is constructed using standards and tools (Unix, X-Windows, Notif, and TCP/IP) that permit its operation in organizations that possess many different hosts, workstations, and platforms. There are no known commercial equivalents to this product at this time. The product has applications in all corporate management environments, particularly those that are information intensive, such as finance, manufacturing, biotechnology, and research and development.

Vector Meson Production at Hera

NASA Astrophysics Data System (ADS)

Szuba, Dorota

The diffractive production of vector mesons ep→eVMY, with VM=ρ0, ω, ϕ, J/ψ, ψ‧ or ϒ and with Y being either the scattered proton or a low mass hadronic system, has been extensively investigated at HERA. HERA offers a unique opportunity to study the dependences of diffractive processes on different scales: the mass of the vector meson, mVM, the centre-of-mass energy of the γp system, W, the photon virtuality, Q2 and the four-momentum transfer squared at the proton vertex, |t|. Strong interactions can be investigated in the transition from the hard to the soft regime, where the confinement of quarks and gluons occurs.

Vector systems for prenatal gene therapy: principles of retrovirus vector design and production.

PubMed

Howe, Steven J; Chandrashekran, Anil

2012-01-01

Vectors derived from the Retroviridae family have several attributes required for successful gene delivery. Retroviral vectors have an adequate payload size for the coding regions of most genes; they are safe to handle and simple to produce. These vectors can be manipulated to target different cell types with low immunogenicity and can permanently insert genetic information into the host cells' genome. Retroviral vectors have been used in gene therapy clinical trials and successfully applied experimentally in vitro, in vivo, and in utero.

Higher-dimensional generalizations of the Watanabe–Strogatz transform for vector models of synchronization

NASA Astrophysics Data System (ADS)

Lohe, M. A.

2018-06-01

We generalize the Watanabe–Strogatz (WS) transform, which acts on the Kuramoto model in d  =  2 dimensions, to a higher-dimensional vector transform which operates on vector oscillator models of synchronization in any dimension , for the case of identical frequency matrices. These models have conserved quantities constructed from the cross ratios of inner products of the vector variables, which are invariant under the vector transform, and have trajectories which lie on the unit sphere S d‑1. Application of the vector transform leads to a partial integration of the equations of motion, leaving independent equations to be solved, for any number of nodes N. We discuss properties of complete synchronization and use the reduced equations to derive a stability condition for completely synchronized trajectories on S d‑1. We further generalize the vector transform to a mapping which acts in and in particular preserves the unit ball , and leaves invariant the cross ratios constructed from inner products of vectors in . This mapping can be used to partially integrate a system of vector oscillators with trajectories in , and for d  =  2 leads to an extension of the Kuramoto system to a system of oscillators with time-dependent amplitudes and trajectories in the unit disk. We find an inequivalent generalization of the Möbius map which also preserves but leaves invariant a different set of cross ratios, this time constructed from the vector norms. This leads to a different extension of the Kuramoto model with trajectories in the complex plane that can be partially integrated by means of fractional linear transformations.

Auto and hetero-associative memory using a 2-D optical logic gate

NASA Technical Reports Server (NTRS)

Chao, Tien-Hsin (Inventor)

1992-01-01

An optical system for auto-associative and hetero-associative recall utilizing Hamming distance as the similarity measure between a binary input image vector V(sup k) and a binary image vector V(sup m) in a first memory array using an optical Exclusive-OR gate for multiplication of each of a plurality of different binary image vectors in memory by the input image vector. After integrating the light of each product V(sup k) x V(sup m), a shortest Hamming distance detection electronics module determines which product has the lowest light intensity and emits a signal that activates a light emitting diode to illuminate a corresponding image vector in a second memory array for display. That corresponding image vector is identical to the memory image vector V(sup m) in the first memory array for auto-associative recall or related to it, such as by name, for hetero-associative recall.

Construction of fusion vectors of corynebacteria: expression of glutathione-S-transferase fusion protein in Corynebacterium acetoacidophilum ATCC 21476.

PubMed

Srivastava, Preeti; Deb, J K

2002-07-02

A series of fusion vectors containing glutathione-S-transferase (GST) were constructed by inserting GST fusion cassette of Escherichia coli vectors pGEX4T-1, -2 and -3 in corynebacterial vector pBK2. Efficient expression of GST driven by inducible tac promoter of E. coli was observed in Corynebacterium acetoacidophilum. Fusion of enhanced green fluorescent protein (EGFP) and streptokinase genes in this vector resulted in the synthesis of both the fusion proteins. The ability of this recombinant organism to produce several-fold more of the product in the extracellular medium than in the intracellular space would make this system quite attractive as far as the downstream processing of the product is concerned.

Helper-Free Foamy Virus Vectors

PubMed Central

TROBRIDGE, GRANT D.; RUSSELL, DAVID W.

2010-01-01

Retroviral vectors based on human foamy virus (HFV) have been developed and show promise as gene therapy vehicles. Here we describe a method for the production of HFV vector stocks free of detectable helper virus. The helper and vector plasmid constructs used both lack the HFV bel genes, so recombination between these constructs cannot create a wild-type virus. A fusion promoter that combines portions of the cytomegalovirus (CMV) immediate-early and HFV long terminal repeat (LTR) promoters was used to drive expression of both the helper and vector constructs. The CMV–LTR fusion promoter allows for HFV vector production in the absence of the Bel-1 trans-activator protein, which would otherwise be necessary for efficient transcription from the HFV LTR. Vector stocks containing either neomycin phosphotransferase or alkaline phosphatase reporter genes were produced by transient transfection at titers greater than 105 transducing units/ml. G418-resistant BHK-21 cells obtained by transduction with neo vectors contained randomly integrated HFV vector proviruses without detectable deletions or rearrangements. The vector stocks generated were free of replication-competent retrovirus (RCR), as determined by assays for LTR trans-activation and a marker rescue assay developed here for the detection of Bel-independent RCR. OVERVIEW SUMMARY Vectors based on human foamy virus have been developed but low titers and the presence of replication-competent retrovirus (RCR) in vector stocks have prevented their use in preclinical animal experiments. We have developed a transient transfection method that can be used to produce replication-incompetent HFV vector stocks at titers greater than 105/ml, and that does not produce contaminating RCR. The use of CMV-HFV LTR fusion promoters in the helper and vector constructs has circumvented the requirement for the HFV Bel-1 trans-activator protein. Consequently, the potential for generating wild-type HFV by recombination between helper and vector constructs during vector production has been eliminated. Here we describe HFV vector production using this Bel-independent system. PMID:9853518

Large-scale adenovirus and poxvirus-vectored vaccine manufacturing to enable clinical trials.

PubMed

Kallel, Héla; Kamen, Amine A

2015-05-01

Efforts to make vaccines against infectious diseases and immunotherapies for cancer have evolved to utilize a variety of heterologous expression systems such as viral vectors. These vectors are often attenuated or engineered to safely deliver genes encoding antigens of different pathogens. Adenovirus and poxvirus vectors are among the viral vectors that are most frequently used to develop prophylactic vaccines against infectious diseases as well as therapeutic cancer vaccines. This mini-review describes the trends and processes in large-scale production of adenovirus and poxvirus vectors to meet the needs of clinical applications. We briefly describe the general principles for the production and purification of adenovirus and poxvirus viral vectors. Currently, adenovirus and poxvirus vector manufacturing methods rely on well-established cell culture technologies. Several improvements have been evaluated to increase the yield and to reduce the overall manufacturing cost, such as cultivation at high cell densities and continuous downstream processing. Additionally, advancements in vector characterization will greatly facilitate the development of novel vectored vaccine candidates. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Robust production of virus-like particles and monoclonal antibodies with geminiviral replicon vectors in lettuce

PubMed Central

Lai, Huafang; He, Junyun; Engle, Michael; Diamond, Michael S.; Chen, Qiang

2011-01-01

Summary Pharmaceutical protein production in plants has been greatly promoted by the development of viral-based vectors and transient expression systems. Tobacco and related Nicotiana species are currently the most common host plants for generation of plant-made pharmaceutical proteins (PMPs). Downstream processing of target PMPs from these plants, however, is hindered by potential technical and regulatory difficulties due to the presence of high levels of phenolics and toxic alkaloids. Here, we explored the use of lettuce, which grows quickly yet produces low levels of secondary metabolites, and viral vector-based transient expression systems to develop a robust PMP production platform. Our results showed that a geminiviral replicon system based on the bean yellow dwarf virus permits high-level expression in lettuce of virus-like particles (VLP) derived from the Norwalk virus capsid protein and therapeutic monoclonal antibodies (mAbs) against Ebola and West Nile viruses. These vaccine and therapeutic candidates can be readily purified from lettuce leaves with scalable processing methods while fully retaining functional activity. Furthermore, this study also demonstrated the feasibility of using commercially produced lettuce for high-level PMP production. This allows our production system to have access to unlimited quantities of inexpensive plant material for large-scale production. These results establish a new production platform for biological pharmaceutical agents that is effective, safe, low-cost, and amenable to large-scale manufacturing. PMID:21883868

Large-scale Clinical-grade Retroviral Vector Production in a Fixed-Bed Bioreactor

PubMed Central

Wang, Xiuyan; Olszewska, Malgorzata; Qu, Jinrong; Wasielewska, Teresa; Bartido, Shirley; Hermetet, Gregory; Sadelain, Michel

2015-01-01

The successful genetic engineering of patient T cells with γ-retroviral vectors expressing chimeric antigen receptors or T-cell receptors for phase II clinical trials and beyond requires the large-scale manufacture of high-titer vector stocks. The production of retroviral vectors from stable packaging cell lines using roller bottles or 10- to 40-layer cell factories is limited by a narrow harvest window, labor intensity, open-system operations, and the requirement for significant incubator space. To circumvent these shortcomings, we optimized the production of vector stocks in a disposable fixed-bed bioreactor using good manufacturing practice–grade packaging cell lines. High-titer vector stocks were harvested over 10 days, representing a much broader harvest window than the 3-day harvest afforded by cell factories. For PG13 and 293Vec packaging cells, the average vector titer and the vector stocks’ yield in the bioreactor were higher by 3.2- to 7.3-fold, and 5.6- to 13.1-fold, respectively, than those obtained in cell factories. The vector production was 10.4 and 18.6 times more efficient than in cell factories for PG13 and 293Vec cells, respectively. Furthermore, the vectors produced from the fixed-bed bioreactors passed the release test assays for clinical applications. Therefore, a single vector lot derived from 293Vec is suitable to transduce up to 500 patients cell doses in the context of large clinical trials using chimeric antigen receptors or T-cell receptors. These findings demonstrate for the first time that a robust fixed-bed bioreactor process can be used to produce γ-retroviral vector stocks scalable up to the commercialization phase. PMID:25751502

Engineered Saccharomyces cerevisiae strain for improved xylose utilization with a three-plasmid SUMO yeast expression system

USDA-ARS?s Scientific Manuscript database

A three-plasmid yeast expression system utilizing the portable small ubiquitin-like modifier (SUMO) vector set combined with the efficient endogenous yeast protease Ulp1 was developed for production of large amounts of soluble functional protein in Saccharomyces cerevisiae. Each vector has a differ...

Construction of two Lactococcus lactis expression vectors combining the Gateway and the NIsin Controlled Expression systems.

PubMed

Douillard, François P; Mahony, Jennifer; Campanacci, Valérie; Cambillau, Christian; van Sinderen, Douwe

2011-09-01

Over the last 10 years, the NIsin Controlled Expression (NICE) system has been extensively used in the food-grade bacterium Lactococcus lactis subsp. cremoris to produce homologous and heterologous proteins for academic and biotechnological purposes. Although various L. lactis molecular tools have been developed, no expression vectors harboring the popular Gateway recombination system are currently available for this widely used cloning host. In this study, we constructed two expression vectors that combine the NICE and the Gateway recombination systems and we tested their applicability by recombining and over-expressing genes encoding structural proteins of lactococcal phages Tuc2009 and TP901-1. Over-expressed phage proteins were analyzed by immunoblotting and purified by His-tag affinity chromatography with protein productions yielding 2.8-3.7 mg/l of culture. This therefore is the first description of L. lactis NICE expression vectors which integrate the Gateway cloning technology and which are suitable for the production of sufficient amounts of proteins to facilitate subsequent structural and functional analyses. Copyright © 2011 Elsevier Inc. All rights reserved.

Gene delivery to skeletal muscle results in sustained expression and systemic delivery of a therapeutic protein.

PubMed

Kessler, P D; Podsakoff, G M; Chen, X; McQuiston, S A; Colosi, P C; Matelis, L A; Kurtzman, G J; Byrne, B J

1996-11-26

Somatic gene therapy has been proposed as a means to achieve systemic delivery of therapeutic proteins. However, there is limited evidence that current methods of gene delivery can practically achieve this goal. In this study, we demonstrate that, following a single intramuscular administration of a recombinant adeno-associated virus (rAAV) vector containing the beta-galactosidase (AAV-lacZ) gene into adult BALB/c mice, protein expression was detected in myofibers for at least 32 weeks. A single intramuscular administration of an AAV vector containing a gene for human erythropoietin (AAV-Epo) into mice resulted in dose-dependent secretion of erythropoietin and corresponding increases in red blood cell production that persisted for up to 40 weeks. Primary human myotubes transduced in vitro with the AAV-Epo vector also showed dose-dependent production of Epo. These results demonstrate that rAAV vectors are able to transduce skeletal muscle and are capable of achieving sustained expression and systemic delivery of a therapeutic protein following a single intramuscular administration. Gene therapy using AAV vectors may provide a practical strategy for the treatment of inherited and acquired protein deficiencies.

Gene delivery to skeletal muscle results in sustained expression and systemic delivery of a therapeutic protein

PubMed Central

Kessler, Paul D.; Podsakoff, Gregory M.; Chen, Xiaojuan; McQuiston, Susan A.; Colosi, Peter C.; Matelis, Laura A.; Kurtzman, Gary J.; Byrne, Barry J.

1996-01-01

Somatic gene therapy has been proposed as a means to achieve systemic delivery of therapeutic proteins. However, there is limited evidence that current methods of gene delivery can practically achieve this goal. In this study, we demonstrate that, following a single intramuscular administration of a recombinant adeno-associated virus (rAAV) vector containing the β-galactosidase (AAV-lacZ) gene into adult BALB/c mice, protein expression was detected in myofibers for at least 32 weeks. A single intramuscular administration of an AAV vector containing a gene for human erythropoietin (AAV-Epo) into mice resulted in dose-dependent secretion of erythropoietin and corresponding increases in red blood cell production that persisted for up to 40 weeks. Primary human myotubes transduced in vitro with the AAV-Epo vector also showed dose-dependent production of Epo. These results demonstrate that rAAV vectors are able to transduce skeletal muscle and are capable of achieving sustained expression and systemic delivery of a therapeutic protein following a single intramuscular administration. Gene therapy using AAV vectors may provide a practical strategy for the treatment of inherited and acquired protein deficiencies. PMID:8943064

A set of ligation-independent in vitro translation vectors for eukaryotic protein production.

PubMed

Bardóczy, Viola; Géczi, Viktória; Sawasaki, Tatsuya; Endo, Yaeta; Mészáros, Tamás

2008-03-27

The last decade has brought the renaissance of protein studies and accelerated the development of high-throughput methods in all aspects of proteomics. Presently, most protein synthesis systems exploit the capacity of living cells to translate proteins, but their application is limited by several factors. A more flexible alternative protein production method is the cell-free in vitro protein translation. Currently available in vitro translation systems are suitable for high-throughput robotic protein production, fulfilling the requirements of proteomics studies. Wheat germ extract based in vitro translation system is likely the most promising method, since numerous eukaryotic proteins can be cost-efficiently synthesized in their native folded form. Although currently available vectors for wheat embryo in vitro translation systems ensure high productivity, they do not meet the requirements of state-of-the-art proteomics. Target genes have to be inserted using restriction endonucleases and the plasmids do not encode cleavable affinity purification tags. We designed four ligation independent cloning (LIC) vectors for wheat germ extract based in vitro protein translation. In these constructs, the RNA transcription is driven by T7 or SP6 phage polymerase and two TEV protease cleavable affinity tags can be added to aid protein purification. To evaluate our improved vectors, a plant mitogen activated protein kinase was cloned in all four constructs. Purification of this eukaryotic protein kinase demonstrated that all constructs functioned as intended: insertion of PCR fragment by LIC worked efficiently, affinity purification of translated proteins by GST-Sepharose or MagneHis particles resulted in high purity kinase, and the affinity tags could efficiently be removed under different reaction conditions. Furthermore, high in vitro kinase activity testified of proper folding of the purified protein. Four newly designed in vitro translation vectors have been constructed which allow fast and parallel cloning and protein purification, thus representing useful molecular tools for high-throughput production of eukaryotic proteins.

Finding Your Literature Match - A Physics Literature Recommender System

NASA Astrophysics Data System (ADS)

Henneken, Edwin; Kurtz, Michael

2010-03-01

A recommender system is a filtering algorithm that helps you find the right match by offering suggestions based on your choices and information you have provided. A latent factor model is a successful approach. Here an item is characterized by a vector describing to what extent a product is described by each of N categories, and a person is characterized by an ``interest'' vector, based on explicit or implicit feedback by this user. The recommender system assigns ratings to new items and suggests items this user might be interested in. Here we present results of a recommender system designed to find recent literature of interest to people working in the field of solid state physics. Since we do not have explicit feedback, our user vector consists of (implicit) ``usage.'' Using a system of N keywords we construct normalized keyword vectors for articles based on the keywords of that article and its bibliography. The normalized ``interest'' vector is created by calculating the normalized frequency of keyword occurrence in the papers cited by the papers read.

The Minkowski sum of a zonotope and the Voronoi polytope of the root lattice E{sub 7}

DOE Office of Scientific and Technical Information (OSTI.GOV)

Grishukhin, Vyacheslav P

2012-11-30

We show that the Minkowski sum P{sub V}(E{sub 7})+Z(U) of the Voronoi polytope P{sub V}(E{sub 7}) of the root lattice E{sub 7} and the zonotope Z(U) is a 7-dimensional parallelohedron if and only if the set U consists of minimal vectors of the dual lattice E{sub 7}{sup *} up to scalar multiplication, and U does not contain forbidden sets. The minimal vectors of E{sub 7} are the vectors r of the classical root system E{sub 7}. If the r{sup 2}-norm of the roots is set equal to 2, then the scalar products of minimal vectors from the dual lattice onlymore » take the values {+-}1/2. A set of minimal vectors is referred to as forbidden if it consists of six vectors, and the directions of some of these vectors can be changed so as to obtain a set of six vectors with all the pairwise scalar products equal to 1/2. Bibliography: 11 titles.« less

Optimization of a one-step heat-inducible in vivo mini DNA vector production system.

PubMed

Nafissi, Nafiseh; Sum, Chi Hong; Wettig, Shawn; Slavcev, Roderick A

2014-01-01

While safer than their viral counterparts, conventional circular covalently closed (CCC) plasmid DNA vectors offer a limited safety profile. They often result in the transfer of unwanted prokaryotic sequences, antibiotic resistance genes, and bacterial origins of replication that may lead to unwanted immunostimulatory responses. Furthermore, such vectors may impart the potential for chromosomal integration, thus potentiating oncogenesis. Linear covalently closed (LCC), bacterial sequence free DNA vectors have shown promising clinical improvements in vitro and in vivo. However, the generation of such minivectors has been limited by in vitro enzymatic reactions hindering their downstream application in clinical trials. We previously characterized an in vivo temperature-inducible expression system, governed by the phage λ pL promoter and regulated by the thermolabile λ CI[Ts]857 repressor to produce recombinant protelomerase enzymes in E. coli. In this expression system, induction of recombinant protelomerase was achieved by increasing culture temperature above the 37°C threshold temperature. Overexpression of protelomerase led to enzymatic reactions, acting on genetically engineered multi-target sites called "Super Sequences" that serve to convert conventional CCC plasmid DNA into LCC DNA minivectors. Temperature up-shift, however, can result in intracellular stress responses and may alter plasmid replication rates; both of which may be detrimental to LCC minivector production. We sought to optimize our one-step in vivo DNA minivector production system under various induction schedules in combination with genetic modifications influencing plasmid replication, processing rates, and cellular heat stress responses. We assessed different culture growth techniques, growth media compositions, heat induction scheduling and temperature, induction duration, post-induction temperature, and E. coli genetic background to improve the productivity and scalability of our system, achieving an overall LCC DNA minivector production efficiency of ∼ 90%.We optimized a robust technology conferring rapid, scalable, one-step in vivo production of LCC DNA minivectors with potential application to gene transfer-mediated therapeutics.

Optimization of a One-Step Heat-Inducible In Vivo Mini DNA Vector Production System

PubMed Central

Wettig, Shawn; Slavcev, Roderick A.

2014-01-01

While safer than their viral counterparts, conventional circular covalently closed (CCC) plasmid DNA vectors offer a limited safety profile. They often result in the transfer of unwanted prokaryotic sequences, antibiotic resistance genes, and bacterial origins of replication that may lead to unwanted immunostimulatory responses. Furthermore, such vectors may impart the potential for chromosomal integration, thus potentiating oncogenesis. Linear covalently closed (LCC), bacterial sequence free DNA vectors have shown promising clinical improvements in vitro and in vivo. However, the generation of such minivectors has been limited by in vitro enzymatic reactions hindering their downstream application in clinical trials. We previously characterized an in vivo temperature-inducible expression system, governed by the phage λ pL promoter and regulated by the thermolabile λ CI[Ts]857 repressor to produce recombinant protelomerase enzymes in E. coli. In this expression system, induction of recombinant protelomerase was achieved by increasing culture temperature above the 37°C threshold temperature. Overexpression of protelomerase led to enzymatic reactions, acting on genetically engineered multi-target sites called “Super Sequences” that serve to convert conventional CCC plasmid DNA into LCC DNA minivectors. Temperature up-shift, however, can result in intracellular stress responses and may alter plasmid replication rates; both of which may be detrimental to LCC minivector production. We sought to optimize our one-step in vivo DNA minivector production system under various induction schedules in combination with genetic modifications influencing plasmid replication, processing rates, and cellular heat stress responses. We assessed different culture growth techniques, growth media compositions, heat induction scheduling and temperature, induction duration, post-induction temperature, and E. coli genetic background to improve the productivity and scalability of our system, achieving an overall LCC DNA minivector production efficiency of ∼90%.We optimized a robust technology conferring rapid, scalable, one-step in vivo production of LCC DNA minivectors with potential application to gene transfer-mediated therapeutics. PMID:24586704

Product demand forecasts using wavelet kernel support vector machine and particle swarm optimization in manufacture system

NASA Astrophysics Data System (ADS)

Wu, Qi

2010-03-01

Demand forecasts play a crucial role in supply chain management. The future demand for a certain product is the basis for the respective replenishment systems. Aiming at demand series with small samples, seasonal character, nonlinearity, randomicity and fuzziness, the existing support vector kernel does not approach the random curve of the sales time series in the space (quadratic continuous integral space). In this paper, we present a hybrid intelligent system combining the wavelet kernel support vector machine and particle swarm optimization for demand forecasting. The results of application in car sale series forecasting show that the forecasting approach based on the hybrid PSOWv-SVM model is effective and feasible, the comparison between the method proposed in this paper and other ones is also given, which proves that this method is, for the discussed example, better than hybrid PSOv-SVM and other traditional methods.

Standard Free Droplet Digital Polymerase Chain Reaction as a New Tool for the Quality Control of High-Capacity Adenoviral Vectors in Small-Scale Preparations

PubMed Central

Boehme, Philip; Stellberger, Thorsten; Solanki, Manish; Zhang, Wenli; Schulz, Eric; Bergmann, Thorsten; Liu, Jing; Doerner, Johannes; Baiker, Armin E.

2015-01-01

Abstract High-capacity adenoviral vectors (HCAdVs) are promising tools for gene therapy as well as for genetic engineering. However, one limitation of the HCAdV vector system is the complex, time-consuming, and labor-intensive production process and the following quality control procedure. Since HCAdVs are deleted for all viral coding sequences, a helper virus (HV) is needed in the production process to provide the sequences for all viral proteins in trans. For the purification procedure of HCAdV, cesium chloride density gradient centrifugation is usually performed followed by buffer exchange using dialysis or comparable methods. However, performing these steps is technically difficult, potentially error-prone, and not scalable. Here, we establish a new protocol for small-scale production of HCAdV based on commercially available adenovirus purification systems and a standard method for the quality control of final HCAdV preparations. For titration of final vector preparations, we established a droplet digital polymerase chain reaction (ddPCR) that uses a standard free-end-point PCR in small droplets of defined volume. By using different probes, this method is capable of detecting and quantifying HCAdV and HV in one reaction independent of reference material, rendering this method attractive for accurately comparing viral titers between different laboratories. In summary, we demonstrate that it is possible to produce HCAdV in a small scale of sufficient quality and quantity to perform experiments in cell culture, and we established a reliable protocol for vector titration based on ddPCR. Our method significantly reduces time and required equipment to perform HCAdV production. In the future the ddPCR technology could be advantageous for titration of other viral vectors commonly used in gene therapy. PMID:25640117

Viral Vectors for In Vivo Gene Transfer in Parkinson’s disease: Properties and Clinical Grade Production

PubMed Central

Burger, Corinna; Snyder, Richard O.

2009-01-01

Because Parkinson’s disease is a progressive degenerative disorder that is mainly confined to the basal ganglia, gene transfer to deliver therapeutic molecules is an attractive treatment avenue. The present review focuses on direct in vivo gene transfer vectors that have been developed to a degree that they have been successfully used in animal model of Parkinson’s disease. Accordingly, the properties of recombinant adenovirus, recombinant adeno-associated virus, herpes simplex virus, and lentivirus are described and contrasted. In order for viral vectors to be developed into clinical grade reagents, they must be manufactured and tested to precise regulatory standards. Indeed, clinical lots of viral vectors can be produced in compliance with current Good Manufacturing Practices (cGMPs) regulations using industry accepted manufacturing methodologies, manufacturing controls, and quality systems. The viral vector properties themselves combined with physiological product formulations facilitate long-term storage and direct in vivo administration. PMID:17916354

In planta expression of HIV-1 p24 protein using an RNA plant virus-based expression vector.

PubMed

Zhang, G; Leung, C; Murdin, L; Rovinski, B; White, K A

2000-02-01

Plant viruses show significant potential as expression vectors for the production of foreign proteins (e.g., antigens) in plants. The HIV-1 p24 nucleocapsid protein is an important early marker of HIV infection and has been used as an antigen in the development of HIV vaccines. Toward developing a plant-based expression system for the production of p24, we have investigated the use of a (positive)-strand RNA plant virus, tomato bushy stunt virus (TBSV), as an expression vector. The HIV p24 open reading frame (ORF) was introduced into a cloned cDNA copy of the TBSV genome as an in-frame fusion with a 5'-terminal portion of the TBSV coat protein ORF. In vitro-generated RNA transcripts corresponding to the engineered virus vector were infectious when inoculated into plant protoplasts; Northern and Western blot analyses verified the accumulation of a predicted p24-encoding viral subgenomic mRNA and the production of p24 fusion product. Whole-plant infections with the viral vector led to the accumulation of p24 fusion protein in inoculated leaves, which cross-reacted with p24-specific antibodies, thus confirming the maintenance of key antigenic determinants. This study is the first to demonstrate that TBSV can be engineered to express a complete foreign protein of clinical importance. Strategies for optimizing protein yield from this viral vector are discussed.

Elucidating the Potential of Plant Rhabdoviruses as Vector Expressions Systems

USDA-ARS?s Scientific Manuscript database

Maize fine streak virus (MFSV) is a member of the genus Nucleorhabdovirus that is transmitted by the leafhopper Graminella nigrifons. The virus replicates in both its maize host and its insect vector. To determine whether Drosophila S2 cells support the production of full-length MFSV proteins, we ...

Insect cells as factories for biomanufacturing.

PubMed

Drugmand, Jean-Christophe; Schneider, Yves-Jacques; Agathos, Spiros N

2012-01-01

Insect cells (IC) and particularly lepidopteran cells are an attractive alternative to mammalian cells for biomanufacturing. Insect cell culture, coupled with the lytic expression capacity of baculovirus expression vector systems (BEVS), constitutes a powerful platform, IC-BEVS, for the abundant and versatile formation of heterologous gene products, including proteins, vaccines and vectors for gene therapy. Such products can be manufactured on a large scale thanks to the development of efficient and scaleable production processes involving the integration of a cell growth stage and a stage of cell infection with the recombinant baculovirus vector. Insect cells can produce multimeric proteins functionally equivalent to the natural ones and engineered vectors can be used for efficient expression. Insect cells can be cultivated easily in serum- and protein-free media. A growing number of companies are currently developing an interest in producing therapeutics using IC-BEVS, and many products are today in clinical trials and on the market for veterinary and human applications. This review summarizes current knowledge on insect cell metabolism, culture conditions and applications. Copyright © 2011 Elsevier Inc. All rights reserved.

An IPSO-SVM algorithm for security state prediction of mine production logistics system

NASA Astrophysics Data System (ADS)

Zhang, Yanliang; Lei, Junhui; Ma, Qiuli; Chen, Xin; Bi, Runfang

2017-06-01

A theoretical basis for the regulation of corporate security warning and resources was provided in order to reveal the laws behind the security state in mine production logistics. Considering complex mine production logistics system and the variable is difficult to acquire, a superior security status predicting model of mine production logistics system based on the improved particle swarm optimization and support vector machine (IPSO-SVM) is proposed in this paper. Firstly, through the linear adjustments of inertia weight and learning weights, the convergence speed and search accuracy are enhanced with the aim to deal with situations associated with the changeable complexity and the data acquisition difficulty. The improved particle swarm optimization (IPSO) is then introduced to resolve the problem of parameter settings in traditional support vector machines (SVM). At the same time, security status index system is built to determine the classification standards of safety status. The feasibility and effectiveness of this method is finally verified using the experimental results.

Exclusive diffractive production of real photons and vector mesons in a factorized Regge-pole model with nonlinear Pomeron trajectory

NASA Astrophysics Data System (ADS)

Fazio, S.; Fiore, R.; Jenkovszky, L.; Lavorini, A.

2012-03-01

Exclusive diffractive production of real photons and vector mesons in ep collisions has been studied at HERA in a wide kinematic range. Here we present and discuss a Regge-type model of real photon production (deeply virtual Compton scattering), as well as production of vector mesons treated on the same footing by using an extension of a factorized Regge-pole model proposed earlier. The model has been fitted to the HERA data. Despite the very small number of the free parameters, the model gives a satisfactory description of the experimental data, both for the total cross section as a function of the photon virtuality Q2 or the energy W in the center of mass of the γ*p system, and the differential cross sections as a function of the squared four-momentum transfer t with fixed Q2 and W.

Neonatal Systemic AAV Induces Tolerance to CNS Gene Therapy in MPS I Dogs and Nonhuman Primates

PubMed Central

Hinderer, Christian; Bell, Peter; Louboutin, Jean-Pierre; Zhu, Yanqing; Yu, Hongwei; Lin, Gloria; Choa, Ruth; Gurda, Brittney L; Bagel, Jessica; O'Donnell, Patricia; Sikora, Tracey; Ruane, Therese; Wang, Ping; Tarantal, Alice F; Casal, Margret L; Haskins, Mark E; Wilson, James M

2015-01-01

The potential host immune response to a nonself protein poses a fundamental challenge for gene therapies targeting recessive diseases. We demonstrate in both dogs and nonhuman primates that liver-directed gene transfer using an adeno-associated virus (AAV) vector in neonates induces a persistent state of immunological tolerance to the transgene product, substantially improving the efficacy of subsequent vector administration targeting the central nervous system (CNS). We applied this approach to a canine model of mucopolysaccharidosis type I (MPS I), a progressive neuropathic lysosomal storage disease caused by deficient activity of the enzyme α-l-iduronidase (IDUA). MPS I dogs treated systemically in the first week of life with a vector expressing canine IDUA did not develop antibodies against the enzyme and exhibited robust expression in the CNS upon intrathecal AAV delivery at 1 month of age, resulting in complete correction of brain storage lesions. Newborn rhesus monkeys treated systemically with AAV vector expressing human IDUA developed tolerance to the transgene, resulting in high cerebrospinal fluid (CSF) IDUA expression and no antibody induction after subsequent CNS gene therapy. These findings suggest that inducing tolerance to the transgene product during a critical period in immunological development can improve the efficacy and safety of gene therapy. PMID:26022732

Neonatal Systemic AAV Induces Tolerance to CNS Gene Therapy in MPS I Dogs and Nonhuman Primates.

PubMed

Hinderer, Christian; Bell, Peter; Louboutin, Jean-Pierre; Zhu, Yanqing; Yu, Hongwei; Lin, Gloria; Choa, Ruth; Gurda, Brittney L; Bagel, Jessica; O'Donnell, Patricia; Sikora, Tracey; Ruane, Therese; Wang, Ping; Tarantal, Alice F; Casal, Margret L; Haskins, Mark E; Wilson, James M

2015-08-01

The potential host immune response to a nonself protein poses a fundamental challenge for gene therapies targeting recessive diseases. We demonstrate in both dogs and nonhuman primates that liver-directed gene transfer using an adeno-associated virus (AAV) vector in neonates induces a persistent state of immunological tolerance to the transgene product, substantially improving the efficacy of subsequent vector administration targeting the central nervous system (CNS). We applied this approach to a canine model of mucopolysaccharidosis type I (MPS I), a progressive neuropathic lysosomal storage disease caused by deficient activity of the enzyme α-l-iduronidase (IDUA). MPS I dogs treated systemically in the first week of life with a vector expressing canine IDUA did not develop antibodies against the enzyme and exhibited robust expression in the CNS upon intrathecal AAV delivery at 1 month of age, resulting in complete correction of brain storage lesions. Newborn rhesus monkeys treated systemically with AAV vector expressing human IDUA developed tolerance to the transgene, resulting in high cerebrospinal fluid (CSF) IDUA expression and no antibody induction after subsequent CNS gene therapy. These findings suggest that inducing tolerance to the transgene product during a critical period in immunological development can improve the efficacy and safety of gene therapy.

Complex matrix multiplication operations with data pre-conditioning in a high performance computing architecture

DOEpatents

Eichenberger, Alexandre E; Gschwind, Michael K; Gunnels, John A

2014-02-11

Mechanisms for performing a complex matrix multiplication operation are provided. A vector load operation is performed to load a first vector operand of the complex matrix multiplication operation to a first target vector register. The first vector operand comprises a real and imaginary part of a first complex vector value. A complex load and splat operation is performed to load a second complex vector value of a second vector operand and replicate the second complex vector value within a second target vector register. The second complex vector value has a real and imaginary part. A cross multiply add operation is performed on elements of the first target vector register and elements of the second target vector register to generate a partial product of the complex matrix multiplication operation. The partial product is accumulated with other partial products and a resulting accumulated partial product is stored in a result vector register.

A new double right border binary vector for producing marker-free transgenic plants

PubMed Central

2013-01-01

Background Once a transgenic plant is developed, the selectable marker gene (SMG) becomes unnecessary in the plant. In fact, the continued presence of the SMG in the transgenic plant may cause unexpected pleiotropic effects as well as environmental or biosafety issues. Several methods for removal of SMGs that have been reported remain inaccessible due to protection by patents, while development of new ones is expensive and cost prohibitive. Here, we describe the development of a new vector for producing marker-free plants by simply adapting an ordinary binary vector to the double right border (DRB) vector design using conventional cloning procedures. Findings We developed the DRB vector pMarkfree5.0 by placing the bar gene (representing genes of interest) between two copies of T-DNA right border sequences. The β-glucuronidase (gus) and nptII genes (representing the selectable marker gene) were cloned next followed by one copy of the left border sequence. When tested in a model species (tobacco), this vector system enabled the generation of 55.6% kanamycin-resistant plants by Agrobacterium-mediated transformation. The frequency of cotransformation of the nptII and bar transgenes using the vector was 66.7%. Using the leaf bleach and Basta assays, we confirmed that the nptII and bar transgenes were coexpressed and segregated independently in the transgenic plants. This enable separation of the transgenes in plants cotransformed using pMarkfree5.0. Conclusions The results suggest that the DRB system developed here is a practical and effective approach for separation of gene(s) of interest from a SMG and production of SMG-free plants. Therefore this system could be instrumental in production of “clean” plants containing genes of agronomic importance. PMID:24207020

A novel packaging system for the generation of helper-free oncolytic MVM vector stocks.

PubMed

Brandenburger, A; Russell, S

1996-10-01

MVM-based autonomous parvoviral vectors have been shown to target the expression of heterologous genes in neoplastic cells and are therefore of interest for cancer gene therapy. The traditional method for production of parvoviral vectors requires the cotransfection of vector and helper plasmids into MVM-permissive cell lines, but recombination between the cotransfected plasmids invariably gives rise to vector stocks that are heavily contaminated with wild-type MVM. Therefore, to minimise recombination between the vector and helper genomes we have utilised a cell line in which the MVM helper functions are expressed inducibly from a modified MVM genome that is stably integrated into the host cell chromosome. Using this MVM packaging cell line, we could reproducibly generate MVM vector stocks that contained no detectable helper virus.

An iterative technique to stabilize a linear time invariant multivariable system with output feedback

NASA Technical Reports Server (NTRS)

Sankaran, V.

1974-01-01

An iterative procedure for determining the constant gain matrix that will stabilize a linear constant multivariable system using output feedback is described. The use of this procedure avoids the transformation of variables which is required in other procedures. For the case in which the product of the output and input vector dimensions is greater than the number of states of the plant, general solution is given. In the case in which the states exceed the product of input and output vector dimensions, a least square solution which may not be stable in all cases is presented. The results are illustrated with examples.

Technique for Solving Electrically Small to Large Structures for Broadband Applications

NASA Technical Reports Server (NTRS)

Jandhyala, Vikram; Chowdhury, Indranil

2011-01-01

Fast iterative algorithms are often used for solving Method of Moments (MoM) systems, having a large number of unknowns, to determine current distribution and other parameters. The most commonly used fast methods include the fast multipole method (FMM), the precorrected fast Fourier transform (PFFT), and low-rank QR compression methods. These methods reduce the O(N) memory and time requirements to O(N log N) by compressing the dense MoM system so as to exploit the physics of Green s Function interactions. FFT-based techniques for solving such problems are efficient for spacefilling and uniform structures, but their performance substantially degrades for non-uniformly distributed structures due to the inherent need to employ a uniform global grid. FMM or QR techniques are better suited than FFT techniques; however, neither the FMM nor the QR technique can be used at all frequencies. This method has been developed to efficiently solve for a desired parameter of a system or device that can include both electrically large FMM elements, and electrically small QR elements. The system or device is set up as an oct-tree structure that can include regions of both the FMM type and the QR type. The system is enclosed with a cube at a 0- th level, splitting the cube at the 0-th level into eight child cubes. This forms cubes at a 1st level, recursively repeating the splitting process for cubes at successive levels until a desired number of levels is created. For each cube that is thus formed, neighbor lists and interaction lists are maintained. An iterative solver is then used to determine a first matrix vector product for any electrically large elements as well as a second matrix vector product for any electrically small elements that are included in the structure. These matrix vector products for the electrically large and small elements are combined, and a net delta for a combination of the matrix vector products is determined. The iteration continues until a net delta is obtained that is within the predefined limits. The matrix vector products that were last obtained are used to solve for the desired parameter. The solution for the desired parameter is then presented to a user in a tangible form; for example, on a display.

Excess Entropy Production in Quantum System: Quantum Master Equation Approach

NASA Astrophysics Data System (ADS)

Nakajima, Satoshi; Tokura, Yasuhiro

2017-12-01

For open systems described by the quantum master equation (QME), we investigate the excess entropy production under quasistatic operations between nonequilibrium steady states. The average entropy production is composed of the time integral of the instantaneous steady entropy production rate and the excess entropy production. We propose to define average entropy production rate using the average energy and particle currents, which are calculated by using the full counting statistics with QME. The excess entropy production is given by a line integral in the control parameter space and its integrand is called the Berry-Sinitsyn-Nemenman (BSN) vector. In the weakly nonequilibrium regime, we show that BSN vector is described by ln \\breve{ρ }_0 and ρ _0 where ρ _0 is the instantaneous steady state of the QME and \\breve{ρ }_0 is that of the QME which is given by reversing the sign of the Lamb shift term. If the system Hamiltonian is non-degenerate or the Lamb shift term is negligible, the excess entropy production approximately reduces to the difference between the von Neumann entropies of the system. Additionally, we point out that the expression of the entropy production obtained in the classical Markov jump process is different from our result and show that these are approximately equivalent only in the weakly nonequilibrium regime.

Generation of infectious recombinant Adeno-associated virus in Saccharomyces cerevisiae.

PubMed

Barajas, Daniel; Aponte-Ubillus, Juan Jose; Akeefe, Hassibullah; Cinek, Tomas; Peltier, Joseph; Gold, Daniel

2017-01-01

The yeast Saccharomyces cerevisiae has been successfully employed to establish model systems for a number of viruses. Such model systems are powerful tools to study the virus biology and in particular for the identification and characterization of host factors playing a role in the viral infection cycle. Adeno-associated viruses (AAV) are heavily studied due to their use as gene delivery vectors. AAV relies on other helper viruses for successful replication and on host factors for several aspects of the viral life cycle. However the role of host and helper viral factors is only partially known. Production of recombinant AAV (rAAV) vectors for gene delivery applications depends on knowledge of AAV biology and the limited understanding of host and helper viral factors may be precluding efficient production, particularly in heterologous systems. Model systems in simpler eukaryotes like the yeast S. cerevisiae would be useful tools to identify and study the role of host factors in AAV biology. Here we show that expression of AAV2 viral proteins VP1, VP2, VP3, AAP, Rep78, Rep52 and an ITR-flanked DNA in yeast leads to capsid formation, DNA replication and encapsidation, resulting in formation of infectious particles. Many of the AAV characteristics observed in yeast resemble those in other systems, making it a suitable model system. Future findings in the yeast system could be translatable to other AAV host systems and aid in more efficient production of rAAV vectors.

In situ pneumococcal vaccine production and delivery through a hybrid biological-biomaterial vector

PubMed Central

Li, Yi; Beitelshees, Marie; Fang, Lei; Hill, Andrew; Ahmadi, Mahmoud Kamal; Chen, Mingfu; Davidson, Bruce A.; Knight, Paul; Smith, Randall J.; Andreadis, Stelios T.; Hakansson, Anders P.; Jones, Charles H.; Pfeifer, Blaine A.

2016-01-01

The type and potency of an immune response provoked during vaccination will determine ultimate success in disease prevention. The basis for this response will be the design and implementation of antigen presentation to the immune system. Whereas direct antigen administration will elicit some form of immunological response, a more sophisticated approach would couple the antigen of interest to a vector capable of broad delivery formats and designed for heightened response. New antigens associated with pneumococcal disease virulence were used to test the delivery and adjuvant capabilities of a hybrid biological-biomaterial vector consisting of a bacterial core electrostatically coated with a cationic polymer. The hybrid design provides (i) passive and active targeting of antigen-presenting cells, (ii) natural and multicomponent adjuvant properties, (iii) dual intracellular delivery mechanisms, and (iv) a simple formulation mechanism. In addition, the hybrid format enables device-specific, or in situ, antigen production and consolidation via localization within the bacterial component of the vector. This capability eliminates the need for dedicated antigen production and purification before vaccination efforts while leveraging the aforementioned features of the overall delivery device. We present the first disease-specific utilization of the vector toward pneumococcal disease highlighted by improved immune responses and protective capabilities when tested against traditional vaccine formulations and a range of clinically relevant Streptococcus pneumoniae strains. More broadly, the results point to similar levels of success with other diseases that would benefit from the production, delivery, and efficacy capabilities offered by the hybrid vector. PMID:27419235

Mechanisms and ecological role of carbon transfer within coastal seascapes.

PubMed

Hyndes, Glenn A; Nagelkerken, Ivan; McLeod, Rebecca J; Connolly, Rod M; Lavery, Paul S; Vanderklift, Mathew A

2014-02-01

Worldwide, coastal systems provide some of the most productive habitats, which potentially influence a range of marine and terrestrial ecosystems through the transfer of nutrients and energy. Several reviews have examined aspects of connectivity within coastal seascapes, but the scope of those reviews has been limited to single systems or single vectors. We use the transfer of carbon to examine the processes of connectivity through multiple vectors in multiple ecosystems using four coastal seascapes as case studies. We discuss and compare the main vectors of carbon connecting different ecosystems, and then the natural and human-induced factors that influence the magnitude of effect for those vectors on recipient systems. Vectors of carbon transfer can be grouped into two main categories: detrital particulate organic carbon (POC) and its associated dissolved organic and inorganic carbon (DOC/DIC) that are transported passively; and mobile consumers that transport carbon actively. High proportions of net primary production can be exported over meters to hundreds of kilometers from seagrass beds, algal reefs and mangroves as POC, with its export dependent on wind-generated currents in the first two of these systems and tidal currents for the last. By contrast, saltmarshes export large quantities of DOC through tidal movement, while land run-off plays a critical role in the transport of terrestrial POC and DOC into temperate fjords. Nekton actively transfers carbon across ecosystem boundaries through foraging movements, ontogenetic migrations, or 'trophic relays', into and out of seagrass beds, mangroves or saltmarshes. The magnitude of these vectors is influenced by: the hydrodynamics and geomorphology of the region; the characteristics of the carbon vector, such as their particle size and buoyancy; and for nekton, the extent and frequency of migrations between ecosystems. Through a risk-assessment process, we have identified the most significant human disturbances that affect the integrity of connectivity among ecosystems. Loss of habitat, net primary production (NPP) and overfishing pose the greatest risks to carbon transfer in temperate saltmarsh and tropical estuaries, particularly through their effects on nekton abundance and movement. In comparison, habitat/NPP loss and climate change are likely to be the major risks to carbon transfer in temperate fjords and temperate open coasts through alteration in the amount of POC and/or DOC/DIC being transported. While we have highlighted the importance of these vectors in coastal seascapes, there is limited quantitative data on the effects of these vectors on recipient systems. It is only through quantifying those subsidies that we can effectively incorporate complex interactions into the management of the marine environment and its resources. © 2013 The Authors. Biological Reviews © 2013 Cambridge Philosophical Society.

Some Correlation Functions in Matrix Product Ground States of One-Dimensional Two-State Chains

NASA Astrophysics Data System (ADS)

Shariati, Ahmad; Aghamohammadi, Amir; Fatollahi, Amir H.; Khorrami, Mohammad

2014-04-01

Consider one-dimensional chains with nearest neighbour interactions, for which to each site correspond two independent states (say up and down), and the ground state is a matrix product state. It has been shown [23] that for such systems, the ground states are linear combinations of specific vectors which are essentially direct products of specific numbers of ups and downs, symmetrized in a generalized manner. By a generalized manner, it is meant that the coefficient corresponding to the interchange of states of two sites, in not necessarily plus one or minus one, but a phase which depends on the Hamiltonian and the position of the two sites. Such vectors are characterized by a phase χ, the N-th power of which is one (where N is the number of sites), and an integer. Corresponding to χ, there is another integer M which is the smallest positive integer that χM is one. Two classes of correlation functions for such systems (basically correlation functions for such vectors) are calculated. The first class consists of correlation functions of tensor products of one-site diagonal observables; the second class consists of correlation functions of tensor products of less than M one-site observables (but not necessarily diagonal).

Viral Vectors for Gene Delivery to the Central Nervous System

PubMed Central

Lentz, Thomas B.; Gray, Steven J.; Samulski, R. Jude

2011-01-01

The potential benefits of gene therapy for neurological diseases such as Parkinson’s, Amyotrophic Lateral Sclerosis (ALS), Epilepsy, and Alzheimer’s are enormous. Even a delay in the onset of severe symptoms would be invaluable to patients suffering from these and other diseases. Significant effort has been placed in developing vectors capable of delivering therapeutic genes to the CNS in order to treat neurological disorders. At the forefront of potential vectors, viral systems have evolved to efficiently deliver their genetic material to a cell. The biology of different viruses offers unique solutions to the challenges of gene therapy, such as cell targeting, transgene expression and vector production. It is important to consider the natural biology of a vector when deciding whether it will be the most effective for a specific therapeutic function. In this review, we outline desired features of the ideal vector for gene delivery to the CNS and discuss how well available viral vectors compare to this model. Adeno-associated virus, retrovirus, adenovirus and herpesvirus vectors are covered. Focus is placed on features of the natural biology that have made these viruses effective tools for gene delivery with emphasis on their application in the CNS. Our goal is to provide insight into features of the optimal vector and which viral vectors can provide these features. PMID:22001604

Imer-product array processor for retrieval of stored images represented by bipolar binary (+1,-1) pixels using partial input trinary pixels represented by (+1,-1)

NASA Technical Reports Server (NTRS)

Liu, Hua-Kuang (Inventor); Awwal, Abdul A. S. (Inventor); Karim, Mohammad A. (Inventor)

1993-01-01

An inner-product array processor is provided with thresholding of the inner product during each iteration to make more significant the inner product employed in estimating a vector to be used as the input vector for the next iteration. While stored vectors and estimated vectors are represented in bipolar binary (1,-1), only those elements of an initial partial input vector that are believed to be common with those of a stored vector are represented in bipolar binary; the remaining elements of a partial input vector are set to 0. This mode of representation, in which the known elements of a partial input vector are in bipolar binary form and the remaining elements are set equal to 0, is referred to as trinary representation. The initial inner products corresponding to the partial input vector will then be equal to the number of known elements. Inner-product thresholding is applied to accelerate convergence and to avoid convergence to a negative input product.

Detection of a sudden change of the field time series based on the Lorenz system.

PubMed

Da, ChaoJiu; Li, Fang; Shen, BingLu; Yan, PengCheng; Song, Jian; Ma, DeShan

2017-01-01

We conducted an exploratory study of the detection of a sudden change of the field time series based on the numerical solution of the Lorenz system. First, the time when the Lorenz path jumped between the regions on the left and right of the equilibrium point of the Lorenz system was quantitatively marked and the sudden change time of the Lorenz system was obtained. Second, the numerical solution of the Lorenz system was regarded as a vector; thus, this solution could be considered as a vector time series. We transformed the vector time series into a time series using the vector inner product, considering the geometric and topological features of the Lorenz system path. Third, the sudden change of the resulting time series was detected using the sliding t-test method. Comparing the test results with the quantitatively marked time indicated that the method could detect every sudden change of the Lorenz path, thus the method is effective. Finally, we used the method to detect the sudden change of the pressure field time series and temperature field time series, and obtained good results for both series, which indicates that the method can apply to high-dimension vector time series. Mathematically, there is no essential difference between the field time series and vector time series; thus, we provide a new method for the detection of the sudden change of the field time series.

Production of SV40-derived vectors.

PubMed

Strayer, David S; Mitchell, Christine; Maier, Dawn A; Nichols, Carmen N

2010-06-01

Recombinant simian virus 40 (rSV40)-derived vectors are particularly useful for gene delivery to bone marrow progenitor cells and their differentiated derivatives, certain types of epithelial cells (e.g., hepatocytes), and central nervous system neurons and microglia. They integrate rapidly into cellular DNA to provide long-term gene expression in vitro and in vivo in both resting and dividing cells. Here we describe a protocol for production and purification of these vectors. These procedures require only packaging cells (e.g., COS-7) and circular vector genome DNA. Amplification involves repeated infection of packaging cells with vector produced by transfection. Cotransfection is not required in any step. Viruses are purified by centrifugation using discontinuous sucrose or cesium chloride (CsCl) gradients and resulting vectors are replication-incompetent and contain no detectable wild-type SV40 revertants. These approaches are simple, give reproducible results, and may be used to generate vectors that are deleted only for large T antigen (Tag), or for all SV40-coding sequences capable of carrying up to 5 kb of foreign DNA. These vectors are best applied to long-term expression of proteins normally encoded by mammalian cells or by viruses that infect mammalian cells, or of untranslated RNAs (e.g., RNA interference). The preparative approaches described facilitate application of these vectors and allow almost any laboratory to exploit their strengths for diverse gene delivery applications.

Acoustic nonreciprocity in Coriolis mean flow systems.

PubMed

Naghdi, Masoud; Farzbod, Farhad

2018-01-01

One way to break acoustic reciprocity is to have a moving wave propagation medium. If the acoustic wave vector and the moving fluid velocity are collinear, the wave vector shift caused by the fluid flow can be used to break. In this paper, an alternative approach is investigated in which the fluid velocity enters the differential equation of the system as a cross product term with the wave vector. A circular field where the fluid velocity increases radially has a Coriolis acceleration term. In such a system, the acoustic wave enters from the central wall and exits from the perimeter wall. In this paper, the differential equation is solved numerically and the effect of fluid velocity on the nonreciprocity factor is examined.

The Cross Product of Two Vectors Is Not Just Another Vector--A Major Misconception Being Perpetuated in Calculus and Vector Analysis Textbooks.

ERIC Educational Resources Information Center

Elk, Seymour B.

1997-01-01

Suggests that the cross product of two vectors can be more easily and accurately explained by starting from the perspective of dyadics because then the concept of vector multiplication has a simple geometrical picture that encompasses both the dot and cross products in any number of dimensions in terms of orthogonal unit vector components. (AIM)

Evaluation of Commercial Agrochemicals as New Tools for Malaria Vector Control.

PubMed

Hoppé, Mark; Hueter, Ottmar F; Bywater, Andy; Wege, Philip; Maienfisch, Peter

2016-10-01

Malaria is a vector-borne and life-threatening disease caused by parasites that are transmitted to people through the bites of infected female Anopheles mosquitoes. The vector control insecticide market represents a small fraction of the crop protection market and is estimated to be valued at up to $500 million at the active ingredient level. Insecticide resistance towards the current WHOPES-approved products urgently requires the development of new tools to protect communities against the transmission of malaria. The evaluation of commercial products for malaria vector control is a viable and cost effective strategy to identify new malaria vector control products. Several examples of such spin-offs from crop protection insecticides are already evidencing the success of this strategy, namely pirimiphos-methyl for indoor residual sprays and spinosad, diflubenzuron, novaluron, and pyriproxifen for mosquito larvae control, a supplementary technology for control of malaria vectors. In our study the adulticidal activities of 81 insecticides representing 23 insecticidal modes of action classes, 34 fungicides from 6 fungicidal mode of action classes and 15 herbicides from 2 herbicidal modes of action classes were tested in a newly developed screening system. WHOPES approved insecticides for malaria vector control consistently caused 80-100% mortality of adult Anopheles stephensi at application rates between 0.2 and 20 mg active ingradient (AI) litre -1 . Chlorfenapyr, fipronil, carbosulfan and endosulfan showed the expected good activity. Four new insecticides and three fungicides with promising activity against adult mosquitoes were identified, namely the insecticides acetamiprid, thiamethoxam, thiocyclam and metaflumizone and the fungicides diflumetorin, picoxystrobin, and fluazinam. Some of these compounds certainly deserve to be further evaluated for malaria vector control. This is the first report describing good activity of commercial fungicides against malaria vectors.

Cost Study of Educational Media Systems and Their Equipment Components. Volume II, Technical Report. Final Report.

ERIC Educational Resources Information Center

General Learning Corp., Washington, DC.

A common instructional task and a set of educational environments are hypothesized for analysis of media cost data. The analytic structure may be conceptulized as a three-dimensional matrix: the first vector separates costs into production, distribution, and reception; the second vector delineates capital (initial) and operating (annual) costs;…

A novel expression system for intracellular production and purification of recombinant affinity-tagged proteins in Aspergillus niger.

PubMed

Roth, Andreas H F J; Dersch, Petra

2010-03-01

A set of different integrative expression vectors for the intracellular production of recombinant proteins with or without affinity tag in Aspergillus niger was developed. Target genes can be expressed under the control of the highly efficient, constitutive pkiA promoter or the novel sucrose-inducible promoter of the beta-fructofuranosidase (sucA) gene of A. niger in the presence or absence of alternative carbon sources. All expression plasmids contain an identical multiple cloning sequence that allows parallel construction of N- or C-terminally His6- and StrepII-tagged versions of the target proteins. Production of two heterologous model proteins, the green fluorescence protein and the Thermobifida fusca hydrolase, proved the functionality of the vector system. Efficient production and easy detection of the target proteins as well as their fast purification by a one-step affinity chromatography, using the His6- or StrepII-tag sequence, was demonstrated.

Optical implementation of inner product neural associative memory

NASA Technical Reports Server (NTRS)

Liu, Hua-Kuang (Inventor)

1995-01-01

An optical implementation of an inner-product neural associative memory is realized with a first spatial light modulator for entering an initial two-dimensional N-tuple vector and for entering a thresholded output vector image after each iteration until convergence is reached, and a second spatial light modulator for entering M weighted vectors of inner-product scalars multiplied with each of the M stored vectors, where the inner-product scalars are produced by multiplication of the initial input vector in the first iterative cycle (and thresholded vectors in subsequent iterative cycles) with each of the M stored vectors, and the weighted vectors are produced by multiplication of the scalars with corresponding ones of the stored vectors. A Hughes liquid crystal light valve is used for the dual function of summing the weighted vectors and thresholding the sum vector. The thresholded vector is then entered through the first spatial light modulator for reiteration of the process cycle until convergence is reached.

Firewalls Prevent Systemic Dissemination of Vectors Derived from Human Adenovirus Type 5 and Suppress Production of Transgene-Encoded Antigen in a Murine Model of Oral Vaccination

PubMed Central

Revaud, Julien; Unterfinger, Yves; Rol, Nicolas; Suleman, Muhammad; Shaw, Julia; Galea, Sandra; Gavard, Françoise; Lacour, Sandrine A.; Coulpier, Muriel; Versillé, Nicolas; Havenga, Menzo; Klonjkowski, Bernard; Zanella, Gina; Biacchesi, Stéphane; Cordonnier, Nathalie; Corthésy, Blaise; Ben Arous, Juliette; Richardson, Jennifer P.

2018-01-01

To define the bottlenecks that restrict antigen expression after oral administration of viral-vectored vaccines, we tracked vectors derived from the human adenovirus type 5 at whole body, tissue, and cellular scales throughout the digestive tract in a murine model of oral delivery. After intragastric administration of vectors encoding firefly luciferase or a model antigen, detectable levels of transgene-encoded protein or mRNA were confined to the intestine, and restricted to delimited anatomical zones. Expression of luciferase in the form of multiple small bioluminescent foci in the distal ileum, cecum, and proximal colon suggested multiple crossing points. Many foci were unassociated with visible Peyer's patches, implying that transduced cells lay in proximity to villous rather than follicle-associated epithelium, as supported by detection of transgene-encoded antigen in villous epithelial cells. Transgene-encoded mRNA but not protein was readily detected in Peyer's patches, suggesting that post-transcriptional regulation of viral gene expression might limit expression of transgene-encoded antigen in this tissue. To characterize the pathways by which the vector crossed the intestinal epithelium and encountered sentinel cells, a fluorescent-labeled vector was administered to mice by the intragastric route or inoculated into ligated intestinal loops comprising a Peyer's patch. The vector adhered selectively to microfold cells in the follicle-associated epithelium, and, after translocation to the subepithelial dome region, was captured by phagocytes that expressed CD11c and lysozyme. In conclusion, although a large number of crossing events took place throughout the intestine within and without Peyer's patches, multiple firewalls prevented systemic dissemination of vector and suppressed production of transgene-encoded protein in Peyer's patches. PMID:29423380

Determination of Anti-Adeno-Associated Virus Vector Neutralizing Antibody Titer with an In Vitro Reporter System

PubMed Central

Meliani, Amine; Leborgne, Christian; Triffault, Sabrina; Jeanson-Leh, Laurence; Veron, Philippe

2015-01-01

Abstract Adeno-associated virus (AAV) vectors are a platform of choice for in vivo gene transfer applications. However, neutralizing antibodies (NAb) to AAV can be found in humans and some animal species as a result of exposure to the wild-type virus, and high-titer NAb develop following AAV vector administration. In some conditions, anti-AAV NAb can block transduction with AAV vectors even when present at low titers, thus requiring prescreening before vector administration. Here we describe an improved in vitro, cell-based assay for the determination of NAb titer in serum or plasma samples. The assay is easy to setup and sensitive and, depending on the purpose, can be validated to support clinical development of gene therapy products based on AAV vectors. PMID:25819687

A Technical and Economic Review of Solar Hydrogen Production Technologies

ERIC Educational Resources Information Center

Wilhelm, Erik; Fowler, Michael

2006-01-01

Hydrogen energy systems are being developed to replace fossil fuels-based systems for transportation and stationary application. One of the challenges facing the widespread adoption of hydrogen as an energy vector is the lack of an efficient, economical, and sustainable method of hydrogen production. In the short term, hydrogen produced from…

Product-Related Impurities in Clinical-Grade Recombinant AAV Vectors: Characterization and Risk Assessment

PubMed Central

Wright, J. Fraser

2014-01-01

Adeno-associated virus (AAV)-based vectors expressing therapeutic genes continue to demonstrate great promise for the treatment of a wide variety of diseases and together with other gene transfer vectors represent an emerging new therapeutic paradigm comparable in potential impact on human health to that achieved by recombinant proteins and vaccines. A challenge for the current pipeline of AAV-based investigational products as they advance through clinical development is the identification, characterization and lot-to-lot control of the process- and product-related impurities present in even highly purified preparations. Especially challenging are AAV vector product-related impurities that closely resemble the vector itself and are, in some cases, without clear precedent in established biotherapeutic products. The determination of acceptable levels of these impurities in vectors prepared for human clinical product development, with the goal of new product licensure, requires careful risk and feasibility assessment. This review focuses primarily on the AAV product-related impurities that have been described in vectors prepared for clinical development. PMID:28548061

A new implementation of the CMRH method for solving dense linear systems

NASA Astrophysics Data System (ADS)

Heyouni, M.; Sadok, H.

2008-04-01

The CMRH method [H. Sadok, Methodes de projections pour les systemes lineaires et non lineaires, Habilitation thesis, University of Lille1, Lille, France, 1994; H. Sadok, CMRH: A new method for solving nonsymmetric linear systems based on the Hessenberg reduction algorithm, Numer. Algorithms 20 (1999) 303-321] is an algorithm for solving nonsymmetric linear systems in which the Arnoldi component of GMRES is replaced by the Hessenberg process, which generates Krylov basis vectors which are orthogonal to standard unit basis vectors rather than mutually orthogonal. The iterate is formed from these vectors by solving a small least squares problem involving a Hessenberg matrix. Like GMRES, this method requires one matrix-vector product per iteration. However, it can be implemented to require half as much arithmetic work and less storage. Moreover, numerical experiments show that this method performs accurately and reduces the residual about as fast as GMRES. With this new implementation, we show that the CMRH method is the only method with long-term recurrence which requires not storing at the same time the entire Krylov vectors basis and the original matrix as in the GMRES algorithmE A comparison with Gaussian elimination is provided.

Single-Step Conversion of Cells to Retrovirus Vector Producers with Herpes Simplex Virus–Epstein-Barr Virus Hybrid Amplicons

PubMed Central

Sena-Esteves, Miguel; Saeki, Yoshinaga; Camp, Sara M.; Chiocca, E. Antonio; Breakefield, Xandra O.

1999-01-01

We report here on the development and characterization of a novel herpes simplex virus type 1 (HSV-1) amplicon-based vector system which takes advantage of the host range and retention properties of HSV–Epstein-Barr virus (EBV) hybrid amplicons to efficiently convert cells to retrovirus vector producer cells after single-step transduction. The retrovirus genes gag-pol and env (GPE) and retroviral vector sequences were modified to minimize sequence overlap and cloned into an HSV-EBV hybrid amplicon. Retrovirus expression cassettes were used to generate the HSV-EBV-retrovirus hybrid vectors, HERE and HERA, which code for the ecotropic and the amphotropic envelopes, respectively. Retrovirus vector sequences encoding lacZ were cloned downstream from the GPE expression unit. Transfection of 293T/17 cells with amplicon plasmids yielded retrovirus titers between 106 and 107 transducing units/ml, while infection of the same cells with amplicon vectors generated maximum titers 1 order of magnitude lower. Retrovirus titers were dependent on the extent of transduction by amplicon vectors for the same cell line, but different cell lines displayed varying capacities to produce retrovirus vectors even at the same transduction efficiencies. Infection of human and dog primary gliomas with this system resulted in the production of retrovirus vectors for more than 1 week and the long-term retention and increase in transgene activity over time in these cell populations. Although the efficiency of this system still has to be determined in vivo, many applications are foreseeable for this approach to gene delivery. PMID:10559361

Viral vector-based influenza vaccines

PubMed Central

de Vries, Rory D.; Rimmelzwaan, Guus F.

2016-01-01

ABSTRACT Antigenic drift of seasonal influenza viruses and the occasional introduction of influenza viruses of novel subtypes into the human population complicate the timely production of effective vaccines that antigenically match the virus strains that cause epidemic or pandemic outbreaks. The development of game-changing vaccines that induce broadly protective immunity against a wide variety of influenza viruses is an unmet need, in which recombinant viral vectors may provide. Use of viral vectors allows the delivery of any influenza virus antigen, or derivative thereof, to the immune system, resulting in the optimal induction of virus-specific B- and T-cell responses against this antigen of choice. This systematic review discusses results obtained with vectored influenza virus vaccines and advantages and disadvantages of the currently available viral vectors. PMID:27455345

Enhancing vector shoreline data using a data fusion approach

NASA Astrophysics Data System (ADS)

Carlotto, Mark; Nebrich, Mark; DeMichele, David

2017-05-01

Vector shoreline (VSL) data is potentially useful in ATR systems that distinguish between objects on land or water. Unfortunately available data such as the NOAA 1:250,000 World Vector Shoreline and NGA Prototype Global Shoreline data cannot be used by themselves to make a land/water determination because of the manner in which the data are compiled. We describe a data fusion approach for creating labeled VSL data using test points from Global 30 Arc-Second Elevation (GTOPO30) data to determine the direction of vector segments; i.e., whether they are in clockwise or counterclockwise order. We show consistently labeled VSL data be used to easily determine whether a point is on land or water using a vector cross product test.

Viral vector-based influenza vaccines.

PubMed

de Vries, Rory D; Rimmelzwaan, Guus F

2016-11-01

Antigenic drift of seasonal influenza viruses and the occasional introduction of influenza viruses of novel subtypes into the human population complicate the timely production of effective vaccines that antigenically match the virus strains that cause epidemic or pandemic outbreaks. The development of game-changing vaccines that induce broadly protective immunity against a wide variety of influenza viruses is an unmet need, in which recombinant viral vectors may provide. Use of viral vectors allows the delivery of any influenza virus antigen, or derivative thereof, to the immune system, resulting in the optimal induction of virus-specific B- and T-cell responses against this antigen of choice. This systematic review discusses results obtained with vectored influenza virus vaccines and advantages and disadvantages of the currently available viral vectors.

Development of Novel Adenoviral Vectors to Overcome Challenges Observed With HAdV-5–based Constructs

PubMed Central

Alonso-Padilla, Julio; Papp, Tibor; Kaján, Győző L; Benkő, Mária; Havenga, Menzo; Lemckert, Angelique; Harrach, Balázs; Baker, Andrew H

2016-01-01

Recombinant vectors based on human adenovirus serotype 5 (HAdV-5) have been extensively studied in preclinical models and clinical trials over the past two decades. However, the thorough understanding of the HAdV-5 interaction with human subjects has uncovered major concerns about its product applicability. High vector-associated toxicity and widespread preexisting immunity have been shown to significantly impede the effectiveness of HAdV-5–mediated gene transfer. It is therefore that the in-depth knowledge attained working on HAdV-5 is currently being used to develop alternative vectors. Here, we provide a comprehensive overview of data obtained in recent years disqualifying the HAdV-5 vector for systemic gene delivery as well as novel strategies being pursued to overcome the limitations observed with particular emphasis on the ongoing vectorization efforts to obtain vectors based on alternative serotypes. PMID:26478249

Iterative-method performance evaluation for multiple vectors associated with a large-scale sparse matrix

NASA Astrophysics Data System (ADS)

Imamura, Seigo; Ono, Kenji; Yokokawa, Mitsuo

2016-07-01

Ensemble computing, which is an instance of capacity computing, is an effective computing scenario for exascale parallel supercomputers. In ensemble computing, there are multiple linear systems associated with a common coefficient matrix. We improve the performance of iterative solvers for multiple vectors by solving them at the same time, that is, by solving for the product of the matrices. We implemented several iterative methods and compared their performance. The maximum performance on Sparc VIIIfx was 7.6 times higher than that of a naïve implementation. Finally, to deal with the different convergence processes of linear systems, we introduced a control method to eliminate the calculation of already converged vectors.

Detection of a sudden change of the field time series based on the Lorenz system

PubMed Central

Li, Fang; Shen, BingLu; Yan, PengCheng; Song, Jian; Ma, DeShan

2017-01-01

We conducted an exploratory study of the detection of a sudden change of the field time series based on the numerical solution of the Lorenz system. First, the time when the Lorenz path jumped between the regions on the left and right of the equilibrium point of the Lorenz system was quantitatively marked and the sudden change time of the Lorenz system was obtained. Second, the numerical solution of the Lorenz system was regarded as a vector; thus, this solution could be considered as a vector time series. We transformed the vector time series into a time series using the vector inner product, considering the geometric and topological features of the Lorenz system path. Third, the sudden change of the resulting time series was detected using the sliding t-test method. Comparing the test results with the quantitatively marked time indicated that the method could detect every sudden change of the Lorenz path, thus the method is effective. Finally, we used the method to detect the sudden change of the pressure field time series and temperature field time series, and obtained good results for both series, which indicates that the method can apply to high-dimension vector time series. Mathematically, there is no essential difference between the field time series and vector time series; thus, we provide a new method for the detection of the sudden change of the field time series. PMID:28141832

The immune response induced by DNA vaccine expressing nfa1 gene against Naegleria fowleri.

PubMed

Kim, Jong-Hyun; Lee, Sang-Hee; Sohn, Hae-Jin; Lee, Jinyoung; Chwae, Yong-Joon; Park, Sun; Kim, Kyongmin; Shin, Ho-Joon

2012-12-01

The pathogenic free-living amoeba, Naegleria fowleri, causes fatal primary amoebic meningoencephalitis in experimental animals and in humans. The nfa1 gene that was cloned from N. fowleri is located on pseudopodia, especially amoebic food cups and plays an important role in the pathogenesis of N. fowleri. In this study, we constructed and characterized retroviral vector and lentiviral vector systems for nfa1 DNA vaccination in mice. We constructed the retroviral vector (pQCXIN) and the lentiviral vector (pCDH) cloned with the egfp-nfa1 gene. The expression of nfa1 gene in Chinese hamster ovary cell and human primary nasal epithelial cell transfected with the pQCXIN/egfp-nfa1 vector or pCDH/egfp-nfa1 vector was observed by fluorescent microscopy and Western blotting analysis. Our viral vector systems effectively delivered the nfa1 gene to the target cells and expressed the Nfa1 protein within the target cells. To evaluate immune responses of nfa1-vaccinated mice, BALB/c mice were intranasally vaccinated with viral particles of each retro- or lentiviral vector expressing nfa1 gene. DNA vaccination using viral vectors expressing nfa1 significantly stimulated the production of Nfa1-specific IgG subclass, as well as IgG levels. In particular, both levels of IgG2a (Th1) and IgG1 (Th2) were significantly increased in mice vaccinated with viral vectors. These results show the nfa1-vaccination induce efficiently Th1 type, as well as Th2 type immune responses. This is the first report to construct viral vector systems and to evaluate immune responses as DNA vaccination in N. fowleri infection. Furthermore, these results suggest that nfal vaccination may be an effective method for treatment of N. fowleri infection.

Virtual world reconstruction using the modeling and simulation extended vector product prototype

DOT National Transportation Integrated Search

1997-05-30

The MS Extended Vector Product (MSEVP) prototype being developed is an extended vector product format-based product containing a continuous surface representation and a consistent view of elevation across the thematic coverages contained within a dat...

Systemic errors in quantitative polymerase chain reaction titration of self-complementary adeno-associated viral vectors and improved alternative methods.

PubMed

Fagone, Paolo; Wright, J Fraser; Nathwani, Amit C; Nienhuis, Arthur W; Davidoff, Andrew M; Gray, John T

2012-02-01

Self-complementary AAV (scAAV) vector genomes contain a covalently closed hairpin derived from a mutated inverted terminal repeat that connects the two monomer single-stranded genomes into a head-to-head or tail-to-tail dimer. We found that during quantitative PCR (qPCR) this structure inhibits the amplification of proximal amplicons and causes the systemic underreporting of copy number by as much as 10-fold. We show that cleavage of scAAV vector genomes with restriction endonuclease to liberate amplicons from the covalently closed terminal hairpin restores quantitative amplification, and we implement this procedure in a simple, modified qPCR titration method for scAAV vectors. In addition, we developed and present an AAV genome titration procedure based on gel electrophoresis that requires minimal sample processing and has low interassay variability, and as such is well suited for the rigorous quality control demands of clinical vector production facilities.

Matrix multiplication operations using pair-wise load and splat operations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eichenberger, Alexandre E.; Gschwind, Michael K.; Gunnels, John A.

Mechanisms for performing a matrix multiplication operation are provided. A vector load operation is performed to load a first vector operand of the matrix multiplication operation to a first target vector register. A pair-wise load and splat operation is performed to load a pair of scalar values of a second vector operand and replicate the pair of scalar values within a second target vector register. An operation is performed on elements of the first target vector register and elements of the second target vector register to generate a partial product of the matrix multiplication operation. The partial product is accumulatedmore » with other partial products and a resulting accumulated partial product is stored. This operation may be repeated for a second pair of scalar values of the second vector operand.« less

Relevance of Assembly-Activating Protein for Adeno-associated Virus Vector Production and Capsid Protein Stability in Mammalian and Insect Cells.

PubMed

Grosse, Stefanie; Penaud-Budloo, Magalie; Herrmann, Anne-Kathrin; Börner, Kathleen; Fakhiri, Julia; Laketa, Vibor; Krämer, Chiara; Wiedtke, Ellen; Gunkel, Manuel; Ménard, Lucie; Ayuso, Eduard; Grimm, Dirk

2017-10-15

The discovery that adeno-associated virus 2 (AAV2) encodes an eighth protein, called assembly-activating protein (AAP), transformed our understanding of wild-type AAV biology. Concurrently, it raised questions about the role of AAP during production of recombinant vectors based on natural or molecularly engineered AAV capsids. Here, we show that AAP is indeed essential for generation of functional recombinant AAV2 vectors in both mammalian and insect cell-based vector production systems. Surprisingly, we observed that AAV2 capsid proteins VP1 to -3 are unstable in the absence of AAP2, likely due to rapid proteasomal degradation. Inhibition of the proteasome led to an increase of intracellular VP1 to -3 but neither triggered assembly of functional capsids nor promoted nuclear localization of the capsid proteins. Together, this underscores the crucial and unique role of AAP in the AAV life cycle, where it rapidly chaperones capsid assembly, thus preventing degradation of free capsid proteins. An expanded analysis comprising nine alternative AAV serotypes (1, 3 to 9, and rh10) showed that vector production always depends on the presence of AAP, with the exceptions of AAV4 and AAV5, which exhibited AAP-independent, albeit low-level, particle assembly. Interestingly, AAPs from all 10 serotypes could cross-complement AAP-depleted helper plasmids during vector production, despite there being distinct intracellular AAP localization patterns. These were most pronounced for AAP4 and AAP5, congruent with their inability to rescue an AAV2/AAP2 knockout. We conclude that AAP is key for assembly of genuine capsids from at least 10 different AAV serotypes, which has implications for vectors derived from wild-type or synthetic AAV capsids. IMPORTANCE Assembly of adeno-associated virus 2 (AAV2) is regulated by the assembly-activating protein (AAP), whose open reading frame overlaps with that of the viral capsid proteins. As the majority of evidence was obtained using virus-like particles composed solely of the major capsid protein VP3, AAP's role in and relevance for assembly of genuine AAV capsids have remained largely unclear. Thus, we established a trans -complementation assay permitting assessment of AAP functionality during production of recombinant vectors based on complete AAV capsids and derived from any serotype. We find that AAP is indeed a critical factor not only for AAV2, but also for generation of vectors derived from nine other AAV serotypes. Moreover, we identify a new role of AAP in maintaining capsid protein stability in mammalian and insect cells. Thereby, our study expands our current understanding of AAV/AAP biology, and it concomitantly provides insights into the importance of AAP for AAV vector production. Copyright © 2017 American Society for Microbiology.

Relevance of Assembly-Activating Protein for Adeno-associated Virus Vector Production and Capsid Protein Stability in Mammalian and Insect Cells

PubMed Central

Grosse, Stefanie; Penaud-Budloo, Magalie; Herrmann, Anne-Kathrin; Börner, Kathleen; Fakhiri, Julia; Laketa, Vibor; Krämer, Chiara; Wiedtke, Ellen; Gunkel, Manuel; Ménard, Lucie; Ayuso, Eduard

2017-01-01

ABSTRACT The discovery that adeno-associated virus 2 (AAV2) encodes an eighth protein, called assembly-activating protein (AAP), transformed our understanding of wild-type AAV biology. Concurrently, it raised questions about the role of AAP during production of recombinant vectors based on natural or molecularly engineered AAV capsids. Here, we show that AAP is indeed essential for generation of functional recombinant AAV2 vectors in both mammalian and insect cell-based vector production systems. Surprisingly, we observed that AAV2 capsid proteins VP1 to -3 are unstable in the absence of AAP2, likely due to rapid proteasomal degradation. Inhibition of the proteasome led to an increase of intracellular VP1 to -3 but neither triggered assembly of functional capsids nor promoted nuclear localization of the capsid proteins. Together, this underscores the crucial and unique role of AAP in the AAV life cycle, where it rapidly chaperones capsid assembly, thus preventing degradation of free capsid proteins. An expanded analysis comprising nine alternative AAV serotypes (1, 3 to 9, and rh10) showed that vector production always depends on the presence of AAP, with the exceptions of AAV4 and AAV5, which exhibited AAP-independent, albeit low-level, particle assembly. Interestingly, AAPs from all 10 serotypes could cross-complement AAP-depleted helper plasmids during vector production, despite there being distinct intracellular AAP localization patterns. These were most pronounced for AAP4 and AAP5, congruent with their inability to rescue an AAV2/AAP2 knockout. We conclude that AAP is key for assembly of genuine capsids from at least 10 different AAV serotypes, which has implications for vectors derived from wild-type or synthetic AAV capsids. IMPORTANCE Assembly of adeno-associated virus 2 (AAV2) is regulated by the assembly-activating protein (AAP), whose open reading frame overlaps with that of the viral capsid proteins. As the majority of evidence was obtained using virus-like particles composed solely of the major capsid protein VP3, AAP's role in and relevance for assembly of genuine AAV capsids have remained largely unclear. Thus, we established a trans-complementation assay permitting assessment of AAP functionality during production of recombinant vectors based on complete AAV capsids and derived from any serotype. We find that AAP is indeed a critical factor not only for AAV2, but also for generation of vectors derived from nine other AAV serotypes. Moreover, we identify a new role of AAP in maintaining capsid protein stability in mammalian and insect cells. Thereby, our study expands our current understanding of AAV/AAP biology, and it concomitantly provides insights into the importance of AAP for AAV vector production. PMID:28768875

Fundamental Principles of Classical Mechanics: a Geometrical Perspectives

NASA Astrophysics Data System (ADS)

Lam, Kai S.

2014-07-01

Classical mechanics is the quantitative study of the laws of motion for oscopic physical systems with mass. The fundamental laws of this subject, known as Newton's Laws of Motion, are expressed in terms of second-order differential equations governing the time evolution of vectors in a so-called configuration space of a system (see Chapter 12). In an elementary setting, these are usually vectors in 3-dimensional Euclidean space, such as position vectors of point particles; but typically they can be vectors in higher dimensional and more abstract spaces. A general knowledge of the mathematical properties of vectors, not only in their most intuitive incarnations as directed arrows in physical space but as elements of abstract linear vector spaces, and those of linear operators (transformations) on vector spaces as well, is then indispensable in laying the groundwork for both the physical and the more advanced mathematical - more precisely topological and geometrical - concepts that will prove to be vital in our subject. In this beginning chapter we will review these properties, and introduce the all-important related notions of dual spaces and tensor products of vector spaces. The notational convention for vectorial and tensorial indices used for the rest of this book (except when otherwise specified) will also be established...

Management of whitefly-transmitted viruses in open-field production systems

USDA-ARS?s Scientific Manuscript database

Whiteflies are a key pest of crops in open field production throughout the tropics and subtropics. This is due in large part to the long and diverse list of devastating plant viruses transmitted by these vectors. Open field production provides many challenges to manage these viruses and in many case...

Hypoxia-inducible vascular endothelial growth factor gene therapy using the oxygen-dependent degradation domain in myocardial ischemia.

PubMed

Kim, Hyun Ah; Lim, Soyeon; Moon, Hyung-Ho; Kim, Sung Wan; Hwang, Ki-Chul; Lee, Minhyung; Kim, Sun Hwa; Choi, Donghoon

2010-10-01

A hypoxia-inducible VEGF expression system with the oxygen-dependent degradation (ODD) domain was constructed and tested to be used in gene therapy for ischemic myocardial disease. Luciferase and VEGF expression vector systems were constructed with or without the ODD domain: pEpo-SV-Luc (or pEpo-SV-VEGF) and pEpo-SV-Luc-ODD (or pEpo-SV-VEGF-ODD). In vitro gene expression efficiency of each vector type was evaluated in HEK 293 cells under both hypoxic and normoxic conditions. The amount of VEGF protein was estimated by ELISA. The VEGF expression vectors with or without the ODD domain were injected into ischemic rat myocardium. Fibrosis, neovascularization, and cardiomyocyte apoptosis were assessed using Masson's trichrome staining, α-smooth muscle actin (α-SMA) immunostaining, and the TUNEL assay, respectively. The plasmid vectors containing ODD significantly improved the expression level of VEGF protein in hypoxic conditions. The enhancement of VEGF protein production was attributed to increased protein stability due to oxygen deficiency. In a rat model of myocardial ischemia, the pEpo-SV-VEGF-ODD group exhibited less myocardial fibrosis, higher microvessel density, and less cardiomyocyte apoptosis compared to the control groups (saline and pEpo-SV-VEGF treatments). An ODD-mediated VEGF expression system that facilitates VEGF-production under hypoxia may be useful in the treatment of ischemic heart disease.

Manga Vectorization and Manipulation with Procedural Simple Screentone.

PubMed

Yao, Chih-Yuan; Hung, Shih-Hsuan; Li, Guo-Wei; Chen, I-Yu; Adhitya, Reza; Lai, Yu-Chi

2017-02-01

Manga are a popular artistic form around the world, and artists use simple line drawing and screentone to create all kinds of interesting productions. Vectorization is helpful to digitally reproduce these elements for proper content and intention delivery on electronic devices. Therefore, this study aims at transforming scanned Manga to a vector representation for interactive manipulation and real-time rendering with arbitrary resolution. Our system first decomposes the patch into rough Manga elements including possible borders and shading regions using adaptive binarization and screentone detector. We classify detected screentone into simple and complex patterns: our system extracts simple screentone properties for refining screentone borders, estimating lighting, compensating missing strokes inside screentone regions, and later resolution independently rendering with our procedural shaders. Our system treats the others as complex screentone areas and vectorizes them with our proposed line tracer which aims at locating boundaries of all shading regions and polishing all shading borders with the curve-based Gaussian refiner. A user can lay down simple scribbles to cluster Manga elements intuitively for the formation of semantic components, and our system vectorizes these components into shading meshes along with embedded Bézier curves as a unified foundation for consistent manipulation including pattern manipulation, deformation, and lighting addition. Our system can real-time and resolution independently render the shading regions with our procedural shaders and drawing borders with the curve-based shader. For Manga manipulation, the proposed vector representation can be not only magnified without artifacts but also deformed easily to generate interesting results.

Scalar products of Bethe vectors in models with {\\mathfrak{gl}}(2| 1) symmetry 1. Super-analog of Reshetikhin formula

NASA Astrophysics Data System (ADS)

Hutsalyuk, A.; Liashyk, A.; Pakuliak, S. Z.; Ragoucy, E.; Slavnov, N. A.

2016-11-01

We study the scalar products of Bethe vectors in integrable models solvable by the nested algebraic Bethe ansatz and possessing {gl}(2| 1) symmetry. Using explicit formulas of the monodromy matrix entries’ multiple actions onto Bethe vectors we obtain a representation for the scalar product in the most general case. This explicit representation appears to be a sum over partitions of the Bethe parameters. It can be used for the analysis of scalar products involving on-shell Bethe vectors. As a by-product, we obtain a determinant representation for the scalar products of generic Bethe vectors in integrable models with {gl}(1| 1) symmetry. Dedicated to the memory of Petr Petrovich Kulish.

Studies of Zγ production in association with a high-mass dijet system in pp collisions at $$ \\sqrt{s}=8$$ TeV with the ATLAS detector

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aaboud, M.; Aad, G.; Abbott, B.

2017-07-21

The production of a Z boson and a photon in association with a high-mass dijet system is studied using 20.2 fb -1 of proton-proton collision data at a centre-of-mass energy ofmore » $$\\sqrt{s}$$ = 8 TeV recorded with the ATLAS detector in 2012 at the Large Hadron Collider. Final states with a photon and a Z boson decaying into a pair of either electrons, muons, or neutrinos are analysed. Electroweak and total pp → Zγjj cross-sections are extracted in two fiducial regions with different sensitivities to electroweak production processes. Quartic couplings of vector bosons are studied in regions of phase space with an enhanced contribution from pure electroweak production, sensitive to vector-boson scattering processes VV → Zγ. Finally, no deviations from Standard Model predictions are observed and constraints are placed on anomalous couplings parameterized by higher-dimensional operators using effective field theory.« less

Studies of Zγ production in association with a high-mass dijet system in pp collisions at $$\\sqrt{s}=8$$ TeV with the ATLAS detector

DOE PAGES

Aaboud, M.; Aad, G.; Abbott, B.; ...

2017-07-21

The production of a Z boson and a photon in association with a high-mass dijet system is studied using 20.2 fb -1 of proton-proton collision data at a centre-of-mass energy ofmore » $$\\sqrt{s}$$ = 8 TeV recorded with the ATLAS detector in 2012 at the Large Hadron Collider. Final states with a photon and a Z boson decaying into a pair of either electrons, muons, or neutrinos are analysed. Electroweak and total pp → Zγjj cross-sections are extracted in two fiducial regions with different sensitivities to electroweak production processes. Quartic couplings of vector bosons are studied in regions of phase space with an enhanced contribution from pure electroweak production, sensitive to vector-boson scattering processes VV → Zγ. Finally, no deviations from Standard Model predictions are observed and constraints are placed on anomalous couplings parameterized by higher-dimensional operators using effective field theory.« less

Analysis of Genetic Algorithm for Rule-Set Production (GARP) modeling approach for predicting distributions of fleas implicated as vectors of plague, Yersinia pestis, in California.

PubMed

Adjemian, Jennifer C Z; Girvetz, Evan H; Beckett, Laurel; Foley, Janet E

2006-01-01

More than 20 species of fleas in California are implicated as potential vectors of Yersinia pestis. Extremely limited spatial data exist for plague vectors-a key component to understanding where the greatest risks for human, domestic animal, and wildlife health exist. This study increases the spatial data available for 13 potential plague vectors by using the ecological niche modeling system Genetic Algorithm for Rule-Set Production (GARP) to predict their respective distributions. Because the available sample sizes in our data set varied greatly from one species to another, we also performed an analysis of the robustness of GARP by using the data available for flea Oropsylla montana (Baker) to quantify the effects that sample size and the chosen explanatory variables have on the final species distribution map. GARP effectively modeled the distributions of 13 vector species. Furthermore, our analyses show that all of these modeled ranges are robust, with a sample size of six fleas or greater not significantly impacting the percentage of the in-state area where the flea was predicted to be found, or the testing accuracy of the model. The results of this study will help guide the sampling efforts of future studies focusing on plague vectors.

Viral vectors for production of recombinant proteins in plants.

PubMed

Lico, Chiara; Chen, Qiang; Santi, Luca

2008-08-01

Global demand for recombinant proteins has steadily accelerated for the last 20 years. These recombinant proteins have a wide range of important applications, including vaccines and therapeutics for human and animal health, industrial enzymes, new materials and components of novel nano-particles for various applications. The majority of recombinant proteins are produced by traditional biological "factories," that is, predominantly mammalian and microbial cell cultures along with yeast and insect cells. However, these traditional technologies cannot satisfy the increasing market demand due to prohibitive capital investment requirements. During the last two decades, plants have been under intensive investigation to provide an alternative system for cost-effective, highly scalable, and safe production of recombinant proteins. Although the genetic engineering of plant viral vectors for heterologous gene expression can be dated back to the early 1980s, recent understanding of plant virology and technical progress in molecular biology have allowed for significant improvements and fine tuning of these vectors. These breakthroughs enable the flourishing of a variety of new viral-based expression systems and their wide application by academic and industry groups. In this review, we describe the principal plant viral-based production strategies and the latest plant viral expression systems, with a particular focus on the variety of proteins produced and their applications. We will summarize the recent progress in the downstream processing of plant materials for efficient extraction and purification of recombinant proteins. (c) 2008 Wiley-Liss, Inc.

Recent Weather Extremes and Impacts on Agricultural Production and Vector-Borne Disease Outbreak Patterns

DTIC Science & Technology

2014-03-21

funding from USDA Foreign Agricultural Service towards the Global Agricultural Monitoring project, DoD Armed Forces Health Surveillance Center’s...Global Emerging Infections Surveillance and Response System (AFHSC/GEIS) under the Human Febrile and Vector -Borne Illnesses (FVBI) Program and USDA ...outbreaks during the 2010?2012 period. We utilized 2000?2012 vegetation index and land surface temperature data from NASA ?s satellitebased Moderate

Evaluation Method for Service Branding Using Word-of-Mouth Data

NASA Astrophysics Data System (ADS)

Shirahada, Kunio; Kosaka, Michitaka

Development and spread of internet technology contributes service firms to obtaining the high capability of brand information transmission as well as relative customer feedback data collection. In this paper, we propose a new evaluation method for service branding using firms and consumers data on the internet. Based on service marketing 7Ps (Product, Price, Place, Promotion, People, Physical evidence, Process) which are the key viewpoints for branding, we develop a brand evaluation system including coding methods for Word-of-Mouth (WoM) and corporate introductory information on the internet to identify both customer's service value recognition vector and firm's service value proposition vector. Our system quantitatively clarify both customer's service value recognition of the firm and firm's strength in service value proposition, thereby analyzing service brand communication gaps between firm and consumers. We applied this system to Japanese Ryokan hotel industry. Using six ryokan-hotels' data on Jyaran-net and Rakuten travel, we made totally 983 codes from WoM information and analyzed their service brand value according to three price based categories. As a result, we found that the characteristics of customers' service value recognition vector differ according to the price categories. In addition, the system clarified that there is a firm that has a different service value proposition vector from customers' recognition vector. This helps to analyze corporate service brand strategy and has a significance as a system technology supporting service management.

Existence of frozen-in coordinate systems

NASA Technical Reports Server (NTRS)

Chertkov, A. D.

1995-01-01

The 'frozen-in' coordinate systems were first introduced in the works on 'reconnection' and 'magnetic barrier' theories (see review by M.l.Pudovkin and V.S.Semenov, Space Sci. Rev. 41,1 1985). The idea was to utilize the mathematical apparatus developed for 'general relativity' theory to simplify obtaining solutions to the ideal MHD equations set. Magnetic field (B), plasma velocity (v), and their vector product were used as coordinate vectors. But there exist no stationary solutions of ideal MHD set that satisfies the required boundary conditions at infinity (A.D.Chertkov, Solar Wind Seven Conf.,Pergamon Press,1992,165) having non-zero vector product of v and B where v and B originate from the same sphere. The existence of a solution is the hidden mine of the mentioned theories. The solution is constructed in the coordinate system, which is unknown and indeterminate before obtaining this solution. A substitution of the final solution must be done directly into the initial MHD set in order to check the method. One can demonstrate that 'solutions' of Petschek's problem, obtained by 'frozen-in' coordinate systems, does not satisfy just the 'frozen-in' equation, i.e. induction equation. It stems from the fact that Petschek's 're-connection' model, treated as a boundary problem, is over determined. This problem was incorrectly formulated.

AN ADA LINEAR ALGEBRA PACKAGE MODELED AFTER HAL/S

NASA Technical Reports Server (NTRS)

Klumpp, A. R.

1994-01-01

This package extends the Ada programming language to include linear algebra capabilities similar to those of the HAL/S programming language. The package is designed for avionics applications such as Space Station flight software. In addition to the HAL/S built-in functions, the package incorporates the quaternion functions used in the Shuttle and Galileo projects, and routines from LINPAK that solve systems of equations involving general square matrices. Language conventions in this package follow those of HAL/S to the maximum extent practical and minimize the effort required for writing new avionics software and translating existent software into Ada. Valid numeric types in this package include scalar, vector, matrix, and quaternion declarations. (Quaternions are fourcomponent vectors used in representing motion between two coordinate frames). Single precision and double precision floating point arithmetic is available in addition to the standard double precision integer manipulation. Infix operators are used instead of function calls to define dot products, cross products, quaternion products, and mixed scalar-vector, scalar-matrix, and vector-matrix products. The package contains two generic programs: one for floating point, and one for integer. The actual component type is passed as a formal parameter to the generic linear algebra package. The procedures for solving systems of linear equations defined by general matrices include GEFA, GECO, GESL, and GIDI. The HAL/S functions include ABVAL, UNIT, TRACE, DET, INVERSE, TRANSPOSE, GET, PUT, FETCH, PLACE, and IDENTITY. This package is written in Ada (Version 1.2) for batch execution and is machine independent. The linear algebra software depends on nothing outside the Ada language except for a call to a square root function for floating point scalars (such as SQRT in the DEC VAX MATHLIB library). This program was developed in 1989, and is a copyrighted work with all copyright vested in NASA.

SCI Hazard Report Methodology

NASA Technical Reports Server (NTRS)

Mitchell, Michael S.

2010-01-01

This slide presentation reviews the methodology in creating a Source Control Item (SCI) Hazard Report (HR). The SCI HR provides a system safety risk assessment for the following Ares I Upper Stage Production Contract (USPC) components (1) Pyro Separation Systems (2) Main Propulsion System (3) Reaction and Roll Control Systems (4) Thrust Vector Control System and (5) Ullage Settling Motor System components.

Secreted expression of Leuconostoc mesenteroides glucansucrase in Lactococcus lactis for the production of insoluble glucans

USDA-ARS?s Scientific Manuscript database

We expressed a glucansucrase, DsrI, from Leuconostoc mesenteroides that catalyzes formation of water-insoluble glucans from sucrose in Lactococcus lactis using a nisin-controlled gene expression system. Production of DsrI was optimized using several different background vectors, signal peptides, str...

Diagnosis of nutrient imbalances with vector analysis in agroforestry systems.

PubMed

Isaac, Marney E; Kimaro, Anthony A

2011-01-01

Agricultural intensification has had unintended environmental consequences, including increased nutrient leaching and surface runoff and other agrarian-derived pollutants. Improved diagnosis of on-farm nutrient dynamics will have the advantage of increasing yields and will diminish financial and environmental costs. To achieve this, a management support system that allows for site-specific rapid evaluation of nutrient production imbalances and subsequent management prescriptions is needed for agroecological design. Vector diagnosis, a bivariate model to depict changes in yield and nutritional response simultaneously in a single graph, facilitates identification of nutritional status such as growth dilution, deficiency, sufficiency, luxury uptake, and toxicity. Quantitative data from cocoa agroforestry systems and pigeonpea intercropping trials in Ghana and Tanzania, respectively, were re-evaluated with vector analysis. Relative to monoculture, biomass increase in cocoa ( L.) under shade (35-80%) was accompanied by a 17 to 25% decline in P concentration, the most limiting nutrient on this site. Similarly, increasing biomass with declining P concentrations was noted for pigeonpea [ (L). Millsp.] in response to soil moisture availability under intercropping. Although vector analysis depicted nutrient responses, the current vector model does not consider non-nutrient resource effects on growth, such as ameliorated light and soil moisture, which were particularly active in these systems. We revisit and develop vector analysis into a framework for diagnosing nutrient and non-nutrient interactions in agroforestry systems. Such a diagnostic technique advances management decision-making by increasing nutrient precision and reducing environmental issues associated with agrarian-derived soil contamination. American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America.

Optoelectronic Inner-Product Neural Associative Memory

NASA Technical Reports Server (NTRS)

Liu, Hua-Kuang

1993-01-01

Optoelectronic apparatus acts as artificial neural network performing associative recall of binary images. Recall process is iterative one involving optical computation of inner products between binary input vector and one or more reference binary vectors in memory. Inner-product method requires far less memory space than matrix-vector method.

A straightforward characterization of non-modal effects from the evolution of linear dynamical systems

NASA Astrophysics Data System (ADS)

Arratia, Cristobal

2014-11-01

A simple construction will be shown, which reveals a general property satisfied by the evolution in time of a state vector composed by a superposition of orthogonal eigenmodes of a linear dynamical system. This property results from the conservation of the inner product between such state vectors evolving forward and backwards in time, and it can be simply evaluated from the state vector and its first and second time derivatives. This provides an efficient way to characterize, instantaneously along any specific phase-space trajectory of the linear system, the relevance of the non-normality of the linearized Navier-Stokes operator on the energy (or any other norm) gain or decay of small perturbations. Examples of this characterization applied to stationary or time dependent base flows will be shown. CONICYT, Concurso de Apoyo al Retorno de Investigadores del Extranjero, folio 821320055.

Upgrades to the NOAA/NESDIS automated Cloud-Motion Vector system

NASA Technical Reports Server (NTRS)

Nieman, Steve; Menzel, W. Paul; Hayden, Christopher M.; Wanzong, Steve; Velden, Christopher S.

1993-01-01

The latest version of the automated cloud motion vector software has yielded significant improvements in the quality of the GOES cloud-drift winds produced operationally by NESDIS. Cloud motion vectors resulting from the automated system are now equal or superior in quality to those which had the benefit of manual quality control a few years ago. The single most important factor in this improvement has been the upgraded auto-editor. Improved tracer selection procedures eliminate targets in difficult regions and allow a higher target density and therefore enhanced coverage in areas of interest. The incorporation of the H2O-intercept height assignment method allows an adequate representation of the heights of semi-transparent clouds in the absence of a CO2-absorption channel. Finally, GOES-8 water-vapor motion winds resulting from the automated system are superior to any done previously by NESDIS and should now be considered as an operational product.

A modular toolset for recombination transgenesis and neurogenetic analysis of Drosophila.

PubMed

Wang, Ji-Wu; Beck, Erin S; McCabe, Brian D

2012-01-01

Transgenic Drosophila have contributed extensively to our understanding of nervous system development, physiology and behavior in addition to being valuable models of human neurological disease. Here, we have generated a novel series of modular transgenic vectors designed to optimize and accelerate the production and analysis of transgenes in Drosophila. We constructed a novel vector backbone, pBID, that allows both phiC31 targeted transgene integration and incorporates insulator sequences to ensure specific and uniform transgene expression. Upon this framework, we have built a series of constructs that are either backwards compatible with existing restriction enzyme based vectors or utilize Gateway recombination technology for high-throughput cloning. These vectors allow for endogenous promoter or Gal4 targeted expression of transgenic proteins with or without fluorescent protein or epitope tags. In addition, we have generated constructs that facilitate transgenic splice isoform specific RNA inhibition of gene expression. We demonstrate the utility of these constructs to analyze proteins involved in nervous system development, physiology and neurodegenerative disease. We expect that these reagents will facilitate the proficiency and sophistication of Drosophila genetic analysis in both the nervous system and other tissues.

Critical design criteria for minimal antibiotic-free plasmid vectors necessary to combine robust RNA Pol II and Pol III-mediated eukaryotic expression with high bacterial production yields

PubMed Central

Carnes, Aaron E.; Luke, Jeremy M.; Vincent, Justin M.; Anderson, Sheryl; Schukar, Angela; Hodgson, Clague P.; Williams, James A.

2010-01-01

Background For safety considerations, regulatory agencies recommend elimination of antibiotic resistance markers and nonessential sequences from plasmid DNA-based gene medicines. In the present study we analyzed antibiotic-free (AF) vector design criteria impacting bacterial production and mammalian transgene expression. Methods Both CMV-HTLV-I R RNA Pol II promoter (protein transgene) and murine U6 RNA Pol III promoter (RNA transgene) vector designs were studied. Plasmid production yield was assessed through inducible fed-batch fermentation. RNA Pol II-directed EGFP and RNA Pol III-directed RNA expression were quantified by fluorometry and quantitative real-time polymerase chain reaction (RT-PCR), respectively, after transfection of human HEK293 cells. Results Sucrose-selectable minimalized protein and therapeutic RNA expression vector designs that combined an RNA-based AF selection with highly productive fermentation manufacturing (>1,000 mg/L plasmid DNA) and high level in vivo expression of encoded products were identified. The AF selectable marker was also successfully applied to convert existing kanamycin-resistant DNA vaccine plasmids gWIZ and pVAX1 into AF vectors, demonstrating a general utility for retrofitting existing vectors. A minimum vector size for high yield plasmid fermentation was identified. A strategy for stable fermentation of plasmid dimers with improved vector potency and fermentation yields up to 1,740 mg/L was developed. Conclusions We report the development of potent high yield AF gene medicine expression vectors for protein or RNA (e.g. short hairpin RNA or microRNA) products. These AF expression vectors were optimized to exceed a newly identified size threshold for high copy plasmid replication and direct higher transgene expression levels than alternative vectors. PMID:20806425

A novel system for the production of high levels of functional human therapeutic proteins in stable cells with a Semliki Forest virus noncytopathic vector.

PubMed

Casales, Erkuden; Aranda, Alejandro; Quetglas, Jose I; Ruiz-Guillen, Marta; Rodriguez-Madoz, Juan R; Prieto, Jesus; Smerdou, Cristian

2010-05-31

Semliki Forest virus (SFV) vectors lead to high protein expression in mammalian cells, but expression is transient due to vector cytopathic effects, inhibition of host cell proteins and RNA-based expression. We have used a noncytopathic SFV mutant (ncSFV) RNA vector to generate stable cell lines expressing two human therapeutic proteins: insulin-like growth factor I (IGF-I) and cardiotrophin-1 (CT-1). Therapeutic genes were fused at the carboxy-terminal end of Puromycin N-acetyl-transferase gene by using as a linker the sequence coding for foot-and-mouth disease virus (FMDV) 2A autoprotease. These cassettes were cloned into the ncSFV vector. Recombinant ncSFV vectors allowed rapid and efficient selection of stable BHK cell lines with puromycin. These cells expressed IGF-I and CT-1 in supernatants at levels reaching 1.4 and 8.6 microg/10(6)cells/24 hours, respectively. Two cell lines generated with each vector were passaged ten times during 30 days, showing constant levels of protein expression. Recombinant proteins expressed at different passages were functional by in vitro signaling assays. Stability at RNA level was unexpectedly high, showing a very low mutation rate in the CT-1 sequence, which did not increase at high passages. CT-1 was efficiently purified from supernatants of ncSFV cell lines, obtaining a yield of approximately 2mg/L/24 hours. These results indicate that the ncSFV vector has a great potential for the production of recombinant proteins in mammalian cells. 2010 Elsevier B.V. All rights reserved.

Matrix multiplication operations with data pre-conditioning in a high performance computing architecture

DOEpatents

Eichenberger, Alexandre E; Gschwind, Michael K; Gunnels, John A

2013-11-05

Mechanisms for performing matrix multiplication operations with data pre-conditioning in a high performance computing architecture are provided. A vector load operation is performed to load a first vector operand of the matrix multiplication operation to a first target vector register. A load and splat operation is performed to load an element of a second vector operand and replicating the element to each of a plurality of elements of a second target vector register. A multiply add operation is performed on elements of the first target vector register and elements of the second target vector register to generate a partial product of the matrix multiplication operation. The partial product of the matrix multiplication operation is accumulated with other partial products of the matrix multiplication operation.

On approximately symmetric informationally complete positive operator-valued measures and related systems of quantum states

NASA Astrophysics Data System (ADS)

Klappenecker, Andreas; Rötteler, Martin; Shparlinski, Igor E.; Winterhof, Arne

2005-08-01

We address the problem of constructing positive operator-valued measures (POVMs) in finite dimension n consisting of n2 operators of rank one which have an inner product close to uniform. This is motivated by the related question of constructing symmetric informationally complete POVMs (SIC-POVMs) for which the inner products are perfectly uniform. However, SIC-POVMs are notoriously hard to construct and, despite some success of constructing them numerically, there is no analytic construction known. We present two constructions of approximate versions of SIC-POVMs, where a small deviation from uniformity of the inner products is allowed. The first construction is based on selecting vectors from a maximal collection of mutually unbiased bases and works whenever the dimension of the system is a prime power. The second construction is based on perturbing the matrix elements of a subset of mutually unbiased bases. Moreover, we construct vector systems in Cn which are almost orthogonal and which might turn out to be useful for quantum computation. Our constructions are based on results of analytic number theory.

A remote sensing and geographic information system approach to sampling malaria vector habitats in Chiapas, Mexico

NASA Astrophysics Data System (ADS)

Beck, L.; Wood, B.; Whitney, S.; Rossi, R.; Spanner, M.; Rodriguez, M.; Rodriguez-Ramirez, A.; Salute, J.; Legters, L.; Roberts, D.; Rejmankova, E.; Washino, R.

1993-08-01

This paper describes a procedure whereby remote sensing and geographic information system (GIS) technologies are used in a sample design to study the habitat of Anopheles albimanus, one of the principle vectors of malaria in Central America. This procedure incorporates Landsat-derived land cover maps with digital elevation and road network data to identify a random selection of larval habitats accessible for field sampling. At the conclusion of the sampling season, the larval counts will be used to determine habitat productivity, and then integrated with information on human settlement to assess where people are at high risk of malaria. This aproach would be appropriate in areas where land cover information is lacking and problems of access constrain field sampling. The use of a GIS also permits other data (such as insecticide spraying data) to the incorporated in the sample design as they arise. This approach would also be pertinent for other tropical vector-borne diseases, particularly where human activities impact disease vector habitat.

Heterologous Protein Secretion in Lactobacilli with Modified pSIP Vectors

PubMed Central

Karlskås, Ingrid Lea; Maudal, Kristina; Axelsson, Lars; Rud, Ida; Eijsink, Vincent G. H.; Mathiesen, Geir

2014-01-01

We describe new variants of the modular pSIP-vectors for inducible gene expression and protein secretion in lactobacilli. The basic functionality of the pSIP system was tested in Lactobacillus strains representing 14 species using pSIP411, which harbors the broad-host-range Lactococcus lactis SH71rep replicon and a β-glucuronidase encoding reporter gene. In 10 species, the inducible gene expression system was functional. Based on these results, three pSIP vectors with different signal peptides were modified by replacing their narrow-host-range L. plantarum 256rep replicon with SH71rep and transformed into strains of five different species of Lactobacillus. All recombinant strains secreted the target protein NucA, albeit with varying production levels and secretion efficiencies. The Lp_3050 derived signal peptide generally resulted in the highest levels of secreted NucA. These modified pSIP vectors are useful tools for engineering a wide variety of Lactobacillus species. PMID:24614815

Vector production in an academic environment: a tool to assess production costs.

PubMed

Boeke, Aaron; Doumas, Patrick; Reeves, Lilith; McClurg, Kyle; Bischof, Daniela; Sego, Lina; Auberry, Alisha; Tatikonda, Mohan; Cornetta, Kenneth

2013-02-01

Generating gene and cell therapy products under good manufacturing practices is a complex process. When determining the cost of these products, researchers must consider the large number of supplies used for manufacturing and the personnel and facility costs to generate vector and maintain a cleanroom facility. To facilitate cost estimates, the Indiana University Vector Production Facility teamed with the Indiana University Kelley School of Business to develop a costing tool that, in turn, provides pricing. The tool is designed in Microsoft Excel and is customizable to meet the needs of other core facilities. It is available from the National Gene Vector Biorepository. The tool allows cost determinations using three different costing methods and was developed in an effort to meet the A21 circular requirements for U.S. core facilities performing work for federally funded projects. The costing tool analysis reveals that the cost of vector production does not have a linear relationship with batch size. For example, increasing the production from 9 to18 liters of a retroviral vector product increases total costs a modest 1.2-fold rather than doubling in total cost. The analysis discussed in this article will help core facilities and investigators plan a cost-effective strategy for gene and cell therapy production.

Vector Production in an Academic Environment: A Tool to Assess Production Costs

PubMed Central

Boeke, Aaron; Doumas, Patrick; Reeves, Lilith; McClurg, Kyle; Bischof, Daniela; Sego, Lina; Auberry, Alisha; Tatikonda, Mohan

2013-01-01

Abstract Generating gene and cell therapy products under good manufacturing practices is a complex process. When determining the cost of these products, researchers must consider the large number of supplies used for manufacturing and the personnel and facility costs to generate vector and maintain a cleanroom facility. To facilitate cost estimates, the Indiana University Vector Production Facility teamed with the Indiana University Kelley School of Business to develop a costing tool that, in turn, provides pricing. The tool is designed in Microsoft Excel and is customizable to meet the needs of other core facilities. It is available from the National Gene Vector Biorepository. The tool allows cost determinations using three different costing methods and was developed in an effort to meet the A21 circular requirements for U.S. core facilities performing work for federally funded projects. The costing tool analysis reveals that the cost of vector production does not have a linear relationship with batch size. For example, increasing the production from 9 to18 liters of a retroviral vector product increases total costs a modest 1.2-fold rather than doubling in total cost. The analysis discussed in this article will help core facilities and investigators plan a cost-effective strategy for gene and cell therapy production. PMID:23360377

Production, concentration and titration of pseudotyped HIV-1-based lentiviral vectors.

PubMed

Kutner, Robert H; Zhang, Xian-Yang; Reiser, Jakob

2009-01-01

Over the past decade, lentiviral vectors have emerged as powerful tools for transgene delivery. The use of lentiviral vectors has become commonplace and applications in the fields of neuroscience, hematology, developmental biology, stem cell biology and transgenesis are rapidly emerging. Also, lentiviral vectors are at present being explored in the context of human clinical trials. Here we describe improved protocols to generate highly concentrated lentiviral vector pseudotypes involving different envelope glycoproteins. In this protocol, vector stocks are prepared by transient transfection using standard cell culture media or serum-free media. Such stocks are then concentrated by ultracentrifugation and/or ion exchange chromatography, or by precipitation using polyethylene glycol 6000, resulting in vector titers of up to 10(10) transducing units per milliliter and above. We also provide reliable real-time PCR protocols to titrate lentiviral vectors based on proviral DNA copies present in genomic DNA extracted from transduced cells or on vector RNA. These production/concentration methods result in high-titer vector preparations that show reduced toxicity compared with lentiviral vectors produced using standard protocols involving ultracentrifugation-based methods. The vector production and titration protocol described here can be completed within 8 d.

Knock-in of Enhanced Green Fluorescent Protein or/and Human Fibroblast Growth Factor 2 Gene into β-Casein Gene Locus in the Porcine Fibroblasts to Produce Therapeutic Protein.

PubMed

Lee, Sang Mi; Kim, Ji Woo; Jeong, Young-Hee; Kim, Se Eun; Kim, Yeong Ji; Moon, Seung Ju; Lee, Ji-Hye; Kim, Keun-Jung; Kim, Min-Kyu; Kang, Man-Jong

2014-11-01

Transgenic animals have become important tools for the production of therapeutic proteins in the domestic animal. Production efficiencies of transgenic animals by conventional methods as microinjection and retrovirus vector methods are low, and the foreign gene expression levels are also low because of their random integration in the host genome. In this study, we investigated the homologous recombination on the porcine β-casein gene locus using a knock-in vector for the β-casein gene locus. We developed the knock-in vector on the porcine β-casein gene locus and isolated knock-in fibroblast for nuclear transfer. The knock-in vector consisted of the neomycin resistance gene (neo) as a positive selectable marker gene, diphtheria toxin-A gene as negative selection marker, and 5' arm and 3' arm from the porcine β-casein gene. The secretion of enhanced green fluorescent protein (EGFP) was more easily detected in the cell culture media than it was by western blot analysis of cell extract of the HC11 mouse mammary epithelial cells transfected with EGFP knock-in vector. These results indicated that a knock-in system using β-casein gene induced high expression of transgene by the gene regulatory sequence of endogenous β-casein gene. These fibroblasts may be used to produce transgenic pigs for the production of therapeutic proteins via the mammary glands.

Knock-in of Enhanced Green Fluorescent Protein or/and Human Fibroblast Growth Factor 2 Gene into β-Casein Gene Locus in the Porcine Fibroblasts to Produce Therapeutic Protein

PubMed Central

Lee, Sang Mi; Kim, Ji Woo; Jeong, Young-Hee; Kim, Se Eun; Kim, Yeong Ji; Moon, Seung Ju; Lee, Ji-Hye; Kim, Keun-Jung; Kim, Min-Kyu; Kang, Man-Jong

2014-01-01

Transgenic animals have become important tools for the production of therapeutic proteins in the domestic animal. Production efficiencies of transgenic animals by conventional methods as microinjection and retrovirus vector methods are low, and the foreign gene expression levels are also low because of their random integration in the host genome. In this study, we investigated the homologous recombination on the porcine β-casein gene locus using a knock-in vector for the β-casein gene locus. We developed the knock-in vector on the porcine β-casein gene locus and isolated knock-in fibroblast for nuclear transfer. The knock-in vector consisted of the neomycin resistance gene (neo) as a positive selectable marker gene, diphtheria toxin-A gene as negative selection marker, and 5′ arm and 3′ arm from the porcine β-casein gene. The secretion of enhanced green fluorescent protein (EGFP) was more easily detected in the cell culture media than it was by western blot analysis of cell extract of the HC11 mouse mammary epithelial cells transfected with EGFP knock-in vector. These results indicated that a knock-in system using β-casein gene induced high expression of transgene by the gene regulatory sequence of endogenous β-casein gene. These fibroblasts may be used to produce transgenic pigs for the production of therapeutic proteins via the mammary glands. PMID:25358326

Construction and production of oncotropic vectors, derived from MVM(p), that share reduced sequence homology with helper plasmids.

PubMed

Clément, Nathalie; Velu, Thierry; Brandenburger, Annick

2002-09-01

The production of currently available vectors derived from autonomous parvoviruses requires the expression of capsid proteins in trans, from helper sequences. Cotransfection of a helper plasmid always generates significant amounts of replication-competent virus (RCV) that can be reduced by the integration of helper sequences into a packaging cell line. Although stocks of minute virus of mice (MVM)-based vectors with no detectable RCV could be produced by transfection into packaging cells; the latter appear after one or two rounds of replication, precluding further amplification of the vector stock. Indeed, once RCVs become detectable, they are efficiently amplified and rapidly take over the culture. Theoretically RCV-free vector stocks could be produced if all homology between vector and helper DNA is eliminated, thus preventing homologous recombination. We constructed new vectors based on the structure of spontaneously occurring defective particles of MVM. Based on published observations related to the size of vectors and the sequence of the viral origin of replication, these vectors were modified by the insertion of foreign DNA sequences downstream of the transgene and by the introduction of a consensus NS-1 nick site near the origin of replication to optimize their production. In one of the vectors the inserted fragment of mouse genomic DNA had a synergistic effect with the modified origin of replication in increasing vector production.

Applications of lentiviral vectors in molecular imaging.

PubMed

Chatterjee, Sushmita; De, Abhijit

2014-06-01

Molecular imaging provides the ability of simultaneous visual and quantitative estimation of long term gene expression directly from living organisms. To reveal the kinetics of gene expression by imaging method, often sustained expression of the transgene is required. Lentiviral vectors have been extensively used over last fifteen years for delivery of a transgene in a wide variety of cell types. Lentiviral vectors have the well known advantages such as sustained transgene delivery through stable integration into the host genome, the capability of infecting non-dividing and dividing cells, broad tissue tropism, a reasonably large carrying capacity for delivering therapeutic and reporter gene combinations. Additionally, they do not express viral proteins during transduction, have a potentially safe integration site profile, and a relatively easy system for vector manipulation and infective viral particle production. As a result, lentiviral vector mediated therapeutic and imaging reporter gene delivery to various target organs holds promise for the future treatment. In this review, we have conducted a brief survey of important lentiviral vector developments in diverse biomedical fields including reproductive biology.

Production of Recombinant Adeno-associated Virus Vectors Using Suspension HEK293 Cells and Continuous Harvest of Vector From the Culture Media for GMP FIX and FLT1 Clinical Vector

PubMed Central

Grieger, Joshua C; Soltys, Stephen M; Samulski, Richard Jude

2016-01-01

Adeno-associated virus (AAV) has shown great promise as a gene therapy vector in multiple aspects of preclinical and clinical applications. Many developments including new serotypes as well as self-complementary vectors are now entering the clinic. With these ongoing vector developments, continued effort has been focused on scalable manufacturing processes that can efficiently generate high-titer, highly pure, and potent quantities of rAAV vectors. Utilizing the relatively simple and efficient transfection system of HEK293 cells as a starting point, we have successfully adapted an adherent HEK293 cell line from a qualified clinical master cell bank to grow in animal component-free suspension conditions in shaker flasks and WAVE bioreactors that allows for rapid and scalable rAAV production. Using the triple transfection method, the suspension HEK293 cell line generates greater than 1 × 105 vector genome containing particles (vg)/cell or greater than 1 × 1014 vg/l of cell culture when harvested 48 hours post-transfection. To achieve these yields, a number of variables were optimized such as selection of a compatible serum-free suspension media that supports both growth and transfection, selection of a transfection reagent, transfection conditions and cell density. A universal purification strategy, based on ion exchange chromatography methods, was also developed that results in high-purity vector preps of AAV serotypes 1–6, 8, 9 and various chimeric capsids tested. This user-friendly process can be completed within 1 week, results in high full to empty particle ratios (>90% full particles), provides postpurification yields (>1 × 1013 vg/l) and purity suitable for clinical applications and is universal with respect to all serotypes and chimeric particles. To date, this scalable manufacturing technology has been utilized to manufacture GMP phase 1 clinical AAV vectors for retinal neovascularization (AAV2), Hemophilia B (scAAV8), giant axonal neuropathy (scAAV9), and retinitis pigmentosa (AAV2), which have been administered into patients. In addition, we report a minimum of a fivefold increase in overall vector production by implementing a perfusion method that entails harvesting rAAV from the culture media at numerous time-points post-transfection. PMID:26437810

The tunable pReX expression vector enables optimizing the T7-based production of membrane and secretory proteins in E. coli.

PubMed

Kuipers, Grietje; Karyolaimos, Alexandros; Zhang, Zhe; Ismail, Nurzian; Trinco, Gianluca; Vikström, David; Slotboom, Dirk Jan; de Gier, Jan-Willem

2017-12-16

To optimize the production of membrane and secretory proteins in Escherichia coli, it is critical to harmonize the expression rates of the genes encoding these proteins with the capacity of their biogenesis machineries. Therefore, we engineered the Lemo21(DE3) strain, which is derived from the T7 RNA polymerase-based BL21(DE3) protein production strain. In Lemo21(DE3), the T7 RNA polymerase activity can be modulated by the controlled co-production of its natural inhibitor T7 lysozyme. This setup enables to precisely tune target gene expression rates in Lemo21(DE3). The t7lys gene is expressed from the pLemo plasmid using the titratable rhamnose promoter. A disadvantage of the Lemo21(DE3) setup is that the system is based on two plasmids, a T7 expression vector and pLemo. The aim of this study was to simplify the Lemo21(DE3) setup by incorporating the key elements of pLemo in a standard T7-based expression vector. By incorporating the gene encoding the T7 lysozyme under control of the rhamnose promoter in a standard T7-based expression vector, pReX was created (ReX stands for Regulated gene eXpression). For two model membrane proteins and a model secretory protein we show that the optimized production yields obtained with the pReX expression vector in BL21(DE3) are similar to the ones obtained with Lemo21(DE3) using a standard T7 expression vector. For another secretory protein, a c-type cytochrome, we show that pReX, in contrast to Lemo21(DE3), enables the use of a helper plasmid that is required for the maturation and hence the production of this heme c protein. Here, we created pReX, a T7-based expression vector that contains the gene encoding the T7 lysozyme under control of the rhamnose promoter. pReX enables regulated T7-based target gene expression using only one plasmid. We show that with pReX the production of membrane and secretory proteins can be readily optimized. Importantly, pReX facilitates the use of helper plasmids. Furthermore, the use of pReX is not restricted to BL21(DE3), but it can in principle be used in any T7 RNAP-based strain. Thus, pReX is a versatile alternative to Lemo21(DE3).

Enhanced production of 3-hydroxypropionic acid from glucose via malonyl-CoA pathway by engineered Escherichia coli.

PubMed

Cheng, Zhuan; Jiang, Jiaqi; Wu, Hui; Li, Zhimin; Ye, Qin

2016-01-01

In this study, production of 3-HP via malonyl-CoA was investigated by using metabolically engineered Escherichia coli carrying heterogeneous acetyl-CoA carboxylase (Acc) from Corynebacterium glutamicum and codon-optimized malonyl-CoA reductase (MCR) from Chloroflexus aurantiacus. Three engineered E. coli strains with different host-vector systems were constructed and investigated. The results indicated that the combination of E. coli BL21(DE3) and pET28a was the most efficient host-vector system for 3-HP production, and the highest concentration of 3-HP attained in shake flask cultivation reached 1.80g/L by the strain BE-MDA with induction at 0.25mM IPTG and 25°C, and supplementation of NaHCO3 and biotin. In fed-batch fermentation performed in a 5-L reactor, the concentration of 3-HP achieved 10.08g/L in 36h. Copyright © 2015 Elsevier Ltd. All rights reserved.

A Single-Vector Force Calibration Method Featuring the Modern Design of Experiments

NASA Technical Reports Server (NTRS)

Parker, P. A.; Morton, M.; Draper, N.; Line, W.

2001-01-01

This paper proposes a new concept in force balance calibration. An overview of the state-of-the-art in force balance calibration is provided with emphasis on both the load application system and the experimental design philosophy. Limitations of current systems are detailed in the areas of data quality and productivity. A unique calibration loading system integrated with formal experimental design techniques has been developed and designated as the Single-Vector Balance Calibration System (SVS). This new concept addresses the limitations of current systems. The development of a quadratic and cubic calibration design is presented. Results from experimental testing are compared and contrasted with conventional calibration systems. Analyses of data are provided that demonstrate the feasibility of this concept and provide new insights into balance calibration.

Decomposition of group-velocity-locked-vector-dissipative solitons and formation of the high-order soliton structure by the product of their recombination.

PubMed

Wang, Xuan; Li, Lei; Geng, Ying; Wang, Hanxiao; Su, Lei; Zhao, Luming

2018-02-01

By using a polarization manipulation and projection system, we numerically decomposed the group-velocity-locked-vector-dissipative solitons (GVLVDSs) from a normal dispersion fiber laser and studied the combination of the projections of the phase-modulated components of the GVLVDS through a polarization beam splitter. Pulses with a structure similar to a high-order vector soliton could be obtained, which could be considered as a pseudo-high-order GVLVDS. It is found that, although GVLVDSs are intrinsically different from group-velocity-locked-vector solitons generated in fiber lasers operated in the anomalous dispersion regime, similar characteristics for the generation of pseudo-high-order GVLVDS are obtained. However, pulse chirp plays a significant role on the generation of pseudo-high-order GVLVDS.

In Vivo Stable Transduction of Humanized Liver Tissue in Chimeric Mice via High-Capacity Adenovirus–Lentivirus Hybrid Vector

PubMed Central

Kataoka, Miho; Tateno, Chise; Yoshizato, Katsutoshi; Kawasaki, Yoshiko; Kimura, Takahiro; Faure-Kumar, Emmanuelle; Palmer, Donna J.; Ng, Philip; Okamura, Haruki; Kasahara, Noriyuki

2010-01-01

Abstract We developed hybrid vectors employing high-capacity adenovirus as a first-stage carrier encoding all the components required for in situ production of a second-stage lentivirus, thereby achieving stable transgene expression in secondary target cells. Such vectors have never previously been tested in normal tissues, because of the scarcity of suitable in vivo systems permissive for second-stage lentivirus assembly. Here we employed a novel murine model in which endogenous liver tissue is extensively reconstituted with engrafted human hepatocytes, and successfully achieved stable transduction by the second-stage lentivirus produced in situ from first-stage adenovirus. This represents the first demonstration of the functionality of adenoviral-lentiviral hybrid vectors in a normal parenchymal organ in vivo. PMID:19725756

A reporter system for replication-competent gammaretroviruses: the inGluc-MLV-DERSE assay

PubMed Central

Aloia, Amanda L.; Duffy, Lisa; Pak, Vladimir; Lee, KyeongEun; Sanchez-Martinez, Silvia; Derse, David; Heidecker, Gisela; Cornetta, Kenneth; Rein, Alan

2012-01-01

While novel retroviral vectors for use in gene-therapy products are reducing the potential for formation of replication-competent retrovirus (RCR), it remains crucial to screen products for RCR for both research and clinical purposes. For clinical grade gammaretrovirus-based vectors, RCR screening is achieved by an extended S+L− or marker rescue assay, while standard methods for replication-competent lentivirus detection are still in development. In this report, we describe a rapid and sensitive method for replication-competent gammaretrovirus detection. We used this assay to detect three members of the gammaretrovirus family and compared the sensitivity of our assay with well-established methods for retrovirus detection, including the extended S+L− assay. Results presented here demonstrate that this assay should be useful for gene-therapy product testing. PMID:22402321

Genome-Based Genetic Tool Development for Bacillus methanolicus: Theta- and Rolling Circle-Replicating Plasmids for Inducible Gene Expression and Application to Methanol-Based Cadaverine Production.

PubMed

Irla, Marta; Heggeset, Tonje M B; Nærdal, Ingemar; Paul, Lidia; Haugen, Tone; Le, Simone B; Brautaset, Trygve; Wendisch, Volker F

2016-01-01

Bacillus methanolicus is a thermophilic methylotroph able to overproduce amino acids from methanol, a substrate not used for human or animal nutrition. Based on our previous RNA-seq analysis a mannitol inducible promoter and a putative mannitol activator gene mtlR were identified. The mannitol inducible promoter was applied for controlled gene expression using fluorescent reporter proteins and a flow cytometry analysis, and improved by changing the -35 promoter region and by co-expression of the mtlR regulator gene. For independent complementary gene expression control, the heterologous xylose-inducible system from B. megaterium was employed and a two-plasmid gene expression system was developed. Four different replicons for expression vectors were compared with respect to their copy number and stability. As an application example, methanol-based production of cadaverine was shown to be improved from 11.3 to 17.5 g/L when a heterologous lysine decarboxylase gene cadA was expressed from a theta-replicating rather than a rolling-circle replicating vector. The current work on inducible promoter systems and compatible theta- or rolling circle-replicating vectors is an important extension of the poorly developed B. methanolicus genetic toolbox, valuable for genetic engineering and further exploration of this bacterium.

Genome-Based Genetic Tool Development for Bacillus methanolicus: Theta- and Rolling Circle-Replicating Plasmids for Inducible Gene Expression and Application to Methanol-Based Cadaverine Production

PubMed Central

Irla, Marta; Heggeset, Tonje M. B.; Nærdal, Ingemar; Paul, Lidia; Haugen, Tone; Le, Simone B.; Brautaset, Trygve; Wendisch, Volker F.

2016-01-01

Bacillus methanolicus is a thermophilic methylotroph able to overproduce amino acids from methanol, a substrate not used for human or animal nutrition. Based on our previous RNA-seq analysis a mannitol inducible promoter and a putative mannitol activator gene mtlR were identified. The mannitol inducible promoter was applied for controlled gene expression using fluorescent reporter proteins and a flow cytometry analysis, and improved by changing the -35 promoter region and by co-expression of the mtlR regulator gene. For independent complementary gene expression control, the heterologous xylose-inducible system from B. megaterium was employed and a two-plasmid gene expression system was developed. Four different replicons for expression vectors were compared with respect to their copy number and stability. As an application example, methanol-based production of cadaverine was shown to be improved from 11.3 to 17.5 g/L when a heterologous lysine decarboxylase gene cadA was expressed from a theta-replicating rather than a rolling-circle replicating vector. The current work on inducible promoter systems and compatible theta- or rolling circle-replicating vectors is an important extension of the poorly developed B. methanolicus genetic toolbox, valuable for genetic engineering and further exploration of this bacterium. PMID:27713731

Multigrid Equation Solvers for Large Scale Nonlinear Finite Element Simulations

DTIC Science & Technology

1999-01-01

purpose of the second partitioning phase , on each SMP, is to minimize the communication within the SMP; even if a multi - threaded matrix vector product...8.7 Comparison of model with experimental data for send phase of matrix vector product on ne grid...140 8.4 Matrix vector product phase times : : : : : : : : : : : : : : : : : : : : : : : 145 9.1 Flat and

Vectorial laws of refraction and reflection using the cross product and dot product.

PubMed

Tkaczyk, Eric R

2012-03-01

We demonstrate that published vectorial laws of reflection and refraction of light based solely on the cross product do not, in general, uniquely determine the direction of the reflected and refracted waves without additional information. This is because the cross product does not have a unique inverse operation, which is explained in this Letter in linear algebra terms. However, a vector is in fact uniquely determined if both the cross product (vector product) and dot product (scalar product) with a known vector are specified, which can be written as a single equation with a left-invertible matrix. It is thus possible to amend the vectorial laws of reflection and refraction to incorporate both the cross and dot products for a complete specification with unique solution. This enables highly efficient, unambiguous computation of reflected and refracted wave vectors from the incident wave and surface normal. © 2012 Optical Society of America

Genetic transformation of Fusarium avenaceum by Agrobacterium tumefaciens mediated transformation and the development of a USER-Brick vector construction system

PubMed Central

2014-01-01

Background The plant pathogenic and saprophytic fungus Fusarium avenaceum causes considerable in-field and post-field losses worldwide due to its infections of a wide range of different crops. Despite its significant impact on the profitability of agriculture production and a desire to characterize the infection process at the molecular biological level, no genetic transformation protocol has yet been established for F. avenaceum. In the current study, it is shown that F. avenaceum can be efficiently transformed by Agrobacterium tumefaciens mediated transformation. In addition, an efficient and versatile single step vector construction strategy relying on Uracil Specific Excision Reagent (USER) Fusion cloning, is developed. Results The new vector construction system, termed USER-Brick, is based on a limited number of PCR amplified vector fragments (core USER-Bricks) which are combined with PCR generated fragments from the gene of interest. The system was found to have an assembly efficiency of 97% with up to six DNA fragments, based on the construction of 55 vectors targeting different polyketide synthase (PKS) and PKS associated transcription factor encoding genes in F. avenaceum. Subsequently, the ΔFaPKS3 vector was used for optimizing A. tumefaciens mediated transformation (ATMT) of F. avenaceum with respect to six variables. Acetosyringone concentration, co-culturing time, co-culturing temperature and fungal inoculum were found to significantly impact the transformation frequency. Following optimization, an average of 140 transformants per 106 macroconidia was obtained in experiments aimed at introducing targeted genome modifications. Targeted deletion of FaPKS6 (FA08709.2) in F. avenaceum showed that this gene is essential for biosynthesis of the polyketide/nonribosomal compound fusaristatin A. Conclusion The new USER-Brick system is highly versatile by allowing for the reuse of a common set of building blocks to accommodate seven different types of genome modifications. New USER-Bricks with additional functionality can easily be added to the system by future users. The optimized protocol for ATMT of F. avenaceum represents the first reported targeted genome modification by double homologous recombination of this plant pathogen and will allow for future characterization of this fungus. Functional linkage of FaPKS6 to the production of the mycotoxin fusaristatin A serves as a first testimony to this. PMID:25048842

Method and apparatus for optimized processing of sparse matrices

DOEpatents

Taylor, Valerie E.

1993-01-01

A computer architecture for processing a sparse matrix is disclosed. The apparatus stores a value-row vector corresponding to nonzero values of a sparse matrix. Each of the nonzero values is located at a defined row and column position in the matrix. The value-row vector includes a first vector including nonzero values and delimiting characters indicating a transition from one column to another. The value-row vector also includes a second vector which defines row position values in the matrix corresponding to the nonzero values in the first vector and column position values in the matrix corresponding to the column position of the nonzero values in the first vector. The architecture also includes a circuit for detecting a special character within the value-row vector. Matrix-vector multiplication is executed on the value-row vector. This multiplication is performed by multiplying an index value of the first vector value by a column value from a second matrix to form a matrix-vector product which is added to a previous matrix-vector product.

Improvements and Advances to the Cross-Calibrated Multi-Platform (CCMP) Ocean Vector Wind Analysis (V2.0 release)

NASA Astrophysics Data System (ADS)

Scott, J. P.; Wentz, F. J.; Hoffman, R. N.; Atlas, R. M.

2016-02-01

Ocean vector wind is a valuable climate data record (CDR) useful in observing and monitoring changes in climate and air-sea interactions. Ocean surface wind stress influences such processes as heat, moisture, and momentum fluxes between the atmosphere and ocean, driving ocean currents and forcing ocean circulation. The Cross-Calibrated Multi-Platform (CCMP) ocean vector wind analysis is a quarter-degree, six-hourly global ocean wind analysis product created using the variational analysis method (VAM) [Atlas et al., 1996; Hoffman et al., 2003]. The CCMP V1.1 wind product is a highly-esteemed, widely-used data set containing the longest gap-free record of satellite-based ocean vector wind data (July 1987 to June 2012). CCMP V1.1 was considered a "first-look" data set that used the most-timely, albeit preliminary, releases of satellite, in situ, and modeled ECMWF-Operational wind background fields. The authors have been working with the original producers of CCMP V1.1 to create an updated, improved, and consistently-reprocessed CCMP V2.0 ocean vector wind analysis data set. With Remote Sensing Systems (RSS) having recently updated all passive microwave satellite instrument calibrations and retrievals to the RSS Version-7 RTM standard, the reprocessing of the CCMP data set into a higher-quality CDR using inter-calibrated satellite inputs became feasible. In addition to the use of SSM/I, SSMIS, TRMM TMI, QuikSCAT, AMSRE, and WindSat instruments, AMSR2, GMI, and ASCAT have been also included in the CCMP V2.0 data set release, which has now been extended to the beginning of 2015. Additionally, the background field has been updated to use six-hourly, quarter-degree ERA-Interim wind vector inputs, and the quality-checks on the in situ data have been carefully reviewed and improved. The goal of the release of the CCMP V2.0 ocean wind vector analysis product is to serve as a merged ocean wind vector data set for climate studies. Diligent effort has been made by the authors to minimize systematic and spurious sources of error. The authors will present a complete discussion of upgrades made to the CCMP V2.0 data set, as well as present validation work that has been completed on the CCMP V2.0 wind analysis product.

The linear combination of vectors implies the existence of the cross and dot products

NASA Astrophysics Data System (ADS)

Pujol, Jose

2018-07-01

Given two vectors u and v, their cross product u × v is a vector perpendicular to u and v. The motivation for this property, however, is never addressed. Here we show that the existence of the cross and dot products and the perpendicularity property follow from the concept of linear combination, which does not involve products of vectors. For our proof we consider the plane generated by a linear combination of uand v. When looking for the coefficients in the linear combination required to reach a desired point on the plane, the solution involves the existence of a normal vector n = u × v. Our results have a bearing on the history of vector analysis, as a product similar to the cross product but without the perpendicularity requirement existed at the same time. These competing products originate in the work of two major nineteen-century mathematicians, W. Hamilton, and H. Grassmann. These historical aspects are discussed in some detail here. We also address certain aspects of the teaching of u × v to undergraduate students, which is known to carry some difficulties. This includes the algebraic and geometric denitions of u × v, the rule for the direction of u × v, and the pseudovectorial nature of u × v.

A Modular Toolset for Recombination Transgenesis and Neurogenetic Analysis of Drosophila

PubMed Central

Wang, Ji-Wu; Beck, Erin S.; McCabe, Brian D.

2012-01-01

Transgenic Drosophila have contributed extensively to our understanding of nervous system development, physiology and behavior in addition to being valuable models of human neurological disease. Here, we have generated a novel series of modular transgenic vectors designed to optimize and accelerate the production and analysis of transgenes in Drosophila. We constructed a novel vector backbone, pBID, that allows both phiC31 targeted transgene integration and incorporates insulator sequences to ensure specific and uniform transgene expression. Upon this framework, we have built a series of constructs that are either backwards compatible with existing restriction enzyme based vectors or utilize Gateway recombination technology for high-throughput cloning. These vectors allow for endogenous promoter or Gal4 targeted expression of transgenic proteins with or without fluorescent protein or epitope tags. In addition, we have generated constructs that facilitate transgenic splice isoform specific RNA inhibition of gene expression. We demonstrate the utility of these constructs to analyze proteins involved in nervous system development, physiology and neurodegenerative disease. We expect that these reagents will facilitate the proficiency and sophistication of Drosophila genetic analysis in both the nervous system and other tissues. PMID:22848718

An improved Red/ET recombineering system and mouse ES cells culture conditions for the generation of targeted mutant mice.

PubMed

Kumagai, Katsuyoshi; Takanashi, Masakatsu; Ohno, Shin-Ichiro; Kuroda, Masahiko; Sudo, Katsuko

2017-05-03

Targeted mutant mice generated on a C57BL/6 background are powerful tools for analysis of the biological functions of genes, and gene targeting technologies using mouse embryonic stem (ES) cells have been used to generate such mice. Recently, a bacterial artificial chromosome (BAC) recombineering system was established for the construction of targeting vectors. However, gene retrieval from BACs for the generation of gene targeting vectors using this system remains difficult. Even when construction of a gene targeting vector is successful, the efficiency of production of targeted mutant mice from ES cells derived from C57BL/6 mice are poor. Therefore, in this study, we first improved the strategy for the retrieval of genes from BACs and their transfer into a DT-A plasmid, for the generation of gene targeting vectors using the BAC recombineering system. Then, we attempted to generate targeted mutant mice from ES cell lines derived from C57BL/6 mice, by culturing in serum-free medium. In conclusion, we established an improved strategy for the efficient generation of targeted mutant mice on a C57BL/6 background, which are useful for the in vivo analysis of gene functions and regulation.

Expression of the core antigen gene of hepatitis B virus (HBV) in Acetobacter methanolicus using broad-host-range vectors.

PubMed

Schröder, R; Maassen, A; Lippoldt, A; Börner, T; von Baehr, R; Dobrowolski, P

1991-08-01

Using the broad-host-range promoter probe vector pRS201 for cloning of phage Acm1 promoters, we established a convenient vector system for expression of heterologous genes in different Gram-negative bacteria. The usefulness of this system was demonstrated by expression of the HBV core gene in Acetobacter methanolicus. Plasmids carrying the HBV core gene downstream of different Acm1-phage promoters were transferred to A. methanolicus, a new potential host for recombinant DNA expression. Using enzyme immunoassay and immunoblot techniques, the amount and composition of core antigen produced in A. methanolicus were compared with that derived from Escherichia coli. The expression of immunoreactive core antigen in A. methanolicus exceeds by sevenfold that in E. coli using an expression system with tandemly arranged promoters. Morphological observations by electron microscopy show that the HBV core gene products isolated from both hosts are assembled into regular spherical particles with a diameter of about 28 nm that are comparable to original viral nucleocapsids.

A Simple And Rapid Minicircle DNA Vector Manufacturing System

PubMed Central

Kay, Mark A; He, Cheng-Yi; Chen, Zhi-Ying

2010-01-01

Minicircle DNA vectors consisting of a circular expression cassette devoid of the bacterial plasmid DNA backbone provides several advantages including sustained transgene expression in quiescent cells/tissues. Their use has been limited by labor-intensive production. We report on a strategy for making multiple genetic modifications in E.coli to construct a producer strain that stably expresses a set of inducible minicircle-assembly enzymes, the øC31-integrase and I-SceI homing-endonuclease. This bacterial strain is capable of producing highly purified minicircle yields in the same time frame as routine plasmid DNA. It is now feasible for minicircle DNA vectors to replace routine plasmids in mammalian transgene expression studies. PMID:21102455

Arbitration in crossbar interconnect for low latency

DOEpatents

Ohmacht, Martin; Sugavanam, Krishnan

2013-02-05

A system and method and computer program product for reducing the latency of signals communicated through a crossbar switch, the method including using at slave arbitration logic devices associated with Slave devices for which access is requested from one or more Master devices, two or more priority vector signals cycled among their use every clock cycle for selecting one of the requesting Master devices and updates the respective priority vector signal used every clock cycle. Similarly, each Master for which access is requested from one or more Slave devices, can have two or more priority vectors and can cycle among their use every clock cycle to further reduce latency and increase throughput performance via the crossbar.

A diagram for evaluating multiple aspects of model performance in simulating vector fields

NASA Astrophysics Data System (ADS)

Xu, Zhongfeng; Hou, Zhaolu; Han, Ying; Guo, Weidong

2016-12-01

Vector quantities, e.g., vector winds, play an extremely important role in climate systems. The energy and water exchanges between different regions are strongly dominated by wind, which in turn shapes the regional climate. Thus, how well climate models can simulate vector fields directly affects model performance in reproducing the nature of a regional climate. This paper devises a new diagram, termed the vector field evaluation (VFE) diagram, which is a generalized Taylor diagram and able to provide a concise evaluation of model performance in simulating vector fields. The diagram can measure how well two vector fields match each other in terms of three statistical variables, i.e., the vector similarity coefficient, root mean square length (RMSL), and root mean square vector difference (RMSVD). Similar to the Taylor diagram, the VFE diagram is especially useful for evaluating climate models. The pattern similarity of two vector fields is measured by a vector similarity coefficient (VSC) that is defined by the arithmetic mean of the inner product of normalized vector pairs. Examples are provided, showing that VSC can identify how close one vector field resembles another. Note that VSC can only describe the pattern similarity, and it does not reflect the systematic difference in the mean vector length between two vector fields. To measure the vector length, RMSL is included in the diagram. The third variable, RMSVD, is used to identify the magnitude of the overall difference between two vector fields. Examples show that the VFE diagram can clearly illustrate the extent to which the overall RMSVD is attributed to the systematic difference in RMSL and how much is due to the poor pattern similarity.

Ecological, biological and social dimensions of dengue vector breeding in five urban settings of Latin America: a multi-country study.

PubMed

Quintero, Juliana; Brochero, Helena; Manrique-Saide, Pablo; Barrera-Pérez, Mario; Basso, César; Romero, Sonnia; Caprara, Andrea; De Lima Cunha, Jane Cris; Beltrán-Ayala, Efraín; Mitchell-Foster, Kendra; Kroeger, Axel; Sommerfeld, Johannnes; Petzold, Max

2014-01-21

Dengue is an increasingly important public health problem in most Latin American countries and more cost-effective ways of reducing dengue vector densities to prevent transmission are in demand by vector control programs. This multi-centre study attempted to identify key factors associated with vector breeding and development as a basis for improving targeted intervention strategies. In each of 5 participant cities in Mexico, Colombia, Ecuador, Brazil and Uruguay, 20 clusters were randomly selected by grid sampling to incorporate 100 contiguous households, non-residential private buildings (businesses) and public spaces. Standardized household surveys, cluster background surveys and entomological surveys specifically targeted to obtain pupal indices for Aedes aegypti, were conducted in the dry and wet seasons. The study clusters included mainly urban low-middle class populations with satisfactory infrastructure and -except for Uruguay- favourable climatic conditions for dengue vector development. Household knowledge about dengue and "dengue mosquitoes" was widespread, mainly through mass media, but there was less awareness around interventions to reduce vector densities. Vector production (measured through pupal indices) was favoured when water containers were outdoor, uncovered, unused (even in Colombia and Ecuador where the large tanks used for household water storage and washing were predominantly productive) and -particularly during the dry season- rainwater filled. Larval infestation did not reflect productive container types. All productive container types, including those important in the dry season, were identified by pupal surveys executed during the rainy season. A number of findings are relevant for improving vector control: 1) there is a need for complementing larval surveys with occasional pupal surveys (to be conducted during the wet season) for identifying and subsequently targeting productive container types; 2) the need to raise public awareness about useful and effective interventions in productive container types specific to their area; and 3) the motivation for control services that-according to this and similar studies in Asia- dedicated, targeted vector management can make a difference in terms of reducing vector abundance.

Development of an optimized tetracycline-inducible expression system to increase the accumulation of interleukin-10 in tobacco BY-2 suspension cells.

PubMed

Bortesi, Luisa; Rademacher, Thomas; Schiermeyer, Andreas; Schuster, Flora; Pezzotti, Mario; Schillberg, Stefan

2012-07-11

Plant cell suspension cultures can be used for the production of valuable pharmaceutical and industrial proteins. When the recombinant protein is secreted into the culture medium, restricting expression to a defined growth phase can improve both the quality and quantity of the recovered product by minimizing proteolytic activity. Temporal restriction is also useful for recombinant proteins whose constitutive expression affects cell growth and viability, such as viral interleukin-10 (vIL-10). We have developed a novel, tetracycline-inducible system suitable for tobacco BY-2 suspension cells which increases the yields of vIL-10. The new system is based on a binary vector that is easier to handle than conventional vectors, contains an enhanced inducible promoter and 5'-UTR to improve yields, and incorporates a constitutively-expressed visible marker gene to allow the rapid and straightforward selection of the most promising transformed clones. Stable transformation of BY-2 cells with this vector, without extensive optimization of the induction conditions, led to a 3.5 fold increase in vIL-10 levels compared to constitutive expression in the same host. We have developed an effective and straightforward molecular farming platform technology that improves both the quality and the quantity of recombinant proteins produced in plant cells, particularly those whose constitutive expression has a negative impact on plant growth and development. Although we tested the platform using vIL-10 produced in BY-2 cells, it can be applied to other host/product combinations and is also useful for basic research requiring strictly controlled transgene expression.

Genetic engineering of cell lines using lentiviral vectors to achieve antibody secretion following encapsulated implantation.

PubMed

Lathuilière, Aurélien; Bohrmann, Bernd; Kopetzki, Erhard; Schweitzer, Christoph; Jacobsen, Helmut; Moniatte, Marc; Aebischer, Patrick; Schneider, Bernard L

2014-01-01

The controlled delivery of antibodies by immunoisolated bioimplants containing genetically engineered cells is an attractive and safe approach for chronic treatments. To reach therapeutic antibody levels there is a need to generate renewable cell lines, which can long-term survive in macroencapsulation devices while maintaining high antibody specific productivity. Here we have developed a dual lentiviral vector strategy for the genetic engineering of cell lines compatible with macroencapsulation, using separate vectors encoding IgG light and heavy chains. We show that IgG expression level can be maximized as a function of vector dose and transgene ratio. This approach allows for the generation of stable populations of IgG-expressing C2C12 mouse myoblasts, and for the subsequent isolation of clones stably secreting high IgG levels. Moreover, we demonstrate that cell transduction using this lentiviral system leads to the production of a functional glycosylated antibody by myogenic cells. Subsequent implantation of antibody-secreting cells in a high-capacity macroencapsulation device enables continuous delivery of recombinant antibodies in the mouse subcutaneous tissue, leading to substantial levels of therapeutic IgG detectable in the plasma.

Rapid production of functionalized recombinant proteins: marrying ligation independent cloning and in vitro protein ligation.

PubMed

Kushnir, Susanna; Marsac, Yoann; Breitling, Reinhard; Granovsky, Igor; Brok-Volchanskaya, Vera; Goody, Roger S; Becker, Christian F W; Alexandrov, Kirill

2006-01-01

Functional genomics and proteomics have been very active fields since the sequencing of several genomes was completed. To assign a physiological role to the newly discovered coding genes with unknown function, new generic methods for protein production, purification, and targeted functionalization are needed. This work presents a new vector, pCYSLIC, that allows rapid generation of Escherichia coli expression constructs via ligation-independent cloning (LIC). The vector is designed to facilitate protein purification by either Ni-NTA or GSH affinity chromatography. Subsequent proteolytic removal of affinity tags liberates an N-terminal cysteine residue that is then used for covalent modification of the target protein with different biophysical probes via protein ligation. The described system has been tested on 36 mammalian Rab GTPases, and it was demonstrated that recombinant GTPases produced with pCYSLIC could be efficiently modified with fluorescein or biotin in vitro. Finally, LIC was compared with the recently developed In-Fusion cloning method, and it was demonstrated that In-Fusion provides superior flexibility in choice of expression vector. By the application of In-Fusion cloning Cys-Rab6A GTPase with an N-terminal cysteine residue was generated employing unmodified pET30a vector and TVMV protease.

Syngeneic AAV pseudo-vectors potentiates full vector transduction

USDA-ARS?s Scientific Manuscript database

An excessive amount of empty capsids are generated during regular AAV vector production process. These pseudo-vectors often remain in final vectors used for animal studies or clinical trials. The potential effects of these pseudo-vectors on AAV transduction have been a major concern. In the current ...

Phytophthora community structure analyses in Oregon nurseries inform systems approaches to disease management

Treesearch

J.L. Parke; B.J. Knaus; V.J. Fieland; C. Lewis; N.J. Grünwald

2014-01-01

Nursery plants are important vectors for plant pathogens. Understanding what pathogens occur in nurseries in different production stages can be useful to the development of integrated systems approaches. Four horticultural nurseries in Oregon were sampled every 2 months for 4 years to determine the identity and community structure of Phytophthora...

A Novel Immunogenic Spore Coat-Associated Protein in Bacillus Anthracis: Characterization via Proteomics Approaches and a Vector-Based Vaccine System

PubMed Central

Liu, Yu-Tsueng; Lin, Shwu-Bin; Huang, Cheng-Po; Huang, Chun-Ming

2007-01-01

New generation anthrax vaccines have been actively explored with the aim of enhancing efficacies and decreasing undesirable side effects that could be caused by licensed vaccines. Targeting novel antigens and/or eliminating the requirements for multiple needle injections and adjuvants are major objectives in the development of new anthrax vaccines. Using proteomics approaches, we identified a spore coat-associated protein (SCAP) in Bacillus anthracis. An E. coli vector-based vaccine system was used to determine the immunogenicity of SCAP. Mice generated detectable SCAP antibodies three weeks after intranasal immunization with an intact particle of ultraviolet (UV)-irradiated E. coli vector overproducing SCAP. The production of SCAP antibodies was detected via western blotting and SCAP-spotted antigen-arrays. The adjuvant effect of a UV-irradiated E. coli vector eliminates the necessity of boosting and the use of other immunomodulators which will foster the screening and manufacturing of new generation anthrax vaccines. More importantly, the immunogenic SCAP may potentially be a new candidate for the development of anthrax vaccines. PMID:18029197

Cationic nanoemulsions as nucleic acids delivery systems.

PubMed

Teixeira, Helder Ferreira; Bruxel, Fernanda; Fraga, Michelle; Schuh, Roselena Silvestri; Zorzi, Giovanni Konat; Matte, Ursula; Fattal, Elias

2017-12-20

Since the first clinical studies, knowledge in the field of gene therapy has advanced significantly, and these advances led to the development and subsequent approval of the first gene medicines. Although viral vectors-based products offer efficient gene expression, problems related to their safety and immune response have limited their clinical use. Thus, design and optimization of nonviral vectors is presented as a promising strategy in this scenario. Nonviral systems are nanotechnology-based products composed of polymers or lipids, which are usually biodegradable and biocompatible. Cationic liposomes are the most studied nonviral carriers and knowledge about these systems has greatly evolved, especially in understanding the role of phospholipids and cationic lipids. However, the search for efficient delivery systems aiming at gene therapy remains a challenge. In this context, cationic nanoemulsions have proved to be an interesting approach, as their ability to protect and efficiently deliver nucleic acids for diverse therapeutic applications has been demonstrated. This review focused on cationic nanoemulsions designed for gene therapy, providing an overview on their composition, physicochemical properties, and their efficacy on biological response in vitro and in vivo. Copyright © 2017 Elsevier B.V. All rights reserved.

Truncated Conjugate Gradient: An Optimal Strategy for the Analytical Evaluation of the Many-Body Polarization Energy and Forces in Molecular Simulations.

PubMed

Aviat, Félix; Levitt, Antoine; Stamm, Benjamin; Maday, Yvon; Ren, Pengyu; Ponder, Jay W; Lagardère, Louis; Piquemal, Jean-Philip

2017-01-10

We introduce a new class of methods, denoted as Truncated Conjugate Gradient(TCG), to solve the many-body polarization energy and its associated forces in molecular simulations (i.e. molecular dynamics (MD) and Monte Carlo). The method consists in a fixed number of Conjugate Gradient (CG) iterations. TCG approaches provide a scalable solution to the polarization problem at a user-chosen cost and a corresponding optimal accuracy. The optimality of the CG-method guarantees that the number of the required matrix-vector products are reduced to a minimum compared to other iterative methods. This family of methods is non-empirical, fully adaptive, and provides analytical gradients, avoiding therefore any energy drift in MD as compared to popular iterative solvers. Besides speed, one great advantage of this class of approximate methods is that their accuracy is systematically improvable. Indeed, as the CG-method is a Krylov subspace method, the associated error is monotonically reduced at each iteration. On top of that, two improvements can be proposed at virtually no cost: (i) the use of preconditioners can be employed, which leads to the Truncated Preconditioned Conjugate Gradient (TPCG); (ii) since the residual of the final step of the CG-method is available, one additional Picard fixed point iteration ("peek"), equivalent to one step of Jacobi Over Relaxation (JOR) with relaxation parameter ω, can be made at almost no cost. This method is denoted by TCG-n(ω). Black-box adaptive methods to find good choices of ω are provided and discussed. Results show that TPCG-3(ω) is converged to high accuracy (a few kcal/mol) for various types of systems including proteins and highly charged systems at the fixed cost of four matrix-vector products: three CG iterations plus the initial CG descent direction. Alternatively, T(P)CG-2(ω) provides robust results at a reduced cost (three matrix-vector products) and offers new perspectives for long polarizable MD as a production algorithm. The T(P)CG-1(ω) level provides less accurate solutions for inhomogeneous systems, but its applicability to well-conditioned problems such as water is remarkable, with only two matrix-vector product evaluations.

Truncated Conjugate Gradient: An Optimal Strategy for the Analytical Evaluation of the Many-Body Polarization Energy and Forces in Molecular Simulations

PubMed Central

2016-01-01

We introduce a new class of methods, denoted as Truncated Conjugate Gradient(TCG), to solve the many-body polarization energy and its associated forces in molecular simulations (i.e. molecular dynamics (MD) and Monte Carlo). The method consists in a fixed number of Conjugate Gradient (CG) iterations. TCG approaches provide a scalable solution to the polarization problem at a user-chosen cost and a corresponding optimal accuracy. The optimality of the CG-method guarantees that the number of the required matrix-vector products are reduced to a minimum compared to other iterative methods. This family of methods is non-empirical, fully adaptive, and provides analytical gradients, avoiding therefore any energy drift in MD as compared to popular iterative solvers. Besides speed, one great advantage of this class of approximate methods is that their accuracy is systematically improvable. Indeed, as the CG-method is a Krylov subspace method, the associated error is monotonically reduced at each iteration. On top of that, two improvements can be proposed at virtually no cost: (i) the use of preconditioners can be employed, which leads to the Truncated Preconditioned Conjugate Gradient (TPCG); (ii) since the residual of the final step of the CG-method is available, one additional Picard fixed point iteration (“peek”), equivalent to one step of Jacobi Over Relaxation (JOR) with relaxation parameter ω, can be made at almost no cost. This method is denoted by TCG-n(ω). Black-box adaptive methods to find good choices of ω are provided and discussed. Results show that TPCG-3(ω) is converged to high accuracy (a few kcal/mol) for various types of systems including proteins and highly charged systems at the fixed cost of four matrix-vector products: three CG iterations plus the initial CG descent direction. Alternatively, T(P)CG-2(ω) provides robust results at a reduced cost (three matrix-vector products) and offers new perspectives for long polarizable MD as a production algorithm. The T(P)CG-1(ω) level provides less accurate solutions for inhomogeneous systems, but its applicability to well-conditioned problems such as water is remarkable, with only two matrix-vector product evaluations. PMID:28068773

Design and construction of targeted AAVP vectors for mammalian cell transduction.

PubMed

Hajitou, Amin; Rangel, Roberto; Trepel, Martin; Soghomonyan, Suren; Gelovani, Juri G; Alauddin, Mian M; Pasqualini, Renata; Arap, Wadih

2007-01-01

Bacteriophage (phage) evolved as bacterial viruses, but can be adapted to transduce mammalian cells through ligand-directed targeting to a specific receptor. We have recently reported a new generation of hybrid prokaryotic-eukaryotic vectors, which are chimeras of genetic cis-elements of recombinant adeno-associated virus and phage (termed AAVP). This protocol describes the design and construction of ligand-directed AAVP vectors, production of AAVP particles and the methodology to transduce mammalian cells in vitro and to target tissues in vivo after systemic administration. Targeted AAVP particles are made in a two-step process. First, a ligand peptide of choice is displayed on the coat protein to generate a targeted backbone phage vector. Then, a recombinant AAV carrying a mammalian transgene cassette is inserted into an intergenomic region. High-titer suspensions (approximately 10(10)-10(11) transducing units per microl) can be produced within 3 days after vector construction. Transgene expression by targeted AAVP usually reaches maximum levels within 1 week.

Music Signal Processing Using Vector Product Neural Networks

NASA Astrophysics Data System (ADS)

Fan, Z. C.; Chan, T. S.; Yang, Y. H.; Jang, J. S. R.

2017-05-01

We propose a novel neural network model for music signal processing using vector product neurons and dimensionality transformations. Here, the inputs are first mapped from real values into three-dimensional vectors then fed into a three-dimensional vector product neural network where the inputs, outputs, and weights are all three-dimensional values. Next, the final outputs are mapped back to the reals. Two methods for dimensionality transformation are proposed, one via context windows and the other via spectral coloring. Experimental results on the iKala dataset for blind singing voice separation confirm the efficacy of our model.

A review of trends in the distribution of vector-borne diseases: is international trade contributing to their spread?

PubMed

de La Rocque, S; Balenghien, T; Halos, L; Dietze, K; Claes, F; Ferrari, G; Guberti, V; Slingenbergh, J

2011-04-01

It is difficult to determine the part that international trade has played in the expansion of vector-borne diseases, because of the multitude of factors that affect the transformation of habitats and the interfaces between vectors and hosts. The introduction of pathogens through trade in live animals or products of animal origin, as well as the arrival of arthropod vectors, is probably quite frequent but the establishment of an efficient transmission system that develops into a disease outbreak remains the exception. In this paper, based on well-documented examples, the authors review the ecological and epidemiological characteristics of vector-borne diseases that may have been affected in their spread and change of distribution by international trade. In addition, they provide a detailed analysis of the risks associated with specific trade routes and recent expansions of vector populations. Finally, the authors highlight the importance, as well as the challenges, of preventive surveillance and regulation. The need for improved monitoring of vector populations and a readiness to face unpredictable epidemiological events are also emphasised, since this will require rapid reaction, not least in the regulatory context.

Use of the piggyBac transposon to create stable packaging cell lines for the production of clinical-grade self-inactivating γ-retroviral vectors.

PubMed

Feldman, Steven A; Xu, Hui; Black, Mary A; Park, Tristen S; Robbins, Paul F; Kochenderfer, James N; Morgan, Richard A; Rosenberg, Steven A

2014-08-01

Efforts to improve the biosafety of γ-retroviral-mediated gene therapy have resulted in a shift toward the use of self-inactivating (SIN) γ-retroviral vectors. However, scale-up and manufacturing of such vectors requires significant optimization of transient transfection-based processes or development of novel platforms for the generation of stable producer cell clones. To that end, we describe the use of the piggybac transposon to generate stable producer cell clones for the production of SIN γ-retroviral vectors. The piggybac transposon is a universal tool allowing for the stable integration of SIN γ-retroviral constructs into murine (PG13) and human 293-based Phoenix (GALV and RD114, respectively) packaging cell lines without reverse transcription. Following transposition, a high-titer clone is selected for manufacture of a master cell bank and subsequent γ-retroviral vector supernatant production. Packaging cell clones created using the piggybac transposon have comparable titers to non-SIN vectors generated via conventional methods. We describe herein the use of the piggybac transposon for the production of stable packaging cell clones for the manufacture of clinical-grade SIN γ-retroviral vectors for ex vivo gene therapy clinical trials.

Adenoviral production of interleukin-2 at the tumor site removes the need for systemic postconditioning in adoptive cell therapy.

PubMed

Santos, Joao Manuel; Havunen, Riikka; Siurala, Mikko; Cervera-Carrascon, Víctor; Tähtinen, Siri; Sorsa, Suvi; Anttila, Marjukka; Karell, Pauliina; Kanerva, Anna; Hemminki, Akseli

2017-10-01

Systemic high dose interleukin-2 (IL-2) postconditioning has long been utilized in boosting the efficacy of T cells in adoptive cell therapy (ACT) of solid tumors. The resulting severe off-target toxicity of these regimens renders local production at the tumor an attractive concept with possible safety gains. We evaluated the efficacy and safety of intratumorally administered IL-2-coding adenoviruses in combination with tumor-infiltrating lymphocyte therapy in syngeneic Syrian hamsters bearing HapT1 pancreatic tumors and with T cell receptor transgenic ACT in B16.OVA melanoma bearing C57BL/6 mice. The models are complementary: hamsters are semi-permissive for human oncolytic adenovirus, whereas detailed immunological analyses are possible in mice. In both models, local production of IL-2 successfully replaced the need for systemic recombinant IL-2 (rIL-2) administration and increased the efficacy of the cell therapy. Furthermore, vectored delivery of IL-2 significantly enhanced the infiltration of CD8+ T cells, M1-like macrophages, and B-cells while systemic rIL-2 increased CD25 + FoxP3+ T cells at the tumor. In contrast with vectored delivery, histopathological analysis of systemic rIL-2-treated animals revealed significant changes in lungs, livers, hearts, spleens, and kidneys. In summary, local IL-2 production results in efficacy and safety gains in the context of ACT. These preclinical assessments provide the rationale for ongoing clinical translation. © 2017 UICC.

Production and purification of non replicative canine adenovirus type 2 derived vectors.

PubMed

Szelechowski, Marion; Bergeron, Corinne; Gonzalez-Dunia, Daniel; Klonjkowski, Bernard

2013-12-03

Adenovirus (Ad) derived vectors have been widely used for short or long-term gene transfer, both for gene therapy and vaccine applications. Because of the frequent pre-existing immunity against the classically used human adenovirus type 5, canine adenovirus type 2 (CAV2) has been proposed as an alternative vector for human gene transfer. The well-characterized biology of CAV2, together with its ease of genetic manipulation, offer major advantages, notably for gene transfer into the central nervous system, or for inducing a wide range of protective immune responses, from humoral to cellular immunity. Nowadays, CAV2 represents one of the most appealing nonhuman adenovirus for use as a vaccine vector. This protocol describes a simple method to construct, produce and titer recombinant CAV2 vectors. After cloning the expression cassette of the gene of interest into a shuttle plasmid, the recombinant genomic plasmid is obtained by homologous recombination in the E. coli BJ5183 bacterial strain. The resulting genomic plasmid is then transfected into canine kidney cells expressing the complementing CAV2-E1 genes (DK-E1). A viral amplification enables the production of a large viral stock, which is purified by ultracentrifugation through cesium chloride gradients and desalted by dialysis. The resulting viral suspension routinely has a titer of over 10(10) infectious particles per ml and can be directly administrated in vivo.

Quantum and electromagnetic propagation with the conjugate symmetric Lanczos method.

PubMed

Acevedo, Ramiro; Lombardini, Richard; Turner, Matthew A; Kinsey, James L; Johnson, Bruce R

2008-02-14

The conjugate symmetric Lanczos (CSL) method is introduced for the solution of the time-dependent Schrodinger equation. This remarkably simple and efficient time-domain algorithm is a low-order polynomial expansion of the quantum propagator for time-independent Hamiltonians and derives from the time-reversal symmetry of the Schrodinger equation. The CSL algorithm gives forward solutions by simply complex conjugating backward polynomial expansion coefficients. Interestingly, the expansion coefficients are the same for each uniform time step, a fact that is only spoiled by basis incompleteness and finite precision. This is true for the Krylov basis and, with further investigation, is also found to be true for the Lanczos basis, important for efficient orthogonal projection-based algorithms. The CSL method errors roughly track those of the short iterative Lanczos method while requiring fewer matrix-vector products than the Chebyshev method. With the CSL method, only a few vectors need to be stored at a time, there is no need to estimate the Hamiltonian spectral range, and only matrix-vector and vector-vector products are required. Applications using localized wavelet bases are made to harmonic oscillator and anharmonic Morse oscillator systems as well as electrodynamic pulse propagation using the Hamiltonian form of Maxwell's equations. For gold with a Drude dielectric function, the latter is non-Hermitian, requiring consideration of corrections to the CSL algorithm.

Reprint of "versatile and stable vectors for efficient gene expression in Ralstonia eutropha H16".

PubMed

Gruber, Steffen; Hagen, Jeremias; Schwab, Helmut; Koefinger, Petra

2014-12-20

The Gram-negative β-proteobacterium Ralstonia eutropha H16 is primarily known for polyhydroxybutyrate (PHB) production and its ability to grow chemolithoautotrophically by using CO2 and H2 as sole carbon and energy sources. The majority of metabolic engineering and heterologous expression studies conducted so far rely on a small number of suitable expression systems. Particularly the plasmid based expression systems already developed for the use in R. eutropha H16 suffer from high segregational instability and plasmid loss after a short time of fermentation. In order to develop efficient and highly stable plasmid expression vectors for the use in R. eutropha H16, a new plasmid design was created including the RP4 partitioning system, as well as various promoters and origins of replication. The application of minireplicons derived from broad-host-range plasmids RSF1010, pBBR1, RP4 and pSa for the construction of expression vectors and the use of numerous, versatile promoters extend the range of feasible expression levels considerably. In particular, the use of promoters derived from the bacteriophage T5 was described for the first time in this work, characterizing the j5 promoter as the strongest promoter yet to be applied in R. eutropha H16. Moreover, the implementation of the RP4 partition sequence in plasmid design increased plasmid stability significantly and enables fermentations with marginal plasmid loss of recombinant R. eutropha H16 for at least 96h. The utility of the new vector family in R. eutropha H16 is demonstrated by providing expression data with different model proteins and consequently further raises the value of this organism as cell factory for biotechnological applications including protein and metabolite production. Copyright © 2014 Elsevier B.V. All rights reserved.

Versatile and stable vectors for efficient gene expression in Ralstonia eutropha H16.

PubMed

Gruber, Steffen; Hagen, Jeremias; Schwab, Helmut; Koefinger, Petra

2014-09-30

The Gram-negative β-proteobacterium Ralstonia eutropha H16 is primarily known for polyhydroxybutyrate (PHB) production and its ability to grow chemolithoautotrophically by using CO2 and H2 as sole carbon and energy sources. The majority of metabolic engineering and heterologous expression studies conducted so far rely on a small number of suitable expression systems. Particularly the plasmid based expression systems already developed for the use in R. eutropha H16 suffer from high segregational instability and plasmid loss after a short time of fermentation. In order to develop efficient and highly stable plasmid expression vectors for the use in R. eutropha H16, a new plasmid design was created including the RP4 partitioning system, as well as various promoters and origins of replication. The application of minireplicons derived from broad-host-range plasmids RSF1010, pBBR1, RP4 and pSa for the construction of expression vectors and the use of numerous, versatile promoters extend the range of feasible expression levels considerably. In particular, the use of promoters derived from the bacteriophage T5 was described for the first time in this work, characterizing the j5 promoter as the strongest promoter yet to be applied in R. eutropha H16. Moreover, the implementation of the RP4 partition sequence in plasmid design increased plasmid stability significantly and enables fermentations with marginal plasmid loss of recombinant R. eutropha H16 for at least 96 h. The utility of the new vector family in R. eutropha H16 is demonstrated by providing expression data with different model proteins and consequently further raises the value of this organism as cell factory for biotechnological applications including protein and metabolite production. Copyright © 2014 Elsevier B.V. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, B. -L.; Chang, L.; Ding, M.

A symmetry-preserving truncation of the two-body light-quark bound-state problem in relativistic quantum field theory is used to calculate the leading-twist parton distribution amplitudes (PDAs) of scalar systems, both ground-state and radial excitations, and the radial excitations of vector mesons. Owing to the fact that the scale-independent leptonic decay constant of a scalar meson constituted from equal-mass valence-constituents vanishes, it is found that the PDA of a given scalar system possesses one more zero than that of an analogous vector meson. Consequently, whereas the mean light-front relative momentum of the valence-constituents within a vector meson is zero, that within a scalarmore » meson is large, an outcome which hints at a greater role for light-front angular momentum in systems classified as P-wave in quantum mechanical models. Values for the scale-dependent decay constants of ground-state scalar and vector systems are a by-product of this analysis, and they turn out to be roughly equal, viz. ≃ 0.2 GeV at an hadronic scale. In addition, it is confirmed that the dilation characterising ground-state PDAs is manifest in the PDAs of radial excitations too. The impact of SU(3)-flavour symmetry breaking is also considered. When compared with pseudoscalar states, it is a little stronger in scalar systems, but the size is nevertheless determined by the flavour-dependence of dynamical chiral symmetry breaking and the PDAs are still skewed toward the heavier valence-quark in asymmetric systems.« less

Simplified production and concentration of HIV-1-based lentiviral vectors using HYPERFlask vessels and anion exchange membrane chromatography

PubMed Central

Kutner, Robert H; Puthli, Sharon; Marino, Michael P; Reiser, Jakob

2009-01-01

Background During the past twelve years, lentiviral (LV) vectors have emerged as valuable tools for transgene delivery because of their ability to transduce nondividing cells and their capacity to sustain long-term transgene expression in target cells in vitro and in vivo. However, despite significant progress, the production and concentration of high-titer, high-quality LV vector stocks is still cumbersome and costly. Methods Here we present a simplified protocol for LV vector production on a laboratory scale using HYPERFlask vessels. HYPERFlask vessels are high-yield, high-performance flasks that utilize a multilayered gas permeable growth surface for efficient gas exchange, allowing convenient production of high-titer LV vectors. For subsequent concentration of LV vector stocks produced in this way, we describe a facile protocol involving Mustang Q anion exchange membrane chromatography. Results Our results show that unconcentrated LV vector stocks with titers in excess of 108 transduction units (TU) per ml were obtained using HYPERFlasks and that these titers were higher than those produced in parallel using regular 150-cm2 tissue culture dishes. We also show that up to 500 ml of an unconcentrated LV vector stock prepared using a HYPERFlask vessel could be concentrated using a single Mustang Q Acrodisc with a membrane volume of 0.18 ml. Up to 5.3 × 1010 TU were recovered from a single HYPERFlask vessel. Conclusion The protocol described here is easy to implement and should facilitate high-titer LV vector production for preclinical studies in animal models without the need for multiple tissue culture dishes and ultracentrifugation-based concentration protocols. PMID:19220915

Silencing Genes in the Heart.

PubMed

Fechner, Henry; Vetter, Roland; Kurreck, Jens; Poller, Wolfgang

2017-01-01

Silencing of cardiac genes by RNA interference (RNAi) has developed into a powerful new method to treat cardiac diseases. Small interfering (si)RNAs are the inducers of RNAi, but cultured primary cardiomyocytes and heart are highly resistant to siRNA transfection. This can be overcome by delivery of small hairpin (sh)RNAs or artificial microRNA (amiRNAs) by cardiotropic adeno-associated virus (AAV) vectors. Here we describe as example of the silencing of a cardiac gene, the generation and cloning of shRNA, and amiRNAs directed against the cardiac protein phospholamban. We further describe the generation of AAV shuttle plasmids with self complementary vector genomes, the production of AAV vectors in roller bottles, and their purification via iodixanol gradient centrifugation and concentration with filter systems. Finally we describe the preparation of primary neonatal rat cardiomyocytes (PNRC), the transduction of PNRC with AAV vectors, and the maintenance of the transduced cell culture.

Cloning and Expression of Major Surface Antigen 1 Gene of Toxoplasma gondii RH Strain Using the Expression Vector pVAX1 in Chinese Hamster Ovary Cells

PubMed Central

Abdizadeh, Rahman; Maraghi, Sharif; Ghadiri, Ata A.; Tavalla, Mehdi; Shojaee, Saeedeh

2015-01-01

Background: Toxoplasmosis is an opportunistic protozoan infection with a high prevalence in a broad range of hosts infecting up to one-third of the world human population. Toxoplasmosis leads to serious medical problems in immunocompromised individuals and fetuses and also induces abortion and mortality in domestic animals. Therefore, there is a huge demand for the development of an effective vaccine. Surface Antigen 1 (SAG1) is one of the important immunodominant surface antigens of Toxoplasma gondii, which interacts with host cells and primarily involved in adhesion, invasion and stimulation of host immune response. Surface antigen 1 is considered as the leading candidate for development of an effective vaccine against toxoplasmosis. Objectives: The purpose of this study was to clone the major surface antigen1 gene (SAG1) from the genotype 1 of T. gondii, RH strain into the eukaryotic expression vector pVAX1 in order to use for a DNA vaccine. Materials and Methods: Genomic DNA was extracted from tachyzoite of the parasite using the QIAamp DNA mini kit. After designing the specific primers, SAG1 gene was amplified by Polymerase Chain Reaction (PCR). The purified PCR products were then cloned into a pPrime plasmid vector. The aforementioned product was subcloned into the pVAX1 eukaryotic expression vector. The recombinant pVAX1-SAG1 was then transfected into Chinese Hamster Ovary (CHO) cells and expression of SAG1 antigen was evaluated using Reverse Transcriptase Polymerase Chain Reaction (RT-PCR), Immunofluorescence Assay (IFA) and Western Blotting (WB). Results: The cloning and subcloning products (pPrime-SAG1 and pVAX1-SAG1 plasmid vectors) of SAG1 gene were verified and confirmed by enzyme digestion and sequencing. A 30 kDa recombinant protein was expressed in CHO cells as shown by IFA and WB methods. Conclusions: The pVAX1 expression vector and CHO cells are a suitable system for high-level recombinant protein production for SAG1 gene from T. gondii parasites and are promising approaches for antigen preparation in vaccine development. PMID:25861441

Commensal Streptococcus mitis is a unique vector for oral mucosal vaccination

PubMed Central

Daifalla, Nada; Cayabyab, Mark J.; Xie, Emily; Kim, Hyeun Bum; Tzipori, Saul; Stashenko, Philip; Duncan, Margaret; Campos-Neto, Antonio

2014-01-01

The development of vaccine approaches that induce mucosal and systemic immune responses is critical for the effective prevention of several infections. Here, we report on the use of the abundant human oral commensal bacterium Streptococcus mitis as a delivery vehicle for mucosal immunization. Using homologous recombination we generated a stable rS. mitis expressing a Mycobacterium tuberculosis protein (Ag85b). Oral administration of rS. mitis in gnotobiotic piglets resulted in efficient oral colonization and production of oral and systemic anti-Ag85b specific IgA and IgG antibodies. These results support that the commensal S. mitis is potentially a useful vector for mucosal vaccination. PMID:25522856

Development of a baculovirus vector carrying a small hairpin RNA for suppression of sf-caspase-1 expression and improvement of recombinant protein production.

PubMed

Zhang, Xiaoyue; Xu, Keyan; Ou, Yanmei; Xu, Xiaodong; Chen, Hongying

2018-05-02

The Baculovirus expression vector system (BEVS) is a transient expression platform for recombinant protein production in insect cells. Baculovirus infection of insect cells will shutoff host translation and induce apoptosis and lead to the termination of protein expression. Previous reports have demonstrated the enhancement of protein yield in BEVS using stable insect cell lines expressing interference RNA to suppress the expression of caspase-1. In this study, short-hairpin RNA (shRNA) expression cassettes targeting Spodoptera frugiperda caspase-1 (Sf-caspase-1) were constructed and inserted into an Autographa californica multiple nucleopolyhedrovirus (AcMNPV) vector. Using the recombinant baculovirus vectors, we detected the suppression of Sf-caspase-1 expression and cell apoptosis. Green fluorescent protein (GFP), Discosoma sp. Red (DsRed) and firefly luciferase were then expressed as reporter proteins. The results showed that suppression of apoptosis enhanced the accumulation of exogenous proteins at 2 and 3 days post infection. After 4 days post infection, the activity of the reporter proteins remained higher in BEVS using the baculovirus carrying shRNA in comparison with the control without shRNA, but the accumulated protein levels showed no obvious difference between them, suggesting that apoptosis suppression resulted in improved protein folding rather than translation efficiency at the very late stage of baculovirus infection. The baculovirus vector developed in this study would be a useful tool for the production of active proteins suitable for structural and functional studies or pharmaceutical applications in Sf9 cells, and it also has the potential to be adapted for the improvement of protein expression in different insect cell lines that can be infected by AcMNPV.

Searches For Pair Production of Heavy Vector-Like Quarks in pp Collisions at √s =13 Tev with the Atlas Detector

NASA Astrophysics Data System (ADS)

Marley, Daniel Edison

A summary of two orthogonal and complementary searches for pair production of heavy vector-like quarks is presented. The first analysis considers final states with 1-lepton and hadronic jets that can be used to reconstruct the vector-like quark system after identifying the boosted decay of W bosons and building the neutrino from the missing transverse momentum. Control and signal regions constrain the uncertainties associated with top quark production that most negatively impact this analysis. The second analysis searches in the fully-hadronic final state where no leptons are considered. An explicit veto on the lepton number ensures orthogonality between the analyses. The analysis uses a deep neural network to reconstruct the decays of heavy objects and a data-driven technique to estimate the dominant multi-jet background. Data were recorded by the ATLAS experiment in sqrt(s)=13 TeV proton-proton collisions delivered by the Large Hadron Collider in 2015 and 2016. No excess above the Standard Model background is observed in either analysis. The results from the 1-lepton analysis are interpreted assuming vector-like quarks decay TT¯ → Wb + X. The 0-lepton analysis assumes both types of pair produced vector-like quarks TT and BB that decay to T → Wb, Ht, Zt and B → Wt, Hb, Zb. The 1-lepton analysis observes a 95% CL lower limit on the T mass of 1090 (810) GeV assuming the scenario BR(T → Wb)=1 (SU(2) singlet). The strongest observed 95% CL lower limits in the 0-lepton analysis are 850 GeV and 903 GeV in the BR(T → Ht) = 1 and BR(B → Hb)=1 decays, respectively.

Systems Biology of Recombinant Protein Production in Bacillus megaterium

NASA Astrophysics Data System (ADS)

Biedendieck, Rebekka; Bunk, Boyke; Fürch, Tobias; Franco-Lara, Ezequiel; Jahn, Martina; Jahn, Dieter

Over the last two decades the Gram-positive bacterium Bacillus megaterium was systematically developed to a useful alternative protein production host. Multiple vector systems for high yield intra- and extracellular protein production were constructed. Strong inducible promoters were combined with DNA sequences for optimised ribosome binding sites, various leader peptides for protein export and N- as well as C-terminal affinity tags for affinity chromatographic purification of the desired protein. High cell density cultivation and recombinant protein production were successfully tested. For further system biology based control and optimisation of the production process the genomes of two B. megaterium strains were completely elucidated, DNA arrays designed, proteome, fluxome and metabolome analyses performed and all data integrated using the bioinformatics platform MEGABAC. Now, solid theoretical and experimental bases for primary modeling attempts of the production process are available.

Ecological, biological and social dimensions of dengue vector breeding in five urban settings of Latin America: a multi-country study

PubMed Central

2014-01-01

Background Dengue is an increasingly important public health problem in most Latin American countries and more cost-effective ways of reducing dengue vector densities to prevent transmission are in demand by vector control programs. This multi-centre study attempted to identify key factors associated with vector breeding and development as a basis for improving targeted intervention strategies. Methods In each of 5 participant cities in Mexico, Colombia, Ecuador, Brazil and Uruguay, 20 clusters were randomly selected by grid sampling to incorporate 100 contiguous households, non-residential private buildings (businesses) and public spaces. Standardized household surveys, cluster background surveys and entomological surveys specifically targeted to obtain pupal indices for Aedes aegypti, were conducted in the dry and wet seasons. Results The study clusters included mainly urban low-middle class populations with satisfactory infrastructure and –except for Uruguay- favourable climatic conditions for dengue vector development. Household knowledge about dengue and “dengue mosquitoes” was widespread, mainly through mass media, but there was less awareness around interventions to reduce vector densities. Vector production (measured through pupal indices) was favoured when water containers were outdoor, uncovered, unused (even in Colombia and Ecuador where the large tanks used for household water storage and washing were predominantly productive) and –particularly during the dry season- rainwater filled. Larval infestation did not reflect productive container types. All productive container types, including those important in the dry season, were identified by pupal surveys executed during the rainy season. Conclusions A number of findings are relevant for improving vector control: 1) there is a need for complementing larval surveys with occasional pupal surveys (to be conducted during the wet season) for identifying and subsequently targeting productive container types; 2) the need to raise public awareness about useful and effective interventions in productive container types specific to their area; and 3) the motivation for control services that-according to this and similar studies in Asia- dedicated, targeted vector management can make a difference in terms of reducing vector abundance. PMID:24447796

A CGMMV genome-replicon vector with partial sequences of coat protein gene efficiently expresses GFP in Nicotiana benthamiana.

PubMed

Jailani, A Abdul Kader; Solanki, Vikas; Roy, Anirban; Sivasudha, T; Mandal, Bikash

2017-04-02

A highly infectious clone of Cucumber green mottle mosaic virus (CGMMV), a cucurbit-infecting tobamovirus was utilized for designing of gene expression vectors. Two versions of vector were examined for their efficacy in expressing the green fluorescent protein (GFP) in Nicotiana benthamiana. When the GFP gene was inserted at the stop codon of coat protein (CP) gene of the CGMMV genome without any read-through codon, systemic expression of GFP, as well as virion formation and systemic symptoms expression were obtained in N. benthamiana. The qRT-PCR analysis showed 23 fold increase of GFP over actin at 10days post inoculation (dpi), which increased to 45 fold at 14dpi and thereafter the GFP expression was significantly declined. Further, we show that when the most of the CP sequence is deleted retaining only the first 105 nucleotides, the shortened vector containing GFP in frame of original CP open reading frame (ORF) resulted in 234 fold increase of GFP expression over actin at 5dpi in N. benthamiana without the formation of virions and disease symptoms. Our study demonstrated that a simple manipulation of CP gene in the CGMMV genome while preserving the translational frame of CP resulted in developing a virus-free, rapid and efficient foreign protein expression system in the plant. The CGMMV based vectors developed in this study may be potentially useful for the production of edible vaccines in cucurbits. Copyright © 2017 Elsevier B.V. All rights reserved.

Production of lentiviral vectors

PubMed Central

Merten, Otto-Wilhelm; Hebben, Matthias; Bovolenta, Chiara

2016-01-01

Lentiviral vectors (LV) have seen considerably increase in use as gene therapy vectors for the treatment of acquired and inherited diseases. This review presents the state of the art of the production of these vectors with particular emphasis on their large-scale production for clinical purposes. In contrast to oncoretroviral vectors, which are produced using stable producer cell lines, clinical-grade LV are in most of the cases produced by transient transfection of 293 or 293T cells grown in cell factories. However, more recent developments, also, tend to use hollow fiber reactor, suspension culture processes, and the implementation of stable producer cell lines. As is customary for the biotech industry, rather sophisticated downstream processing protocols have been established to remove any undesirable process-derived contaminant, such as plasmid or host cell DNA or host cell proteins. This review compares published large-scale production and purification processes of LV and presents their process performances. Furthermore, developments in the domain of stable cell lines and their way to the use of production vehicles of clinical material will be presented. PMID:27110581

In vivo production of recombinant proteins using occluded recombinant AcMNPV-derived baculovirus vectors.

PubMed

Guijarro-Pardo, Eva; Gómez-Sebastián, Silvia; Escribano, José M

2017-12-01

Trichoplusia ni insect larvae infected with vectors derived from the Autographa californica multiple nucleopolyhedrovirus (AcMNPV), are an excellent alternative to insect cells cultured in conventional bioreactors to produce recombinant proteins because productivity and cost-efficiency reasons. However, there is still a lot of work to do to reduce the manual procedures commonly required in this production platform that limit its scalability. To increase the scalability of this platform technology, a current bottleneck to be circumvented in the future is the need of injection for the inoculation of larvae with polyhedrin negative baculovirus vectors (Polh-) because of the lack of oral infectivity of these viruses, which are commonly used for production in insect cell cultures. In this work we have developed a straightforward alternative to obtain orally infective vectors derived from AcMNPV and expressing recombinant proteins that can be administered to the insect larvae (Trichoplusia ni) by feeding, formulated in the insect diet. The approach developed was based on the use of a recombinant polyhedrin protein expressed by a recombinant vector (Polh+), able to co-occlude any recombinant Polh- baculovirus vector expressing a recombinant protein. A second alternative was developed by the generation of a dual vector co-expressing the recombinant polyhedrin protein and the foreign gene of interest to obtain the occluded viruses. Additionally, by the incorporation of a reporter gene into the helper Polh+ vector, it was possible the follow-up visualization of the co-occluded viruses infection in insect larvae and will help to homogenize infection conditions. By using these methodologies, the production of recombinant proteins in per os infected larvae, without manual infection procedures, was very similar in yield to that obtained by manual injection of recombinant Polh- AcMNPV-based vectors expressing the same proteins. However, further analyses will be required for a detailed comparison of production yields reached by injection vs oral infections for different recombinant proteins. In conclusion, these results open the possibility of future industrial scaling-up production of recombinant proteins in insect larvae by reducing manual operations. Copyright © 2017 Elsevier B.V. All rights reserved.

Efficient production of human acidic fibroblast growth factor in pea (Pisum sativum L.) plants by agroinfection of germinated seeds

PubMed Central

2011-01-01

Background For efficient and large scale production of recombinant proteins in plants transient expression by agroinfection has a number of advantages over stable transformation. Simple manipulation, rapid analysis and high expression efficiency are possible. In pea, Pisum sativum, a Virus Induced Gene Silencing System using the pea early browning virus has been converted into an efficient agroinfection system by converting the two RNA genomes of the virus into binary expression vectors for Agrobacterium transformation. Results By vacuum infiltration (0.08 Mpa, 1 min) of germinating pea seeds with 2-3 cm roots with Agrobacteria carrying the binary vectors, expression of the gene for Green Fluorescent Protein as marker and the gene for the human acidic fibroblast growth factor (aFGF) was obtained in 80% of the infiltrated developing seedlings. Maximal production of the recombinant proteins was achieved 12-15 days after infiltration. Conclusions Compared to the leaf injection method vacuum infiltration of germinated seeds is highly efficient allowing large scale production of plants transiently expressing recombinant proteins. The production cycle of plants for harvesting the recombinant protein was shortened from 30 days for leaf injection to 15 days by applying vacuum infiltration. The synthesized aFGF was purified by heparin-affinity chromatography and its mitogenic activity on NIH 3T3 cells confirmed to be similar to a commercial product. PMID:21548923

Transgene Expression and Host Cell Responses to Replication-Defective, Single-Cycle, and Replication-Competent Adenovirus Vectors.

PubMed

Crosby, Catherine M; Barry, Michael A

2017-02-18

Most adenovirus (Ad) vectors are E1 gene deleted replication defective (RD-Ad) vectors that deliver one transgene to the cell and all expression is based on that one gene. In contrast, E1-intact replication-competent Ad (RC-Ad) vectors replicate their DNA and their transgenes up to 10,000-fold, amplifying transgene expression markedly higher than RD-Ad vectors. While RC-Ad are more potent, they run the real risk of causing adenovirus infections in vector recipients and those that administer them. To gain the benefits of transgene amplification, but avoid the risk of Ad infections, we developed "single cycle" Ad (SC-Ad) vectors. SC-Ads amplify transgene expression and generated markedly stronger and more persistent immune responses than RD-Ad as expected. However, they also unexpectedly generated stronger immune responses than RC-Ad vectors. To explore the basis of this potency here, we compared gene expression and the cellular responses to infection to these vectors in vitro and in vivo. In vitro, in primary human lung epithelial cells, SC- and RC-Ad amplified their genomes more than 400-fold relative to RD-Ad with higher replication by SC-Ad. This replication translated into higher green fluorescent protein (GFP) expression for 48 h by SC- and RC-Ad than by RD-Ad. In vitro, in the absence of an immune system, RD-Ad expression became higher by 72 h coincident with cell death mediated by SC- and RC-Ad and release of transgene product from the dying cells. When the vectors were compared in human THP-1 Lucia- interferon-stimulated gene (ISG) cells, which are a human monocyte cell line that have been modified to quantify ISG activity, RC-Ad6 provoked significantly stronger ISG responses than RD- or SC-Ad. In mice, intravenous or intranasal injection produced up to 100-fold genome replication. Under these in vivo conditions in the presence of the immune system, luciferase expression by RC and SC-Ad was markedly higher than that by RD-Ad. In immunodeficient mice, SC-Ad drove stronger luciferase expression than RC- or RD-Ad. These data demonstrate better transgene expression by SC- and RC-Ad in vitro and in vivo than RD-Ad. This higher expression by the replicating vectors results in a peak of expression within 1 to 2 days followed by cell death of infected cells and release of transgene products. While SC- and RC-Ad expression were similar in mice and in Syrian hamsters, RC-Ad provoked much stronger ISG induction which may explain in part SC-Ad's ability to generate stronger and more persistent immune responses than RC-Ad in Ad permissive hamsters.

Development of an optimized tetracycline-inducible expression system to increase the accumulation of interleukin-10 in tobacco BY-2 suspension cells

PubMed Central

2012-01-01

Background Plant cell suspension cultures can be used for the production of valuable pharmaceutical and industrial proteins. When the recombinant protein is secreted into the culture medium, restricting expression to a defined growth phase can improve both the quality and quantity of the recovered product by minimizing proteolytic activity. Temporal restriction is also useful for recombinant proteins whose constitutive expression affects cell growth and viability, such as viral interleukin-10 (vIL-10). Results We have developed a novel, tetracycline-inducible system suitable for tobacco BY-2 suspension cells which increases the yields of vIL-10. The new system is based on a binary vector that is easier to handle than conventional vectors, contains an enhanced inducible promoter and 5′-UTR to improve yields, and incorporates a constitutively-expressed visible marker gene to allow the rapid and straightforward selection of the most promising transformed clones. Stable transformation of BY-2 cells with this vector, without extensive optimization of the induction conditions, led to a 3.5 fold increase in vIL-10 levels compared to constitutive expression in the same host. Conclusions We have developed an effective and straightforward molecular farming platform technology that improves both the quality and the quantity of recombinant proteins produced in plant cells, particularly those whose constitutive expression has a negative impact on plant growth and development. Although we tested the platform using vIL-10 produced in BY-2 cells, it can be applied to other host/product combinations and is also useful for basic research requiring strictly controlled transgene expression. PMID:22784336

[Prokaryotic expression systems].

PubMed

Porowińska, Dorota; Wujak, Magdalena; Roszek, Katarzyna; Komoszyński, Michał

2013-03-01

For overproduction of recombinant proteins both eukaryotic and prokaryotic expression systems are used. Choosing the right system depends, among other things, on the growth rate and culture of host cells, level of the target gene expression and posttranslational processing of the synthesized protein. Regardless of the type of expression system, its basic elements are the vector and the expression host. The most widely used system for protein overproduction, both on a laboratory and industrial scale, is the prokaryotic system. This system is based primarily on the bacteria E. coli, although increasingly often Bacillus species are used. The prokaryotic system allows one to obtain large quantities of recombinant proteins in a short time. A simple and inexpensive bacterial cell culture and well-known mechanisms of transcription and translation facilitate the use of these microorganisms. The simplicity of genetic modifications and the availability of many bacterial mutants are additional advantages of the prokaryotic system. In this article we characterize the structural elements of prokaryotic expression vectors. Also strategies for preparation of the target protein gene that increase productivity, facilitate detection and purification of recombinant protein and provide its activity are discussed. Bacterial strains often used as host cells in expression systems as well as the potential location of heterologous proteins are characterized. Knowledge of the basic elements of the prokaryotic expression system allows for production of biologically active proteins in a short time and in satisfactory quantities. 

Insecticide resistance profile of Anopheles gambiae from a phase II field station in Cové, southern Benin: implications for the evaluation of novel vector control products.

PubMed

Ngufor, Corine; N'Guessan, Raphael; Fagbohoun, Josias; Subramaniam, Krishanthi; Odjo, Abibatou; Fongnikin, Augustin; Akogbeto, Martin; Weetman, David; Rowland, Mark

2015-11-18

Novel indoor residual spraying (IRS) and long-lasting insecticidal net (LLIN) products aimed at improving the control of pyrethroid-resistant malaria vectors have to be evaluated in Phase II semi-field experimental studies against highly pyrethroid-resistant mosquitoes. To better understand their performance it is necessary to fully characterize the species composition, resistance status and resistance mechanisms of the vector populations in the experimental hut sites. Bioassays were performed to assess phenotypic insecticide resistance in the malaria vector population at a newly constructed experimental hut site in Cové, a rice growing area in southern Benin, being used for WHOPES Phase II evaluation of newly developed LLIN and IRS products. The efficacy of standard WHOPES-approved pyrethroid LLIN and IRS products was also assessed in the experimental huts. Diagnostic genotyping techniques and microarray studies were performed to investigate the genetic basis of pyrethroid resistance in the Cové Anopheles gambiae population. The vector population at the Cové experimental hut site consisted of a mixture of Anopheles coluzzii and An. gambiae s.s. with the latter occurring at lower frequencies (23 %) and only in samples collected in the dry season. There was a high prevalence of resistance to pyrethroids and DDT (>90 % bioassay survival) with pyrethroid resistance intensity reaching 200-fold compared to the laboratory susceptible An. gambiae Kisumu strain. Standard WHOPES-approved pyrethroid IRS and LLIN products were ineffective in the experimental huts against this vector population (8-29 % mortality). The L1014F allele frequency was 89 %. CYP6P3, a cytochrome P450 validated as an efficient metabolizer of pyrethroids, was over-expressed. Characterizing pyrethroid resistance at Phase II field sites is crucial to the accurate interpretation of the performance of novel vector control products. The strong levels of pyrethroid resistance at the Cové experimental hut station make it a suitable site for Phase II experimental hut evaluations of novel vector control products, which aim for improved efficacy against pyrethroid-resistant malaria vectors to WHOPES standards. The resistance genes identified can be used as markers for further studies investigating the resistance management potential of novel mixture LLIN and IRS products tested at the site.

New signals for vector-like down-type quark in U(1) of E_6

NASA Astrophysics Data System (ADS)

Das, Kasinath; Li, Tianjun; Nandi, S.; Rai, Santosh Kumar

2018-01-01

We consider the pair production of vector-like down-type quarks in an E_6 motivated model, where each of the produced down-type vector-like quark decays into an ordinary Standard Model light quark and a singlet scalar. Both the vector-like quark and the singlet scalar appear naturally in the E_6 model with masses at the TeV scale with a favorable choice of symmetry breaking pattern. We focus on the non-standard decay of the vector-like quark and the new scalar which decays to two photons or two gluons. We analyze the signal for the vector-like quark production in the 2γ +≥ 2j channel and show how the scalar and vector-like quark masses can be determined at the Large Hadron Collider.

Vector-Boson Fusion Higgs Production at Three Loops in QCD.

PubMed

Dreyer, Frédéric A; Karlberg, Alexander

2016-08-12

We calculate the next-to-next-to-next-to-leading-order (N^{3}LO) QCD corrections to inclusive vector-boson fusion Higgs production at proton colliders, in the limit in which there is no color exchange between the hadronic systems associated with the two colliding protons. We also provide differential cross sections for the Higgs transverse momentum and rapidity distributions. We find that the corrections are at the 1‰-2‰ level, well within the scale uncertainty of the next-to-next-to-leading-order calculation. The associated scale uncertainty of the N^{3}LO calculation is typically found to be below the 2‰ level. We also consider theoretical uncertainties due to missing higher order parton distribution functions, and provide an estimate of their importance.

pPCV, a versatile vector for cloning PCR products.

PubMed

Janner, Christiane R; Brito, Ana Lívia P; Moraes, Lidia Maria P; Reis, Viviane Cb; Torres, Fernando Ag

2013-01-01

The efficiency of PCR product cloning depends on the nature of the DNA polymerase employed because amplicons may have blunt-ends or 3' adenosines overhangs. Therefore, for amplicon cloning, available commercial vectors are either blunt-ended or have a single 3' overhanging thymidine. The aim of this work was to offer in a single vector the ability to clone both types of PCR products. For that purpose, a minimal polylinker was designed to include restriction sites for EcoRV and XcmI which enable direct cloning of amplicons bearing blunt-ends or A-overhangs, respectively, still offering blue/white selection. When tested, the resulting vector, pPCV, presented high efficiency cloning of both types of amplicons.

Water deficit enhances the transmission of plant viruses by insect vectors

PubMed Central

Yvon, Michel; Vile, Denis; Dader, Beatriz; Fereres, Alberto

2017-01-01

Drought is a major threat to crop production worldwide and is accentuated by global warming. Plant responses to this abiotic stress involve physiological changes overlapping, at least partially, the defense pathways elicited both by viruses and their herbivore vectors. Recently, a number of theoretical and empirical studies anticipated the influence of climate changes on vector-borne viruses of plants and animals, mainly addressing the effects on the virus itself or on the vector population dynamics, and inferring possible consequences on virus transmission. Here, we directly assess the effect of a severe water deficit on the efficiency of aphid-transmission of the Cauliflower mosaic virus (CaMV) or the Turnip mosaic virus (TuMV). For both viruses, our results demonstrate that the rate of vector-transmission is significantly increased from water-deprived source plants: CaMV transmission reproducibly increased by 34% and that of TuMV by 100%. In both cases, the enhanced transmission rate could not be explained by a higher virus accumulation, suggesting a more complex drought-induced process that remains to be elucidated. The evidence that infected plants subjected to drought are much better virus sources for insect vectors may have extensive consequences for viral epidemiology, and should be investigated in a wide range of plant-virus-vector systems. PMID:28467423

Water deficit enhances the transmission of plant viruses by insect vectors.

PubMed

van Munster, Manuella; Yvon, Michel; Vile, Denis; Dader, Beatriz; Fereres, Alberto; Blanc, Stéphane

2017-01-01

Drought is a major threat to crop production worldwide and is accentuated by global warming. Plant responses to this abiotic stress involve physiological changes overlapping, at least partially, the defense pathways elicited both by viruses and their herbivore vectors. Recently, a number of theoretical and empirical studies anticipated the influence of climate changes on vector-borne viruses of plants and animals, mainly addressing the effects on the virus itself or on the vector population dynamics, and inferring possible consequences on virus transmission. Here, we directly assess the effect of a severe water deficit on the efficiency of aphid-transmission of the Cauliflower mosaic virus (CaMV) or the Turnip mosaic virus (TuMV). For both viruses, our results demonstrate that the rate of vector-transmission is significantly increased from water-deprived source plants: CaMV transmission reproducibly increased by 34% and that of TuMV by 100%. In both cases, the enhanced transmission rate could not be explained by a higher virus accumulation, suggesting a more complex drought-induced process that remains to be elucidated. The evidence that infected plants subjected to drought are much better virus sources for insect vectors may have extensive consequences for viral epidemiology, and should be investigated in a wide range of plant-virus-vector systems.

Invasive mutualisms between a plant pathogen and insect vectors in the Middle East and Brazil

PubMed Central

Queiroz, Renan Batista; Silva, Fábio Nascimento; Al-Mahmmoli, Issa Hashil; Al-Sadi, Abdullah Mohammed; Carvalho, Claudine Márcia; Elliot, Simon L.

2016-01-01

Complex multi-trophic interactions in vectorborne diseases limit our understanding and ability to predict outbreaks. Arthropod-vectored pathogens are especially problematic, with the potential for novel interspecific interactions during invasions. Variations and novelties in plant–arthropod–pathogen triumvirates present significant threats to global food security. We examined aspects of a phytoplasma pathogen of citrus across two continents. ‘Candidatus Phytoplasma aurantifolia’ causes Witches' Broom Disease of Lime (WBDL) and has devastated citrus production in the Middle East. A variant of this phytoplasma currently displays asymptomatic or ‘silent’ infections in Brazil. We first studied vector capacity and fitness impacts of the pathogen on its vectors. The potential for co-occurring weed species to act as pathogen reservoirs was analysed and key transmission periods in the year were also studied. We demonstrate that two invasive hemipteran insects—Diaphorina citri and Hishimonus phycitis—can vector the phytoplasma. Feeding on phytoplasma-infected hosts greatly increased reproduction of its invasive vector D. citri both in Oman and Brazil; suggesting that increased fitness of invasive insect vectors thereby further increases the pathogen's capacity to spread. Based on our findings, this is a robust system for studying the effects of invasions on vectorborne diseases and highlights concerns about its spread to warmer, drier regions of Brazil. PMID:28083099

Coherent states for the relativistic harmonic oscillator

NASA Technical Reports Server (NTRS)

Aldaya, Victor; Guerrero, J.

1995-01-01

Recently we have obtained, on the basis of a group approach to quantization, a Bargmann-Fock-like realization of the Relativistic Harmonic Oscillator as well as a generalized Bargmann transform relating fock wave functions and a set of relativistic Hermite polynomials. Nevertheless, the relativistic creation and annihilation operators satisfy typical relativistic commutation relations of the Lie product (vector-z, vector-z(sup dagger)) approximately equals Energy (an SL(2,R) algebra). Here we find higher-order polarization operators on the SL(2,R) group, providing canonical creation and annihilation operators satisfying the Lie product (vector-a, vector-a(sup dagger)) = identity vector 1, the eigenstates of which are 'true' coherent states.

Production of first generation adenoviral vectors for preclinical protocols: amplification, purification and functional titration.

PubMed

Armendáriz-Borunda, Juan; Bastidas-Ramírez, Blanca Estela; Sandoval-Rodríguez, Ana; González-Cuevas, Jaime; Gómez-Meda, Belinda; García-Bañuelos, Jesús

2011-11-01

Gene therapy represents a promising approach in the treatment of several diseases. Currently, the ideal vector has yet to be designed; though, adenoviral vectors (Ad-v) have provided the most utilized tool for gene transfer due principally to their simple production, among other specific characteristics. Ad-v viability represents a critical variable that may be affected by storage or shipping conditions and therefore it is advisable to be assessed previously to protocol performance. The present work is unique in this matter, as the complete detailed process to obtain Ad-v of preclinical grade is explained. Amplification in permissive HEK-293 cells, purification in CsCl gradients in a period of 10 h, spectrophotometric titration of viral particles (VP) and titration of infectious units (IU), yielding batches of AdβGal, AdGFP, AdHuPA and AdMMP8, of approximately 10¹³-10¹⁴ VP and 10¹²-10¹³ IU were carried out. In vivo functionality of therapeutic AdHuPA and AdMMP8 was evidenced in rats presenting CCl₄-induced fibrosis, as more than 60% of fibrosis was eliminated in livers after systemic delivery through iliac vein in comparison with irrelevant AdβGal. Time required to accomplish the whole Ad-v production steps, including IU titration was 20 to 30 days. We conclude that production of Ad-v following standard operating procedures assuring vector functionality and the possibility to effectively evaluate experimental gene therapy results, leaving aside the use of high-cost commercial kits or sophisticated instrumentation, can be performed in a conventional laboratory of cell culture. Copyright © 2011 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Automatic River Network Extraction from LIDAR Data

NASA Astrophysics Data System (ADS)

Maderal, E. N.; Valcarcel, N.; Delgado, J.; Sevilla, C.; Ojeda, J. C.

2016-06-01

National Geographic Institute of Spain (IGN-ES) has launched a new production system for automatic river network extraction for the Geospatial Reference Information (GRI) within hydrography theme. The goal is to get an accurate and updated river network, automatically extracted as possible. For this, IGN-ES has full LiDAR coverage for the whole Spanish territory with a density of 0.5 points per square meter. To implement this work, it has been validated the technical feasibility, developed a methodology to automate each production phase: hydrological terrain models generation with 2 meter grid size and river network extraction combining hydrographic criteria (topographic network) and hydrological criteria (flow accumulation river network), and finally the production was launched. The key points of this work has been managing a big data environment, more than 160,000 Lidar data files, the infrastructure to store (up to 40 Tb between results and intermediate files), and process; using local virtualization and the Amazon Web Service (AWS), which allowed to obtain this automatic production within 6 months, it also has been important the software stability (TerraScan-TerraSolid, GlobalMapper-Blue Marble , FME-Safe, ArcGIS-Esri) and finally, the human resources managing. The results of this production has been an accurate automatic river network extraction for the whole country with a significant improvement for the altimetric component of the 3D linear vector. This article presents the technical feasibility, the production methodology, the automatic river network extraction production and its advantages over traditional vector extraction systems.

Prediction of B-cell linear epitopes with a combination of support vector machine classification and amino acid propensity identification.

PubMed

Wang, Hsin-Wei; Lin, Ya-Chi; Pai, Tun-Wen; Chang, Hao-Teng

2011-01-01

Epitopes are antigenic determinants that are useful because they induce B-cell antibody production and stimulate T-cell activation. Bioinformatics can enable rapid, efficient prediction of potential epitopes. Here, we designed a novel B-cell linear epitope prediction system called LEPS, Linear Epitope Prediction by Propensities and Support Vector Machine, that combined physico-chemical propensity identification and support vector machine (SVM) classification. We tested the LEPS on four datasets: AntiJen, HIV, a newly generated PC, and AHP, a combination of these three datasets. Peptides with globally or locally high physicochemical propensities were first identified as primitive linear epitope (LE) candidates. Then, candidates were classified with the SVM based on the unique features of amino acid segments. This reduced the number of predicted epitopes and enhanced the positive prediction value (PPV). Compared to four other well-known LE prediction systems, the LEPS achieved the highest accuracy (72.52%), specificity (84.22%), PPV (32.07%), and Matthews' correlation coefficient (10.36%).

Variational Iterative Methods for Nonsymmetric Systems of Linear Equations.

DTIC Science & Technology

1981-08-01

With a third matrix-vector product, b(i) can be computed as i j ( ATAr i+l’pj)/ApjpApj), and the previous (Apj) need not be saved. Page 8 I OCR I Orthomin... Economics and Mathematical Systems, Volume 134, Springer-Verlag, Berlin, 1976. [51 Paul Concus, Gene H. Golub, and Dianne P. O’Leary. A generalized

Estimating dengue vector abundance in the wet and dry season: implications for targeted vector control in urban and peri-urban Asia

PubMed Central

Wai, Khin Thet; Arunachalam, Natarajan; Tana, Susilowati; Espino, Fe; Kittayapong, Pattamaporn; Abeyewickreme, W; Hapangama, Dilini; Tyagi, Brij Kishore; Htun, Pe Than; Koyadun, Surachart; Kroeger, Axel; Sommerfeld, Johannes; Petzold, Max

2012-01-01

Background Research has shown that the classical Stegomyia indices (or “larval indices”) of the dengue vector Aedes aegypti reflect the absence or presence of the vector but do not provide accurate measures of adult mosquito density. In contrast, pupal indices as collected in pupal productivity surveys are a much better proxy indicator for adult vector abundance. However, it is unknown when it is most optimal to conduct pupal productivity surveys, in the wet or in the dry season or in both, to inform control services about the most productive water container types and if this pattern varies among different ecological settings. Methods A multi-country study in randomly selected twelve to twenty urban and peri-urban neighborhoods (“clusters”) of six Asian countries, in which all water holding containers were examined for larvae and pupae of Aedes aegypti during the dry season and the wet season and their productivity was characterized by water container types. In addition, meteorological data and information on reported dengue cases were collected. Findings The study reconfirmed the association between rainfall and dengue cases (“dengue season”) and underlined the importance of determining through pupal productivity surveys the “most productive containers types”, responsible for the majority (>70%) of adult dengue vectors. The variety of productive container types was greater during the wet than during the dry season, but included practically all container types productive in the dry season. Container types producing pupae were usually different from those infested by larvae indicating that containers with larval infestations do not necessarily foster pupal development and thus the production of adult Aedes mosquitoes. Conclusion Pupal productivity surveys conducted during the wet season will identify almost all of the most productive container types for both the dry and wet seasons and will therefore facilitate cost-effective targeted interventions. PMID:23318235

A novel double fine guide sensor design on space telescope

NASA Astrophysics Data System (ADS)

Zhang, Xu-xu; Yin, Da-yi

2018-02-01

To get high precision attitude for space telescope, a double marginal FOV (field of view) FGS (Fine Guide Sensor) is proposed. It is composed of two large area APS CMOS sensors and both share the same lens in main light of sight. More star vectors can be get by two FGS and be used for high precision attitude determination. To improve star identification speed, the vector cross product in inter-star angles for small marginal FOV different from traditional way is elaborated and parallel processing method is applied to pyramid algorithm. The star vectors from two sensors are then used to attitude fusion with traditional QUEST algorithm. The simulation results show that the system can get high accuracy three axis attitudes and the scheme is feasibility.

Production of recombinant antigens and antibodies in Nicotiana benthamiana using 'magnifection' technology: GMP-compliant facilities for small- and large-scale manufacturing.

PubMed

Klimyuk, Victor; Pogue, Gregory; Herz, Stefan; Butler, John; Haydon, Hugh

2014-01-01

This review describes the adaptation of the plant virus-based transient expression system, magnICON(®) for the at-scale manufacturing of pharmaceutical proteins. The system utilizes so-called "deconstructed" viral vectors that rely on Agrobacterium-mediated systemic delivery into the plant cells for recombinant protein production. The system is also suitable for production of hetero-oligomeric proteins like immunoglobulins. By taking advantage of well established R&D tools for optimizing the expression of protein of interest using this system, product concepts can reach the manufacturing stage in highly competitive time periods. At the manufacturing stage, the system offers many remarkable features including rapid production cycles, high product yield, virtually unlimited scale-up potential, and flexibility for different manufacturing schemes. The magnICON system has been successfully adaptated to very different logistical manufacturing formats: (1) speedy production of multiple small batches of individualized pharmaceuticals proteins (e.g. antigens comprising individualized vaccines to treat NonHodgkin's Lymphoma patients) and (2) large-scale production of other pharmaceutical proteins such as therapeutic antibodies. General descriptions of the prototype GMP-compliant manufacturing processes and facilities for the product formats that are in preclinical and clinical testing are provided.

Gene Overexpression and RNA Silencing Tools for the Genetic Manipulation of the S-(+)-Abscisic Acid Producing Ascomycete Botrytis cinerea

PubMed Central

Ding, Zhong-Tao; Zhang, Zhi; Luo, Di; Zhou, Jin-Yan; Zhong, Juan; Yang, Jie; Xiao, Liang; Shu, Dan; Tan, Hong

2015-01-01

The phytopathogenic ascomycete Botrytis cinerea produces several secondary metabolites that have biotechnical significance and has been particularly used for S-(+)-abscisic acid production at the industrial scale. To manipulate the expression levels of specific secondary metabolite biosynthetic genes of B. cinerea with Agrobacterium tumefaciens-mediated transformation system, two expression vectors (pCBh1 and pCBg1 with different selection markers) and one RNA silencing vector, pCBSilent1, were developed with the In-Fusion assembly method. Both expression vectors were highly effective in constitutively expressing eGFP, and pCBSilent1 effectively silenced the eGFP gene in B. cinerea. Bcaba4, a gene suggested to participate in ABA biosynthesis in B. cinerea, was then targeted for gene overexpression and RNA silencing with these reverse genetic tools. The overexpression of bcaba4 dramatically induced ABA formation in the B. cinerea wild type strain Bc-6, and the gene silencing of bcaba4 significantly reduced ABA-production in an ABA-producing B. cinerea strain. PMID:25955649

Pure state consciousness and its local reduction to neuronal space

NASA Astrophysics Data System (ADS)

Duggins, A. J.

2013-01-01

The single neuronal state can be represented as a vector in a complex space, spanned by an orthonormal basis of integer spike counts. In this model a scalar element of experience is associated with the instantaneous firing rate of a single sensory neuron over repeated stimulus presentations. Here the model is extended to composite neural systems that are tensor products of single neuronal vector spaces. Depiction of the mental state as a vector on this tensor product space is intended to capture the unity of consciousness. The density operator is introduced as its local reduction to the single neuron level, from which the firing rate can again be derived as the objective correlate of a subjective element. However, the relational structure of perceptual experience only emerges when the non-local mental state is considered. A metric of phenomenal proximity between neuronal elements of experience is proposed, based on the cross-correlation function of neurophysiology, but constrained by the association of theoretical extremes of correlation/anticorrelation in inseparable 2-neuron states with identical and opponent elements respectively.

New Constructions of Orthogonal Product Basis Quantum States

NASA Astrophysics Data System (ADS)

Zuo, Huijuan; Liu, Shuxia; Yang, Yinghui

2018-02-01

An orthogonal basis B9 for the Hilbert space C 3 × C 3 was presented by Bennett et al. (Phys. Rev. A 59, 1070, 1999) which was illustrated in a visual figure in their report. The character of the construction is that each base vector is a product state, thus any distinguishing operator cannot create entanglement. In this paper, we mainly focus on some new constructions of orthogonal product basis quantum states in the high-dimensional quantum systems. Especially, as for the quantum system of (2m + 1) ⊗ (2m + 1), where m ∈ Z and m ≥ 2, we have provided the direct construction in mathematical method.

75 FR 51043 - Pesticide Experimental Use Permit; Receipt of Application; Comment Request

Federal Register 2010, 2011, 2012, 2013, 2014

2010-08-18

... (vector pCOT1) in event COT102 cotton, Bt Cry1Ac protein and the genetic material necessary for its production (vector PV-GHBK04) in event MON 15985 cotton, and Bt Cry2Ab2 protein and the genetic material necessary for its production (vector PV-GHBK11) in event MON 15985 cotton. The Agency has determined that...

A Preliminary Examination of the Second Generation CMORPH Real-time Production

NASA Astrophysics Data System (ADS)

Joyce, R.; Xie, P.; Wu, S.

2017-12-01

The second generation CMORPH (CMORPH2) has started test real-time production of 30-minute precipitation estimates on a 0.05olat/lon grid over the entire globe, from pole-to-pole. The CMORPH2 is built upon the Kalman Filter based CMORPH algorithm of Joyce and Xie (2011). Inputs to the system include rainfall and snowfall rate retrievals from passive microwave (PMW) measurements aboard all available low earth orbit (LEO) satellites, precipitation estimates derived from infrared (IR) observations of geostationary (GEO) and LEO platforms, and precipitation simulations from the NCEP operational global forecast system (GFS). Inputs from the various sources are first inter-calibrated to ensure quantitative consistencies in representing precipitation events of different intensities through PDF calibration against a common reference standard. The inter-calibrated PMW retrievals and IR-based precipitation estimates are then propagated from their respective observation times to the target analysis time along the motion vectors of the precipitating clouds. Motion vectors are first derived separately from the satellite IR based precipitation estimates and the GFS precipitation fields. These individually derived motion vectors are then combined through a 2D-VAR technique to form an analyzed field of cloud motion vectors over the entire globe. The propagated PMW and IR based precipitation estimates are finally integrated into a single field of global precipitation through the Kalman Filter framework. A set of procedures have been established to examine the performance of the CMORPH2 real-time production. CMORPH2 satellite precipitation estimates are compared against the CPC daily gauge analysis, Stage IV radar precipitation over the CONUS, and numerical model forecasts to discover potential shortcomings and quantify improvements against the first generation CMORPH. Special attention has been focused on the CMORPH behavior over high-latitude areas beyond the coverage of the first generation CMORPH. Detailed results will be reported at the AGU.

Genetic modification of cells for transplantation.

PubMed

Lai, Yi; Drobinskaya, Irina; Kolossov, Eugen; Chen, Chunguang; Linn, Thomas

2008-01-14

Progress in gene therapy has produced promising results that translate experimental research into clinical treatment. Gene modification has been extensively employed in cell transplantation. The main barrier is an effective gene delivery system. Several viral vectors were utilized in end-stage differentiated cells. Recently, successful applications were described with adenovirus-associated vectors. As an alternative, embryonic stem cell- and stem cell-like systems were established for generation of tissue-specified gene-modified cells. Owing to the feasibility for genetic manipulations and the self-renewing potency of these cells they can be used in a way enabling large-scale in vitro production. This approach offers the establishment of in vitro cell culture systems that will deliver sufficient amounts of highly purified, immunoautologous cells suitable for application in regenerative medicine. In this review, the current technology of gene delivery systems to cells is recapitulated and the latest developments for cell transplantation are discussed.

Energy-Efficient Underwater Surveillance by Means of Hybrid Aquacopters

DTIC Science & Technology

2014-12-01

life-cycle analysis, photovoltaic device maximum power point tracking (MPPT), and surface treatments for antifouling of the solar cells can be...108 3. Power Conversion and Storage...15 Figure 10. Shallow Water Analysis and Forecast System product, displaying regional ocean current vectors overlaying a sea surface

The Helioseismic and Magnetic Imager (HMI) Vector Magnetic Field Pipeline: SHARPs - Space-Weather HMI Active Region Patches

NASA Astrophysics Data System (ADS)

Bobra, M. G.; Sun, X.; Hoeksema, J. T.; Turmon, M.; Liu, Y.; Hayashi, K.; Barnes, G.; Leka, K. D.

2014-09-01

A new data product from the Helioseismic and Magnetic Imager (HMI) onboard the Solar Dynamics Observatory (SDO) called Space-weather HMI Active Region Patches ( SHARPs) is now available. SDO/HMI is the first space-based instrument to map the full-disk photospheric vector magnetic field with high cadence and continuity. The SHARP data series provide maps in patches that encompass automatically tracked magnetic concentrations for their entire lifetime; map quantities include the photospheric vector magnetic field and its uncertainty, along with Doppler velocity, continuum intensity, and line-of-sight magnetic field. Furthermore, keywords in the SHARP data series provide several parameters that concisely characterize the magnetic-field distribution and its deviation from a potential-field configuration. These indices may be useful for active-region event forecasting and for identifying regions of interest. The indices are calculated per patch and are available on a twelve-minute cadence. Quick-look data are available within approximately three hours of observation; definitive science products are produced approximately five weeks later. SHARP data are available at jsoc.stanford.edu and maps are available in either of two different coordinate systems. This article describes the SHARP data products and presents examples of SHARP data and parameters.

Gene Delivery into Plant Cells for Recombinant Protein Production

PubMed Central

Chen, Qiang

2015-01-01

Recombinant proteins are primarily produced from cultures of mammalian, insect, and bacteria cells. In recent years, the development of deconstructed virus-based vectors has allowed plants to become a viable platform for recombinant protein production, with advantages in versatility, speed, cost, scalability, and safety over the current production paradigms. In this paper, we review the recent progress in the methodology of agroinfiltration, a solution to overcome the challenge of transgene delivery into plant cells for large-scale manufacturing of recombinant proteins. General gene delivery methodologies in plants are first summarized, followed by extensive discussion on the application and scalability of each agroinfiltration method. New development of a spray-based agroinfiltration and its application on field-grown plants is highlighted. The discussion of agroinfiltration vectors focuses on their applications for producing complex and heteromultimeric proteins and is updated with the development of bridge vectors. Progress on agroinfiltration in Nicotiana and non-Nicotiana plant hosts is subsequently showcased in context of their applications for producing high-value human biologics and low-cost and high-volume industrial enzymes. These new advancements in agroinfiltration greatly enhance the robustness and scalability of transgene delivery in plants, facilitating the adoption of plant transient expression systems for manufacturing recombinant proteins with a broad range of applications. PMID:26075275

MISR Level 3 Cloud Motion Vector Versioning

Atmospheric Science Data Center

2016-11-04

... Versioning   Cloud Motion Vector Product (CMV) - Monthly, Quarterly, Yearly products Processing Status ... MI3MCMVN, MI3QCMVN, MI3YCMVN MISR_AM1_CMV Stage 1 Validated:  All parameters MISR maturity ...

Earth observation in support of malaria control and epidemiology: MALAREO monitoring approaches.

PubMed

Franke, Jonas; Gebreslasie, Michael; Bauwens, Ides; Deleu, Julie; Siegert, Florian

2015-06-03

Malaria affects about half of the world's population, with the vast majority of cases occuring in Africa. National malaria control programmes aim to reduce the burden of malaria and its negative, socioeconomic effects by using various control strategies (e.g. vector control, environmental management and case tracking). Vector control is the most effective transmission prevention strategy, while environmental factors are the key parameters affecting transmission. Geographic information systems (GIS), earth observation (EO) and spatial modelling are increasingly being recognised as valuable tools for effective management and malaria vector control. Issues previously inhibiting the use of EO in epidemiology and malaria control such as poor satellite sensor performance, high costs and long turnaround times, have since been resolved through modern technology. The core goal of this study was to develop and implement the capabilities of EO data for national malaria control programmes in South Africa, Swaziland and Mozambique. High- and very high resolution (HR and VHR) land cover and wetland maps were generated for the identification of potential vector habitats and human activities, as well as geoinformation on distance to wetlands for malaria risk modelling, population density maps, habitat foci maps and VHR household maps. These products were further used for modelling malaria incidence and the analysis of environmental factors that favour vector breeding. Geoproducts were also transferred to the staff of national malaria control programmes in seven African countries to demonstrate how EO data and GIS can support vector control strategy planning and monitoring. The transferred EO products support better epidemiological understanding of environmental factors related to malaria transmission, and allow for spatio-temporal targeting of malaria control interventions, thereby improving the cost-effectiveness of interventions.

A Generic Protocol for Intracellular Expression of Recombinant Proteins in Bacillus subtilis.

PubMed

Phan, Trang; Huynh, Phuong; Truong, Tuom; Nguyen, Hoang

2017-01-01

Bacillus subtilis (B. subtilis) is a potential and attractive host for the production of recombinant proteins. Different expression systems for B. subtilis have been developed recently, and various target proteins have been recombinantly synthesized and purified using this host. In this chapter, we introduce a generic protocol to express a recombinant protein in B. subtilis. It includes protocols for (1) using our typical expression vector (plasmid pHT254) to introduce a target gene, (2) transformation of the target vector into B. subtilis, and (3) evaluation of the actual expression of a recombinant protein.

Characteristics of Minimally Oversized Adeno-Associated Virus Vectors Encoding Human Factor VIII Generated Using Producer Cell Lines and Triple Transfection.

PubMed

Nambiar, Bindu; Cornell Sookdeo, Cathleen; Berthelette, Patricia; Jackson, Robert; Piraino, Susan; Burnham, Brenda; Nass, Shelley; Souza, David; O'Riordan, Catherine R; Vincent, Karen A; Cheng, Seng H; Armentano, Donna; Kyostio-Moore, Sirkka

2017-02-01

Several ongoing clinical studies are evaluating recombinant adeno-associated virus (rAAV) vectors as gene delivery vehicles for a variety of diseases. However, the production of vectors with genomes >4.7 kb is challenging, with vector preparations frequently containing truncated genomes. To determine whether the generation of oversized rAAVs can be improved using a producer cell-line (PCL) process, HeLaS3-cell lines harboring either a 5.1 or 5.4 kb rAAV vector genome encoding codon-optimized cDNA for human B-domain deleted Factor VIII (FVIII) were isolated. High-producing "masterwells" (MWs), defined as producing >50,000 vg/cell, were identified for each oversized vector. These MWs provided stable vector production for >20 passages. The quality and potency of the AAVrh8R/FVIII-5.1 and AAVrh8R/FVIII-5.4 vectors generated by the PCL method were then compared to those prepared via transient transfection (TXN). Southern and dot blot analyses demonstrated that both production methods resulted in packaging of heterogeneously sized genomes. However, the PCL-derived rAAV vector preparations contained some genomes >4.7 kb, whereas the majority of genomes generated by the TXN method were ≤4.7 kb. The PCL process reduced packaging of non-vector DNA for both the AAVrh8R/FVIII-5.1 and the AAVrh8R/FVIII-5.4 kb vector preparations. Furthermore, more DNA-containing viral particles were obtained for the AAVrh8R/FVIII-5.1 vector. In a mouse model of hemophilia A, animals administered a PCL-derived rAAV vector exhibited twofold higher plasma FVIII activity and increased levels of vector genomes in the liver than mice treated with vector produced via TXN did. Hence, the quality of oversized vectors prepared using the PCL method is greater than that of vectors generated using the TXN process, and importantly this improvement translates to enhanced performance in vivo.

Remote sensing and GIS integration: Towards intelligent imagery within a spatial data infrastructure

NASA Astrophysics Data System (ADS)

Abdelrahim, Mohamed Mahmoud Hosny

2001-11-01

In this research, an "Intelligent Imagery System Prototype" (IISP) was developed. IISP is an integration tool that facilitates the environment for active, direct, and on-the-fly usage of high resolution imagery, internally linked to hidden GIS vector layers, to query the real world phenomena and, consequently, to perform exploratory types of spatial analysis based on a clear/undisturbed image scene. The IISP was designed and implemented using the software components approach to verify the hypothesis that a fully rectified, partially rectified, or even unrectified digital image can be internally linked to a variety of different hidden vector databases/layers covering the end user area of interest, and consequently may be reliably used directly as a base for "on-the-fly" querying of real-world phenomena and for performing exploratory types of spatial analysis. Within IISP, differentially rectified, partially rectified (namely, IKONOS GEOCARTERRA(TM)), and unrectified imagery (namely, scanned aerial photographs and captured video frames) were investigated. The system was designed to handle four types of spatial functions, namely, pointing query, polygon/line-based image query, database query, and buffering. The system was developed using ESRI MapObjects 2.0a as the core spatial component within Visual Basic 6.0. When used to perform the pre-defined spatial queries using different combinations of image and vector data, the IISP provided the same results as those obtained by querying pre-processed vector layers even when the image used was not orthorectified and the vector layers had different parameters. In addition, the real-time pixel location orthorectification technique developed and presented within the IKONOS GEOCARTERRA(TM) case provided a horizontal accuracy (RMSE) of +/- 2.75 metres. This accuracy is very close to the accuracy level obtained when purchasing the orthorectified IKONOS PRECISION products (RMSE of +/- 1.9 metre). The latter cost approximately four times as much as the IKONOS GEOCARTERRA(TM) products. The developed IISP is a step closer towards the direct and active involvement of high-resolution remote sensing imagery in querying the real world and performing exploratory types of spatial analysis. (Abstract shortened by UMI.)

Definition of Contravariant Velocity Components

NASA Technical Reports Server (NTRS)

Hung, Ching-moa; Kwak, Dochan (Technical Monitor)

2002-01-01

In this paper we have reviewed the basics of tensor analysis in an attempt to clarify some misconceptions regarding contravariant and covariant vector components as used in fluid dynamics. We have indicated that contravariant components are components of a given vector expressed as a unique combination of the covariant base vector system and, vice versa, that the covariant components are components of a vector expressed with the contravariant base vector system. Mathematically, expressing a vector with a combination of base vector is a decomposition process for a specific base vector system. Hence, the contravariant velocity components are decomposed components of velocity vector along the directions of coordinate lines, with respect to the covariant base vector system. However, the contravariant (and covariant) components are not physical quantities. Their magnitudes and dimensions are controlled by their corresponding covariant (and contravariant) base vectors.

Product Quality Modelling Based on Incremental Support Vector Machine

NASA Astrophysics Data System (ADS)

Wang, J.; Zhang, W.; Qin, B.; Shi, W.

2012-05-01

Incremental Support vector machine (ISVM) is a new learning method developed in recent years based on the foundations of statistical learning theory. It is suitable for the problem of sequentially arriving field data and has been widely used for product quality prediction and production process optimization. However, the traditional ISVM learning does not consider the quality of the incremental data which may contain noise and redundant data; it will affect the learning speed and accuracy to a great extent. In order to improve SVM training speed and accuracy, a modified incremental support vector machine (MISVM) is proposed in this paper. Firstly, the margin vectors are extracted according to the Karush-Kuhn-Tucker (KKT) condition; then the distance from the margin vectors to the final decision hyperplane is calculated to evaluate the importance of margin vectors, where the margin vectors are removed while their distance exceed the specified value; finally, the original SVs and remaining margin vectors are used to update the SVM. The proposed MISVM can not only eliminate the unimportant samples such as noise samples, but also can preserve the important samples. The MISVM has been experimented on two public data and one field data of zinc coating weight in strip hot-dip galvanizing, and the results shows that the proposed method can improve the prediction accuracy and the training speed effectively. Furthermore, it can provide the necessary decision supports and analysis tools for auto control of product quality, and also can extend to other process industries, such as chemical process and manufacturing process.

Opportunities for Improved Chagas Disease Vector Control Based on Knowledge, Attitudes and Practices of Communities in the Yucatan Peninsula, Mexico

PubMed Central

Rosecrans, Kathryn; Cruz-Martin, Gabriela; King, Ashley; Dumonteil, Eric

2014-01-01

Background Chagas disease is a vector-borne parasitic disease of major public health importance. Current prevention efforts are based on triatomine vector control to reduce transmission to humans. Success of vector control interventions depends on their acceptability and value to affected communities. We aimed to identify opportunities for and barriers to improved vector control strategies in the Yucatan peninsula, Mexico. Methodology/principal findings We employed a sequence of qualitative and quantitative research methods to investigate knowledge, attitudes and practices surrounding Chagas disease, triatomines and vector control in three rural communities. Our combined data show that community members are well aware of triatomines and are knowledgeable about their habits. However, most have a limited understanding of the transmission dynamics and clinical manifestations of Chagas disease. While triatomine control is not a priority for community members, they frequently use domestic insecticide products including insecticide spray, mosquito coils and plug-in repellents. Families spend about $32 US per year on these products. Alternative methods such as yard cleaning and window screens are perceived as desirable and potentially more effective. Screens are nonetheless described as unaffordable, in spite of a cost comparable to the average annual spending on insecticide products. Conclusion/Significance Further education campaigns and possibly financing schemes may lead families to redirect their current vector control spending from insecticide products to window screens. Also, synergism with mosquito control efforts should be further explored to motivate community involvement and ensure sustainability of Chagas disease vector control. PMID:24676038

Mineral Replacement Reactions as a Precursor to Strain Localisation: an (HR-)EBSD approach

NASA Astrophysics Data System (ADS)

Gardner, J.; Wheeler, J.; Wallis, D.; Hansen, L. N.; Mariani, E.

2017-12-01

Much remains to be learned about the links between metamorphism and deformation. Our work investigates the behaviour of fluid-mediated mineral replacement reaction products when exposed to subsequent shear stresses. We focus on albite from a metagabbro that has experienced metamorphism and subsequent deformation at greenschist facies, resulting in a reduction in grain size and associated strain localisation. EBSD maps show that prior to grain size reduction, product grains are highly distorted, yet they formed, and subsequently deformed, at temperatures at which extensive dislocation creep is unlikely. The Weighted Burgers Vector can be used to quantitatively describe the types of Burgers vectors present in geometrically necessary dislocation (GND) populations derived from 2-D EBSD map data. Application of this technique to the distorted product grains reveals the prominence of, among others, dislocations with apparent [010] Burgers vectors. This supports (with some caveats) the idea that dislocation creep is not responsible for the observed lattice distortion, as there are no known slip systems in plagioclase with a [010] Burgers vector. Distortion in a replacement microstructure has also been attributed to the presence of nanoscale product grains, which share very similar, but not identical, orientations due to topotactic nucleation from adjacent sites on the same substrate. As a precipitate, the product grains should be expected to be largely free of elastic strain. However, high angular resolution EBSD results demonstrate that product grains contain both elastic strains (> 10-3) and residual stresses (several hundred MPa), as well as GND densities on the order of 1014-1015 m-2. Thus we suggest the observed distortion (elastic strain plus rotations) in the lattice is produced during the mineral replacement reaction by a lattice mismatch and volume change between parent and product. Stored strain energy then provides a driving force for recovery and recrystallization. Recrystallization produces smaller grains with high angle boundaries, reducing the strength of, and allowing deformation to localise in, the albite phase. Grain size reduction in turn facilitates shear deformation to high strains by a grain size sensitive mechanism (fluid-assisted diffusion creep).

Manufacture of Third-Generation Lentivirus for Preclinical Use, with Process Development Considerations for Translation to Good Manufacturing Practice.

PubMed

Gándara, Carolina; Affleck, Valerie; Stoll, Elizabeth Ann

2018-02-01

Lentiviral vectors are used in laboratories around the world for in vivo and ex vivo delivery of gene therapies, and increasingly clinical investigation as well as preclinical applications. The third-generation lentiviral vector system has many advantages, including high packaging capacity, stable gene expression in both dividing and post-mitotic cells, and low immunogenicity in the recipient organism. Yet, the manufacture of these vectors is challenging, especially at high titers required for direct use in vivo, and further challenges are presented by the process of translating preclinical gene therapies toward manufacture of products for clinical investigation. The goals of this paper are to report the protocol for manufacturing high-titer third-generation lentivirus for preclinical testing and to provide detailed information on considerations for translating preclinical viral vector manufacture toward scaled-up platforms and processes in order to make gene therapies under Good Manufacturing Practice that are suitable for clinical trials.

Manufacture of Third-Generation Lentivirus for Preclinical Use, with Process Development Considerations for Translation to Good Manufacturing Practice

PubMed Central

Gándara, Carolina; Affleck, Valerie; Stoll, Elizabeth Ann

2018-01-01

Lentiviral vectors are used in laboratories around the world for in vivo and ex vivo delivery of gene therapies, and increasingly clinical investigation as well as preclinical applications. The third-generation lentiviral vector system has many advantages, including high packaging capacity, stable gene expression in both dividing and post-mitotic cells, and low immunogenicity in the recipient organism. Yet, the manufacture of these vectors is challenging, especially at high titers required for direct use in vivo, and further challenges are presented by the process of translating preclinical gene therapies toward manufacture of products for clinical investigation. The goals of this paper are to report the protocol for manufacturing high-titer third-generation lentivirus for preclinical testing and to provide detailed information on considerations for translating preclinical viral vector manufacture toward scaled-up platforms and processes in order to make gene therapies under Good Manufacturing Practice that are suitable for clinical trials. PMID:29212357

Generating an Open Reading Frame (ORF) Entry Clone and Destination Clone.

PubMed

Reece-Hoyes, John S; Walhout, Albertha J M

2018-01-02

This protocol describes using the Gateway recombinatorial cloning system to create an Entry clone carrying an open reading frame (ORF) and then to transfer the ORF into a Destination vector. In this example, BP recombination is used to clone an ORF from a cDNA source into the Donor vector pDONR 221. The ORF from the resulting Entry clone is then transferred into the Destination vector pDEST-15; the product (the Destination clone) will express the ORF as an amino-terminal GST-fusion. The technique can be used as a guide for cloning any other DNA fragment of interest-a promoter sequence or 3' untranslated region (UTR), for example-with substitutions of different genetic material such as genomic DNA, att sites, and vectors as required. The series of constructions and transformations requires 9-15 d, not including time that may be required for sequence confirmation, if desired/necessary. © 2018 Cold Spring Harbor Laboratory Press.

Will integrated surveillance systems for vectors and vector-borne diseases be the future of controlling vector-borne diseases? A practical example from China.

PubMed

Wu, Y; Ling, F; Hou, J; Guo, S; Wang, J; Gong, Z

2016-07-01

Vector-borne diseases are one of the world's major public health threats and annually responsible for 30-50% of deaths reported to the national notifiable disease system in China. To control vector-borne diseases, a unified, effective and economic surveillance system is urgently needed; all of the current surveillance systems in China waste resources and/or information. Here, we review some current surveillance systems and present a concept for an integrated surveillance system combining existing vector and vector-borne disease monitoring systems. The integrated surveillance system has been tested in pilot programmes in China and led to a 21·6% cost saving in rodent-borne disease surveillance. We share some experiences gained from these programmes.

Lentiviral vectors encoding shRNAs efficiently transduce and knockdown LINGO-1 but induce an interferon response and cytotoxicity in CNS neurons

PubMed Central

Hutson, Thomas H.; Foster, Edmund; Dawes, John M.; Hindges, Robert; Yáñez-Muñoz, Rafael J.; Moon, Lawrence D.F.

2017-01-01

Background Knocking down neuronal LINGO-1 using short hairpin RNAs (shRNAs) might enhance axon regeneration in the CNS. Integration-deficient lentiviral vectors have great potential as a therapeutic delivery system for CNS injuries. However, recent studies have revealed that shRNAs can induce an interferon response resulting in off-target effects and cytotoxicity. Methods CNS neurons were transduced with integration-deficient lentiviral vectors in vitro. The transcriptional effect of shRNA expression was analysed using qRT-PCR and northern blots were used to assess shRNA production. Results Integration-deficient lentiviral vectors efficiently transduced CNS neurons and knocked down LINGO-1 mRNA in vitro. However, an increase in cell death was observed when lentiviral vectors encoding an shRNA were applied or when high vector concentrations were used. We demonstrate that high doses of vector or the use of vectors encoding shRNAs can induce an up-regulation of interferon stimulated genes (OAS1 and PKR) and a down-regulation of off- target genes (including p75NTR and NgR1). Furthermore, the northern blot demonstrated that these negative consequences occur even when lentiviral vectors express low levels of shRNAs. Together, these results may explain why neurite outgrowth was not enhanced on an inhibitory substrate after transduction with lentiviral vectors encoding an shRNA targeting LINGO-1. Conclusions These findings highlight the importance of including appropriate controls to verify silencing specificity and the requirement to check for an interferon response when conducting RNA interference experiments. However, the potential benefits that RNA interference and viral vectors offer to gene-based therapies to CNS injuries cannot be overlooked and demand further investigation. PMID:22499506

Organic livestock production in Uganda: potentials, challenges and prospects.

PubMed

Nalubwama, Sylvia Muwanga; Mugisha, Anthony; Vaarst, Mette

2011-04-01

Development in organic farming has been stimulated by farmers and consumers becoming interested in healthy food products and sustainable environment. Organic agriculture is a holistic production management system which is based on the principles of health, ecology, care, and fairness. Organic development in Uganda has focused more on the crop sector than livestock sector and has primarily involved the private sector, like organic products export companies and non-governmental organizations. Agriculture in Uganda and many African countries is predominantly traditional, less mechanized, and is usually associated with minimum use of chemical fertilizers, pesticides, and drugs. This low external input agriculture also referred to as "organic by default" can create basis for organic farming where agroecological methods are introduced and present an alternative in terms of intensification to the current low-input/low-output systems. Traditional farming should not be confused with organic farming because in some cases, the existing traditional practices have consequences like overstocking and less attention to soil improvement as well as to animal health and welfare, which is contrary to organic principles of ecology, fairness, health, and care. Challenges of implementing sustainable organic practices in the Ugandan livestock sector threaten its future development, such as vectors and vector-borne diseases, organic feed insufficiency, limited education, research, and support to organic livestock production. The prospects of organic livestock development in Uganda can be enhanced with more scientific research in organic livestock production under local conditions and strengthening institutional support.

Non-viral delivery systems for CRISPR/Cas9-based genome editing: Challenges and opportunities.

PubMed

Li, Ling; Hu, Shuo; Chen, Xiaoyuan

2018-07-01

In recent years, CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) genome editing systems have become one of the most robust platforms in basic biomedical research and therapeutic applications. To date, efficient in vivo delivery of the CRISPR/Cas9 system to the targeted cells remains a challenge. Although viral vectors have been widely used in the delivery of the CRISPR/Cas9 system in vitro and in vivo, their fundamental shortcomings, such as the risk of carcinogenesis, limited insertion size, immune responses and difficulty in large-scale production, severely limit their further applications. Alternative non-viral delivery systems for CRISPR/Cas9 are urgently needed. With the rapid development of non-viral vectors, lipid- or polymer-based nanocarriers have shown great potential for CRISPR/Cas9 delivery. In this review, we analyze the pros and cons of delivering CRISPR/Cas9 systems in the form of plasmid, mRNA, or protein and then discuss the limitations and challenges of CRISPR/Cas9-based genome editing. Furthermore, current non-viral vectors that have been applied for CRISPR/Cas9 delivery in vitro and in vivo are outlined in details. Finally, critical obstacles for non-viral delivery of CRISPR/Cas9 system are highlighted and promising strategies to overcome these barriers are proposed. Published by Elsevier Ltd.

Vector-borne viruses of pulse crops, with a particular emphasis on North American cropping systems

USDA-ARS?s Scientific Manuscript database

Pulse crop production in the USA has increased dramatically over the past decade, in part due to their nutritional value and ability to form symbiotic associations with rhizobacteria that fix atmospheric nitrogen. There are several insect-transmitted viruses that are prevalent and periodically quite...

Maize transformation technology development for commercial event generation.

PubMed

Que, Qiudeng; Elumalai, Sivamani; Li, Xianggan; Zhong, Heng; Nalapalli, Samson; Schweiner, Michael; Fei, Xiaoyin; Nuccio, Michael; Kelliher, Timothy; Gu, Weining; Chen, Zhongying; Chilton, Mary-Dell M

2014-01-01

Maize is an important food and feed crop in many countries. It is also one of the most important target crops for the application of biotechnology. Currently, there are more biotech traits available on the market in maize than in any other crop. Generation of transgenic events is a crucial step in the development of biotech traits. For commercial applications, a high throughput transformation system producing a large number of high quality events in an elite genetic background is highly desirable. There has been tremendous progress in Agrobacterium-mediated maize transformation since the publication of the Ishida et al. (1996) paper and the technology has been widely adopted for transgenic event production by many labs around the world. We will review general efforts in establishing efficient maize transformation technologies useful for transgenic event production in trait research and development. The review will also discuss transformation systems used for generating commercial maize trait events currently on the market. As the number of traits is increasing steadily and two or more modes of action are used to control key pests, new tools are needed to efficiently transform vectors containing multiple trait genes. We will review general guidelines for assembling binary vectors for commercial transformation. Approaches to increase transformation efficiency and gene expression of large gene stack vectors will be discussed. Finally, recent studies of targeted genome modification and transgene insertion using different site-directed nuclease technologies will be reviewed.

Convection-enhanced delivery of AAV vector in parkinsonian monkeys; in vivo detection of gene expression and restoration of dopaminergic function using pro-drug approach.

PubMed

Bankiewicz, K S; Eberling, J L; Kohutnicka, M; Jagust, W; Pivirotto, P; Bringas, J; Cunningham, J; Budinger, T F; Harvey-White, J

2000-07-01

Using an approach that combines gene therapy with aromatic l-amino acid decarboxylase (AADC) gene and a pro-drug (l-dopa), dopamine, the neurotransmitter involved in Parkinson's disease, can be synthesized and regulated. Striatal neurons infected with the AADC gene by an adeno-associated viral vector can convert peripheral l-dopa to dopamine and may therefore provide a buffer for unmetabolized l-dopa. This approach to treating Parkinson's disease may reduce the need for l-dopa/carbidopa, thus providing a better clinical response with fewer side effects. In addition, the imbalance in dopamine production between the nigrostriatal and mesolimbic dopaminergic systems can be corrected by using AADC gene delivery to the striatum. We have also demonstrated that a fundamental obstacle in the gene therapy approach to the central nervous system, i.e., the ability to deliver viral vectors in sufficient quantities to the whole brain, can be overcome by using convection-enhanced delivery. Finally, this study demonstrates that positron emission tomography and the AADC tracer, 6-[(18)F]fluoro-l-m-tyrosine, can be used to monitor gene therapy in vivo. Our therapeutic approach has the potential to restore dopamine production, even late in the disease process, at levels that can be maintained during continued nigrostriatal degeneration. Copyright 2000 Academic Press.

Maize transformation technology development for commercial event generation

PubMed Central

Que, Qiudeng; Elumalai, Sivamani; Li, Xianggan; Zhong, Heng; Nalapalli, Samson; Schweiner, Michael; Fei, Xiaoyin; Nuccio, Michael; Kelliher, Timothy; Gu, Weining; Chen, Zhongying; Chilton, Mary-Dell M.

2014-01-01

Maize is an important food and feed crop in many countries. It is also one of the most important target crops for the application of biotechnology. Currently, there are more biotech traits available on the market in maize than in any other crop. Generation of transgenic events is a crucial step in the development of biotech traits. For commercial applications, a high throughput transformation system producing a large number of high quality events in an elite genetic background is highly desirable. There has been tremendous progress in Agrobacterium-mediated maize transformation since the publication of the Ishida et al. (1996) paper and the technology has been widely adopted for transgenic event production by many labs around the world. We will review general efforts in establishing efficient maize transformation technologies useful for transgenic event production in trait research and development. The review will also discuss transformation systems used for generating commercial maize trait events currently on the market. As the number of traits is increasing steadily and two or more modes of action are used to control key pests, new tools are needed to efficiently transform vectors containing multiple trait genes. We will review general guidelines for assembling binary vectors for commercial transformation. Approaches to increase transformation efficiency and gene expression of large gene stack vectors will be discussed. Finally, recent studies of targeted genome modification and transgene insertion using different site-directed nuclease technologies will be reviewed. PMID:25140170

Genetic engineering in Actinoplanes sp. SE50/110 - development of an intergeneric conjugation system for the introduction of actinophage-based integrative vectors.

PubMed

Gren, Tetiana; Ortseifen, Vera; Wibberg, Daniel; Schneiker-Bekel, Susanne; Bednarz, Hanna; Niehaus, Karsten; Zemke, Till; Persicke, Marcus; Pühler, Alfred; Kalinowski, Jörn

2016-08-20

The α-glucosidase inhibitor acarbose is used for treatment of diabetes mellitus type II, and is manufactured industrially with overproducing derivatives of Actinoplanes sp. SE50/110, reportedly obtained by conventional mutagenesis. Despite of high industrial significance, only limited information exists regarding acarbose metabolism, function and regulation of these processes, due to the absence of proper genetic engineering methods and tools developed for this strain. Here, a basic toolkit for genetic engineering of Actinoplanes sp. SE50/110 was developed, comprising a standardized protocol for a DNA transfer through Escherichia coli-Actinoplanes intergeneric conjugation and applied for the transfer of ϕC31, ϕBT1 and VWB actinophage-based integrative vectors. Integration sites, occurring once per genome for all vectors, were sequenced and characterized for the first time in Actinoplanes sp. SE50/110. Notably, in case of ϕC31 based vector pSET152, the integration site is highly conserved, while for ϕBT1 and the VWB based vectors pRT801 and pSOK804, respectively, no sequence similarities to those in other bacteria were detected. The studied plasmids were proven to be stable and neutral with respect to strain morphology and acarbose production, enabling future use for genetic manipulations of Actinoplanes sp. SE50/110. To further broaden the spectrum of available tools, a GUS reporter system, based on the pSET152 derived vector, was also established in Actinoplanes sp. SE50/110. Copyright © 2016 Elsevier B.V. All rights reserved.

Study of Equatorial Ionospheric irregularities and Mapping of Electron Density Profiles and Ionograms

DTIC Science & Technology

2012-03-09

equation is a product of a complex basis vector in Jackson and a linear combination of plane wave functions. We convert both the amplitudes and the...wave function arguments from complex scalars to complex vectors . This conversion allows us to separate the electric field vector and the imaginary...magnetic field vector , because exponentials of imaginary scalars convert vectors to imaginary vectors and vice versa, while ex- ponentials of imaginary

Study of The Vector Product using Three Dimensions Vector Card of Engineering in Pathumwan Institute of Technology

NASA Astrophysics Data System (ADS)

Mueanploy, Wannapa

2015-06-01

The objective of this research was to offer the way to improve engineering students in Physics topic of vector product. The sampling of this research was the engineering students at Pathumwan Institute of Technology during the first semester of academic year 2013. 1) Select 120 students by random sampling are asked to fill in a satisfaction questionnaire scale, to select size of three dimensions vector card in order to apply in the classroom. 2) Select 60 students by random sampling to do achievement test and take the test to be used in the classroom. The methods used in analysis of achievement test by the Kuder-Richardson Method (KR- 20). The results show that 12 items of achievement test are appropriate to be applied in the classroom. The achievement test gets Difficulty (P) = 0.40-0.67, Discrimination = 0.33-0.73 and Reliability (r) = 0.70.The experimental in the classroom. 3) Select 60 students by random sampling divide into two groups; group one (the controlled group) with 30 students was chosen to study in the vector product lesson by the regular teaching method. Group two (the experimental group) with 30 students was chosen to learn the vector product lesson with three dimensions vector card. 4) Analyzed data between the controlled group and the experimental group, the result showed that experimental group got higher achievement test than the controlled group significant at .01 level.

Expression of the human blood coagulation protein factor XIIIa in Saccharomyces cerevisiae: dependence of the expression levels from host-vector systems and medium conditions.

PubMed

Bröker, M; Bäuml, O; Göttig, A; Ochs, J; Bodenbenner, M; Amann, E

1991-03-01

The human blood coagulation protein Factor XIIIa (FXIIIa) was expressed in Saccharomyces cerevisiae employing Escherichia coli-yeast shuttle vectors based on a 2-mu plasmid. Several factors affecting high production yield of recombinant FXIIIa were analysed. The use of the regulatable GAL-CYC1 hybrid promoter resulted in higher FXIIIa expression when compared with the constitutive ADCI promoter. Screening for suitable yeast strains for expression of FXIIIa under the transcriptional control of the GAL-CYC1 hybrid promoter revealed a broad spectrum of productivity. No obvious correlation between the expression rate and the genetic markers of the strains could be identified. The medium composition markedly influenced the FXIIIa expression rates. The expression of FXIIIa was strictly regulated by the carbon source. Glucose as the only sugar and energy source repressed the synthesis of FXIIIa, whereas addition of galactose induced FXIIIa expression. Special feeding schemes resulted in a productivity of up to 100 mg FXIIIa/l in shake flasks.

pSW2, a Novel Low-Temperature-Inducible Gene Expression Vector Based on a Filamentous Phage of the Deep-Sea Bacterium Shewanella piezotolerans WP3.

PubMed

Yang, Xin-Wei; Jian, Hua-Hua; Wang, Feng-Ping

2015-08-15

A low-temperature-inducible protein expression vector (pSW2) based on a filamentous phage (SW1) of the deep-sea bacterium Shewanella piezotolerans WP3 was constructed. This vector replicated stably in Escherichia coli and Shewanella species, and its copy number increased at low temperatures. The pSW2 vector can be utilized as a complementation plasmid in WP3, and it can also be used for the production of complex cytochromes with multiple heme groups, which has the potential for application for metal ion recovery or bioremediation. Promoters of low-temperature-inducible genes in WP3 were fused into the vector to construct a series of vectors for enhancing protein expression at low temperature. The maximum green fluorescent protein intensity was obtained when the promoter for the hfq gene was used. The WP3/pSW2 system can efficiently produce a patatin-like protein (PLP) from a metagenomic library that tends to form inclusion bodies in E. coli. The yields of PLP in the soluble fraction were 8.3 mg/liter and 4.7 mg/liter of culture at 4°C and 20°C, respectively. Moreover, the pSW2 vector can be broadly utilized in other Shewanella species, such as S. oneidensis and S. psychrophila. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Dissociative Recombination Chemistry and Plasma Dynamics

DTIC Science & Technology

2008-06-16

the fractional square of product momentum with product momentum vectors . Qx and Qy denote the degenerate bend two-body dissociation limits...with product momentum vectors . Qx and Qy denote the degenerate bend normal modes for C2v symmetry H2D and HD2 isomers of H3. symmetry for the Qx...heavy (D atom) products in general receive a greater partitioning of energy than the light product. This may have important implications for

The application of vector concepts on two skew lines

NASA Astrophysics Data System (ADS)

Alghadari, F.; Turmudi; Herman, T.

2018-01-01

The purpose of this study is knowing how to apply vector concepts on two skew lines in three-dimensional (3D) coordinate and its utilization. Several mathematical concepts have a related function for the other, but the related between the concept of vector and 3D have not applied in learning classroom. In fact, there are studies show that female students have difficulties in learning of 3D than male. It is because of personal spatial intelligence. The relevance of vector concepts creates both learning achievement and mathematical ability of male and female students enables to be balanced. The distance like on a cube, cuboid, or pyramid whose are drawn on the rectangular coordinates of a point in space. Two coordinate points of the lines can be created a vector. The vector of two skew lines has the shortest distance and the angle. Calculating of the shortest distance is started to create two vectors as a representation of line by vector position concept, next to determining a norm-vector of two vector which was obtained by cross-product, and then to create a vector from two combination of pair-points which was passed by two skew line, the shortest distance is scalar orthogonal projection of norm-vector on a vector which is a combination of pair-points. While calculating the angle are used two vectors as a representation of line to dot-product, and the inverse of cosine is yield. The utilization of its application on mathematics learning and orthographic projection method.

Naturally enveloped AAV vectors for shielding neutralizing antibodies and robust gene delivery in vivo

PubMed Central

György, Bence; Fitzpatrick, Zachary; Crommentuijn, Matheus HW; Mu, Dakai; Maguire, Casey A.

2014-01-01

Recently adeno-associated virus (AAV) became the first clinically approved gene therapy product in the western world. To develop AAV for future clinical application in a widespread patient base, particularly in therapies which require intravenous (i.v.) administration of vector, the virus must be able to evade pre-existing antibodies to the wild type virus. Here we demonstrate that in mice, AAV vectors associated with extracellular vesicles (EVs) can evade human anti-AAV neutralizing antibodies. We observed different antibody evasion and gene transfer abilities with populations of EVs isolated by different centrifugal forces. EV-associated AAV vector (ev-AAV) was up to 136-fold more resistant over a range of neutralizing antibody concentrations relative to standard AAV vector in vitro. Importantly in mice, at a concentration of passively transferred human antibodies which decreased i.v. administered standard AAV transduction of brain by 80%, transduction of ev-AAV transduction was not reduced and was 4,000-fold higher. Finally, we show that expressing a brain targeting peptide on the EV surface allowed significant enhancement of transduction compared to untargeted ev-AAV. Using ev-AAV represents an effective, clinically relevant approach to evade human neutralizing anti-AAV antibodies after systemic administration of vector. PMID:24917028

Cyanobacterium sp. host cell and vector for production of chemical compounds in cyanobacterial cultures

DOEpatents

Piven, Irina; Friedrich, Alexandra; Duhring, Ulf; Uliczka, Frank; Baier, Kerstin; Inaba, Masami; Shi, Tuo; Wang, Kui; Enke, Heike; Kramer, Dan

2014-09-30

A cyanobacterial host cell, Cyanobacterium sp., that harbors at least one recombinant gene for the production of a chemical compounds is provided, as well as vectors derived from an endogenous plasmid isolated from the cell.

Cyanobacterium sp. host cell and vector for production of chemical compounds in Cyanobacterial cultures

DOEpatents

Piven, Irina; Friedrich, Alexandra; Duhring, Ulf; Uliczka, Frank; Baier, Kerstin; Inaba, Masami; Shi, Tuo; Wang, Kui; Enke, Heike; Kramer, Dan

2016-04-19

A cyanobacterial host cell, Cyanobacterium sp., that harbors at least one recombinant gene for the production of a chemical compounds is provided, as well as vectors derived from an endogenous plasmid isolated from the cell.

Cloning strategy for producing brush-forming protein-based polymers.

PubMed

Henderson, Douglas B; Davis, Richey M; Ducker, William A; Van Cott, Kevin E

2005-01-01

Brush-forming polymers are being used in a variety of applications, and by using recombinant DNA technology, there exists the potential to produce protein-based polymers that incorporate unique structures and functions in these brush layers. Despite this potential, production of protein-based brush-forming polymers is not routinely performed. For the design and production of new protein-based polymers with optimal brush-forming properties, it would be desirable to have a cloning strategy that allows an iterative approach wherein the protein based-polymer product can be produced and evaluated, and then if necessary, it can be sequentially modified in a controlled manner to obtain optimal surface density and brush extension. In this work, we report on the development of a cloning strategy intended for the production of protein-based brush-forming polymers. This strategy is based on the assembly of modules of DNA that encode for blocks of protein-based polymers into a commercially available expression vector; there is no need for custom-modified vectors and no need for intermediate cloning vectors. Additionally, because the design of new protein-based biopolymers can be an iterative process, our method enables sequential modification of a protein-based polymer product. With at least 21 bacterial expression vectors and 11 yeast expression vectors compatible with this strategy, there are a number of options available for production of protein-based polymers. It is our intent that this strategy will aid in advancing the production of protein-based brush-forming polymers.

An Elementary Treatment of General Inner Products

ERIC Educational Resources Information Center

Graver, Jack E.

2011-01-01

A typical first course on linear algebra is usually restricted to vector spaces over the real numbers and the usual positive-definite inner product. Hence, the proof that dim(S)+ dim(S[perpendicular]) = dim("V") is not presented in a way that is generalizable to non-positive?definite inner products or to vector spaces over other fields. In this…

Engineering Software Suite Validates System Design

NASA Technical Reports Server (NTRS)

2007-01-01

EDAptive Computing Inc.'s (ECI) EDAstar engineering software tool suite, created to capture and validate system design requirements, was significantly funded by NASA's Ames Research Center through five Small Business Innovation Research (SBIR) contracts. These programs specifically developed Syscape, used to capture executable specifications of multi-disciplinary systems, and VectorGen, used to automatically generate tests to ensure system implementations meet specifications. According to the company, the VectorGen tests considerably reduce the time and effort required to validate implementation of components, thereby ensuring their safe and reliable operation. EDASHIELD, an additional product offering from ECI, can be used to diagnose, predict, and correct errors after a system has been deployed using EDASTAR -created models. Initial commercialization for EDASTAR included application by a large prime contractor in a military setting, and customers include various branches within the U.S. Department of Defense, industry giants like the Lockheed Martin Corporation, Science Applications International Corporation, and Ball Aerospace and Technologies Corporation, as well as NASA's Langley and Glenn Research Centers

Estimating chlorophyll with thermal and broadband multispectral high resolution imagery from an unmanned aerial system using relevance vector machines for precision agriculture

NASA Astrophysics Data System (ADS)

Elarab, Manal; Ticlavilca, Andres M.; Torres-Rua, Alfonso F.; Maslova, Inga; McKee, Mac

2015-12-01

Precision agriculture requires high-resolution information to enable greater precision in the management of inputs to production. Actionable information about crop and field status must be acquired at high spatial resolution and at a temporal frequency appropriate for timely responses. In this study, high spatial resolution imagery was obtained through the use of a small, unmanned aerial system called AggieAirTM. Simultaneously with the AggieAir flights, intensive ground sampling for plant chlorophyll was conducted at precisely determined locations. This study reports the application of a relevance vector machine coupled with cross validation and backward elimination to a dataset composed of reflectance from high-resolution multi-spectral imagery (VIS-NIR), thermal infrared imagery, and vegetative indices, in conjunction with in situ SPAD measurements from which chlorophyll concentrations were derived, to estimate chlorophyll concentration from remotely sensed data at 15-cm resolution. The results indicate that a relevance vector machine with a thin plate spline kernel type and kernel width of 5.4, having LAI, NDVI, thermal and red bands as the selected set of inputs, can be used to spatially estimate chlorophyll concentration with a root-mean-squared-error of 5.31 μg cm-2, efficiency of 0.76, and 9 relevance vectors.

A rapid generation of adenovirus vector with a genetic modification in hexon protein.

PubMed

Di, Bingyan; Mao, Qinwen; Zhao, Junli; Li, Xing; Wang, Dongyang; Xia, Haibin

2012-02-10

The generation of hexon-modified adenovirus vector has proven difficult. In this paper, we developed a novel method for rapid generation of hexon-modified adenoviral vector via one step ligation in vitro followed by quick white/blue color screening. The new system has the following features. First, eGFP expression driven by the CMV promoter in E1 region functions as a reporter to evaluate the tropism of hexon-modified adenovirus in vitro. Second, it has two unique restriction enzyme sites with sticky ends located in the hexon HVR5 region. Third, a lacZ expression cassette under the control of plac promoter is placed between the two restriction enzyme sites, which allows recombinants to be selected using blue/white screening. To prove the principle of the method, genetically modified adenoviruses were successfully produced by insertion of NGR, RGD or Tat PTD peptide into hexon HVR5. Furthermore, the transduction efficiency of the Tat PTD modified virus was shown to be a significant enhancement in A172 and CHO-K1 cells. In conclusion, the novel system makes the production of truly retargeted vectors more promising, which would be of substantial benefit for cancer gene therapy. Copyright © 2012 Elsevier B.V. All rights reserved.

Forces Associated with Nonlinear Nonholonomic Constraint Equations

NASA Technical Reports Server (NTRS)

Roithmayr, Carlos M.; Hodges, Dewey H.

2010-01-01

A concise method has been formulated for identifying a set of forces needed to constrain the behavior of a mechanical system, modeled as a set of particles and rigid bodies, when it is subject to motion constraints described by nonholonomic equations that are inherently nonlinear in velocity. An expression in vector form is obtained for each force; a direction is determined, together with the point of application. This result is a consequence of expressing constraint equations in terms of dot products of vectors rather than in the usual way, which is entirely in terms of scalars and matrices. The constraint forces in vector form are used together with two new analytical approaches for deriving equations governing motion of a system subject to such constraints. If constraint forces are of interest they can be brought into evidence in explicit dynamical equations by employing the well-known nonholonomic partial velocities associated with Kane's method; if they are not of interest, equations can be formed instead with the aid of vectors introduced here as nonholonomic partial accelerations. When the analyst requires only the latter, smaller set of equations, they can be formed directly; it is not necessary to expend the labor to form the former, larger set first and subsequently perform matrix multiplications.

[Public health pests. Arthropods and rodents as causative disease agents as well as reservoirs and vectors of pathogens].

PubMed

Faulde, M; Freise, J

2014-05-01

Globally, infectious diseases pose the most important cause of death. Among known human pathogenic diseases, approximately 50 % are zoonoses. When considering emerging infectious diseases separately 73 % currently belong to the group of zoonoses. In Central Europe, hard ticks show by far the biggest potential as vectors of agents of human disease. Lyme borreliosis, showing an estimated annual incidence between 60,000 and 214,000 cases is by far the most frequent tick-borne disease in Germany. Continually, formerly unknown disease agents could be discovered in endemic vector species. Additionally, introduction of new arthropod vectors and/or agents of disease occur constantly. Recently, five mosquito species of the genus Aedes have been newly introduced to Europe where they are currently spreading in different regions. Uncommon autochthonous transmission of dengue and chikungunya fever viruses in Southern Europe could be directly linked to these vector species and of these Ae. albopictus and Ae. japonicus are currently reported to occur in Germany. The German Protection against Infection Act only covers the control of public health pests which are either active hematophagous vectors or mechanical transmitters of agents of diseases. Use of officially recommended biocidal products aiming to interrupt transmission cycles of vector-borne diseases, is confined to infested buildings only, including sewage systems in the case of Norway rat control. Outdoor vectors, such as hard ticks and mosquitoes, are currently not taken into consideration. Additionally, adjustments of national public health regulations, detailed arthropod vector and rodent reservoir mapping, including surveillance of vector-borne disease agents, are necessary in order to mitigate future disease risks.

Kinematic sensitivity of robot manipulators

NASA Technical Reports Server (NTRS)

Vuskovic, Marko I.

1989-01-01

Kinematic sensitivity vectors and matrices for open-loop, n degrees-of-freedom manipulators are derived. First-order sensitivity vectors are defined as partial derivatives of the manipulator's position and orientation with respect to its geometrical parameters. The four-parameter kinematic model is considered, as well as the five-parameter model in case of nominally parallel joint axes. Sensitivity vectors are expressed in terms of coordinate axes of manipulator frames. Second-order sensitivity vectors, the partial derivatives of first-order sensitivity vectors, are also considered. It is shown that second-order sensitivity vectors can be expressed as vector products of the first-order sensitivity vectors.

Novel strategies to construct complex synthetic vectors to produce DNA molecular weight standards.

PubMed

Chen, Zhe; Wu, Jianbing; Li, Xiaojuan; Ye, Chunjiang; Wenxing, He

2009-05-01

DNA molecular weight standards (DNA markers, nucleic acid ladders) are commonly used in molecular biology laboratories as references to estimate the size of various DNA samples in electrophoresis process. One method of DNA marker production is digestion of synthetic vectors harboring multiple DNA fragments of known sizes by restriction enzymes. In this article, we described three novel strategies-sequential DNA fragment ligation, screening of ligation products by polymerase chain reaction (PCR) with end primers, and "small fragment accumulation"-for constructing complex synthetic vectors and minimizing the mass differences between DNA fragments produced from restrictive digestion of synthetic vectors. The strategy could be applied to construct various complex synthetic vectors to produce any type of low-range DNA markers, usually available commercially. In addition, the strategy is useful for single-step ligation of multiple DNA fragments for construction of complex synthetic vectors and other applications in molecular biology field.

Wavemill Product Assessment- Defining Products and Evaluating Potential Performance from a Novel Spaceborne Interferometric SAR

NASA Astrophysics Data System (ADS)

Cotton, P. D.; Gommenginger, C.; Martin, A.; Marquez, J.; Burbidge, G.; Quilfen, Y.; Chapron, B.; Reppucci, A.; Buck, C.

2016-08-01

Ocean Surface Currents are one of the most important ocean properties for oceanographers and operators in the maritime domain. Improved monitoring of ocean currents is systematically the number one requirement that emerges from any science or end user requirement surveys.Wavemill is a novel hybrid interferometric SAR system first proposed by ESA/ESTEC [Buck, 2005]. It offers the possibility of generating two-dimensional wide swath, high resolution, high precision maps of surface current vectors and ocean topography [Buck et al., 2009]. Based on a single spacecraft, it avoids the difficulties of synchronisation and baseline estimation associated with other interferometric SAR systems based on two or more satellites (e.g. the "cartwheel" or "helix" concept).The Wavemill concept has developed steadily since its first inception in 2005. A number of Wavemill studies in recent years have gradually put together facts and figures to support the case for Wavemill as a possible space-borne mission.The Wavemill Product Assessment study (WaPA) aimed to define the scientific capabilities and limitations of a spaceborne Wavemill instrument in preparation for a possible submission of the Wavemill concept as a candidate Earth Explorer Core mission. The WaPA project team brought together expert scientists and engineers in the field of SAR imaging of ocean currents, and included the National Oceanography Centre (UK), Starlab (Spain), IFREMER (France) and Airbus Defence and Space (UK). Overall project management was provided by Satellite Oceanographic Consultants (UK). The approach taken included:- A review of SAR imaging of ocean currents in along-track interferometric mode to learn from previous experiments and modelling what key phenomena need to be accounted for to determine the true performance of a spaceborne Wavemill system- Validation of proposed Wavemill primary products based on Wavemill airborne proof-of-concept data and numerical simulations to determine the capabilities and limitations of a spaceborne Wavemill instrument for ocean current vector and sea surface height mapping.- An analysis of the potential for ocean wind vector retrieval from a spaceborne Wavemill instrument.- An investigation of possible secondary products from Wavemill relating to rivers, ocean/atmosphere interactions, ocean swell and cryospheric applications.An assessment of the synergy between Wavemill and ocean surface current products derived from other remote sensing techniques, accounting for the nature and variability of the measured properties, to identify any additional requirements on a future Wavemill mission.

MISR Level 3 Cloud Motion Vector

Atmospheric Science Data Center

2013-07-10

... 2012 A new version, F02_0002, of the MISR L3 CMV (Cloud Motion Vector) data product is now available. This new release ... CMV products for the full mission time period of March 2000 - September 2012 are now available for ordering.   Information ...

9 CFR 122.1 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Definitions. 122.1 Section 122.1..., SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS ORGANISMS AND VECTORS § 122.1 Definitions... regulations. (g) Person. Any individual, firm, partnership, corporation, company, society, association, or...

(abstract) Satellite Physical Oceanography Data Available From an EOSDIS Archive

NASA Technical Reports Server (NTRS)

Digby, Susan A.; Collins, Donald J.

1996-01-01

The Physical Oceanography Distributed Active Archive Center (PO.DAAC) at the Jet Propulsion Laboratory archives and distributes data as part of the Earth Observing System Data and Information System (EOSDIS). Products available from JPL are largely satellite derived and include sea-surface height, surface-wind speed and vectors, integrated water vapor, atmospheric liquid water, sea-surface temperature, heat flux, and in-situ data as it pertains to satellite data. Much of the data is global and spans fourteen years.There is email access, a WWW site, product catalogs, and FTP capabilities. Data is free of charge.

Definition of Contravariant Velocity Components

NASA Technical Reports Server (NTRS)

Hung, Ching-Mao; Kwak, Dochan (Technical Monitor)

2002-01-01

This is an old issue in computational fluid dynamics (CFD). What is the so-called contravariant velocity or contravariant velocity component? In the article, we review the basics of tensor analysis and give the contravariant velocity component a rigorous explanation. For a given coordinate system, there exist two uniquely determined sets of base vector systems - one is the covariant and another is the contravariant base vector system. The two base vector systems are reciprocal. The so-called contravariant velocity component is really the contravariant component of a velocity vector for a time-independent coordinate system, or the contravariant component of a relative velocity between fluid and coordinates, for a time-dependent coordinate system. The contravariant velocity components are not physical quantities of the velocity vector. Their magnitudes, dimensions, and associated directions are controlled by their corresponding covariant base vectors. Several 2-D (two-dimensional) linear examples and 2-D mass-conservation equation are used to illustrate the details of expressing a vector with respect to the covariant and contravariant base vector systems, respectively.

Bacteriophage Mediates Efficient Gene Transfer in Combination with Conventional Transfection Reagents

PubMed Central

Donnelly, Amanda; Yata, Teerapong; Bentayebi, Kaoutar; Suwan, Keittisak; Hajitou, Amin

2015-01-01

The development of commercially available transfection reagents for gene transfer applications has revolutionized the field of molecular biology and scientific research. However, the challenge remains in ensuring that they are efficient, safe, reproducible and cost effective. Bacteriophage (phage)-based viral vectors have the potential to be utilized for general gene transfer applications within research and industry. Yet, they require adaptations in order to enable them to efficiently enter cells and overcome mammalian cellular barriers, as they infect bacteria only; furthermore, limited progress has been made at increasing their efficiency. The production of a novel hybrid nanocomplex system consisting of two different nanomaterial systems, phage vectors and conventional transfection reagents, could overcome these limitations. Here we demonstrate that the combination of cationic lipids, cationic polymers or calcium phosphate with M13 bacteriophage-derived vectors, engineered to carry a mammalian transgene cassette, resulted in increased cellular attachment, entry and improved transgene expression in human cells. Moreover, addition of a targeting ligand into the nanocomplex system, through genetic engineering of the phage capsid further increased gene expression and was effective in a stable cell line generation application. Overall, this new hybrid nanocomplex system (i) provides enhanced phage-mediated gene transfer; (ii) is applicable for laboratory transfection processes and (iii) shows promise within industry for large-scale gene transfer applications. PMID:26670247

Bacteriophage Mediates Efficient Gene Transfer in Combination with Conventional Transfection Reagents.

PubMed

Donnelly, Amanda; Yata, Teerapong; Bentayebi, Kaoutar; Suwan, Keittisak; Hajitou, Amin

2015-12-08

The development of commercially available transfection reagents for gene transfer applications has revolutionized the field of molecular biology and scientific research. However, the challenge remains in ensuring that they are efficient, safe, reproducible and cost effective. Bacteriophage (phage)-based viral vectors have the potential to be utilized for general gene transfer applications within research and industry. Yet, they require adaptations in order to enable them to efficiently enter cells and overcome mammalian cellular barriers, as they infect bacteria only; furthermore, limited progress has been made at increasing their efficiency. The production of a novel hybrid nanocomplex system consisting of two different nanomaterial systems, phage vectors and conventional transfection reagents, could overcome these limitations. Here we demonstrate that the combination of cationic lipids, cationic polymers or calcium phosphate with M13 bacteriophage-derived vectors, engineered to carry a mammalian transgene cassette, resulted in increased cellular attachment, entry and improved transgene expression in human cells. Moreover, addition of a targeting ligand into the nanocomplex system, through genetic engineering of the phage capsid further increased gene expression and was effective in a stable cell line generation application. Overall, this new hybrid nanocomplex system (i) provides enhanced phage-mediated gene transfer; (ii) is applicable for laboratory transfection processes and (iii) shows promise within industry for large-scale gene transfer applications.

Virus diseases of peppers (Capsicum spp.) and their control.

PubMed

Kenyon, Lawrence; Kumar, Sanjeet; Tsai, Wen-Shi; Hughes, Jacqueline d'A

2014-01-01

The number of virus species infecting pepper (Capsicum spp.) crops and their incidences has increased considerably over the past 30 years, particularly in tropical and subtropical pepper production systems. This is probably due to a combination of factors, including the expansion and intensification of pepper cultivation in these regions, the increased volume and speed of global trade of fresh produce (including peppers) carrying viruses and vectors to new locations, and perhaps climate change expanding the geographic range suitable for the viruses and vectors. With the increased incidences of diverse virus species comes increased incidences of coinfection with two or more virus species in the same plant. There is then greater chance of synergistic interactions between virus species, increasing symptom severity and weakening host resistance, as well as the opportunity for genetic recombination and component exchange and a possible increase in aggressiveness, virulence, and transmissibility. The main virus groups infecting peppers are transmitted by aphids, whiteflies, or thrips, and a feature of many populations of these vector groups is that they can develop resistance to some of the commonly used insecticides relatively quickly. This, coupled with the increasing concern over the impact of over- or misuse of insecticides on the environment, growers, and consumers, means that there should be less reliance on insecticides to control the vectors of viruses infecting pepper crops. To improve the durability of pepper crop protection measures, there should be a shift away from the broadscale use of insecticides and the use of single, major gene resistance to viruses. Instead, integrated and pragmatic virus control measures should be sought that combine (1) cultural practices that reduce sources of virus inoculum and decrease the rate of spread of viruliferous vectors into the pepper crop, (2) synthetic insecticides, which should be used judiciously and only when the plants are young and most susceptible to infection, (3) appropriate natural products and biocontrol agents to induce resistance in the plants, affect the behavior of the vector insects, or augment the local populations of parasites or predators of the virus vectors, and (4) polygenic resistances against viruses and vector insects with pyramided single-gene virus resistances to improve resistance durability.

Mapping of courses on vector biology and vector-borne diseases systems: time for a worldwide effort.

PubMed

Casas, Jérôme; Lazzari, Claudio; Insausti, Teresita; Launois, Pascal; Fouque, Florence

2016-11-01

Major emergency efforts are being mounted for each vector-borne disease epidemiological crisis anew, while knowledge about the biology of arthropods vectors is dwindling slowly but continuously, as is the number of field entomologists. The discrepancy between the rates of production of knowledge and its use and need for solving crises is widening, in particular due to the highly differing time spans of the two concurrent processes. A worldwide web based search using multiple key words and search engines of onsite and online courses in English, Spanish, Portuguese, French, Italian and German concerned with the biology of vectors identified over 140 courses. They are geographically and thematically scattered, the vast majority of them are on-site, with very few courses using the latest massive open online course (MOOC) powerfulness. Over two third of them is given in English and Western Africa is particularity poorly represented. The taxonomic groups covered are highly unbalanced towards mosquitoes. A worldwide unique portal to guide students of all grades and levels of expertise, in particular those in remote locations, is badly needed. This is the objective a new activity supported by the Special Programme for Research and Training in Tropical Diseases (TDR).

Mapping of courses on vector biology and vector-borne diseases systems: time for a worldwide effort

PubMed Central

Casas, Jérôme; Lazzari, Claudio; Insausti, Teresita; Launois, Pascal; Fouque, Florence

2016-01-01

Major emergency efforts are being mounted for each vector-borne disease epidemiological crisis anew, while knowledge about the biology of arthropods vectors is dwindling slowly but continuously, as is the number of field entomologists. The discrepancy between the rates of production of knowledge and its use and need for solving crises is widening, in particular due to the highly differing time spans of the two concurrent processes. A worldwide web based search using multiple key words and search engines of onsite and online courses in English, Spanish, Portuguese, French, Italian and German concerned with the biology of vectors identified over 140 courses. They are geographically and thematically scattered, the vast majority of them are on-site, with very few courses using the latest massive open online course (MOOC) powerfulness. Over two third of them is given in English and Western Africa is particularity poorly represented. The taxonomic groups covered are highly unbalanced towards mosquitoes. A worldwide unique portal to guide students of all grades and levels of expertise, in particular those in remote locations, is badly needed. This is the objective a new activity supported by the Special Programme for Research and Training in Tropical Diseases (TDR). PMID:27759770

Fast software-based volume rendering using multimedia instructions on PC platforms and its application to virtual endoscopy

NASA Astrophysics Data System (ADS)

Mori, Kensaku; Suenaga, Yasuhito; Toriwaki, Jun-ichiro

2003-05-01

This paper describes a software-based fast volume rendering (VolR) method on a PC platform by using multimedia instructions, such as SIMD instructions, which are currently available in PCs' CPUs. This method achieves fast rendering speed through highly optimizing software rather than an improved rendering algorithm. In volume rendering using a ray casting method, the system requires fast execution of the following processes: (a) interpolation of voxel or color values at sample points, (b) computation of normal vectors (gray-level gradient vectors), (c) calculation of shaded values obtained by dot-products of normal vectors and light source direction vectors, (d) memory access to a huge area, and (e) efficient ray skipping at translucent regions. The proposed software implements these fundamental processes in volume rending by using special instruction sets for multimedia processing. The proposed software can generate virtual endoscopic images of a 3-D volume of 512x512x489 voxel size by volume rendering with perspective projection, specular reflection, and on-the-fly normal vector computation on a conventional PC without any special hardware at thirteen frames per second. Semi-translucent display is also possible.

Photon and vector meson exchanges in the production of light meson pairs and elementary atoms

NASA Astrophysics Data System (ADS)

Gevorkyan, S. R.; Kuraev, E. A.; Volkov, M. K.

2013-01-01

The production of pseudoscalar and scalar meson pairs ππ, ηη, η‧η‧, σσ as well as bound states in high energy γγ collisions are considered. The exchange by a vector particle in the binary process γ + γ → ha + hb with hadronic states ha, hb in fragmentation regions of the initial particle leads to nondecreasing cross sections with increasing energy, that is a priority of peripheral kinematics. Unlike the photon exchange the vector meson exchange needs a reggeization leading to fall with energy growth. Nevertheless, due to the peripheral kinematics beyond very forward production angles the vector meson exchanges dominate over all possible exchanges. The proposed approach allows one to express the matrix elements of the considered processes through impacting factors, which can be calculated in perturbation models like chiral perturbation theory (ChPT) or the Nambu-Jona-Lasinio (NJL) model. In particular cases the impact factors can be determined from relevant γγ sub-processes or the vector meson radiative decay width. The pionium atom production in the collisions of high energy electrons and pions with protons is considered and the relevant cross sections have been estimated.

A method of determining where to target surveillance efforts in heterogeneous epidemiological systems

PubMed Central

van den Bosch, Frank; Gottwald, Timothy R.; Alonso Chavez, Vasthi

2017-01-01

The spread of pathogens into new environments poses a considerable threat to human, animal, and plant health, and by extension, human and animal wellbeing, ecosystem function, and agricultural productivity, worldwide. Early detection through effective surveillance is a key strategy to reduce the risk of their establishment. Whilst it is well established that statistical and economic considerations are of vital importance when planning surveillance efforts, it is also important to consider epidemiological characteristics of the pathogen in question—including heterogeneities within the epidemiological system itself. One of the most pronounced realisations of this heterogeneity is seen in the case of vector-borne pathogens, which spread between ‘hosts’ and ‘vectors’—with each group possessing distinct epidemiological characteristics. As a result, an important question when planning surveillance for emerging vector-borne pathogens is where to place sampling resources in order to detect the pathogen as early as possible. We answer this question by developing a statistical function which describes the probability distributions of the prevalences of infection at first detection in both hosts and vectors. We also show how this method can be adapted in order to maximise the probability of early detection of an emerging pathogen within imposed sample size and/or cost constraints, and demonstrate its application using two simple models of vector-borne citrus pathogens. Under the assumption of a linear cost function, we find that sampling costs are generally minimised when either hosts or vectors, but not both, are sampled. PMID:28846676

V+jets Background and Systematic Uncertainties in Top Quark Analyses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adomeit, Stefanie; Peters, Reinhild Yvonne

2014-12-01

Vector boson production in association with jets is an important process to test perturbative quantum chromodynamics and also a background process in top quark analyses. Measurements on vector boson production in association with light and heavy flavour jets are presented, performed by the D0 and CDF collaborations at the Tevatron as well as the ATLAS and CMS experiments at LHC. Techniques applied in top quark analyses to estimate the vector boson+jets background are also discussed.

Mifepristone-inducible transgene expression in neural progenitor cells in vitro and in vivo

PubMed Central

Hjelm, BE; Grunseich, C; Gowing, G; Avalos, P; Tian, J; Shelley, BC; Mooney, M; Narwani, K; Shi, Y; Svendsen, CN; Wolfe, JH; Fischbeck, KH; Pierson, TM

2016-01-01

Numerous gene and cell therapy strategies are being developed for the treatment of neurodegenerative disorders. Many of these strategies use constitutive expression of therapeutic transgenic proteins, and although functional in animal models of disease, this method is less likely to provide adequate flexibility for delivering therapy to humans. Ligand-inducible gene expression systems may be more appropriate for these conditions, especially within the central nervous system (CNS). Mifepristone’s ability to cross the blood–brain barrier makes it an especially attractive ligand for this purpose. We describe the production of a mifepristone-inducible vector system for regulated expression of transgenes within the CNS. Our inducible system used a lentivirus-based vector platform for the ex vivo production of mifepristone-inducible murine neural progenitor cells that express our transgenes of interest. These cells were processed through a series of selection steps to ensure that the cells exhibited appropriate transgene expression in a dose-dependent and temporally controlled manner with minimal background activity. Inducible cells were then transplanted into the brains of rodents, where they exhibited appropriate mifepristone-inducible expression. These studies detail a strategy for regulated expression in the CNS for use in the development of safe and efficient gene therapy for neurological disorders. PMID:26863047

Engineering HSV-1 vectors for gene therapy.

PubMed

Goins, William F; Huang, Shaohua; Cohen, Justus B; Glorioso, Joseph C

2014-01-01

Virus vectors have been employed as gene transfer vehicles for various preclinical and clinical gene therapy applications, and with the approval of Glybera (alipogene tiparvovec) as the first gene therapy product as a standard medical treatment (Yla-Herttuala, Mol Ther 20: 1831-1832, 2013), gene therapy has reached the status of being a part of standard patient care. Replication-competent herpes simplex virus (HSV) vectors that replicate specifically in actively dividing tumor cells have been used in Phase I-III human trials in patients with glioblastoma multiforme, a fatal form of brain cancer, and in malignant melanoma. In fact, T-VEC (talimogene laherparepvec, formerly known as OncoVex GM-CSF) displayed efficacy in a recent Phase III trial when compared to standard GM-CSF treatment alone (Andtbacka et al. J Clin Oncol 31: sLBA9008, 2013) and may soon become the second FDA-approved gene therapy product used in standard patient care. In addition to the replication-competent oncolytic HSV vectors like T-VEC, replication-defective HSV vectors have been employed in Phase I-II human trials and have been explored as delivery vehicles for disorders such as pain, neuropathy, and other neurodegenerative conditions. Research during the last decade on the development of HSV vectors has resulted in the engineering of recombinant vectors that are totally replication defective, nontoxic, and capable of long-term transgene expression in neurons. This chapter describes methods for the construction of recombinant genomic HSV vectors based on the HSV-1 replication-defective vector backbones, steps in their purification, and their small-scale production for use in cell culture experiments as well as preclinical animal studies.

[Adeno-associated viral vectors: methods for production and purification for gene therapy applications].

PubMed

Mena-Enriquez, Mayra; Flores-Contreras, Lucia; Armendáriz-Borunda, Juan

2012-01-01

Viral vectors based on adeno-associated virus (AAV) are widely used in gene therapy protocols, because they have characteristics that make them valuable for the treatment of genetic and chronic degenerative diseases. AAV2 serotype had been the best characterized to date. However, the AAV vectors developed from other serotypes is of special interest, since they have organ-specific tropism which increases their potential for transgene delivery to target cells for performing their therapeutic effects. This article summarizes AAV generalities, methods for their production and purification. It also discusses the use of these vectors in vitro, in vivo and their application in gene therapy clinical trials.

QCD next-to-leading-order predictions matched to parton showers for vector-like quark models.

PubMed

Fuks, Benjamin; Shao, Hua-Sheng

2017-01-01

Vector-like quarks are featured by a wealth of beyond the Standard Model theories and are consequently an important goal of many LHC searches for new physics. Those searches, as well as most related phenomenological studies, however, rely on predictions evaluated at the leading-order accuracy in QCD and consider well-defined simplified benchmark scenarios. Adopting an effective bottom-up approach, we compute next-to-leading-order predictions for vector-like-quark pair production and single production in association with jets, with a weak or with a Higgs boson in a general new physics setup. We additionally compute vector-like-quark contributions to the production of a pair of Standard Model bosons at the same level of accuracy. For all processes under consideration, we focus both on total cross sections and on differential distributions, most these calculations being performed for the first time in our field. As a result, our work paves the way to precise extraction of experimental limits on vector-like quarks thanks to an accurate control of the shapes of the relevant observables and emphasise the extra handles that could be provided by novel vector-like-quark probes never envisaged so far.

High-capacity 'gutless' adenoviral vectors.

PubMed

Kochanek, S; Schiedner, G; Volpers, C

2001-10-01

Adenoviral vectors are promising gene transfer vehicles for different gene therapy applications. High-capacity adenoviral (HC-Ad) vectors address some of the problems that have been observed with replication-defective, E1-deleted first-generation adenoviral vectors: toxicity and immunogenicity due to viral gene expression and 7 to 8 kb capacity limit for the transport of therapeutic DNA. This review summarizes HC-Ad vector-related publications from the past 18 months that are mainly concerned with vector design/production and in vivo applications in different murine models.

Stability of Lentiviral Vector-Mediated Transgene Expression in the Brain in the Presence of Systemic Antivector Immune Responses

PubMed Central

ABORDO-ADESIDA, EVELYN; FOLLENZI, ANTONIA; BARCIA, CARLOS; SCIASCIA, SANDRA; CASTRO, MARIA G.; NALDINI, LUIGI; LOWENSTEIN, PEDRO R.

2009-01-01

Lentiviral vectors are promising tools for gene therapy in the CNS. It is therefore important to characterize their interactions with the immune system in the CNS. This work characterizes transgene expression and brain inflammation in the presence or absence of immune responses generated after systemic immunization with lentiviral vectors. We characterized transduction with SIN-LV vectors in the CNS. A dose—response curve using SIN-LV-GFP demonstrated detectable transgene expression in the striatum at a dose of 102, and maximum expression at 106, transducing units of lentiviral vector, with minimal increase in inflammatory markers between the lowest and highest dose of vector injected. Our studies demonstrate that injection of a lentiviral vector into the CNS did not cause a measurable inflammatory response. Systemic immunization after CNS injection, with the lentiviral vector expressing the same transgene as a vector injected into the CNS, caused a decrease in transgene expression in the CNS, concomitantly with an infiltration of inflammatory cells into the CNS parenchyma at the injection site. However, peripheral immunization with a lentiviral vector carrying a different transgene did not diminish transgene expression, or cause CNS inflammation. Systemic immunization preceding injection of lentiviral vectors into the CNS determined that preexisting antilentiviral immunity, regardless of the transgene, did not affect transgene expression. Furthermore, we showed that the transgene, but not the virion or vector components, is responsible for providing antigenic epitopes to the activated immune system, on systemic immunization with lentivirus. Low immunogenicity and prolonged transgene expression in the presence of preexisting lentiviral immunity are encouraging data for the future use of lentiviral vectors in CNS gene therapy. In summary, the lentiviral vectors tested induced undetectable activation of innate immune responses, and stimulation of adaptive immune responses against lentiviral vectors was effective in causing a decrease in transgene expression only if the immune response was directed against the transgene. A systemic immune response against vector components alone did not cause brain inflammation, possibly because vector-derived epitopes were not being presented in the CNS. PMID:15960605

A method for rapid production of heteromultimeric protein complexes in plants: assembly of protective bluetongue virus-like particles.

PubMed

Thuenemann, Eva C; Meyers, Ann E; Verwey, Jeanette; Rybicki, Edward P; Lomonossoff, George P

2013-09-01

Plant expression systems based on nonreplicating virus-based vectors can be used for the simultaneous expression of multiple genes within the same cell. They therefore have great potential for the production of heteromultimeric protein complexes. This work describes the efficient plant-based production and assembly of Bluetongue virus-like particles (VLPs), requiring the simultaneous expression of four distinct proteins in varying amounts. Such particles have the potential to serve as a safe and effective vaccine against Bluetongue virus (BTV), which causes high mortality rates in ruminants and thus has a severe effect on the livestock trade. Here, VLPs produced and assembled in Nicotiana benthamiana using the cowpea mosaic virus-based HyperTrans (CPMV-HT) and associated pEAQ plant transient expression vector system were shown to elicit a strong antibody response in sheep. Furthermore, they provided protective immunity against a challenge with a South African BTV-8 field isolate. The results show that transient expression can be used to produce immunologically relevant complex heteromultimeric structures in plants in a matter of days. The results have implications beyond the realm of veterinary vaccines and could be applied to the production of VLPs for human use or the coexpression of multiple enzymes for the manipulation of metabolic pathways. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Cascading biomethane energy systems for sustainable green gas production in a circular economy.

PubMed

Wall, David M; McDonagh, Shane; Murphy, Jerry D

2017-11-01

Biomethane is a flexible energy vector that can be used as a renewable fuel for both the heat and transport sectors. Recent EU legislation encourages the production and use of advanced, third generation biofuels with improved sustainability for future energy systems. The integration of technologies such as anaerobic digestion, gasification, and power to gas, along with advanced feedstocks such as algae will be at the forefront in meeting future sustainability criteria and achieving a green gas supply for the gas grid. This paper explores the relevant pathways in which an integrated biomethane industry could potentially materialise and identifies and discusses the latest biotechnological advances in the production of renewable gas. Three scenarios of cascading biomethane systems are developed. Copyright © 2017 Elsevier Ltd. All rights reserved.

Wavefront analysis from its slope data

NASA Astrophysics Data System (ADS)

Mahajan, Virendra N.; Acosta, Eva

2017-08-01

In the aberration analysis of a wavefront over a certain domain, the polynomials that are orthogonal over and represent balanced wave aberrations for this domain are used. For example, Zernike circle polynomials are used for the analysis of a circular wavefront. Similarly, the annular polynomials are used to analyze the annular wavefronts for systems with annular pupils, as in a rotationally symmetric two-mirror system, such as the Hubble space telescope. However, when the data available for analysis are the slopes of a wavefront, as, for example, in a Shack- Hartmann sensor, we can integrate the slope data to obtain the wavefront data, and then use the orthogonal polynomials to obtain the aberration coefficients. An alternative is to find vector functions that are orthogonal to the gradients of the wavefront polynomials, and obtain the aberration coefficients directly as the inner products of these functions with the slope data. In this paper, we show that an infinite number of vector functions can be obtained in this manner. We show further that the vector functions that are irrotational are unique and propagate minimum uncorrelated additive random noise from the slope data to the aberration coefficients.

Characteristics of a dry, pulsating microburst at Denver Stapleton Airport

NASA Technical Reports Server (NTRS)

Proctor, Fred H.

1994-01-01

This study examines the influence of ambient vertical wind shear on microburst intensity, asymmetry, and translation. Results show that microburst asymmetry is influenced by the magnitude of the low-level ambient vertical shear. The microburst outflow elongates in the direction of the shear vector (which is not necessarily in the direction of translation), and generates the greatest hazard (for commercial jet transports) along paths orthogonal to the shear vector. The model results also show that the asymmetry increases with increasing shear magnitude. One implication of these results concerns the detection of a microburst by a ground-based doppler systems. These systems may underestimate the hazard for landing and departing aircraft that are on trajectories orthogonal to both the sensor beam and shear vector, especially if the magnitude of the shear is large. Another implication is that microburst are more likely to be asymmetrical in regions (seasons) where there is climatologically a significant low-level shear. The model results also show that the rotor microbursts and severe wind damage can be a product of the microburst interaction with strong ambient wind shear.

Paratransgenesis applied for control of tsetse transmitted sleeping sickness.

PubMed

Aksoy, Serap; Weiss, Brian; Attardo, Geoffrey

2008-01-01

African trypanosomiasis (sleeping sickness) is a major cause of morbidity and mortality in Subsaharan Africa for human and animal health. In the absence of effective vaccines and efficacious drugs, vector control is an alternative intervention tool to break the disease cycle. This chapter describes the vectorial and symbiotic biology of tsetse with emphasis on the current knowledge on tsetse symbiont genomics and functional biology, and tsetse's trypanosome transmission capability. The ability to culture one of tsetse's commensal symbiotic microbes, Sodalis in vitro has allowed for the development of a genetic transformation system for this organism. Tsetse can be repopulated with the modified Sodalis symbiont, which can express foreign gene products (an approach we refer to as paratransgenic expression system). Expanding knowledge on tsetse immunity effectors, on genomics of tsetse symbionts and on tsetse's parasite transmission biology stands to enhance the development and potential application of paratransgenesis as a new vector-control strategy. We describe the hallmarks of the paratransgenic transformation technology where the modified symbionts expressing trypanocidal compounds can be used to manipulate host functions and lead to the control of trypanosomiasis by blocking trypanosome transmission in the tsetse vector.

Production of adenovirus vectors and their use as a delivery system for influenza vaccines

PubMed Central

Vemula, Sai V.; Mittal, Suresh K.

2010-01-01

IMPORTANCE OF THE FIELD With the emergence of highly pathogenic avian influenza H5N1 viruses that have crossed species barriers and are responsible for lethal infections in humans in many countries, there is an urgent need for the development of effective vaccines which can be produced in large quantities at a short notice and confer broad protection against these H5N1 variants. In order to meet the potential global vaccine demand in a pandemic scenario, new vaccine-production strategies must be explored in addition to the currently used egg-based technology for seasonal influenza. AREAS COVERED IN THIS REVIEW Adenovirus (Ad) based influenza vaccines represent an attractive alternative/supplement to the currently licensed egg-based influenza vaccines. Ad-based vaccines are relatively inexpensive to manufacture, and their production process does not require either chicken eggs or labor intensive and time-consuming processes necessitating enhanced biosafety facilities. Most importantly, in a pandemic situation, this vaccine strategy could offer a stockpiling option to reduce the response time before a strain-matched vaccine could be developed. WHAT THE READER WILL GAIN This review discusses Ad-vector technology and the current progress in the development of Ad-based influenza vaccines. TAKE HOME MESSAGE Ad vector-based influenza vaccines for pandemic preparedness are under development to meet the global vaccine demand. PMID:20822477

Comparison of the ocean surface vector winds from atmospheric reanalysis and scatterometer-based wind products over the Nordic Seas and the northern North Atlantic and their application for ocean modeling

NASA Astrophysics Data System (ADS)

Dukhovskoy, Dmitry S.; Bourassa, Mark A.; Petersen, Gudrún Nína; Steffen, John

2017-03-01

Ocean surface vector wind fields from reanalysis data sets and scatterometer-derived gridded products are analyzed over the Nordic Seas and the northern North Atlantic for the time period from 2000 to 2009. The data sets include the National Center for Environmental Prediction Reanalysis 2 (NCEPR2), Climate Forecast System Reanalysis (CFSR), Arctic System Reanalysis (ASR), Cross-Calibrated Multiplatform (CCMP) wind product version 1.1 and recently released version 2.0, and QuikSCAT. The goal of the study is to assess discrepancies across the wind vector fields in the data sets and demonstrate possible implications of these differences for ocean modeling. Large-scale and mesoscale characteristics of winds are compared at interannual, seasonal, and synoptic timescales. A cyclone tracking methodology is developed and applied to the wind fields to compare cyclone characteristics in the data sets. Additionally, the winds are evaluated against observations collected from meteorological buoys deployed in the Iceland and Irminger Seas. The agreement among the wind fields is better for longer time and larger spatial scales. The discrepancies are clearly apparent for synoptic timescales and mesoscales. CCMP, ASR, and CFSR show the closest overall agreement with each other. Substantial biases are found in the NCEPR2 winds. Numerical sensitivity experiments are conducted with a coupled ice-ocean model forced by different wind fields. The experiments demonstrate differences in the net surface heat fluxes during storms. In the experiment forced by NCEPR2 winds, there are discrepancies in the large-scale wind-driven ocean dynamics compared to the other experiments.

Supplier Short Term Load Forecasting Using Support Vector Regression and Exogenous Input

NASA Astrophysics Data System (ADS)

Matijaš, Marin; Vukićcević, Milan; Krajcar, Slavko

2011-09-01

In power systems, task of load forecasting is important for keeping equilibrium between production and consumption. With liberalization of electricity markets, task of load forecasting changed because each market participant has to forecast their own load. Consumption of end-consumers is stochastic in nature. Due to competition, suppliers are not in a position to transfer their costs to end-consumers; therefore it is essential to keep forecasting error as low as possible. Numerous papers are investigating load forecasting from the perspective of the grid or production planning. We research forecasting models from the perspective of a supplier. In this paper, we investigate different combinations of exogenous input on the simulated supplier loads and show that using points of delivery as a feature for Support Vector Regression leads to lower forecasting error, while adding customer number in different datasets does the opposite.

Fully Differential Vector-Boson-Fusion Higgs Production at Next-to-Next-to-Leading Order.

PubMed

Cacciari, Matteo; Dreyer, Frédéric A; Karlberg, Alexander; Salam, Gavin P; Zanderighi, Giulia

2015-08-21

We calculate the fully differential next-to-next-to-leading-order (NNLO) corrections to vector-boson fusion (VBF) Higgs boson production at proton colliders, in the limit in which there is no cross talk between the hadronic systems associated with the two protons. We achieve this using a new "projection-to-Born" method that combines an inclusive NNLO calculation in the structure-function approach and a suitably factorized next-to-leading-order VBF Higgs plus three-jet calculation, using appropriate Higgs plus two-parton counterevents. An earlier calculation of the fully inclusive cross section had found small NNLO corrections, at the 1% level. In contrast, the cross section after typical experimental VBF cuts receives NNLO contributions of about (5-6)%, while differential distributions show corrections of up to (10-12)% for some standard observables. The corrections are often outside the next-to-leading-order scale-uncertainty band.

Assessing Undergraduate Curriculum Through Student Exit Vectors

NASA Astrophysics Data System (ADS)

Keane, C. M.; Gonzales, L.; Martinez, C.

2008-12-01

One aspect of assessing the undergraduate curriculum is recognizing that the exit vector of the student is a metric in the absence of a structured assessment program. Detailed knowledge across all geosciences departments regarding the disposition of their recent baccalaureate recipients has been at best inconsistent, and in the case of about half of geoscience programs, non-existent. However, through examining of multiple datasets, a pattern of disposition of geosciences BS recipients emerges, providing a snapshot of the system- wide response to the system-wide "average" program. This pattern can also be juxtaposed against several frameworks of desired skill sets for recent graduates and the employment sectors likely to hire them. The question remains is can one deduce the effectiveness of the undergraduate program in placing graduates in their next step, whether in graduate school or the workplace. Likewise, with an increasing scrutiny on the "value" of an education, is the resulting economic gain sufficient for the student, such that programs will be viewed as sustainable. A factor in answering this question is the importance of the undergraduate program in the ultimate destination of the professional. Clear pathways exist for "optimal" schools for the production of new faculty and new industry professionals, but is it possible to identify those trends further up the educational pipeline? One major mechanism to examine the undergraduate program effectiveness related to exit vectors is to look at hiring trends witnessed related to markedly different program structures, such as those at universities outside of the United States. Rectifying academic programs between the United States and other national systems is often a challenge, but even given the substantial differences between depth of technical knowledge and breadth of education across these programs, in the end, the sum product is often viewed as roughly comparable. This paper will look at end-of-baccalaureate vectors in several countries, including Australia and South Africa, and how it reflects on the structure of their programs, how the programs align with the country's professional needs, and the ability for the undergraduate geosciences system to provide the key intellectual feedstock for sustaining the geosciences discipline in these countries.

Recombinase-mediated cassette exchange (RMCE) for monoclonal antibody expression in the commercially relevant CHOK1SV cell line.

PubMed

Zhang, Lin; Inniss, Mara C; Han, Shu; Moffat, Mark; Jones, Heather; Zhang, Baohong; Cox, Wendy L; Rance, James R; Young, Robert J

2015-01-01

To meet product quality and cost parameters for therapeutic monoclonal antibody (mAb) production, cell lines are required to have excellent growth, stability, and productivity characteristics. In particular, cell line generation stability is critical to the success of a program, especially where high cell line generation numbers are required for large in-market supply. However, a typical process for developing such cell lines is laborious, lengthy, and costly. In this study, we applied a FLP/FRT recombinase-mediated cassette exchange (RMCE) system to build a site-specific integration (SSI) system for mAb expression in the commercially relevant CHOK1SV cell line. Using a vector with a FRT-flanked mAb expression cassette, we generated a clonal cell line with good productivity, long-term production stability, and low mAb gene-copy number indicating the vector was located in a 'hot-spot.' A SSI host cell line was made by removing the mAb genes from the 'hot-spot' by RMCE, creating a 'landing pad' containing two recombination cassettes that allow targeting of one or two copies of recombinant genes. Cell lines made from this host exhibited excellent growth and productivity profiles, and stability for at least 100 generations in the absence of selection agents. Importantly, while clones containing two copies had higher productivity than single copy clones, both were stable over many generations. Taken together, this study suggests the use of FLP-based RMCE to develop SSI host cells for mAb production in CHOK1SV offers significant savings in both resources and overall cell line development time, leading to a shortened 'time-to-clinic' for therapeutic mAbs. © 2015 American Institute of Chemical Engineers.

Wheel speed management control system for spacecraft

NASA Technical Reports Server (NTRS)

Goodzeit, Neil E. (Inventor); Linder, David M. (Inventor)

1991-01-01

A spacecraft attitude control system uses at least four reaction wheels. In order to minimize reaction wheel speed and therefore power, a wheel speed management system is provided. The management system monitors the wheel speeds and generates a wheel speed error vector. The error vector is integrated, and the error vector and its integral are combined to form a correction vector. The correction vector is summed with the attitude control torque command signals for driving the reaction wheels.

Efficient and stable production of Modified Vaccinia Ankara virus in two-stage semi-continuous and in continuous stirred tank cultivation systems.

PubMed

Tapia, Felipe; Jordan, Ingo; Genzel, Yvonne; Reichl, Udo

2017-01-01

One important aim in cell culture-based viral vaccine and vector production is the implementation of continuous processes. Such a development has the potential to reduce costs of vaccine manufacturing as volumetric productivity is increased and the manufacturing footprint is reduced. In this work, continuous production of Modified Vaccinia Ankara (MVA) virus was investigated. First, a semi-continuous two-stage cultivation system consisting of two shaker flasks in series was established as a small-scale approach. Cultures of the avian AGE1.CR.pIX cell line were expanded in the first shaker, and MVA virus was propagated and harvested in the second shaker over a period of 8-15 days. A total of nine small-scale cultivations were performed to investigate the impact of process parameters on virus yields. Harvest volumes of 0.7-1 L with maximum TCID50 titers of up to 1.0×109 virions/mL were obtained. Genetic analysis of control experiments using a recombinant MVA virus containing green-fluorescent-protein suggested that the virus was stable over at least 16 d of cultivation. In addition, a decrease or fluctuation of infectious units that may indicate an excessive accumulation of defective interfering particles was not observed. The process was automated in a two-stage continuous system comprising two connected 1 L stirred tank bioreactors. Stable MVA virus titers, and a total production volume of 7.1 L with an average TCID50 titer of 9×107 virions/mL was achieved. Because titers were at the lower range of the shake flask cultivations potential for further process optimization at large scale will be discussed. Overall, MVA virus was efficiently produced in continuous and semi-continuous cultivations making two-stage stirred tank bioreactor systems a promising platform for industrial production of MVA-derived recombinant vaccines and viral vectors.

Solving periodic block tridiagonal systems using the Sherman-Morrison-Woodbury formula

NASA Technical Reports Server (NTRS)

Yarrow, Maurice

1989-01-01

Many algorithms for solving the Navier-Stokes equations require the solution of periodic block tridiagonal systems of equations. By applying a splitting to the matrix representing this system of equations, it may first be reduced to a block tridiagonal matrix plus an outer product of two block vectors. The Sherman-Morrison-Woodbury formula is then applied. The algorithm thus reduces a periodic banded system to a non-periodic banded system with additional right-hand sides and is of higher efficiency than standard Thomas algorithm/LU decompositions.

Electromagnetic energy flux vector for a dispersive linear medium.

PubMed

Crenshaw, Michael E; Akozbek, Neset

2006-05-01

The electromagnetic energy flux vector in a dispersive linear medium is derived from energy conservation and microscopic quantum electrodynamics and is found to be of the Umov form as the product of an electromagnetic energy density and a velocity vector.

Epidemiology of East Coast fever (Theileria parva infection) in Kenya: past, present and the future.

PubMed

Gachohi, John; Skilton, Rob; Hansen, Frank; Ngumi, Priscilla; Kitala, Philip

2012-09-07

In this article, we review the epidemiology of East Coast fever (ECF), a tick-borne infection of cattle, in Kenya. The major factors associated with epidemiology of ECF include the agro-ecological zone (AEZ), livestock production system (LPS) and both animal breed and age. These factors appear to influence the epidemiology of ECF through structured gradients. We further show that the gradients are dynamically shaped by socio-demographic and environmental processes. For a vector-borne disease whose transmission depends on environmental characteristics that influence vector dynamics, a change in the environment implies a change in the epidemiology of the disease. The review recommends that future ECF epidemiological studies should account for these factors and the dynamic interactions between them. In Kenya, ECF control has previously relied predominantly on tick control using acaricides and chemotherapy while ECF immunization is steadily being disseminated. We highlight the contribution of ECF epidemiology and economics in the design of production system and/or geographical area-specific integrated control strategies based on both the dynamic epidemiological risk of the disease and economic impacts of control strategies. In all production systems (except marginal areas), economic analyses demonstrate that integrated control in which ECF immunization is always an important component, can play an important role in the overall control of the disease. Indeed, Kenya has recently approved ECF immunization in all production systems (except in marginal areas). If the infrastructure of the vaccine production and distribution can be heightened, large ECF endemic areas are expected to be endemically stable and the disease controlled. Finally, the review points the way for future research by identifying scenario analyses as a critical methodology on which to base future investigations on how both dynamic livestock management systems and patterns of land use influence the dynamics and complexity of ECF epidemiology and the implications for control.

Epidemiology of East Coast fever (Theileria parva infection) in Kenya: past, present and the future

PubMed Central

2012-01-01

In this article, we review the epidemiology of East Coast fever (ECF), a tick-borne infection of cattle, in Kenya. The major factors associated with epidemiology of ECF include the agro-ecological zone (AEZ), livestock production system (LPS) and both animal breed and age. These factors appear to influence the epidemiology of ECF through structured gradients. We further show that the gradients are dynamically shaped by socio-demographic and environmental processes. For a vector-borne disease whose transmission depends on environmental characteristics that influence vector dynamics, a change in the environment implies a change in the epidemiology of the disease. The review recommends that future ECF epidemiological studies should account for these factors and the dynamic interactions between them. In Kenya, ECF control has previously relied predominantly on tick control using acaricides and chemotherapy while ECF immunization is steadily being disseminated. We highlight the contribution of ECF epidemiology and economics in the design of production system and/or geographical area-specific integrated control strategies based on both the dynamic epidemiological risk of the disease and economic impacts of control strategies. In all production systems (except marginal areas), economic analyses demonstrate that integrated control in which ECF immunization is always an important component, can play an important role in the overall control of the disease. Indeed, Kenya has recently approved ECF immunization in all production systems (except in marginal areas). If the infrastructure of the vaccine production and distribution can be heightened, large ECF endemic areas are expected to be endemically stable and the disease controlled. Finally, the review points the way for future research by identifying scenario analyses as a critical methodology on which to base future investigations on how both dynamic livestock management systems and patterns of land use influence the dynamics and complexity of ECF epidemiology and the implications for control. PMID:22958352

Insect cells-baculovirus system for the production of difficult to express proteins.

PubMed

Osz-Papai, Judit; Radu, Laura; Abdulrahman, Wassim; Kolb-Cheynel, Isabelle; Troffer-Charlier, Nathalie; Birck, Catherine; Poterszman, Arnaud

2015-01-01

The production of sufficient quantities of homogenous protein not only is an essential prelude for structural investigations but also represents a rate-limiting step for many human functional studies. Although technologies for expression of recombinant proteins and complexes have been improved tremendously, in many cases, protein production remains a challenge and can be associated with considerable investment. This chapter describes simple and efficient protocols for expression screening and optimization of protein production in insect cells using the baculovirus expression system. We describe the procedure, starting from the cloning of a gene of interest into an expression transfer baculovirus vector, followed by generation of the recombinant virus by homologous recombination, evaluation of protein expression, and scale-up. Handling of insect cell cultures and preparation of bacmid for co-transfection are also detailed.

Thyra Abstract Interface Package

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bartlett, Roscoe A.

2005-09-01

Thrya primarily defines a set of abstract C++ class interfaces needed for the development of abstract numerical atgorithms (ANAs) such as iterative linear solvers, transient solvers all the way up to optimization. At the foundation of these interfaces are abstract C++ classes for vectors, vector spaces, linear operators and multi-vectors. Also included in the Thyra package is C++ code for creating concrete vector, vector space, linear operator, and multi-vector subclasses as well as other utilities to aid in the development of ANAs. Currently, very general and efficient concrete subclass implementations exist for serial and SPMD in-core vectors and multi-vectors. Codemore » also currently exists for testing objects and providing composite objects such as product vectors.« less

Aerosol polarization effects on atmospheric correction and aerosol retrievals in ocean color remote sensing.

PubMed

Wang, Menghua

2006-12-10

The current ocean color data processing system for the Sea-viewing Wide Field-of-View Sensor (SeaWiFS) and the moderate resolution imaging spectroradiometer (MODIS) uses the Rayleigh lookup tables that were generated using the vector radiative transfer theory with inclusion of the polarization effects. The polarization effects, however, are not accounted for in the aerosol lookup tables for the ocean color data processing. I describe a study of the aerosol polarization effects on the atmospheric correction and aerosol retrieval algorithms in the ocean color remote sensing. Using an efficient method for the multiple vector radiative transfer computations, aerosol lookup tables that include polarization effects are generated. Simulations have been carried out to evaluate the aerosol polarization effects on the derived ocean color and aerosol products for all possible solar-sensor geometries and the various aerosol optical properties. Furthermore, the new aerosol lookup tables have been implemented in the SeaWiFS data processing system and extensively tested and evaluated with SeaWiFS regional and global measurements. Results show that in open oceans (maritime environment), the aerosol polarization effects on the ocean color and aerosol products are usually negligible, while there are some noticeable effects on the derived products in the coastal regions with nonmaritime aerosols.

LentiPro26: novel stable cell lines for constitutive lentiviral vector production.

PubMed

Tomás, H A; Rodrigues, A F; Carrondo, M J T; Coroadinha, A S

2018-03-27

Lentiviral vectors (LVs) are excellent tools to promote gene transfer and stable gene expression. Their potential has been already demonstrated in gene therapy clinical trials for the treatment of diverse disorders. For large scale LV production, a stable producer system is desirable since it allows scalable and cost-effective viral productions, with increased reproducibility and safety. However, the development of stable systems has been challenging and time-consuming, being the selection of cells presenting high expression levels of Gag-Pro-Pol polyprotein and the cytotoxicity associated with some viral components, the main limitations. Hereby is described the establishment of a new LV producer cell line using a mutated less active viral protease to overcome potential cytotoxic limitations. The stable transfection of bicistronic expression cassettes with re-initiation of the translation mechanism enabled the generation of LentiPro26 packaging populations supporting high titers. Additionally, by skipping intermediate clone screening steps and performing only one final clone screening, it was possible to save time and generate LentiPro26-A59 cell line, that constitutively produces titers above 10 6 TU.mL -1 .day -1 , in less than six months. This work constitutes a step forward towards the development of improved LV producer cell lines, aiming to efficiently supply the clinical expanding gene therapy applications.

Developmental neurogenetics of sexual dimorphism in Aedes aegypti

PubMed Central

Duman-Scheel, Molly; Syed, Zainulabeuddin

2015-01-01

Sexual dimorphism, a poorly understood but crucial aspect of vector mosquito biology, encompasses sex-specific physical, physiological, and behavioral traits related to mosquito reproduction. The study of mosquito sexual dimorphism has largely focused on analysis of the differences between adult female and male mosquitoes, particularly with respect to sex-specific behaviors related to disease transmission. However, sexually dimorphic behaviors are the products of differential gene expression that initiates during development and therefore must also be studied during development. Recent technical advancements are facilitating functional genetic studies in the dengue vector Aedes aegypti, an emerging model for mosquito development. These methodologies, many of which could be extended to other non-model insect species, are facilitating analysis of the development of sexual dimorphism in neural tissues, particularly the olfactory system. These studies are providing insight into the neurodevelopmental genetic basis for sexual dimorphism in vector mosquitoes. PMID:26949699

On real-space Density Functional Theory for non-orthogonal crystal systems: Kronecker product formulation of the kinetic energy operator

NASA Astrophysics Data System (ADS)

Sharma, Abhiraj; Suryanarayana, Phanish

2018-05-01

We present an accurate and efficient real-space Density Functional Theory (DFT) framework for the ab initio study of non-orthogonal crystal systems. Specifically, employing a local reformulation of the electrostatics, we develop a novel Kronecker product formulation of the real-space kinetic energy operator that significantly reduces the number of operations associated with the Laplacian-vector multiplication, the dominant cost in practical computations. In particular, we reduce the scaling with respect to finite-difference order from quadratic to linear, thereby significantly bridging the gap in computational cost between non-orthogonal and orthogonal systems. We verify the accuracy and efficiency of the proposed methodology through selected examples.

Elements of the quality management in the materials' industry

NASA Astrophysics Data System (ADS)

Ioana, Adrian; Semenescu, Augustin; Costoiu, Mihnea; Marcu, Dragoş

2017-12-01

The criteria function concept consists of transforming the criteria function (CF) in a quality-economical matrix math MQE. The levels of prescribing the criteria function was obtained by using a composition algorithm for three vectors: T¯ vector - technical parameters' vector (ti); Ē vector - economical parameters' vector (ej) and P¯ vector - weight vector (p1). For each product or service, the area of the circle represents the value of its sales. The BCG Matrix thus offers a very useful map of the organization's service strengths and weaknesses, at least in terms of current profitability, as well as the likely cash flows.

The myeloid-binding peptide adenoviral vector enables multi-organ vascular endothelial gene targeting.

PubMed

Lu, Zhi Hong; Kaliberov, Sergey; Zhang, Jingzhu; Muz, Barbara; Azab, Abdel K; Sohn, Rebecca E; Kaliberova, Lyudmila; Du, Yingqiu; Curiel, David T; Arbeit, Jeffrey M

2014-08-01

Vascular endothelial cells (ECs) are ideal gene therapy targets as they provide widespread tissue access and are the first contact surfaces following intravenous vector administration. Human recombinant adenovirus serotype 5 (Ad5) is the most frequently used gene transfer system because of its appreciable transgene payload capacity and lack of somatic mutation risk. However, standard Ad5 vectors predominantly transduce liver but not the vasculature following intravenous administration. We recently developed an Ad5 vector with a myeloid cell-binding peptide (MBP) incorporated into the knob-deleted, T4 fibritin chimeric fiber (Ad.MBP). This vector was shown to transduce pulmonary ECs presumably via a vector handoff mechanism. Here we tested the body-wide tropism of the Ad.MBP vector, its myeloid cell necessity, and vector-EC expression dose response. Using comprehensive multi-organ co-immunofluorescence analysis, we discovered that Ad.MBP produced widespread EC transduction in the lung, heart, kidney, skeletal muscle, pancreas, small bowel, and brain. Surprisingly, Ad.MBP retained hepatocyte tropism albeit at a reduced frequency compared with the standard Ad5. While binding specifically to myeloid cells ex vivo, multi-organ Ad.MBP expression was not dependent on circulating monocytes or macrophages. Ad.MBP dose de-escalation maintained full lung-targeting capacity but drastically reduced transgene expression in other organs. Swapping the EC-specific ROBO4 for the CMV promoter/enhancer abrogated hepatocyte expression but also reduced gene expression in other organs. Collectively, our multilevel targeting strategy could enable therapeutic biological production in previously inaccessible organs that pertain to the most debilitating or lethal human diseases.

76 FR 66032 - Availability of an Environmental Assessment for Field Testing Avian Influenza-Marek's Disease...

Federal Register 2010, 2011, 2012, 2013, 2014

2011-10-25

... Avian Influenza-Marek's Disease Vaccine, H5 Subtype, Serotype 3, Live Marek's Disease Vector AGENCY...-Marek's Disease Vaccine, H5 Subtype, Serotype 3, Live Marek's Disease Vector. The environmental... product: Requester: Biomune Company. Product: Avian Influenza-Marek's Disease Vaccine, H5 Subtype...

Control of cucurbit viruses.

PubMed

Lecoq, Hervé; Katis, Nikolaos

2014-01-01

More than 70 well-characterized virus species transmitted by a diversity of vectors may infect cucurbit crops worldwide. Twenty of those cause severe epidemics in major production areas, occasionally leading to complete crop failures. Cucurbit viruses' control is based on three major axes: (i) planting healthy seeds or seedlings in a clean environment, (ii) interfering with vectors activity, and (iii) using resistant cultivars. Seed disinfection and seed or seedling quality controls guarantee growers on the sanitary status of their planting material. Removal of virus or vector sources in the crop environment can significantly delay the onset of viral epidemics. Insecticide or oil application may reduce virus spread in some situations. Diverse cultural practices interfere with or prevent vector reaching the crop. Resistance can be obtained by grafting for soil-borne viruses, by cross-protection, or generally by conventional breeding or genetic engineering. The diversity of the actions that may be taken to limit virus spread in cucurbit crops and their limits will be discussed. The ultimate goal is to provide farmers with technical packages that combine these methods within an integrated disease management program and are adapted to different countries and cropping systems.

Quantum speed limit time in a magnetic resonance

NASA Astrophysics Data System (ADS)

Ivanchenko, E. A.

2017-12-01

A visualization for dynamics of a qudit spin vector in a time-dependent magnetic field is realized by means of mapping a solution for a spin vector on the three-dimensional spherical curve (vector hodograph). The obtained results obviously display the quantum interference of precessional and nutational effects on the spin vector in the magnetic resonance. For any spin the bottom bounds of the quantum speed limit time (QSL) are found. It is shown that the bottom bound goes down when using multilevel spin systems. Under certain conditions the non-nil minimal time, which is necessary to achieve the orthogonal state from the initial one, is attained at spin S = 2. An estimation of the product of two and three standard deviations of the spin components are presented. We discuss the dynamics of the mutual uncertainty, conditional uncertainty and conditional variance in terms of spin standard deviations. The study can find practical applications in the magnetic resonance, 3D visualization of computational data and in designing of optimized information processing devices for quantum computation and communication.

Simplified process for the production of anti-CD19-CAR engineered T cells

PubMed Central

Tumaini, Barbara; Lee, Daniel W.; Lin, Tasha; Castiello, Luciano; Stroncek, David F.; Mackall, Crystal; Wayne, Alan; Sabatino, Marianna

2014-01-01

Background Adoptive Immunotherapy using chimeric antigen receptor (CAR) engineered T cells specific for CD19 has shown promising results for the treatment of B cell lymphomas and leukemia. This therapy involves the transduction of autologous T cells with a viral vector and the subsequent cell expansion. Here, we describe a new, simplified method to produce anti-CD19-CAR T cells. Methods T cells were isolated from peripheral blood mononuclear cell (PBMC) with anti-CD3/anti-CD28 paramagnetic beads. After 2 days, the T cells were added to culture bags pre-treated with RetroNectin and loaded with the retroviral anti-CD19 CAR vector. The cells, beads and vector were incubated for 24 hours and then a second transduction was performed. No spinoculation was used. Cells were then expanded for an additional 9 days. Results The method was validated using 2 PBMC products from a patient with B-CLL and one PBMC product from a healthy subject. The 2 PBMC products from the B-CLL patient contained 11.4% and 12.9% T cells. The manufacture process led to final products highly enriched in T cells with a mean CD3+ cell content of 98%, a mean expansion of 10.6 fold and a mean transduction efficiency of 68%. Similar results were obtained from the PBMCs of the first 4 ALL patients treated at our institution. Discussion We developed a simplified semi-closed system for the initial selection, activation, transduction and expansion of T cells using anti-CD3/anti-CD28 beads and bags, to produce autologous anti-CD19 CAR transduced T cells to support an ongoing clinical trial. PMID:23992830

Use of Bibliometric Analysis to Assess the Scientific Productivity and Impact of the Global Emerging Infections Surveillance and Response System Program, 2006-2012.

PubMed

Reaves, Erik J; Valle, Ruben; Chandrasekera, Ruvani M; Soto, Giselle; Burke, Ronald L; Cummings, James F; Bausch, Daniel G; Kasper, Matthew R

2017-05-01

Scientific publication in academic literature is a key venue in which the U.S. Department of Defense's Global Emerging Infections Surveillance and Response System (GEIS) program disseminates infectious disease surveillance data. Bibliometric analyses are tools to evaluate scientific productivity and impact of published research, yet are not routinely used for disease surveillance. Our objective was to incorporate bibliometric indicators to measure scientific productivity and impact of GEIS-funded infectious disease surveillance, and assess their utility in the management of the GEIS surveillance program. Metrics on GEIS program scientific publications, project funding, and countries of collaborating institutions from project years 2006 to 2012 were abstracted from annual reports and program databases and organized by the six surveillance priority focus areas: respiratory infections, gastrointestinal infections, febrile and vector-borne infections, antimicrobial resistance, sexually transmitted infections, and capacity building and outbreak response. Scientific productivity was defined as the number of scientific publications in peer-reviewed literature derived from GEIS-funded projects. Impact was defined as the number of citations of a GEIS-funded publication by other peer-reviewed publications, and the Thomson Reuters 2-year journal impact factor. Indicators were retrieved from the Web of Science and Journal Citation Report. To determine the global network of international collaborations between GEIS partners, countries were organized by the locations of collaborating institutions. Between 2006 and 2012, GEIS distributed approximately US $330 million to support 921 total projects. On average, GEIS funded 132 projects (range 96-160) with $47 million (range $43 million-$53 million), annually. The predominant surveillance focus areas were respiratory infections with 317 (34.4%) projects and $225 million, and febrile and vector-borne infections with 274 (29.8%) projects and $45 million. The number of annual respiratory infections-related projects peaked in 2006 and 2009. The number of febrile and vector-borne infections projects increased from 29 projects in 2006 to 58 in 2012. There were 651 articles published in 147 different peer-reviewed journals, with an average Thomson Reuters 2-year journal impact factor of 4.2 (range 0.3-53.5). On average, 93 articles were published per year (range 67-117) with $510,000 per publication. Febrile and vector-borne, respiratory, and gastrointestinal infections had 287, 167, and 73 articles published, respectively. Of the 651 articles published, 585 (89.9%) articles were cited at least once (range 1-1,045). Institutions from 90 countries located in all six World Health Organization regions collaborated with surveillance projects. These findings summarize the GEIS-funded surveillance portfolio between 2006 and 2012, and demonstrate the scientific productivity and impact of the program in each of the six disease surveillance priority focus areas. GEIS might benefit from further financial investment in both the febrile and vector-borne and sexually transmitted infections surveillance priority focus areas and increasing peer-reviewed publications of surveillance data derived from respiratory infections projects. Bibliometric indicators are useful to measure scientific productivity and impact in surveillance systems; and this methodology can be utilized as a management tool to assess future changes to GEIS surveillance priorities. Additional metrics should be developed when peer-reviewed literature is not used to disseminate noteworthy accomplishments. Reprint & Copyright © 2017 Association of Military Surgeons of the U.S.

pYEMF, a pUC18-derived XcmI T-vector for efficient cloning of PCR products.

PubMed

Gu, Jingsong; Ye, Chunjiang

2011-03-01

A 1330-bp DNA sequence with two XcmI cassettes was inserted into pUC18 to construct an efficient XcmI T-vector parent plasmid, pYEMF. The large size of the inserted DNA fragment improved T-vector cleavage efficiency, and guaranteed good separation of the molecular components after restriction digestion. The pYEMF-T-vector generated from parent plasmid pYEMF permits blue/white colony screening; cloning efficiency analysis showed that most white colonies (>75%) were putative transformants which carried the cloning product. The sequence analysis and design approach presented here will facilitate applications in the fields of molecular biology and genetic engineering.

Strategies for vectorizing the sparse matrix vector product on the CRAY XMP, CRAY 2, and CYBER 205

NASA Technical Reports Server (NTRS)

Bauschlicher, Charles W., Jr.; Partridge, Harry

1987-01-01

Large, randomly sparse matrix vector products are important in a number of applications in computational chemistry, such as matrix diagonalization and the solution of simultaneous equations. Vectorization of this process is considered for the CRAY XMP, CRAY 2, and CYBER 205, using a matrix of dimension of 20,000 with from 1 percent to 6 percent nonzeros. Efficient scatter/gather capabilities add coding flexibility and yield significant improvements in performance. For the CYBER 205, it is shown that minor changes in the IO can reduce the CPU time by a factor of 50. Similar changes in the CRAY codes make a far smaller improvement.

Human gene therapy: novel approaches to improve the current gene delivery systems.

PubMed

Cucchiarini, Magali

2016-06-01

Even though gene therapy made its way through the clinics to treat a number of human pathologies since the early years of experimental research and despite the recent approval of the first gene-based product (Glybera) in Europe, the safe and effective use of gene transfer vectors remains a challenge in human gene therapy due to the existence of barriers in the host organism. While work is under active investigation to improve the gene transfer systems themselves, the use of controlled release approaches may offer alternative, convenient tools of vector delivery to achieve a performant gene transfer in vivo while overcoming the various physiological barriers that preclude its wide use in patients. This article provides an overview of the most significant contributions showing how the principles of controlled release strategies may be adapted for human gene therapy.

Peak Locations and Relative Phase of Different Decay Modes of the a 1 Axial Vector Resonance in Diffractive Production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Basdevant, Jean-Louis; Berger, Edmond L.

2015-05-01

We show that a single I = 1 spin-parity J(PC) = 1(++) a(1) resonance can manifest itself as two separated mass peaks, one decaying into an S-wave rho pi system and the second decaying into a P-wave f(0)(980)pi system, with a rapid increase of the phase difference between their amplitudes arising mainly from the structure of the diffractive production process. This study clarifies questions related to the mass, width, and decay rates of the a(1) resonance raised by the recent high statistics data of the COMPASS Collaboration on a 1 production in pi N -> pi pi pi N atmore » high energies.« less

Management of whitefly-transmitted viruses in open-field production systems.

PubMed

Lapidot, Moshe; Legg, James P; Wintermantel, William M; Polston, Jane E

2014-01-01

Whiteflies are a key pest of crops in open-field production throughout the tropics and subtropics. This is due in large part to the long and diverse list of devastating plant viruses transmitted by these vectors. Open-field production provides many challenges to manage these viruses and in many cases adequate management has not been possible. Diseases caused by whitefly-transmitted viruses have become limiting factors in open-field production of a wide range of crops, i.e., bean golden mosaic disease in beans, tomato yellow leaf curl disease in tomato, cassava mosaic disease and cassava brown streak disease in cassava, and cotton leaf crumple disease in cotton. While host resistance has proven to be the most cost-effective management solution, few examples of host resistance have been developed to date. The main strategy to limit the incidence of virus-infected plants has been the application of insecticides to reduce vector populations aided to some extent by the use of selected cultural practices. However, due to concerns about the effect of insecticides on pollinators, consumer demand for reduced pesticide use, and the ability of the whitefly vectors to develop insecticide-resistance, there is a growing need to develop and deploy strategies that do not rely on insecticides. The reduction in pesticide use will greatly increase the need for genetic resistance to more viruses in more crop plants. Resistance combined with selected IPM strategies could become a viable means to increase yields in crops produced in open fields despite the presence of whitefly-transmitted viruses.

Parasite killing in malaria non-vector mosquito Anopheles culicifacies species B: implication of nitric oxide synthase upregulation.

PubMed

Vijay, Sonam; Rawat, Manmeet; Adak, Tridibes; Dixit, Rajnikant; Nanda, Nutan; Srivastava, Harish; Sharma, Joginder K; Prasad, Godavarthi B K S; Sharma, Arun

2011-04-04

Anopheles culicifacies, the main vector of human malaria in rural India, is a complex of five sibling species. Despite being phylogenetically related, a naturally selected subgroup species B of this sibling species complex is found to be a poor vector of malaria. We have attempted to understand the differences between vector and non-vector Anopheles culicifacies mosquitoes in terms of transcriptionally activated nitric oxide synthase (AcNOS) physiologies to elucidate the mechanism of refractoriness. Identification of the differences between genes and gene products that may impart refractory phenotype can facilitate development of novel malaria transmission blocking strategies. We conducted a study on phylogenetically related susceptible (species A) and refractory (species B) sibling species of An. culicifacies mosquitoes to characterize biochemical and molecular differences in AcNOS gene and gene elements and their ability to inhibit oocyst growth. We demonstrate that in species B, AcNOS specific activity and nitrite/nitrates in mid-guts and haemolymph were higher as compared to species A after invasion of the mid-gut by P. vivax at the beginning and during the course of blood feeding. Semiquantitative RT-PCR and real time PCR data of AcNOS concluded that this gene is more abundantly expressed in midgut of species B than in species A and is transcriptionally upregulated post blood meals. Dietary feeding of L-NAME along with blood meals significantly inhibited midgut AcNOS activity leading to an increase in oocyst production in An. culicifacies species B. We hypothesize that upregulation of mosquito innate cytotoxicity due to NOS in refractory strain to Plasmodium vivax infection may contribute to natural refractoriness in An. culicifacies mosquito population. This innate capacity of refractory mosquitoes could represent the ancestral function of the mosquito immune system against the parasite and could be utilized to understand the molecular basis of refractoriness in planning effective vector control strategies.

Parasite Killing in Malaria Non-Vector Mosquito Anopheles culicifacies Species B: Implication of Nitric Oxide Synthase Upregulation

PubMed Central

Vijay, Sonam; Rawat, Manmeet; Adak, Tridibes; Dixit, Rajnikant; Nanda, Nutan; Srivastava, Harish; Sharma, Joginder K.; Prasad, Godavarthi B. K. S.; Sharma, Arun

2011-01-01

Background Anopheles culicifacies, the main vector of human malaria in rural India, is a complex of five sibling species. Despite being phylogenetically related, a naturally selected subgroup species B of this sibling species complex is found to be a poor vector of malaria. We have attempted to understand the differences between vector and non-vector Anopheles culicifacies mosquitoes in terms of transcriptionally activated nitric oxide synthase (AcNOS) physiologies to elucidate the mechanism of refractoriness. Identification of the differences between genes and gene products that may impart refractory phenotype can facilitate development of novel malaria transmission blocking strategies. Methodology/Principal Findings We conducted a study on phylogenetically related susceptible (species A) and refractory (species B) sibling species of An. culicifacies mosquitoes to characterize biochemical and molecular differences in AcNOS gene and gene elements and their ability to inhibit oocyst growth. We demonstrate that in species B, AcNOS specific activity and nitrite/nitrates in mid-guts and haemolymph were higher as compared to species A after invasion of the mid-gut by P. vivax at the beginning and during the course of blood feeding. Semiquantitative RT-PCR and real time PCR data of AcNOS concluded that this gene is more abundantly expressed in midgut of species B than in species A and is transcriptionally upregulated post blood meals. Dietary feeding of L-NAME along with blood meals significantly inhibited midgut AcNOS activity leading to an increase in oocyst production in An. culicifacies species B. Conclusions/Significance We hypothesize that upregulation of mosquito innate cytotoxicity due to NOS in refractory strain to Plasmodium vivax infection may contribute to natural refractoriness in An. culicifacies mosquito population. This innate capacity of refractory mosquitoes could represent the ancestral function of the mosquito immune system against the parasite and could be utilized to understand the molecular basis of refractoriness in planning effective vector control strategies. PMID:21483693

Analysis of Particle Content of Recombinant Adeno-Associated Virus Serotype 8 Vectors by Ion-Exchange Chromatography

PubMed Central

Lock, Martin; Alvira, Mauricio R.

2012-01-01

Abstract Advances in adeno-associated virus (AAV)-mediated gene therapy have brought the possibility of commercial manufacturing of AAV vectors one step closer. To realize this prospect, a parallel effort with the goal of ever-increasing sophistication for AAV vector production technology and supporting assays will be required. Among the important release assays for a clinical gene therapy product, those monitoring potentially hazardous contaminants are most critical for patient safety. A prominent contaminant in many AAV vector preparations is vector particles lacking a genome, which can substantially increase the dose of AAV capsid proteins and lead to possible unwanted immunological consequences. Current methods to determine empty particle content suffer from inconsistency, are adversely affected by contaminants, or are not applicable to all serotypes. Here we describe the development of an ion-exchange chromatography-based assay that permits the rapid separation and relative quantification of AAV8 empty and full vector particles through the application of shallow gradients and a strong anion-exchange monolith chromatography medium. PMID:22428980

Killing vector fields in three dimensions: a method to solve massive gravity field equations

NASA Astrophysics Data System (ADS)

Gürses, Metin

2010-10-01

Killing vector fields in three dimensions play an important role in the construction of the related spacetime geometry. In this work we show that when a three-dimensional geometry admits a Killing vector field then the Ricci tensor of the geometry is determined in terms of the Killing vector field and its scalars. In this way we can generate all products and covariant derivatives at any order of the Ricci tensor. Using this property we give ways to solve the field equations of topologically massive gravity (TMG) and new massive gravity (NMG) introduced recently. In particular when the scalars of the Killing vector field (timelike, spacelike and null cases) are constants then all three-dimensional symmetric tensors of the geometry, the Ricci and Einstein tensors, their covariant derivatives at all orders, and their products of all orders are completely determined by the Killing vector field and the metric. Hence, the corresponding three-dimensional metrics are strong candidates for solving all higher derivative gravitational field equations in three dimensions.

Plasmid-encoded biosynthetic genes alleviate metabolic disadvantages while increasing glucose conversion to shikimate in an engineered Escherichia coli strain.

PubMed

Rodriguez, Alberto; Martínez, Juan A; Millard, Pierre; Gosset, Guillermo; Portais, Jean-Charles; Létisse, Fabien; Bolivar, Francisco

2017-06-01

Metabolic engineering strategies applied over the last two decades to produce shikimate (SA) in Escherichia coli have resulted in a battery of strains bearing many expression systems. However, the effects that these systems have on the host physiology and how they impact the production of SA are still not well understood. In this work we utilized an engineered E. coli strain to determine the consequences of carrying a vector that promotes SA production from glucose with a high-yield but that is also expected to impose a significant cellular burden. Kinetic comparisons in fermentors showed that instead of exerting a negative effect, the sole presence of the plasmid increased glucose consumption without diminishing the growth rate. By constitutively expressing a biosynthetic operon from this vector, the more active glycolytic metabolism was exploited to redirect intermediates toward the production of SA, which further increased the glucose consumption rate and avoided excess acetate production. Fluxomics and metabolomics experiments revealed a global remodeling of the carbon and energy metabolism in the production strain, where the increased SA production reduced the carbon available for oxidative and fermentative pathways. Moreover, the results showed that the production of SA relies on a specific setup of the pentose phosphate pathway, where both its oxidative and non-oxidative branches are strongly activated to supply erythrose-4-phosphate and balance the NADPH requirements. This work improves our understanding of the metabolic reorganization observed in E. coli in response to the plasmid-based expression of the SA biosynthetic pathway. Biotechnol. Bioeng. 2017;114: 1319-1330. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Wave Phase-Sensitive Transformation of 3d-Straining of Mechanical Fields

NASA Astrophysics Data System (ADS)

Smirnov, I. N.; Speranskiy, A. A.

2015-11-01

It is the area of research of oscillatory processes in elastic mechanical systems. Technical result of innovation is creation of spectral set of multidimensional images which reflect time-correlated three-dimensional vector parameters of metrological, and\\or estimated, and\\or design parameters of oscillations in mechanical systems. Reconstructed images of different dimensionality integrated in various combinations depending on their objective function can be used as homeostatic profile or cybernetic image of oscillatory processes in mechanical systems for an objective estimation of current operational conditions in real time. The innovation can be widely used to enhance the efficiency of monitoring and research of oscillation processes in mechanical systems (objects) in construction, mechanical engineering, acoustics, etc. Concept method of vector vibrometry based on application of vector 3D phase- sensitive vibro-transducers permits unique evaluation of real stressed-strained states of power aggregates and loaded constructions and opens fundamental innovation opportunities: conduct of continuous (on-line regime) reliable monitoring of turboagregates of electrical machines, compressor installations, bases, supports, pipe-lines and other objects subjected to damaging effect of vibrations; control of operational safety of technical systems at all the stages of life cycle including design, test production, tuning, testing, operational use, repairs and resource enlargement; creation of vibro-diagnostic systems of authentic non-destructive control of anisotropic characteristics of materials resistance of power aggregates and loaded constructions under outer effects and operational flaws. The described technology is revolutionary, universal and common for all branches of engineering industry and construction building objects.

Development and Implementation of Flood Risk Mapping, Water Bodies Monitoring and Climate Information for Human Health

NASA Astrophysics Data System (ADS)

Ceccato, P.; McDonald, K. C.; Jensen, K.; Podest, E.; De La Torre Juarez, M.

2013-12-01

Public health professionals are increasingly concerned about the potential impact that climate variability and change can have on infectious disease. The International Research Institute for Climate and Society (IRI), City College of New York (CCNY) and NASA Jet Propulsion Laboratory (JPL) are developing new products to increase the public health community's capacity to understand, use, and demand the appropriate climate data and climate information to mitigate the public health impacts of climate on vector-borne diseases such as malaria, leishmaniasis, rift valley fever. In this poster we present the new and improved products that have been developed for monitoring water bodies for monitoring and forecasting risks of vector-borne disease epidemics. The products include seasonal inundation patterns in the East African region based on the global mappings of inundated water fraction derived at the 25-km scale from both active and passive microwave instruments QuikSCAT, AMSR-E, SSM/I, ERS, ASCAT, and MODIS and LANDSAT data. We also present how the products are integrated into a knowledge system (IRI Data Library Map room, SERVIR) to support the use of climate and environmental information in climate-sensitive health decision-making.

Genetic characterization and construction of an auxotrophic strain of Saccharomyces cerevisiae JP1, a Brazilian industrial yeast strain for bioethanol production.

PubMed

Reis, Viviane Castelo Branco; Nicola, André Moraes; de Souza Oliveira Neto, Osmar; Batista, Vinícius Daniel Ferreira; de Moraes, Lidia Maria Pepe; Torres, Fernando Araripe Gonçalves

2012-11-01

Used for millennia to produce beverages and food, Saccharomyces cerevisiae also became a workhorse in the production of biofuels, most notably bioethanol. Yeast strains have acquired distinct characteristics that are the result of evolutionary adaptation to the stresses of industrial ethanol production. JP1 is a dominant industrial S. cerevisiae strain isolated from a sugarcane mill and is becoming increasingly popular for bioethanol production in Brazil. In this work, we carried out the genetic characterization of this strain and developed a set of tools to permit its genetic manipulation. Using flow cytometry, mating type, and sporulation analysis, we verified that JP1 is diploid and homothallic. Vectors with dominant selective markers for G418, hygromycin B, zeocin, and ρ-fluoro-DL-phenylalanine were used to successfully transform JP1 cells. Also, an auxotrophic ura3 mutant strain of JP1 was created by gene disruption using integration cassettes with dominant markers flanked by loxP sites. Marker excision was accomplished by the Cre/loxP system. The resulting auxotrophic strain was successfully transformed with an episomal vector that allowed green fluorescent protein expression.

Gene transfer of high-mobility group box 1 box-A domain in a rat acute liver failure model.

PubMed

Tanaka, Masayuki; Shinoda, Masahiro; Takayanagi, Atsushi; Oshima, Go; Nishiyama, Ryo; Fukuda, Kazumasa; Yagi, Hiroshi; Hayashida, Tetsu; Masugi, Yohei; Suda, Koichi; Yamada, Shingo; Miyasho, Taku; Hibi, Taizo; Abe, Yuta; Kitago, Minoru; Obara, Hideaki; Itano, Osamu; Takeuchi, Hiroya; Sakamoto, Michiie; Tanabe, Minoru; Maruyama, Ikuro; Kitagawa, Yuko

2015-04-01

High-mobility group box 1 (HMGB1) has recently been identified as an important mediator of various kinds of acute and chronic inflammation. The protein encoded by the box-A domain of the HMGB1 gene is known to act as a competitive inhibitor of HMGB1. In this study, we investigated whether box-A gene transfer results in box-A protein production in rats and assessed therapeutic efficacy in vivo using an acute liver failure (ALF) model. Three types of adenovirus vectors were constructed-a wild type and two mutants-and a mutant vector was then selected based on the secretion from HeLa cells. The secreted protein was subjected to a tumor necrosis factor (TNF) production inhibition test in vitro. The vector was injected via the portal vein in healthy Wistar rats to confirm box-A protein production in the liver. The vector was then injected via the portal vein in rats with ALF. Western blot analysis showed enhanced expression of box-A protein in HeLa cells transfected with one of the mutant vectors. The culture supernatant from HeLa cells transfected with the vector inhibited TNF-α production from macrophages. Expression of box-A protein was confirmed in the transfected liver at 72 h after transfection. Transfected rats showed decreased hepatic enzymes, plasma HMGB1, and hepatic TNF-α messenger RNA levels, and histologic findings and survival were significantly improved. HMGB1 box-A gene transfer results in box-A protein production in the liver and appears to have a beneficial effect on ALF in rats. Copyright © 2015 Elsevier Inc. All rights reserved.

Adaptation of retrovirus producer cells to serum deprivation: Implications in lipid biosynthesis and vector production.

PubMed

Rodrigues, A F; Amaral, A I; Veríssimo, V; Alves, P M; Coroadinha, A S

2012-05-01

The manufacture of enveloped virus, particularly retroviral (RV) and lentiviral (LV) vectors, faces the challenge of low titers that are aggravated under serum deprivation culture conditions. Also, the scarce knowledge on the biochemical pathways related with virus production is still limiting the design of rational strategies for improved production yields. This work describes the adaptation to serum deprivation of two human RV packaging cell lines, 293 FLEX and Te Fly and its effects on lipid biosynthetic pathways and infectious vector production. Total lipid content as well as cellular cholesterol were quantified and lipid biosynthesis was assessed by (13)C-NMR spectroscopy; changes in gene expression of lipid biosynthetic enzymes were also evaluated. The effects of adaptation to serum deprivation in lipid biosynthesis were cell line specific and directly correlated with infectious virus titers: 293 FLEX cells faced severe lipid starvation-up to 50% reduction in total lipid content-along with a 68-fold reduction in infectious vector titers; contrarily, Te Fly cells were able to maintain identical levels of total lipid content by rising de novo lipid biosynthesis, particularly for cholesterol-50-fold increase-with the consequent recovery of infectious vector productivities. Gene expression analysis of lipid biosynthetic enzymes further confirmed cholesterol production pathway to be prominently up-regulated under serum deprivation conditions for Te Fly cells, providing a genotype-phenotype validation for enhanced cholesterol synthesis. These results highlight lipid metabolism dynamics and the ability to activate lipid biosynthesis under serum deprivation as an important feature for high retroviral titers. Mechanisms underlying virus production and its relationship with lipid biosynthesis, with special focus on cholesterol, are discussed as potential targets for cellular metabolic engineering. Copyright © 2011 Wiley Periodicals, Inc.

Disclosing the Parameters Leading to High Productivity of Retroviral Producer Cells Lines: Evaluating Random Versus Targeted Integration.

PubMed

Bandeira, Vanessa S; Tomás, Hélio A; Alici, Evren; Carrondo, Manuel J T; Coroadinha, Ana S

2017-04-01

Gammaretrovirus and lentivirus are the preferred viral vectors to genetically modify T and natural killer cells to be used in immune cell therapies. The transduction efficiency of hematopoietic and T cells is more efficient using gibbon ape leukemia virus (GaLV) pseudotyping. In this context gammaretroviral vector producer cells offer competitive higher titers than transient lentiviral vectors productions. The main aim of this work was to identify the key parameters governing GaLV-pseudotyped gammaretroviral vector productivity in stable producer cells, using a retroviral vector expression cassette enabling positive (facilitating cell enrichment) and negative cell selection (allowing cell elimination). The retroviral vector contains a thymidine kinase suicide gene fused with a ouabain-resistant Na + ,K + -ATPase gene, a potential safer and faster marker. The establishment of retroviral vector producer cells is traditionally performed by randomly integrating the retroviral vector expression cassette codifying the transgene. More recently, recombinase-mediated cassette exchange methodologies have been introduced to achieve targeted integration. Herein we compared random and targeted integration of the retroviral vector transgene construct. Two retroviral producer cell lines, 293 OuaS and 293 FlexOuaS, were generated by random and targeted integration, respectively, producing high titers (on the order of 10 7 infectious particles·ml -1 ). Results showed that the retroviral vector transgene cassette is the key retroviral vector component determining the viral titers notwithstanding, single-copy integration is sufficient to provide high titers. The expression levels of the three retroviral constructs (gag-pol, GaLV env, and retroviral vector transgene) were analyzed. Although gag-pol and GaLV env gene expression levels should surpass a minimal threshold, we found that relatively modest expression levels of these two expression cassettes are required. Their levels of expression should not be maximized. We concluded, to establish a high producer retroviral vector cell line only the expression level of the genomic retroviral RNA, that is, the retroviral vector transgene cassette, should be maximized, both through (1) the optimization of its design (i.e., genetic elements composition) and (2) the selection of high expressing chromosomal locus for its integration. The use of methodologies identifying and promoting integration into high-expression loci, as targeted integration or high-throughput screening are in this perspective highly valuable.

Syngeneic AAV pseudo-particles potentiate gene transduction of AAV vectors

USDA-ARS?s Scientific Manuscript database

Gene delivery vectors based on adeno-associated virus (AAV) have emerged as safe and efficient therapeutic platform for numerous diseases. Excessive empty particles were generated as impurities during AAV vector production, but their effects on clinical outcome of AAV gene therapy are unclear. Here,...

Non-viral vectors based on magnetoplexes, lipoplexes and polyplexes for VEGF gene delivery into central nervous system cells.

PubMed

Villate-Beitia, Ilia; Puras, Gustavo; Soto-Sánchez, Cristina; Agirre, Mireia; Ojeda, Edilberto; Zarate, Jon; Fernández, Eduardo; Pedraz, José Luis

2017-04-15

Nanotechnology based non-viral vectors hold great promise to deliver therapeutic genes into the central nervous system (CNS) in a safe and controlled way. Vascular endothelial growth factor (VEGF) is a potential therapeutic gene candidate for CNS disorders due to its specific roles in brain angiogenesis and neuroprotection. In this work, we elaborated three different non-viral vectors based on magnetic, cationic lipid and polymeric nanoparticles complexed to the phVEGF165aIRESGFP plasmid, which codifies the VEGF protein -extracellular- and the green fluorescent protein (GFP) -intracellular-. Nanoparticles and corresponding nanoplexes -magnetoplexes, lipoplexes and polyplexes- were characterized in terms of size, zeta potential, polydispersity index, morphology and ability to bind, release and protect DNA. Transfection efficiencies of nanoplexes were measured in terms of percentage of GFP expressing cells, mean fluorescent intensity (MFI) and VEGF (ng/ml) production in HEK293, C6 and primary neuronal culture cells. Magnetoplexes showed the highest transfection efficiencies in C6, followed by lipoplexes, and in primary neuronal culture cells, followed by polyplexes. Lipoplexes were the most efficient in HEK293 cells, followed by magnetoplexes. The biological activity of VEGF was confirmed by its proliferative effect in HUVEC cells. Overall, these results provide new insights for VEGF gene delivery into CNS cells using non-viral vectors. Copyright © 2017. Published by Elsevier B.V.

Broad-host-range vector system for synthetic biology and biotechnology in cyanobacteria

PubMed Central

Taton, Arnaud; Unglaub, Federico; Wright, Nicole E.; Zeng, Wei Yue; Paz-Yepes, Javier; Brahamsha, Bianca; Palenik, Brian; Peterson, Todd C.; Haerizadeh, Farzad; Golden, Susan S.; Golden, James W.

2014-01-01

Inspired by the developments of synthetic biology and the need for improved genetic tools to exploit cyanobacteria for the production of renewable bioproducts, we developed a versatile platform for the construction of broad-host-range vector systems. This platform includes the following features: (i) an efficient assembly strategy in which modules released from 3 to 4 donor plasmids or produced by polymerase chain reaction are assembled by isothermal assembly guided by short GC-rich overlap sequences. (ii) A growing library of molecular devices categorized in three major groups: (a) replication and chromosomal integration; (b) antibiotic resistance; (c) functional modules. These modules can be assembled in different combinations to construct a variety of autonomously replicating plasmids and suicide plasmids for gene knockout and knockin. (iii) A web service, the CYANO-VECTOR assembly portal, which was built to organize the various modules, facilitate the in silico construction of plasmids, and encourage the use of this system. This work also resulted in the construction of an improved broad-host-range replicon derived from RSF1010, which replicates in several phylogenetically distinct strains including a new experimental model strain Synechocystis sp. WHSyn, and the characterization of nine antibiotic cassettes, four reporter genes, four promoters, and a ribozyme-based insulator in several diverse cyanobacterial strains. PMID:25074377

Hyaluronic acid synthase-2 gene transfer into the joints of Beagles by use of recombinant adeno-associated viral vectors.

PubMed

Kyostio-Moore, Sirkka; Berthelette, Patricia; Cornell, Cathleen Sookdeo; Nambiar, Bindu; Figueiredo, Monica Dias

2018-05-01

OBJECTIVE To evaluate gene transfer of recombinant adeno-associated viral (rAAV) vectors with AAV2 or AAV5 capsid and encoding hyaluronic acid (HA) synthase-2 (HAS2) into joints of healthy dogs. ANIMALS 22 purpose-bred Beagles. PROCEDURES Plasmid expression cassettes encoding canine HAS2 (cHAS2) were assessed in vitro for concentration and molecular size of secreted HA. Thereafter, rAAV2-cHAS2 vectors at 3 concentrations and rAAV5-cHAS2 vectors at 1 concentration were each administered intra-articularly into the left stifle joint of 5 dogs; 2 dogs received PBS solution instead. Synovial fluid HA concentration and serum and synovial fluid titers of neutralizing antibodies against AAV capsids were measured at various points. Dogs were euthanized 28 days after treatment, and cartilage and synovium samples were collected for vector DNA and mRNA quantification and histologic examination. RESULTS Cell transfection with plasmids encoding cHAS2 resulted in an increase in production and secretion of HA in vitro. In vivo, the rAAV5-cHAS2 vector yielded uniform genome transfer and cHAS2 expression in collected synovium and cartilage samples. In contrast, rAAV2-cHAS2 vectors were detected inconsistently in synovium and cartilage samples and failed to produce clear dose-related responses. Histologic examination revealed minimal synovial inflammation in joints injected with rAAV vectors. Neutralizing antibodies against AAV capsids were detected in serum and synovial fluid samples from all vector-treated dogs. CONCLUSIONS AND CLINICAL RELEVANCE rAAV5-mediated transfer of the gene for cHAS2 into healthy joints of dogs by intra-articular injection appeared safe and resulted in vector-derived cHAS2 production by synoviocytes and chondrocytes. Whether this treatment may increase HA production by synoviocytes and chondrocytes in osteoarthritic joints remains to be determined.

Generation of Envelope-Modified Baculoviruses for Gene Delivery into Mammalian Cells.

PubMed

Hofmann, Christian

2016-01-01

Genetically modified baculoviruses can efficiently deliver and express genes in mammalian cells. The major prerequisite for the expression of a gene transferred by baculovirus is its control by a promoter that is active in mammalian cells. This chapter describes methods for producing second generation baculovirus vectors through modification of their envelope. Envelope modified baculoviruses offer additional new applications of the system, such as their use in in vivo gene delivery, targeting, and vaccination. Methods of generating a recombinant baculovirus vector with a modified envelope and its amplification and purification, including technical scale production, are discussed. A variety of notes give clues regarding specific technical procedures. Finally, methods to analyze the virus and transduction procedures are presented.

Implementing a vector surveillance-response system for chagas disease control: a 4-year field trial in Nicaragua.

PubMed

Yoshioka, Kota; Tercero, Doribel; Pérez, Byron; Nakamura, Jiro; Pérez, Lenin

2017-03-06

Chagas disease is one of the neglected tropical diseases (NTDs). International goals for its control involve elimination of vector-borne transmission. Central American countries face challenges in establishing sustainable vector control programmes, since the main vector, Triatoma dimidiata, cannot be eliminated. In 2012, the Ministry of Health in Nicaragua started a field test of a vector surveillance-response system to control domestic vector infestation. This paper reports the main findings from this pilot study. This study was carried out from 2012 to 2015 in the Municipality of Totogalpa. The Japan International Cooperation Agency provided technical cooperation in designing and monitoring the surveillance-response system until 2014. This system involved 1) vector reports by householders to health facilities, 2) data analysis and planning of responses at the municipal health centre and 3) house visits or insecticide spraying by health personnel as a response. We registered all vector reports and responses in a digital database. The collected data were used to describe and analyse the system performance in terms of amount of vector reports as well as rates and timeliness of responses. During the study period, T. dimidiata was reported 396 times. Spatiotemporal analysis identified some high-risk clusters. All houses reported to be infested were visited by health personnel in 2013 and this response rate dropped to 39% in 2015. Rates of insecticide spraying rose above 80% in 2013 but no spraying was carried out in the following 2 years. The timeliness of house visits improved significantly after the responsibility was transferred from a vector control technician to primary health care staff. We argue that the proposed vector surveillance-response system is workable within the resource-constrained health system in Nicaragua. Integration to the primary health care services was a key to improve the system performance. Continual efforts are necessary to keep adapting the surveillance-response system to the dynamic health systems. We also discuss that the goal of eliminating vector-borne transmission remains unachievable. This paper provides lessons not only for Chagas disease control in Central America, but also for control efforts for other NTDs that need a sustainable surveillance-response system to support elimination.

An improved ternary vector system for Agrobacterium-mediated rapid maize transformation.

PubMed

Anand, Ajith; Bass, Steven H; Wu, Emily; Wang, Ning; McBride, Kevin E; Annaluru, Narayana; Miller, Michael; Hua, Mo; Jones, Todd J

2018-05-01

A simple and versatile ternary vector system that utilizes improved accessory plasmids for rapid maize transformation is described. This system facilitates high-throughput vector construction and plant transformation. The super binary plasmid pSB1 is a mainstay of maize transformation. However, the large size of the base vector makes it challenging to clone, the process of co-integration is cumbersome and inefficient, and some Agrobacterium strains are known to give rise to spontaneous mutants resistant to tetracycline. These limitations present substantial barriers to high throughput vector construction. Here we describe a smaller, simpler and versatile ternary vector system for maize transformation that utilizes improved accessory plasmids requiring no co-integration step. In addition, the newly described accessory plasmids have restored virulence genes found to be defective in pSB1, as well as added virulence genes. Testing of different configurations of the accessory plasmids in combination with T-DNA binary vector as ternary vectors nearly doubles both the raw transformation frequency and the number of transformation events of usable quality in difficult-to-transform maize inbreds. The newly described ternary vectors enabled the development of a rapid maize transformation method for elite inbreds. This vector system facilitated screening different origins of replication on the accessory plasmid and T-DNA vector, and four combinations were identified that have high (86-103%) raw transformation frequency in an elite maize inbred.

A Guide to Approaching Regulatory Considerations for Lentiviral-Mediated Gene Therapies.

PubMed

White, Michael; Whittaker, Roger; Gándara, Carolina; Stoll, Elizabeth A

2017-08-01

Lentiviral vectors are increasingly the gene transfer tool of choice for gene or cell therapies, with multiple clinical investigations showing promise for this viral vector in terms of both safety and efficacy. The third-generation vector system is well characterized, effectively delivers genetic material and maintains long-term stable expression in target cells, delivers larger amounts of genetic material than other methods, is nonpathogenic, and does not cause an inflammatory response in the recipient. This report aims to help academic scientists and regulatory managers negotiate the governance framework to achieve successful translation of a lentiviral vector-based gene therapy. The focus is on European regulations and how they are administered in the United Kingdom, although many of the principles will be similar for other regions, including the United States. The report justifies the rationale for using third-generation lentiviral vectors to achieve gene delivery for in vivo and ex vivo applications; briefly summarizes the extant regulatory guidance for gene therapies, categorized as advanced therapeutic medicinal products (ATMPs); provides guidance on specific regulatory issues regarding gene therapies; presents an overview of the key stakeholders to be approached when pursuing clinical trials authorization for an ATMP; and includes a brief catalogue of the documentation required to submit an application for regulatory approval of a new gene therapy.

A Guide to Approaching Regulatory Considerations for Lentiviral-Mediated Gene Therapies

PubMed Central

White, Michael; Whittaker, Roger; Gándara, Carolina; Stoll, Elizabeth A.

2017-01-01

Lentiviral vectors are increasingly the gene transfer tool of choice for gene or cell therapies, with multiple clinical investigations showing promise for this viral vector in terms of both safety and efficacy. The third-generation vector system is well characterized, effectively delivers genetic material and maintains long-term stable expression in target cells, delivers larger amounts of genetic material than other methods, is nonpathogenic, and does not cause an inflammatory response in the recipient. This report aims to help academic scientists and regulatory managers negotiate the governance framework to achieve successful translation of a lentiviral vector-based gene therapy. The focus is on European regulations and how they are administered in the United Kingdom, although many of the principles will be similar for other regions, including the United States. The report justifies the rationale for using third-generation lentiviral vectors to achieve gene delivery for in vivo and ex vivo applications; briefly summarizes the extant regulatory guidance for gene therapies, categorized as advanced therapeutic medicinal products (ATMPs); provides guidance on specific regulatory issues regarding gene therapies; presents an overview of the key stakeholders to be approached when pursuing clinical trials authorization for an ATMP; and includes a brief catalogue of the documentation required to submit an application for regulatory approval of a new gene therapy. PMID:28817344

Back to basics: pBR322 and protein expression systems in E. coli.

PubMed

Balbás, Paulina; Bolívar, Francisco

2004-01-01

The extensive variety of plasmid-based expression systems in E. coli resulted from the fact that there is no single strategy for achieving maximal expression of every cloned gene. Although a number of strategies have been implemented to deal with problems associated to gene transcription and translation, protein folding, secretion, location, posttranslational modifications, particularities of different strains, and the like and more integrated processes have been developed, the basic plasmid-borne elements and their interaction with the particular host strain will influence the overall expression system and final productivity. Plasmid vector pBR322 is a well-established multipurpose cloning vector in laboratories worldwide, and a large number of derivatives have been created for specific applications and research purposes, including gene expression in its natural host, E. coli, and few other bacteria. The early characterization of the molecule, including its nucleotide sequence, replication and maintenance mechanisms, and determination of its coding regions, accounted for its success, not only as a universal cloning vector, but also as a provider of genes and an origin of replication for other intraspecies vectors. Since the publication of the aforementioned reviews, novel discoveries pertaining to these issues have appeared in the literature that deepen the understanding of the plasmid's features, behavior, and impact in gene expression systems, as well as some important strain characteristics that affect plasmid replication and stability. The objectives of this review include updating and discussing the new information about (1) the replication and maintenance of pBR322; (2) the host-related modulation mechanisms of plasmid replication; (3) the effects of growth rate on replication control, stability, and recombinant gene expression; (4) ways for plasmid amplification and elimination. Finally, (5) a summary of novel ancillary studies about pBR322 is presented.

Fat grafting as a vehicle for the delivery of recombinant adenoassociated viral vectors to achieve gene modification of muscle flaps.

PubMed

Carruthers, Katherine H; During, Matthew J; Muravlev, Alexander; Wang, Chuansong; Kocak, Ergun

2013-06-01

The combination of gene therapy and plastic surgery may have the potential to improve the specificity that is needed to achieve clinically applicable treatment regimens. Our goal was to develop a method for gene modification that would yield sustainable production of gene products but would be less time consuming than existing protocols. An adenoassociated virus was used to deliver gene products to pectoralis muscle flaps. Gene modification was accomplished via either direct injection or novel fat grafting techniques. The production of gene product was observable by both in vivo imaging and immunohistochemical staining. Gene products were not detected in tissues that were not in contact with the fat grafts that were incubated with the viral vector, indicating that the transduction stayed local to the flap. Using novel recombinant adenoassociated virus vectors, we have developed a method for gene delivery that is highly efficient and applicable to muscle flaps.

Improved annotation of the insect vector of citrus greening disease: Biocuration by a diverse genomics community

USDA-ARS?s Scientific Manuscript database

The Asian citrus psyllid (Diaphorina citri Kuwayama) is the insect vector of the bacterium Candidatus Liberibacter asiaticus (CLas), the pathogen associated with citrus Huanglongbing (HLB, citrus greening). HLB threatens citrus production worldwide. Suppression or reduction of the insect vector usin...

Transient Expression of Lumbrokinase (PI239) in Tobacco (Nicotiana tabacum) Using a Geminivirus-Based Single Replicon System Dissolves Fibrin and Blood Clots.

PubMed

Dickey, Alexia; Wang, Nan; Cooper, Edwin; Tull, Lauren; Breedlove, Drew; Mason, Hugh; Liu, Dehu; Wang, Kevin Yueju

2017-01-01

Lumbrokinases, a group of fibrinolytic enzymes extracted from earthworm, have been widely used to prevent and treat various cardiovascular diseases. They specifically target fibrin to effectively degrade thrombi without major side effects. Plant expression systems are becoming potential alternative expression platforms for producing pharmaceutical proteins. In this work, a lumbrokinase (PI239) was produced from a plant system. Both wild-type (WT) and plant codon-optimized (OP) PI239 gene sequences were synthesized and cloned into a geminivirus-based single-vector DNA replicon system. Both vectors were independently expressed in tobacco (Nicotiana tabacum) leaves transiently by agroinfiltration. Overexpressed PI239 resulted in sudden tissue necrosis 3 days after infiltration. Remaining proteins were purified through His-tag affinity chromatography and analyzed with SDS-PAGE and Western blot methods. Purified PI239 successfully degraded artificial fibrin with relative activity of 13,400 U/mg when compared with commercial lumbrokinase product. In vitro tests demonstrated that plant-derived PI239 dissolved human blood clots and that the plant expression system is capable of producing functional PI239.

The p40 Subunit of Interleukin (IL)-12 Promotes Stabilization and Export of the p35 Subunit

PubMed Central

Jalah, Rashmi; Rosati, Margherita; Ganneru, Brunda; Pilkington, Guy R.; Valentin, Antonio; Kulkarni, Viraj; Bergamaschi, Cristina; Chowdhury, Bhabadeb; Zhang, Gen-Mu; Beach, Rachel Kelly; Alicea, Candido; Broderick, Kate E.; Sardesai, Niranjan Y.; Pavlakis, George N.; Felber, Barbara K.

2013-01-01

IL-12 is a 70-kDa heterodimeric cytokine composed of the p35 and p40 subunits. To maximize cytokine production from plasmid DNA, molecular steps controlling IL-12p70 biosynthesis at the posttranscriptional and posttranslational levels were investigated. We show that the combination of RNA/codon-optimized gene sequences and fine-tuning of the relative expression levels of the two subunits within a cell resulted in increased production of the IL-12p70 heterodimer. We found that the p40 subunit plays a critical role in enhancing the stability, intracellular trafficking, and export of the p35 subunit. This posttranslational regulation mediated by the p40 subunit is conserved in mammals. Based on these findings, dual gene expression vectors were generated, producing an optimal ratio of the two subunits, resulting in a ∼1 log increase in human, rhesus, and murine IL-12p70 production compared with vectors expressing the wild type sequences. Such optimized DNA plasmids also produced significantly higher levels of systemic bioactive IL-12 upon in vivo DNA delivery in mice compared with plasmids expressing the wild type sequences. A single therapeutic injection of an optimized murine IL-12 DNA plasmid showed significantly more potent control of tumor development in the B16 melanoma cancer model in mice. Therefore, the improved IL-12p70 DNA vectors have promising potential for in vivo use as molecular vaccine adjuvants and in cancer immunotherapy. PMID:23297419

Detection of osteoclastic cell-cell fusion through retroviral vector packaging.

PubMed

Kondo, Takako; Ikeda, Kyoji; Matsuo, Koichi

2004-11-01

Cell-cell fusion generates multinucleated cells such as osteoclasts in bone, myotubes in muscle, and trophoblasts in placenta. Molecular details governing these fusion processes are still largely unknown. As a step toward identification of fusogenic genes, we tested the concept that retroviral vectors can be packaged as a result of cell-cell fusion. First, we introduced replication-deficient retroviral vectors expressing mCAT-1, which mediates fusogenic interaction with the retroviral envelope protein Env, into Chinese hamster ovary (CHO) cells to generate vector cells. Plasmids expressing virion proteins Gag, Pol, and Env were introduced into a separate culture of CHO cells to generate packaging cells. Co-culturing vector and packaging cells resulted in production of infectious retroviruses carrying the mCAT-1 gene as a consequence of cell-cell fusion. Second, we introduced a retroviral vector into primary osteoclast precursors and co-cultured them with established osteoclast precursor RAW264.7 cells, which turned out to harbor packaging activity. Packaged retroviral vector was detected in culture supernatants only where the osteoclast differentiation factor receptor activator for NF-kappaB ligand (RANKL) induced fusion between these two cell types. These data suggest that retrovirus production can occur as a result of cell-cell fusion. This provides a novel approach for isolating and characterizing fusogenic genes using retroviral expression vectors.

A Partial E3 Deletion in Replication-Defective Adenoviral Vectors Allows for Stable Expression of Potentially Toxic Transgene Products.

PubMed

Haut, Larissa H; Gill, Amanda L; Kurupati, Raj K; Bian, Ang; Li, Yan; Giles-Davis, Wynetta; Xiang, Zhiquan; Zhou, Xiang Yang; Ertl, Hildegund C J

2016-10-01

Adenovirus (Ad) is used extensively for construction of viral vectors, most commonly with deletion in its E1 and/or E3 genomic regions. Previously, our attempts to insert envelope proteins (Env) of HIV-1 into such vectors based on chimpanzee-derived Ad (AdC) viruses were thwarted. Here, we describe that genetic instability of an E1- and E3-deleted AdC vector of serotype C6 expressing Env of HIV-1 can be overcome by reinsertion of E3 sequences with anti-apoptotic activities. This partial E3 deletion presumably delays premature death of HEK-293 packaging cell lines due to Env-induced cell apoptosis. The same partial E3 deletion also allows for the generation of stable glycoprotein 140 (gp140)- and gp160-expressing Ad vectors based on AdC7, a distinct AdC serotype. Env-expressing AdC vectors containing the partial E3 deletion are genetically stable upon serial cell culture passaging, produce yields comparable to those of other AdC vectors, and induce transgene product-specific antibody responses in mice. A partial E3 deletion thereby allows expansion of the repertoire of transgenes that can be expressed by Ad vectors.

Influence of vector dose on factor IX-specific T and B cell responses in muscle-directed gene therapy.

PubMed

Herzog, Roland W; Fields, Paul A; Arruda, Valder R; Brubaker, Jeff O; Armstrong, Elina; McClintock, Darryl; Bellinger, Dwight A; Couto, Linda B; Nichols, Timothy C; High, Katherine A

2002-07-20

Intramuscular injection of an adeno-associated virus (AAV) vector has resulted in vector dose-dependent, stable expression of canine factor IX (cF.IX) in hemophilia B dogs with an F.IX missense mutation (Herzog et al., Nat. Med. 1999;5:56-63). The use of a species-specific transgene allowed us to study risks and characteristics of antibody formation against the therapeutic transgene product. We analyzed seven dogs that had been injected at a single time point at multiple intramuscular sites with varying vector doses (dose per kilogram, dose per animal, dose per site). Comparison of individual animals suggests an increased likelihood of inhibitory anti-cF.IX (inhibitor) development with increased vector doses, with dose per site showing the strongest correlation with the risk of inhibitor formation. In six of seven animals, such immune responses were either absent or transient, and therefore did not prevent sustained systemic expression of cF.IX. Transient inhibitory/neutralizing anti-cF.IX responses occurred at vector doses of 2 x 10(12)/site, whereas a 6-fold higher dose resulted in a longer lasting, higher titer inhibitor. Anti-cF.IX was efficiently blocked in an eighth animal that was injected with a high vector dose per site, but in addition received transient immune suppression. Inhibitor formation was characterized by synthesis of two IgG subclasses and in vitro proliferation of lymphocytes to cF.IX antigen, indicating a helper T cell-dependent mechanism. Anti-cF.IX formation is likely influenced by the extent of local antigen presentation and may be avoided by limited vector doses or by transient immune modulation.

Recent Developments In Theory Of Balanced Linear Systems

NASA Technical Reports Server (NTRS)

Gawronski, Wodek

1994-01-01

Report presents theoretical study of some issues of controllability and observability of system represented by linear, time-invariant mathematical model of the form. x = Ax + Bu, y = Cx + Du, x(0) = xo where x is n-dimensional vector representing state of system; u is p-dimensional vector representing control input to system; y is q-dimensional vector representing output of system; n,p, and q are integers; x(0) is intial (zero-time) state vector; and set of matrices (A,B,C,D) said to constitute state-space representation of system.

A static investigation of the thrust vectoring system of the F/A-18 high-alpha research vehicle

NASA Technical Reports Server (NTRS)

Mason, Mary L.; Capone, Francis J.; Asbury, Scott C.

1992-01-01

A static (wind-off) test was conducted in the static test facility of the Langley 16-foot Transonic Tunnel to evaluate the vectoring capability and isolated nozzle performance of the proposed thrust vectoring system of the F/A-18 high alpha research vehicle (HARV). The thrust vectoring system consisted of three asymmetrically spaced vanes installed externally on a single test nozzle. Two nozzle configurations were tested: A maximum afterburner-power nozzle and a military-power nozzle. Vane size and vane actuation geometry were investigated, and an extensive matrix of vane deflection angles was tested. The nozzle pressure ratios ranged from two to six. The results indicate that the three vane system can successfully generate multiaxis (pitch and yaw) thrust vectoring. However, large resultant vector angles incurred large thrust losses. Resultant vector angles were always lower than the vane deflection angles. The maximum thrust vectoring angles achieved for the military-power nozzle were larger than the angles achieved for the maximum afterburner-power nozzle.

Formation of AAV Single Stranded DNA Genome from a Circular Plasmid in Saccharomyces cerevisiae

PubMed Central

Cervelli, Tiziana; Backovic, Ana; Galli, Alvaro

2011-01-01

Adeno-associated virus (AAV)-based vectors are promising tools for targeted transfer in gene therapy studies. Many efforts have been accomplished to improve production and purification methods. We thought to develop a simple eukaryotic system allowing AAV replication which could provide an excellent opportunity for studying AAV biology and, more importantly, for AAV vector production. It has been shown that yeast Saccharomyces cerevisiae is able to replicate and form the capsid of many viruses. We investigated the ability of the yeast Saccharomyces cerevisiae to carry out the replication of a recombinant AAV (rAAV). When a plasmid containing a rAAV genome in which the cap gene was replaced with the S. cerevisiae URA3 gene, was co-transformed in yeast with a plasmid expressing Rep68, a significant number of URA3+ clones were scored (more than 30-fold over controls). Molecular analysis of low molecular weight DNA by Southern blotting revealed that single stranded DNA is formed and that the plasmid is entirely replicated. The ssDNA contains the ITRs, URA3 gene and also vector sequences suggesting the presence of two distinct molecules. Its formation was dependent on Rep68 expression and ITR. These data indicate that DNA is not obtained by the canonical AAV replication pathway. PMID:21853137

Formation of AAV single stranded DNA genome from a circular plasmid in Saccharomyces cerevisiae.

PubMed

Cervelli, Tiziana; Backovic, Ana; Galli, Alvaro

2011-01-01

Adeno-associated virus (AAV)-based vectors are promising tools for targeted transfer in gene therapy studies. Many efforts have been accomplished to improve production and purification methods. We thought to develop a simple eukaryotic system allowing AAV replication which could provide an excellent opportunity for studying AAV biology and, more importantly, for AAV vector production. It has been shown that yeast Saccharomyces cerevisiae is able to replicate and form the capsid of many viruses. We investigated the ability of the yeast Saccharomyces cerevisiae to carry out the replication of a recombinant AAV (rAAV). When a plasmid containing a rAAV genome in which the cap gene was replaced with the S. cerevisiae URA3 gene, was co-transformed in yeast with a plasmid expressing Rep68, a significant number of URA3(+) clones were scored (more than 30-fold over controls). Molecular analysis of low molecular weight DNA by Southern blotting revealed that single stranded DNA is formed and that the plasmid is entirely replicated. The ssDNA contains the ITRs, URA3 gene and also vector sequences suggesting the presence of two distinct molecules. Its formation was dependent on Rep68 expression and ITR. These data indicate that DNA is not obtained by the canonical AAV replication pathway.

Shoelace Formula: Connecting the Area of a Polygon and the Vector Cross Product

ERIC Educational Resources Information Center

Lee, Younhee; Lim, Woong

2017-01-01

Understanding how one representation connects to another and how the essential ideas in that relationship are generalized can result in a mathematical theorem or a formula. In this article, the authors demonstrate this process by connecting a vector cross product in algebraic form to a geometric representation and applying a key mathematical idea…

Genetic modification of hematopoietic stem cells with nonviral systems: past progress and future prospects.

PubMed

Papapetrou, E P; Zoumbos, N C; Athanassiadou, A

2005-10-01

Serious unwanted complications provoked by retroviral gene transfer into hematopoietic stem cells (HSCs) have recently raised the need for the development and assessment of alternative gene transfer vectors. Within this context, nonviral gene transfer systems are attracting increasing interest. Their main advantages include low cost, ease of handling and large-scale production, large packaging capacity and, most importantly, biosafety. While nonviral gene transfer into HSCs has been restricted in the past by poor transfection efficiency and transient maintenance, in recent years, biotechnological developments are converting nonviral transfer into a realistic approach for genetic modification of cells of hematopoietic origin. Herein we provide an overview of past accomplishments in the field of nonviral gene transfer into hematopoietic progenitor/stem cells and we point at future challenges. We argue that episomally maintained self-replicating vectors combined with physical methods of delivery show the greatest promise among nonviral gene transfer strategies for the treatment of disorders of the hematopoietic system.

An efficient numerical method for the solution of the problem of elasticity for 3D-homogeneous elastic medium with cracks and inclusions

NASA Astrophysics Data System (ADS)

Kanaun, S.; Markov, A.

2017-06-01

An efficient numerical method for solution of static problems of elasticity for an infinite homogeneous medium containing inhomogeneities (cracks and inclusions) is developed. Finite number of heterogeneous inclusions and planar parallel cracks of arbitrary shapes is considered. The problem is reduced to a system of surface integral equations for crack opening vectors and volume integral equations for stress tensors inside the inclusions. For the numerical solution of these equations, a class of Gaussian approximating functions is used. The method based on these functions is mesh free. For such functions, the elements of the matrix of the discretized system are combinations of explicit analytical functions and five standard 1D-integrals that can be tabulated. Thus, the numerical integration is excluded from the construction of the matrix of the discretized problem. For regular node grids, the matrix of the discretized system has Toeplitz's properties, and Fast Fourier Transform technique can be used for calculation matrix-vector products of such matrices.

Parvovirus-like particles as vaccine vectors.

PubMed

Casal, J I; Rueda, P; Hurtado, A

1999-09-01

A wide array of systems have been developed to improve "classic" vaccines. The use of small polypeptides able to elicit potent antibody and cytotoxic responses seems to have enormous potential in the design of safer vaccines. While peptide coupling to large soluble proteins such as keyhole limpet hemocyanin is the current method of choice for eliciting antibody responses and insertion in live viruses for cytotoxic T-lymphocyte responses, alternative cheaper and/or safer methods will clearly be required in the future. Virus-like particles constitute very immunogenic molecules that allow for covalent coupling of the epitopes of interest in a simple way. In this article, we detail the methodology employed for the preparation of efficient virus vectors as delivery systems. We used parvovirus as the model for the design of new vaccine vectors. Recently parvovirus-like particles have been engineered to express foreign polypeptides in certain positions, resulting in the production of large quantities of highly immunogenic peptides, and to induce strong antibody, helper-T-cell, and cytotoxic T-lymphocyte responses. We discuss the different alternatives and the necessary steps to carry out this process, placing special emphasis on the flow of decisions that need to be made during the project. Copyright 1999 Academic Press.

Distribution of AAV8 particles in cell lysates and culture media changes with time and is dependent on the recombinant vector

PubMed Central

Piras, Bryan A; Drury, Jason E; Morton, Christopher L; Spence, Yunyu; Lockey, Timothy D; Nathwani, Amit C; Davidoff, Andrew M; Meagher, Michael M

2016-01-01

With clinical trials ongoing, efficient clinical production of adeno-associated virus (AAV) to treat large numbers of patients remains a challenge. We compared distribution of AAV8 packaged with Factor VIII (FVIII) in cell culture media and lysates on days 3, 5, 6, and 7 post-transfection and found increasing viral production through day 6, with the proportion of viral particles in the media increasing from 76% at day 3 to 94% by day 7. Compared to FVIII, AAV8 packaged with Factor IX and Protective Protein/Cathepsin A vectors demonstrated a greater shift from lysate towards media from day 3 to 6, implying that particle distribution is dependent on recombinant vector. Larger-scale productions showed that the ratio of full-to-empty AAV particles is similar in media and lysate, and that AAV harvested on day 6 post-transfection provides equivalent function in mice compared to AAV harvested on day 3. This demonstrates that AAV8 production can be optimized by prolonging the duration of culture post-transfection, and simplified by allowing harvest of media only, with disposal of cells that contain 10% or less of total vector yield. Additionally, the difference in particle distribution with different expression cassettes implies a recombinant vector-dependent processing mechanism which should be taken into account during process development. PMID:27069949

Effects of energy conservation on equilibrium properties of hot asymmetric nuclear matter

NASA Astrophysics Data System (ADS)

Zhang, Zhen; Ko, Che Ming

2018-01-01

Based on the relativistic Vlasov-Uehling-Uhlenbeck transport model, which includes relativistic scalar and vector potentials on baryons, we consider an N -Δ -π system in a box with periodic boundary conditions to study the effects of energy conservation in particle production and absorption processes on the equilibrium properties of the system. The density and temperature of the matter in the box are taken to be similar to the hot dense matter formed in heavy ion collisions at intermediate energies. We find that to maintain the equilibrium numbers of N ,Δ , and π , which depend on the mean-field potentials of N and Δ , we must include these potentials in the energy conservation condition that determines the momenta of outgoing particles after a scattering or decay process. We further find that the baryon scalar potentials mainly affect the Δ and pion equilibrium numbers, while the baryon vector potentials have considerable effect on the effective charged pion ratio at equilibrium. Our results thus indicate that it is essential to include in the transport model the effect of potentials in the energy conservation of a scattering or decay process, which is ignored in most transport models, for studying pion production in heavy ion collisions.

The generalized formula for angular velocity vector of the moving coordinate system

NASA Astrophysics Data System (ADS)

Ermolin, Vladislav S.; Vlasova, Tatyana V.

2018-05-01

There are various ways for introducing the concept of the instantaneous angular velocity vector. In this paper we propose a method based on introducing of this concept by construction of the solution for the system of kinematic equations. These equations connect the function vectors defining the motion of the basis, and their derivatives. Necessary and sufficient conditions for the existence and uniqueness of the solution of this system are established. The instantaneous angular velocity vector is a solution of the algebraic system of equations. It is built explicitly. The derived formulas for the angular velocity vector generalize the earlier results, both for a basis of an affine oblique coordinate system and for an orthonormal basis.

Efficient Single Tobamoviral Vector-Based Bioproduction of Broadly Neutralizing Anti-HIV-1 Monoclonal Antibody VRC01 in Nicotiana benthamiana Plants and Utility of VRC01 in Combination Microbicides

PubMed Central

Hamorsky, Krystal Teasley; Grooms-Williams, Tiffany W.; Husk, Adam S.; Bennett, Lauren J.; Palmer, Kenneth E.

2013-01-01

Broadly neutralizing monoclonal antibodies (bnMAbs) may offer powerful tools for HIV-1 preexposure prophylaxis, such as topical microbicides. However, this option is hampered due to expensive MAb biomanufacturing based on mammalian cell culture. To address this issue, we developed a new production system for bnMAb VRC01 in Nicotiana benthamiana plants using a tobamovirus replicon vector. Unlike conventional two-vector-based expression, this system was designed to overexpress full-length IgG1 from a single polypeptide by means of kex2p-like enzyme recognition sites introduced between the heavy and light chains. An enzyme-linked immunosorbent assay (ELISA) revealed that gp120-binding VRC01 IgG1 was maximally accumulated on 5 to 7 days following vector inoculation, yielding ∼150 mg of the bnMAb per kg of fresh leaf material. The plant-made VRC01 (VRC01p) was efficiently purified by protein A affinity followed by hydrophobic-interaction chromatography. ELISA, surface plasmon resonance, and an HIV-1 neutralization assay demonstrated that VRC01p has gp120-binding affinity and HIV-1-neutralization capacity virtually identical to the human-cell-produced counterpart. To advance VRC01p's use in topical microbicides, we analyzed combinations of the bnMAb with other microbicide candidates holding distinct antiviral mechanisms in an HIV-1 neutralization assay. VRC01p exhibited clear synergy with the antiviral lectin griffithsin, the CCR5 antagonist maraviroc, and the reverse transcriptase inhibitor tenofovir in multiple CCR5-tropic HIV-1 strains from clades A, B, and C. In summary, VRC01p is amenable to robust, rapid, and large-scale production and may be developed as an active component in combination microbicides with other anti-HIV agents such as antiviral lectins, CCR5 antagonists, and reverse transcriptase inhibitors. PMID:23403432

Search for vector-like quark production in the lepton+jets and dilepton+jets final states using 5.4 fb -1 of Run II data

DOE Office of Scientific and Technical Information (OSTI.GOV)

Caughron, Seth

2011-01-01

The Standard Model of particle physics provides an excellent description of particle interactions at energies up to ~1 TeV, but it is expected to fail above that scale. Multiple models developed to describe phenomena above the TeV scale predict the existence of very massive, vector-like quarks. A search for single electroweak production of such particles in pmore » $$\\bar{p}$$ collisions at a center-of-mass energy of 1.96 TeV is performed in the W+jets and Z+jets channels. The data were collected by the DØ detector at the Fermilab Tevatron Collider and correspond to an integrated luminosity of 5.4 fb -1. Events consistent with a heavy object decaying to a vector boson and a jet are selected. We observe no significant excess in comparison to the background prediction and set 95% confidence level upper limits on production cross sections for vector-like quarks decaying to W+jet and Z+jet. Assuming a vector-like quark -- standard model quark coupling parameter $$\\tilde{κ}$$ qQ of unity, we exclude vector-like quarks with mass below 693 GeV for decays to W+jet and mass below 449 GeV for decays to Z+jet. These represent the most sensitive limits to date.« less

Amino acid "little Big Bang": representing amino acid substitution matrices as dot products of Euclidian vectors.

PubMed

Zimmermann, Karel; Gibrat, Jean-François

2010-01-04

Sequence comparisons make use of a one-letter representation for amino acids, the necessary quantitative information being supplied by the substitution matrices. This paper deals with the problem of finding a representation that provides a comprehensive description of amino acid intrinsic properties consistent with the substitution matrices. We present a Euclidian vector representation of the amino acids, obtained by the singular value decomposition of the substitution matrices. The substitution matrix entries correspond to the dot product of amino acid vectors. We apply this vector encoding to the study of the relative importance of various amino acid physicochemical properties upon the substitution matrices. We also characterize and compare the PAM and BLOSUM series substitution matrices. This vector encoding introduces a Euclidian metric in the amino acid space, consistent with substitution matrices. Such a numerical description of the amino acid is useful when intrinsic properties of amino acids are necessary, for instance, building sequence profiles or finding consensus sequences, using machine learning algorithms such as Support Vector Machine and Neural Networks algorithms.

Design and construction of 2A peptide-linked multicistronic vectors.

PubMed

Szymczak-Workman, Andrea L; Vignali, Kate M; Vignali, Dario A A

2012-02-01

The need for reliable, multicistronic vectors for multigene delivery is at the forefront of biomedical technology. This article describes the design and construction of 2A peptide-linked multicistronic vectors, which can be used to express multiple proteins from a single open reading frame (ORF). The small 2A peptide sequences, when cloned between genes, allow for efficient, stoichiometric production of discrete protein products within a single vector through a novel "cleavage" event within the 2A peptide sequence. Expression of more than two genes using conventional approaches has several limitations, most notably imbalanced protein expression and large size. The use of 2A peptide sequences alleviates these concerns. They are small (18-22 amino acids) and have divergent amino-terminal sequences, which minimizes the chance for homologous recombination and allows for multiple, different 2A peptide sequences to be used within a single vector. Importantly, separation of genes placed between 2A peptide sequences is nearly 100%, which allows for stoichiometric and concordant expression of the genes, regardless of the order of placement within the vector.

Higgs constraints from vector boson fusion and scattering

DOE PAGES

Campbell, John M.; Ellis, R. Keith

2015-04-07

We present results on 4-lepton + 2-jet production, the partonic processes most commonly described as vector boson pair production in the Vector Boson Fusion (VBF) mode. That final state contains diagrams that are mediated by Higgs boson exchange. We focus particularly on the high-mass behaviour of the Higgs boson mediated diagrams, which unlike on-shell production, gives information about the Higgs couplings without assumptions on the Higgs boson total width. We assess the sensitivity of the high-mass region to Higgs coupling strengths, considering all vector boson pair channels, W - W +, W ± W ±, W ± Z and ZZ.more » Because of the small background, the most promising mode is W + W + which has sensitivity to Higgs couplings because of Higgs boson exchange in the t-channel. Furthermore, using the Caola-Melnikov (CM) method, the off-shell couplings can be interpreted as bounds on the Higgs boson total width. We estimate the bound that can be obtained with current data, as well as the bounds that could be obtained at √s=13 TeV in the VBF channel for data samples of 100 and 300 fb -1. The CM method has already been successfully applied in the gluon fusion (GGF) production channel. The VBF production channel gives important complementary information, because both production and decay of the Higgs boson occur already at tree graph level.« less

AAV vector-mediated secretion of chondroitinase provides a sensitive tracer for axonal arborisations.

PubMed

Alves, João Nuno; Muir, Elizabeth M; Andrews, Melissa R; Ward, Anneliese; Michelmore, Nicholas; Dasgupta, Debayan; Verhaagen, Joost; Moloney, Elizabeth B; Keynes, Roger J; Fawcett, James W; Rogers, John H

2014-04-30

As part of a project to express chondroitinase ABC (ChABC) in neurons of the central nervous system, we have inserted a modified ChABC gene into an adeno-associated viral (AAV) vector and injected it into the vibrissal motor cortex in adult rats to determine the extent and distribution of expression of the enzyme. A similar vector for expression of green fluorescent protein (GFP) was injected into the same location. For each vector, two versions with minor differences were used, giving similar results. After 4 weeks, the brains were stained to show GFP and products of chondroitinase digestion. Chondroitinase was widely expressed, and the AAV-ChABC and AAV-GFP vectors gave similar expression patterns in many respects, consistent with the known projections from the directly transduced neurons in vibrissal motor cortex and adjacent cingulate cortex. In addition, diffusion of vector to deeper neuronal populations led to labelling of remote projection fields which was much more extensive with AAV-ChABC than with AAV-GFP. The most notable of these populations are inferred to be neurons of cortical layer 6, projecting widely in the thalamus, and neurons of the anterior pole of the hippocampus, projecting through most of the hippocampus. We conclude that, whereas GFP does not label the thinnest axonal branches of some neuronal types, chondroitinase is efficiently secreted from these arborisations and enables their extent to be sensitively visualised. After 12 weeks, chondroitinase expression was undiminished. Copyright © 2014 Elsevier B.V. All rights reserved.

Modification of the Creator recombination system for proteomics applications--improved expression by addition of splice sites.

PubMed

Colwill, Karen; Wells, Clark D; Elder, Kelly; Goudreault, Marilyn; Hersi, Kadija; Kulkarni, Sarang; Hardy, W Rod; Pawson, Tony; Morin, Gregg B

2006-03-06

Recombinational systems have been developed to rapidly shuttle Open Reading Frames (ORFs) into multiple expression vectors in order to analyze the large number of cDNAs available in the post-genomic era. In the Creator system, an ORF introduced into a donor vector can be transferred with Cre recombinase to a library of acceptor vectors optimized for different applications. Usability of the Creator system is impacted by the ability to easily manipulate DNA, the number of acceptor vectors for downstream applications, and the level of protein expression from Creator vectors. To date, we have developed over 20 novel acceptor vectors that employ a variety of promoters and epitope tags commonly employed for proteomics applications and gene function analysis. We also made several enhancements to the donor vectors including addition of different multiple cloning sites to allow shuttling from pre-existing vectors and introduction of the lacZ alpha reporter gene to allow for selection. Importantly, in order to ameliorate any effects on protein expression of the loxP site between a 5' tag and ORF, we introduced a splicing event into our expression vectors. The message produced from the resulting 'Creator Splice' vector undergoes splicing in mammalian systems to remove the loxP site. Upon analysis of our Creator Splice constructs, we discovered that protein expression levels were also significantly increased. The development of new donor and acceptor vectors has increased versatility during the cloning process and made this system compatible with a wider variety of downstream applications. The modifications introduced in our Creator Splice system were designed to remove extraneous sequences due to recombination but also aided in downstream analysis by increasing protein expression levels. As a result, we can now employ epitope tags that are detected less efficiently and reduce our assay scale to allow for higher throughput. The Creator Splice system appears to be an extremely useful tool for proteomics.

Modification of the Creator recombination system for proteomics applications – improved expression by addition of splice sites

PubMed Central

Colwill, Karen; Wells, Clark D; Elder, Kelly; Goudreault, Marilyn; Hersi, Kadija; Kulkarni, Sarang; Hardy, W Rod; Pawson, Tony; Morin, Gregg B

2006-01-01

Background Recombinational systems have been developed to rapidly shuttle Open Reading Frames (ORFs) into multiple expression vectors in order to analyze the large number of cDNAs available in the post-genomic era. In the Creator system, an ORF introduced into a donor vector can be transferred with Cre recombinase to a library of acceptor vectors optimized for different applications. Usability of the Creator system is impacted by the ability to easily manipulate DNA, the number of acceptor vectors for downstream applications, and the level of protein expression from Creator vectors. Results To date, we have developed over 20 novel acceptor vectors that employ a variety of promoters and epitope tags commonly employed for proteomics applications and gene function analysis. We also made several enhancements to the donor vectors including addition of different multiple cloning sites to allow shuttling from pre-existing vectors and introduction of the lacZ alpha reporter gene to allow for selection. Importantly, in order to ameliorate any effects on protein expression of the loxP site between a 5' tag and ORF, we introduced a splicing event into our expression vectors. The message produced from the resulting 'Creator Splice' vector undergoes splicing in mammalian systems to remove the loxP site. Upon analysis of our Creator Splice constructs, we discovered that protein expression levels were also significantly increased. Conclusion The development of new donor and acceptor vectors has increased versatility during the cloning process and made this system compatible with a wider variety of downstream applications. The modifications introduced in our Creator Splice system were designed to remove extraneous sequences due to recombination but also aided in downstream analysis by increasing protein expression levels. As a result, we can now employ epitope tags that are detected less efficiently and reduce our assay scale to allow for higher throughput. The Creator Splice system appears to be an extremely useful tool for proteomics. PMID:16519801

Verification of 2A peptide cleavage.

PubMed

Szymczak-Workman, Andrea L; Vignali, Kate M; Vignali, Dario A A

2012-02-01

The need for reliable, multicistronic vectors for multigene delivery is at the forefront of biomedical technology. It is now possible to express multiple proteins from a single open reading frame (ORF) using 2A peptide-linked multicistronic vectors. These small sequences, when cloned between genes, allow for efficient, stoichiometric production of discrete protein products within a single vector through a novel "cleavage" event within the 2A peptide sequence. The easiest and most effective way to assess 2A cleavage is to perform transient transfection of 293T cells (human embryonic kidney cells) followed by western blot analysis, as described in this protocol. 293T cells are easy to grow and can be efficiently transfected with a variety of vectors. Cleavage can be assessed by detection with antibodies against the target proteins or anti-2A serum.

Quick and clean cloning.

PubMed

Thieme, Frank; Marillonnet, Sylvestre

2014-01-01

Identification of unknown sequences that flank known sequences of interest requires PCR amplification of DNA fragments that contain the junction between the known and unknown flanking sequences. Since amplified products often contain a mixture of specific and nonspecific products, the quick and clean (QC) cloning procedure was developed to clone specific products only. QC cloning is a ligation-independent cloning procedure that relies on the exonuclease activity of T4 DNA polymerase to generate single-stranded extensions at the ends of the vector and insert. A specific feature of QC cloning is the use of vectors that contain a sequence called catching sequence that allows cloning specific products only. QC cloning is performed by a one-pot incubation of insert and vector in the presence of T4 DNA polymerase at room temperature for 10 min followed by direct transformation of the incubation mix in chemo-competent Escherichia coli cells.

Portable implementation model for CFD simulations. Application to hybrid CPU/GPU supercomputers

NASA Astrophysics Data System (ADS)

Oyarzun, Guillermo; Borrell, Ricard; Gorobets, Andrey; Oliva, Assensi

2017-10-01

Nowadays, high performance computing (HPC) systems experience a disruptive moment with a variety of novel architectures and frameworks, without any clarity of which one is going to prevail. In this context, the portability of codes across different architectures is of major importance. This paper presents a portable implementation model based on an algebraic operational approach for direct numerical simulation (DNS) and large eddy simulation (LES) of incompressible turbulent flows using unstructured hybrid meshes. The strategy proposed consists in representing the whole time-integration algorithm using only three basic algebraic operations: sparse matrix-vector product, a linear combination of vectors and dot product. The main idea is based on decomposing the nonlinear operators into a concatenation of two SpMV operations. This provides high modularity and portability. An exhaustive analysis of the proposed implementation for hybrid CPU/GPU supercomputers has been conducted with tests using up to 128 GPUs. The main objective consists in understanding the challenges of implementing CFD codes on new architectures.

Solution of nonlinear time-dependent PDEs through componentwise approximation of matrix functions

NASA Astrophysics Data System (ADS)

Cibotarica, Alexandru; Lambers, James V.; Palchak, Elisabeth M.

2016-09-01

Exponential propagation iterative (EPI) methods provide an efficient approach to the solution of large stiff systems of ODEs, compared to standard integrators. However, the bulk of the computational effort in these methods is due to products of matrix functions and vectors, which can become very costly at high resolution due to an increase in the number of Krylov projection steps needed to maintain accuracy. In this paper, it is proposed to modify EPI methods by using Krylov subspace spectral (KSS) methods, instead of standard Krylov projection methods, to compute products of matrix functions and vectors. Numerical experiments demonstrate that this modification causes the number of Krylov projection steps to become bounded independently of the grid size, thus dramatically improving efficiency and scalability. As a result, for each test problem featured, as the total number of grid points increases, the growth in computation time is just below linear, while other methods achieved this only on selected test problems or not at all.

Vector Doppler: spatial sampling analysis and presentation techniques for real-time systems

NASA Astrophysics Data System (ADS)

Capineri, Lorenzo; Scabia, Marco; Masotti, Leonardo F.

2001-05-01

The aim of the vector Doppler (VD) technique is the quantitative reconstruction of a velocity field independently of the ultrasonic probe axis to flow angle. In particular vector Doppler is interesting for studying vascular pathologies related to complex blood flow conditions. Clinical applications require a real-time operating mode and the capability to perform Doppler measurements over a defined volume. The combination of these two characteristics produces a real-time vector velocity map. In previous works the authors investigated the theory of pulsed wave (PW) vector Doppler and developed an experimental system capable of producing off-line 3D vector velocity maps. Afterwards, for producing dynamic velocity vector maps, we realized a new 2D vector Doppler system based on a modified commercial echograph. The measurement and presentation of a vector velocity field requires a correct spatial sampling that must satisfy the Shannon criterion. In this work we tackled this problem, establishing a relationship between sampling steps and scanning system characteristics. Another problem posed by the vector Doppler technique is the data representation in real-time that should be easy to interpret for the physician. With this in mine we attempted a multimedia solution that uses both interpolated images and sound to represent the information of the measured vector velocity map. These presentation techniques were experimented for real-time scanning on flow phantoms and preliminary measurements in vivo on a human carotid artery.

Single Vector Calibration System for Multi-Axis Load Cells and Method for Calibrating a Multi-Axis Load Cell

NASA Technical Reports Server (NTRS)

Parker, Peter A. (Inventor)

2003-01-01

A single vector calibration system is provided which facilitates the calibration of multi-axis load cells, including wind tunnel force balances. The single vector system provides the capability to calibrate a multi-axis load cell using a single directional load, for example loading solely in the gravitational direction. The system manipulates the load cell in three-dimensional space, while keeping the uni-directional calibration load aligned. The use of a single vector calibration load reduces the set-up time for the multi-axis load combinations needed to generate a complete calibration mathematical model. The system also reduces load application inaccuracies caused by the conventional requirement to generate multiple force vectors. The simplicity of the system reduces calibration time and cost, while simultaneously increasing calibration accuracy.

Field applications of stand-off sensing using visible/NIR multivariate optical computing

NASA Astrophysics Data System (ADS)

Eastwood, DeLyle; Soyemi, Olusola O.; Karunamuni, Jeevanandra; Zhang, Lixia; Li, Hongli; Myrick, Michael L.

2001-02-01

12 A novel multivariate visible/NIR optical computing approach applicable to standoff sensing will be demonstrated with porphyrin mixtures as examples. The ultimate goal is to develop environmental or counter-terrorism sensors for chemicals such as organophosphorus (OP) pesticides or chemical warfare simulants in the near infrared spectral region. The mathematical operation that characterizes prediction of properties via regression from optical spectra is a calculation of inner products between the spectrum and the pre-determined regression vector. The result is scaled appropriately and offset to correspond to the basis from which the regression vector is derived. The process involves collecting spectroscopic data and synthesizing a multivariate vector using a pattern recognition method. Then, an interference coating is designed that reproduces the pattern of the multivariate vector in its transmission or reflection spectrum, and appropriate interference filters are fabricated. High and low refractive index materials such as Nb2O5 and SiO2 are excellent choices for the visible and near infrared regions. The proof of concept has now been established for this system in the visible and will later be extended to chemicals such as OP compounds in the near and mid-infrared.

A nonproliferating parvovirus vaccine vector elicits sustained, protective humoral immunity following a single intravenous or intranasal inoculation.

PubMed

Palmer, Gene A; Brogdon, Jennifer L; Constant, Stephanie L; Tattersall, Peter

2004-02-01

An ideal vaccine delivery system would elicit persistent protection following a single administration, preferably by a noninvasive route, and be safe even in the face of immunosuppression, either inherited or acquired, of the recipient. We have exploited the unique life cycle of the autonomous parvoviruses to develop a nonproliferating vaccine platform that appears to both induce priming and continually boost a protective immune response following a single inoculation. A crippled parvovirus vector was constructed, based on a chimera between minute virus of mice (MVM) and LuIII, which expresses Borrelia burgdorferi outer surface protein A (OspA) instead of its coat protein. The vector was packaged into an MVM lymphotropic capsid and inoculated into naive C3H/HeNcr mice. Vaccination with a single vector dose, either intravenously or intranasally, elicited high-titer anti-OspA-specific antibody that provided protection from live spirochete challenge and was sustained over the lifetime of the animal. Both humoral and cell-mediated Th(1) immunity was induced, as shown by anti-OspA immunoglobulin G2a antibody and preferential gamma interferon production by OspA-specific CD4(+) T cells.

A Nonproliferating Parvovirus Vaccine Vector Elicits Sustained, Protective Humoral Immunity following a Single Intravenous or Intranasal Inoculation

PubMed Central

Palmer, Gene A.; Brogdon, Jennifer L.; Constant, Stephanie L.; Tattersall, Peter

2004-01-01

An ideal vaccine delivery system would elicit persistent protection following a single administration, preferably by a noninvasive route, and be safe even in the face of immunosuppression, either inherited or acquired, of the recipient. We have exploited the unique life cycle of the autonomous parvoviruses to develop a nonproliferating vaccine platform that appears to both induce priming and continually boost a protective immune response following a single inoculation. A crippled parvovirus vector was constructed, based on a chimera between minute virus of mice (MVM) and LuIII, which expresses Borrelia burgdorferi outer surface protein A (OspA) instead of its coat protein. The vector was packaged into an MVM lymphotropic capsid and inoculated into naive C3H/HeNcr mice. Vaccination with a single vector dose, either intravenously or intranasally, elicited high-titer anti-OspA-specific antibody that provided protection from live spirochete challenge and was sustained over the lifetime of the animal. Both humoral and cell-mediated Th1 immunity was induced, as shown by anti-OspA immunoglobulin G2a antibody and preferential gamma interferon production by OspA-specific CD4+ T cells. PMID:14722265

Metabolic flux profiling of MDCK cells during growth and canine adenovirus vector production.

PubMed

Carinhas, Nuno; Pais, Daniel A M; Koshkin, Alexey; Fernandes, Paulo; Coroadinha, Ana S; Carrondo, Manuel J T; Alves, Paula M; Teixeira, Ana P

2016-03-23

Canine adenovirus vector type 2 (CAV2) represents an alternative to human adenovirus vectors for certain gene therapy applications, particularly neurodegenerative diseases. However, more efficient production processes, assisted by a greater understanding of the effect of infection on producer cells, are required. Combining [1,2-(13)C]glucose and [U-(13)C]glutamine, we apply for the first time (13)C-Metabolic flux analysis ((13)C-MFA) to study E1-transformed Madin-Darby Canine Kidney (MDCK) cells metabolism during growth and CAV2 production. MDCK cells displayed a marked glycolytic and ammoniagenic metabolism, and (13)C data revealed a large fraction of glutamine-derived labelling in TCA cycle intermediates, emphasizing the role of glutamine anaplerosis. (13)C-MFA demonstrated the importance of pyruvate cycling in balancing glycolytic and TCA cycle activities, as well as occurrence of reductive alphaketoglutarate (AKG) carboxylation. By turn, CAV2 infection significantly upregulated fluxes through most central metabolism, including glycolysis, pentose-phosphate pathway, glutamine anaplerosis and, more prominently, reductive AKG carboxylation and cytosolic acetyl-coenzyme A formation, suggestive of increased lipogenesis. Based on these results, we suggest culture supplementation strategies to stimulate nucleic acid and lipid biosynthesis for improved canine adenoviral vector production.

Development of A Flexible System for the Simultaneous Conversion of Biomass to Industrial Chemicals and the Production of Industrial Biocatalysts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Johnway; Hooker, Brian S.; Skeen, R S.

2002-01-01

A flexible system was developed for the simultaneous conversion of biomass to industrial chemicals and the production of industrial biocatalysts. In particular, the expression of a bacterial enzyme, beta-glucuronidase (GUS), was investigated using a genetically modified starch-degrading Saccharomyces strain in suspension cultures in starch media. Different sources of starch including corn and waste potato starch were used for yeast biomass accumulation and GUS expression studies under controls of inducible and constitutive promoters. A thermostable bacterial cellulase, Acidothermus cellulolyticus E1 endoglucanase gene was also cloned into an episomal plasmid expression vector and expressed in the starch-degrading Saccharomyces strain.

Cloning and characterization of an adenoviral vector for highly efficient and doxycycline – suppressible expression of bioactive human single – chain interleukin 12 in colon cancer

PubMed Central

Wulff, Holger; Krieger, Thorsten; Krüger, Karen; Stahmer, Ingrid; Thaiss, Friedrich; Schäfer, Hansjörg; Block, Andreas

2007-01-01

Background Interleukin-12 (IL-12) is well characterized to induce cellular antitumoral immunity by activation of NK-cells and T-lymphocytes. However, systemic administration of recombinant human IL-12 resulted in severe toxicity without perceptible therapeutic benefit. Even though intratumoral expression of IL-12 leads to tumor regression and long-term survival in a variety of animal models, clinical trials have not yet shown a significant therapeutic benefit. One major obstacle in the treatment with IL-12 is to overcome the relatively low expression of the therapeutic gene without compromising the safety of such an approach. Our objective was to generate an adenoviral vector system enabling the regulated expression of very high levels of bioactive, human IL-12. Results High gene expression was obtained utilizing the VP16 herpes simplex transactivator. Strong regulation of gene expression was realized by fusion of the VP16 to a tetracycline repressor with binding of the fusion protein to a flanking tetracycline operator and further enhanced by auto-regulated expression of its fusion gene within a bicistronic promoter construct. Infection of human colon cancer cells (HT29) at a multiplicity of infection (m.o.i.) of 10 resulted in the production of up to 8000 ng/106 cells in 48 h, thus exceeding any published vector system so far. Doxycycline concentrations as low as 30 ng/ml resulted in up to 5000-fold suppression, enabling significant reduction of gene expression in a possible clinical setting. Bioactivity of the human single-chain IL-12 was similar to purified human heterodimeric IL-12. Frozen sections of human colon cancer showed high expression of the coxsackie adenovirus receptor with significant production of human single chain IL-12 in colon cancer biopsies after infection with 3*107 p.f.u. Ad.3r-scIL12. Doxycycline mediated suppression of gene expression was up to 9000-fold in the infected colon cancer tissue. Conclusion VP16 transactivator-mediated and doxycycline-regulated expression of the human interleukin-12 gene enables highly efficient and tightly controlled cytokine expression in human cancer. These data illustrate the potential of the described adenoviral vector system for the safe and superior expression of therapeutic genes in the treatment of colorectal cancer and other malignancies. PMID:17594499

Real-time individualized training vectors for experiential learning.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Willis, Matt; Tucker, Eilish Marie; Raybourn, Elaine Marie

2011-01-01

Military training utilizing serious games or virtual worlds potentially generate data that can be mined to better understand how trainees learn in experiential exercises. Few data mining approaches for deployed military training games exist. Opportunities exist to collect and analyze these data, as well as to construct a full-history learner model. Outcomes discussed in the present document include results from a quasi-experimental research study on military game-based experiential learning, the deployment of an online game for training evidence collection, and results from a proof-of-concept pilot study on the development of individualized training vectors. This Lab Directed Research & Development (LDRD)more » project leveraged products within projects, such as Titan (Network Grand Challenge), Real-Time Feedback and Evaluation System, (America's Army Adaptive Thinking and Leadership, DARWARS Ambush! NK), and Dynamic Bayesian Networks to investigate whether machine learning capabilities could perform real-time, in-game similarity vectors of learner performance, toward adaptation of content delivery, and quantitative measurement of experiential learning.« less

Method and system for operating an electric motor

DOEpatents

Gallegos-Lopez, Gabriel; Hiti, Silva; Perisic, Milun

2013-01-22

Methods and systems for operating an electric motor having a plurality of windings with an inverter having a plurality of switches coupled to a voltage source are provided. A first plurality of switching vectors is applied to the plurality of switches. The first plurality of switching vectors includes a first ratio of first magnitude switching vectors to second magnitude switching vectors. A direct current (DC) current associated with the voltage source is monitored during the applying of the first plurality of switching vectors to the plurality of switches. A second ratio of the first magnitude switching vectors to the second magnitude switching vectors is selected based on the monitoring of the DC current associated with the voltage source. A second plurality of switching vectors is applied to the plurality of switches. The second plurality of switching vectors includes the second ratio of the first magnitude switching vectors to the second magnitude switching vectors.

Influence of sequence and size of DNA on packaging efficiency of parvovirus MVM-based vectors.

PubMed

Brandenburger, A; Coessens, E; El Bakkouri, K; Velu, T

1999-05-01

We have derived a vector from the autonomous parvovirus MVM(p), which expresses human IL-2 specifically in transformed cells (Russell et al., J. Virol 1992;66:2821-2828). Testing the therapeutic potential of these vectors in vivo requires high-titer stocks. Stocks with a titer of 10(9) can be obtained after concentration and purification (Avalosse et al., J. Virol. Methods 1996;62:179-183), but this method requires large culture volumes and cannot easily be scaled up. We wanted to increase the production of recombinant virus at the initial transfection step. Poor vector titers could be due to inadequate genome amplification or to inefficient packaging. Here we show that intracellular amplification of MVM vector genomes is not the limiting factor for vector production. Several vector genomes of different size and/or structure were amplified to an equal extent. Their amplification was also equivalent to that of a cotransfected wild-type genome. We did not observe any interference between vector and wild-type genomes at the level of DNA amplification. Despite equivalent genome amplification, vector titers varied greatly between the different genomes, presumably owing to differences in packaging efficiency. Genomes with a size close to 100% that of wild type were packaged most efficiently with loss of efficiency at lower and higher sizes. However, certain genomes of identical size showed different packaging efficiencies, illustrating the importance of the DNA sequence, and probably its structure.

A Fiber-Optic Borehole Seismic Vector Sensor System for Geothermal Site Characterization and Monitoring

DOE Office of Scientific and Technical Information (OSTI.GOV)

Paulsson, Bjorn N.P.; Thornburg, Jon A.; He, Ruiqing

2015-04-21

Seismic techniques are the dominant geophysical techniques for the characterization of subsurface structures and stratigraphy. The seismic techniques also dominate the monitoring and mapping of reservoir injection and production processes. Borehole seismology, of all the seismic techniques, despite its current shortcomings, has been shown to provide the highest resolution characterization and most precise monitoring results because it generates higher signal to noise ratio and higher frequency data than surface seismic techniques. The operational environments for borehole seismic instruments are however much more demanding than for surface seismic instruments making both the instruments and the installation much more expensive. The currentmore » state-of-the-art borehole seismic instruments have not been robust enough for long term monitoring compounding the problems with expensive instruments and installations. Furthermore, they have also not been able to record the large bandwidth data available in boreholes or having the sensitivity allowing them to record small high frequency micro seismic events with high vector fidelity. To reliably achieve high resolution characterization and long term monitoring of Enhanced Geothermal Systems (EGS) sites a new generation of borehole seismic instruments must therefore be developed and deployed. To address the critical site characterization and monitoring needs for EGS programs, US Department of Energy (DOE) funded Paulsson, Inc. in 2010 to develop a fiber optic based ultra-large bandwidth clamped borehole seismic vector array capable of deploying up to one thousand 3C sensor pods suitable for deployment into ultra-high temperature and high pressure boreholes. Tests of the fiber optic seismic vector sensors developed on the DOE funding have shown that the new borehole seismic sensor technology is capable of generating outstanding high vector fidelity data with extremely large bandwidth: 0.01 – 6,000 Hz. Field tests have shown that the system can record events at magnitudes much smaller than M-2.6 at frequencies up to 2,000 Hz. The sensors have also proved to be about 100 times more sensitive than the regular coil geophones that are used in borehole seismic systems today. The fiber optic seismic sensors have furthermore been qualified to operate at temperatures over 300°C (572°F). Simultaneously with the fiber optic based seismic 3C vector sensors we are using the lead-in fiber to acquire Distributed Acoustic Sensor (DAS) data from the surface to the bottom of the vector array. While the DAS data is of much lower quality than the vector sensor data it provides a 1 m spatial sampling of the downgoing wavefield which will be used to build the high resolution velocity model which is an essential component in high resolution imaging and monitoring.« less

Sequential cloning of chromosomes

DOEpatents

Lacks, Sanford A.

1995-07-18

A method for sequential cloning of chromosomal DNA of a target organism is disclosed. A first DNA segment homologous to the chromosomal DNA to be sequentially cloned is isolated. The first segment has a first restriction enzyme site on either side. A first vector product is formed by ligating the homologous segment into a suitably designed vector. The first vector product is circularly integrated into the target organism's chromosomal DNA. The resulting integrated chromosomal DNA segment includes the homologous DNA segment at either end of the integrated vector segment. The integrated chromosomal DNA is cleaved with a second restriction enzyme and ligated to form a vector-containing plasmid, which is replicated in a host organism. The replicated plasmid is then cleaved with the first restriction enzyme. Next, a DNA segment containing the vector and a segment of DNA homologous to a distal portion of the previously isolated DNA segment is isolated. This segment is then ligated to form a plasmid which is replicated within a suitable host. This plasmid is then circularly integrated into the target chromosomal DNA. The chromosomal DNA containing the circularly integrated vector is treated with a third, retrorestriction (class IIS) enzyme. The cleaved DNA is ligated to give a plasmid that is used to transform a host permissive for replication of its vector. The sequential cloning process continues by repeated cycles of circular integration and excision. The excision is carried out alternately with the second and third enzymes.

A robust gene-stacking method utilizing yeast assembly for plant synthetic biology

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shih, Patrick M.; Vuu, Khanh; Mansoori, Nasim

The advent and growth of synthetic biology has demonstrated its potential as a promising avenue of research to address many societal needs. But, plant synthetic biology efforts have been hampered by a dearth of DNA part libraries, versatile transformation vectors and efficient assembly strategies. We describe a versatile system (named jStack) utilizing yeast homologous recombination to efficiently assemble DNA into plant transformation vectors. We also demonstrate how this method can facilitate pathway engineering of molecules of pharmaceutical interest, production of potential biofuels and shuffling of disease-resistance traits between crop species. Our approach provides a powerful alternative to conventional strategies formore » stacking genes and traits to address many impending environmental and agricultural challenges.« less

A robust gene-stacking method utilizing yeast assembly for plant synthetic biology

DOE PAGES

Shih, Patrick M.; Vuu, Khanh; Mansoori, Nasim; ...

2016-10-26

The advent and growth of synthetic biology has demonstrated its potential as a promising avenue of research to address many societal needs. But, plant synthetic biology efforts have been hampered by a dearth of DNA part libraries, versatile transformation vectors and efficient assembly strategies. We describe a versatile system (named jStack) utilizing yeast homologous recombination to efficiently assemble DNA into plant transformation vectors. We also demonstrate how this method can facilitate pathway engineering of molecules of pharmaceutical interest, production of potential biofuels and shuffling of disease-resistance traits between crop species. Our approach provides a powerful alternative to conventional strategies formore » stacking genes and traits to address many impending environmental and agricultural challenges.« less

A multi-dimensional Smolyak collocation method in curvilinear coordinates for computing vibrational spectra

DOE Office of Scientific and Technical Information (OSTI.GOV)

Avila, Gustavo, E-mail: Gustavo-Avila@telefonica.net; Carrington, Tucker, E-mail: Tucker.Carrington@queensu.ca

In this paper, we improve the collocation method for computing vibrational spectra that was presented in Avila and Carrington, Jr. [J. Chem. Phys. 139, 134114 (2013)]. Using an iterative eigensolver, energy levels and wavefunctions are determined from values of the potential on a Smolyak grid. The kinetic energy matrix-vector product is evaluated by transforming a vector labelled with (nondirect product) grid indices to a vector labelled by (nondirect product) basis indices. Both the transformation and application of the kinetic energy operator (KEO) scale favorably. Collocation facilitates dealing with complicated KEOs because it obviates the need to calculate integrals of coordinatemore » dependent coefficients of differential operators. The ideas are tested by computing energy levels of HONO using a KEO in bond coordinates.« less

Equivalent Vectors

ERIC Educational Resources Information Center

Levine, Robert

2004-01-01

The cross-product is a mathematical operation that is performed between two 3-dimensional vectors. The result is a vector that is orthogonal or perpendicular to both of them. Learning about this for the first time while taking Calculus-III, the class was taught that if AxB = AxC, it does not necessarily follow that B = C. This seemed baffling. The…

Intramuscular injection of AAV8 in mice and macaques is associated with substantial hepatic targeting and transgene expression.

PubMed

Greig, Jenny A; Peng, Hui; Ohlstein, Jason; Medina-Jaszek, C Angelica; Ahonkhai, Omua; Mentzinger, Anne; Grant, Rebecca L; Roy, Soumitra; Chen, Shu-Jen; Bell, Peter; Tretiakova, Anna P; Wilson, James M

2014-01-01

Intramuscular (IM) administration of adeno-associated viral (AAV) vectors has entered the early stages of clinical development with some success, including the first approved gene therapy product in the West called Glybera. In preparation for broader clinical development of IM AAV vector gene therapy, we conducted detailed pre-clinical studies in mice and macaques evaluating aspects of delivery that could affect performance. We found that following IM administration of AAV8 vectors in mice, a portion of the vector reached the liver and hepatic gene expression contributed significantly to total expression of secreted transgenes. The contribution from liver could be controlled by altering injection volume and by the use of traditional (promoter) and non-traditional (tissue-specific microRNA target sites) expression control elements. Hepatic distribution of vector following IM injection was also noted in rhesus macaques. These pre-clinical data on AAV delivery should inform safe and efficient development of future AAV products.

Naval Observatory Vector Astrometry Software (NOVAS) Version 3.1, Introducing a Python Edition

NASA Astrophysics Data System (ADS)

Barron, Eric G.; Kaplan, G. H.; Bangert, J.; Bartlett, J. L.; Puatua, W.; Harris, W.; Barrett, P.

2011-01-01

The Naval Observatory Vector Astrometry Software (NOVAS) is a source-code library that provides common astrometric quantities and transformations. NOVAS calculations are accurate at the sub-milliarcsecond level. The library can supply, in one or two subroutine or function calls, the instantaneous celestial position of any star or planet in a variety of coordinate systems. NOVAS also provides access to all of the building blocks that go into such computations. NOVAS Version 3.1 introduces a Python edition alongside the Fortran and C editions. The Python edition uses the computational code from the C edition and, currently, mimics the function calls of the C edition. Future versions will expand the functionality of the Python edition to harness the object-oriented nature of the Python language, and will implement the ability to handle large quantities of objects or observers using the array functionality in NumPy (a third-party scientific package for Python). NOVAS 3.1 also adds a module to transform GCRS vectors to the ITRS; the ITRS to GCRS transformation was already provided in NOVAS 3.0. The module that corrects an ITRS vector for polar motion has been modified to undo that correction upon demand. In the C edition, the ephemeris-access functions have been revised for use on 64-bit systems and for improved performance in general. NOVAS, including documentation, is available from the USNO website (http://www.usno.navy.mil/USNO/astronomical-applications/software-products/novas).

A versatile expression vector for the growth and amplification of unmodified phage display polypeptides.

PubMed

Winton, Alexander J; Baptiste, Janae L; Allen, Mark A

2018-09-01

Proteins and polypeptides represent nature's most complex and versatile polymer. They provide complicated shapes, diverse chemical functionalities, and tightly regulated and controlled sizes. Several disease states are related to the misfolding or overproduction of polypeptides and yet polypeptides are present in several therapeutic molecules. In addition to biological roles; short chain polypeptides have been shown to interact with and drive the bio-inspired synthesis or modification of inorganic materials. This paper outlines the development of a versatile cloning vector which allows for the expression of a short polypeptide by controlling the incorporation of a desired DNA coding insert. As a demonstration of the efficacy of the expression system, a solid binding polypeptide identified from M13 phage display was expressed and purified. The solid binding polypeptide was expressed as a soluble 6xHis-SUMO tagged construct. Expression was performed in E. coli using auto-induction followed by Ni-NTA affinity chromatography and ULP1 protease cleavage. Methodology demonstrates the production of greater than 8 mg of purified polypeptide per liter of E. coli culture. Isotopic labeling of the peptide is also demonstrated. The versatility of the designed cloning vector, use of the 6xHis-SUMO solubility partner, bacterial expression in auto-inducing media and the purification methodology make this expressionun vector a readily scalable and user-friendly system for the creation of desired peptide domains. Copyright © 2018. Published by Elsevier Inc.

Cloning-independent plasmid construction for genetic studies in streptococci

PubMed Central

Xie, Zhoujie; Qi, Fengxia; Merritt, Justin

2013-01-01

Shuttle plasmids are among the few routinely utilized tools in the Streptococcus mutans genetic system that still require the use of classical cloning methodologies and intermediate hosts for genetic manipulation. Accordingly, it typically requires considerably less time and effort to introduce mutations onto the S. mutans chromosome than it does to construct shuttle vectors for expressing genes in trans. Occasionally, shuttle vector constructs also exhibit toxicity in E. coli, which prevents their proper assembly. To circumvent these limitations, we modified a prolonged overlap extension PCR (POE-PCR) protocol to facilitate direct plasmid assembly in S. mutans. Using solely PCR, we created the reporter vector pZX7, which contains a single minimal streptococcal replication origin and harbors a spectinomycin resistance cassette and the gusA gene encoding β-glucuronidase. We compared the efficiency of pZX7 assembly using multiple strains of S. mutans and were able to obtain from 5×103 – 2×105 CFU/μg PCR product. Likewise, we used pZX7 to further demonstrate that Streptococcus sanguinis and Streptococcus gordonii are also excellent hosts for cloning-independent plasmid assembly, which suggests that this system is likely to function in numerous other streptococci. Consequently, it should be possible to completely forgo the use of E. coli – Streptococcus shuttle vectors in many streptococcal species, thereby decreasing the time and effort required to assemble constructs and eliminating any toxicity issues associated with intermediate hosts. PMID:23673081

Cloning-independent plasmid construction for genetic studies in streptococci.

PubMed

Xie, Zhoujie; Qi, Fengxia; Merritt, Justin

2013-08-01

Shuttle plasmids are among the few routinely utilized tools in the Streptococcus mutans genetic system that still require the use of classical cloning methodologies and intermediate hosts for genetic manipulation. Accordingly, it typically requires considerably less time and effort to introduce mutations onto the S. mutans chromosome than it does to construct shuttle vectors for expressing genes in trans. Occasionally, shuttle vector constructs also exhibit toxicity in Escherichia coli, which prevents their proper assembly. To circumvent these limitations, we modified a prolonged overlap extension PCR (POE-PCR) protocol to facilitate direct plasmid assembly in S. mutans. Using solely PCR, we created the reporter vector pZX7, which contains a single minimal streptococcal replication origin and harbors a spectinomycin resistance cassette and the gusA gene encoding β-glucuronidase. We compared the efficiency of pZX7 assembly using multiple strains of S. mutans and were able to obtain from 5 × 10³ to 2 × 10⁵ CFU/μg PCR product. Likewise, we used pZX7 to further demonstrate that Streptococcus sanguinis and Streptococcus gordonii are also excellent hosts for cloning-independent plasmid assembly, which suggests that this system is likely to function in numerous other streptococci. Consequently, it should be possible to completely forgo the use of E. coli-Streptococcus shuttle vectors in many streptococcal species, thereby decreasing the time and effort required to assemble constructs and eliminating any toxicity issues associated with intermediate hosts. Copyright © 2013 Elsevier B.V. All rights reserved.

Rapid construction of a Bacterial Artificial Chromosomal (BAC) expression vector using designer DNA fragments.

PubMed

Chen, Chao; Zhao, Xinqing; Jin, Yingyu; Zhao, Zongbao Kent; Suh, Joo-Won

2014-11-01

Bacterial artificial chromosomal (BAC) vectors are increasingly being used in cloning large DNA fragments containing complex biosynthetic pathways to facilitate heterologous production of microbial metabolites for drug development. To express inserted genes using Streptomyces species as the production hosts, an integration expression cassette is required to be inserted into the BAC vector, which includes genetic elements encoding a phage-specific attachment site, an integrase, an origin of transfer, a selection marker and a promoter. Due to the large sizes of DNA inserted into the BAC vectors, it is normally inefficient and time-consuming to assemble these fragments by routine PCR amplifications and restriction-ligations. Here we present a rapid method to insert fragments to construct BAC-based expression vectors. A DNA fragment of about 130 bp was designed, which contains upstream and downstream homologous sequences of both BAC vector and pIB139 plasmid carrying the whole integration expression cassette. In-Fusion cloning was performed using the designer DNA fragment to modify pIB139, followed by λ-RED-mediated recombination to obtain the BAC-based expression vector. We demonstrated the effectiveness of this method by rapid construction of a BAC-based expression vector with an insert of about 120 kb that contains the entire gene cluster for biosynthesis of immunosuppressant FK506. The empty BAC-based expression vector constructed in this study can be conveniently used for construction of BAC libraries using either microbial pure culture or environmental DNA, and the selected BAC clones can be directly used for heterologous expression. Alternatively, if a BAC library has already been constructed using a commercial BAC vector, the selected BAC vectors can be manipulated using the method described here to get the BAC-based expression vectors with desired gene clusters for heterologous expression. The rapid construction of a BAC-based expression vector facilitates heterologous expression of large gene clusters for drug discovery. Copyright © 2014 Elsevier Inc. All rights reserved.

Effective matrix-free preconditioning for the augmented immersed interface method

NASA Astrophysics Data System (ADS)

Xia, Jianlin; Li, Zhilin; Ye, Xin

2015-12-01

We present effective and efficient matrix-free preconditioning techniques for the augmented immersed interface method (AIIM). AIIM has been developed recently and is shown to be very effective for interface problems and problems on irregular domains. GMRES is often used to solve for the augmented variable(s) associated with a Schur complement A in AIIM that is defined along the interface or the irregular boundary. The efficiency of AIIM relies on how quickly the system for A can be solved. For some applications, there are substantial difficulties involved, such as the slow convergence of GMRES (particularly for free boundary and moving interface problems), and the inconvenience in finding a preconditioner (due to the situation that only the products of A and vectors are available). Here, we propose matrix-free structured preconditioning techniques for AIIM via adaptive randomized sampling, using only the products of A and vectors to construct a hierarchically semiseparable matrix approximation to A. Several improvements over existing schemes are shown so as to enhance the efficiency and also avoid potential instability. The significance of the preconditioners includes: (1) they do not require the entries of A or the multiplication of AT with vectors; (2) constructing the preconditioners needs only O (log ⁡ N) matrix-vector products and O (N) storage, where N is the size of A; (3) applying the preconditioners needs only O (N) flops; (4) they are very flexible and do not require any a priori knowledge of the structure of A. The preconditioners are observed to significantly accelerate the convergence of GMRES, with heuristical justifications of the effectiveness. Comprehensive tests on several important applications are provided, such as Navier-Stokes equations on irregular domains with traction boundary conditions, interface problems in incompressible flows, mixed boundary problems, and free boundary problems. The preconditioning techniques are also useful for several other problems and methods.

Job Management Requirements for NAS Parallel Systems and Clusters

NASA Technical Reports Server (NTRS)

Saphir, William; Tanner, Leigh Ann; Traversat, Bernard

1995-01-01

A job management system is a critical component of a production supercomputing environment, permitting oversubscribed resources to be shared fairly and efficiently. Job management systems that were originally designed for traditional vector supercomputers are not appropriate for the distributed-memory parallel supercomputers that are becoming increasingly important in the high performance computing industry. Newer job management systems offer new functionality but do not solve fundamental problems. We address some of the main issues in resource allocation and job scheduling we have encountered on two parallel computers - a 160-node IBM SP2 and a cluster of 20 high performance workstations located at the Numerical Aerodynamic Simulation facility. We describe the requirements for resource allocation and job management that are necessary to provide a production supercomputing environment on these machines, prioritizing according to difficulty and importance, and advocating a return to fundamental issues.

Different applications of virus-like particles in biology and medicine: Vaccination and delivery systems.

PubMed

Shirbaghaee, Zeinab; Bolhassani, Azam

2016-03-01

Virus-like particles (VLPs) mimic the whole construct of virus particles devoid of viral genome as used in subunit vaccine design. VLPs can elicit efficient protective immunity as direct immunogens compared to soluble antigens co-administered with adjuvants in several booster injections. Up to now, several prokaryotic and eukaryotic systems such as insect, yeast, plant, and E. coli were used to express recombinant proteins, especially for VLP production. Recent studies are also generating VLPs in plants using different transient expression vectors for edible vaccines. VLPs and viral particles have been applied for different functions such as gene therapy, vaccination, nanotechnology, and diagnostics. Herein, we describe VLP production in different systems as well as its applications in biology and medicine. © 2015 Wiley Periodicals, Inc.

Development of inducer-free expression plasmids based on IPTG-inducible promoters for Bacillus subtilis.

PubMed

Tran, Dinh Thi Minh; Phan, Trang Thi Phuong; Huynh, Thanh Kieu; Dang, Ngan Thi Kim; Huynh, Phuong Thi Kim; Nguyen, Tri Minh; Truong, Tuom Thi Tinh; Tran, Thuoc Linh; Schumann, Wolfgang; Nguyen, Hoang Duc

2017-07-25

Besides Escherichia coli, Bacillus subtilis is an important bacterial species for the production of recombinant proteins. Recombinant genes are inserted into shuttle expression vectors which replicate in both E. coli and in B. subtilis. The ligation products are first transformed into E. coli cells, analyzed for correct insertions, and the correct recombinant plasmids are then transformed into B. subtilis. A major problem using E. coli cells can be the strong basal level of expression of the recombinant protein which may interfere with the stability of the cells. To minimize this problem, we developed strong expression vectors being repressed in E. coli and inducer-free in B. subtilis. In general, induction of IPTG-inducible expression vectors is determined by the regulatory lacI gene encoding the LacI repressor in combination with the lacO operator on the promoter. To investigate the inducer-free properties of the vectors, we constructed inducer-free expression plasmids by removing the lacI gene and characterized their properties. First, we examined the ability to repress a reporter gene in E. coli, which is a prominent property facilitating the construction of the expression vectors carrying a target gene. The β-galactosidase (bgaB gene) basal levels expressed from Pgrac01-bgaB could be repressed at least twice in the E. coli cloning strain. Second, the inducer-free production of BgaB from four different plasmids with the Pgrac01 promoter in B. subtilis was investigated. As expected, BgaB expression levels of inducer-free constructs are at least 37 times higher than that of the inducible constructs in the absence of IPTG, and comparable to those in the presence of the inducer. Third, using efficient IPTG-inducible expression vectors containing the strong promoter Pgrac100, we could convert them into inducer-free expression plasmids. The BgaB production levels from the inducer-free plasmid in the absence of the inducer were at least 4.5 times higher than that of the inducible vector using the same promoter. Finally, we used gfp as a reporter gene in combination with the two promoters Pgrac01 and Pgrac100 to test the new vector types. The GFP expression levels could be repressed at least 1.5 times for the Pgrac01-gfp+ inducer-free construct in E. coli. The inducer-free constructs Pgrac01-gfp+ and Pgrac100-gfp+ allowed GFP expression at high levels from 23 × 10 4 to 32 × 10 4 RFU units and 9-13% of total intracellular proteins. We could reconfirm the two major advantages of the new inducer-free expression plasmids: (1) Strong repression of the target gene expression in the E. coli cloning strain, and (2) production of the target protein at high levels in B. subtilis in the absence of the inducer. We propose a general strategy to generate inducer-free expression vector by using IPTG-inducible vectors, and more specifically we developed inducer-free expression plasmids using IPTG-inducible promoters in the absence of the LacI repressor. These plasmids could be an excellent choice for high-level production of recombinant proteins in B. subtilis without the addition of inducer and at the same time maintaining a low basal level of the recombinant proteins in E. coli. The repression of the recombinant gene expression would facilitate cloning of genes that potentially inhibit the growth of E. coli cloning strains. The inducer-free expression plasmids will be extended versions of the current available IPTG-inducible expression vectors for B. subtilis, in which all these vectors use the same cognate promoters. These inducer-free and previously developed IPTG-inducible expression plasmids will be a useful cassette to study gene expression at a small scale up to a larger scale up for the production of recombinant proteins.

Combinatorial vector fields and the valley structure of fitness landscapes.

PubMed

Stadler, Bärbel M R; Stadler, Peter F

2010-12-01

Adaptive (downhill) walks are a computationally convenient way of analyzing the geometric structure of fitness landscapes. Their inherently stochastic nature has limited their mathematical analysis, however. Here we develop a framework that interprets adaptive walks as deterministic trajectories in combinatorial vector fields and in return associate these combinatorial vector fields with weights that measure their steepness across the landscape. We show that the combinatorial vector fields and their weights have a product structure that is governed by the neutrality of the landscape. This product structure makes practical computations feasible. The framework presented here also provides an alternative, and mathematically more convenient, way of defining notions of valleys, saddle points, and barriers in landscape. As an application, we propose a refined approximation for transition rates between macrostates that are associated with the valleys of the landscape.

Double promoter expression systems for recombinant protein production by industrial microorganisms.

PubMed

Öztürk, Sibel; Ergün, Burcu Gündüz; Çalık, Pınar

2017-10-01

Using double promoter expression systems is a promising approach to increase heterologous protein production. In this review, current double promoter expression systems for the production of recombinant proteins (r-proteins) by industrially important bacteria, Bacillus subtilis and Escherichia coli; and yeasts, Saccharomyces cerevisiae and Pichia pastoris, are discussed by assessing their potentials and drawbacks. Double promoter expression systems need to be designed to maintain a higher specific product formation rate within the production domain. While bacterial double promoter systems have been constructed as chimeric tandem promoters, yeast dual promoter systems have been developed as separate expression cassettes. To increase production and productivity, the optimal transcriptional activity should be justified either by simultaneously satisfying the requirements of both promoters, or by consecutively stimulating the changeover from one to another in a biphasic process or via successive-iterations. Thus, considering the dynamics of a fermentation process, double promoters can be classified according to their operational mechanisms, as: i) consecutively operating double promoter systems, and ii) simultaneously operating double promoter systems. Among these metabolic design strategies, extending the expression period with two promoters activated under different conditions, or enhancing the transcriptional activity with two promoters activated under similar conditions within the production domain, can be applied independently from the host. Novel studies with new insights, which aim a rational systematic design and construction of dual promoter expression vectors with tailored transcriptional activity, will empower r-protein production with enhanced production and productivity. Finally, the current state-of-the-art review emphasizes the advantages of double promoter systems along with the necessity for discovering new promoters for the development of more effective and adaptive processes to meet the increasing demand of r-protein industry.

A Discriminant Distance Based Composite Vector Selection Method for Odor Classification

PubMed Central

Choi, Sang-Il; Jeong, Gu-Min

2014-01-01

We present a composite vector selection method for an effective electronic nose system that performs well even in noisy environments. Each composite vector generated from a electronic nose data sample is evaluated by computing the discriminant distance. By quantitatively measuring the amount of discriminative information in each composite vector, composite vectors containing informative variables can be distinguished and the final composite features for odor classification are extracted using the selected composite vectors. Using the only informative composite vectors can be also helpful to extract better composite features instead of using all the generated composite vectors. Experimental results with different volatile organic compound data show that the proposed system has good classification performance even in a noisy environment compared to other methods. PMID:24747735

Recombinant host cells and media for ethanol production

DOEpatents

Wood, Brent E; Ingram, Lonnie O; Yomano, Lorraine P; York, Sean W

2014-02-18

Disclosed are recombinant host cells suitable for degrading an oligosaccharide that have been optimized for growth and production of high yields of ethanol, and methods of making and using these cells. The invention further provides minimal media comprising urea-like compounds for economical production of ethanol by recombinant microorganisms. Recombinant host cells in accordance with the invention are modified by gene mutation to eliminate genes responsible for the production of unwanted products other than ethanol, thereby increasing the yield of ethanol produced from the oligosaccharides, relative to unmutated parent strains. The new and improved strains of recombinant bacteria are capable of superior ethanol productivity and yield when grown under conditions suitable for fermentation in minimal growth media containing inexpensive reagents. Systems optimized for ethanol production combine a selected optimized minimal medium with a recombinant host cell optimized for use in the selected medium. Preferred systems are suitable for efficient ethanol production by simultaneous saccharification and fermentation (SSF) using lignocellulose as an oligosaccharide source. The invention also provides novel isolated polynucleotide sequences, polypeptide sequences, vectors and antibodies.

An affine projection algorithm using grouping selection of input vectors

NASA Astrophysics Data System (ADS)

Shin, JaeWook; Kong, NamWoong; Park, PooGyeon

2011-10-01

This paper present an affine projection algorithm (APA) using grouping selection of input vectors. To improve the performance of conventional APA, the proposed algorithm adjusts the number of the input vectors using two procedures: grouping procedure and selection procedure. In grouping procedure, the some input vectors that have overlapping information for update is grouped using normalized inner product. Then, few input vectors that have enough information for for coefficient update is selected using steady-state mean square error (MSE) in selection procedure. Finally, the filter coefficients update using selected input vectors. The experimental results show that the proposed algorithm has small steady-state estimation errors comparing with the existing algorithms.

Production of non viral DNA vectors.

PubMed

Schleef, Martin; Blaesen, Markus; Schmeer, Marco; Baier, Ruth; Marie, Corinne; Dickson, George; Scherman, Daniel

2010-12-01

After some decades of research, development and first clinical approaches to use DNA vectors in gene therapy, cell therapy and DNA vaccination, the requirements for the pharmaceutical manufacturing of gene vectors has improved significantly step by step. Even the expression level and specificity of non viral DNA vectors were significantly modified and followed the success of viral vectors. The strict separation of "viral" and "non viral" gene transfer are historic borders between scientist and we will show that both fields together are able to allow the next step towards successful prevention and therapy. Here we summarize the features of producing and modifying these non-viral gene vectors to ensure the required quality to modify cells and to treat human and animals.

Using Chebyshev polynomials and approximate inverse triangular factorizations for preconditioning the conjugate gradient method

NASA Astrophysics Data System (ADS)

Kaporin, I. E.

2012-02-01

In order to precondition a sparse symmetric positive definite matrix, its approximate inverse is examined, which is represented as the product of two sparse mutually adjoint triangular matrices. In this way, the solution of the corresponding system of linear algebraic equations (SLAE) by applying the preconditioned conjugate gradient method (CGM) is reduced to performing only elementary vector operations and calculating sparse matrix-vector products. A method for constructing the above preconditioner is described and analyzed. The triangular factor has a fixed sparsity pattern and is optimal in the sense that the preconditioned matrix has a minimum K-condition number. The use of polynomial preconditioning based on Chebyshev polynomials makes it possible to considerably reduce the amount of scalar product operations (at the cost of an insignificant increase in the total number of arithmetic operations). The possibility of an efficient massively parallel implementation of the resulting method for solving SLAEs is discussed. For a sequential version of this method, the results obtained by solving 56 test problems from the Florida sparse matrix collection (which are large-scale and ill-conditioned) are presented. These results show that the method is highly reliable and has low computational costs.

Targeted systemic gene therapy and molecular imaging of cancer contribution of the vascular-targeted AAVP vector.

PubMed

Hajitou, Amin

2010-01-01

Gene therapy and molecular-genetic imaging have faced a major problem: the lack of an efficient systemic gene delivery vector. Unquestionably, eukaryotic viruses have been the vectors of choice for gene delivery to mammalian cells; however, they have had limited success in systemic gene therapy. This is mainly due to undesired uptake by the liver and reticuloendothelial system, broad tropism for mammalian cells causing toxicity, and their immunogenicity. On the other hand, prokaryotic viruses such as bacteriophage (phage) have no tropism for mammalian cells, but can be engineered to deliver genes to these cells. However, phage-based vectors have inherently been considered poor vectors for mammalian cells. We have reported a new generation of vascular-targeted systemic hybrid prokaryotic-eukaryotic vectors as chimeras between an adeno-associated virus (AAV) and targeted bacteriophage (termed AAV/phage; AAVP). In this hybrid vector, the targeted bacteriophage serves as a shuttle to deliver the AAV transgene cassette inserted in an intergenomic region of the phage DNA genome. As a proof of concept, we assessed the in vivo efficacy of vector in animal models of cancer by displaying on the phage capsid the cyclic Arg-Gly-Asp (RGD-4C) ligand that binds to alphav integrin receptors specifically expressed on the angiogenic blood vessels of tumors. The ligand-directed vector was able to specifically deliver imaging and therapeutic transgenes to tumors in mice, rats, and dogs while sparing the normal organs. This chapter reviews some gene transfer strategies and the potential of the vascular-targeted AAVP vector for enhancing the effectiveness of existing systemic gene delivery and genetic-imaging technologies. Copyright (c) 2010 Elsevier Inc. All rights reserved.

A potyvirus vector efficiently targets recombinant proteins to chloroplasts, mitochondria and nuclei in plant cells when expressed at the amino terminus of the polyprotein.

PubMed

Majer, Eszter; Navarro, José-Antonio; Daròs, José-Antonio

2015-09-01

Plant virus-based expression systems allow quick and efficient production of recombinant proteins in plant biofactories. Among them, a system derived from tobacco etch virus (TEV; genus potyvirus) permits coexpression of equimolar amounts of several recombinant proteins. This work analyzed how to target recombinant proteins to different subcellular localizations in the plant cell using this system. We constructed TEV clones in which green fluorescent protein (GFP), with a chloroplast transit peptide (cTP), a nuclear localization signal (NLS) or a mitochondrial targeting peptide (mTP) was expressed either as the most amino-terminal product or embedded in the viral polyprotein. Results showed that cTP and mTP mediated efficient translocation of GFP to the corresponding organelle only when present at the amino terminus of the viral polyprotein. In contrast, the NLS worked efficiently at both positions. Viruses expressing GFP in the amino terminus of the viral polyprotein produced milder symptoms. Untagged GFPs and cTP and NLS tagged amino-terminal GFPs accumulated to higher amounts in infected tissues. Finally, viral progeny from clones with internal GFPs maintained the extra gene better. These observations will help in the design of potyvirus-based vectors able to coexpress several proteins while targeting different subcellular localizations, as required in plant metabolic engineering. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Advanced technologies for improved expression of recombinant proteins in bacteria: perspectives and applications.

PubMed

Gupta, Sanjeev K; Shukla, Pratyoosh

2016-12-01

Prokaryotic expression systems are superior in producing valuable recombinant proteins, enzymes and therapeutic products. Conventional microbial technology is evolving gradually and amalgamated with advanced technologies in order to give rise to improved processes for the production of metabolites, recombinant biopharmaceuticals and industrial enzymes. Recently, several novel approaches have been employed in a bacterial expression platform to improve recombinant protein expression. These approaches involve metabolic engineering, use of strong promoters, novel vector elements such as inducers and enhancers, protein tags, secretion signals, high-throughput devices for cloning and process screening as well as fermentation technologies. Advancement of the novel technologies in E. coli systems led to the production of "difficult to express" complex products including small peptides, antibody fragments, few proteins and full-length aglycosylated monoclonal antibodies in considerable large quantity. Wacker's secretion technologies, Pfenex system, inducers, cell-free systems, strain engineering for post-translational modification, such as disulfide bridging and bacterial N-glycosylation, are still under evaluation for the production of complex proteins and peptides in E. coli in an efficient manner. This appraisal provides an impression of expression technologies developed in recent times for enhanced production of heterologous proteins in E. coli which are of foremost importance for diverse applications in microbiology and biopharmaceutical production.

Metabolic biotinylation of lentiviral pseudotypes for scalable paramagnetic microparticle-dependent manipulation.

PubMed

Nesbeth, Darren; Williams, Sharon L; Chan, Lucas; Brain, Tony; Slater, Nigel K H; Farzaneh, Farzin; Darling, David

2006-04-01

Nonviral, host-derived proteins on lentiviral vector surfaces can have a profound effect on the vector's biology as they can both promote infection and provide resistance to complement inactivation. We have exploited this to engineer a specific posttranslational modification of a "nonenvelope," virally associated protein. The bacterial biotin ligase (BirA) and a modified human DeltaLNGFR have been introduced into HEK293T cells and their protein products directed to the lumen of the endoplasmic reticulum. The BirA then couples biotin to an acceptor peptide that has been fused to the DeltaLNGFR. This results in the covalent linkage of biotin to the extracellular domain of the DeltaLNGFR expressed on the cell surface. Lentiviral vectors from these cells are metabolically labeled with biotin in the presence of free biotin. These biotinylated lentiviral vectors have a high affinity for streptavidin paramagnetic particles and, once captured, are easily manipulated in vitro. This is illustrated by the concentration of lentiviral vectors pseudotyped with either the VSV-G or an amphotropic envelope in excess of 4500-fold. This new cell line has the potential for widespread application to envelope pseudotypes compatible with lentiviral vector production.

Generation of 2A-linked multicistronic cassettes by recombinant PCR.

PubMed

Szymczak-Workman, Andrea L; Vignali, Kate M; Vignali, Dario A A

2012-02-01

The need for reliable, multicistronic vectors for multigene delivery is at the forefront of biomedical technology. It is now possible to express multiple proteins from a single open reading frame (ORF) using 2A peptide-linked multicistronic vectors. These small sequences, when cloned between genes, allow for efficient, stoichiometric production of discrete protein products within a single vector through a novel "cleavage" event within the 2A peptide sequence. Expression of more than two genes using conventional approaches has several limitations, most notably imbalanced protein expression and large size. The use of 2A peptide sequences alleviates these concerns. They are small (18-22 amino acids) and have divergent amino-terminal sequences, which minimizes the chance for homologous recombination and allows for multiple, different 2A peptide sequences to be used within a single vector. Importantly, separation of genes placed between 2A peptide sequences is nearly 100%, which allows for stoichiometric and concordant expression of the genes, regardless of the order of placement within the vector. This protocol describes the use of recombinant polymerase chain reaction (PCR) to connect multiple 2A-linked protein sequences. The final construct is subcloned into an expression vector.

Statistical distribution of wind speeds and directions globally observed by NSCAT

NASA Astrophysics Data System (ADS)

Ebuchi, Naoto

1999-05-01

In order to validate wind vectors derived from the NASA scatterometer (NSCAT), statistical distributions of wind speeds and directions over the global oceans are investigated by comparing with European Centre for Medium-Range Weather Forecasts (ECMWF) wind data. Histograms of wind speeds and directions are calculated from the preliminary and reprocessed NSCAT data products for a period of 8 weeks. For wind speed of the preliminary data products, excessive low wind distribution is pointed out through comparison with ECMWF winds. A hump at the lower wind speed side of the peak in the wind speed histogram is discernible. The shape of the hump varies with incidence angle. Incompleteness of the prelaunch geophysical model function, SASS 2, tentatively used to retrieve wind vectors of the preliminary data products, is considered to cause the skew of the wind speed distribution. On the contrary, histograms of wind speeds of the reprocessed data products show consistent features over the whole range of incidence angles. Frequency distribution of wind directions relative to spacecraft flight direction is calculated to assess self-consistency of the wind directions. It is found that wind vectors of the preliminary data products exhibit systematic directional preference relative to antenna beams. This artificial directivity is also considered to be caused by imperfections in the geophysical model function. The directional distributions of the reprocessed wind vectors show less directivity and consistent features, except for very low wind cases.

A novel protocol for the production of recombinant LL-37 expressed as a thioredoxin fusion protein.

PubMed

Li, Yifeng

2012-02-01

LL-37 is the only cathelicidin-derived antimicrobial peptide found in humans and it has a multifunctional role in host defense. The peptide has been shown to possess immunomodulatory functions in addition to antimicrobial activity. To provide sufficient material for biological and structural characterization of this important peptide, various systems were developed to produce recombinant LL-37 in Escherichia coli. In one previous approach, LL-37 coding sequence was cloned into vector pET-32a, allowing the peptide to be expressed as a thioredoxin fusion. The fusion protein contains two thrombin cleavage sites: a vector-encoded one that is 30-residue upstream of the insert and an engineered one that is immediately adjacent to LL-37. Cleavage at these two sites shall generate three fragments, one of which is the target peptide. However, when the fusion protein was treated with thrombin, cleavage only occurred at the remote upstream site. A plausible explanation is that the thrombin site adjacent to LL-37 is less accessible due to the peptide's aggregation tendency and cleavage at the remote site generates a fragment, which forms a large aggregate that buries the intended site. In this study, I deleted the vector-encoded thrombin site and S tag in pET-32a, and then inserted the coding sequence for LL-37 plus a thrombin site into the modified vector. Although removing the S tag did not change the oligomeric state of the fusion protein, deletion of the vector-encoded thrombin site allowed the fusion to be cleaved at the engineered site to release LL-37. The released peptide was separated from the carrier and cleavage enzyme by size-exclusion chromatography. This new approach enables a quick production of high quality active LL-37 with a decent amount. Copyright © 2011 Elsevier Inc. All rights reserved.

Progress in developing cationic vectors for non-viral systemic gene therapy against cancer.

PubMed

Morille, Marie; Passirani, Catherine; Vonarbourg, Arnaud; Clavreul, Anne; Benoit, Jean-Pierre

2008-01-01

Initially, gene therapy was viewed as an approach for treating hereditary diseases, but its potential role in the treatment of acquired diseases such as cancer is now widely recognized. The understanding of the molecular mechanisms involved in cancer and the development of nucleic acid delivery systems are two concepts that have led to this development. Systemic gene delivery systems are needed for therapeutic application to cells inaccessible by percutaneous injection and for multi-located tumor sites, i.e. metastases. Non-viral vectors based on the use of cationic lipids or polymers appear to have promising potential, given the problems of safety encountered with viral vectors. Using these non-viral vectors, the current challenge is to obtain a similarly effective transfection to viral ones. Based on the advantages and disadvantages of existing vectors and on the hurdles encountered with these carriers, the aim of this review is to describe the "perfect vector" for systemic gene therapy against cancer.

Cloning and expression of soluble truncated variants of Borrelia OspA, OspB and Vmp7

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dunn, J.J.; Barbour, A.G.

1996-11-05

A method is provided for preparing soluble recombinant variations of Borrelia lipoproteins such as Borrelia burgdorferi outer surface protein A (OspA) and outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The method includes synthesizing a set of oligonucleotide primers, amplifying the template DNA utilizing the PCR, purifying the amplification products, cloning the amplification products into a suitable expression vector, transforming a suitable host utilizing the cloned expression vector, cultivating the transformed host for protein production and subsequently isolating and purifying the resulting protein. Also provided are soluble, recombinant variations of Borrelia burgdorferi outer surface proteinmore » A (OspA), outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The expression vectors harboring DNA encoding the recombinant variations, pET9-OspA, pET9-OspB and pET9-Vmp7, as well as the E. coli host BL21(DE3)/pLysS transformed with each of these vectors, are also disclosed. 38 figs.« less

Optical modular arithmetic

NASA Astrophysics Data System (ADS)

Pavlichin, Dmitri S.; Mabuchi, Hideo

2014-06-01

Nanoscale integrated photonic devices and circuits offer a path to ultra-low power computation at the few-photon level. Here we propose an optical circuit that performs a ubiquitous operation: the controlled, random-access readout of a collection of stored memory phases or, equivalently, the computation of the inner product of a vector of phases with a binary selector" vector, where the arithmetic is done modulo 2pi and the result is encoded in the phase of a coherent field. This circuit, a collection of cascaded interferometers driven by a coherent input field, demonstrates the use of coherence as a computational resource, and of the use of recently-developed mathematical tools for modeling optical circuits with many coupled parts. The construction extends in a straightforward way to the computation of matrix-vector and matrix-matrix products, and, with the inclusion of an optical feedback loop, to the computation of a weighted" readout of stored memory phases. We note some applications of these circuits for error correction and for computing tasks requiring fast vector inner products, e.g. statistical classification and some machine learning algorithms.

Cloning and expression of soluble truncated variants of Borrelia OspA, OspB and Vmp7

DOEpatents

Dunn, John J.; Barbour, Alan G.

1996-11-05

A method is provided herein for preparing soluble recombinant variations of Borrelia lipoproteins such as Borrelia burgdorferi outer surface protein A (OspA) and outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The method includes synthesizing a set of oligonucleotide primers, amplifying the template DNA utilizing the PCR, purifying the amplification products, cloning the amplification products into a suitable expression vector, transforming a suitable host utilizing the cloned expression vector, cultivating the transformed host for protein production and subsequently isolating and purifying the resulting protein. Also provided are soluble, recombinant variations of Borrelia burgdorferi outer surface protein A (OspA), outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The expression vectors harboring DNA encoding the recombinant variations, pET9-OspA, pET9-OspB and pET9-Vmp7, as well as the E. coli host BL21(DE3)/pLysS transformed with each of these vectors, are also disclosed.

Cloning and expression of soluble truncated variants of Borrelia OspA, OspB and Vmp7

DOEpatents

Dunn, J.J.; Barbour, A.G.

1996-11-05

A method is provided for preparing soluble recombinant variations of Borrelia lipoproteins such as Borrelia burgdorferi outer surface protein A (OspA) and outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The method includes synthesizing a set of oligonucleotide primers, amplifying the template DNA utilizing the PCR, purifying the amplification products, cloning the amplification products into a suitable expression vector, transforming a suitable host utilizing the cloned expression vector, cultivating the transformed host for protein production and subsequently isolating and purifying the resulting protein. Also provided are soluble, recombinant variations of Borrelia burgdorferi outer surface protein A (OspA), outer surface protein B (OspB), and B. hermsii variable major protein 7 (Vmp7). The expression vectors harboring DNA encoding the recombinant variations, pET9-OspA, pET9-OspB and pET9-Vmp7, as well as the E. coli host BL21(DE3)/pLysS transformed with each of these vectors, are also disclosed. 38 figs.

Doubly resonant three-wave-mixing spectroscopy of a chiral coupled-chromophore system in solution: coherent two-dimensional optical activity spectroscopy.

PubMed

Cheon, Sangheon; Lee, Hochan; Choi, Jun-Ho; Cho, Minhaeng

2007-02-07

Theoretical descriptions of doubly resonant two-dimensional (2D) sum-frequency-generation (SFG) and difference-frequency-generation (DFG) spectroscopies of coupled-chromophore systems are presented. Despite that each electronic or vibrational chromophore is achiral, the interaction-induced chirality of a coupled multichromophore system in solution can be measured by using the doubly resonant 2D three-wave-mixing (3WM) spectroscopic method. An electronically coupled dimer, where each monomer is modeled as a simple two-level system, can have nonvanishing SFG (or DFG) properties, e.g., susceptibility in frequency domain or nonlinear response function in time domain, if the induced dipole vector of the dimer is not orthogonal to the vector product of the two monomer electronic transition dipole vectors. In order to demonstrate that these 2D 3WM spectroscopic methods can be used to determine the solution structure of a polypeptide, the authors carried out quantum chemistry calculations for an alanine dipeptide and obtained first- and second-order dipole derivatives associated with the amide I vibrational transitions of the dipeptide. It is shown that the numerically simulated 2D IR-IR SFG spectrum is highly sensitive to the dipeptide secondary structure and provides rich information on the one- and two-exciton states. It is believed that the theoretically proposed doubly resonant 2D 3WM spectroscopy, which can be considered to be an optical activity spectroscopy, will be of use in studying both structural and dynamical aspects of coupled multichromophore systems, such as proteins, nucleic acids, nanoparticle aggregates etc.

Development and Application of Modern Optimal Controllers for a Membrane Structure Using Vector Second Order Form

NASA Astrophysics Data System (ADS)

Ferhat, Ipar

With increasing advancement in material science and computational power of current computers that allows us to analyze high dimensional systems, very light and large structures are being designed and built for aerospace applications. One example is a reflector of a space telescope that is made of membrane structures. These reflectors are light and foldable which makes the shipment easy and cheaper unlike traditional reflectors made of glass or other heavy materials. However, one of the disadvantages of membranes is that they are very sensitive to external changes, such as thermal load or maneuvering of the space telescope. These effects create vibrations that dramatically affect the performance of the reflector. To overcome vibrations in membranes, in this work, piezoelectric actuators are used to develop distributed controllers for membranes. These actuators generate bending effects to suppress the vibration. The actuators attached to a membrane are relatively thick which makes the system heterogeneous; thus, an analytical solution cannot be obtained to solve the partial differential equation of the system. Therefore, the Finite Element Model is applied to obtain an approximate solution for the membrane actuator system. Another difficulty that arises with very flexible large structures is the dimension of the discretized system. To obtain an accurate result, the system needs to be discretized using smaller segments which makes the dimension of the system very high. This issue will persist as long as the improving technology will allow increasingly complex and large systems to be designed and built. To deal with this difficulty, the analysis of the system and controller development to suppress the vibration are carried out using vector second order form as an alternative to vector first order form. In vector second order form, the number of equations that need to be solved are half of the number equations in vector first order form. Analyzing the system for control characteristics such as stability, controllability and observability is a key step that needs to be carried out before developing a controller. This analysis determines what kind of system is being modeled and the appropriate approach for controller development. Therefore, accuracy of the system analysis is very crucial. The results of the system analysis using vector second order form and vector first order form show the computational advantages of using vector second order form. Using similar concepts, LQR and LQG controllers, that are developed to suppress the vibration, are derived using vector second order form. To develop a controller using vector second order form, two different approaches are used. One is reducing the size of the Algebraic Riccati Equation to half by partitioning the solution matrix. The other approach is using the Hamiltonian method directly in vector second order form. Controllers are developed using both approaches and compared to each other. Some simple solutions for special cases are derived for vector second order form using the reduced Algebraic Riccati Equation. The advantages and drawbacks of both approaches are explained through examples. System analysis and controller applications are carried out for a square membrane system with four actuators. Two different systems with different actuator locations are analyzed. One system has the actuators at the corners of the membrane, the other has the actuators away from the corners. The structural and control effect of actuator locations are demonstrated with mode shapes and simulations. The results of the controller applications and the comparison of the vector first order form with the vector second order form demonstrate the efficacy of the controllers.

Evolving lessons on nanomaterial-coated viral vectors for local and systemic gene therapy

PubMed Central

Kasala, Dayananda; Yoon, A-Rum; Hong, Jinwoo; Kim, Sung Wan; Yun, Chae-Ok

2016-01-01

Viral vectors are promising gene carriers for cancer therapy. However, virus-mediated gene therapies have demonstrated insufficient therapeutic efficacy in clinical trials due to rapid dissemination to nontarget tissues and to the immunogenicity of viral vectors, resulting in poor retention at the disease locus and induction of adverse inflammatory responses in patients. Further, the limited tropism of viral vectors prevents efficient gene delivery to target tissues. In this regard, modification of the viral surface with nanomaterials is a promising strategy to augment vector accumulation at the target tissue, circumvent the host immune response, and avoid nonspecific interactions with the reticuloendothelial system or serum complement. In the present review, we discuss various chemical modification strategies to enhance the therapeutic efficacy of viral vectors delivered either locally or systemically. We conclude by highlighting the salient features of various nanomaterial-coated viral vectors and their prospects and directions for future research. PMID:27348247

Models for discrete-time self-similar vector processes with application to network traffic

NASA Astrophysics Data System (ADS)

Lee, Seungsin; Rao, Raghuveer M.; Narasimha, Rajesh

2003-07-01

The paper defines self-similarity for vector processes by employing the discrete-time continuous-dilation operation which has successfully been used previously by the authors to define 1-D discrete-time stochastic self-similar processes. To define self-similarity of vector processes, it is required to consider the cross-correlation functions between different 1-D processes as well as the autocorrelation function of each constituent 1-D process in it. System models to synthesize self-similar vector processes are constructed based on the definition. With these systems, it is possible to generate self-similar vector processes from white noise inputs. An important aspect of the proposed models is that they can be used to synthesize various types of self-similar vector processes by choosing proper parameters. Additionally, the paper presents evidence of vector self-similarity in two-channel wireless LAN data and applies the aforementioned systems to simulate the corresponding network traffic traces.

Development and Validation of Remote Sensing-Based Surface Inundation Products for Vector-Borne Disease Risk in East Africa

NASA Astrophysics Data System (ADS)

Jensen, K.; McDonald, K. C.; Ceccato, P.; Schroeder, R.; Podest, E.

2014-12-01

The potential impact of climate variability and change on the spread of infectious disease is of increasingly critical concern to public health. Newly-available remote sensing datasets may be combined with predictive modeling to develop new capabilities to mitigate risks of vector-borne diseases such as malaria, leishmaniasis, and rift valley fever. We have developed improved remote sensing-based products for monitoring water bodies and inundation dynamics that have potential utility for improving risk forecasts of vector-borne disease epidemics. These products include daily and seasonal surface inundation based on the global mappings of inundated area fraction derived at the 25-km scale from active and passive microwave instruments ERS, QuikSCAT, ASCAT, and SSM/I data - the Satellite Water Microwave Product Series (SWAMPS). Focusing on the East African region, we present validation of this product using multi-temporal classification of inundated areas in this region derived from high resolution PALSAR (100m) and Landsat (30m) observations. We assess historical occurrence of malaria in the east African country of Eritrea with respect to the time series SWAMPS datasets, and we aim to construct a framework for use of these new datasets to improve prediction of future malaria risk in this region. This work is supported through funding from the NASA Applied Sciences Program, the NASA Terrestrial Ecology Program, and the NASA Making Earth System Data Records for Use in Research Environments (MEaSUREs) Program. This study is also supported and monitored by National Oceanic and Atmospheric Administration (NOAA) under Grant - CREST Grant # NA11SEC4810004. The statements contained within the manuscript/research article are not the opinions of the funding agency or the U.S. government, but reflect the authors' opinions. This work was conducted in part under the framework of the ALOS Kyoto and Carbon Initiative. ALOS PALSAR data were provided by JAXA EORC.

HIV-1-based defective lentiviral vectors efficiently transduce human monocytes-derived macrophages and suppress replication of wild-type HIV-1

PubMed Central

Zeng, Lingbing; Planelles, Vicente; Sui, Ziye; Gartner, Suzanne; Maggirwar, Sanjay B.; Dewhurst, Stephen; Ye, Linbai; Nerurkar, Vivek R.; Yanagihara, Richard; Lu, Yuanan

2010-01-01

Background Human monocytes play an important role in mediating human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system (CNS), and monocytes-derived macrophages (MDM) represent a major viral reservoir within the brain and other target organs. Current gene transduction of MDM is hindered by a limited efficiency. In this study we established a lentiviral vector-based technique for improved gene transfer into human MDM cultures in vitro and demonstrated significant protection of transduced MDM from super-infection with wild-type HIV-1. Methods HIV-1-based lentiviral vector stocks were prepared in 293T cells by the established calcium phosphate transfection method. Human monocytes were isolated from donors' blood by Ficoll-Paque separation and cultured in vitro. To establish an effective technique for vector-mediated gene transfer, primary cultures of human MDM were transduced at varying multiplicities of infection (MOI) and at a range of time points following initial isolation of cells (time-in-culture). Transduced cells were then examined for transgene (green fluorescent protein (GFP)) expression by fluorescent microscopy and reverse transcription polymerase chain reaction (RT-PCR). These cultures were then exposed to wild-type HIV-1, and viral replication was quantitated by p24 assay; production of neurotoxic effector molecules by the transduced MDM was also examined, using indicator neurons. Results We have demonstrated that primary human MDM could be efficiently transduced (>50%) with concentrated HIV-1-based defective lentiviral vectors (DLV). Furthermore, DLV-mediated gene transduction was stable, and the transduced cells exhibited no apparent difference from normal MDM in terms of their morphology, viability and neurotoxin secretion. Challenge of DLV-transduced MDM cultures with HIV-1Ba-L revealed a 4- to 5-fold reduction in viral replication, as measured by p24 antigen production. This effect was associated with the mobilization of the GFP-expressing DLV construct by the wild-type virus. Conclusions These data demonstrate the inhibition of HIV-1 replication in primary MDM, by a DLV vector that lacks any anti-HIV-1 transgene. These findings lay the initial groundwork for future studies on the ability of DLV-modified monocytes to introduce anti-HIV-1 genes into the CNS. Lentiviral vector-mediated gene delivery to the CNS by monocytes/macrophages is a promising, emerging strategy for treating neuro-AIDS. PMID:16142830

Production of recombinant allergens in plants.

PubMed

Schmidt, Georg; Gadermaier, Gabriele; Pertl, Heidi; Siegert, Marc; Oksman-Caldentey, Kirsi-Marja; Ritala, Anneli; Himly, Martin; Obermeyer, Gerhard; Ferreira, Fatima

2008-10-01

A large percentage of allergenic proteins are of plant origin. Hence, plant-based expression systems are considered ideal for the recombinant production of certain allergens. First attempts to establish production of plant-derived allergens in plants focused on transient expression in Nicotiana benthamiana infected with recombinant viral vectors. Accordingly, allergens from birch and mugwort pollen, as well as from apple have been expressed in plants. Production of house dust mite allergens has been achieved by Agrobacterium-mediated transformation of tobacco plants. Beside the use of plants as production systems, other approaches have focused on the development of edible vaccines expressing allergens or epitopes thereof, which bypasses the need of allergen purification. The potential of this approach has been convincingly demonstrated for transgenic rice seeds expressing seven dominant human T cell epitopes derived from Japanese cedar pollen allergens. Parallel to efforts in developing recombinant-based diagnostic and therapeutic reagents, different gene-silencing approaches have been used to decrease the expression of allergenic proteins in allergen sources. In this way hypoallergenic ryegrass, soybean, rice, apple, and tomato were developed.

Decomposition-aggregation stability analysis. [for large scale dynamic systems with application to spinning Skylab control system

NASA Technical Reports Server (NTRS)

Siljak, D. D.; Weissenberger, S.; Cuk, S. M.

1973-01-01

This report presents the development and description of the decomposition aggregation approach to stability investigations of high dimension mathematical models of dynamic systems. The high dimension vector differential equation describing a large dynamic system is decomposed into a number of lower dimension vector differential equations which represent interconnected subsystems. Then a method is described by which the stability properties of each subsystem are aggregated into a single vector Liapunov function, representing the aggregate system model, consisting of subsystem Liapunov functions as components. A linear vector differential inequality is then formed in terms of the vector Liapunov function. The matrix of the model, which reflects the stability properties of the subsystems and the nature of their interconnections, is analyzed to conclude over-all system stability characteristics. The technique is applied in detail to investigate the stability characteristics of a dynamic model of a hypothetical spinning Skylab.

A study of single-meson production in neutrino and antineutrino charged-current interactions on protons

NASA Astrophysics Data System (ADS)

Allen, P.; Grässler, H.; Schulte, R.; Jones, G. T.; Kennedy, B. W.; O'Neale, S. W.; Gebel, W.; Hofmann, E.; Klein, H.; Mittendorfer, J.; Morrison, D. R. O.; Schmid, P.; Wachsmuth, H.; Barnham, K. W. J.; Clayton, E. F.; Hamisi, F.; Miller, D. B.; Mobayyen, M. M.; Aderholz, M.; Deck, L.; Schmitz, N.; Wittek, W.; Corrigan, G.; Myatt, G.; Radojicic, D.; Saitta, B.; Shotton, P. N.; Towers, S. J.; Aachen-Birmingham-Bonn-CERN-London IC-Munich (MPI)-Oxford Collaboration

1986-01-01

We present results on exclusive single-charged pion and kaon production in neutrino and antineutrino interactions on protons in the energy range from 5 to 120 GeV. The data were obtained from exposures of BEBC to wide band beams at the CERN SPS. For invariant masses of the (pπ) system below 2 GeV, the pions originate predominantly from decays of baryon resonances excited by the weak charged current. Similarly, we observe the production of Λ(1520) decaying into p and K -. For invariant masses above 2 GeV pion production becomes peripheral by interaction of the weak current with a virtual π0. We establish a contribution of longitudinally polarised intermediate vector bosons to this process.

Vector breather-to-soliton transitions and nonlinear wave interactions induced by higher-order effects in an erbium-doped fiber

NASA Astrophysics Data System (ADS)

Sun, Wen-Rong; Wang, Lei; Xie, Xi-Yang

2018-06-01

Vector breather-to-soliton transitions for the higher-order nonlinear Schrödinger-Maxwell-Bloch (NLS-MB) system with sextic terms are investigated. The Lax pair and Darboux transformation (DT) of such system are constructed. With the DT, analytic vector breather solutions up to the second order are obtained. With appropriate choices of the spectra parameters, vector breather-to-soliton transitions happen. Interaction mechanisms of vector nonlinear waves (breather-soliton or soliton-soliton interactions) are displayed.

The Helioseismic and Magnetic Imager (HMI) Vector Magnetic Field Pipeline: Overview and Performance

NASA Astrophysics Data System (ADS)

Hoeksema, J. Todd; Liu, Yang; Hayashi, Keiji; Sun, Xudong; Schou, Jesper; Couvidat, Sebastien; Norton, Aimee; Bobra, Monica; Centeno, Rebecca; Leka, K. D.; Barnes, Graham; Turmon, Michael

2014-09-01

The Helioseismic and Magnetic Imager (HMI) began near-continuous full-disk solar measurements on 1 May 2010 from the Solar Dynamics Observatory (SDO). An automated processing pipeline keeps pace with observations to produce observable quantities, including the photospheric vector magnetic field, from sequences of filtergrams. The basic vector-field frame list cadence is 135 seconds, but to reduce noise the filtergrams are combined to derive data products every 720 seconds. The primary 720 s observables were released in mid-2010, including Stokes polarization parameters measured at six wavelengths, as well as intensity, Doppler velocity, and the line-of-sight magnetic field. More advanced products, including the full vector magnetic field, are now available. Automatically identified HMI Active Region Patches (HARPs) track the location and shape of magnetic regions throughout their lifetime. The vector field is computed using the Very Fast Inversion of the Stokes Vector (VFISV) code optimized for the HMI pipeline; the remaining 180∘ azimuth ambiguity is resolved with the Minimum Energy (ME0) code. The Milne-Eddington inversion is performed on all full-disk HMI observations. The disambiguation, until recently run only on HARP regions, is now implemented for the full disk. Vector and scalar quantities in the patches are used to derive active region indices potentially useful for forecasting; the data maps and indices are collected in the SHARP data series, hmi.sharp_720s. Definitive SHARP processing is completed only after the region rotates off the visible disk; quick-look products are produced in near real time. Patches are provided in both CCD and heliographic coordinates. HMI provides continuous coverage of the vector field, but has modest spatial, spectral, and temporal resolution. Coupled with limitations of the analysis and interpretation techniques, effects of the orbital velocity, and instrument performance, the resulting measurements have a certain dynamic range and sensitivity and are subject to systematic errors and uncertainties that are characterized in this report.

Geographical distribution of Culicoides (DIPTERA: CERATOPOGONIDAE) in mainland Portugal: Presence/absence modelling of vector and potential vector species.

PubMed

Ramilo, David W; Nunes, Telmo; Madeira, Sara; Boinas, Fernando; da Fonseca, Isabel Pereira

2017-01-01

Vector-borne diseases are not only accounted responsible for their burden on human health-care systems, but also known to cause economic constraints to livestock and animal production. Animals are affected directly by the transmitted pathogens and indirectly when animal movement is restricted. Distribution of such diseases depends on climatic and social factors, namely, environmental changes, globalization, trade and unplanned urbanization. Culicoides biting midges are responsible for the transmission of several pathogenic agents with relevant economic impact. Due to a fragmentary knowledge of their ecology, occurrence is difficult to predict consequently, limiting the control of these arthropod vectors. In order to understand the distribution of Culicoides species, in mainland Portugal, data collected during the National Entomologic Surveillance Program for Bluetongue disease (2005-2013), were used for statistical evaluation. Logistic regression analysis was preformed and prediction maps (per season) were obtained for vector and potentially vector species. The variables used at the present study were selected from WorldClim (two climatic variables) and CORINE databases (twenty-two land cover variables). This work points to an opposite distribution of C. imicola and species from the Obsoletus group within mainland Portugal. Such findings are evidenced in autumn, with the former appearing in Central and Southern regions. Although appearing northwards, on summer and autumn, C. newsteadi reveals a similar distribution to C. imicola. The species C. punctatus appears in all Portuguese territory throughout the year. Contrary, C. pulicaris is poorly caught in all areas of mainland Portugal, being paradoxical present near coastal areas and higher altitude regions.

Geographical distribution of Culicoides (DIPTERA: CERATOPOGONIDAE) in mainland Portugal: Presence/absence modelling of vector and potential vector species

PubMed Central

Madeira, Sara; Boinas, Fernando; da Fonseca, Isabel Pereira

2017-01-01

Vector-borne diseases are not only accounted responsible for their burden on human health-care systems, but also known to cause economic constraints to livestock and animal production. Animals are affected directly by the transmitted pathogens and indirectly when animal movement is restricted. Distribution of such diseases depends on climatic and social factors, namely, environmental changes, globalization, trade and unplanned urbanization. Culicoides biting midges are responsible for the transmission of several pathogenic agents with relevant economic impact. Due to a fragmentary knowledge of their ecology, occurrence is difficult to predict consequently, limiting the control of these arthropod vectors. In order to understand the distribution of Culicoides species, in mainland Portugal, data collected during the National Entomologic Surveillance Program for Bluetongue disease (2005–2013), were used for statistical evaluation. Logistic regression analysis was preformed and prediction maps (per season) were obtained for vector and potentially vector species. The variables used at the present study were selected from WorldClim (two climatic variables) and CORINE databases (twenty-two land cover variables). This work points to an opposite distribution of C. imicola and species from the Obsoletus group within mainland Portugal. Such findings are evidenced in autumn, with the former appearing in Central and Southern regions. Although appearing northwards, on summer and autumn, C. newsteadi reveals a similar distribution to C. imicola. The species C. punctatus appears in all Portuguese territory throughout the year. Contrary, C. pulicaris is poorly caught in all areas of mainland Portugal, being paradoxical present near coastal areas and higher altitude regions. PMID:28683145

Development of the gateway recycling cloning system for multiple linking of expression cassettes in a defined order, and direction on gateway compatible binary vectors.

PubMed

Kimura, Tetsuya; Nakao, Akihide; Murata, Sachiko; Kobayashi, Yasuyuki; Tanaka, Yuji; Shibahara, Kenta; Kawazu, Tetsu; Nakagawa, Tsuyoshi

2013-01-01

We developed the Gateway recycling cloning system, which allows multiple linking of expression cassettes by multiple rounds of the Gateway LR reaction. Employing this system, the recycling donor vector pRED419 was subjected to the first LR reaction with an attR1-attR2 type destination vector. Then conversion vector pCON was subjected to an LR reaction to restore the attR1-attR2 site on the destination vector for the next cloning cycle. By repetition of these two simple steps, we linked four expression cassettes of a reporter gene in Gateway binary vector pGWB1, introduced the constructs into tobacco BY-2 cells, and observed the expression of transgenes.

Word-level recognition of multifont Arabic text using a feature vector matching approach

NASA Astrophysics Data System (ADS)

Erlandson, Erik J.; Trenkle, John M.; Vogt, Robert C., III

1996-03-01

Many text recognition systems recognize text imagery at the character level and assemble words from the recognized characters. An alternative approach is to recognize text imagery at the word level, without analyzing individual characters. This approach avoids the problem of individual character segmentation, and can overcome local errors in character recognition. A word-level recognition system for machine-printed Arabic text has been implemented. Arabic is a script language, and is therefore difficult to segment at the character level. Character segmentation has been avoided by recognizing text imagery of complete words. The Arabic recognition system computes a vector of image-morphological features on a query word image. This vector is matched against a precomputed database of vectors from a lexicon of Arabic words. Vectors from the database with the highest match score are returned as hypotheses for the unknown image. Several feature vectors may be stored for each word in the database. Database feature vectors generated using multiple fonts and noise models allow the system to be tuned to its input stream. Used in conjunction with database pruning techniques, this Arabic recognition system has obtained promising word recognition rates on low-quality multifont text imagery.

Sequential cloning of chromosomes

DOEpatents

Lacks, S.A.

1995-07-18

A method for sequential cloning of chromosomal DNA of a target organism is disclosed. A first DNA segment homologous to the chromosomal DNA to be sequentially cloned is isolated. The first segment has a first restriction enzyme site on either side. A first vector product is formed by ligating the homologous segment into a suitably designed vector. The first vector product is circularly integrated into the target organism`s chromosomal DNA. The resulting integrated chromosomal DNA segment includes the homologous DNA segment at either end of the integrated vector segment. The integrated chromosomal DNA is cleaved with a second restriction enzyme and ligated to form a vector-containing plasmid, which is replicated in a host organism. The replicated plasmid is then cleaved with the first restriction enzyme. Next, a DNA segment containing the vector and a segment of DNA homologous to a distal portion of the previously isolated DNA segment is isolated. This segment is then ligated to form a plasmid which is replicated within a suitable host. This plasmid is then circularly integrated into the target chromosomal DNA. The chromosomal DNA containing the circularly integrated vector is treated with a third, retrorestriction (class IIS) enzyme. The cleaved DNA is ligated to give a plasmid that is used to transform a host permissive for replication of its vector. The sequential cloning process continues by repeated cycles of circular integration and excision. The excision is carried out alternately with the second and third enzymes. 9 figs.

Practical utilization of recombinant AAV vector reference standards: focus on vector genomes titration by free ITR qPCR.

PubMed

D'Costa, Susan; Blouin, Veronique; Broucque, Frederic; Penaud-Budloo, Magalie; François, Achille; Perez, Irene C; Le Bec, Christine; Moullier, Philippe; Snyder, Richard O; Ayuso, Eduard

2016-01-01

Clinical trials using recombinant adeno-associated virus (rAAV) vectors have demonstrated efficacy and a good safety profile. Although the field is advancing quickly, vector analytics and harmonization of dosage units are still a limitation for commercialization. AAV reference standard materials (RSMs) can help ensure product safety by controlling the consistency of assays used to characterize rAAV stocks. The most widely utilized unit of vector dosing is based on the encapsidated vector genome. Quantitative polymerase chain reaction (qPCR) is now the most common method to titer vector genomes (vg); however, significant inter- and intralaboratory variations have been documented using this technique. Here, RSMs and rAAV stocks were titered on the basis of an inverted terminal repeats (ITRs) sequence-specific qPCR and we found an artificial increase in vg titers using a widely utilized approach. The PCR error was introduced by using single-cut linearized plasmid as the standard curve. This bias was eliminated using plasmid standards linearized just outside the ITR region on each end to facilitate the melting of the palindromic ITR sequences during PCR. This new "Free-ITR" qPCR delivers vg titers that are consistent with titers obtained with transgene-specific qPCR and could be used to normalize in-house product-specific AAV vector standards and controls to the rAAV RSMs. The free-ITR method, including well-characterized controls, will help to calibrate doses to compare preclinical and clinical data in the field.

A Vector Approach to Euclidean Geometry: Inner Product Spaces, Euclidean Geometry and Trigonometry, Volume 2. Teacher's Edition.

ERIC Educational Resources Information Center

Vaughan, Herbert E.; Szabo, Steven

This is the teacher's edition of a text for the second year of a two-year high school geometry course. The course bases plane and solid geometry and trigonometry on the fact that the translations of a Euclidean space constitute a vector space which has an inner product. Congruence is a geometric topic reserved for Volume 2. Volume 2 opens with an…

Spin asymmetries for vector boson production in polarized p + p collisions

DOE PAGES

Huang, Jin; Kang, Zhong-Bo; Vitev, Ivan; ...

2016-01-28

We study the cross section for vector boson (W ±/Z 0/γ more » $$\\star$$) production in polarized nucleon-nucleon collisions for low transverse momentum of the observed vector boson. For the case where one measures the transverse momentum and azimuthal angle of the vector bosons, we present the cross sections and the associated spin asymmetries in terms of transverse momentum dependent parton distribution functions (TMDs) at tree level within the TMD factorization formalism. To assess the feasibility of experimental measurements, we estimate the spin asymmetries forW ±/Z 0 boson production in polarized proton-proton collisions at the Relativistic Heavy Ion Collider by using current knowledge of the relevant TMDs. Here, we find that some of these asymmetries can be sizable if the suppression effect from TMD evolution is not too strong. The W program at RHIC can, thus, test and constrain spin theory by providing unique information on the universality properties of TMDs, TMD evolution, and the nucleon structure. For example, the single transverse spin asymmetries could be used to probe the well-known Sivers function f$$⊥q\\atop{1T}$$, as well as the transversal helicity distribution g$$q\\atop{1T}$$ via the parity-violating nature of W production.« less

Interaction of the Lyme disease spirochete with its tick vector.

PubMed

Caimano, Melissa J; Drecktrah, Dan; Kung, Faith; Samuels, D Scott

2016-07-01

Borrelia burgdorferi, the causative agent of Lyme disease (along with closely related genospecies), is in the deeply branching spirochete phylum. The bacterium is maintained in nature in an enzootic cycle that involves transmission from a tick vector to a vertebrate host and acquisition from a vertebrate host to a tick vector. During its arthropod sojourn, B. burgdorferi faces a variety of stresses, including nutrient deprivation. Here, we review some of the spirochetal factors that promote persistence, maintenance and dissemination of B. burgdorferi in the tick, and then focus on the utilization of available carbohydrates as well as the exquisite regulatory systems invoked to adapt to the austere environment between blood meals and to signal species transitions as the bacteria traverse their enzootic cycle. The spirochetes shift their source of carbon and energy from glucose in the vertebrate to glycerol in the tick. Regulation of survival under limiting nutrients requires the classic stringent response in which RelBbu controls the levels of the alarmones guanosine tetraphosphate and guanosine pentaphosphate (collectively termed (p)ppGpp), while regulation at the tick-vertebrate interface as well as regulation of protective responses to the blood meal require the two-component system Hk1/Rrp1 to activate production of the second messenger cyclic-dimeric-GMP (c-di-GMP). © 2016 John Wiley & Sons Ltd.

Evaluating the Operational Content of Contestable Market Theory.

DTIC Science & Technology

1986-12-01

inputs needed for production would be avialable to any firm at the market price ; and ii) The cost of collusion amoung the producers is prohibitively high... advertising costs or increase the quality (and hence the costs) of his product to produce a perfect 9 substitute for the entrant’s good. Thus, free entry alone...n x 1) vector, where n is the number of different products produced by the firm. A firm’s output is denoted by X. Similarly, price is a (n x 1) vector

Measurement of Vector Boson Plus $$D^{*}(2010)^+$$ Meson Production in $$\\bar{p}p$$ Collisions at $$\\sqrt{s}=1.96\\, {\\rm TeV}$$

DOE PAGES

Aaltonen, T.

2016-03-21

Our study of vector boson (V ) production in conjunction with a D*(2010) + meson is presented. Using a data sample correponding to 9.7 fb -1 p of proton-antiproton collisions at center-of-mass energy √s = 1:96 TeV produced by the Fermilab Tevatron, we reconstruct V +D *+ samples with the CDF II detector. The D *+ is fully reconstructed in the D*(2010) + → D 0(→ K - π +)π + decay mode. This technique is sensitive to the associated production of vector boson plus charm or bottom mesons. We measure the ratio of production cross sections σ(W +D *)/more » σ(W) = [1.75±0.13(stat)±0:09(syst)]% and σ(Z +D *)/ σ(Z) = [1:5±0:4(stat)_0:2(syst)]%. Event properties are utilized to determine the fraction of V +D *(2010) + events originating from different production processes. Our results are in agreement with the predictions obtained with the pythia program, limiting possible contribution from non-standard-model physics processes.« less

Measurement of Vector Boson Plus $$D^{*}(2010)^+$$ Meson Production in $$\\bar{p}p$$ Collisions at $$\\sqrt{s}=1.96\\, {\\rm TeV}$$

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aaltonen, T.

Our study of vector boson (V ) production in conjunction with a D*(2010) + meson is presented. Using a data sample correponding to 9.7 fb -1 p of proton-antiproton collisions at center-of-mass energy √s = 1:96 TeV produced by the Fermilab Tevatron, we reconstruct V +D *+ samples with the CDF II detector. The D *+ is fully reconstructed in the D*(2010) + → D 0(→ K - π +)π + decay mode. This technique is sensitive to the associated production of vector boson plus charm or bottom mesons. We measure the ratio of production cross sections σ(W +D *)/more » σ(W) = [1.75±0.13(stat)±0:09(syst)]% and σ(Z +D *)/ σ(Z) = [1:5±0:4(stat)_0:2(syst)]%. Event properties are utilized to determine the fraction of V +D *(2010) + events originating from different production processes. Our results are in agreement with the predictions obtained with the pythia program, limiting possible contribution from non-standard-model physics processes.« less

A flippon related singlet at the LHC II

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Tianjun; Maxin, James A.; Mayes, Van E.

2016-06-28

Here, we consider the 750 GeV diphoton resonance at the 13 TeV LHC in the ℱ-SU(5) model with a Standard Model (SM) singlet field which couples to TeV-scale vector-like particles, dubbed flippons. This singlet field assumes the role of the 750 GeV resonance, with production via gluon fusion and subsequent decay to a diphoton via the vector-like particle loops. We present a numerical analysis showing that the observed 8 TeV and 13 TeV diphoton production cross-sections can be generated in the model space with realistic electric charges and Yukawa couplings for light vector-like masses. We further discuss the experimental viabilitymore » of light vector-like masses in a General No-Scale ℱ-SU(5) model, offering a few benchmark scenarios in this consistent GUT that can satisfy all experimental constraints imposed by the LHC and other essential experiments.« less

Coherent vector meson photoproduction from deuterium at intermediate energies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rogers, T.C.; Strikman, M.I.; Sargsian, M.M.

2006-04-15

We analyze the cross section for vector meson photoproduction off a deuteron for the intermediate range of photon energies starting at a few giga-electron-volts above the threshold and higher. We reproduce the steps in the derivation of the conventional nonrelativistic Glauber expression based on an effective diagrammatic method while making corrections for Fermi motion and intermediate-energy kinematic effects. We show that, for intermediate-energy vector meson production, the usual Glauber factorization breaks down, and we derive corrections to the usual Glauber method to linear order in longitudinal nucleon momentum. The purpose of our analysis is to establish methods for probing interestingmore » physics in the production mechanism for {phi} mesons and heavier vector mesons. We demonstrate how neglecting the breakdown of Glauber factorization can lead to errors in measurements of basic cross sections extracted from nuclear data.« less

Vector solitons in femtosecond fibre lasers

NASA Astrophysics Data System (ADS)

Chen, W. C.; Xu, W. C.; Song, F.; Shen, M. C.; Han, D. A.; Chen, L. B.

2008-07-01

Experimental observation of spectral sideband suppression of mode-locked pulses is obtained in an erbium-doped fibre ring laser with nonlinear polarization rotation techniques. This effect may indicate the formation of a vector soliton in accordance with the theoretical work of reference [Phys. Rev. E 74, 046605 (2006)]. The 3 dB spectral bandwidth, the central wavelength and the repetition rate of the vector solitons are 24.41 nm, 1565.14 nm and 12.15 MHz, respectively. Based on the experimental observations, we propose an experimental criterion for the production of vector solitons, with spectral sideband suppression as a sign of the generation of vector solitons.

Agroinfiltration: a rapid and reliable method to select suitable rose cultivars for blue flower production.

PubMed

Zeinipour, Masoume; Azadi, Pejman; Majd, Ahmad; Kermani, Maryam Jafarkhani; Irian, Saeed; Hosseini, Seyed Mohammad; Mii, Masahiro

2018-05-01

Rose cultivars with blue flower color are among the most attractive breeding targets in floriculture. However, they are difficult to produce due to the low efficiency of transformation systems, interactive effects of hosts and vectors, and lengthy processes. In this study, agroinfiltration-mediated transient expression was investigated as a tool to assess the function of flower color genes and to determine appropriate host cultivars for stable transformation in Rosa hybrida . To induce delphinidin accumulation and consequently to produce blue hue, the petals of 30 rose cultivars were infiltrated with three different expression vectors namely pBIH-35S-CcF3'5'H, pBIH-35S-Del2 and pBIH-35S-Del8, harbouring different sets of flower color genes. The results obtained showed that the ectopic expression of the genes was only detected in three cultivars with dark pink petals (i.e. 'Purple power', 'High & Mora' and 'Marina') after 6-8 days. The high performance liquid chromatography analyses confirmed delphinidin accumulation in the infiltrated petals caused by transient expression of CcF3'5'H gene. Moreover, there were significant differences in the amounts of delphinidin among the three cultivars infiltrated with the three different expression vectors. More specifically, the highest delphinidin content was detected in the cultivar 'Purple power' (4.67 µg g -1 FW), infiltrated with the pBIH-35S-Del2 vector. The expression of CcF3'5'H gene in the infiltrated petals was also confirmed by real time PCR. In conclusion and based on the findings of the present study, the agroinfiltration could be regarded as a reliable method to identify suitable rose cultivars in blue rose flower production programs.

Brucella lipopolysaccharide reinforced Salmonella delivering Brucella immunogens protects mice against virulent challenge.

PubMed

Lalsiamthara, Jonathan; Lee, John Hwa

2017-06-01

Intracellular pathogen Salmonella exhibits natural infection broadly analogous to Brucella, this phenomenon makes Salmonella a pragmatic choice for an anti-Brucella vaccine delivery platform. In this study we developed and formulated a combination of four attenuated Salmonella Typhimurium live vector strains delivering heterologous Brucella antigens (rBs), namely lumazine synthase, proline racemase subunit A, lipoprotein outer membrane protein-19, and Cu-Zn superoxide dismutase. With an aim to develop a cross-protecting vaccine, Brucella pan-species conserved rBs were selected. The present study compared the efficacy of smooth and rough variants of Salmonella delivery vector and also evaluated the inclusion of purified Brucella lipopolysaccharide (LPS) in the formulation. Immunization of SPF-BALB/c mice with the vaccine combinations significantly (P≤0.05) reduced splenic wild-type Brucella abortus 544 colonization as compared to non-immunized mice as well as Salmonella only immunized mice. Increased induction of Brucella specific-IgG, sIgA production, and antigen-specific splenocyte proliferative responses were observed in the mice immunized with the formulations as compared to naïve or vector only immunized mice. Modulatory effects of rB and LPS on production of interleukin (IL)-4, IL-12, and interferon-γ were detected in splenocytes of mice immunized with the formulation. Rough Salmonella variant in combination with LPS could further enhance the efficacy of the delivery when applied intraperitoneally. Taken together, it is compelling that Brucella LPS-augmented Salmonella vector delivering immunogenic Brucella proteins may be more suitable than the current non-ideal live Brucella abortus vaccine. The vaccine system also provides a basis for the development of cross-protecting vaccine capable of preventing multispecies brucellosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Gene Editing Vectors for Studying Nicotinic Acetylcholine Receptors in Cholinergic Transmission.

PubMed

Peng, Can; Yan, Yijin; Kim, Veronica J; Engle, Staci E; Berry, Jennifer N; McIntosh, J Michael; Neve, Rachael L; Drenan, Ryan M

2018-05-19

Nicotinic acetylcholine receptors (nAChRs), prototype members of the cys-loop ligand gated ion channel family, are key mediators of cholinergic transmission in the central nervous system. Despite their importance, technical gaps exist in our ability to dissect the function of individual subunits in the brain. To overcome these barriers, we designed CRISPR/Cas9 small guide RNA sequences (sgRNAs) for production of loss-of-function alleles in mouse nAChR genes. These sgRNAs were validated in vitro via deep sequencing. We subsequently targeted candidate nAChR genes in vivo by creating herpes simplex virus (HSV) vectors delivering sgRNAs and Cas9 expression to mouse brain. Production of loss-of-function insertions or deletions (indels) by these "all-in-one" HSV vectors was confirmed using brain slice patch clamp electrophysiology coupled with pharmacological analysis. Next, we developed a scheme for cell type-specific gene editing in mouse brain. Knockin mice expressing Cas9 in a Cre-dependent manner were validated using viral microinjections and genetic crosses to common Cre-driver mouse lines. We subsequently confirmed functional Cas9 activity by targeting the ubiquitous neuronal protein, NeuN, using adeno associated virus (AAV) delivery of sgRNAs. Finally, the mouse β2 nAChR gene was successfully targeted in dopamine transporter (DAT) positive neurons via CRISPR/Cas9. The sgRNA sequences and viral vectors, including our scheme for Cre-dependent gene editing, should be generally useful to the scientific research community. These tools could lead to new discoveries related to the function of nAChRs in neurotransmission and behavioral processes. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Reverse Genetics for Mammalian Orthoreovirus.

PubMed

Stuart, Johnasha D; Phillips, Matthew B; Boehme, Karl W

2017-01-01

Reverse genetics allows introduction of specific alterations into a viral genome. Studies performed with mutant viruses generated using reverse genetics approaches have contributed immeasurably to our understanding of viral replication and pathogenesis, and also have led to development of novel vaccines and virus-based vectors. Here, we describe the reverse genetics system that allows for production and recovery of mammalian orthoreovirus, a double-stranded (ds) RNA virus, from plasmids that encode the viral genome.

The components of shear stress affecting insect cells used with the baculovirus expression vector system.

PubMed

Weidner, Tobias; Druzinec, Damir; Mühlmann, Martina; Buchholz, Rainer; Czermak, Peter

2017-09-26

Insect-based expression platforms such as the baculovirus expression vector system (BEVS) are widely used for the laboratory- and industrial-scale production of recombinant proteins. Thereby, major drawbacks to gain high-quality proteins are the lytic infection cycle and the shear sensitivity of infected insect cells due to turbulence and aeration. Smaller bubbles were formerly assumed to be more harmful than larger ones, but we found that cell damage is also dependent on the concentration of protective agents such as Pluronic®. At the appropriate concentration, Pluronic forms a layer around air bubbles and hinders the attachment of cells, thus limiting the damage. In this context, we used microaeration to vary bubble sizes and confirmed that size is not the most important factor, but the total gas surface area in the reactor is. If the surface area exceeds a certain threshold, the concentration of Pluronic is no longer sufficient for cell protection. To investigate the significance of shear forces, a second study was carried out in which infected insect cells were cultivated in a hollow fiber module to protect them from shear forces. Both model studies revealed important aspects of the design and scale-up of BEVS processes for the production of recombinant proteins.

Imaging chemical reactions - 3D velocity mapping

NASA Astrophysics Data System (ADS)

Chichinin, A. I.; Gericke, K.-H.; Kauczok, S.; Maul, C.

Visualising a collision between an atom or a molecule or a photodissociation (half-collision) of a molecule on a single particle and single quantum level is like watching the collision of billiard balls on a pool table: Molecular beams or monoenergetic photodissociation products provide the colliding reactants at controlled velocity before the reaction products velocity is imaged directly with an elaborate camera system, where one should keep in mind that velocity is, in general, a three-dimensional (3D) vectorial property which combines scattering angles and speed. If the processes under study have no cylindrical symmetry, then only this 3D product velocity vector contains the full information of the elementary process under study.

Resonance Production in Heavy-Ion Collisions

NASA Astrophysics Data System (ADS)

Knospe, Anders G.

2018-02-01

Hadronic resonances are unique probes that allow the properties of heavyion collisions to be studied. Topics that can be studied include modification of spectral shapes, in-medium energy loss of parsons, vector-meson spin alignment, hydrodynamic flow, recombination, strangeness production, and the properties of the hadronic phase. Measurements of resonances in p+p, p+A, and d+A collisions serve as baselines for heavy-ion studies and also permit searches for possible collective effects in these smaller systems. These proceedings present a selection of results related to these topics from experiments at RHIC, LHC, and other facilities, as well as comparisons to theoretical models.

Tightly regulated, high-level expression from controlled copy number vectors based on the replicon of temperate phage N15.

PubMed

Mardanov, Andrey V; Strakhova, Taisia S; Smagin, Vladimir A; Ravin, Nikolai V

2007-06-15

A new Escherichia coli host/vector system has been developed to allow a dual regulation of both the plasmid copy number and gene expression. The new pN15E vectors are low copy number plasmids based on the replicon of temperate phage N15, comprising the repA replicase gene and cB repressor gene, controlling the plasmid copy number. Regulation of pN15E copy number is achieved through arabinose-inducible expression of phage N15 antirepressor protein, AntA, whose gene was integrated into the chromosome of the host strain under control of the PBAD promoter. The host strain also carried phage N15 partition operon, sop, allowing stable inheritance of pN15E vectors in the absence of selection pressure. In the first vector, pN15E4, the same PBAD promoter controls expression of a cloned gene. The second vector, pN15E6, carries the phage T5 promoter with a double lac operator repression module thus allowing independent regulation of promoter activity and copy number. Using the lacZ gene to monitor expression in these vectors, we show that the ratio of induction/repression can be about 7600-fold for pN15E4 and more than 15,000-fold for pN15E6. The low copy number of these vectors ensures very low basal level of expression allowing cloning genes encoding toxic products that was demonstrated by the stable maintenance of a gene encoding a restriction endonuclease in pN15E4. The tight control of transcription and the potential to regulate gene activities quantitatively over wide ranges will open up new approaches in the study of gene function in vivo and controlled expression of heterologous genes.

Heterologous expression of pikromycin biosynthetic gene cluster using Streptomyces artificial chromosome system.

PubMed

Pyeon, Hye-Rim; Nah, Hee-Ju; Kang, Seung-Hoon; Choi, Si-Sun; Kim, Eung-Soo

2017-05-31

Heterologous expression of biosynthetic gene clusters of natural microbial products has become an essential strategy for titer improvement and pathway engineering of various potentially-valuable natural products. A Streptomyces artificial chromosomal conjugation vector, pSBAC, was previously successfully applied for precise cloning and tandem integration of a large polyketide tautomycetin (TMC) biosynthetic gene cluster (Nah et al. in Microb Cell Fact 14(1):1, 2015), implying that this strategy could be employed to develop a custom overexpression scheme of natural product pathway clusters present in actinomycetes. To validate the pSBAC system as a generally-applicable heterologous overexpression system for a large-sized polyketide biosynthetic gene cluster in Streptomyces, another model polyketide compound, the pikromycin biosynthetic gene cluster, was preciously cloned and heterologously expressed using the pSBAC system. A unique HindIII restriction site was precisely inserted at one of the border regions of the pikromycin biosynthetic gene cluster within the chromosome of Streptomyces venezuelae, followed by site-specific recombination of pSBAC into the flanking region of the pikromycin gene cluster. Unlike the previous cloning process, one HindIII site integration step was skipped through pSBAC modification. pPik001, a pSBAC containing the pikromycin biosynthetic gene cluster, was directly introduced into two heterologous hosts, Streptomyces lividans and Streptomyces coelicolor, resulting in the production of 10-deoxymethynolide, a major pikromycin derivative. When two entire pikromycin biosynthetic gene clusters were tandemly introduced into the S. lividans chromosome, overproduction of 10-deoxymethynolide and the presence of pikromycin, which was previously not detected, were both confirmed. Moreover, comparative qRT-PCR results confirmed that the transcription of pikromycin biosynthetic genes was significantly upregulated in S. lividans containing tandem clusters of pikromycin biosynthetic gene clusters. The 60 kb pikromycin biosynthetic gene cluster was isolated in a single integration pSBAC vector. Introduction of the pikromycin biosynthetic gene cluster into the pikromycin non-producing strains resulted in higher pikromycin production. The utility of the pSBAC system as a precise cloning tool for large-sized biosynthetic gene clusters was verified through heterologous expression of the pikromycin biosynthetic gene cluster. Moreover, this pSBAC-driven heterologous expression strategy was confirmed to be an ideal approach for production of low and inconsistent natural products such as pikromycin in S. venezuelae, implying that this strategy could be employed for development of a custom overexpression scheme of natural product biosynthetic gene clusters in actinomycetes.

Polarization ellipse and Stokes parameters in geometric algebra.

PubMed

Santos, Adler G; Sugon, Quirino M; McNamara, Daniel J

2012-01-01

In this paper, we use geometric algebra to describe the polarization ellipse and Stokes parameters. We show that a solution to Maxwell's equation is a product of a complex basis vector in Jackson and a linear combination of plane wave functions. We convert both the amplitudes and the wave function arguments from complex scalars to complex vectors. This conversion allows us to separate the electric field vector and the imaginary magnetic field vector, because exponentials of imaginary scalars convert vectors to imaginary vectors and vice versa, while exponentials of imaginary vectors only rotate the vector or imaginary vector they are multiplied to. We convert this expression for polarized light into two other representations: the Cartesian representation and the rotated ellipse representation. We compute the conversion relations among the representation parameters and their corresponding Stokes parameters. And finally, we propose a set of geometric relations between the electric and magnetic fields that satisfy an equation similar to the Poincaré sphere equation.

Support vector machine in machine condition monitoring and fault diagnosis

NASA Astrophysics Data System (ADS)

Widodo, Achmad; Yang, Bo-Suk

2007-08-01

Recently, the issue of machine condition monitoring and fault diagnosis as a part of maintenance system became global due to the potential advantages to be gained from reduced maintenance costs, improved productivity and increased machine availability. This paper presents a survey of machine condition monitoring and fault diagnosis using support vector machine (SVM). It attempts to summarize and review the recent research and developments of SVM in machine condition monitoring and diagnosis. Numerous methods have been developed based on intelligent systems such as artificial neural network, fuzzy expert system, condition-based reasoning, random forest, etc. However, the use of SVM for machine condition monitoring and fault diagnosis is still rare. SVM has excellent performance in generalization so it can produce high accuracy in classification for machine condition monitoring and diagnosis. Until 2006, the use of SVM in machine condition monitoring and fault diagnosis is tending to develop towards expertise orientation and problem-oriented domain. Finally, the ability to continually change and obtain a novel idea for machine condition monitoring and fault diagnosis using SVM will be future works.

Reversible vector ratchets for skyrmion systems

NASA Astrophysics Data System (ADS)

Ma, X.; Reichhardt, C. J. Olson; Reichhardt, C.

2017-03-01

We show that ac driven skyrmions interacting with an asymmetric substrate provide a realization of a class of ratchet system which we call a vector ratchet that arises due to the effect of the Magnus term on the skyrmion dynamics. In a vector ratchet, the dc motion induced by the ac drive can be described as a vector that can be rotated clockwise or counterclockwise relative to the substrate asymmetry direction. Up to a full 360∘ rotation is possible for varied ac amplitudes or skyrmion densities. In contrast to overdamped systems, in which ratchet motion is always parallel to the substrate asymmetry direction, vector ratchets allow the ratchet motion to be in any direction relative to the substrate asymmetry. It is also possible to obtain a reversal in the direction of rotation of the vector ratchet, permitting the creation of a reversible vector ratchet. We examine vector ratchets for ac drives applied parallel or perpendicular to the substrate asymmetry direction, and show that reverse ratchet motion can be produced by collective effects. No reversals occur for an isolated skyrmion on an asymmetric substrate. Since a vector ratchet can produce motion in any direction, it could represent a method for controlling skyrmion motion for spintronic applications.

Multiaxis Thrust-Vectoring Characteristics of a Model Representative of the F-18 High-Alpha Research Vehicle at Angles of Attack From 0 deg to 70 deg

NASA Technical Reports Server (NTRS)

Asbury, Scott C.; Capone, Francis J.

1995-01-01

An investigation was conducted in the Langley 16-Foot Transonic Tunnel to determine the multiaxis thrust-vectoring characteristics of the F-18 High-Alpha Research Vehicle (HARV). A wingtip supported, partially metric, 0.10-scale jet-effects model of an F-18 prototype aircraft was modified with hardware to simulate the thrust-vectoring control system of the HARV. Testing was conducted at free-stream Mach numbers ranging from 0.30 to 0.70, at angles of attack from O' to 70', and at nozzle pressure ratios from 1.0 to approximately 5.0. Results indicate that the thrust-vectoring control system of the HARV can successfully generate multiaxis thrust-vectoring forces and moments. During vectoring, resultant thrust vector angles were always less than the corresponding geometric vane deflection angle and were accompanied by large thrust losses. Significant external flow effects that were dependent on Mach number and angle of attack were noted during vectoring operation. Comparisons of the aerodynamic and propulsive control capabilities of the HARV configuration indicate that substantial gains in controllability are provided by the multiaxis thrust-vectoring control system.

Computing energy levels of CH4, CHD3, CH3D, and CH3F with a direct product basis and coordinates based on the methyl subsystem.

PubMed

Zhao, Zhiqiang; Chen, Jun; Zhang, Zhaojun; Zhang, Dong H; Wang, Xiao-Gang; Carrington, Tucker; Gatti, Fabien

2018-02-21

Quantum mechanical calculations of ro-vibrational energies of CH 4 , CHD 3 , CH 3 D, and CH 3 F were made with two different numerical approaches. Both use polyspherical coordinates. The computed energy levels agree, confirming the accuracy of the methods. In the first approach, for all the molecules, the coordinates are defined using three Radau vectors for the CH 3 subsystem and a Jacobi vector between the remaining atom and the centre of mass of CH 3 . Euler angles specifying the orientation of a frame attached to CH 3 with respect to a frame attached to the Jacobi vector are used as vibrational coordinates. A direct product potential-optimized discrete variable vibrational basis is used to build a Hamiltonian matrix. Ro-vibrational energies are computed using a re-started Arnoldi eigensolver. In the second approach, the coordinates are the spherical coordinates associated with four Radau vectors or three Radau vectors and a Jacobi vector, and the frame is an Eckart frame. Vibrational basis functions are products of contracted stretch and bend functions, and eigenvalues are computed with the Lanczos algorithm. For CH 4 , CHD 3 , and CH 3 D, we report the first J > 0 energy levels computed on the Wang-Carrington potential energy surface [X.-G. Wang and T. Carrington, J. Chem. Phys. 141(15), 154106 (2014)]. For CH 3 F, the potential energy surface of Zhao et al. [J. Chem. Phys. 144, 204302 (2016)] was used. All the results are in good agreement with experimental data.

Improved dual AAV vectors with reduced expression of truncated proteins are safe and effective in the retina of a mouse model of Stargardt disease

PubMed Central

Trapani, Ivana; Toriello, Elisabetta; de Simone, Sonia; Colella, Pasqualina; Iodice, Carolina; Polishchuk, Elena V.; Sommella, Andrea; Colecchi, Linda; Rossi, Settimio; Simonelli, Francesca; Giunti, Massimo; Bacci, Maria L.; Polishchuk, Roman S.; Auricchio, Alberto

2015-01-01

Stargardt disease (STGD1) due to mutations in the large ABCA4 gene is the most common inherited macular degeneration in humans. We have shown that dual adeno-associated viral (AAV) vectors effectively transfer ABCA4 to the retina of Abca4−/− mice. However, they express both lower levels of transgene compared with a single AAV and truncated proteins. To increase productive dual AAV concatemerization, which would overcome these limitations, we have explored the use of either various regions of homology or heterologous inverted terminal repeats (ITR). In addition, we tested the ability of various degradation signals to decrease the expression of truncated proteins. We found the highest levels of transgene expression using regions of homology based on either alkaline phosphatase or the F1 phage (AK). The use of heterologous ITR does not decrease the levels of truncated proteins relative to full-length ABCA4 and impairs AAV vector production. Conversely, the inclusion of the CL1 degradation signal results in the selective degradation of truncated proteins from the 5′-half without affecting full-length protein production. Therefore, we developed dual AAV hybrid ABCA4 vectors including homologous ITR2, the photoreceptor-specific G protein-coupled receptor kinase 1 promoter, the AK region of homology and the CL1 degradation signal. We show that upon subretinal administration these vectors are both safe in pigs and effective in Abca4−/− mice. Our data support the use of improved dual AAV vectors for gene therapy of STGD1. PMID:26420842

Computing energy levels of CH4, CHD3, CH3D, and CH3F with a direct product basis and coordinates based on the methyl subsystem

NASA Astrophysics Data System (ADS)

Zhao, Zhiqiang; Chen, Jun; Zhang, Zhaojun; Zhang, Dong H.; Wang, Xiao-Gang; Carrington, Tucker; Gatti, Fabien

2018-02-01

Quantum mechanical calculations of ro-vibrational energies of CH4, CHD3, CH3D, and CH3F were made with two different numerical approaches. Both use polyspherical coordinates. The computed energy levels agree, confirming the accuracy of the methods. In the first approach, for all the molecules, the coordinates are defined using three Radau vectors for the CH3 subsystem and a Jacobi vector between the remaining atom and the centre of mass of CH3. Euler angles specifying the orientation of a frame attached to CH3 with respect to a frame attached to the Jacobi vector are used as vibrational coordinates. A direct product potential-optimized discrete variable vibrational basis is used to build a Hamiltonian matrix. Ro-vibrational energies are computed using a re-started Arnoldi eigensolver. In the second approach, the coordinates are the spherical coordinates associated with four Radau vectors or three Radau vectors and a Jacobi vector, and the frame is an Eckart frame. Vibrational basis functions are products of contracted stretch and bend functions, and eigenvalues are computed with the Lanczos algorithm. For CH4, CHD3, and CH3D, we report the first J > 0 energy levels computed on the Wang-Carrington potential energy surface [X.-G. Wang and T. Carrington, J. Chem. Phys. 141(15), 154106 (2014)]. For CH3F, the potential energy surface of Zhao et al. [J. Chem. Phys. 144, 204302 (2016)] was used. All the results are in good agreement with experimental data.

A Phase Field Model of Deformation Twinning: Nonlinear Theory and Numerical Simulations

DTIC Science & Technology

2011-03-01

the system. Concepts for mod- eling multiphase systems were advanced by Steinbach et al. [3] and Steinbach and Apel [4]. Fried and Gurtin [5] and...a3b3 in a three- dimensional vector space. The outer product is (a ⊗ b) AB = aAbB. Juxtaposition implies summation over one set of adjacent indices...e.g., ( AB ) AB = AACBCB. The colon denotes summation over two sets of indices; e.g., A : B = AABBAB and (C : E) AB = CABCDECD. The transpose of amatrix is

Basic linear algebra subprograms for FORTRAN usage

NASA Technical Reports Server (NTRS)

Lawson, C. L.; Hanson, R. J.; Kincaid, D. R.; Krogh, F. T.

1977-01-01

A package of 38 low level subprograms for many of the basic operations of numerical linear algebra is presented. The package is intended to be used with FORTRAN. The operations in the package are dot products, elementary vector operations, Givens transformations, vector copy and swap, vector norms, vector scaling, and the indices of components of largest magnitude. The subprograms and a test driver are available in portable FORTRAN. Versions of the subprograms are also provided in assembly language for the IBM 360/67, the CDC 6600 and CDC 7600, and the Univac 1108.

New perspectives in tracing vector-borne interaction networks.

PubMed

Gómez-Díaz, Elena; Figuerola, Jordi

2010-10-01

Disentangling trophic interaction networks in vector-borne systems has important implications in epidemiological and evolutionary studies. Molecular methods based on bloodmeal typing in vectors have been increasingly used to identify hosts. Although most molecular approaches benefit from good specificity and sensitivity, their temporal resolution is limited by the often rapid digestion of blood, and mixed bloodmeals still remain a challenge for bloodmeal identification in multi-host vector systems. Stable isotope analyses represent a novel complementary tool that can overcome some of these problems. The utility of these methods using examples from different vector-borne systems are discussed and the extents to which they are complementary and versatile are highlighted. There are excellent opportunities for progress in the study of vector-borne transmission networks resulting from the integration of both molecular and stable isotope approaches. Copyright © 2010 Elsevier Ltd. All rights reserved.

Replicating Single-Cycle Adenovirus Vectors Generate Amplified Influenza Vaccine Responses.

PubMed

Crosby, Catherine M; Matchett, William E; Anguiano-Zarate, Stephanie S; Parks, Christopher A; Weaver, Eric A; Pease, Larry R; Webby, Richard J; Barry, Michael A

2017-01-15

Head-to-head comparisons of conventional influenza vaccines with adenovirus (Ad) gene-based vaccines demonstrated that these viral vectors can mediate more potent protection against influenza virus infection in animal models. In most cases, Ad vaccines are engineered to be replication-defective (RD-Ad) vectors. In contrast, replication-competent Ad (RC-Ad) vaccines are markedly more potent but risk causing adenovirus diseases in vaccine recipients and health care workers. To harness antigen gene replication but avoid production of infectious virions, we developed "single-cycle" adenovirus (SC-Ad) vectors. Previous work demonstrated that SC-Ads amplify transgene expression 100-fold and produce markedly stronger and more persistent immune responses than RD-Ad vectors in Syrian hamsters and rhesus macaques. To test them as potential vaccines, we engineered RD and SC versions of adenovirus serotype 6 (Ad6) to express the hemagglutinin (HA) gene from influenza A/PR/8/34 virus. We show here that it takes approximately 33 times less SC-Ad6 than RD-Ad6 to produce equal amounts of HA antigen in vitro SC-Ad produced markedly higher HA binding and hemagglutination inhibition (HAI) titers than RD-Ad in Syrian hamsters. SC-Ad-vaccinated cotton rats had markedly lower influenza titers than RD-Ad-vaccinated animals after challenge with influenza A/PR/8/34 virus. These data suggest that SC-Ads may be more potent vaccine platforms than conventional RD-Ad vectors and may have utility as "needle-free" mucosal vaccines. Most adenovirus vaccines that are being tested are replication-defective adenoviruses (RD-Ads). This work describes testing newer single-cycle adenovirus (SC-Ad) vectors that replicate transgenes to amplify protein production and immune responses. We show that SC-Ads generate markedly more influenza virus hemagglutinin protein and require substantially less vector to generate the same immune responses as RD-Ad vectors. SC-Ads therefore hold promise to be more potent vectors and vaccines than current RD-Ad vectors. Copyright © 2017 Crosby et al.

Development of oral CTL vaccine using a CTP-integrated Sabin 1 poliovirus-based vector system.

PubMed

Han, Seung-Soo; Lee, Jinjoo; Jung, Yideul; Kang, Myeong-Ho; Hong, Jung-Hyub; Cha, Min-Suk; Park, Yu-Jin; Lee, Ezra; Yoon, Cheol-Hee; Bae, Yong-Soo

2015-09-11

We developed a CTL vaccine vector by modification of the RPS-Vax system, a mucosal vaccine vector derived from a poliovirus Sabin 1 strain, and generated an oral CTL vaccine against HIV-1. A DNA fragment encoding a cytoplasmic transduction peptide (CTP) was integrated into the RPS-Vax system to generate RPS-CTP, a CTL vaccine vector. An HIV-1 p24 cDNA fragment was introduced into the RPS-CTP vector system and a recombinant poliovirus (rec-PV) named vRPS-CTP/p24 was produced. vRPS-CTP/p24 was genetically stable and efficiently induced Th1 immunity and p24-specific CTLs in immunized poliovirus receptor-transgenic (PVR-Tg) mice. In challenge experiments, PVR-Tg mice that were pre-immunized orally with vRPS-CTP/p24 were resistant to challenge with a lethal dose of p24-expressing recombinant vaccinia virus (rMVA-p24). These results suggested that the RPS-CTP vector system had potential for developing oral CTL vaccines against infectious diseases. Copyright © 2015 Elsevier Ltd. All rights reserved.

Gene inactivation in the plant pathogen Glomerella cingulata: three strategies for the disruption of the pectin lyase gene pnlA.

PubMed

Bowen, J K; Templeton, M D; Sharrock, K R; Crowhurst, R N; Rikkerink, E H

1995-01-20

The feasibility of performing routine transformation-mediated mutagenesis in Glomerella cingulata was analysed by adopting three one-step gene disruption strategies targeted at the pectin lyase gene pnlA. The efficiencies of disruption following transformation with gene replacement- or gene truncation-disruption vectors were compared. To effect replacement-disruption, G. cingulata was transformed with a vector carrying DNA from the pnlA locus in which the majority of the coding sequence had been replaced by the gene for hygromycin B resistance. Two of the five transformants investigated contained an inactivated pnlA gene (pnlA-); both also contained ectopically integrated vector sequences. The efficacy of gene disruption by transformation with two gene truncation-disruption vectors was also assessed. Both vectors carried at 5' and 3' truncated copy of the pnlA coding sequence, adjacent to the gene for hygromycin B resistance. The promoter sequences controlling the selectable marker differed in the two vectors. In one vector the homologous G. cingulata gpdA promoter controlled hygromycin B phosphotransferase expression (homologous truncation vector), whereas in the second vector promoter elements were from the Aspergillus nidulans gpdA gene (heterologous truncation vector). Following transformation with the homologous truncation vector, nine transformants were analysed by Southern hybridisation; no transformants contained a disrupted pnlA gene. Of nineteen heterologous truncation vector transformants, three contained a disrupted pnlA gene; Southern analysis revealed single integrations of vector sequence at pnlA in two of these transformants. pnlA mRNA was not detected by Northern hybridisation in pnlA- transformants. pnlA- transformants failed to produce a PNLA protein with a pI identical to one normally detected in wild-type isolates by silver and activity staining of isoelectric focussing gels. Pathogenesis on Capsicum and apple was unaffected by disruption of the pnlA gene, indicating that the corresponding gene product, PNLA, is not essential for pathogenicity. Gene disruption is a feasible method for selectively mutating defined loci in G. cingulata for functional analysis of the corresponding gene products.

Constraints on models of scalar and vector leptoquarks decaying to a quark and a neutrino at $$\\sqrt{s}=$$ 13 TeV

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sirunyan, Albert M; et al.

The results of a previous search by the CMS Collaboration for squarks and gluinos are reinterpreted to constrain models of leptoquark (LQ) production. The search considers jets in association with a transverse momentum imbalance, using themore » $$M_\\mathrm{T2}$$ variable. The analysis uses proton-proton collision data at $$\\sqrt{s}=$$ 13 TeV, recorded with the CMS detector at the LHC in 2016 and corresponding to an integrated luminosity of 35.9 fb$$^{-1}$$. Leptoquark pair production is considered with LQ decays to a neutrino and a top, bottom, or light quark. This reinterpretation considers higher mass values than the original CMS search to constrain both scalar and vector LQs. Limits on the cross section for LQ pair production are derived at the 95% confidence level depending on the LQ decay mode. A vector LQ decaying with a 50% branching fraction to t$$\

Search for single production of vector-like top partner decaying to Wb at eγ collision

NASA Astrophysics Data System (ADS)

Yang, Bingfang; Shao, Hongbo; Han, Jinzhong

2018-03-01

In a simplified model including an SU(2) singlet T quark with charge 2/3, we investigate the single vector-like T production at the high energy eγ collision. We study the observability of the vector-like T focusing on the T→ Wb decay channel with W→ l\\bar{ν } at √{s}=2.0 TeV. In this analysis, only two free parameters are involved, namely the T quark coupling strength for single production g^{*} and the mass mT. We scan the parameter space and find that the correlation region of g^{*}\\in [0.24, 0.5] and mT\\in [800, 1360] GeV can be excluded with integrated luminosity L=100 fb^{-1} and the correlation region of g^{*}\\in [0.13, 0.5] and mT\\in [800, 1620] GeV can be excluded with integrated luminosity L=1000 fb^{-1} at 2σ level.

A pepper mottle virus-based vector enables systemic expression of endoglucanase D in non-transgenic plants.

PubMed

Song, Eun Gyeong; Ryu, Ki Hyun

2017-12-01

Plant-virus-based expression vectors have been used as an alternative to the creation of transgenic plants. Using a virus-based vector, we investigated the feasibility of producing the endoglucanase D (EngD) from Clostridium cellulovorans in Nicotiana benthamiana. This protein has endoglucanase, xylanase, and exoglucanase activities and may be of value for cellulose digestion in the generation of biofuels from plant biomass. The EngD gene was cloned between the nuclear inclusion b (NIb)- and coat protein (CP)-encoding sequences of pSP6PepMoV-Vb1. In vitro transcripts derived from the clone (pSP6PepMoV-Vb1/EngD) were infectious in N. benthamiana but caused milder symptoms than wild-type PepMoV-Vb1. RT-PCR amplification of total RNA from non-inoculated upper leaves infected with PepMoV-Vb1/EngD produced the target band for the CP, partial NIb and EngD-CP regions of PepMoV-V1/EngD, in addition to nonspecific bands. Western blot analysis showed the CP target bands of PepMoV-Vb1/EngD as well as non-target bands. EngD enzymatic activity in infected plants was detected using a glucose assay. The plant leaves showed increased senescence compared with healthy and PepMoV-Vb1-infected plants. Our study suggests the feasibility of using a viral vector for systemic infection of plants for expression of heterologous engD for the purpose of digesting a cellulose substrate in plant cells for biomass production.

Methods, systems and apparatus for controlling third harmonic voltage when operating a multi-space machine in an overmodulation region

DOEpatents

Perisic, Milun; Kinoshita, Michael H; Ranson, Ray M; Gallegos-Lopez, Gabriel

2014-06-03

Methods, system and apparatus are provided for controlling third harmonic voltages when operating a multi-phase machine in an overmodulation region. The multi-phase machine can be, for example, a five-phase machine in a vector controlled motor drive system that includes a five-phase PWM controlled inverter module that drives the five-phase machine. Techniques for overmodulating a reference voltage vector are provided. For example, when the reference voltage vector is determined to be within the overmodulation region, an angle of the reference voltage vector can be modified to generate a reference voltage overmodulation control angle, and a magnitude of the reference voltage vector can be modified, based on the reference voltage overmodulation control angle, to generate a modified magnitude of the reference voltage vector. By modifying the reference voltage vector, voltage command signals that control a five-phase inverter module can be optimized to increase output voltages generated by the five-phase inverter module.

A Novel Dual Expression Platform for High Throughput Functional Screening of Phage Libraries in Product like Format.

PubMed

Xiao, Xiaodong; Chen, Yan; Mugabe, Sheila; Gao, Changshou; Tkaczyk, Christine; Mazor, Yariv; Pavlik, Peter; Wu, Herren; Dall'Acqua, William; Chowdhury, Partha Sarathi

2015-01-01

High throughput screenings of single chain Fv (scFv) antibody phage display libraries are currently done as soluble scFvs produced in E.coli. Due to endotoxin contaminations from bacterial cells these preparations cannot be reliably used in mammalian cell based assays. The monovalent nature and lack of Fc in soluble scFvs prevent functional assays that are dependent on target cross linking and/or Fc functions. A convenient approach is to convert scFvs into scFv.Fc fusion proteins and express them in mammalian cell lines for screening. This approach is low throughput and is only taken after primary screening of monovalent scFvs that are expressed in bacteria. There is no platform at present that combines the benefits of both bacterial and mammalian expression system for screening phage library output. We have, therefore, developed a novel dual expression vector, called pSplice, which can be used to express scFv.Fc fusion proteins both in E.coli and mammalian cell lines. The hallmark of the vector is an engineered intron which houses the bacterial promoter and signal peptide for expression and secretion of scFv.Fc in E.coli. When the vector is transfected into a mammalian cell line, the intron is efficiently spliced out resulting in a functional operon for expression and secretion of the scFv.Fc fusion protein into the culture medium. By applying basic knowledge of mammalian introns and splisosome, we designed this vector to enable screening of phage libraries in a product like format. Like IgG, the scFv.Fc fusion protein is bi-valent for the antigen and possesses Fc effector functions. Expression in E.coli maintains the speed of the bacterial expression platform and is used to triage clones based on binding and other assays that are not sensitive to endotoxin. Triaged clones are then expressed in a mammalian cell line without the need for any additional cloning steps. Conditioned media from the mammalian cell line containing the fusion proteins are then used for different types of cell based assays. Thus this system retains the speed of the current screening system for phage libraries and adds additional functionality to it.

Functional characterization of adenoviral/retroviral chimeric vectors and their use for efficient screening of retroviral producer cell lines.

PubMed

Duisit, G; Salvetti, A; Moullier, P; Cosset, F L

1999-01-20

We have generated three different E1-deleted replication-defective adenoviral vectors expressing either Moloney murine leukemia virus (Mo-MuLV) Gag-Pol core particle proteins, gibbon ape leukemia virus (GALV) envelope glycoproteins, or an MuLV-derived retroviral vector genome encoding mCD2 antigen, a murine cell surface marker easily detectable by flow cytometry. Each of the three vectors was first characterized individually by infection of cells providing the complementary retroviral function(s) and able to induce the production of retroviral vectors with an efficiency similar to or higher than that of FLY stable retroviral packaging cells [Cosset, F.-L., Takeuchi, Y., Battini, J.-L., Weiss, R.A., and Collins, M.K.L., (1995). J. Virol. 69, 7430-7436]. In small-scale pilot experiments, TE671 cells simultaneously coinfected with the three adenoviral vectors efficiently released helper-free retroviral vectors in their supernatant, with titers greater than 10(6) infectious particles per milliliter by end-point titrations. Our results also indicated that in contrast to retroviral vector-packageable RNAs, the adenovirus-mediated overexpression of both Gag-Pol and Env packaging functions had limited impact on retroviral titers. The primary mechanism suspected is the premature intracellular cleavage of the Pr65gag precursor that we found in gag-pol-expressing cells, which in turn may impair the normal incorporation of high loads of functional Env. Last, the characterization of the adenoviral/retroviral chimeric vectors allowed the screening of various primate cells for retroviral production and we found that three hepatocyte-derived cell lines were highly efficient in the assembly and release of infectious retroviral particles.

Compute Server Performance Results

NASA Technical Reports Server (NTRS)

Stockdale, I. E.; Barton, John; Woodrow, Thomas (Technical Monitor)

1994-01-01

Parallel-vector supercomputers have been the workhorses of high performance computing. As expectations of future computing needs have risen faster than projected vector supercomputer performance, much work has been done investigating the feasibility of using Massively Parallel Processor systems as supercomputers. An even more recent development is the availability of high performance workstations which have the potential, when clustered together, to replace parallel-vector systems. We present a systematic comparison of floating point performance and price-performance for various compute server systems. A suite of highly vectorized programs was run on systems including traditional vector systems such as the Cray C90, and RISC workstations such as the IBM RS/6000 590 and the SGI R8000. The C90 system delivers 460 million floating point operations per second (FLOPS), the highest single processor rate of any vendor. However, if the price-performance ration (PPR) is considered to be most important, then the IBM and SGI processors are superior to the C90 processors. Even without code tuning, the IBM and SGI PPR's of 260 and 220 FLOPS per dollar exceed the C90 PPR of 160 FLOPS per dollar when running our highly vectorized suite,

Progress and challenges in viral vector manufacturing

PubMed Central

van der Loo, Johannes C.M.; Wright, J. Fraser

2016-01-01

Promising results in several clinical studies have emphasized the potential of gene therapy to address important medical needs and initiated a surge of investments in drug development and commercialization. This enthusiasm is driven by positive data in clinical trials including gene replacement for Hemophilia B, X-linked Severe Combined Immunodeficiency, Leber's Congenital Amaurosis Type 2 and in cancer immunotherapy trials for hematological malignancies using chimeric antigen receptor T cells. These results build on the recent licensure of the European gene therapy product Glybera for the treatment of lipoprotein lipase deficiency. The progress from clinical development towards product licensure of several programs presents challenges to gene therapy product manufacturing. These include challenges in viral vector-manufacturing capacity, where an estimated 1–2 orders of magnitude increase will likely be needed to support eventual commercial supply requirements for many of the promising disease indications. In addition, the expanding potential commercial product pipeline and the continuously advancing development of recombinant viral vectors for gene therapy require that products are well characterized and consistently manufactured to rigorous tolerances of purity, potency and safety. Finally, there is an increase in regulatory scrutiny that affects manufacturers of investigational drugs for early-phase clinical trials engaged in industry partnerships. Along with the recent increase in biopharmaceutical funding in gene therapy, industry partners are requiring their academic counterparts to meet higher levels of GMP compliance at earlier stages of clinical development. This chapter provides a brief overview of current progress in the field and discusses challenges in vector manufacturing. PMID:26519140

Efficient production of D-tagatose using a food-grade surface display system.

PubMed

Liu, Yi; Li, Sha; Xu, Hong; Wu, Lingtian; Xu, Zheng; Liu, Jing; Feng, Xiaohai

2014-07-16

D-tagatose, a functional sweetener, is commonly transformed from D-galactose by L-arabinose isomerase (L-AI). In this study, a novel type of biocatalyst, L-AI from Lactobacillus fermentum CGMCC2921 displayed on the spore surface of Bacillus subtilis 168, was developed for producing D-tagatose. The anchored L-AI, exhibiting the relatively high bioactivity, suggested that the surface display system using CotX as the anchoring protein was successfully constructed. The stability of the anchored L-AI was significantly improved. Specifically, the consolidation of thermal stability representing 87% of relative activity was retained even at 80 °C for 30 min, which remarkably favored the production of D-tagatose. Under the optimal conditions, the robust spores can convert 75% D-galactose (100 g/L) into D-tagatose after 24 h, and the conversion rate remained at 56% at the third cycle. Therefore, this biocatalysis system, which could express the target enzyme on the food-grade vector, was an alternative method for the value-added production of D-tagatose.

Design of an ion thruster movable grid thrust vectoring system

NASA Astrophysics Data System (ADS)

Kural, Aleksander; Leveque, Nicolas; Welch, Chris; Wolanski, Piotr

2004-08-01

Several reasons justify the development of an ion propulsion system thrust vectoring system. Spacecraft launched to date have used ion thrusters mounted on gimbals to control the thrust vector within a range of about ±5°. Such devices have large mass and dimensions, hence the need exists for a more compact system, preferably mounted within the thruster itself. Since the 1970s several thrust vectoring systems have been developed, with the translatable accelerator grid electrode being considered the most promising. Laboratory models of this system have already been built and successfully tested, but there is still room for improvement in their mechanical design. This work aims to investigate possibilities of refining the design of such movable grid thrust vectoring systems. Two grid suspension designs and three types of actuators were evaluated. The actuators examined were a micro electromechanical system, a NanoMuscle shape memory alloy actuator and a piezoelectric driver. Criteria used for choosing the best system included mechanical simplicity (use of the fewest mechanical parts), accuracy, power consumption and behaviour in space conditions. Designs of systems using these actuators are proposed. In addition, a mission to Mercury using the system with piezoelectric drivers has been modelled and its performance presented.

A new preconditioner update strategy for the solution of sequences of linear systems in structural mechanics: application to saddle point problems in elasticity

NASA Astrophysics Data System (ADS)

Mercier, Sylvain; Gratton, Serge; Tardieu, Nicolas; Vasseur, Xavier

2017-12-01

Many applications in structural mechanics require the numerical solution of sequences of linear systems typically issued from a finite element discretization of the governing equations on fine meshes. The method of Lagrange multipliers is often used to take into account mechanical constraints. The resulting matrices then exhibit a saddle point structure and the iterative solution of such preconditioned linear systems is considered as challenging. A popular strategy is then to combine preconditioning and deflation to yield an efficient method. We propose an alternative that is applicable to the general case and not only to matrices with a saddle point structure. In this approach, we consider to update an existing algebraic or application-based preconditioner, using specific available information exploiting the knowledge of an approximate invariant subspace or of matrix-vector products. The resulting preconditioner has the form of a limited memory quasi-Newton matrix and requires a small number of linearly independent vectors. Numerical experiments performed on three large-scale applications in elasticity highlight the relevance of the new approach. We show that the proposed method outperforms the deflation method when considering sequences of linear systems with varying matrices.

Using remote sensing, ecological niche modeling, and Geographic Information Systems for Rift Valley fever risk assessment in the United States

NASA Astrophysics Data System (ADS)

Tedrow, Christine Atkins

The primary goal in this study was to explore remote sensing, ecological niche modeling, and Geographic Information Systems (GIS) as aids in predicting candidate Rift Valley fever (RVF) competent vector abundance and distribution in Virginia, and as means of estimating where risk of establishment in mosquitoes and risk of transmission to human populations would be greatest in Virginia. A second goal in this study was to determine whether the remotely-sensed Normalized Difference Vegetation Index (NDVI) can be used as a proxy variable of local conditions for the development of mosquitoes to predict mosquito species distribution and abundance in Virginia. As part of this study, a mosquito surveillance database was compiled to archive the historical patterns of mosquito species abundance in Virginia. In addition, linkages between mosquito density and local environmental and climatic patterns were spatially and temporally examined. The present study affirms the potential role of remote sensing imagery for species distribution prediction, and it demonstrates that ecological niche modeling is a valuable predictive tool to analyze the distributions of populations. The MaxEnt ecological niche modeling program was used to model predicted ranges for potential RVF competent vectors in Virginia. The MaxEnt model was shown to be robust, and the candidate RVF competent vector predicted distribution map is presented. The Normalized Difference Vegetation Index (NDVI) was found to be the most useful environmental-climatic variable to predict mosquito species distribution and abundance in Virginia. However, these results indicate that a more robust prediction is obtained by including other environmental-climatic factors correlated to mosquito densities (e.g., temperature, precipitation, elevation) with NDVI. The present study demonstrates that remote sensing and GIS can be used with ecological niche and risk modeling methods to estimate risk of virus establishment in mosquitoes and transmission to humans. Maps delineating the geographic areas in Virginia with highest risk for RVF establishment in mosquito populations and RVF disease transmission to human populations were generated in a GIS using human, domestic animal, and white-tailed deer population estimates and the MaxEnt potential RVF competent vector species distribution prediction. The candidate RVF competent vector predicted distribution and RVF risk maps presented in this study can help vector control agencies and public health officials focus Rift Valley fever surveillance efforts in geographic areas with large co-located populations of potential RVF competent vectors and human, domestic animal, and wildlife hosts. Keywords. Rift Valley fever, risk assessment, Ecological Niche Modeling, MaxEnt, Geographic Information System, remote sensing, Pearson's Product-Moment Correlation Coefficient, vectors, mosquito distribution, mosquito density, mosquito surveillance, United States, Virginia, domestic animals, white-tailed deer, ArcGIS

Recent Weather Extremes and Impacts on Agricultural Production and Vector-Borne Disease Outbreak Patterns

NASA Technical Reports Server (NTRS)

Anyamba, Assaf; Small, Jennifer L.; Britch, Seth C.; Tucker, Compton J.; Pak, Edwin W.; Reynolds, Curt A.; Crutchfield, James; Linthicum, Kenneth J.

2014-01-01

We document significant worldwide weather anomalies that affected agriculture and vector-borne disease outbreaks during the 2010-2012 period. We utilized 2000-2012 vegetation index and land surface temperature data from NASA's satellite-based Moderate Resolution Imaging Spectroradiometer (MODIS) to map the magnitude and extent of these anomalies for diverse regions including the continental United States, Russia, East Africa, Southern Africa, and Australia. We demonstrate that shifts in temperature and/or precipitation have significant impacts on vegetation patterns with attendant consequences for agriculture and public health. Weather extremes resulted in excessive rainfall and flooding as well as severe drought, which caused,10 to 80% variation in major agricultural commodity production (including wheat, corn, cotton, sorghum) and created exceptional conditions for extensive mosquito-borne disease outbreaks of dengue, Rift Valley fever, Murray Valley encephalitis, and West Nile virus disease. Analysis of MODIS data provided a standardized method for quantifying the extreme weather anomalies observed during this period. Assessments of land surface conditions from satellite-based systems such as MODIS can be a valuable tool in national, regional, and global weather impact determinations.

Recent weather extremes and impacts on agricultural production and vector-borne disease outbreak patterns.

PubMed

Anyamba, Assaf; Small, Jennifer L; Britch, Seth C; Tucker, Compton J; Pak, Edwin W; Reynolds, Curt A; Crutchfield, James; Linthicum, Kenneth J

2014-01-01

We document significant worldwide weather anomalies that affected agriculture and vector-borne disease outbreaks during the 2010-2012 period. We utilized 2000-2012 vegetation index and land surface temperature data from NASA's satellite-based Moderate Resolution Imaging Spectroradiometer (MODIS) to map the magnitude and extent of these anomalies for diverse regions including the continental United States, Russia, East Africa, Southern Africa, and Australia. We demonstrate that shifts in temperature and/or precipitation have significant impacts on vegetation patterns with attendant consequences for agriculture and public health. Weather extremes resulted in excessive rainfall and flooding as well as severe drought, which caused ∼10 to 80% variation in major agricultural commodity production (including wheat, corn, cotton, sorghum) and created exceptional conditions for extensive mosquito-borne disease outbreaks of dengue, Rift Valley fever, Murray Valley encephalitis, and West Nile virus disease. Analysis of MODIS data provided a standardized method for quantifying the extreme weather anomalies observed during this period. Assessments of land surface conditions from satellite-based systems such as MODIS can be a valuable tool in national, regional, and global weather impact determinations.

Intra-Amniotic rAAV-Mediated Microdystrophin Gene Transfer Improves Canine X-Linked Muscular Dystrophy and May Induce Immune Tolerance

PubMed Central

Hayashita-Kinoh, Hiromi; Yugeta, Naoko; Okada, Hironori; Nitahara-Kasahara, Yuko; Chiyo, Tomoko; Okada, Takashi; Takeda, Shin'ichi

2015-01-01

Duchenne muscular dystrophy (DMD) is a severe congenital disease due to mutations in the dystrophin gene. Supplementation of dystrophin using recombinant adenoassociated virus vector has promise as a treatment of DMD, although therapeutic benefit of the truncated dystrophin still remains to be elucidated. Besides, host immune responses against the vector as well as transgene products have been denoted in the clinical gene therapy studies. Here, we transduced dystrophic dogs fetuses to investigate the therapeutic effects of an AAV vector expressing microdystrophin under conditions of immune tolerance. rAAV-CMV-microdystrophin and a rAAV-CAG-luciferase were injected into the amniotic fluid surrounding fetuses. We also reinjected rAAV9-CMV-microdystrophin into the jugular vein of an infant dystrophic dog to induce systemic expression of microdystrophin. Gait and cardiac function significantly improved in the rAAV-microdystrophin-injected dystrophic dog, suggesting that an adequate treatment of rAAV-microdystrophin with immune modulation induces successful long-term transgene expression to analyze improved dystrophic phenotype. PMID:25586688

MISR CMV New Data

Atmospheric Science Data Center

2016-10-31

Cloud Motion Vector (CMV) Product The MISR Level 3 Products are global or ... field campaigns at daily and monthly time scales. The CMV product provides conveniently organized, high quality retrievals of cloud ...

Production of transgenic Korean native cattle expressing enhanced green fluorescent protein using a FIV-based lentiviral vector injected into MII oocytes.

PubMed

Xu, Yong-Nan; Uhm, Sang-Jun; Koo, Bon-Chul; Kwon, Mo-Sun; Roh, Ji-Yeol; Yang, Jung-Seok; Choi, Hyun-Yong; Heo, Young-Tae; Cui, Xiang-Shun; Yoon, Joon-Ho; Ko, Dae-Hwan; Kim, Teoan; Kim, Nam-Hyung

2013-01-20

The potential benefits of generating and using transgenic cattle range from improvements in agriculture to the production of large quantities of pharmaceutically relevant proteins. Previous studies have attempted to produce transgenic cattle and other livestock by pronuclear injection and somatic cell nuclear transfer, but these approaches have been largely ineffective; however, a third approach, lentivirus-mediated transgenesis, has successfully produced transgenic livestock. In this study, we generated transgenic (TG) Korean native cattle using perivitelline space injection of viral vectors, which expressed enhanced green fluorescent protein (EGFP) systemically. Two different types of lentiviral vectors derived from feline immunodeficiency virus (FIV) and human immunodeficiency virus (HIV) carrying EGFP were injected into the perivitelline space of MII oocytes. EGFP expression at 8-cell stage was significantly higher in the FIV group compared to the HIV group (47.5%±2.2% v.s. 22.9%±2.9%). Eight-cell embryos that expressed EGFP were cultured into blastocysts and then transferred into 40 heifers. Ten heifers were successfully impregnated and delivered 10 healthy calves. All of these calves expressed EGFP as detected by in vivo imaging, PCR and Southern blotting. In addition, we established an EGFP-expressing cell line from TG calves, which was followed by nuclear transfer (NT). Recloned 8-cell embryos also expressed EGFP, and there were no differences in the rates of fusion, cleavage and development between cells derived from TG and non-TG calves, which were subsequently used for NT. These results illustrate that FIV-based lentiviruses are useful for the production of TG cattle. Moreover, our established EGFP cell line can be used for additional studies that involve induced pluripotent stem cells. Copyright © 2013. Published by Elsevier Ltd.

Attitude-Independent Magnetometer Calibration for Spin-Stabilized Spacecraft

NASA Technical Reports Server (NTRS)

Natanson, Gregory

2005-01-01

The paper describes a three-step estimator to calibrate a Three-Axis Magnetometer (TAM) using TAM and slit Sun or star sensor measurements. In the first step, the Calibration Utility forms a loss function from the residuals of the magnitude of the geomagnetic field. This loss function is minimized with respect to biases, scale factors, and nonorthogonality corrections. The second step minimizes residuals of the projection of the geomagnetic field onto the spin axis under the assumption that spacecraft nutation has been suppressed by a nutation damper. Minimization is done with respect to various directions of the body spin axis in the TAM frame. The direction of the spin axis in the inertial coordinate system required for the residual computation is assumed to be unchanged with time. It is either determined independently using other sensors or included in the estimation parameters. In both cases all estimation parameters can be found using simple analytical formulas derived in the paper. The last step is to minimize a third loss function formed by residuals of the dot product between the geomagnetic field and Sun or star vector with respect to the misalignment angle about the body spin axis. The method is illustrated by calibrating TAM for the Fast Auroral Snapshot Explorer (FAST) using in-flight TAM and Sun sensor data. The estimated parameters include magnetic biases, scale factors, and misalignment angles of the spin axis in the TAM frame. Estimation of the misalignment angle about the spin axis was inconclusive since (at least for the selected time interval) the Sun vector was about 15 degrees from the direction of the spin axis; as a result residuals of the dot product between the geomagnetic field and Sun vectors were to a large extent minimized as a by-product of the second step.

Student Understanding of Cross Product Direction and Use of Right-hand Rules: An Exploration of Representation and Context-dependence

NASA Astrophysics Data System (ADS)

Kustusch, Mary Bridget

2011-12-01

Students in introductory physics struggle with vector algebra and with cross product direction in particular. Some have suggested that this may be due to misapplied right-hand rules, but there are few studies that have had the resolution to explore this. Additionally, previous research on student understanding has noted several kinds of representation-dependence of student performance with vector algebra in both physics and non-physics (or math) contexts (e.g. Hawkins et al., 2009; Van Deventer, 2008). Yet with few exceptions (e.g. Scaife and Heckler, 2010), these findings have not been applied to cross product direction questions or the use of right-hand rules. Also, the extensive work in spatial cognition is particularly applicable to cross product direction due to the spatial and kinesthetic nature of the right-hand rule. A synthesis of the literature from these various fields reveals four categories of problem features likely to impact the understanding of cross product direction: (1) the type of reasoning required, (2) the orientation of the vectors, (3) the need for parallel transport, and (4) the physics context and features (or lack thereof). These features formed the basis of the present effort to systematically explore the context-dependence and representation- dependence of student performance on cross product direction questions. This study used a mix of qualitative and quantitative techniques to analyze twenty-seven individual think-aloud interviews. During these interviews, second semester introductory physics students answered 80-100 cross product direction questions in different contexts and with varying problem features. These features were then used as the predictors in regression analyses for correctness and response time. In addition, each problem was coded for the methods used and the errors made to gain a deeper understanding of student behavior and the impact of these features. The results revealed a wide variety of methods (including six different right-hand rules), many different types of errors, and significant context-dependence and representation-dependence for the features mentioned above. Problems that required reasoning backward to find A⃗ (for C⃗=A⃗ xB⃗ ) presented the biggest challenge for students. Participants who recognized the non-commutativity of the cross product would often reverse the order ( B⃗xA⃗ ) on these problems. Also, this error occurred less frequently when a Guess and Check method was used in addition to the right-hand rule. Three different aspects of orientation had a significant impact on performance: (1) the physical discomfort of using a right-hand rule, (2) the plane of the given vectors, and to a lesser extent, (3) the angle between the vectors. One participant was more likely to switch the order of the vectors for the physically awkward orientations than for the physically easy orientations; and there was evidence that some of the difficulty with vector orientations that were not in the xy-plane was due to misinterpretations of the into and out of the page symbols. The impact of both physical discomfort and the plane of the vectors was reduced when participants rotated the paper. Unlike other problem features, the issue of parallel transport did not appear to be nearly as prevalent for cross product direction as it is for vector addition and subtraction. In addition to these findings, this study confirmed earlier findings regarding physics difficulties with magnetic field and magnetic force, such as differences in performance based on the representation of magnetic field (Scaife and Heckler, 2010) and confusion between electric and magnetic fields (Maloney et al., 2001). It also provided evidence of physics difficulties with magnetic field and magnetic force that have been suspected but never explored, specifically the impact of the sign of the charge and the observation location. This study demonstrated that student difficulty with cross product direction is not as simple as misapplied right-hand rules, although this is an issue. Student behavior on cross product direction questions is significantly dependent on both the context of the question and the representation of various problem features. Although more research is necessary, particularly in regard to individual differences, this study represents a significant step forward in our understanding of student difficulties with cross product direction.

Efficient Kriging via Fast Matrix-Vector Products

NASA Technical Reports Server (NTRS)

Memarsadeghi, Nargess; Raykar, Vikas C.; Duraiswami, Ramani; Mount, David M.

2008-01-01

Interpolating scattered data points is a problem of wide ranging interest. Ordinary kriging is an optimal scattered data estimator, widely used in geosciences and remote sensing. A generalized version of this technique, called cokriging, can be used for image fusion of remotely sensed data. However, it is computationally very expensive for large data sets. We demonstrate the time efficiency and accuracy of approximating ordinary kriging through the use of fast matrixvector products combined with iterative methods. We used methods based on the fast Multipole methods and nearest neighbor searching techniques for implementations of the fast matrix-vector products.

Knock-in fibroblasts and transgenic blastocysts for expression of human FGF2 in the bovine β-casein gene locus using CRISPR/Cas9 nuclease-mediated homologous recombination.

PubMed

Jeong, Young-Hee; Kim, Yeong Ji; Kim, Eun Young; Kim, Se Eun; Kim, Jiwoo; Park, Min Jee; Lee, Hong-Gu; Park, Se Pill; Kang, Man-Jong

2016-06-01

Many transgenic domestic animals have been developed to produce therapeutic proteins in the mammary gland, and this approach is one of the most important methods for agricultural and biomedical applications. However, expression and secretion of a protein varies because transgenes are integrated at random sites in the genome. In addition, distal enhancers are very important for transcriptional gene regulation and tissue-specific gene expression. Development of a vector system regulated accurately in the genome is needed to improve production of therapeutic proteins. The objective of this study was to develop a knock-in system for expression of human fibroblast growth factor 2 (FGF2) in the bovine β-casein gene locus. The F2A sequence was fused to the human FGF2 gene and inserted into exon 3 of the β-casein gene. We detected expression of human FGF2 mRNA in the HC11 mouse mammary epithelial cells by RT-PCR and human FGF2 protein in the culture media using western blot analysis when the knock-in vector was introduced. We transfected the knock-in vector into bovine ear fibroblasts and produced knock-in fibroblasts using the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system. Moreover, the CRISPR/Cas9 system was more efficient than conventional methods. In addition, we produced knock-in blastocysts by somatic cell nuclear transfer using the knock-in fibroblasts. Our knock-in fibroblasts may help to create cloned embryos for development of transgenic dairy cattle expressing human FGF2 protein in the mammary gland via the expression system of the bovine β-casein gene.

Cartographic services contract...for everything geographic

USGS Publications Warehouse

,

2003-01-01

The U.S. Geological Survey's (USGS) Cartographic Services Contract (CSC) is used to award work for photogrammetric and mapping services under the umbrella of Architect-Engineer (A&E) contracting. The A&E contract is broad in scope and can accommodate any activity related to standard, nonstandard, graphic, and digital cartographic products. Services provided may include, but are not limited to, photogrammetric mapping and aerotriangulation; orthophotography; thematic mapping (for example, land characterization); analog and digital imagery applications; geographic information systems development; surveying and control acquisition, including ground-based and airborne Global Positioning System; analog and digital image manipulation, analysis, and interpretation; raster and vector map digitizing; data manipulations (for example, transformations, conversions, generalization, integration, and conflation); primary and ancillary data acquisition (for example, aerial photography, satellite imagery, multispectral, multitemporal, and hyperspectral data); image scanning and processing; metadata production, revision, and creation; and production or revision of standard USGS products defined by formal and informal specification and standards, such as those for digital line graphs, digital elevation models, digital orthophoto quadrangles, and digital raster graphics.

Approaches for Language Identification in Mismatched Environments

DTIC Science & Technology

2016-09-08

different i-vector systems are considered, which differ in their feature extraction mechanism. The first, which we refer to as the standard i-vector, or...both conversational telephone speech and narrowband broadcast speech. Multiple experiments are conducted to assess the performance of the system in...bottleneck features using i-vectors. The proposed system results in a 30% improvement over the baseline result. Index Terms: language identification

Reversible vector ratchets for skyrmion systems

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ma, Xiu; Reichhardt, Cynthia Jane Olson; Reichhardt, Charles

In this paper, we show that ac driven skyrmions interacting with an asymmetric substrate provide a realization of a class of ratchet system which we call a vector ratchet that arises due to the effect of the Magnus term on the skyrmion dynamics. In a vector ratchet, the dc motion induced by the ac drive can be described as a vector that can be rotated clockwise or counterclockwise relative to the substrate asymmetry direction. Up to a full 360° rotation is possible for varied ac amplitudes or skyrmion densities. In contrast to overdamped systems, in which ratchet motion is alwaysmore » parallel to the substrate asymmetry direction, vector ratchets allow the ratchet motion to be in any direction relative to the substrate asymmetry. It is also possible to obtain a reversal in the direction of rotation of the vector ratchet, permitting the creation of a reversible vector ratchet. We examine vector ratchets for ac drives applied parallel or perpendicular to the substrate asymmetry direction, and show that reverse ratchet motion can be produced by collective effects. No reversals occur for an isolated skyrmion on an asymmetric substrate. Finally, since a vector ratchet can produce motion in any direction, it could represent a method for controlling skyrmion motion for spintronic applications.« less

Reversible vector ratchets for skyrmion systems

DOE PAGES

Ma, Xiu; Reichhardt, Cynthia Jane Olson; Reichhardt, Charles

2017-03-03

In this paper, we show that ac driven skyrmions interacting with an asymmetric substrate provide a realization of a class of ratchet system which we call a vector ratchet that arises due to the effect of the Magnus term on the skyrmion dynamics. In a vector ratchet, the dc motion induced by the ac drive can be described as a vector that can be rotated clockwise or counterclockwise relative to the substrate asymmetry direction. Up to a full 360° rotation is possible for varied ac amplitudes or skyrmion densities. In contrast to overdamped systems, in which ratchet motion is alwaysmore » parallel to the substrate asymmetry direction, vector ratchets allow the ratchet motion to be in any direction relative to the substrate asymmetry. It is also possible to obtain a reversal in the direction of rotation of the vector ratchet, permitting the creation of a reversible vector ratchet. We examine vector ratchets for ac drives applied parallel or perpendicular to the substrate asymmetry direction, and show that reverse ratchet motion can be produced by collective effects. No reversals occur for an isolated skyrmion on an asymmetric substrate. Finally, since a vector ratchet can produce motion in any direction, it could represent a method for controlling skyrmion motion for spintronic applications.« less

Systematics of strong nuclear amplification of gluon saturation from exclusive vector meson production in high energy electron-nucleus collisions

NASA Astrophysics Data System (ADS)

Mäntysaari, Heikki; Venugopalan, Raju

2018-06-01

We show that gluon saturation gives rise to a strong modification of the scaling in both the nuclear mass number A and the virtuality Q2 of the vector meson production cross-section in exclusive deep-inelastic scattering off nuclei. We present qualitative analytic expressions for how the scaling exponents are modified as well as quantitative predictions that can be tested at an Electron-Ion Collider.

9 CFR 103.2 - Disposition of animals administered experimental biological products or live organisms.

Code of Federal Regulations, 2010 CFR

2010-01-01

... PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS EXPERIMENTAL PRODUCTION, DISTRIBUTION, AND EVALUATION OF BIOLOGICAL...

Bluetongue outbreaks: Looking for effective control strategies against Culicoides vectors.

PubMed

Benelli, Giovanni; Buttazzoni, Luca; Canale, Angelo; D'Andrea, Armando; Del Serrone, Paola; Delrio, Gavino; Foxi, Cipriano; Mariani, Susanna; Savini, Giovanni; Vadivalagan, Chithravel; Murugan, Kadarkarai; Toniolo, Chiara; Nicoletti, Marcello; Serafini, Mauro

2017-12-01

Several arthropod-borne diseases are now rising with increasing impact and risks for public health, due to environmental changes and resistance to pesticides currently marketed. In addition to community surveillance programs and a careful management of herds, a next-generation of effective products is urgently needed to control the spread of these diseases, with special reference to arboviral ones. Natural product research can afford alternative solutions. Recently, a re-emerging of bluetongue disease is ongoing in Italy. Bluetongue is a viral disease that affects ruminants and is spread through the bite of bloodsucking insects, especially Culicoides species. In this review, we focused on the importance of vector control programs for prevention or bluetongue outbreaks, outlining the lack of effective tools in the fight against Culicoides vectors. Then, we analyzed a field case study in Sardinia (Italy) concerning the utilization of the neem cake (Azadirachta indica), to control young instar populations of Culicoides biting midges, the vectors of bluetongue virus. Neem cake is a cheap and eco-friendly by-product obtained from the extraction of neem oil. Overall, we propose that the employ of neem extraction by-products as aqueous formulations in muddy sites close to livestock grazing areas may represent an effective tool in the fight against the spread of bluetongue virus in the Mediterranean areas. Copyright © 2017 Elsevier Ltd. All rights reserved.

Detection of ferromagnetic target based on mobile magnetic gradient tensor system

NASA Astrophysics Data System (ADS)

Gang, Y. I. N.; Yingtang, Zhang; Zhining, Li; Hongbo, Fan; Guoquan, Ren

2016-03-01

Attitude change of mobile magnetic gradient tensor system critically affects the precision of gradient measurements, thereby increasing ambiguity in target detection. This paper presents a rotational invariant-based method for locating and identifying ferromagnetic targets. Firstly, unit magnetic moment vector was derived based on the geometrical invariant, such that the intermediate eigenvector of the magnetic gradient tensor is perpendicular to the magnetic moment vector and the source-sensor displacement vector. Secondly, unit source-sensor displacement vector was derived based on the characteristic that the angle between magnetic moment vector and source-sensor displacement is a rotational invariant. By introducing a displacement vector between two measurement points, the magnetic moment vector and the source-sensor displacement vector were theoretically derived. To resolve the problem of measurement noises existing in the realistic detection applications, linear equations were formulated using invariants corresponding to several distinct measurement points and least square solution of magnetic moment vector and source-sensor displacement vector were obtained. Results of simulation and principal verification experiment showed the correctness of the analytical method, along with the practicability of the least square method.

Effect of temperature post viral vector inoculation on the amount of hemagglutinin transiently expressed in Nicotiana benthamiana leaves.

PubMed

Matsuda, Ryo; Abe, Tatsuki; Fujiuchi, Naomichi; Matoba, Nobuyuki; Fujiwara, Kazuhiro

2017-09-01

Transient gene expression in whole plants by using viral vectors is promising as a rapid, mass production system for biopharmaceutical proteins. Recent studies have indicated that plant growth conditions such as air temperature markedly influence the accumulation levels of target proteins. Here, we investigated time course of the amount of recombinant hemagglutinin (HA), a vaccine antigen of influenza virus, in leaves of Nicotiana benthamiana plants grown at 20°C or 25°C post viral vector inoculation. The HA content per unit of leaf biomass increased and decreased from 4 to 6 days post inoculation at 20°C and 25°C, respectively, irrespective of the subcellular localization of HA. The overall HA contents were higher when HA was targeted to the endoplasmic reticulum (ER) rather than the apoplast. Necrosis of leaf tissues was specifically observed in plants inoculated with the ER-targeting vector and grown at 25°C. With the ER-targeting vector, the maximum HA contents at 20°C and 25°C were recorded at 6 and 4 days post inoculation, respectively, and were comparable to each other. HA contents thereafter decreased at both temperatures; the rate of reduction appeared faster at 25°C than at 20°C. From a practical point of view, our results indicate that the strategy of targeting HA to the ER, growing plants at a lower temperature of 20°C, and harvesting leaves at around a week after vector inoculation should be implemented to obtain a high HA yield stably and efficiently. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Extreme Weather Events and Impacts on Vector-borne Diseases and Agriculture

USDA-ARS?s Scientific Manuscript database

Extreme weather events during the period 2010-2012 impacted agriculture and vector-borne disease throughout the world. We evaluated specific weather events with satellite remotely sensed environmental data and evaluated crop production and diseases associated with these events. Significant droughts ...

Method and apparatus for second-rank tensor generation

NASA Technical Reports Server (NTRS)

Liu, Hua-Kuang (Inventor)

1991-01-01

A method and apparatus are disclosed for generation of second-rank tensors using a photorefractive crystal to perform the outer-product between two vectors via four-wave mixing, thereby taking 2n input data to a control n squared output data points. Two orthogonal amplitude modulated coherent vector beams x and y are expanded and then parallel sides of the photorefractive crystal in exact opposition. A beamsplitter is used to direct a coherent pumping beam onto the crystal at an appropriate angle so as to produce a conjugate beam that is the matrix product of the vector beam that propagates in the exact opposite direction from the pumping beam. The conjugate beam thus separated is the tensor output xy (sup T).

WEBGIS based CropWatch online agriculture monitoring system

NASA Astrophysics Data System (ADS)

Zhang, X.; Wu, B.; Zeng, H.; Zhang, M.; Yan, N.

2015-12-01

CropWatch, which was developed by the Institute of Remote Sensing and Digital Earth (RADI), Chinese Academy of Sciences (CAS), has achieved breakthrough results in the integration of methods, independence of the assessments and support to emergency response by periodically releasing global agricultural information. Taking advantages of the multi-source remote sensing data and the openness of the data sharing policies, CropWatch group reported their monitoring results by publishing four bulletins one year. In order to better analysis and generate the bulletin and provide an alternative way to access agricultural monitoring indicators and results in CropWatch, The CropWatch online system based on the WEBGIS techniques has been developed. Figure 1 shows the CropWatch online system structure and the system UI in Clustering mode. Data visualization is sorted into three different modes: Vector mode, Raster mode and Clustering mode. Vector mode provides the statistic value for all the indicators over each monitoring units which allows users to compare current situation with historical values (average, maximum, etc.). Users can compare the profiles of each indicator over the current growing season with the historical data in a chart by selecting the region of interest (ROI). Raster mode provides pixel based anomaly of CropWatch indicators globally. In this mode, users are able to zoom in to the regions where the notable anomaly was identified from statistic values in vector mode. Data from remote sensing image series at high temporal and low spatial resolution provide key information in agriculture monitoring. Clustering mode provides integrated information on different classes in maps, the corresponding profiles for each class and the percentage of area of each class to the total area of all classes. The time series data is categorized into limited types by the ISODATA algorithm. For each clustering type, pixels on the map, profiles, and percentage legend are all linked together. All the three visualization methods are applied to four scales including 65 monitoring and reporting units (MRUs), 7 major production zones (MPZs), 173 countries and sub-countries for 9 large countries. Agro-Climatic information, Agronomic information and indicators related with crop area, crop yield and crop production are provided.

The role of diatom resting spores in pelagic-benthic coupling in the Southern Ocean

NASA Astrophysics Data System (ADS)

Rembauville, Mathieu; Blain, Stéphane; Manno, Clara; Tarling, Geraint; Thompson, Anu; Wolff, George; Salter, Ian

2018-05-01

Natural iron fertilization downstream of Southern Ocean island plateaus supports large phytoplankton blooms and promotes carbon export from the mixed layer. In addition to sequestering atmospheric CO2, the biological carbon pump also supplies organic matter (OM) to deep-ocean ecosystems. Although the total flux of OM arriving at the seafloor sets the energy input to the system, the chemical nature of OM is also of significance. However, a quantitative framework linking ecological flux vectors to OM composition is currently lacking. In the present study we report the lipid composition of export fluxes collected by five moored sediment traps deployed in contrasting productivity regimes of Southern Ocean island systems (Kerguelen, Crozet and South Georgia) and compile them with quantitative data on diatom and faecal pellet fluxes. At the three naturally iron-fertilized sites, the relative contribution of labile lipids (mono- and polyunsaturated fatty acids, unsaturated fatty alcohols) is 2-4 times higher than at low productivity sites. There is a strong attenuation of labile components as a function of depth, irrespective of productivity. The three island systems also display regional characteristics in lipid export. An enrichment of zooplankton dietary sterols, such as C27Δ5, at South Georgia is consistent with high zooplankton and krill biomass in the region and the importance of faecal pellets to particulate organic carbon (POC) flux. There is a strong association of diatom resting spore fluxes that dominate productive flux regimes with energy-rich unsaturated fatty acids. At the Kerguelen Plateau we provide a statistical framework to link seasonal variation in ecological flux vectors and lipid composition over a complete annual cycle. Our analyses demonstrate that ecological processes in the upper ocean, e.g. resting spore formation and grazing, not only impact the magnitude and stoichiometry of the Southern Ocean biological pump, but also regulate the composition of exported OM and the nature of pelagic-benthic coupling.

Development of two bacterial artificial chromosome shuttle vectors for a recombination-based cloning and regulated expression of large genes in mammalian cells.

PubMed

Hong, Y K; Kim, D H; Beletskii, A; Lee, C; Memili, E; Strauss, W M

2001-04-01

Most conditional expression vectors designed for mammalian cells have been valuable systems for studying genes of interest by regulating their expressions. The available vectors, however, are reliable for the short-length cDNA clones and not optimal for relatively long fragments of genomic DNA or long cDNAs. Here, we report the construction of two bacterial artificial chromosome (BAC) vectors, capable of harboring large inserts and shuttling among Escherichia coli, yeast, and mammalian cells. These two vectors, pEYMT and pEYMI, contain conditional expression systems which are designed to be regulated by tetracycline and mouse interferons, respectively. To test the properties of the vectors, we cloned in both vectors the green fluorescence protein (GFP) through an in vitro ligation reaction and the 17.8-kb-long X-inactive-specific transcript (Xist) cDNA through homologous recombination in yeast. Subsequently, we characterized their regulated expression properties using real-time quantitative RT-PCR (TaqMan) and RNA-fluorescent in situ hybridization (FISH). We demonstrate that these two BAC vectors are good systems for recombination-based cloning and regulated expression of large genes in mammalian cells. Copyright 2001 Academic Press.

Techniques utilized in the simulated altitude testing of a 2D-CD vectoring and reversing nozzle

NASA Technical Reports Server (NTRS)

Block, H. Bruce; Bryant, Lively; Dicus, John H.; Moore, Allan S.; Burns, Maureen E.; Solomon, Robert F.; Sheer, Irving

1988-01-01

Simulated altitude testing of a two-dimensional, convergent-divergent, thrust vectoring and reversing exhaust nozzle was accomplished. An important objective of this test was to develop test hardware and techniques to properly operate a vectoring and reversing nozzle within the confines of an altitude test facility. This report presents detailed information on the major test support systems utilized, the operational performance of the systems and the problems encountered, and test equipment improvements recommended for future tests. The most challenging support systems included the multi-axis thrust measurement system, vectored and reverse exhaust gas collection systems, and infrared temperature measurement systems used to evaluate and monitor the nozzle. The feasibility of testing a vectoring and reversing nozzle of this type in an altitude chamber was successfully demonstrated. Supporting systems performed as required. During reverser operation, engine exhaust gases were successfully captured and turned downstream. However, a small amount of exhaust gas spilled out the collector ducts' inlet openings when the reverser was opened more than 60 percent. The spillage did not affect engine or nozzle performance. The three infrared systems which viewed the nozzle through the exhaust collection system worked remarkably well considering the harsh environment.

Purification of adenoviral vectors by combined anion exchange and gel filtration chromatography.

PubMed

Eglon, Marc N; Duffy, Aoife M; O'Brien, Timothy; Strappe, Padraig M

2009-11-01

Adenoviral vectors are used extensively in human gene therapy trials and in vaccine development. Large-scale GMP production requires a downstream purification process, and liquid chromatography is emerging as the most powerful mode of purification, enabling the production of vectors at a clinically relevant scale and quality. The present study describes the development of a two-step high-performance liquid chromatography (HPLC) process combining anion exchange (AIEX) and gel filtration (GF) in comparison with the caesium chloride density gradient method. HEK-293 cells were cultured in ten-layer CellStacks() and infected with 10 pfu/cell of adenoviral vector expressing green fluorescent protein (Ad5-GFP). Cell-bound virus was harvested and benzonase added to digest DNA, crude lysate was clarified by centrifugation and filtration prior to HPLC. Chromatography fractions were added to HEK-293 cells and GFP expression measured using a fluorescent plate reader. Using AIEX then GF resulted in an adenoviral vector with purity comparable to Ad5-GFP purified by CsCl, whereas the reverse process (GF-AIEX) showed a reduced purity by electrophoresis and required further buffer exchange of the product. The optimal process (AIEX-GF) resulted in a vector yield of 2.3 x 10(7) pfu/cm(2) of cell culture harvested compared to 3.3 x 10(7) pfu/cm(2) for CsCl. The process recovery for the HPLC process was 36% compared to 27.5% for CsCl and total virion to infectious particle ratios of 18 and 11, respectively, were measured. We present a simple two-step chromatography process that is capable of producing high-quality adenovirus at a titre suitable for scale-up and clinical translation.

Improved dual AAV vectors with reduced expression of truncated proteins are safe and effective in the retina of a mouse model of Stargardt disease.

PubMed

Trapani, Ivana; Toriello, Elisabetta; de Simone, Sonia; Colella, Pasqualina; Iodice, Carolina; Polishchuk, Elena V; Sommella, Andrea; Colecchi, Linda; Rossi, Settimio; Simonelli, Francesca; Giunti, Massimo; Bacci, Maria L; Polishchuk, Roman S; Auricchio, Alberto

2015-12-01

Stargardt disease (STGD1) due to mutations in the large ABCA4 gene is the most common inherited macular degeneration in humans. We have shown that dual adeno-associated viral (AAV) vectors effectively transfer ABCA4 to the retina of Abca4-/- mice. However, they express both lower levels of transgene compared with a single AAV and truncated proteins. To increase productive dual AAV concatemerization, which would overcome these limitations, we have explored the use of either various regions of homology or heterologous inverted terminal repeats (ITR). In addition, we tested the ability of various degradation signals to decrease the expression of truncated proteins. We found the highest levels of transgene expression using regions of homology based on either alkaline phosphatase or the F1 phage (AK). The use of heterologous ITR does not decrease the levels of truncated proteins relative to full-length ABCA4 and impairs AAV vector production. Conversely, the inclusion of the CL1 degradation signal results in the selective degradation of truncated proteins from the 5'-half without affecting full-length protein production. Therefore, we developed dual AAV hybrid ABCA4 vectors including homologous ITR2, the photoreceptor-specific G protein-coupled receptor kinase 1 promoter, the AK region of homology and the CL1 degradation signal. We show that upon subretinal administration these vectors are both safe in pigs and effective in Abca4-/- mice. Our data support the use of improved dual AAV vectors for gene therapy of STGD1. © The Author 2015. Published by Oxford University Press.

A support vector machine approach for classification of welding defects from ultrasonic signals

NASA Astrophysics Data System (ADS)

Chen, Yuan; Ma, Hong-Wei; Zhang, Guang-Ming

2014-07-01

Defect classification is an important issue in ultrasonic non-destructive evaluation. A layered multi-class support vector machine (LMSVM) classification system, which combines multiple SVM classifiers through a layered architecture, is proposed in this paper. The proposed LMSVM classification system is applied to the classification of welding defects from ultrasonic test signals. The measured ultrasonic defect echo signals are first decomposed into wavelet coefficients by the wavelet packet transform. The energy of the wavelet coefficients at different frequency channels are used to construct the feature vectors. The bees algorithm (BA) is then used for feature selection and SVM parameter optimisation for the LMSVM classification system. The BA-based feature selection optimises the energy feature vectors. The optimised feature vectors are input to the LMSVM classification system for training and testing. Experimental results of classifying welding defects demonstrate that the proposed technique is highly robust, precise and reliable for ultrasonic defect classification.

A tetracycline inducible expression vector for Corynebacterium glutamicum allowing tightly regulable gene expression.

PubMed

Lausberg, Frank; Chattopadhyay, Ava Rebecca; Heyer, Antonia; Eggeling, Lothar; Freudl, Roland

2012-09-01

Here we report on the construction of a tetracycline inducible expression vector that allows a tightly regulable gene expression in Corynebacterium glutamicum which is used in industry for production of small molecules such as amino acids. Using the green fluorescent protein (GFP) as a reporter protein we show that this vector, named pCLTON1, is characterized by tight repression under non-induced conditions as compared to a conventional IPTG inducible expression vector, and that it allows gradual GFP synthesis upon gradual increase of anhydrotetracycline addition. Copyright © 2012 Elsevier Inc. All rights reserved.

Comparing Aedes vigilax Eggshell Densities in Saltmarsh and Mangrove Systems with Implications for Management.

PubMed

Dale, Pat; Knight, Jon; Griffin, Lachlan

2014-12-12

Aedes vigilax (Skuse), a nuisance and disease vector, is prolific in intertidal wetlands in Australia. Aedine mosquitoes oviposit directly onto substrate. The eggshells are relatively stable spatially and temporally, providing an estimate of mosquito larval production. The aims of the research were to compare, at a general level, oviposition in mangroves and saltmarshes, and to compare oviposition between different habitats within mangroves and saltmarshes. The results indicated that there were no significant differences between production in mangrove and saltmarsh overall. However, within each system there were significant differences between habitat classes, with mangrove hummocks being the most productive. All classes, except for fringing mangrove forests, produced sufficient densities of eggshells (>0.05/cc) to warrant concern. While mosquito production in mangroves is known, the significantly higher production rates in the mangrove hummock habitats had not been demonstrated. This warrants improved management strategies that both specifically target these parts of mangrove systems and, secondly, addresses the longer-term potential for mangrove hummock habitats developing in the future; such as, in response to sea level rise and mangrove encroachment into saltmarsh. A strategy to increase tidal flushing within the systems would improve water quality and mitigate adverse impacts while providing a source reduction outcome.

Hyperbolic-symmetry vector fields.

PubMed

Gao, Xu-Zhen; Pan, Yue; Cai, Meng-Qiang; Li, Yongnan; Tu, Chenghou; Wang, Hui-Tian

2015-12-14

We present and construct a new kind of orthogonal coordinate system, hyperbolic coordinate system. We present and design a new kind of local linearly polarized vector fields, which is defined as the hyperbolic-symmetry vector fields because the points with the same polarization form a series of hyperbolae. We experimentally demonstrate the generation of such a kind of hyperbolic-symmetry vector optical fields. In particular, we also study the modified hyperbolic-symmetry vector optical fields with the twofold and fourfold symmetric states of polarization when introducing the mirror symmetry. The tight focusing behaviors of these vector fields are also investigated. In addition, we also fabricate micro-structures on the K9 glass surfaces by several tightly focused (modified) hyperbolic-symmetry vector fields patterns, which demonstrate that the simulated tightly focused fields are in good agreement with the fabricated micro-structures.

Measurement of vector boson plus D*(2010)+ meson production in p ¯p collisions at √{s }=1.96 TeV

NASA Astrophysics Data System (ADS)

Aaltonen, T.; Amerio, S.; Amidei, D.; Anastassov, A.; Annovi, A.; Antos, J.; Apollinari, G.; Appel, J. A.; Arisawa, T.; Artikov, A.; Asaadi, J.; Ashmanskas, W.; Auerbach, B.; Aurisano, A.; Azfar, F.; Badgett, W.; Bae, T.; Barbaro-Galtieri, A.; Barnes, V. E.; Barnett, B. A.; Barria, P.; Bartos, P.; Bauce, M.; Bedeschi, F.; Behari, S.; Bellettini, G.; Bellinger, J.; Benjamin, D.; Beretvas, A.; Bhatti, A.; Bland, K. R.; Blumenfeld, B.; Bocci, A.; Bodek, A.; Bortoletto, D.; Boudreau, J.; Boveia, A.; Brigliadori, L.; Bromberg, C.; Brucken, E.; Budagov, J.; Budd, H. S.; Burkett, K.; Busetto, G.; Bussey, P.; Butti, P.; Buzatu, A.; Calamba, A.; Camarda, S.; Campanelli, M.; Canelli, F.; Carls, B.; Carlsmith, D.; Carosi, R.; Carrillo, S.; Casal, B.; Casarsa, M.; Castro, A.; Catastini, P.; Cauz, D.; Cavaliere, V.; Cerri, A.; Cerrito, L.; Chen, Y. C.; Chertok, M.; Chiarelli, G.; Chlachidze, G.; Cho, K.; Chokheli, D.; Clark, A.; Clarke, C.; Convery, M. E.; Conway, J.; Corbo, M.; Cordelli, M.; Cox, C. A.; Cox, D. J.; Cremonesi, M.; Cruz, D.; Cuevas, J.; Culbertson, R.; d'Ascenzo, N.; Datta, M.; de Barbaro, P.; Demortier, L.; Deninno, M.; D'Errico, M.; Devoto, F.; Di Canto, A.; Di Ruzza, B.; Dittmann, J. R.; Donati, S.; D'Onofrio, M.; Dorigo, M.; Driutti, A.; Ebina, K.; Edgar, R.; Elagin, A.; Erbacher, R.; Errede, S.; Esham, B.; Farrington, S.; Fernández Ramos, J. P.; Field, R.; Flanagan, G.; Forrest, R.; Franklin, M.; Freeman, J. C.; Frisch, H.; Funakoshi, Y.; Galloni, C.; Garfinkel, A. F.; Garosi, P.; Gerberich, H.; Gerchtein, E.; Giagu, S.; Giakoumopoulou, V.; Gibson, K.; Ginsburg, C. M.; Giokaris, N.; Giromini, P.; Glagolev, V.; Glenzinski, D.; Gold, M.; Goldin, D.; Golossanov, A.; Gomez, G.; Gomez-Ceballos, G.; Goncharov, M.; González López, O.; Gorelov, I.; Goshaw, A. T.; Goulianos, K.; Gramellini, E.; Grosso-Pilcher, C.; Group, R. C.; Guimaraes da Costa, J.; Hahn, S. R.; Han, J. Y.; Happacher, F.; Hara, K.; Hare, M.; Harr, R. F.; Harrington-Taber, T.; Hatakeyama, K.; Hays, C.; Heinrich, J.; Herndon, M.; Hocker, A.; Hong, Z.; Hopkins, W.; Hou, S.; Hughes, R. E.; Husemann, U.; Hussein, M.; Huston, J.; Introzzi, G.; Iori, M.; Ivanov, A.; James, E.; Jang, D.; Jayatilaka, B.; Jeon, E. J.; Jindariani, S.; Jones, M.; Joo, K. K.; Jun, S. Y.; Junk, T. R.; Kambeitz, M.; Kamon, T.; Karchin, P. E.; Kasmi, A.; Kato, Y.; Ketchum, W.; Keung, J.; Kilminster, B.; Kim, D. H.; Kim, H. S.; Kim, J. E.; Kim, M. J.; Kim, S. H.; Kim, S. B.; Kim, Y. J.; Kim, Y. K.; Kimura, N.; Kirby, M.; Knoepfel, K.; Kondo, K.; Kong, D. J.; Konigsberg, J.; Kotwal, A. V.; Kreps, M.; Kroll, J.; Kruse, M.; Kuhr, T.; Kurata, M.; Laasanen, A. T.; Lammel, S.; Lancaster, M.; Lannon, K.; Latino, G.; Lee, H. S.; Lee, J. S.; Leo, S.; Leone, S.; Lewis, J. D.; Limosani, A.; Lipeles, E.; Lister, A.; Liu, H.; Liu, Q.; Liu, T.; Lockwitz, S.; Loginov, A.; Lucchesi, D.; Lucà, A.; Lueck, J.; Lujan, P.; Lukens, P.; Lungu, G.; Lys, J.; Lysak, R.; Madrak, R.; Maestro, P.; Malik, S.; Manca, G.; Manousakis-Katsikakis, A.; Marchese, L.; Margaroli, F.; Marino, P.; Matera, K.; Mattson, M. E.; Mazzacane, A.; Mazzanti, P.; McNulty, R.; Mehta, A.; Mehtala, P.; Mesropian, C.; Miao, T.; Mietlicki, D.; Mitra, A.; Miyake, H.; Moed, S.; Moggi, N.; Moon, C. S.; Moore, R.; Morello, M. J.; Mukherjee, A.; Muller, Th.; Murat, P.; Mussini, M.; Nachtman, J.; Nagai, Y.; Naganoma, J.; Nakano, I.; Napier, A.; Nett, J.; Neu, C.; Nigmanov, T.; Nodulman, L.; Noh, S. Y.; Norniella, O.; Oakes, L.; Oh, S. H.; Oh, Y. D.; Oksuzian, I.; Okusawa, T.; Orava, R.; Ortolan, L.; Pagliarone, C.; Palencia, E.; Palni, P.; Papadimitriou, V.; Parker, W.; Pauletta, G.; Paulini, M.; Paus, C.; Phillips, T. J.; Piacentino, G.; Pianori, E.; Pilot, J.; Pitts, K.; Plager, C.; Pondrom, L.; Poprocki, S.; Potamianos, K.; Pranko, A.; Prokoshin, F.; Ptohos, F.; Punzi, G.; Redondo Fernández, I.; Renton, P.; Rescigno, M.; Rimondi, F.; Ristori, L.; Robson, A.; Rodriguez, T.; Rolli, S.; Ronzani, M.; Roser, R.; Rosner, J. L.; Ruffini, F.; Ruiz, A.; Russ, J.; Rusu, V.; Sakumoto, W. K.; Sakurai, Y.; Santi, L.; Sato, K.; Saveliev, V.; Savoy-Navarro, A.; Schlabach, P.; Schmidt, E. E.; Schwarz, T.; Scodellaro, L.; Scuri, F.; Seidel, S.; Seiya, Y.; Semenov, A.; Sforza, F.; Shalhout, S. Z.; Shears, T.; Shepard, P. F.; Shimojima, M.; Shochet, M.; Shreyber-Tecker, I.; Simonenko, A.; Sliwa, K.; Smith, J. R.; Snider, F. D.; Song, H.; Sorin, V.; Denis, R. St.; Stancari, M.; Stentz, D.; Strologas, J.; Sudo, Y.; Sukhanov, A.; Suslov, I.; Takemasa, K.; Takeuchi, Y.; Tang, J.; Tecchio, M.; Teng, P. K.; Thom, J.; Thomson, E.; Thukral, V.; Toback, D.; Tokar, S.; Tollefson, K.; Tomura, T.; Tonelli, D.; Torre, S.; Torretta, D.; Totaro, P.; Trovato, M.; Ukegawa, F.; Uozumi, S.; Vázquez, F.; Velev, G.; Vellidis, C.; Vernieri, C.; Vidal, M.; Vilar, R.; Vizán, J.; Vogel, M.; Volpi, G.; Wagner, P.; Wallny, R.; Wang, S. M.; Waters, D.; Wester, W. C.; Whiteson, D.; Wicklund, A. B.; Wilbur, S.; Williams, H. H.; Wilson, J. S.; Wilson, P.; Winer, B. L.; Wittich, P.; Wolbers, S.; Wolfe, H.; Wright, T.; Wu, X.; Wu, Z.; Yamamoto, K.; Yamato, D.; Yang, T.; Yang, U. K.; Yang, Y. C.; Yao, W.-M.; Yeh, G. P.; Yi, K.; Yoh, J.; Yorita, K.; Yoshida, T.; Yu, G. B.; Yu, I.; Zanetti, A. M.; Zeng, Y.; Zhou, C.; Zucchelli, S.; CDF Collaboration

2016-03-01

A measurement of vector boson (V ) production in conjunction with a D*(2010)+meson is presented. Using a data sample corresponding to 9.7 fb-1 of proton-antiproton collisions at center-of-mass energy √{s }=1.96 TeV produced by the Fermilab Tevatron, we reconstruct V +D*+ samples with the CDF II detector. The D*+ is fully reconstructed in the D*(2010)+→D0(→K-π+)π+ decay mode. This technique is sensitive to the associated production of vector boson plus charm or bottom mesons. We measure the ratio of production cross sections σ (W +D*)/σ (W )=[1.75 ±0.13 (stat ) ±0.09 (stat ) ]% and σ (Z +D*)/σ (Z )=[1.5 ±0.4 (stat ) ±0.2 (stat ) ]% and perform a differential measurement of d σ (W +D*)/d pT(D*). Event properties are utilized to determine the fraction of V +D*(2010)+ events originating from different production processes. The results are in agreement with the predictions obtained with the pythia program, limiting possible contribution from non-standard-model physics processes.

A ocean bottom vector magnetometer

NASA Astrophysics Data System (ADS)

Wang, Xiaomei; Teng, Yuntian; Wang, Chen; Ma, Jiemei

2017-04-01

The new development instrument with a compact spherical coil system and Overhauser magnetometer for measuring the total strength of the magnetic field and the vectors of strength, Delta inclination - Delta declination, meanwhile we also use a triaxial fluxgate instrument of the traditional instrument for geomagnetic vector filed measurement. The advantages of this method are be calibrated by each other and get good performances with automatic operation, good stability and high resolution. Firstly, a brief description of the instrument measurement principles and the key technologies are given. The instrument used a spherical coil system with 34 coils to product the homogeneous volume inside the coils which is large enough to accommodate the sensor of Overhauser total field sensor; the rest of the footlocker-sized ocean-bottom vector magnetometer consists of equipment to run the sensors and records its data (batteries and a data logger), weight to sink it to the sea floor, a remote-controlled acoustic release and flotation to bring the instrument back to the surface. Finally, the accuracy of the instrument was tested in the Geomagnetic station, and the measurement accuracies of total strength and components were better than 0.2nT and 1nT respectively. The figure 1 shows the development instrument structure. it includes six thick glass spheres which protect the sensor, data logger and batteries from the pressures of the deep sea, meanwhile they also provide recycling positive buoyancy; To cushion the glass, the spheres then go inside yellow plastic "hardhats". The triaxial fluxgate is inside No.1 glass spheres, data logger and batteries are inside No.2 glass spheres, the new vector sensor is inside No.3 glass spheres, acoustic communication unit is inside No.4 glass spheres, No.5 and No.6 glass spheres are empty which only provide recycling positive buoyancy. The figure 2 shows the development instrument Physical photo.

Vector-boson-tagged jet production in heavy ion collisions at energies available at the CERN Large Hadron Collider [Vector boson-tagged jet production in heavy ion collisions at the LHC

DOE PAGES

Kang, Zhong -Bo; Vitev, Ivan Mateev; Xing, Hongxi

2017-07-25

Here, vector-boson-tagged jet production in collisions of heavy nuclei opens new opportunities to study parton shower formation and propagation in strongly interacting matter. It has been argued to provide a golden channel that can constrain the energy loss of jets in the quark-gluon plasma created in heavy ion reactions. We present theoretical results for isolated-photon-tagged and Z 0-boson-tagged jet production in Pb + Pb collisions with √s NN = 5.02TeV at the LHC. Specifically, we evaluate the transverse momentum imbalance x JV distribution and nuclear modification factor I AA of tagged jets and compare our theoretical calculations to recent experimentalmore » measurements by ATLAS and CMS collaborations. Our analysis, which includes both collisional and radiative energy losses, sheds light on their relative importance versus the strength of jet-medium interactions and helps quantify the amount of out-of-cone radiation of predominantly prompt quark-initiated jets.« less

Justice foundations for the Comprehensive Law Movement.

PubMed

Dewhurst, Dale

2010-01-01

Authors examining the developing dispute resolution alternatives to the adversarial system have identified nine converging "vectors" or alternatives in what has been termed the Comprehensive Law Movement. These authors have sought to understand how the developing vectors can remain separate and vibrant movements while sharing common ground. Some analyze these developments as being within law and legal practice, others see them as alternative approaches to law, and still others take a combined approach. It will be impossible to understand how these vectors have meaningful differences from law and legal practice if the search is limited to looking within law and legal practice. It will be impossible to understand how these vectors have meaningful commonalities with law and legal practice if the search is limited to looking external to law and legal practice. Instead of comparing the vectors with the adversarial system, higher order criteria are required. What is needed is a comprehensive and internally consistent super-system of norms; one that can be used to evaluate the adversarial system and the evolving vectors on an equal footing. An Aristotelian natural law virtue theory of justice can: (a) provide a functional guiding definition of justice; (b) serve as a comprehensive and internally consistent super-system of norms; and (c) provide the theoretical and evaluative foundation required to clarify the relationships among the adversarial system and the developing vectors. Finally, it will become clear why the Comprehensive Law Movement might be more appropriately conceptualized as the Comprehensive Justice Movement. Copyright © 2010 Elsevier Ltd. All rights reserved.

A multiplex TaqMan qPCR assay for sensitive and rapid detection of phytoplasmas infecting Rubus species.

PubMed

Linck, Holger; Krüger, Erika; Reineke, Annette

2017-01-01

Rubus stunt is an economically important disease in the production of raspberries, blackberries, and loganberries. A fast, sensitive, and reliable diagnosis of phytoplasmas, the causal agent of the disease, is of prime importance to stop its spread by vegetative propagation and by insect vectors. Therefore, multiplex qPCR assays using TaqMan probes with different kinds of fluorophores in one reaction were developed, allowing the detection of phytoplasmas in general as well as a more specific detection of phytoplasmas belonging to group 16SrV and host DNA (either plant or insect). This assay now provides a practical tool for the screening of motherplants and monitoring the presence and distribution of phytoplasmas in Rubus plants of different geographic origins, cultivars, and cultivation systems, as well as in putative insect vectors like leafhoppers.

A multiplex TaqMan qPCR assay for sensitive and rapid detection of phytoplasmas infecting Rubus species

PubMed Central

Krüger, Erika; Reineke, Annette

2017-01-01

Rubus stunt is an economically important disease in the production of raspberries, blackberries, and loganberries. A fast, sensitive, and reliable diagnosis of phytoplasmas, the causal agent of the disease, is of prime importance to stop its spread by vegetative propagation and by insect vectors. Therefore, multiplex qPCR assays using TaqMan probes with different kinds of fluorophores in one reaction were developed, allowing the detection of phytoplasmas in general as well as a more specific detection of phytoplasmas belonging to group 16SrV and host DNA (either plant or insect). This assay now provides a practical tool for the screening of motherplants and monitoring the presence and distribution of phytoplasmas in Rubus plants of different geographic origins, cultivars, and cultivation systems, as well as in putative insect vectors like leafhoppers. PMID:28545043

Can different quantum state vectors correspond to the same physical state? An experimental test

NASA Astrophysics Data System (ADS)

Nigg, Daniel; Monz, Thomas; Schindler, Philipp; Martinez, Esteban A.; Hennrich, Markus; Blatt, Rainer; Pusey, Matthew F.; Rudolph, Terry; Barrett, Jonathan

2016-01-01

A century after the development of quantum theory, the interpretation of a quantum state is still discussed. If a physicist claims to have produced a system with a particular quantum state vector, does this represent directly a physical property of the system, or is the state vector merely a summary of the physicist’s information about the system? Assume that a state vector corresponds to a probability distribution over possible values of an unknown physical or ‘ontic’ state. Then, a recent no-go theorem shows that distinct state vectors with overlapping distributions lead to predictions different from quantum theory. We report an experimental test of these predictions using trapped ions. Within experimental error, the results confirm quantum theory. We analyse which kinds of models are ruled out.

Classification of subsurface objects using singular values derived from signal frames

DOEpatents

Chambers, David H; Paglieroni, David W

2014-05-06

The classification system represents a detected object with a feature vector derived from the return signals acquired by an array of N transceivers operating in multistatic mode. The classification system generates the feature vector by transforming the real-valued return signals into complex-valued spectra, using, for example, a Fast Fourier Transform. The classification system then generates a feature vector of singular values for each user-designated spectral sub-band by applying a singular value decomposition (SVD) to the N.times.N square complex-valued matrix formed from sub-band samples associated with all possible transmitter-receiver pairs. The resulting feature vector of singular values may be transformed into a feature vector of singular value likelihoods and then subjected to a multi-category linear or neural network classifier for object classification.

Plasmid Vectors for Proteomic Analyses in Giardia: Purification of Virulence Factors and Analysis of the Proteasome

PubMed Central

Stadelmann, Britta; Birkestedt, Sandra; Hellman, Ulf; Svärd, Staffan G.

2012-01-01

In recent years, proteomics has come of age with the development of efficient tools for purification, identification, and characterization of gene products predicted by genome projects. The intestinal protozoan Giardia intestinalis can be transfected, but there is only a limited set of vectors available, and most of them are not user friendly. This work delineates the construction of a suite of cassette-based expression vectors for use in Giardia. Expression is provided by the strong constitutive ornithine carbamoyltransferase (OCT) promoter, and tagging is possible in both N- and C-terminal configurations. Taken together, the vectors are capable of providing protein localization and production of recombinant proteins, followed by efficient purification by a novel affinity tag combination, streptavidin binding peptide–glutathione S-transferase (SBP-GST). The option of removing the tags from purified proteins was provided by the inclusion of a PreScission protease site. The efficiency and feasibility of producing and purifying endogenous recombinant Giardia proteins with the developed vectors was demonstrated by the purification of active recombinant arginine deiminase (ADI) and OCT from stably transfected trophozoites. Moreover, we describe the tagging, purification by StrepTactin affinity chromatography, and compositional analysis by mass spectrometry of the G. intestinalis 26S proteasome by employing the Strep II-FLAG–tandem affinity purification (SF-TAP) tag. This is the first report of efficient production and purification of recombinant proteins in and from Giardia, which will allow the study of specific parasite proteins and protein complexes. PMID:22611020

Analyzing angular distributions for two-step dissociation mechanisms in velocity map imaging.

PubMed

Straus, Daniel B; Butler, Lynne M; Alligood, Bridget W; Butler, Laurie J

2013-08-15

Increasingly, velocity map imaging is becoming the method of choice to study photoinduced molecular dissociation processes. This paper introduces an algorithm to analyze the measured net speed, P(vnet), and angular, β(vnet), distributions of the products from a two-step dissociation mechanism, where the first step but not the second is induced by absorption of linearly polarized laser light. Typically, this might be the photodissociation of a C-X bond (X = halogen or other atom) to produce an atom and a momentum-matched radical that has enough internal energy to subsequently dissociate (without the absorption of an additional photon). It is this second step, the dissociation of the unstable radicals, that one wishes to study, but the measured net velocity of the final products is the vector sum of the velocity imparted to the radical in the primary photodissociation (which is determined by taking data on the momentum-matched atomic cophotofragment) and the additional velocity vector imparted in the subsequent dissociation of the unstable radical. The algorithm allows one to determine, from the forward-convolution fitting of the net velocity distribution, the distribution of velocity vectors imparted in the second step of the mechanism. One can thus deduce the secondary velocity distribution, characterized by a speed distribution P(v1,2°) and an angular distribution I(θ2°), where θ2° is the angle between the dissociating radical's velocity vector and the additional velocity vector imparted to the product detected from the subsequent dissociation of the radical.

Transfected babesia bovis expressing a tick GST as a live vector vaccine

USDA-ARS?s Scientific Manuscript database

The Rhipicephalus microplus tick is a notorious blood-feeding ectoparasite of livestock, especially cattle, responsible for massive losses in animal production. It is the main vector for transmission of pathogenic bacteria and parasites, including Babesia bovis, an intraerythrocytic apicomplexan pro...

Recent weather extremes and impact agricultural production and vector-borne disease patterns

USDA-ARS?s Scientific Manuscript database

We document significant worldwide weather anomalies that affected agriculture and vector-borne disease outbreaks during the 2010-2012 period. We utilized 2000-2012 vegetation index and land surface temperature data from NASA’s satellite-based Moderate Resolution Imaging Spectroradiometer (MODIS) to ...

Search for single production of a heavy vector-like T quark decaying to a Higgs boson and a top quark with a lepton and jets in the final state

DOE Office of Scientific and Technical Information (OSTI.GOV)

Khachatryan, V.; Sirunyan, A. M.; Tumasyan, A.

A search for single production of vector-like top quark partners (T) decaying into a Higgs boson and a top quark is performed using data from pp collisions at a centre-of-mass energy of 13 TeV collected by the CMS experiment at the CERN LHC, corresponding to an integrated luminosity of 2.3 inverse femtobarns. The top quark decay includes an electron or a muon while the Higgs boson decays into a pair of b quarks. No significant excess over standard model backgrounds is observed. Exclusion limits on the product of the production cross section and the branching fraction are derived in themore » T quark mass range 700 to 1800 GeV. For a mass of 1000 GeV, values of the product of the production cross section and the branching fraction greater than 0.8 and 0.7 pb are excluded at 95% confidence level, assuming left- and right-handed coupling of the T quark to standard model particles, respectively. This is the first analysis setting exclusion limits on the cross section of singly produced vector-like T quarks at a centre-of-mass energy of 13 TeV.« less