Science.gov

Sample records for acid constructs vectors

  1. Genetic engineering techniques for lactic acid bacteria: construction of a stable shuttle vector and expression vector for β-glucuronidase.

    PubMed

    Chang, Shiao-Ming; Yan, Tsong-Rong

    2014-02-01

    The shuttle vector, pUL6erm, was constructed by using a replicon from pL2, a multiple cloning site, colE1 ori, the ori of Gram-negative bacteria from vector pUC19, and the erythromycin resistance gene from pVA838 as a selection marker. pUL6erm could be transformed easily and maintained stably in Lactococcus lactis, Streptococcus thermophilus, Lactobacillus plantarum and Lactobacillus casei. Transformation assays of pUL6erm indicated that it had a narrow host range. β-Glucuronidase was induced in the presence of 0.3 M NaCl and 50 mM glutamate and expressed at 2.4 U mg(-1) with the expression vector (pUL6erm-gadR-GUS) constructed based on pUL6erm carrying β-glucuronidase gene wuth a chloride-inducible (gadR) expression cassette using Pgad as promoter. Therefore, pUL6erm and pUL6erm-gadR-GUS might be a safe and useful genetic tool for the improvement of lactic acid bacteria. PMID:24101246

  2. [Methods for construction of transgenic plant expression vector: a review].

    PubMed

    Zhang, Yangpu; Yang, Shushen

    2015-03-01

    Construction of recombinant plasmid vector for gene expression is a key step in making transgenic plants and important to study gene function and plant genetic engineering. A right choice of gene construction method can be cost-effective and achieve more diverse recombinant plasmids. In addition to the traditional methods in construction of plant gene expression vectors, such as Gateway technology, three DNA method and one step cloning, a few novel methods have been developed in recent years. These methods include oligonucleotide synthesis-based construction of small fragment gene expression vectors via competitive connection; construction of small RNA expression vector using pre-microRNA; recombination-fusion PCR method which inserts DNA fragments of multiple restriction sites into the target vector; and insertion of a DNA fragment into any region of a linear vector via In-Fusion Kit. Construction of complex vectors with many fragments uses sequence and ligation-independent cloning method, Gibson isothermal assembly or Golden Gate assembly. This paper summarizes our working experience in the area of recombinant vector construction and reports from others with an intention to disseminate ideas about currently widely used DNA recombination methods for plant transformation. PMID:26204753

  3. Cationic Lipid-Based Nucleic Acid Vectors.

    PubMed

    Jubeli, Emile; Goldring, William P D; Pungente, Michael D

    2016-01-01

    The delivery of nucleic acids into cells remains an important laboratory cell culture technique and potential clinical therapy, based upon the initial cellular uptake, then translation into protein (in the case of DNA), or gene deletion by RNA interference (RNAi). Although viral delivery vectors are more efficient, the high production costs, limited cargo capacity, and the potential for clinical adverse events make nonviral strategies attractive. Cationic lipids are the most widely applied and studied nonviral vectors; however, much remains to be solved to overcome limitations of these systems. Advances in the field of cationic lipid-based nucleic acid (lipoplex) delivery rely upon the development of robust and reproducible lipoplex formulations, together with the use of cell culture assays. This chapter provides detailed protocols towards the formulation, delivery, and assessment of in vitro cationic lipid-based delivery of DNA. PMID:27436310

  4. Automatic selection of a subset size at vector Helds construction

    NASA Astrophysics Data System (ADS)

    Panin, S. V.; Luybutin, P. S.; Titkov, V. V.; Cherepanova, M. V.

    2015-10-01

    An algorithm for selection of the size of a correlation kernel at displacement vector field construction by the method of digital image correlation has been proposed. The algorithm has been tested on simulated and experimental optical images having different texture. The influence of the correlation kernel size and image texture on nose immunity at determining displacements has been studied. It is shown that the proposed algorithm allows to find this size providing the minimum error when determination of displacements and estimation of deformation.

  5. Self-assembling multimeric nucleic acid constructs

    DOEpatents

    Cantor, Charles R.; Niemeyer, Christof M.; Smith, Cassandra L.; Sano, Takeshi; Hnatowich, Donald J.; Rusckowski, Mary

    1996-01-01

    The invention is directed to constructs and compositions containing multimeric forms of nucleic acid. Multimeric nucleic acids comprise single-stranded nucleic acids attached via biotin to streptavidin and bound with a functional group. These constructs can be utilized in vivo to treat or identify diseased tissue or cells. Repeated administrations of multimeric nucleic acid compositions produce a rapid and specific amplification of nucleic acid constructs and their attached functional groups. For treatment purposes, functional groups may be toxins, radioisotopes, genes or enzymes. Diagnostically, labeled multimeric constructs may be used to identify specific targets in vivo or in vitro. Multimeric nucleic acids may also be used in nanotechnology and to create self-assembling polymeric aggregates such as membranes of defined porosity, microcircuits and many other products.

  6. Self-assembling multimeric nucleic acid constructs

    DOEpatents

    Cantor, Charles R.; Niemeyer, Christof M.; Smith, Cassandra L.; Sano, Takeshi; Hnatowich, Donald J.; Rusckowski, Mary

    1999-10-12

    The invention is directed to constructs and compositions containing multimeric forms of nucleic acid. Multimeric nucleic acids comprise single-stranded nucleic acids attached via biotin to streptavidin and bound with a functional group. These constructs can be utilized in vivo to treat or identify diseased tissue or cells. Repeated administrations of multimeric nucleic acid compositions produce a rapid and specific amplification of nucleic acid constructs and their attached functional groups. For treatment purposes, functional groups may be toxins, radioisotopes, genes or enzymes. Diagnostically, labeled multimeric constructs may be used to identify specific targets in vivo or in vitro. Multimeric nucleic acids may also be used in nanotechnology and to create self-assembling polymeric aggregates such as membranes of defined porosity, microcircuits and many other products.

  7. Self-assembling multimeric nucleic acid constructs

    DOEpatents

    Cantor, C.R.; Niemeyer, C.M.; Smith, C.L.; Sano, Takeshi; Hnatowich, D.J.; Rusckowski, M.

    1996-10-01

    The invention is directed to constructs and compositions containing multimeric forms of nucleic acid. Multimeric nucleic acids comprise single-stranded nucleic acids attached via biotin to streptavidin and bound with a functional group. These constructs can be utilized in vivo to treat or identify diseased tissue or cells. Repeated administrations of multimeric nucleic acid compositions produce a rapid and specific amplification of nucleic acid constructs and their attached functional groups. For treatment purposes, functional groups may be toxins, radioisotopes, genes or enzymes. Diagnostically, labeled multimeric constructs may be used to identify specific targets in vivo or in vitro. Multimeric nucleic acids may also be used in nanotechnology and to create self-assembling polymeric aggregates such as membranes of defined porosity, microcircuits and many other products. 5 figs.

  8. Molecular cloning and expression vector construction of bovine TRIM28.

    PubMed

    Ma, X; Zhai, Z C; Zhang, M L; Song, B H; Zhu, Y R; Yang, S B; Dong, X Q; Su, L Y; Wang, C F; Ma, H X; Luan, W M

    2016-01-01

    The bovine TRIM28 gene was amplified from ovary tissue by using RT-PCR. The TRIM28 gene was inserted into the eukaryotic expression vector pIRES2-EGFP and transfected into bovine fetal fibroblasts by using Lipofectamine 3000. TRIM28 mRNA and protein were detected by fluorescence microscope and western blotting. The results showed that the full length of TRIM28 was cloned and pIRES2-EGFP-TRIM28 was constructed successfully. EGFP expression was observed, and the pIRES2-EGFP-TRIM28 transfected group expressed more TRIM28 protein than that by the pIRES2-EGFP group. The TIMR28 gene has been successfully transferred into bovine fetal fibroblasts. PMID:27420979

  9. Construction and Evaluation of Novel Rhesus Monkey Adenovirus Vaccine Vectors

    DOE PAGESBeta

    Abbink, Peter; Maxfield, Lori F.; Ng'ang'a, David; Borducchi, Erica N.; Iampietro, M. Justin; Bricault, Christine A.; Teigler, Jeffrey E.; Blackmore, Stephen; Parenteau, Lily; Wagh, Kshitij; et al

    2014-11-19

    Adenovirus vectors are widely used as vaccine candidates for a variety of pathogens, including HIV-1. To date, human and chimpanzee adenoviruses have been explored in detail as vaccine vectors. Furthermore, the phylogeny of human and chimpanzee adenoviruses is overlapping, and preexisting humoral and cellular immunity to both are exhibited in human populations worldwide. More distantly related adenoviruses may therefore offer advantages as vaccine vectors. We describe the primary isolation and vectorization of three novel adenoviruses from rhesus monkeys. The seroprevalence of these novel rhesus monkey adenovirus vectors was extremely low in sub-Saharan Africa human populations, and these vectors proved tomore » have immunogenicity comparable to that of human and chimpanzee adenovirus vaccine vectors in mice. These rhesus monkey adenoviruses phylogenetically clustered with the poorly described adenovirus species G and robustly stimulated innate immune responses. These novel adenoviruses represent a new class of candidate vaccine vectors.« less

  10. Construction and Evaluation of Novel Rhesus Monkey Adenovirus Vaccine Vectors

    SciTech Connect

    Abbink, Peter; Maxfield, Lori F.; Ng'ang'a, David; Borducchi, Erica N.; Iampietro, M. Justin; Bricault, Christine A.; Teigler, Jeffrey E.; Blackmore, Stephen; Parenteau, Lily; Wagh, Kshitij; Handley, Scott A.; Zhao, Guoyan; Virgin, Herbert W.; Korber, Bette; Barouch, Dan H.

    2014-11-19

    Adenovirus vectors are widely used as vaccine candidates for a variety of pathogens, including HIV-1. To date, human and chimpanzee adenoviruses have been explored in detail as vaccine vectors. Furthermore, the phylogeny of human and chimpanzee adenoviruses is overlapping, and preexisting humoral and cellular immunity to both are exhibited in human populations worldwide. More distantly related adenoviruses may therefore offer advantages as vaccine vectors. We describe the primary isolation and vectorization of three novel adenoviruses from rhesus monkeys. The seroprevalence of these novel rhesus monkey adenovirus vectors was extremely low in sub-Saharan Africa human populations, and these vectors proved to have immunogenicity comparable to that of human and chimpanzee adenovirus vaccine vectors in mice. These rhesus monkey adenoviruses phylogenetically clustered with the poorly described adenovirus species G and robustly stimulated innate immune responses. These novel adenoviruses represent a new class of candidate vaccine vectors.

  11. Lead-acid battery construction

    NASA Technical Reports Server (NTRS)

    Rowlette, John J. (Inventor)

    1988-01-01

    The power characteristics of a lead-acid battery are improved by incorporating a dispersion of 1 to 10% by weight of a thermodynamically stable conductivity additive, such as conductive tin oxide coated glass fibers (34) of filamentary glass wool (42) in the positive active layer (32) carried on the grid (30) of the positive plate (16). The avoiding of positive plate reversal to prevent reduction of the tin oxide is accomplished by (a) employing an oversized positive plate and pre-charging it; (b) by pre-discharging the negative plate; and/or (c) by placing a circuit breaker (26) in combination with the plates (16, 18) and terminals (22, 24) to remove the load when the voltage of the positive plate falls below a pre-selected level.

  12. Construction of efficient and effective transformation vectors for palmitoyl-acyl carrier protein thioesterase gene silencing in oil palm

    PubMed Central

    Bhore, Subhash Janardhan; Shah, Farida Habib

    2011-01-01

    Palm oil obtained from E. guineensis Jacq. Tenera is known to have about 44% of palmitic acid (C16:0). Palmitoyl-Acyl Carrier Protein Thioesterase (PATE) is one of the key enzymes involved in plastidial fatty acid biosynthesis; and it determines the level of the C16:0 assimilation in oilseeds. This enzyme's activity in oil palm is responsible for high (> 44 % in E. guineensis Jacq. Tenera and 25 % in E. oleifera) content of C16:0 in its oil. By post-transcriptional PATE gene silencing, C16:0 content can be minimized for nutritional value improvement of the palm oil. The objective of this study was the construction of novel transformation vectors for PATE gene silencing. Six different transformation vectors targeted against PATE gene were constructed using 619 bp long PATE gene (5' region) fragment (from GenBank AF507115). In one set of three transformation vectors, PATE gene fragment was fused with CaMV 35S promoter in antisense, intron-spliced inverted repeat (ISIR), and inverted repeat (IR) orientations to generate antisense mRNA and hair-pin RNAs (hpRNA). In another set of three transformation vectors with same design, CaMV 35S was replaced with Oil palm mesocarp tissue-specific promoter (MSP). The expression cassette of antisense, ISIR, and IR of PATE gene fragments were constructed in primary cloning vector, pHANNIBAL or its derivative/s. Finally, all 6 expression cassettes were sub-cloned into pCAMBIA 1301 which contains the Hygromycinr and the GUS reporter genes for transformant selection and transformation detection respectively. The results of the RE analyses of the constructs and sequence analyses of PATE and MSP shows and confirms the orientation, size and locations of all the components from constructs. We hypothesize that 4 (pISIRPATE-PC, pIRPATE-PC, pMISIRPATE-PC and pMIRPATE-PC) out of 6 transformation vectors constructed in this study will be efficient and effective in palmitoyl-ACP thioesterase gene silencing in oil palm. Abbreviations anti

  13. 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

  14. High yield of functional metagenomic library from mangroves constructed in fosmid vector.

    PubMed

    Gonçalves, A C S; dos Santos, A C F; dos Santos, T F; Pessoa, T B A; Dias, J C T; Rezende, R P

    2015-01-01

    In the present study, metagenomic technique and fosmid vectors were used to construct a library of clones for exploring the biotechnological potential of mangrove soils by isolation of functional genes encoding hydrolytic enzymes. The library was built with genomic DNA from the soil samples of mangrove sediments and the functional screening of 1824 clones (~64 Mbp) was performed to detect the hydrolytic activity specific for cellulases, amylases (at acidic, neutral and basic pH), lipases/esterases, proteases, and nitrilases. Significant numbers of clones, positive for the tested enzyme activities were obtained. Our results indicate the importance and biotechnological potential of mangrove soils especially when compared to those obtained using other soil metagenomic libraries. PMID:26436508

  15. Expression vectors for the construction of hybrid Ty-VLPs.

    PubMed

    Adams, S E; Richardson, S M; Kingsman, S M; Kingsman, A J

    1994-04-01

    Purification of expressed proteins can be facilitated by expressing the recombinant protein as a fusion with a carrier protein that assembles into particulate structures. This article describes the use of expression vectors in producing a hybrid of the yeast retrotransposon Ty, which self-assembles into virus-like particles (VLPs). Hybrid VLPs can be used in such laboratory applications as the production of polyclonal and monoclonal antibodies, structure/function analyses, the detection of important antigenic determinants, and epitope mapping of monoclonal antibodies. PMID:7859156

  16. Prediction of protein subcellular localization by support vector machines using multi-scale energy and pseudo amino acid composition.

    PubMed

    Shi, J-Y; Zhang, S-W; Pan, Q; Cheng, Y-M; Xie, J

    2007-07-01

    As more and more genomes have been discovered in recent years, there is an urgent need to develop a reliable method to predict the subcellular localization for the explosion of newly found proteins. However, many well-known prediction methods based on amino acid composition have problems utilizing the sequence-order information. Here, based on the concept of Chou's pseudo amino acid composition (PseAA), a new feature extraction method, the multi-scale energy (MSE) approach, is introduced to incorporate the sequence-order information. First, a protein sequence was mapped to a digital signal using the amino acid index. Then, by wavelet transform, the mapped signal was broken down into several scales in which the energy factors were calculated and further formed into an MSE feature vector. Following this, combining this MSE feature vector with amino acid composition (AA), we constructed a series of MSEPseAA feature vectors to represent the protein subcellular localization sequences. Finally, according to a new kind of normalization approach, the MSEPseAA feature vectors were normalized to form the improved MSEPseAA vectors, named as IEPseAA. Using the technique of IEPseAA, C-support vector machine (C-SVM) and three multi-class SVMs strategies, quite promising results were obtained, indicating that MSE is quite effective in reflecting the sequence-order effects and might become a useful tool for predicting the other attributes of proteins as well. PMID:17235454

  17. A simple method for construction of artificial microRNA vector in plant.

    PubMed

    Li, Yang; Li, Yang; Zhao, Sunping; Zhong, Sheng; Wang, Zhaohai; Ding, Bo; Li, Yangsheng

    2014-10-01

    Artificial microRNA (amiRNA) is a powerful tool for silencing genes in many plant species. Here we provide an easy method to construct amiRNA vectors that reinvents the Golden Gate cloning approach and features a novel system called top speed amiRNA construction (TAC). This speedy approach accomplishes one restriction-ligation step in only 5 min, allowing easy and high-throughput vector construction. Three primers were annealed to be a specific adaptor, then digested and ligated on our novel vector pTAC. Importantly, this method allows the recombined amiRNA constructs to maintain the precursor of osa-miR528 with exception of the desired amiRNA/amiRNA* sequences. Using this method, our results showed the expected decrease of targeted genes in Nicotiana benthamiana and Oryza sativa. PMID:24930106

  18. Construction of filter vectors for the information-efficient spectral imaging sensor

    SciTech Connect

    Stallard, B.R.; Gentry, S.M.

    1998-12-01

    The information-efficient spectral imaging sensor (ISIS) seeks to improve system performance by processing hyperspectral information in the optical hardware. Its output may be a gray scale image in which one attempts to maximize the contrast between a given target and the background. Alternatively, its output may be a small number of images, rather than a full data cube, that retain the essential information required in the application. The principal advantage of ISIS is that it offers the discrimination of hyperspectral imaging while achieving the signal-to-noise ratio of multispectral imaging. The paper focuses on construction of the filter vectors that are needed to program ISIS. The instrument produces an image which is essentially a dot product of the scene and the filter vector. Both single vector and multiple vector approaches are considered. Also, they discuss some subtle points related to optimizing the filter vectors.

  19. DIY series of genetic cassettes useful in construction of versatile vectors specific for Alphaproteobacteria.

    PubMed

    Dziewit, Lukasz; Adamczuk, Marcin; Szuplewska, Magdalena; Bartosik, Dariusz

    2011-08-01

    We have developed a DIY (Do It Yourself) series of genetic cassettes, which facilitate construction of novel versatile vectors for Alphaproteobacteria. All the cassettes are based on defined genetic modules derived from three natural plasmids of Paracoccus aminophilus JCM 7686. We have constructed over 50 DIY cassettes, which differ in structure and specific features. All of them are functional in eight strains representing three orders of Alphaproteobacteria: Rhodobacterales, Rhizobiales and Caulobacterales. Besides various replication and stabilization systems, many of the cassettes also contain selective markers appropriate for Alphaproteobacteria (40 cassettes) and genetic modules responsible for mobilization for conjugal transfer (24 cassettes). All the DIY cassettes are bordered by different types of polylinkers, which facilitate vector construction. Using these DIY cassettes, we have created a set of compatible Escherichia coli-Alphaproteobacteria mobilizable shuttle vectors (high or low copy number in E. coli), which will greatly assist the genetic manipulation of Alphaproteobacteria. PMID:21569803

  20. Construction of Yeast Recombinant Expression Vector Containing Human Epidermal Growth Factor (hEGF)

    PubMed Central

    Mohammadian, Jamal; Mansoori-Derakhshan, Sima; Mohammadian, Masood; Shekari-Khaniani, Mahmoud

    2013-01-01

    Purpose: The objective of this study was construction of recombinant hEGF-pPIC9 which may be used for expression of recombinant hEGF in following studies. Methods: EGF cDNA was purchased from Genecopoeia Company and used for PCR amplification. Prior to ligation, the PCR product and pPIC9 vector was digested with EcoRI and XhoI and ligated in pPIC9 vector and subjected to colony PCR screening and sequencing analysis. Results: PCR amplification of EGF cDNA using recombinant hEGF-pPIC9 vector as template was concluded in amplification of 197bp fragment. Construction of recombinant hEGF-pPIC9 of EGf gene was verified by PCR and sequencing. Conclusion: Construction of Recombinant hEGF-pPIC9 was the primary stage for production and expression of EFG in the future study. PMID:24312882

  1. Construction of vectors for inducible and constitutive gene expression in Lactobacillus.

    PubMed

    Duong, Tri; Miller, Michael J; Barrangou, Rodolphe; Azcarate-Peril, M Andrea; Klaenhammer, Todd R

    2011-05-01

    Microarray analysis of the genome of Lactobacillus acidophilus identified a number of operons that were differentially expressed in response to carbohydrate source or constitutively expressed regardless of carbohydrate source. These included operons implicated in the transport and catabolism of fructooligosaccharides (FOS), lactose (lac), trehalose (tre) and genes directing glycolysis. Analysis of these operons identified a number of putative promoter and repressor elements, which were used to construct a series of expression vectors for use in lactobacilli, based on the broad host range pWV01 replicon. A β-glucuronidase (GusA3) reporter gene was cloned into each vector to characterize expression from each promoter. GUS reporter assays showed FOS, lac and tre based vectors to be highly inducible by their specific carbohydrate and repressed by glucose. Additionally, a construct based on the phosphoglycerate mutase (pgm) promoter was constitutively highly expressed. To demonstrate the potential utility of these vectors, we constructed a plasmid for the overexpression of the oxalate degradation pathway (Frc and Oxc) of L. acidophilus NCFM. This construct was able to improve oxalate degradation by L. gasseri ATCC 33323 and compliment a L. acidophilus oxalate-deficient mutant. Development of these expression vectors could support several novel applications, including the expression of enzymes, proteins, vaccines and biotherapeutics by intestinal lactobacilli. PMID:21375708

  2. [Construction and identification of mammary expressional vector for cDNA of human lactoferrin].

    PubMed

    Meng, Li; Zhang, Yanli; Xu, Xin; Wang, Ziyu; Yan, Yibo; Pang, Xunsheng; Zhong, Bushuai; Huang, Rong; Song, Yang; Wang, Jinyu; Wang, Feng

    2011-02-01

    The aim of this study was to construct a mammary gland-specific expressional vector pBC1-hLF-Neo for Human Lactoferrin (hLF) gene and then investigate its expression in the mammary gland epithelium cells. The constructed vector contained the 6.2 kb long 5' flank regulation region including promoter, other elements and the 7.1 kb long 3' flank regulation region including transcriptional ending signal of a goat's beta-casein gene. A cassette of Neo gene was also inserted into the vector which gave a total length of 26.736 kb identified by restriction fragment analysis and partial DNA sequencing. The results revealed that the structure of the final constructed vector accords with the designed plasmid map. In order to analyze the bioactivity of the vector, we transfected the lined vector DNA into the dairy goat's mammary gland epithelium cells and C127 cells of a mouse's mammary epithelium by Lipofectamine. After selection with G418 for 8-10 days, G418-risistant clones were obtained. PCR analysis demonstrated that hLF gene cassette had been integrated into the genomic DNA of G418-risistant clones. After proliferation culture, the two kinds of transgenic cells were cultured in serum-free DMEM-F12 medium with prolactin, insulin and hydrocortisone- a medium capable of inducing recombinant hLF expression. RT-PCR, Western blotting and anti-bacteria bioactivity experiments demonstrated that the constructed mammary gland specific vector pBC1-hLF-Neo possessed the desirable bioactivity to efficiently express and could secrete hLF in both mammary gland cells and have the effect of E. coli proliferation inhibition. Paramount to everything, this study laid a firm foundation for preparing the hLF gene transgenic goat fetal-derived fibroblast cells. PMID:21650051

  3. A Universal Vector for High-Efficiency Multi-Fragment Recombineering of BACs and Knock-In Constructs

    PubMed Central

    Miller-Hodges, Eve; Slight, Joan; Thornburn, Anna; Devenney, Paul S.; Hohenstein, Peter

    2013-01-01

    There is an increasing need for more efficient generation of transgenic constructs. Here we present a universal multi-site Gateway vector for use in recombineering reactions. Using transgenic mouse models, we show its use for the generation of BAC transgenics and targeting vectors. The modular nature of the vector allows for rapid modification of constructs to generate different versions of the same construct. As such it will help streamline the generation of series of related transgenic models. PMID:23637962

  4. Construction and characterization of a recombinant human adenovirus vector expressing bone morphogenetic protein 2.

    PubMed

    Zhang, Zheng; Wang, Guoxian; Li, Chen; Liu, Danping

    2013-08-01

    The aim of this study was to construct and characterize a novel recombinant human adenovirus vector expressing bone morphogenetic protein 2 (BMP2) and green fluorescent protein (GFP). The BMP2 gene in the plasmid pcDNA3-BMP2 was sequenced and the restriction enzyme recognition sites were analyzed. Following mutagenesis using polymerase chain reaction (PCR), the gene sequence after the translation termination codon was removed and new restriction sites were added. The mutated BMP2 gene (BMP2(+) gene) was cloned into an adenovirus shuttle vector to obtain pShuttle cytomegalovirus (CMV)-BMP2(+)-internal ribosome entry site (IRES)-hrGFP-1. The adenovirus plasmid pAd CMV-BMP2(+)-IRES-hrGFP-1 was constructed by homologous recombination and was transfected into HEK293A cells, followed by adenovirus packaging. pAd CMV-BMP2 was used as the control. The two types of adenovirus were transfected into marrow stromal cells (MSCs). The expression of BMP2 and GFP, as well as the alkaline phosphatase (ALP) activity of expressed BMP2 were detected. Following mutagenesis, the BMP2 gene sequence and recombinant adenovirus vector were as predicted. The novel adenovirus vector expressed both BMP2 and GFP, indicating that a novel recombinant human adenovirus vector expressing BMP2 had been successfully constructed. PMID:24137184

  5. Plasmids for Increased Efficiency of Vector Construction and Genetic Engineering in Filamentous Fungi

    PubMed Central

    Schoberle, Taylor J.; Nguyen-Coleman, C. Kim; May, Gregory S.

    2013-01-01

    Fungal species are continuously being studied to not only understand disease in humans and plants but also to identify novel antibiotics and other metabolites of industrial importance. Genetic manipulations, such as gene deletion, gene complementation, and gene over-expression, are common techniques to investigate fungal gene functions. Although advances in transformation efficiency and promoter usage have improved genetic studies, some basic steps in vector construction are still laborious and time-consuming. Gateway cloning technology solves this problem by increasing the efficiency of vector construction through the use of λ phage integrase proteins and att recombination sites. We developed a series of Gateway-compatible vectors for use in genetic studies in a range of fungal species. They contain nutritional and drug-resistance markers and can be utilized to manipulate different filamentous fungal genomes. PMID:23867711

  6. Efficient construction of recombinant adenovirus expression vector of the Qinchuan cattle LYRM1 gene.

    PubMed

    Li, Y K; Fu, C Z; Zhang, Y R; Zan, L S

    2015-01-01

    In this study, we cloned the coding DNA sequence (CDS) region of Qinchuan cattle LYR motif-containing 1 (LYRM1) and constructed a recombinant adenovirus expression vector to examine the function of LYRM1 on the cellular level. Total RNA was extracted from the adipose tissue of Qinchuan cattle, cDNA was obtained by reverse transcription, and polymerase chain reaction was used to amplify the CDS region of the LYRM1 gene. The CDS-containing fragment was inserted into the shuttle vector pAdTrack-CMV to construct pAdTrack-CMV-LYRM1 vector. After linearization of pAdTrack-CMV-LYRM1 and the negative control vector pAdTrack-CMV by restriction endonuclease PmeI, the vectors were transformed into Escherichia coli BJ5183 containing pAdEasy-1 to obtain the recombinant adenovirus vector pAd-LYRM1 and pAd-CMV through homologous recombination. pAd-LYRM1 and pAd-CMV were then digested by PacI and transfected into the 293A cell line. The recombinant adenovirus Ad-LYRM1 and Ad-CMV was obtained at a concentration of 7 x 108 and 1.3 x 109 green fluorescent units/mL, respectively. Preadipocytes derived from Qinchuan cattle were separately infected with Ad-LYRM1 and Ad- CMV. Quantitative real-time polymerase chain reaction demonstrated that the expression of LYRM1 was increased by approximate 28,000-folds after the infection with recombinant adenovirus for 48 h. In conclusion, we successfully cloned the CDS region of the Qinchuan cattle LYRM1 gene, constructed the recombinant adenovirus expression vector, and obtained the adenovirus with high titer, providing valuable materials for studying the function of LYRM1 at the cellular level. PMID:26345880

  7. Construction and application of the vectors to identify genes encoding exported proteins of Escherichia coli.

    PubMed

    Niu, Dong; Shen, Qinfang; Zhu, Junli; Liu, Jiangmei; Yuan, Jiajie; Tan, Shuang; Yu, Xuping

    2013-10-01

    In order to clone genes having signal sequences of Escherichia coli, four vectors with or without Lac or Ara promoter were constructed using a leaderless β-lactamase as reporter. Fragments of tetracycline resistance gene (Tet) with or without promoter were used to confirm the vectors' ability to clone and report signal sequences. The minimum inhibitory concentration of ampicillin of the transformants was measured to detect the expression and secretion efficiency of the vectors. The results showed that the β-lactamase could be co-expressed and secreted with Tet protein. The Lac or Ara promoter in the vectors could be regulated by different inducers, and the Ara promoter showed higher regulative efficiency than the Lac. The best induction dose of L-arabinose for the Ara promoter is 1.25 %. All the four vectors were stably maintained in host after being inoculated for 20 passages in antibiotics-free media. Genomic library of an avian pathogenic strain, E. coli O2, was constructed using the pMB-Ara-T vector we developed. 318 clones were obtained from the genomic library of E. coli strain O2, and the inserts in these clones represented 276 genes based on sequence analysis. Among the 276 cloned fragments, only 128 had complete promoter sequence. For the 128 fragments with promoter, only 27 could be expressed under LB culture condition without inducer, the other 101 were only expressed under induction. The results showed our constructed vectors could efficiently capture all kinds of exported protein genes in vitro, including the ones without promoter or with inactive promoter. PMID:24052231

  8. Semilinear degenerate evolution inequalities with singular potential constructed from the generalized Greiner vector fields

    NASA Astrophysics Data System (ADS)

    Yuan, Zixia; Niu, Pengcheng

    2007-03-01

    We study the existence and nonexistence of global solutions to the degenerate evolution inequalities with singular potential constructed from the generalized Greiner vector fields. For the proof of the existence results, we use the method of supersolution and the modified Bessel function. The nonexistence results are established by the test function method.

  9. Yeast recombination-based cloning as an efficient way of constructing vectors for Zymoseptoria tritici.

    PubMed

    Kilaru, S; Steinberg, G

    2015-06-01

    Many pathogenic fungi are genetically tractable. Analysis of their cellular organization and invasion mechanisms underpinning virulence determinants profits from exploiting such molecular tools as fluorescent fusion proteins or conditional mutant protein alleles. Generation of these tools requires efficient cloning methods, as vector construction is often a rate-limiting step. Here, we introduce an efficient yeast recombination-based cloning (YRBC) method to construct vectors for the fungus Zymoseptoria tritici. This method is of low cost and avoids dependency on the availability of restriction enzyme sites in the DNA sequence, as needed in more conventional restriction/ligation-based cloning procedures. Furthermore, YRBC avoids modification of the DNA of interest, indeed this potential risk limits the use of site-specific recombination systems, such as Gateway cloning. Instead, in YRBC, multiple DNA fragments, with 30bp overlap sequences, are transformed into Saccharomyces cerevisiae, whereupon homologous recombination generates the vector in a single step. Here, we provide a detailed experimental protocol and four vectors, each encoding a different dominant selectable marker cassette, that enable YRBC of constructs to be used in the wheat pathogen Z. tritici. PMID:26092792

  10. Yeast recombination-based cloning as an efficient way of constructing vectors for Zymoseptoria tritici

    PubMed Central

    Kilaru, S.; Steinberg, G.

    2015-01-01

    Many pathogenic fungi are genetically tractable. Analysis of their cellular organization and invasion mechanisms underpinning virulence determinants profits from exploiting such molecular tools as fluorescent fusion proteins or conditional mutant protein alleles. Generation of these tools requires efficient cloning methods, as vector construction is often a rate-limiting step. Here, we introduce an efficient yeast recombination-based cloning (YRBC) method to construct vectors for the fungus Zymoseptoria tritici. This method is of low cost and avoids dependency on the availability of restriction enzyme sites in the DNA sequence, as needed in more conventional restriction/ligation-based cloning procedures. Furthermore, YRBC avoids modification of the DNA of interest, indeed this potential risk limits the use of site-specific recombination systems, such as Gateway cloning. Instead, in YRBC, multiple DNA fragments, with 30 bp overlap sequences, are transformed into Saccharomyces cerevisiae, whereupon homologous recombination generates the vector in a single step. Here, we provide a detailed experimental protocol and four vectors, each encoding a different dominant selectable marker cassette, that enable YRBC of constructs to be used in the wheat pathogen Z. tritici. PMID:26092792

  11. Construction and experimental application of a highly efficient temperature-selection T-vector.

    PubMed

    Ma, Yue; Li, Anxing; Wang, Yongcai; Xie, Mingquan; Li, Jing; Zhang, Sen; Wang, Qiuju

    2009-08-01

    T-A cloning is the most commonly used molecular cloning technique in which T-vector is the core tool. T-vectors commonly used now are blue-white positive-selection vectors based on mechanism of alpha-complement of beta-galactose of Escherichia coli. Disadvantages of these traditional T-vectors in application include insufficient positive rate and the existence of false-positive clones, which, especially the latter, makes screening of transformant clones via colony PCR necessary. In the present study, a novel T-vector, based on the strategy of inhibiting the growth of negative transformants by means of a lethal gene and innovative design of insertion site, was developed. The innovative design of the insertion site for PCR fragments not only guarantees complete blocking of the expression of the lethal gene when PCR fragments are inserted, but also eliminates the formation of false-positive clones. All (100%) of 20 randomly chosen grown colonies were shown to be positive via colony PCR in four separate experimental T-A clonings of PCR fragments with lengths between 219 and 2100 bp. The novel T-vector developed in the present study makes colony PCR screening unnecessary, because of its extremely high efficiency of negative transformant inhibition and the mechanism for elimination of false-positive clones. Temperature (42 degrees C)-dependent positive selection greatly simplifies the procedure and lowers the cost of T-A cloning. Furthermore, it is an ideal T-vector for constructing libraries of PCR-amplified DNA fragments such as SSH (suppressive subtraction hybridization) libraries because of its zero negative background performance. PMID:19327129

  12. Alternative methods for the efficient construction of short hairpin RNA expression vectors.

    PubMed

    Xu, Kun; Zhang, Tingting; Guo, Lijun; Xin, Ying; Zhang, Long; Zhang, Zhiying

    2015-06-01

    Short hairpin RNA (shRNA)-mediated RNA interference has become a basic technique in modern molecular biology and biochemistry for studying gene function and biological pathways. Here, we report two alternative and efficient methods to construct shRNA expression vectors based respectively on multiple-step sequential PCR and primer extension-homologous recombination (PE-HR). Neither method requires synthesizing long oligonucleotides containing hairpin sequences as used in traditional approaches. The hairpin sequences may produce mutations during oligo synthesis, pose problems in annealing, and lead to inefficient cloning. The PE-HR method further provides rapid and economical construction of shRNA expression vectors without needing the ligation procedure. PMID:25794926

  13. Construction of a bovine enterovirus-based vector expressing a foot-and-mouth disease virus epitope.

    PubMed

    Chu, Jia-Qi; Lee, Yeo-Joo; Park, Jeong-Nam; Kim, Su-Mi; Lee, Kwang-Nyeong; Ko, Young-Joon; Lee, Hyang-Sim; Cho, In-Soo; Kim, Byounghan; Park, Jong-Hyeon

    2013-04-01

    A recombinant infectious bovine enterovirus (BEV) vector was constructed to express a foot-and-mouth disease virus (FMDV) capsid protein (VP1) epitope. Sequences encoding the VP1 epitope (amino acid residues 141-160) of FMDV (vaccine strain O1/Manisa/Turkey/69) were inserted into pBLUBEV at the VP1/2A junction. The growth characteristics of the parental virus and viruses derived from recombinant plasmids (pBLUBEV, pBLUBEV-Manisa-epi) were determined by plaque assay and one-step growth curve analysis. There were no significant differences in the growth kinetics and plaque morphologies between transfectant viruses and their parental virus. The expressed VP1 epitope was detected successfully by using indirect immunofluorescence assay with a polyclonal antibody against the FMDV VP1 epitope from Madin Darby bovine kidney (MDBK) cells infected with BEV-Manisa-epi transfectant virus. This study demonstrated a novel alternative live viral vector that may be utilized as a candidate vaccine vector for veterinary applications. PMID:23391822

  14. Modular construction of plasmids through ligation-free assembly of vector components with oligonucleotide linkers.

    PubMed

    Vroom, Jonathan A; Wang, Clifford L

    2008-06-01

    We have developed a modular method of plasmid construction that can join multiple DNA components in a single reaction. A nicking enzyme is used to create 5' and 3' overhangs on PCR-generated DNA components. Without the use of ligase or restriction enzymes, components are joined using oligonucleotide linkers that recognize the overhangs. By specifying the sequences of the linkers, desired components can be assembled in any combination and order to generate different plasmid vectors. PMID:18533903

  15. Subcloning plus insertion (SPI)--a novel recombineering method for the rapid construction of gene targeting vectors.

    PubMed

    Reddy, Thimma R; Kelsall, Emma J; Fevat, Léna M S; Munson, Sarah E; Cowley, Shaun M

    2015-01-01

    Gene targeting refers to the precise modification of a genetic locus using homologous recombination. The generation of novel cell lines and transgenic mouse models using this method necessitates the construction of a 'targeting' vector, which contains homologous DNA sequences to the target gene, and has for many years been a limiting step in the process. Vector construction can be performed in vivo in Escherichia coli cells using homologous recombination mediated by phage recombinases using a technique termed recombineering. Recombineering is the preferred technique to subclone the long homology sequences (>4 kb) and various targeting elements including selection markers that are required to mediate efficient allelic exchange between a targeting vector and its cognate genomic locus. Typical recombineering protocols follow an iterative scheme of step-wise integration of the targeting elements and require intermediate purification and transformation steps. Here, we present a novel recombineering methodology of vector assembly using a multiplex approach. Plasmid gap repair is performed by the simultaneous capture of genomic sequence from mouse Bacterial Artificial Chromosome libraries and the insertion of dual bacterial and mammalian selection markers. This subcloning plus insertion method is highly efficient and yields a majority of correct recombinants. We present data for the construction of different types of conditional gene knockout, or knock-in, vectors and BAC reporter vectors that have been constructed using this method. SPI vector construction greatly extends the repertoire of the recombineering toolbox and provides a simple, rapid and cost-effective method of constructing these highly complex vectors. PMID:25590226

  16. [Construction of recombinant yellow fever virus 17D containing 2A fragment as a vaccine vector].

    PubMed

    Xiaowu, Pang; Fu, Wen-Chuan; Guo, Yin-Han; Zhang, Li-Shu; Xie, Tian-Pei; Xinbin, Gu

    2006-05-01

    The Yellow Fever (YF) vaccine, an attenuated yellow fever 17D (YF-17D) live vaccine, is one of the most effective and safest vaccines in the world and is regarded as one of the best candidates for viral expression vector. We here first reported in China the construction and characterization of the recombinant expression vector of yellow fever 17D which contained the proteinase 2A fragment of foot-and-mouth disease virus (FMDV). Three cDNA fragments representing the full-length YF-17D genome, named 5'-end cDNA (A), 3'-end cDNA (B) and middle cDNA (C), were obtained by reverse transcription polymerase chain reaction (RT-PCR), together with the introduction of SP6 enhancer, necessary restriction sites and overlaps for homologous recombination in yeast. Fragment A and B were then introduced into pRS424 in turn by DNA recombination, followed by transfection of fragment C and the recombinant pRS424 containing A and B (pRS-A-B) into yeast. A recombinant vector containing full length cDNA of YF-17D (pRS-YF) was obtained by screening on medium lack of tryptophan and uracil. A recombinant YF-17D expression vector containing FMDV-2A gene fragment (pRS-YF-2A1) was then constructed by methods of DNA recombination and homologous recombination in yeast described above. In vitro transcription of the recombinant vector pRS-YF-2A1 was then carried out and introduced into BHK-21 cells by electroporation. Results of indirect immunofluorescence assay (IFA) and titer determination showed a stable infectious recombinant virus was gotten, whose features such as growth curve were similar to those of the parental YF-17D. The results suggest that the recombinant vector pRS-YF-2A1, by introduction of heterogenous genes via 2A region, is potential to be an effective live vaccine expression vector. PMID:16755933

  17. Construction and Validation of a Dual-Transgene Vector System for Stable Transformation in Plants.

    PubMed

    He, Zhimin; Liu, Bin; Wang, Xu; Bian, Mingdi; He, Reqing; Yan, Jindong; Zhong, Ming; Zhao, Xiaoying; Liu, Xuanming

    2016-04-20

    In this study, we constructed dual-transgene vectors (pDT1, pDT7, and pDT7G) that simultaneously co-expressed two genes in plants. ACTIN2 and UBQ10 promoters were used to control the expression of these two genes. The 4×Myc, 3×HA, and 3×Flag reporter genes allowed for the convenient identification of a tunable co-expression system in plants, whereas the dexamethasone (Dex) inducible reporter gene C-terminus of the glucocorticoid receptor (cGR) provided Dex-dependent translocation of the fusion gene between the nucleus and cytoplasm. The function of pDT vectors was validated using four pairwise genes in Nicotiana benthamiana or Arabidopsis thaliana. The co-expression efficiency of two genes from the pDT1 and pDT7G vectors was 35% and 42%, respectively, which ensured the generation of sufficient transgenic materials. These pDT vectors are simple, reliable, efficient, and time-saving tools for the co-expression of two genes through a single transformation event and can be used in the study of protein-protein interactions or multi-component complexes. PMID:27157807

  18. Vectorization of Nucleic Acids for Therapeutic Approach: Tutorial Review.

    PubMed

    Geinguenaud, Frederic; Guenin, Erwann; Lalatonne, Yoann; Motte, Laurence

    2016-05-20

    Oligonucleotides present a high therapeutic potential for a wide variety of diseases. However, their clinical development is limited by their degradation by nucleases and their poor blood circulation time. Depending on the administration mode and the cellular target, these macromolecules will have to cross the vascular endothelium, to diffuse through the extracellular matrix, to be transported through the cell membrane, and finally to reach the cytoplasm. To overcome these physiological barriers, many strategies have been developed. Here, we review different methods of DNA vectorization, discuss limitations and advantages of the various vectors, and provide new perspectives for future development. PMID:26950048

  19. Construction of an adenovirus type 7a E1A- vector.

    PubMed Central

    Abrahamsen, K; Kong, H L; Mastrangeli, A; Brough, D; Lizonova, A; Crystal, R G; Falck-Pedersen, E

    1997-01-01

    A strategy for constructing replication-defective adenovirus vectors from non-subgroup C viruses has been successfully demonstrated with adenovirus type 7 strain a (Ad7a) as the prototype. An E1A-deleted Ad7a reporter virus expressing the chloramphenicol acetyltransferase (CAT) gene from the cytomegalovirus promoter enhancer was constructed with DNA fragments isolated from Ad7a, an Ad7a recombination reporter plasmid, and the 293 cell line. The Ad7a-CAT virus particle transduces A549 cells as efficiently as Ad5-based vectors. Intravenous infections in a murine model indicate that the Ad7a-CAT virus infects a variety of tissues, with maximal levels of CAT gene expression found in the liver. The duration of Ad7a-CAT transgene expression in the liver was maximally maintained 2 weeks postinfection, with a decline to baseline activity by the week 4 postinfection. Ad7a-CAT represents the first example of a non-subgroup C E1A- adenovirus gene transfer vector. PMID:9343264

  20. Construction of a 3He magnetic force microscope with a vector magnet

    NASA Astrophysics Data System (ADS)

    Yang, Jinho; Yang, Ilkyu; Kim, Yun Won; Shin, Dongwoo; Jeong, Juyoung; Wulferding, Dirk; Yeom, Han Woong; Kim, Jeehoon

    2016-02-01

    We constructed a 3He magnetic force microscope operating at the base temperature of 300 mK under a vector magnetic field of 2-2-9 T in the x-y-z direction. Fiber optic interferometry as a detection scheme is employed in which two home-built fiber walkers are used for the alignment between the cantilever and the optical fiber. The noise level of the laser interferometer is close to its thermodynamic limit. The capabilities of the sub-Kelvin and vector field are demonstrated by imaging the coexistence of magnetism and superconductivity in a ferromagnetic superconductor (ErNi2B2C) at T = 500 mK and by probing a dipole shape of a single Abrikosov vortex with an in-plane tip magnetization.

  1. Construction of Actinobacillus pleuropneumoniae-Escherichia coli shuttle vectors: expression of antibiotic-resistance genes.

    PubMed

    West, S E; Romero, M J; Regassa, L B; Zielinski, N A; Welch, R A

    1995-07-01

    We constructed several cloning vectors, designated pGZRS-18/19 and pGZRS-38/39, which were based on an endogenous Actinobacillus pleuropneumoniae (Apl) 4.3-kb plasmid. They carry the lacZ alpha-complementation fragment and MCS from pUC18/19, and either the bla gene under the control of a putative Apl promoter or the KmR gene from Tn903. These vectors replicate in representative strains of Apl serotypes 1 and 7, Escherichia coli, Pasteurella haemolytica (Ph) and Haemophilus (Actinobacillus) actinomycetemcomitans. We also found that Apl and Ph did not express genes under the control of the lacZ or bla promoters, suggesting that their RNA polymerases may not utilize these promoters. PMID:7628722

  2. [Construction and function identification of luciferase reporter gene vectors containing SNPs in NFKBIA gene 3'UTR].

    PubMed

    Yang, Shuo; Li, Jia-li; Bi, Hui-chang; Zhou, Shou-ning; Liu, Xiao-man; Zeng, Hang; Hu, Bing-fang; Huang, Min

    2016-01-01

    This study aims to investigate the function of two SNPs (rs8904C > T and rs696G >A) in 3' untranslated region (3'UTR) of NFKBIA gene by constructing luciferase reporter gene. A patient's genomic DNA with rs8904 CC and rs696 GA genotype was used as the PCR template. Full-length 3'UTR of NFKBIA gene was amplified by different primers. After sequencing validation, these fragments were inserted to the luciferase reporter vector, pGL3-promoter to construct recombinant plasmids containing four kinds of haplotypes, pGL3-rs8904C/rs696G, pGL3-rs8904C/rs696A, pGL3-rs8904T/rs696G and pGL3-rs8904T/rs696A. Then these plasmids were transfected into LS174T cells and the luciferase activity was detected. Compared with pGL3-vector transfected cells (negative control), the luciferase activity of the four kinds of recombinant plasmids was significantly decreased (P < 0.001). For rs696G > A, the luciferase activity of the recombinant plasmids containing A allele (pGL3-rs8904C/rs696A and pGL3-rs8904T/rs696A) was about 45.1% (P < 0.05) and 56.1% (P < 0.001) lower than those containing G allele (pGL3-rs8904C/rs696G and pGL3-rs8904T/rs696G), respectively. For rs8904C > T, there were no significant differences in the luciferase activity between the recombinant plasmids containing T allele and those with C allele. Together, the luciferase reporter gene vectors containing SNPs in NFKBIA gene 3'UTR were constructed successfully and rs696G > A could decrease the luciferase activity while rs8904C >T didn't have much effect on the luciferase activity. PMID:27405166

  3. HANDBOOK FOR CONSTRUCTED WETLANDS RECEIVING ACID MINE DRAINAGE

    EPA Science Inventory

    In the summer of 1987, a pilot constructed wetland was built at the Big Five Tunnel in Idaho Springs, Colorado. This report details the theory, design and construction of wetlands receiving acid mine drainages, based on the second and third year of operation of this wetland, whic...

  4. HANDBOOK FOR CONSTRUCTED WETLANDS RECEIVING ACID MINE DRAINAGE

    EPA Science Inventory

    In the summer of 1987, a pilot constructed wetland was built at the Big Five Tunnel in Idaho Springs, Colorado. his report details the theory, design and construction of wetlands receiving acid mine drainages, based on the second and third year of operation of this wetland, which...

  5. Construction of novel shuttle expression vectors for gene expression in Bacillus subtilis and Bacillus pumilus.

    PubMed

    Shao, Huanhuan; Cao, Qinghua; Zhao, Hongyan; Tan, Xuemei; Feng, Hong

    2015-01-01

    A native plasmid (pSU01) was detected by genome sequencing of Bacillus subtilis strain S1-4. Two pSU01-based shuttle expression vectors pSU02-AP and pSU03-AP were constructed enabling stable replication in B. subtilis WB600. These vectors contained the reporter gene aprE, encoding an alkaline protease from Bacillus pumilus BA06. The expression vector pSU03-AP only possessed the minimal replication elements (rep, SSO, DSO) and exhibited more stability on structure, suggesting that the rest of the genes in pSU01 (ORF1, ORF2, mob, hsp) were unessential for the structural stability of plasmid in B. subtilis. In addition, recombinant production of the alkaline protease was achieved more efficiently with pSU03-AP whose copy number was estimated to be more than 100 per chromosome. Furthermore, pSU03-AP could also be used to transform and replicate in B. pumilus BA06 under selective pressure. In conclusion, pSU03-AP is expected to be a useful tool for gene expression in Bacillus subtilis and B. pumilus. PMID:26377132

  6. A new procedure for constructing basis vectors of SU(3)⊃SO(3)

    NASA Astrophysics Data System (ADS)

    Pan, Feng; Yuan, Shuli; Launey, Kristina D.; Draayer, Jerry P.

    2016-08-01

    A simple and effective algebraic angular momentum projection procedure for constructing basis vectors of SU (3) ⊃ SO (3) ⊃ SO (2) from the canonical U (3) ⊃ U (2) ⊃ U (1) basis vectors is outlined. The expansion coefficients are components of the null-space vectors of a projection matrix with, in general, four nonzero elements in each row, where the projection matrix is derived from known matrix elements of the U (3) generators in the canonical basis. The advantage of the new procedure lies in the fact that the Hill-Wheeler integral involved in the Elliott's projection operator method used previously is avoided, thereby achieving faster numerical calculations with improved accuracy. Selected analytical expressions of the expansion coefficients for the SU (3) irreps [n13 ,n23 ], or equally, (λ , μ) = (n13 -n23 ,n23) with λ and μ the SU (3) labels familiar from the Elliott model, are presented as examples for n23 ≤ 4. Explicit formulae for evaluating SO (3)-reduced matrix elements of SU (3) generators are derived. A general formula for evaluating the SU (3) ⊃ SO (3) Wigner coefficients is given, which is expressed in terms of the expansion coefficients and known U (3) ⊃ U (2) and U (2) ⊃ U (1) Wigner coefficients. Formulae for evaluating the elementary Wigner coefficients of SU (3) ⊃ SO (3), i.e., for the SU (3) coupling [n13 ,n23 ] ⊗ [ 1 , 0 ], are explicitly given with some analytical examples shown to check the validity of the results. However, the Gram-Schmidt orthonormalization is still needed in order to provide orthonormalized basis vectors.

  7. Prediction of water quality index in constructed wetlands using support vector machine.

    PubMed

    Mohammadpour, Reza; Shaharuddin, Syafiq; Chang, Chun Kiat; Zakaria, Nor Azazi; Ab Ghani, Aminuddin; Chan, Ngai Weng

    2015-04-01

    Poor water quality is a serious problem in the world which threatens human health, ecosystems, and plant/animal life. Prediction of surface water quality is a main concern in water resource and environmental systems. In this research, the support vector machine and two methods of artificial neural networks (ANNs), namely feed forward back propagation (FFBP) and radial basis function (RBF), were used to predict the water quality index (WQI) in a free constructed wetland. Seventeen points of the wetland were monitored twice a month over a period of 14 months, and an extensive dataset was collected for 11 water quality variables. A detailed comparison of the overall performance showed that prediction of the support vector machine (SVM) model with coefficient of correlation (R(2)) = 0.9984 and mean absolute error (MAE) = 0.0052 was either better or comparable with neural networks. This research highlights that the SVM and FFBP can be successfully employed for the prediction of water quality in a free surface constructed wetland environment. These methods simplify the calculation of the WQI and reduce substantial efforts and time by optimizing the computations. PMID:25408070

  8. Optimized Transformation of Newly Constructed Escherichia coli-Clostridia Shuttle Vectors into Clostridium beijerinckii.

    PubMed

    Oh, Young Hoon; Eom, Gyeong Tae; Kang, Kyoung Hee; Choi, Jae Woo; Song, Bong Keun; Lee, Seung Hwan; Park, Si Jae

    2015-09-01

    Three Escherichia coli-Clostridia shuttle vectors, pKBA411-MCS, pKBE411-MCS, and pKBM411-MCS, which contain p15A, ColE1, and pMB1 origins for replication in E. coli, respectively, along with the pAMB origin for replication in C. beijerinckii, were constructed and examined for their transformation efficiencies into Clostridium beijerinckii NCIMB8052. The transformation condition of pKBM411-MCS, which was optimized by varying resistance, buffer composition, and DNA concentration, was further employed for the transformation of the other plasmids, pKBA411-MCS and pKBE411-MCS into C. beijerinckii. It was found out that transformation efficiency is highly dependent on the origin of replication. The highest transformation efficiency of 7.44 × 10(3) colony-forming units per microgram of DNA was obtained at 5.0 kV cm(-1) field strength, 200 Ω resistance, 270 mM sucrose concentration, 150 ng μg(-1), and 3.0 μg DNA using pKBM411-MCS having pMB1 and pAMB origins of replication. The application of the newly constructed vector system was also investigated by introducing the putative alcohol dehydrogenase gene of C. beijerinckii. PMID:26152821

  9. Construction of small plasmid vectors for use in genetic improvement of the extremely acidophilic Acidithiobacillus caldus.

    PubMed

    Meng, Jianzhou; Wang, Huiyan; Liu, Xiangmei; Lin, Jianqun; Pang, Xin; Lin, Jianqiang

    2013-10-01

    The genetic improvement of biomining bacteria including Acidithiobacillus caldus could facilitate the bioleaching process of sulfur-containing minerals. However, the available vectors for use in A. caldus are very scanty and limited to relatively large broad-host-range IncQ plasmids. In this study, a set of small, mobilizable plasmid vectors (pBBR1MCS-6, pMSD1 and pMSD2) were constructed based on plasmid pBBR1MCS-2, which does not belong to the IncQ, IncW, or IncP groups. The function of the tac promoter on 5.8-kb pMSD2 was determined by inserting a kanamycin-resistant reporter gene. The resulting recombinant pMSD2-Km was successfully transferred by conjugation into A. caldus MTH-04 with transfer frequency of 1.38±0.64×10(-5). The stability and plasmid copy number of pMSD2-Km in A. caldus MTH-04 were 75±2.7% and 5-6 copies per cell, respectively. By inserting an arsABC operon into pMSD2, an arsenic-resistant recombinant pMSD2-As was constructed and transferred into A. caldus MTH-04 by conjugation. The arsenic tolerance of A. caldus MTH-04 containing pMSD2-As was obviously increased up to 45mM of NaAsO2. These vectors could be applied in genetic improvement of A. caldus as well as other bioleaching bacteria. PMID:23639949

  10. Construction and evaluation of a plasmid vector for the expression of recombinant lipoproteins in Escherichia coli.

    PubMed

    Cullen, Paul A; Lo, Miranda; Bulach, Dieter M; Cordwell, Stuart J; Adler, Ben

    2003-01-01

    Outer membrane lipoproteins are emerging as key targets for protective immunity to many bacterial pathogens. Heterologous expression of lipoproteins in Escherichia coli does not always result in high level expression of acylated recombinant protein. Thus, these proteins do not take up their correct membrane topology and are lacking the immunostimulatory properties endowed by the lipid. To this end, we have designed a lipoprotein expression vector (pDUMP) that results in the production of fusion proteins containing the E. coli major outer membrane lipoprotein (Lpp) signal sequence, lipoprotein signal peptidase recognition site, and the +2 outer membrane sorting signal at their N termini. To test the ability of pDUMP to express lipoproteins from heterologous hosts, the surface lipoprotein PsaA from the Gram-positive organism Streptococcus pneumoniae and the outer membrane lipoproteins MlpA from the Gram-negative Pasteurella multocida and BlpA from the spirochete Brachyspira hyodysenteriae were cloned into both hexahistidine fusion vectors and pDUMP. High level expression of antigenically active protein from both the hexahistidine fusion vectors and pDUMP resulted in abundant bands of the predicted molecular masses when analyzed by SDS-PAGE. When grown in the presence of 3[H]palmitic acid, proteins encoded by pDUMP were observed to incorporate palmitic acid whilst the hexahistidine fusion proteins did not. Using mass spectrometry and image analysis we determined the efficiency of lipidation between the three clones to vary from 31.7 to 100%. In addition, lipidated, but not hexahistidine, forms of the proteins were presented on the E. coli surface. PMID:12583997

  11. A marker-free system for highly efficient construction of vaccinia virus vectors using CRISPR Cas9

    PubMed Central

    Yuan, Ming; Gao, Xuefei; Chard, Louisa S; Ali, Zarah; Ahmed, Jahangir; Li, Yunqing; Liu, Pentao; Lemoine, Nick R; Wang, Yaohe

    2015-01-01

    The current method for creation of vaccinia virus (VACV) vectors involves using a selection and purification marker, however inclusion of a gene without therapeutic value in the resulting vector is not desirable for clinical use. The Cre-LoxP system has been used to make marker-free Poxviruses, but the efficiency was very low. To obtain a marker-free VACV vector, we developed marker gene excision systems to modify the thymidine kinase (TK) region and N1L regions using Cre-Loxp and Flp-FRET systems respectively. CRISPR-Cas9 system significantly resulted in a high efficiency (~90%) in generation of marker gene-positive TK-mutant VACV vector. The marker gene (RFP) could be excised from the recombinant virus using Cre recombinase. To make a marker-free VV vector with double gene deletions targeting the TK and N1L gene, we constructed a donor repair vector targeting the N1L gene, which can carry a therapeutic gene and the marker (RFP) that could be excised from the recombinant virus using Flp recombinase. The marker-free system developed here can be used to efficiently construct VACV vectors armed with any therapeutic genes in the TK region or N1L region without marker genes. Our marker-free system platform has significant potential for development of new marker-free VACV vectors for clinical application. PMID:26417609

  12. [Construction of human growth hormone lentiviral vector and its expression in murine skeletal myoblasts].

    PubMed

    Liu, Xiang-Yang; Lu, Yong-Xin; Xu, Yu-Lan; Li, Xiao-Qing; Liu, Juan; Li, Ai-Hua; Luo, Ping; Wan, Jian-Ping

    2006-03-01

    The aim of this study is to construct a lentiviral vector encoding human growth hormone, and to achieve the long, efficient and stable expression in murine skeletal myoblasts. Primary skeletal myoblasts were isolated from Sprague-Dawley rats and cultured by enzymatic digestion. We tested them by Desmin immunohistochemistry stains and found their viability was up to 94% by Trypan blue. Human growth hormone (hGH) cDNA was subcloned into expression vector pLenti6/V5-D-TOPO to construct recombinant pLenti6/V5-hGH. The pLenti6/V5-hGH and the contructed pLenti6/V5-EGFP were transfected into murine skeletal myoblasts by the Lipofectamin 2000. Through counting by the Confocal Laser Scanning Microscope, we identified the transfection efficency. We added the blasticidin to the 6-well plate with lids and obtained stable myoblasts expressing hGH. The concentration of human growth hormone (hGH) in cell culture medium was detected by Radioimmunoassay (RIA). Polymerase Chain Reaction (PCR) and DNA sequence showed hGH cDNA had been correctly inserted into pLenti6/V5-D-TOPO vector. Bright green fluorescence of the transfected cells could be observed under the Confocal Laser Scanning Microscope after 24 h transfection with pLenti6/V5-EGFP plasmids, and the transfection rate reached 40%. The difference was distinct (P < 0.01) between the pLenti6/V5- hGH groups and control groups in the secretive level of human growth hormone. After 8 weeks, the expression of human growth hormone was still stable. Then, we validated the biological characterization of the rhGH by the enzyme-link immunosorbent assay (ELISA) of the Insulin-like growth factor I (IGF-1). These results demonstrate we have successfully constructed the recombinant pLenti6/V5-hGH plasmids and accomplished rhGH long, efficient and stable expression ectopic in skeletal muscle myoblasts. PMID:16607951

  13. Optimized and enhanced DNA plasmid vector based in vivo construction of a neutralizing anti-HIV-1 envelope glycoprotein Fab.

    PubMed

    Muthumani, Kar; Flingai, Seleeke; Wise, Megan; Tingey, Colleen; Ugen, Kenneth E; Weiner, David B

    2013-10-01

    Monoclonal antibody preparations have demonstrated considerable clinical utility in the treatment of specific malignancies, as well as inflammatory and infectious diseases. Antibodies are conventionally delivered by passive administration, typically requiring costly large-scale laboratory development and production. Additional limitations include the necessity for repeat administrations, and the length of in vivo potency. Therefore, the development of methods to generate therapeutic antibodies and antibody like molecules in vivo, distinct from an active antigen-based immunization strategy, would have considerable clinical utility. In fact, adeno-associated viral (AAV) vector mediated delivery of immunoglobulin genes with subsequent generation of functional antibodies has recently been developed. As well, anon-viral vector mediated nucleic acid based delivery technology could permit the generation of therapeutic/prophylactic antibodies in vivo, obviating potential safety issues associated with viral vector based gene delivery. This delivery strategy has limitations as well, mainly due to very low in vivo production and expression of protein from the delivered gene. In the study reported here we have constructed an "enhanced and optimized" DNA plasmid technology to generate immunoglobulin heavy and light chains (i.e., Fab fragments) from an established neutralizing anti-HIV envelope glycoprotein monoclonal antibody (VRC01). This "enhanced" DNA (E-DNA) plasmid technology includes codon/RNA optimization, leader sequence utilization, as well as targeted potentiation of delivery and expression of the Fab immunoglobulin genes through use of "adaptive" in vivo electroporation. The results demonstrate that delivery by this method of a single administration of the optimized Fab expressing constructs resulted in generation of Fab molecules in mouse sera possessing high antigen specific binding and HIV neutralization activity for at least 7 d after injection, against diverse

  14. Optimized and enhanced DNA plasmid vector based in vivo construction of a neutralizing anti-HIV-1 envelope glycoprotein Fab

    PubMed Central

    Muthumani, Kar; Flingai, Seleeke; Wise, Megan; Tingey, Colleen; Ugen, Kenneth E; Weiner, David B

    2013-01-01

    Monoclonal antibody preparations have demonstrated considerable clinical utility in the treatment of specific malignancies, as well as inflammatory and infectious diseases. Antibodies are conventionally delivered by passive administration, typically requiring costly large-scale laboratory development and production. Additional limitations include the necessity for repeat administrations, and the length of in vivo potency. Therefore, the development of methods to generate therapeutic antibodies and antibody like molecules in vivo, distinct from an active antigen-based immunization strategy, would have considerable clinical utility. In fact, adeno-associated viral (AAV) vector mediated delivery of immunoglobulin genes with subsequent generation of functional antibodies has recently been developed. As well, anon-viral vector mediated nucleic acid based delivery technology could permit the generation of therapeutic/prophylactic antibodies in vivo, obviating potential safety issues associated with viral vector based gene delivery. This delivery strategy has limitations as well, mainly due to very low in vivo production and expression of protein from the delivered gene. In the study reported here we have constructed an “enhanced and optimized” DNA plasmid technology to generate immunoglobulin heavy and light chains (i.e., Fab fragments) from an established neutralizing anti-HIV envelope glycoprotein monoclonal antibody (VRC01). This “enhanced” DNA (E-DNA) plasmid technology includes codon/RNA optimization, leader sequence utilization, as well as targeted potentiation of delivery and expression of the Fab immunoglobulin genes through use of “adaptive” in vivo electroporation. The results demonstrate that delivery by this method of a single administration of the optimized Fab expressing constructs resulted in generation of Fab molecules in mouse sera possessing high antigen specific binding and HIV neutralization activity for at least 7 d after injection

  15. A protocol for construction of gene targeting vectors and generation of homologous recombinant ES cells

    PubMed Central

    Bouabe, Hicham; Okkenhaug, Klaus

    2015-01-01

    Summary The completion of human and mouse genome sequencing has confronted us with huge amount of data sequences that certainly need decades and many generations of scientists to be reasonably interpreted and assigned to physiological functions, and subsequently fruitfully translated into medical application. A means to assess the function of genes provides gene targeting in mouse embryonic stem (ES) cells that enables to introduce site-specific modifications in the mouse genome, and analyze their physiological consequences. Gene targeting enables almost any type of genetic modifications of interest, ranging from gene insertion (e.g. insertion of human-specific genes or reporter genes), gene disruption, point mutations, short and long range deletions, inversions. Site-specific modification into the genome of ES cells can be reached by homologous recombination using targeting vectors. Here, we describe a protocol to generate targeting constructs and homologous recombinant ES cells. PMID:23996269

  16. Construction and analysis of a secreting expression vector for fish cells.

    PubMed

    Collet, Bertrand; Secombes, Christopher J

    2005-02-18

    A new expression plasmid (pcDNA3-LP) was designed to produce and secrete proteins in fish cells by fusion with the rainbow trout TGF-beta leader peptide. The luciferase reporter gene was used to test the secreting ability of this vector. Secreting (pcDNA3-LP-LUC) and non-secreting (pcDNA3-LUC) constructs were made and compared in transient transfection experiments in salmonid (RTG-2) and cyprinid (EPC) cell lines. The amount of luciferase secreted into the supernatants of RTG-2 or EPC cells transiently transfected with pcDNA3-LP-LUC relative to cells transfected with pcDNA3-LUC was 7- and 85-fold, respectively. Two stable clones of EPC transfected with pcDNA3-LUC and four clones transfected with pcDNA3-LP-LUC were isolated. Approximately 90% of the total luciferase activity produced was secreted by stable EPC clones containing pcDNA3-LP-LUC whereas only 5% of total activity was secreted by clones containing pcDNA3-LUC. The two constructs were injected intra-muscularly into rainbow trout and the luciferase activity present in the serum of fish determined. The luciferase activity in serum from fish injected with pcDNA3-LP-LUC was 2.7-fold higher (P<0.05) than that fish injected with pcDNA3-LUC. This new vector opens up opportunities in fish DNA vaccinology and in the production of fish recombinant proteins. PMID:15694505

  17. High-throughput CRISPR Vector Construction and Characterization of DNA Modifications by Generation of Tomato Hairy Roots.

    PubMed

    Jacobs, Thomas B; Martin, Gregory B

    2016-01-01

    Targeted DNA mutations generated by vectors with clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology have proven useful for functional genomics studies. While most cloning strategies are simple to perform, they generally use multiple steps and can require several days to generate the ultimate constructs. The method presented here is based on DNA assembly and can produce fully functional CRISPR vectors in a single cloning reaction. Vector construction can also be pooled, further increasing the efficiency and utility of the process. A modification of the method is used to create CRISPR vectors with multiple gene targets. CRISPR vectors are then transformed into tomato hairy roots to generate transgenic materials with targeted DNA modifications. Hairy roots are a useful system for testing vector functionality as they are technically simple to generate and amenable to large-scale production. The methods presented here will have wide application as they can be used to generate a variety of CRISPR vectors and be used in a wide range of plant species. PMID:27167304

  18. Construction and application of an expression vector from the new plasmid pLAtc1 of Acidithiobacillus caldus.

    PubMed

    Zhang, Ming-Jiang; Jiang, Cheng-Ying; You, Xiao-Yan; Liu, Shuang-Jiang

    2014-05-01

    In this study, a recently sequenced 9.8-kb plasmid, pLAtc1, from Acidithiobacillus caldus strain SM-1 was characterized and developed into an expression vector. The pLAtc1 backbone carried an oriV, three rep genes, five mob genes, a Nic site, and an addiction system. Multilocus sequence analysis indicated that pLAtc1 was phylogenetically more related to the IncQ-like broad host range plasmids than to other IncQ plasmids. pLAtc1 was able to replicate and reside in Gram-negative Escherichia coli, Comamonas testosteroni, but not in Gram-positive Corynebacterium glutamicum. pLAtc1 was mobilized via conjugation into E. coli BL21 and A. caldus SM-1 from E. coli S17-1. Quantitative PCR revealed seven and four copies of plasmid in A. caldus and E. coli cells, respectively. The expression vector pLAtcE was constructed from pLAtc1 by introducing a regulatable promoter (P tetH ), a transcriptional terminator, a multiple cloning site, a kanamycin resistance gene, and a streptomycin resistance gene. The functionality of pLAtcE was demonstrated by expressing a gene encoding enhanced green fluorescence protein in E. coli and in A. caldus. pLAtcE was used to express α-ketoglutarate dehydrogenase (sucAB) and succinate dehydrogenase (sdhA) genes in A. caldus. The newly engineered strain that harbored sucAB and sdhA on a plasmid pLAtcE-sucA-sucB-sdhA grew better than the parent strain SM-1/pLAtcE in tetrathionate and glucose-supplemented medium and produced more acidity and resulted in a more oxidative environment. This study created a useful molecular tool for genetic manipulation of the thermoacidophilic and autotrophic A. caldus. PMID:24445921

  19. Production of Retrovirus-Based Vectors in Mildly Acidic pH Conditions.

    PubMed

    Holic, Nathalie; Fenard, David

    2016-01-01

    Gene transfer vectors based on retroviridae are increasingly becoming a tool of choice for biomedical research and for the development of biotherapies in rare diseases or cancers. To meet the challenges of preclinical and clinical production, different steps of the production process of self-inactivating γ-retroviral (RVs) and lentiviral vectors (LVs) have been improved (e.g., transfection, media optimization, cell culture conditions). However, the increasing need for mass production of such vectors is still a challenge and could hamper their availability for therapeutic use. Recently, we observed that the use of a neutral pH during vector production is not optimal. The use of mildly acidic pH conditions (pH 6) can increase by two- to threefold the production of RVs and LVs pseudotyped with the vesicular stomatitis virus G (VSV-G) or gibbon ape leukemia virus (GALV) glycoproteins. Here, we describe the production protocol in mildly acidic pH conditions of GALVTR- and VSV-G-pseudotyped LVs using the transient transfection of HEK293T cells and the production protocol of GALV-pseudotyped RVs produced from a murine producer cell line. These protocols should help to achieve higher titers of vectors, thereby facilitating experimental research and therapeutic applications. PMID:27317171

  20. HomeRun Vector Assembly System: A Flexible and Standardized Cloning System for Assembly of Multi-Modular DNA Constructs

    PubMed Central

    Li, Ming V.; Shukla, Dip; Rhodes, Brian H.; Lall, Anjali; Shu, Jingmin; Moriarity, Branden S.; Largaespada, David A.

    2014-01-01

    Advances in molecular and synthetic biology call for efficient assembly of multi-modular DNA constructs. We hereby present a novel modular cloning method that obviates the need for restriction endonucleases and significantly improves the efficiency in the design and construction of complex DNA molecules by standardizing all DNA elements and cloning reactions. Our system, named HomeRun Vector Assembly System (HVAS), employs a three-tiered vector series that utilizes both multisite gateway cloning and homing endonucleases, with the former building individual functional modules and the latter linking modules into the final construct. As a proof-of-principle, we first built a two-module construct that supported doxycycline-induced expression of green fluorescent protein (GFP). Further, with a three-module construct we showed quantitatively that there was minimal promoter leakage between neighbouring modules. Finally, we developed a method, in vitro Cre recombinase-mediated cassette exchange (RMCE) cloning, to regenerate a gateway destination vector from a previous multisite gateway cloning reaction, allowing access to existing DNA element libraries in conventional gateway entry clones, and simple creation of constructs ready for in vivo RMCE. We believe these methods constitute a useful addition to the standard molecular cloning techniques that could potentially support industrial scale synthesis of DNA constructs. PMID:24959875

  1. Advances in polymeric and inorganic vectors for nonviral nucleic acid delivery

    PubMed Central

    Sunshine, Joel C; Bishop, Corey J; Green, Jordan J

    2014-01-01

    Nonviral systems for nucleic acid delivery offer a host of potential advantages compared with viruses, including reduced toxicity and immunogenicity, increased ease of production and less stringent vector size limitations, but remain far less efficient than their viral counterparts. In this article we review recent advances in the delivery of nucleic acids using polymeric and inorganic vectors. We discuss the wide range of materials being designed and evaluated for these purposes while considering the physical requirements and barriers to entry that these agents face and reviewing recent novel approaches towards improving delivery with respect to each of these barriers. Furthermore, we provide a brief overview of past and ongoing nonviral gene therapy clinical trials. We conclude with a discussion of multifunctional nucleic acid carriers and future directions. PMID:22826857

  2. Construction of a new shuttle vector for DNA delivery into mammalian cells using non-invasive Lactococcus lactis.

    PubMed

    Yagnik, Bhrugu; Padh, Harish; Desai, Priti

    2016-04-01

    Use of food grade Lactococcus lactis (L. lactis) is fast emerging as a safe alternative for delivery of DNA vaccine. To attain efficient DNA delivery, L. lactis, a non-invasive bacterium is converted to invasive strain either by expressing proteins like Internalin A (InlA) or Fibronectin binding protein A (FnBPA) or through chemical treatments. However the safety status of invasive L. lactis is questionable. In the present report, we have shown that non-invasive L. lactis efficiently delivered the newly constructed reporter plasmid pPERDBY to mammalian cells without any chemical enhancers. The salient features of the vector are; I) Ability to replicate in two different hosts; Escherichia coli (E. coli) and Lactic Acid Bacteria (LAB), II) One of the smallest reporter plasmid for DNA vaccine, III) Enhanced Green Fluorescence Protein (EGFP) linked to Multiple Cloning Site (MCS), IV) Immunostimulatory CpG motifs functioning as an adjuvant. Expression of EGFP in pPERDBY transfected CHO-K1 and Caco-2 cells demonstrates its functionality. Non-invasive r-L. lactis was found efficient in delivering pPERDBY to Caco-2 cells. The in vitro data presented in this article supports the hypothesis that in the absence of invasive proteins or relevant chemical treatment, L. lactis was found efficient in delivering DNA to mammalian cells. PMID:26655884

  3. Gateway-assisted vector construction to facilitate expression of foreign proteins in the chloroplast of single celled algae.

    PubMed

    Oey, Melanie; Ross, Ian L; Hankamer, Ben

    2014-01-01

    With a rising world population, demand will increase for food, energy and high value products. Renewable production systems, including photosynthetic microalgal biotechnologies, can produce biomass for foods, fuels and chemical feedstocks and in parallel allow the production of high value protein products, including recombinant proteins. Such high value recombinant proteins offer important economic benefits during startup of industrial scale algal biomass and biofuel production systems, but the limited markets for individual recombinant proteins will require a high throughput pipeline for cloning and expression in microalgae, which is currently lacking, since genetic engineering of microalgae is currently complex and laborious. We have introduced the recombination based Gateway® system into the construction process of chloroplast transformation vectors for microalgae. This simplifies the vector construction and allows easy, fast and flexible vector design for the high efficiency protein production in microalgae, a key step in developing such expression pipelines. PMID:24523866

  4. Handbook for constructed wetlands receiving acid mine drainage

    SciTech Connect

    Wildeman, T.; Dietz, J.; Gusek, J.; Morea, S.

    1993-09-01

    In the summer of 1987, a pilot constructed wetland was built at the Big Five Tunnel in Idaho Springs, Colorado. The report details the theory, design and construction of wetlands receiving acid mine drainages, based on the second and third year of operation of this wetland, which was funded by the U.S. Environmental Protection Agency under the SITE Emerging Technologies Program. The text is divided into two broad sections: Part A - Theoretical Development, and Part B - Design Consideration. In the latter sections of Part A and through all of Part B the focus is on removal of metals by precipitation of sulfides through the activity of sulfate reducing bacteria.

  5. Recent progress in gene therapy to deliver nucleic acids with multivalent cationic vectors.

    PubMed

    Junquera, Elena; Aicart, Emilio

    2016-07-01

    Due to the potential use as transfecting agents of nucleic acids (DNA or RNA), multivalent cationic non-viral vectors have received special attention in the last decade. Much effort has been addressed to synthesize more efficient and biocompatible gene vectors able to transport nucleic acids into the cells without provoking an immune response. Among them, the mostly explored to compact and transfect nucleic acids are: (a) gemini and multivalent cationic lipids, mixed with a helper lipid, by forming lipoplexes; and (b) cationic polymers, polycations, and polyrotaxanes, by forming polyplexes. This review is focused on the progress and recent advances experimented in this area, mainly during the present decade, devoting special attention to the lipoplexes and polyplexes, as follows: (a) to its biophysical characterization (mainly electrostatics, structure, size and morphology) using a wide variety of experimental methods; and (b) to its biological activity (transfection efficacy and cytotoxicity) addressed to confirm the optimum formulations and viability of these complexes as very promising gene vectors of nucleic acids in nanomedicine. PMID:26265376

  6. Preparation, Physicochemical Properties, and Transfection Activities of Tartaric Acid-Based Cationic Lipids as Effective Nonviral Gene Delivery Vectors.

    PubMed

    Wan, Ning; Jia, Yi-Yang; Hou, Yi-Lin; Ma, Xi-Xi; He, Yong-Sheng; Li, Chen; Zhou, Si-Yuan; Zhang, Bang-Le

    2016-07-01

    In this work two novel cationic lipids using natural tartaric acid as linking backbone were synthesized. These cationic lipids were simply constructed by tartaric acid backbone using head group 6-aminocaproic acid and saturated hydrocarbon chains dodecanol (T-C12-AH) or hexadecanol (T-C16-AH). The physicochemical properties, gel electrophoresis, transfection activities, and cytotoxicity of cationic liposomes were tested. The optimum formulation for T-C12-AH and T-C16-AH was at cationic lipid/dioleoylphosphatidylethanolamine (DOPE) molar ratio of 1 : 0.5 and 1 : 2, respectively, and N/P charge molar ratio of 1 : 1 and 1 : 1, respectively. Under optimized conditions, T-C12-AH and T-C16-AH showed effective gene transfection capabilities, superior or comparable to that of commercially available transfecting reagent 3β-[N-(N',N'-dimethylaminoethyl)carbamoyl]cholesterol (DC-Chol) and N-[2,3-dioleoyloxypropyl]-N,N,N-trimethylammonium chloride (DOTAP). The results demonstrated that the two novel tartaric acid-based cationic lipids exhibited low toxicity and efficient transfection performance, offering an excellent prospect as nonviral vectors for gene delivery. PMID:27118165

  7. Characterization of engineered cartilage constructs using multiexponential T₂ relaxation analysis and support vector regression.

    PubMed

    Irrechukwu, Onyi N; Reiter, David A; Lin, Ping-Chang; Roque, Remigio A; Fishbein, Kenneth W; Spencer, Richard G

    2012-06-01

    Increased sensitivity in the characterization of cartilage matrix status by magnetic resonance (MR) imaging, through the identification of surrogate markers for tissue quality, would be of great use in the noninvasive evaluation of engineered cartilage. Recent advances in MR evaluation of cartilage include multiexponential and multiparametric analysis, which we now extend to engineered cartilage. We studied constructs which developed from chondrocytes seeded in collagen hydrogels. MR measurements of transverse relaxation times were performed on samples after 1, 2, 3, and 4 weeks of development. Corresponding biochemical measurements of sulfated glycosaminoglycan (sGAG) were also performed. sGAG per wet weight increased from 7.74±1.34 μg/mg in week 1 to 21.06±4.14 μg/mg in week 4. Using multiexponential T₂ analysis, we detected at least three distinct water compartments, with T₂ values and weight fractions of (45 ms, 3%), (200 ms, 4%), and (500 ms, 97%), respectively. These values are consistent with known properties of engineered cartilage and previous studies of native cartilage. Correlations between sGAG and MR measurements were examined using conventional univariate analysis with T₂ data from monoexponential fits with individual multiexponential compartment fractions and sums of these fractions, through multiple linear regression based on linear combinations of fractions, and, finally, with multivariate analysis using the support vector regression (SVR) formalism. The phenomenological relationship between T₂ from monoexponential fitting and sGAG exhibited a correlation coefficient of r²=0.56, comparable to the more physically motivated correlations between individual fractions or sums of fractions and sGAG; the correlation based on the sum of the two proteoglycan-associated fractions was r²=0.58. Correlations between measured sGAG and those calculated using standard linear regression were more modest, with r² in the range 0

  8. [Construction of RNAi vectors for SmNAC1 transcription factors of Salvia miltiorrhiza using Gateway cloning technology].

    PubMed

    Zhao, Rong; Rong, Qi-Xian; Liu, Yu-Zhong; Shen, Ye; Huang, Lu-Qi

    2014-05-01

    NAC transcription factors involved in plant growth and development, as well as responses to biotic and abiotic stress. RNAi Vectors for SmNAC transcription factors of Salvia miltiorrhiza was constructed by using Gateway cloning technology, in order to further study the function of SmNAC1 transcription factor. According to Gateway cloning technology, the specific fragments of SmNAC1 containing attB adapter was amplified by PCR using ultra-fideling phusion polymerase of NEB. By the BP recombination reaction, the PCR product containing attB was transferred to an donor vector (pENTR/SD/D-TOPO). Finally, SmNACi specific gene was cloned into pK7GWIWG2D plant expression vectors by LR recombination reaction. Experimental results showed that Gateway cloning technology provide a rapid and highly efficient way to clone the interested gene. PMID:25095362

  9. [Construction of ADAMTS13-pEGFP-N1 vector and its expression in HeLa cells].

    PubMed

    Ling, Jing; Ma, Zhen-Ni; Su, Jian; Ruan, Chang-Geng

    2013-02-01

    This study was aimed to construct a pEGFP-N1 vector of von Willebrand factor cleaving protease (ADAMTS13, a disintegrin and metalloprotease with a thrombospondin type 1 motifs 13) so as to pave the way for further studying its synthesis and secretion. Human full-length cDNA sequence of ADAMTS13 was acquired by polymerase chain reaction (PCR) with Phusion(®) High-Fidelity (NEB), then the PCR product was double digested with EcoRI and XhoI. After digestion, the ADAMTS13 cDNA sequence was purified and recombined with the pEGFP-N1 vector. The DNA sequence analysis showed that ADAMTS13 was ligated to the pEGFP-N1 vector correctly. After transient expression in HeLa cells, the expression of EGFP could be detected by fluorescent microscopy, and the expression of ADAMTS13 protein could be detected by SDS-PAGE and Western blot. It is concluded that the ADAMTS13-pEGFP-N1 vector is successfully constructed, and it can be widely used in further research on the mechanism of the synthesis and secretion of ADAMTS13. PMID:23484705

  10. Design and construction of two yeast shuttle vectors containing human procollagen genes expression cassette for expression in yeast.

    PubMed

    Abdemami, Baharak; Shokrgozar, Mohammad Ali; Shahreza, Hossein Khanahmad; Ghavami, Mehdi

    2011-01-01

    Collagens are the most abundant proteins in the human body. Their main function is to provide structural and mechanical support for the tissues, but they are also involved in a number of other biological functions including cell attachment, migration and differentiation. Collagens and gelatins are widely used in pharmaceutical and medical applications. Every year, more than 50,000 tons of collagen and gelatin are used in medical applications. These materials may have some viral and prion impurity and/or stimulate allergic response in human body. Therefore, scientists have produced human collagen in recombinant systems. In this study we have constructed two yeast shuttle vectors containing human procollagen genes expression cassette for expression in yeast. Total RNA was extracted from human skin fibroblast cell line, and cDNA synthesis was done by oligo dt. Then gene fragments were amplified from the cDNA with the necessary changes by Polymerase Chain Reaction (PCR). Finally they were cloned in yeast vector pPICZαA containing regulatory sequences for expressing and secreting the polypeptide product. Two yeast shuttle vectors containing human COL1A1 and COL1A2 expression cassettes were created. Final constructs were confirmed by enzymatic digestion, PCR of desired fragment and sequencing. The yeast shuttle vectors containing human COL1A1 and COL1A2 can be transferred into the yeast in the later stages to determine the scale of expression. PMID:23407617

  11. Acid-Labile Poly(glycidyl methacrylate)-Based Star Gene Vectors.

    PubMed

    Yang, Yan-Yu; Hu, Hao; Wang, Xing; Yang, Fei; Shen, Hong; Xu, Fu-Jian; Wu, De-Cheng

    2015-06-10

    It was recently reported that ethanolamine-functionalized poly(glycidyl methacrylate) (PGEA) possesses great potential applications in gene therapy due to its good biocompatibility and high transfection efficiency. Importing responsivity into PGEA vectors would further improve their performances. Herein, a series of responsive star-shaped vectors, acetaled β-cyclodextrin-PGEAs (A-CD-PGEAs) consisting of a β-CD core and five PGEA arms linked by acid-labile acetal groups, were proposed and characterized as therapeutic pDNA vectors. The A-CD-PGEAs owned abundant hydroxyl groups to shield extra positive charges of A-CD-PGEAs/pDNA complexes, and the star structure could decrease charge density. The incorporation of acetal linkers endowed A-CD-PGEAs with pH responsivity and degradation. In weakly acidic endosome, the broken acetal linkers resulted in decomposition of A-CD-PGEAs and morphological transformation of A-CD-PGEAs/pDNA complexes, lowering cytotoxicity and accelerating release of pDNA. In comparison with control CD-PGEAs without acetal linkers, A-CD-PGEAs exhibited significantly better transfection performances. PMID:25993557

  12. Construction of a pIX-modified Adenovirus Vector Able to Effectively Bind to Nanoantibodies for Targeting.

    PubMed

    Garas, M N; Tillib, S V; Zubkova, O V; Rogozhin, V N; Ivanova, T I; Vasilev, L A; Logunov, D Yu; Shmarov, M M; Tutykhina, I L; Esmagambetov, I B; Gribova, I Yu; Bandelyuk, A S; Naroditsky, B S; Gintsburg, A L

    2014-04-01

    Current targeting strategies for genetic vectors imply the creation of a specific vector for every targeted receptor, which is time-consuming and expensive. Therefore, the development of a universal vector system whose surface can specifically bind molecules to provide efficient targeting is of particular interest. In this study, we propose a new approach in creating targeted vectors based on the genome of human adenovirus serotype 5 carrying the modified gene of the capsid protein pIX (Ad5-EGFP-pIX-ER): recombinant pseudoadenoviral nanoparticles (RPANs). The surfaces of such RPANs are able to bind properly modified chimeric nanoantibodies that specifically recognize a particular target antigen (carcinoembryonic antigen (CEA)) with high affinity. The efficient binding of nanoantibodies (aCEA-RE) to the RPAN capsid surfaces has been demonstrated by ELISA. The ability of the constructed vector to deliver target genes has been confirmed by experiments with the tumor cell lines A549 and Lim1215 expressing CEA. It has been shown that Ad5-EGFP-pIX-ER carrying aCEA-RE on its surface penetrates into the tumor cell lines A549 and Lim1215 via the CAR-independent pathway three times more efficiently than unmodified RPAN and Ad5-EGFP-pIX-ER without nanoantibodies on the capsid surface. Thus, RPAN Ad5-EGFP-pIX-ER is a universal platform that may be useful for targeted gene delivery in specific cells due to "nanoantibody-modified RPAN" binding. PMID:25093116

  13. Construction and functional screening of a metagenomic library using a T7 RNA polymerase-based expression cosmid vector.

    PubMed

    Lussier, François-Xavier; Chambenoit, Olivier; Côté, Amélie; Hupé, Jean-François; Denis, François; Juteau, Pierre; Beaudet, Réjean; Shareck, François

    2011-09-01

    The metagenomic approach has greatly accelerated the discovery of new enzymes by giving access to the genetic potential of microorganisms from various environments. Function-based screening depends on adequate expression of the foreign genes in the heterologous host, which can be challenging in large-insert libraries. In this study, the shuttle cosmid vector pFX583 was used for the construction and screening of a metagenomic library. This vector allows T7 RNA polymerase-directed transcription of the cloned DNA and can be used in Escherichia coli and Streptomyces lividans. The DNA used for the library construction was obtained from an enriched biomass. The library was screened for lipolytic and proteolytic activities using E. coli and S. lividans as hosts. Numerous E. coli clones with lipolytic activity were detected. Unfortunately, proteases could not be detected in both hosts. From the lipolytic activity screen, a gene coding for a new lipase was isolated, and partial characterization was conducted. PMID:21108039

  14. Use of Green Fluorescent Protein To Tag Lactic Acid Bacterium Strains under Development as Live Vaccine Vectors

    PubMed Central

    Geoffroy, Marie-Claude; Guyard, Cyril; Quatannens, Brigitte; Pavan, Sonia; Lange, Marc; Mercenier, Annick

    2000-01-01

    The lactic acid bacteria (LAB) are safe microorganisms which are mainly used for the preparation of fermented foods and for probiotic applications. The potential of LAB as live vehicles for the production and delivery of therapeutic molecules such as antigens is also being actively investigated today. However, very little is known about the fate of live LAB when administered in vivo and about the interaction of these microorganisms with the nasal or gastrointestinal ecosystem. For future applications, it is essential to be able to discriminate the biotherapeutic strain from the endogenous microflora and to unravel the mechanisms underlying the postulated health-beneficial effect. We therefore started to investigate both aspects in a mouse model with two LAB species presently under development as live vaccine vectors, i.e., Lactococcus lactis and Lactobacillus plantarum. We have constructed different expression vectors carrying the gfp (green fluorescent protein [GFP]) gene from the jellyfish Aequoria victoria, and we found that this visible marker was best expressed when placed under the control of the inducible strong nisA promoter from L. lactis. Notably, a threshold amount of GFP was necessary to obtain a bright fluorescent phenotype. We further demonstrated that fluorescent L. plantarum NCIMB8826 can be enumerated and sorted by flow cytometry. Moreover, tagging of this strain with GFP allowed us to visualize its phagocytosis by macrophages in vitro and ex vivo and to trace it in the gastrointestinal tract of mice upon oral administration. PMID:10618252

  15. A series of vectors to construct lacZ fusions for the study of gene expression in Schizosaccharomyces pombe.

    PubMed

    Lafuente, M J; Petit, T; Gancedo, C

    1997-12-22

    We have constructed a series of plasmids to facilitate the fusion of promoters with or without coding regions of genes of Schizosaccharomyces pombe to the lacZ gene of Escherichia coli. These vectors carry a multiple cloning region in which fission yeast DNA may be inserted in three different reading frames with respect to the coding region of lacZ. The plasmids were constructed with the ura4+ or the his3+ marker of S. pombe. Functionality of the plasmids was tested measuring in parallel the expression of fructose 1,6-bisphosphatase and beta-galactosidase under the control of the fbp1+ promoter in different conditions. PMID:9450546

  16. [Construction of a set of secreting expression vectors for Saccharomyces cerevisiea].

    PubMed

    Zhao, Yingyi; Liang, Shizhong; Huang, Kun; Huang, Ribo

    2002-08-01

    The DNA fragment ecoding the Signal peptide of inulinase of Kluyveromyces smarxianu was synthesized chemically. This fragment was cloned in-frame in the expression vector pYES2 of Saccharomyces cerevisiae, resulting in a set of new secreting expression vectors pYES2 I, pYES2 II, pYES2 III. The L-Asparaginase gene (ASN) of E. coli and alpha-acetylactate decarboxylase gene (ALDC) of B. brevis which were amplified by PCR and cloned into the new vectors respectively were transformed into Saccharomyces cerevisia, and most of enzyme activities were secreted into the medium. The new secreting expression vectors still have excellent segregational stability even after growth for 100 h in the absence of selective pressure. PMID:12557548

  17. MAC-T cells as a tool to evaluate lentiviral vector construction targeting recombinant protein expression in milk.

    PubMed

    Monzani, Paulo S; Guemra, Samuel; Adona, Paulo R; Ohashi, Otavio M; Meirelles, Flávio V; Wheeler, Matthew B

    2015-01-01

    Prior to generating transgenic animals for bioreactors, it is important to evaluate the vector constructed to avoid poor protein expression. Mammary epithelial cells cultured in vitro have been proposed as a model to reproduce the biology of the mammary gland. In the present work, three lentiviral vectors were constructed for the human growth hormone (GH), interleukin 2 (IL2), and granulocyte colony-stimulating factor 3 (CSF3) genes driven by the bovine β-casein promoter. The lentiviruses were used to transduce mammary epithelial cells (MAC-T), and the transformed cells were cultured on polystyrene in culture medium with and without prolactin. The gene expression of transgenes was evaluated by PCR using cDNA, and recombinant protein expression was evaluated by Western-blotting using concentrated medium and cellular extracts. The gene expression, of the three introduced genes, was detected in both induced and non induced MAC-T cells. The human GH protein was detected in the concentrated medium, whereas CSF3 was detected in the cellular extract. Apparently, the cellular extract is more appropriate than the concentrated medium to detect recombinant protein, principally because concentrated medium has a high concentration of bovine serum albumin. The results suggest that MAC-T cells may be a good system to evaluate vector construction targeting recombinant protein expression in milk. PMID:25380466

  18. Construction of a lentiviral T/A vector for direct analysis of PCR-amplified promoters.

    PubMed

    Yu, Fu-xian; Zhu, Zhi-wei; Chen, Xiao-yu; Huang, Jing; Shi, Tuan-yuan; Li, Jun-xing; Pan, Jian-zhi

    2014-11-01

    The promoter plays an important role in the regulation of gene expression. To analyze a promoter's activity, we developed a novel lentiviral T/A vector that contains two reporter genes, a luciferase (Luc2) gene and a green fluorescent protein (Venus) gene, that are linked via an internal ribosome entry site (IRES2). To test the performance of this vector, phosphoglycerate kinase-1 (PGK) and elongation factor-1α (EF1α) promoters were amplified by PCR and inserted into this lentiviral T/A vector using T4 DNA ligase, yielding two promoter-reporter vectors: pLent-T-PGK and pLent-T-EF1α. When these vectors were transfected into 293T cells, we observed a higher level of Venus expression under a fluorescence microscopy in the case of pLent-T-EF1α as compared to pLent-T-PGK. The results of the luciferase reporter assay showed that the ratio of the promoter activities of EF1α and PGK was approximately 9:1. The two promoter-reporter vectors were also packaged as lentiviral particles to conduct promoter activity assay in cultured cells. The ratio of the promoter activities of EF1α and PGK was 4.23:1 when they were infected into 293T cells at a multiplicity of infection of 1. This value is comparable to that of a parallel experiment using the commercial luciferase reporter vector pGL4.10 with an activity ratio of 5.99:1 for EF1α and PGK. These results indicate that lentiviral T/A vector will be a useful tool for analysis of promoter activity and specificity. PMID:25091945

  19. [Construction of PPENK-MIDGE-NLS gene vector and the expression in rat].

    PubMed

    Chen, Xi; Xu, Xuemin; Peng, Xijuan; Jiang, Wei; Yao, Linong

    2015-02-01

    Increasing the production and secretion of endogenous opioid peptide by immune cell can significantly induce myocardial protective effects against ischemia-reperfusion injury. Gene therapy is promising to increase endogenous enkephalin (ENK). However, classical viral and plasmid vectors for gene delivery are hampered by immunogenicity, gene recombination, oncogene activation, the production of antibacterial antibody and changes in physiological gene expression. Minimalistic immunologically defined gene expression (MIDGE) can overcome all the deficients of viral and plasmid vectors. The exon of rat's preproenkephalin (PPENK) gene was amplified by PCR and the fragments were cloned into pEGFP-N1 plasmids. The recombined plasmids were digested with enzymes to obtain a linear vector contained promoter, preproenkephalin gene, RNA stable sequences and oligodesoxy nucleotides (ODNs) added to both ends of the gene vector to protect gene vector from exonuclease degradation. A nuclear localization sequence (NLS) was attached to an ODN to ensure the effective transport to the nucleus and transgene expression. Flow cytometry, laser confocal microscopy and Western blotting demonstrated that PPENK-MIDGE-NLS can transfect leukocyte of rat in vivo, increase the expression of proenkephalin (PENK) in tissue, and the transfection efficiency depends on gene vector's dosage. These results indicate that PPENK-MIDGE-NLS could be an innovative method to protect and treatment of myocardial ischemia-reperfusion injury. PMID:26062347

  20. Polyamidoamine dendrimer and oleic acid-functionalized graphene as biocompatible and efficient gene delivery vectors.

    PubMed

    Liu, Xiahui; Ma, Dongmei; Tang, Hao; Tan, Liang; Xie, Qingji; Zhang, Youyu; Ma, Ming; Yao, Shouzhuo

    2014-06-11

    Functionalized graphene has good potential in biomedical applications. To address a better and multiplex design of graphene-based gene vectors, the graphene-oleate-polyamidoamine (PAMAM) dendrimer hybrids were synthesized by the oleic acid adsorption and covalent linkage of PAMAM dendrimers. The micromorphology, electrical charge property, and amount of free amine groups of the graphene-oleate-PAMAM hybrids were characterized, and the peripheral functional groups were identified. The PAMAM dendrimers could be tethered onto graphene surface in high density. The graphene-oleate-PAMAM hybrids exhibit relatively good dispersity and stability in aqueous solutions. To evaluate the potential application of the hybrids in gene delivery vectors, cytotoxicity to HeLa and MG-63 cells and gene (plasmid DNA of enhanced green fluorescent protein) transfection capacity of the hybrids were investigated in detail. The graphene-oleate-PAMAM hybrids show mammalian cell type- and dose-dependent in vitro cytotoxicity. Under the optimal condition, the hybrids possess good biocompatibility and gene transfection capacity. The surface modification of graphene with oleic acid and PAMAM improves the gene transfection efficiency 13 times in contrast to the ultrasonicated graphene. Moreover, the hybrids show better transfection efficiency than the graphene oxide-PAMAM without the oleic acid modification. PMID:24836601

  1. [Expression of NANOG gene in acute lymphoblastic leukemia cells and construction of lentiviral vector carrying NANOG specific shRNA].

    PubMed

    Cao, Jiang; Meng, Fan-Jing; Li, Li; Lu, Chao; Zhou, Jun; Cheng, Hai; Chen, Wei; Chen, Chong; Xu, Kai-Lin

    2014-04-01

    The aim of this study was to detect the expression of NANOG gene in acute lymphoblastic leukemia (ALL) cells, and to construct the lentiviral vector carrying NANOG specific shRNA. The expression of NANOG was detected by RT-PCR and Western blot in MOLT-4, CCRF-HSB2, Jurkat cells and bone marrow cells from 15 patients with ALL in our hospital. The lentiviral vector carrying NANOG specific shRNA was constructed. After infection of MOLT-4 cells with the lentivirus constructs, GFP (+) cells were harvested by flow cytometry. The efficiency of RNA interference was detected by real-time quantitative PCR and Western blot. The results showed that the expression of NANOG mRNA and protein was detected in MOLT-4, CCRF-HSB2 cells and 33.3% samples of bone marrow from patients with ALL. The sequencing results demonstrated that the mRNAs amplified from these leukemic cells showed higher homology to NANOGP8 than NANOG1. The lentiviral vector pLB-shNANOG-1, pLB-shNANOG-2 and pLB-shcontrol were constructed. The viral particles were harvested and concentrated by ultracentrifugation. The virus titers were (1.83-3.12) ×10(8) IU/ml. After infection of MOLT-4 cells with the lentivirus, flow cytometry detection indicated that the GFP(+) cells were harvested by real-time quantitative PCR and Western blot, the assays showed that the 2 designed shRNA could significantly down-regulate expression of NANOG gene and protein. It is concluded that NANOGP8 is expressed in various types of ALL cells and in 33.3% of marrow cell samples obtained from ALL patients. After infection with the lentivirus constructs, MOLT-4 cells which stably down-regulate the expression of NANOG mRNA are obtained. PMID:24762991

  2. Colonization of a newly constructed urban wetland by mosquitoes in England: implications for nuisance and vector species.

    PubMed

    Medlock, Jolyon M; Vaux, Alexander G C

    2014-12-01

    Urban wetlands are being created in the UK as part of sustainable urban drainage strategies, to create wetland habitats lost during development, to provide a habitat for protected species, and to increase the public's access to 'blue-space' for the improvement of health and well-being. Sewage treatment reedbeds are also being incorporated into newly constructed wetlands to offer an alternative approach to dealing with sewage. This field study aims to provide the first UK evidence of how such newly constructed aquatic habitats are colonized by mosquitoes. A number of new aquatic habitats were surveyed for immature mosquitoes every fortnight over the first two years following wetland construction. The majority of mosquitoes collected were Culex sp. and were significantly associated with the sewage treatment reedbed system, particularly following storm events and sewage inflow. Other more natural aquatic habitats that were subject to cycles of drying and re-wetting contributed the majority of the remaining mosquitoes colonizing. Colonization of permanent habitats was slow, particularly where fluctuations in water levels inhibited emergent vegetation growth. It is recommended that during the planning process for newly constructed wetlands consideration is given on a case-by-case basis to the impact of mosquitoes, either as a cause of nuisance or as potential vectors. Although ornithophagic Culex dominated in this wetland, their potential role as enzootic West Nile virus vectors should not be overlooked. PMID:25424253

  3. The construction and use of divergence free vector expansions for incompressible fluid flow calculations

    NASA Technical Reports Server (NTRS)

    Mhuiris, N. M. G.

    1986-01-01

    For incompressible fluids the law of mass conservation reduces to a constraint on the velocity vector, namely that it be divergence free. This constraint has long been a source of great difficulty to the numericist seeking to discretize the Navier-Stokes and Euler equations. A spectral method is discussed which overcomes this difficulty. Its efficacy is demonstrated on some simple problems. The velocity is approximated by a finite sum of divergence free vectors, each of which satisfies the same boundary conditions as the velocity. Projecting the governing equation onto the space of inviscid vector fields eliminates the pressure term and produces a set of ordinary differential equations that must be solved for the coefficents in the velocity. The pressure can then be recovered if it is needed.

  4. New Invertebrate Vectors for PST, Spirolides and Okadaic Acid in the North Atlantic

    PubMed Central

    Silva, Marisa; Barreiro, Aldo; Rodriguez, Paula; Otero, Paz; Azevedo, Joana; Alfonso, Amparo; Botana, Luis M.; Vasconcelos, Vitor

    2013-01-01

    The prevalence of poisoning events due to harmful algal blooms (HABs) has declined during the last two decades through monitoring programs and legislation, implemented mainly for bivalves. However, new toxin vectors and emergent toxins pose a challenge to public health. Several locations on the Portuguese coast were surveyed between 2009 and 2010 for three distinct biotoxin groups [saxitoxin (PST), spirolide (SPX) and okadaic acid (OA)], in 14 benthic species of mollusks and echinoderms. Our main goals were to detect new vectors and unravel the seasonal and geographical patterns of these toxins. PSTs were analyzed by the Lawrence method, SPXs by LC-MS/MS, and OA by LC-MS/MS and UPLC-MS/MS. We report 16 new vectors for these toxins in the North Atlantic. There were differences in toxin contents among species, but no significant geographical or seasonal patterns were found. Our results suggest that legislation should be adjusted to extend the monitoring of marine toxins to a wider range of species besides edible bivalves. PMID:23739043

  5. Development of a Propionibacterium-Escherichia coli Shuttle Vector for Metabolic Engineering of Propionibacterium jensenii, an Efficient Producer of Propionic Acid

    PubMed Central

    Zhuge, Xin; Liu, Long; Shin, Hyun-dong; Chen, Rachel R.; Chen, Jian

    2013-01-01

    Propionic acid (PA) is an important chemical building block and is widely applied for organic synthesis, food, feedstuff, and pharmaceuticals. To date, the strains that can efficiently produce PA have included Propionibacterium thoenii, P. freudenreichii, and P. acidipropionici. In this report, we show that P. jensenii ATCC 4868 is also able to produce PA in much higher yields than the previously reported strains. To further improve the production capacity, a P. jensenii-Escherichia coli shuttle vector was developed for the metabolic engineering of P. jensenii. Specifically, a 6.9-kb endogenous plasmid, pZGX01, was isolated from P. acidipropionici ATCC 4875 and sequenced. Since the sequencing analysis indicated that pZGX01 could encode 11 proteins, the transcriptional levels of the corresponding genes were also investigated. Then, a P. jensenii-Escherichia coli shuttle vector was constructed using the pZGX01 plasmid, the E. coli pUC18 plasmid, and a chloramphenicol resistance gene. Interestingly, not only could the developed shuttle vector be transformed into P. jensenii ATCC 4868 and 4870, but it also could be transformed into freudenreichii ATCC 6207 subspecies of P. freudenreichii. Finally, the glycerol dehydrogenase gene (gldA) from Klebsiella pneumoniae was expressed in P. jensenii ATCC 4868 with the constructed shuttle vector. In a 3-liter batch culture, the PA production by the engineered P. jensenii ATCC 4868 strain reached 28.23 ± 1.0 g/liter, which was 26.07% higher than that produced by the wild-type strain (22.06 ± 1.2 g/liter). This result indicated that the constructed vector can be used a useful tool for metabolic engineering of P. jensenii. PMID:23709505

  6. Construction of a broad host range shuttle vector for gene cloning and expression in Actinobacillus pleuropneumoniae and other Pasteurellaceae.

    PubMed

    Frey, J

    1992-01-01

    We have constructed a pair of broad host range expression vectors, pJFF224-NX and pJFF224-XN, based on plasmid RSF1010, which enable cloning and efficient expression of genes in Actinobacillus pleuropneumoniae and Pasteurella haemolytica and in Escherichia coli. The vectors consist of the minimal autonomous replicon of the broad host range plasmid RSF1010 and a type II chloramphenicol acetyl transferase gene for chloramphenicol resistance selection. In addition, they contain a gene expression cassette based on the E. coli bacteriophage T4 gene 32 promoter region and a transcription stop signal, which are separated by a segment of multiple cloning sites in both orientations. Electroporation and subsequent selection for chloramphenicol resistance was used for the introduction of the vectors in A. pleuropneumoniae and P. haemolytica. A promoterless xy/E gene from the Pseudomonas putida TOL plasmid was cloned onto pJFF224-NX. This plasmid enabled efficient expression of active catechol2,3oxygenase in A. pleuropneumoniae and P. haemolytica. It was stably maintained in A. pleuropneumoniae without antibiotic selection, showing less than 0.1% loss after 100 generations, while native RSF1010 and other RSF1010-based vectors were unstable in this host. PMID:1448612

  7. A PCR-Based Method to Construct Lentiviral Vector Expressing Double Tough Decoy for miRNA Inhibition.

    PubMed

    Qiu, Huiling; Zhong, Jiasheng; Luo, Lan; Liu, Nian; Kang, Kang; Qu, Junle; Peng, Wenda; Gou, Deming

    2015-01-01

    DNA vector-encoded Tough Decoy (TuD) miRNA inhibitor is attracting increased attention due to its high efficiency in miRNA suppression. The current methods used to construct TuD vectors are based on synthesizing long oligonucleotides (~90 mer), which have been costly and problematic because of mutations during synthesis. In this study, we report a PCR-based method for the generation of double Tough Decoy (dTuD) vector in which only two sets of shorter oligonucleotides (< 60 mer) were used. Different approaches were employed to test the inhibitory potency of dTuDs. We demonstrated that dTuD is the most efficient method in miRNA inhibition in vitro and in vivo. Using this method, a mini dTuD library against 88 human miRNAs was constructed and used for a high-throughput screening (HTS) of AP-1 pathway-related miRNAs. Seven miRNAs (miR-18b-5p, -101-3p, -148b-3p, -130b-3p, -186-3p, -187-3p and -1324) were identified as candidates involved in AP-1 pathway regulation. This novel method allows for an accurate and cost-effective generation of dTuD miRNA inhibitor, providing a powerful tool for efficient miRNA suppression in vitro and in vivo. PMID:26624995

  8. [Construction of IK6 recombinant lentiviral vector and its expression and biologic feature in THP1 cells].

    PubMed

    Zhang, Na; Liu, Ya-Nan; Xiao, Min; Ding, Xiao-Yi; Zhou, Jian-Feng; Li, Chun-Rui

    2014-08-01

    The purpose of this study was to construct a lentiviral vector carrying IK6 gene and to observe the expression of IK6 as well as related biologic feature in THP1 cells, so as to provide an effective method to further investigate the role of this gene in leukemia. The IK6 gene was obtained by using reverse transcription polymerase chain reaction (RT-PCR). Then IK6 was recombined with the pGC-FU vector to construct a recombinant lentiviral vector named pGC-FU-IK6 gene-GFP,which was confirmed by PCR and sequencing. The 293T cells were transfected with pGC-FU- IK6-GFP by using Lipofectamine 2000. After examining the titer of the virus, pGC-FU- IK6-GFP was used to transfect THP1 cells. The transfection efficiency was detected by flow cytometry, and the expression level of mRNA and IK6-GFP fusion protein were confirmed by RT-PCR and Western blot respectively. Then the impact of IK6 on apoptosis and cell cycle was analyzed. The results showed that the IK6 gene was obtained by RT-PCR and connected into the linearized lentiviral vector to successfully constructed target plasmid named pGC-FU-IK6-GFP with Amp resistant. The target plasmid was transfected into 293T cells and the virus titer was 2.0×10(9)TU/ml. Next, THP1 cells were transfected with pGC-FU-IK6-GFP and the efficiency was up to 90%. The detection of the IK6 mRNA and IK6-GFP fusion protein in target cells showed that IK6 could promote target cell clone formation and inhibit apoptosis, but had no significant effect on the cell cycle. It is concluded that virus vector carrying IK6 gene had been successfully constructed and expressed in THP1 stably. Biology studies of target THP1 cell shows that the IK6 is likely to interfere with the function of normal Ikaros protein as tumor suppressor, and it exerts a potential anti-apoptotic effect. Thus, IK6 can promote leukemia cell growth. However, there is no significant effect on the cell cycle. It provides an effective method for exploring the function of IK6 in acute

  9. Construction of an infectious clone of a plant RNA virus in a binary vector using one-step Gibson Assembly.

    PubMed

    Blawid, Rosana; Nagata, Tatsuya

    2015-09-15

    The construction of full-length infectious clones of RNA viruses is often laborious due to the many cloning steps required and the DNA exclusion within the plasmid during Escherichia coli transformation. We demonstrate single-step cloning procedure of an infectious cDNA of the tomato blistering mosaic virus (ToBMV) using Gibson Assembly (GA), which drastically reduces the number of cloning steps. By agro-inoculation with the construct obtained by this procedure, ToBMV was recovered six days post-inoculation in Nicotiana benthamiana plants. The symptoms induced by the recovered virus were indistinguishable from those caused by the wild-type virus. We conclude that the GA is very useful method particularly to construct a full-length cDNA clone of a plant RNA virus in a binary vector. PMID:25986144

  10. [Full-length cDNA cloning of flavonol synthase genes of Carthamus tinctorius and construction plant expression vector].

    PubMed

    Yang, Wen-ting; Liu, Xiu-ming; Wan, Qiu; Yao, Na; Wang, Nan; Zhang, Xue-meng; Jiao, Zhong-da; Li, Hai-yan; Li, Xiao-kun

    2015-02-01

    Flavonol synthase (FLS) is one of the key enzymes in flavonoids metabolic pathways. In this study, middle sequence was obtained from Carthamus tinctorius transcriptome sequencing results. Full-length cDNAs of FLS was cloned from petals of C. tinctorius to FLS by using RT-PCR and RACE technology. Its full-length cDNA was 1,201 bp, with an open reading frame of 1,101 bp and 336 encoded amino acids. The phylogenetic analysis showed that, FLS gene encoded amino acids in C. tinctorius were highly homologous with amino acids in congeneric Compositae species, especially Rudbeckia laciniata. The pBASTA-FLS plant expression vector was successfully built by the molecular biology method, which lays a foundation for further studying biology functions of the gene and biosynthesis mechanism of flavonoids. PMID:26137682

  11. Construction of p66Shc gene interfering lentivirus vectors and its effects on alveolar epithelial cells apoptosis induced by hyperoxia

    PubMed Central

    Zhang, Chan; Dong, Wen-Bin; Zhao, Shuai; Li, Qing-Ping; Kang, Lan; Lei, Xiao-Ping; Guo, Lin; Zhai, Xue-Song

    2016-01-01

    Background The aim of this study is to observe the inhibitive effects of p66Shc gene interfering lentivirus vectors on the expression of p66Shc, and to explore its effects on alveolar epithelial cells apoptosis induced by hyperoxia. Methods The gene sequences were cloned into the pLenR-GPH-shRNA lentiviral vector, which was selected by Genebank searches. The pLenR-GPH-shRNA and lentiviral vector packaging plasmid mix were cotransfected into 293T cells to package lentiviral particles. Culture virus supernatant was harvested, and then the virus titer was determined by serial dilution assay. A549 cells were transduced with the constructed lentiviral vectors, and real-time polymerase chain reaction (RT-PCR) and Western blot were used to evaluate p66Shc expression. This study is divided into a control group, a hyperoxia group, an A549-p66ShcshRNA hyperoxia group, and a negative lentivirus group. Cell apoptosis was detected by flow cytometry after 24 hours; the expression of X-linked inhibitor of apoptosis protein (XIAP) and caspase-9 were detected by immunohistochemistry assay. The production of reactive oxygen species and cellular mitochondria membrane potential (ΔΨm) were determined by fluorescence microscopy. Results We successfully established the p66Shc gene interfering lentivirus vectors, A549-p66ShcshRNA. The A549-p66ShcshRNA was transfected into alveolar epithelial cells, and the inhibitive effects on the expression of p66Shc were observed. Both RT-PCR and Western blot demonstrated downregulation of p66Shc expression in A549 cells. In the A549-p66ShcshRNA hyperoxia group, we found dampened oxidative stress. A549-p66ShcshRNA can cause p66Shc gene silencing, reduce mitochondrial reactive oxygen species generation, reduce membrane potential decrease, reduce the apoptosis of A549 cells, and reduce alveolar epithelial cell injury, while the lentiviral empty vector group had no such changes. Conclusion p66Shc gene interfering lentivirus vector can affect the

  12. Exploring the limits of vector construction based on Citrus tristeza virus.

    PubMed

    El-Mohtar, Choaa; Dawson, William O

    2014-01-01

    We examined the limits of manipulation of the Citrus tristeza virus (CTV) genome for expressing foreign genes in plants. We previously created a vector with a foreign gene cassette inserted between the major and minor coat protein genes, which is position 6 from the 3' terminus. Yet, this virus has 10 3'-genes with several other potential locations for expression of foreign genes. Since genes positioned closer to the 3' terminus tend to be expressed in greater amounts, there were opportunities for producing greater amounts of foreign protein. We found that the virus tolerated insertions of an extra gene in most positions within the 3' region of the genome with substantially increased levels of gene product produced throughout citrus trees. CTV was amazingly tolerant to manipulation resulting in a suite of stable transient expression vectors, each with advantages for specific uses and sizes of foreign genes in citrus trees. PMID:24314658

  13. [Construction of recombinant adenoviral vector expressing genes of the conservative influenza proteins M2 and nucleoprotein].

    PubMed

    Esmagambetov, I B; Sedova, E S; Shcherbinin, D N; Lysenko, A A; Garas, M N; Shmarov, M M; Logunov, D Iu

    2014-01-01

    Influenza is a highly contagious and one of the most massive infection diseases. General epidemiological significance has a strain, which belongs to subtype A. A high degree of genetic variety leads to the permanent changes in the antigenic structure of the influenza virus. Therefore, the current influenza vaccines require periodic updating of the composition of strains. Presently, it is important to develop a universal vaccine that can protect against different strains of influenza A virus at the same time and is based on the conserved antigens of the influenza virus. The recombinant adenovirus vectors expressing genes of conserved viral antigenes may be a promising candidate vaccine against influenza A. Using the method of the homologous recombination, we developed in this study recombinant adenovirus of fifth serotype that expresses genes of the ion channel M2 and nucleoprotein NP of the influenza virus A. Genes of the consensus protein M2 and NP of human influenza A virus were included into the structure of the viral genome. The expression of the antigens M2 and NP using recombinant adenovirus vector was detected by a Western blot assay. The immunogenicity of the developed recombinant adenovirus vector was demonstrated by the intranasal immunization of laboratory mice. PMID:25080815

  14. A simplified approach to construct infectious cDNA clones of a tobamovirus in a binary vector.

    PubMed

    Junqueira, Bruna Rayane Teodoro; Nicolini, Cícero; Lucinda, Natalia; Orílio, Anelise Franco; Nagata, Tatsuya

    2014-03-01

    Infectious cDNA clones of RNA viruses are important tools to study molecular processes such as replication and host-virus interactions. However, the cloning steps necessary for construction of cDNAs of viral RNA genomes in binary vectors are generally laborious. In this study, a simplified method of producing an agro-infectious Pepper mild mottle virus (PMMoV) clone is described in detail. Initially, the complete genome of PMMoV was amplified by a single-step RT-PCR, cloned, and subcloned into a small plasmid vector under the T7 RNA polymerase promoter to confirm the infectivity of the cDNA clone through transcript inoculation. The complete genome was then transferred to a binary vector using a single-step, overlap-extension PCR. The selected clones were agro-infiltrated to Nicotiana benthamiana plants and showed to be infectious, causing typical PMMoV symptoms. No differences in host responses were observed when the wild-type PMMoV isolate, the T7 RNA polymerase-derived transcripts and the agroinfiltration-derived viruses were inoculated to N. benthamiana, Capsicum chinense PI 159236 and Capsicum annuum plants. PMID:24388933

  15. Construction of Hermes shuttle vectors: a versatile system useful for genetic complementation of transformable and non-transformable Neisseria mutants.

    PubMed

    Kupsch, E M; Aubel, D; Gibbs, C P; Kahrs, A F; Rudel, T; Meyer, T F

    1996-03-20

    A versatile shuttle system has been developed for genetic complementation with cloned genes of transformable and non-transformable Neisseria mutants. By random insertion of a selectable marker into the conjugative Neisseria plasmid ptetM25.2, a site within this plasmid was identified that is compatible with plasmid replication and with conjugative transfer of plasmid. Regions flanking the permissive insertion site of ptetM25.2 were cloned in Escherichia coli and served as a basis for the construction of the Hermes vectors. Hermes vectors are composed of an E. coli replicon that does not support autonomous replication in Neisseria, e.g. ColE1, p15A, or ori(fd), fused with a shuttle consisting of a selectable marker and a multiple cloning site flanked by the integration region of ptetM25.2. Complementation of a non-transformable Neisseria strain involves a three-step process: (i) insertion of the desired gene into a +Hermes vector; (ii) transformation of Hermes into a Neisseria strain containing ptetM25.2 to create a hybrid ptetM25.2 via gene replacement by the Hermes shuttle cassette; and (iii) conjugative transfer of the hybrid ptetM25.2 into the final Neisseria recipient. Several applications for the genetic manipulation of pathogenic Neisseriae are described. PMID:8676859

  16. Construction of expression vectors for metabolic engineering of the vanillin-producing actinomycete Amycolatopsis sp. ATCC 39116.

    PubMed

    Fleige, Christian; Steinbüchel, Alexander

    2014-01-01

    Amycolatopsis sp. ATCC 39116 is able to synthesize the important flavoring agent vanillin from cheap natural substrates. The bacterium is therefore of great interest for the industry and used for the fermentative production of vanillin. In order to improve the production of natural vanillin with Amycolatopsis sp. ATCC 39116, the strain has been genetically engineered to optimize the metabolic flux towards the desired product. Extensive metabolic engineering was hitherto hampered, due to the lack of genetic tools like functional promoters and expression vectors. In this study, we report the establishment of a plasmid-based gene expression system for Amycolatopsis sp. ATCC 39116 that allows a further manipulation of the genotype. Four new Escherichia coli-Amycolatopsis shuttle vectors harboring different promoter elements were constructed, and the functionality of these regulatory elements was proven by the expression of the reporter gene gusA, encoding a β-glucuronidase. Glucuronidase activity was detected in all plasmid-harboring strains, and remarkable differences in the expression strength of the reporter gene depending on the used promoter were observed. The new expression vectors will promote the further genetic engineering of Amycolatopsis sp. ATCC 39116 to get insight into the metabolic network and to improve the strain for a more efficient industrial use. PMID:24743982

  17. Construction and characterization of plasmid and lambda phage vector systems for study of transcriptional control in Escherichia coli.

    PubMed

    Hirano, M; Shigesada, K; Imai, M

    1987-01-01

    We constructed a family of lambda phage and plasmid vectors which facilitate cloning and quantitative analysis of transcriptional regulator in both single and multiple copies. Their expression system was modified from the ara-trp-lac fusion operon of plasmid pMC81 [Casadaban and Cohen, J. Mol. Biol. 138 (1980) 179-207], which is designed to assay both promoters and terminators with a single vehicle. To eliminate transcriptional and translational polar effects liable to occur in the original fusion operon upon insertion of a foreign nucleotide sequence, intracistronic Rho-dependent terminators, that are present within the trpB gene and distal to the cloning site were deleted, and DNA spacers containing stop codons were introduced immediately before and after the cloning site. In analysis of the cloned trp regulatory region, the lambda phage system faithfully reproduced the tight regulation by tryptophan characteristic to the natural trp operon on the E. coli chromosome, whereas the plasmid counterpart exhibited a substantially relaxed response. Comparative studies on the relative strengths of various promoters and terminators have further demonstrated that the lambda phage vector system permits accurate assays of exceptionally strong promoters like Ptrp and lambda pL without disturbing the bacterial growth, while being sensitive enough for detecting low-level transcription under the control of weak promoters or potent terminators. Cloning with the lambda phage vector can be greatly facilitated by transferring the target regulatory site precloned with the plasmid onto the phage genome through in vivo recombination. PMID:2828183

  18. Development of series of gateway binary vectors, pGWBs, for realizing efficient construction of fusion genes for plant transformation.

    PubMed

    Nakagawa, Tsuyoshi; Kurose, Takayuki; Hino, Takeshi; Tanaka, Katsunori; Kawamukai, Makoto; Niwa, Yasuo; Toyooka, Kiminori; Matsuoka, Ken; Jinbo, Tetsuro; Kimura, Tetsuya

    2007-07-01

    We developed a new series of binary vectors useful for Gateway cloning to facilitate transgenic experiments in plant biotechnology. The new system, Gateway Binary Vectors (pGWBs) realized efficient cloning, constitutive expression using the cauliflower mosaic virus (CaMV) 35S promoter and the construction of fusion genes by simple clonase reaction with an entry clone. The reporters employable in this system are beta-glucuronidase (GUS), synthetic green fluorescent protein with S65T mutation (sGFP), luciferase (LUC), enhanced yellow fluorescent protein (EYFP), and enhanced cyan fluorescent protein (ECFP). The tags available are 6xHis, FLAG, 3xHA, 4xMyc, 10xMyc, GST, T7-epitope, and tandem affinity purification (TAP). In total, 13 kinds of reporter or tag were arranged and were almost applicable to both N- and C-fusions. The pGWBs could be used for many purposes, such as promoter::reporter analysis, observation of subcellular localization by the expression of proteins fused to a reporter or tag, and analysis of protein-protein interaction by copurification and immunodetection experiments. The pGWBs were constructed with modified pBI101 containing a CaMV35S promoter-driven hygromycin phosphotransferase (HPT) gene as the second selection marker. We also constructed pGWBs with the marker HPT driven by the nopaline synthase promoter. By using the pGWB system, the expression of tagged proteins, and the localization of GFP-fused proteins were easily analyzed. Moreover, tissue-specific and inducible gene expression using a promoter was also monitored with pGWBs. It is expected that, the pGWB system will serve as a powerful tool for plasmid construction in plant research. PMID:17697981

  19. Polyethylenimine: A versatile, multifunctional non-viral vector for nucleic acid delivery.

    PubMed

    Pandey, Abhijeet P; Sawant, Krutika K

    2016-11-01

    Polyethylenimine (PEI) has recently been widely studied for the design of nucleic acid delivery vehicles. Gene delivery using PEI involves condensation of DNA into compact particles, uptake into the cells, release from the endosomal compartment into the cytoplasm, and uptake of the DNA into the nucleus. PEIs being positively charged, linear or branched polymers are able to form nanoscale complexes with small RNAs, leading to RNA protection, cellular delivery, and intracellular release. This review highlights the important properties of various PEIs with regard to their use for nucleic acid delivery. Brief discussion on cellular uptake mechanism of non-viral vector is included to understand its utility for gene delivery. Applications of modified PEI for increased efficacy, altered pharmacokinetic properties; improved biocompatibility and targeted delivery have also been discussed. An overview of simulation studies which can help in understanding the underlying complexation mechanism has also been included. The review provides a brief discussion about clinical trials and patents related to nucleic acid delivery using PEI based systems. PMID:27524093

  20. Characterization of an Oncolytic Adenovirus Vector Constructed to Target the cMet Receptor

    PubMed Central

    Sakr, Hany I; Coleman, David T; Cardelli, James A; Mathis, J Michael

    2015-01-01

    The cMet receptor is a homodimer with tyrosine kinase activity. Upon stimulation with its ligand, hepatocyte growth factor (HGF), the receptor mediates wide physiologic actions. The HGF-cMet signaling pathway is dysregulated in many cancers, which makes cMet an important target for novel therapeutic interventions. Oncolytic adenoviruses (Ads) have been used for the past three decades as a promising therapeutic approach for a wide array of neoplastic diseases. To date, achieving cancer-specific replication of oncolytic Ads has been accomplished by either viral genome deletions or by incorporating tumor selective promoters. To achieve novel specificity of oncolytic Ad infection of cancer cells that overexpress cMet, we inserted the HGF NK2 sequence, corresponding to a competitive antagonist of HGF binding to the cMet receptor, into the Ad serotype 5 (Ad5) fiber gene. The resulting vector, Ad5-pIX-RFP-FF/NK2, was rescued, amplified in HEK293 cells, and characterized. Binding specificity and viral infectivity were tested in various cancer cell lines that express varying levels of cMet and hCAR (the Ad5 receptor). We found that Ad5-pIX-RFP-FF/NK2 demonstrated binding specificity to the cMet receptor. In addition, there was enhanced viral infectivity and virus replication compared with a non-targeted Ad vector. Although NK2 weakly induces cMet receptor activation, our results showed no receptor phosphorylation in the context of an oncolytic Ad virus. In summary, these results suggest that an oncolytic Ad retargeted to the cMet receptor is a promising vector for developing a novel cancer therapeutic agent. PMID:26866014

  1. Construction of minicircle DNA vectors capable of correcting familial hypercholesterolemia phenotype in a LDLR-deficient mouse model.

    PubMed

    Hou, X; Jiao, R; Guo, X; Wang, T; Chen, P; Wang, D; Chen, Y; He, C-Y; Chen, Z-Y

    2016-08-01

    Familial hypercholesterolemia (FH) caused by defect in low-density lipoprotein receptor (LDLR) is a life-threatening disease with poor response to conventional treatments. Earlier gene therapy studies have generated promising results, but further development is hampered because the cells harboring the viral vectors were eliminated by host immune system soon after delivery, whereas the nonviral vectors were too bulky to be delivered to target cells. To overcome these problems, we constructed multiple minicircle (MC) DNA vectors to express the therapeutic LDLR. MC is an optimized nonviral vector that is capable of expressing high level of transgene product persistently. We found that among the seven MCs tested, the best is MC5 with multiple advanced features. First, the LDLr gene was placed under the control of sterol regulatory element (SRE) using LDLr gene promoter or apoprotein E (ApoE) promoter, allowing the transcription of the LDLr gene to be regulated by serum low-density lipoprotein (LDL) cholesterol as its functional gene counterpart. Second, a hepatic control region (HCR) was placed upstream of the promoter that serves as a controller to ensure liver-specific expression. Third, the modified Kozak sequence was placed in front of the LDLr gene start codon to enhance its translation efficiency. MC5 was 5.23 kb in size, and was capable of tight physiological control in intracellular LDL cholesterol level even when challenged with high dose of sterols in vitro. Importantly, it was able to correct the phenotype of LDLR-deficient mice C57BL/6 LDLR(-/-) for more than 105 days without detectable toxicity. Therefore, this MC has the clinical application potential for treating FH. PMID:27092942

  2. Construction of a new shuttle vector and its use for cloning and expression of two plasmid-encoded bacteriocins from Lactobacillus paracasei subsp. paracasei BGSJ2-8.

    PubMed

    Kojic, Milan; Lozo, Jelena; Jovcic, Branko; Strahinic, Ivana; Fira, Djordje; Topisirovic, Ljubisa

    2010-06-15

    A new shuttle-cloning vector, pA13, was constructed and successfully introduced into Escherichia coli, Lactobacillus and Lactococcus strains. It showed high segregational and structural stability in all three hosts. The natural plasmid pSJ2-8 from L. paracasei subsp. paracasei BGSJ2-8 was cloned into pA13 using BamHI to obtain the construct, pB5. Sequencing and in silico analysis of pB5 revealed fifteen open reading frames (ORF). Plasmid pSJ2-8 harbours genes encoding the production of two bacteriocins, BacSJ and acidocin 8912. Combined N-terminal amino acid sequencing of BacSJ in combination with DNA sequencing of the bacSJ2-8 gene enabled determination of the primary structure of bacteriocin BacSJ. The bacSJ2-8 gene encodes 68-amino-acid peptide with a double-glycine leader peptide consisting of 18 amino acids, followed by the orf2 (bacSJ2-8i) which encodes the immunity protein of BacSJ. The production and functional expression of BacSJ in homologous and heterologous hosts suggest that bacSJ2-8 and bacSJ2-8i together with the genes encoding the ABC transporter and accessory protein are the minimal requirements for production of BacSJ. Biochemical and genetic analyses showed that BacSJ belongs to class II bacteriocins. PMID:20439125

  3. Identification, characterization and use of two tick promoters for construction of a dual luciferase reporter vector

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dual luciferase reporter systems are valuable tools for functional genomic studies, but have not previously been developed for use in tick cell culture. We evaluated expression of available luciferase constructs in tick cell cultures derived from Rhipicephalus (Boophilus) microplus, an important vec...

  4. Isolation of a novel IS3 group insertion element and construction of an integration vector for Lactobacillus spp.

    PubMed Central

    Walker, D C; Klaenhammer, T R

    1994-01-01

    An insertion sequence (IS) element from Lactobacillus johnsonii was isolated, characterized, and exploited to construct an IS-based integration vector. L. johnsonii NCK61, a high-frequency conjugal donor of bacteriocin production (Laf+) and immunity (Lafr), was transformed to erythromycin resistance (Emr) with the shuttle vector pSA3. The NCK61 conjugative functions were used to mobilize pSA3 into a Laf- Lafs EMs recipient. DNA from the Emr transconjugants transformed into Escherichia coli MC1061 yielded a resolution plasmid with the same size as that of pSA3 with a 1.5-kb insertion. The gram-positive replication region of the resolution plasmid was removed to generate a pSA3-based suicide vector (pTRK327) bearing the 1.5-kb insert of Lactobacillus origin. Plasmid pTRK327 inserted randomly into the chromosomes of both Lactobacillus gasseri ATCC 33323 and VPI 11759. No homology was detected between plasmid and total host DNAs, suggesting a Rec-independent insertion. The DNA sequence of the 1.5-kb region revealed the characteristics of an IS element (designated IS1223): a length of 1,492 bp; flanking, 25-bp, imperfect inverted repeats; and two overlapping open reading frames (ORFs). Sequence comparisons revealed 71.1% similarity, including 35.7% identity, between the deduced ORFB protein of the E. coli IS element IS150 and the putative ORFB protein encoded by the Lactobacillus IS element. A putative frameshift site was detected between the overlapping ORFs of the Lactobacillus IS element. It is proposed that, similar to IS150, IS1223 produces an active transposase via translational frameshifting between two tandem, overlapping ORFs. Images PMID:8071209

  5. [Construction and identification of recombinant lentiviral vector of hNoc4L gene].

    PubMed

    Wang, Tingting; Wang, Shujuan; Yan, Jinghua

    2010-11-01

    Formation and nuclear export of pre-ribosomes requires many nucleolar complexes, hNoc4L which contains a conserved Noc doman is a homolog of nucleolar complex associated 4 (S. cerevisiae), but its function is completely unclear. Here, we successfully got the recombinant lentiviral vector p113.7-EF1-hNoc4L-Flag by replacing the U6 promoter in p113.7 with EF1alpha promoter, and then inserted hNoc4L to down-stream of the EF1alpha prompter. We determined the transduction efficiency in different mammalian cell lines based on lentiviral packaging system. Subsequently, we analyzed the immunogenicity of the recombinant lentivirus and stable expression of hNoc4L in RAW264.7 cells. The results showed that the recombinant lentivirus characterized a high transduction efficiency, long-term expression and low immunogenicity. Therefore, we pave the way for further identification of the biological activity of hNoc4L protein during ribosome biogenesis in mammalian. PMID:21284218

  6. The construction of support vector machine classifier using the firefly algorithm.

    PubMed

    Chao, Chih-Feng; Horng, Ming-Huwi

    2015-01-01

    The setting of parameters in the support vector machines (SVMs) is very important with regard to its accuracy and efficiency. In this paper, we employ the firefly algorithm to train all parameters of the SVM simultaneously, including the penalty parameter, smoothness parameter, and Lagrangian multiplier. The proposed method is called the firefly-based SVM (firefly-SVM). This tool is not considered the feature selection, because the SVM, together with feature selection, is not suitable for the application in a multiclass classification, especially for the one-against-all multiclass SVM. In experiments, binary and multiclass classifications are explored. In the experiments on binary classification, ten of the benchmark data sets of the University of California, Irvine (UCI), machine learning repository are used; additionally the firefly-SVM is applied to the multiclass diagnosis of ultrasonic supraspinatus images. The classification performance of firefly-SVM is also compared to the original LIBSVM method associated with the grid search method and the particle swarm optimization based SVM (PSO-SVM). The experimental results advocate the use of firefly-SVM to classify pattern classifications for maximum accuracy. PMID:25802511

  7. The Construction of Support Vector Machine Classifier Using the Firefly Algorithm

    PubMed Central

    Chao, Chih-Feng; Horng, Ming-Huwi

    2015-01-01

    The setting of parameters in the support vector machines (SVMs) is very important with regard to its accuracy and efficiency. In this paper, we employ the firefly algorithm to train all parameters of the SVM simultaneously, including the penalty parameter, smoothness parameter, and Lagrangian multiplier. The proposed method is called the firefly-based SVM (firefly-SVM). This tool is not considered the feature selection, because the SVM, together with feature selection, is not suitable for the application in a multiclass classification, especially for the one-against-all multiclass SVM. In experiments, binary and multiclass classifications are explored. In the experiments on binary classification, ten of the benchmark data sets of the University of California, Irvine (UCI), machine learning repository are used; additionally the firefly-SVM is applied to the multiclass diagnosis of ultrasonic supraspinatus images. The classification performance of firefly-SVM is also compared to the original LIBSVM method associated with the grid search method and the particle swarm optimization based SVM (PSO-SVM). The experimental results advocate the use of firefly-SVM to classify pattern classifications for maximum accuracy. PMID:25802511

  8. A polymerase chain reaction-based method for constructing a linear vector with site-specific DNA methylation.

    PubMed

    Arakawa, Toshiya; Ohta, Tohru; Abiko, Yoshihiro; Okayama, Miki; Mizoguchi, Itaru; Takuma, Taishin

    2011-09-15

    DNA methylation is an important epigenetic modification that leads to a wide variety of biological functions, including transcription, growth and development, and diseases associated with altered gene expression such as cancers. However, tools to insert site-specific methylation into DNA for analyzing epigenetic functions are limited. Here we describe a novel polymerase chain reaction (PCR)-based approach to provide site-specific DNA methylation at any site, including CpG or CpNpG islands. This method is simple and versatile, and it consists of four steps to construct the DNA methylation vector: (I) design and synthesis of methylated primers, (II) PCR amplification, (III) isolation of single-stranded DNA, and (IV) annealing and ligation of isolated single-stranded DNAs. First we produced and validated a linear green fluorescence protein (GFP) vector by this method. Next we applied this method to introduce methyl groups into the promoter of the cyclooxygenase-2 (COX-2) gene and found that site-specific DNA methylation at the CRE element significantly altered COX-2 gene expression. These results demonstrate that this PCR-based approach is useful for the analysis of biological functions that depend on DNA methylation. PMID:21669180

  9. Application of the extracellular alpha-amylase gene from Streptococcus bovis 148 to construction of a secretion vector for yogurt starter strains.

    PubMed

    Satoh, E; Ito, Y; Sasaki, Y; Sasaki, T

    1997-11-01

    Streptococcus thermophilus ATCC 19258, Lactobacillus delbrueckii subsp. bulgaricus T-11, and Lactococcus lactis subsp. lactis IL1403 were transformed with the alpha-amylase gene (amyA) from Streptococcus bovis 148 by using a wide host-range vector, and all the transformants secreted the alpha-amylase successfully. Since the promoter and the secretion signal of the amyA gene were functional in these strains, we constructed a secretion vector using the expression elements of amyA. Trials to secrete foreign enzymes in yogurt starter strains were performed using this novel secretion vector. PMID:9361445

  10. Application of the extracellular alpha-amylase gene from Streptococcus bovis 148 to construction of a secretion vector for yogurt starter strains.

    PubMed Central

    Satoh, E; Ito, Y; Sasaki, Y; Sasaki, T

    1997-01-01

    Streptococcus thermophilus ATCC 19258, Lactobacillus delbrueckii subsp. bulgaricus T-11, and Lactococcus lactis subsp. lactis IL1403 were transformed with the alpha-amylase gene (amyA) from Streptococcus bovis 148 by using a wide host-range vector, and all the transformants secreted the alpha-amylase successfully. Since the promoter and the secretion signal of the amyA gene were functional in these strains, we constructed a secretion vector using the expression elements of amyA. Trials to secrete foreign enzymes in yogurt starter strains were performed using this novel secretion vector. PMID:9361445

  11. Characterisation of Zygosaccharomyces rouxii centromeres and construction of first Z. rouxii centromeric vectors.

    PubMed

    Pribylova, Lenka; Straub, Marie-Laure; Sychrova, Hana; de Montigny, Jacky

    2007-01-01

    Zygosaccharomyces rouxii is a hemiascomycetous yeast known for its high osmotolerance, the basis of which still remains unknown. By exploring the Génolevures I database, four Z. rouxii fragments homologous to Saccharomyces cerevisiae centromeres were identified. Two of them were subjected to further analysis. Their function as centromeres in Z. rouxii was proved, and they were localized to Z. rouxii chromosomes II and VII, respectively. The species-specificity of centromeres was observed; plasmids with a Z. rouxii centromere were not recognized as centromeric in S. cerevisiae, and a S. cerevisiae centromere did not function as a centromere in Z. rouxii. Constructed plasmids bearing Z. rouxii centromeres serve as the first specific centromeric plasmids, and thus contribute to the so-far limited set of genetic tools needed to study the Z. rouxii specific features. PMID:17487563

  12. AFREET: HUMAN-INSPIRED SPATIO-SPECTRAL FEATURE CONSTRUCTION FOR IMAGE CLASSIFICATION WITH SUPPORT VECTOR MACHINES

    SciTech Connect

    S. PERKINS; N. HARVEY

    2001-02-01

    The authors examine the task of pixel-by-pixel classification of the multispectral and grayscale images typically found in remote-sensing and medical applications. Simple machine learning techniques have long been applied to remote-sensed image classification, but almost always using purely spectral information about each pixel. Humans can often outperform these systems, and make extensive use of spatial context to make classification decisions. They present AFREET: an SVM-based learning system which attempts to automatically construct and refine spatio-spectral features in a somewhat human-inspired fashion. Comparisons with traditionally used machine learning techniques show that AFREET achieves significantly higher performance. The use of spatial context is particularly useful for medical imagery, where multispectral images are still rare.

  13. Lactic acid bacteria: reviewing the potential of a promising delivery live vector for biomedical purposes.

    PubMed

    Cano-Garrido, Olivia; Seras-Franzoso, Joaquin; Garcia-Fruitós, Elena

    2015-01-01

    Lactic acid bacteria (LAB) have a long history of safe exploitation by humans, being used for centuries in food production and preservation and as probiotic agents to promote human health. Interestingly, some species of these Gram-positive bacteria, which are generally recognized as safe organisms by the US Food and Drug Administration (FDA), are able to survive through the gastrointestinal tract (GIT), being capable to reach and colonize the intestine, where they play an important role. Besides, during the last decades, an important effort has been done for the development of tools to use LAB as microbial cell factories for the production of proteins of interest. Given the need to develop effective strategies for the delivery of prophylactic and therapeutic molecules, LAB have appeared as an appealing option for the oral, intranasal and vaginal delivery of such molecules. So far, these genetically modified organisms have been successfully used as vehicles for delivering functional proteins to mucosal tissues in the treatment of many different pathologies including GIT related pathologies, diabetes, cancer and viral infections, among others. Interestingly, the administration of such microorganisms would suppose a significant decrease in the production cost of the treatments agents since being live organisms, such vectors would be able to autonomously amplify and produce and deliver the protein of interest. In this context, this review aims to provide an overview of the use of LAB engineered as a promising alternative as well as a safety delivery platform of recombinant proteins for the treatment of a wide range of diseases. PMID:26377321

  14. Construction of a genomic DNA library with a TA vector and its application in cloning of the phytoene synthase gene from the cyanobacterium Spirulina platensis M-135

    NASA Astrophysics Data System (ADS)

    Yoshikazu, Kawata; Shin-Ichi, Yano; Hiroyuki, Kojima

    1998-03-01

    An efficient and simple method for constructing a genomic DNA library using a TA cloning vector is presented. It is based on the sonicative cleavage of genomic DNA and modification of fragment ends with Taq DNA polymerase, followed by ligation using a TA vector. This method was applied for cloning of the phytoene synthase gene crt B from Spirulina platensis. This method is useful when genomic DNA cannot be efficiently digested with restriction enzymes, a problem often encountered during the construction of a genomic DNA library of cyanobacteria.

  15. Construction the model on the breast cancer survival analysis use support vector machine, logistic regression and decision tree.

    PubMed

    Chao, Cheng-Min; Yu, Ya-Wen; Cheng, Bor-Wen; Kuo, Yao-Lung

    2014-10-01

    The aim of the paper is to use data mining technology to establish a classification of breast cancer survival patterns, and offers a treatment decision-making reference for the survival ability of women diagnosed with breast cancer in Taiwan. We studied patients with breast cancer in a specific hospital in Central Taiwan to obtain 1,340 data sets. We employed a support vector machine, logistic regression, and a C5.0 decision tree to construct a classification model of breast cancer patients' survival rates, and used a 10-fold cross-validation approach to identify the model. The results show that the establishment of classification tools for the classification of the models yielded an average accuracy rate of more than 90% for both; the SVM provided the best method for constructing the three categories of the classification system for the survival mode. The results of the experiment show that the three methods used to create the classification system, established a high accuracy rate, predicted a more accurate survival ability of women diagnosed with breast cancer, and could be used as a reference when creating a medical decision-making frame. PMID:25119239

  16. Construction of an Escherichia coli-rhodococcus shuttle vector and plasmid transformation in Rhodococcus spp

    SciTech Connect

    Singer, M.E.V.; Finnerty, W.R.

    1988-02-01

    A plasmid transformation system for Rhodococcus sp. strain H13-A was developed by using an Escherichia coli-Rhodococcus shuttle plasmid constructed in this study. Rhodococcus sp. strain H13-A contains three cryptic indigenous plasmids, designated pMVS100, pMVS200, and pMVS300, of 75, 19.5, and 13.4 kilobases (kb), respectively. A 3.8-kb restriction fragment of pMVS300 was cloned into pIJ30, a 6.3-kb pBR322 derivative, containing the E. coli origin of replication (ori) and ampicillin resistance determinant (bla), as well as a Streptomyces gene for thiostrepton resistance, tsr. The resulting 10.1-kb recombinant plasmid, designated pMVS301, was isolated from E. coli DH1 (pMVS301) and transformed into Rhodococccus sp. strain AS-50, a derivative of strain H13-A. The cloned 3.8-kb fragment of Rhodococcus DNA in pMVS301 contains a Rhodococcus origin of replication, since the hybrid plasmid was capable of replication in both genera. The plasmid was identical in E. coli and Rhodococcus transformants as determined by restriction analysis and was maintained as a stable, independent replicon in both organisms. A restriction map demonstrated 14 unique restriction sites in pMVS301, some of which are potentially useful for molecular cloning in Rhodococcus spp. and other actinomycetes. This is the first report of plasmid transformation and of heterologous gene expression in a Rhodococcus sp.

  17. [Creation of DNA vaccine vector based on codon-optimized gene of rabies virus glycoprotein (G protein) with consensus amino acid sequence].

    PubMed

    Starodubova, E S; Kuzmenko, Y V; Latanova, A A; Preobrazhenskaya, O V; Karpov, V L

    2016-01-01

    An optimized design of the rabies virus glycoprotein (G protein) for use within DNA vaccines has been suggested. The design represents a territorially adapted antigen constructed taking into account glycoprotein amino acid sequences of the rabies viruses registered in the Russian Federation and the vaccine Vnukovo-32 strain. Based on the created consensus amino acid sequence, the nucleotide codon-optimized sequence of this modified glycoprotein was obtained and cloned into the pVAX1 plasmid (a vector of the last generation used in the creation of DNA vaccines). A twofold increase in this gene expression compared to the expression of the Vnukovo-32 strain viral glycoprotein gene in a similar vector was registered in the transfected cell culture. It has been demonstrated that the accumulation of modified G protein exceeds the number of the control protein synthesized using the plasmid with the Vnukovo-32 strain viral glycoprotein gene by 20 times. Thus, the obtained modified rabies virus glycoprotein can be considered to be a promising DNA vaccine antigen. PMID:27239860

  18. thyA as a Selection Marker in Construction of Food-Grade Host-Vector and Integration Systems for Streptococcus thermophilus

    PubMed Central

    Sasaki, Yasuko; Ito, Yoshiyuki; Sasaki, Takashi

    2004-01-01

    We constructed food-grade host-vector and integration systems for Streptococcus thermophilus by using a thymidylate synthase gene (thyA) as the selection marker. Two thyA genes, thyASt and thyALb, were cloned from S. thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, respectively. Thymidine-requiring mutants of S. thermophilus were obtained after successive cultures in the presence of trimethoprim, and one of them, TM1-1, was used as the host. Food-grade vectors were constructed by using either thyASt or thyALb as the selection marker. Transformants of TM1-1 created by using these vectors were selected for thymidine autotrophy as efficiently as for erythromycin resistance. By using the host-vector system developed in this way, a foreign amylase gene (amyA) was expressed in TM1-1 and was also integrated into the chromosome by use of a temperature-sensitive integration vector constructed with thyALb as the selection marker via a double-crossover event. The results obtained show that thyA is an efficient and safe selection marker for S. thermophilus that is suitable for food applications. PMID:15006818

  19. Enhanced efficacy of an AAV vector encoding chimeric, highly secreted acid alpha-glucosidase in glycogen storage disease type II.

    PubMed

    Sun, Baodong; Zhang, Haoyue; Benjamin, Daniel K; Brown, Talmage; Bird, Andrew; Young, Sarah P; McVie-Wylie, Alison; Chen, Y-T; Koeberl, Dwight D

    2006-12-01

    Glycogen storage disease type II (GSD-II; Pompe disease; MIM 232300) is an inherited muscular dystrophy caused by deficiency in the activity of the lysosomal enzyme acid alpha-glucosidase (GAA). We hypothesized that chimeric GAA containing an alternative signal peptide could increase the secretion of GAA from transduced cells and enhance the receptor-mediated uptake of GAA in striated muscle. The relative secretion of chimeric GAA from transfected 293 cells increased up to 26-fold. Receptor-mediated uptake of secreted, chimeric GAA corrected cultured GSD-II patient cells. High-level hGAA was sustained in the plasma of GSD-II mice for 24 weeks following administration of an AAV2/8 vector encoding chimeric GAA; furthermore, GAA activity was increased and glycogen content was significantly reduced in striated muscle and in the brain. Administration of only 1 x 10(10) vector particles increased GAA activity in the heart and diaphragm for >18 weeks, whereas 3 x 10(10) vector particles increased GAA activity and reduced glycogen content in the heart, diaphragm, and quadriceps. Furthermore, an AAV2/2 vector encoding chimeric GAA produced secreted hGAA for >12 weeks in the majority of treated GSD-II mice. Thus, chimeric, highly secreted GAA enhanced the efficacy of AAV vector-mediated gene therapy in GSD-II mice. PMID:16987711

  20. Indole diterpene synthetic studies. Total synthesis of (+)-nodulisporic acid F and construction of the heptacyclic cores of (+)-nodulisporic acids A and B and (-)-nodulisporic acid D.

    PubMed

    Smith, Amos B; Davulcu, Akin H; Cho, Young Shin; Ohmoto, Kazuyuki; Kürti, László; Ishiyama, Haruaki

    2007-06-22

    A first-generation strategy for construction of (+)-nodulisporic acids A (1) and B (2) is described. The strategy entails union of the eastern and western hemisphere subtargets via the indole synthesis protocol developed in our laboratory. Subsequent elaboration of rings E and F, however, revealed the considerable acid instability of the C(24) hydroxyl, thereby preventing further advancement. Nonetheless, preparation of the heptacyclic core of (+)-nodulisporic acids A and B, the total synthesis of (+)-nodulisporic acid F, the simplest member of the nodulisporic acid family, and elaboration of the heptacyclic core of (-)-nodulisporic acid D were achieved. PMID:17511507

  1. Construction of the mammalian expressing vector pEGFP-N1-P53 and its expression successful in chicken fibroblast cells and blastoderm.

    PubMed

    Song, Z; Li, Z H; Lei, X Q; Xu, T S; Zhang, X H; Li, Y J; Zhang, G M; Xi, S M; Yang, Y B; Wei, Z G

    2015-01-01

    The enhanced green fluorescent protein (EGFP) pEGFP-N1-P53 eukaryotic expression vector, which contains the human tumor suppressor p53, was constructed and transfected into chicken fibroblast cells and stage-X blastoderm to analyze the transfection efficiency. The complementary DNA of the human p53 gene was cloned by reverse transcription-polymerase chain reaction from human peripheral blood and inserted into the pEGFP-N1 vector by HindIII and BamHI double digestion. The pEGFP-N1-P53 vector was transfected into chicken embryo fibroblasts by Lipofectamine 2000 liposomes, and the transfection efficiency was analyzed by fluorescence microscope after 36 h of transfection. The stage-X blastoderm was also transfected by blastoderm injection using Lipofectamine 2000 liposomes at room temperature after 12-24 h; then hatching occurred until seventh day, and the transfection efficiency was analyzed by fluorescence microscope in the dead embryo. A total of 90 hatching eggs were transfected by the pEGFP-N1-P53 vector, and 20 chicken embryos expressed the reporter gene, which indicated that recombinant pEGFP-N1-P53 could be transfected and expressed in stage-X blastoderm by liposomes. Chicken embryo fibroblasts were transfected and expressed the reporter gene. The pEGFP-N1-P53 vector was constructed successfully and could be transfected and expressed in chicken embryo fibroblasts and stage-X blastoderms efficiently. PMID:25730031

  2. 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

  3. Biosynthesis of high molecular weight hyaluronic acid by Streptococcus zooepidemicus using oxygen vector and optimum impeller tip speed.

    PubMed

    Lai, Zee-Wei; Rahim, Raha Abdul; Ariff, Arbakariya B; Mohamad, Rosfarizan

    2012-09-01

    The potential use of n-dodecane and n-hexadecane as oxygen vectors for enhancing hyaluronic acid (HA) biosynthesis by Streptococcus zooepidemicus ATCC 39920 was investigated using a 2-L stirred-tank bioreactor equipped with helical ribbon or Rushton turbine impellers. The volumetric fraction of the oxygen vector influenced the gas-liquid volumetric oxygen transfer coefficient (K(L)a) positively. Batch HA fermentation with 1% (v/v) n-dodecane or 0.5% (v/v) n-hexadecane addition was carried out at different impeller tip speeds. Even though cell growth was lower in the fermentation with oxygen vector addition, the HA productivity and molecular weight were higher when compared to the fermentation without oxygen vector at low impeller tip speed. The highest HA concentration (4.25 gHA/l) and molecular weight (1.54 × 10(7) Da) were obtained when 0.5% (v/v) n-hexadecane and 0.785 m/s impeller tip speed of helical ribbon were used. PMID:22608992

  4. Lactic acid bacteria--20 years exploring their potential as live vectors for mucosal vaccination.

    PubMed

    Wyszyńska, Agnieszka; Kobierecka, Patrycja; Bardowski, Jacek; Jagusztyn-Krynicka, Elżbieta Katarzyna

    2015-04-01

    Lactic acid bacteria (LAB) are a diverse group of Gram-positive, nonsporulating, low G + C content bacteria. Many of them have been given generally regarded as safe status. Over the past two decades, intensive genetic and molecular research carried out on LAB, mainly Lactococcus lactis and some species of the Lactobacillus genus, has revealed new, potential biomedical LAB applications, including the use of LAB as adjuvants, immunostimulators, or therapeutic drug delivery systems, or as factories to produce therapeutic molecules. LAB enable immunization via the mucosal route, which increases effectiveness against pathogens that use the mucosa as the major route of entry into the human body. In this review, we concentrate on the encouraging application of Lactococcus and Lactobacillus genera for the development of live mucosal vaccines. First, we present the progress that has recently been made in the field of developing tools for LAB genetic manipulations, which has resulted in the successful expression of many bacterial, parasitic, and viral antigens in LAB strains. Next, we discuss the factors influencing the efficacy of the constructed vaccine prototypes that have been tested in various animal models. Apart from the research focused on an application of live LABs as carriers of foreign antigens, a lot of work has been recently done on the potential usage of nonliving, nonrecombinant L. lactis designated as Gram-positive enhancer matrix (GEM), as a delivery system for mucosal vaccination. The advantages and disadvantages of both strategies are also presented. PMID:25750046

  5. Static corrosion of construction materials exposed to superphosphoric acid made from various sources of phosphate rock

    SciTech Connect

    Nguyen, D.T.; McDonald, C.L.; McGill, K.E.

    1994-10-01

    Corrosion tests were performed with various construction materials, such as carbon steel, cast iron, stainless steels, nickel and nickel-based alloys, copper and its alloys, aluminum alloy, zirconium alloy, and tantalum, exposed to wet-process superphosphoric acids (approximately 70% P{sub 2}O{sub 5}) from all the suppliers in the United States and to a technical-grade (55% P{sub 2}O{sub 5}) acid made by the electric furnace process. The study was conducted in response to reports from pipe-reactor users of excessive corrosion by superphosphoric acids and electric furnace acid. Test temperatures were ambient (approximately 21{degrees}C or 70{degrees}F), 66{degrees}C (150{degrees}F), and 93{degrees}C (200{degrees}F). Test results showed that temperature was a significant factor in acid corrosivity. Electric furnace acid was more corrosive than the superphosphoric acids. Carbon steel, cast iron, and aluminum alloy were not resistant to either the superphosphoric acids or the electric furnace acid. Nickel-chromium (Ni-Cr) and nickel-molybdenum (Ni-Mo) based alloys and tantalum exhibited adequate corrosion resistance in the superphosphoric acids and the electric furnace acid. Stainless steels performed well in all test acids at all test temperatures with some exceptions in the electric furnace acid at 93{degrees}C. Zirconium alloy, copper and its alloys, pure nickel, and Monel 400 provided adequate corrosion resistance to all test acids at ambient temperature only.

  6. New Invertebrate Vectors of Okadaic Acid from the North Atlantic Waters--Portugal (Azores and Madeira) and Morocco.

    PubMed

    Silva, Marisa; Rodriguez, Inés; Barreiro, Aldo; Kaufmann, Manfred; Isabel Neto, Ana; Hassouani, Meryem; Sabour, Brahim; Alfonso, Amparo; Botana, Luis M; Vasconcelos, Vitor

    2015-12-01

    Okadaic acid and its analogues are potent phosphatase inhibitors that cause Diarrheic Shellfish Poisoning (DSP) through the ingestion of contaminated shellfish by humans. This group of toxins is transmitted worldwide but the number of poisoning incidents has declined over the last 20 years due to legislation and monitoring programs that were implemented for bivalves. In the summer of 2012 and 2013, we collected a total of 101 samples of 22 different species that were made up of benthic and subtidal organisms such echinoderms, crustaceans, bivalves and gastropods from Madeira, São Miguel Island (Azores archipelago) and the northwestern coast of Morocco. The samples were analyzed by UPLC-MS/MS. Our main objective was to detect new vectors for these biotoxins. We can report nine new vectors for these toxins in the North Atlantic: Astropecten aranciacus, Arbacia lixula, Echinaster sepositus, Holothuria sanctori, Ophidiaster ophidianus, Onchidella celtica, Aplysia depilans, Patella spp., and Stramonita haemostoma. Differences in toxin contents among the species were found. Even though low concentrations were detected, the levels of toxins that were present, especially in edible species, indicate the importance of these types of studies. Routine monitoring should be extended to comprise a wider number of vectors other than for bivalves of okadaic acid and its analogues. PMID:26670254

  7. New Invertebrate Vectors of Okadaic Acid from the North Atlantic Waters—Portugal (Azores and Madeira) and Morocco

    PubMed Central

    Silva, Marisa; Rodriguez, Inés; Barreiro, Aldo; Kaufmann, Manfred; Neto, Ana Isabel; Hassouani, Meryem; Sabour, Brahim; Alfonso, Amparo; Botana, Luis M.; Vasconcelos, Vitor

    2015-01-01

    Okadaic acid and its analogues are potent phosphatase inhibitors that cause Diarrheic Shellfish Poisoning (DSP) through the ingestion of contaminated shellfish by humans. This group of toxins is transmitted worldwide but the number of poisoning incidents has declined over the last 20 years due to legislation and monitoring programs that were implemented for bivalves. In the summer of 2012 and 2013, we collected a total of 101 samples of 22 different species that were made up of benthic and subtidal organisms such echinoderms, crustaceans, bivalves and gastropods from Madeira, São Miguel Island (Azores archipelago) and the northwestern coast of Morocco. The samples were analyzed by UPLC-MS/MS. Our main objective was to detect new vectors for these biotoxins. We can report nine new vectors for these toxins in the North Atlantic: Astropecten aranciacus, Arbacia lixula, Echinaster sepositus, Holothuria sanctori, Ophidiaster ophidianus, Onchidella celtica, Aplysia depilans, Patella spp., and Stramonita haemostoma. Differences in toxin contents among the species were found. Even though low concentrations were detected, the levels of toxins that were present, especially in edible species, indicate the importance of these types of studies. Routine monitoring should be extended to comprise a wider number of vectors other than for bivalves of okadaic acid and its analogues. PMID:26670254

  8. Lesion-dependent regulation of transgene expression in the rat brain using a human glial fibrillary acidic protein-lentiviral vector.

    PubMed

    Jakobsson, Johan; Georgievska, Biljana; Ericson, Cecilia; Lundberg, Cecilia

    2004-02-01

    The ability to regulate transgene expression will be crucial for development of gene therapy to the brain. The most commonly used systems are based on a transactivator in combination with a drug, e.g. the tetracycline-regulated system. Here we describe a different method of transgene regulation by the use of the human glial fibrillary acidic protein (GFAP) promoter. We constructed a lentiviral vector that directs transgene expression to astrocytes. Using toxin-induced lesions we investigated to what extent transgene expression could be regulated in accordance with the activation of the endogenous GFAP gene. In animals receiving excitotoxic lesions of the striatum we detected an eightfold increase of green fluorescent protein (GFP)-expressing cells. The vast majority of these cells did not divide, suggesting that the transgene was indeed regulated in a similar fashion as the endogenous GFAP gene. This finding will lead to the development of lentiviral vectors with autoregulatory capacities that may be very useful for gene therapy to the brain. PMID:14984426

  9. Construction of interference vector targeting Ep-CAM gene and its effects on colorectal cancer cell proliferation

    PubMed Central

    Qi, Yanmei; Zhou, Fengqiang; Zhang, Lu; Liu, Lei; Xu, Hong; Guo, Huiguang

    2015-01-01

    Background Prior study indicates that abnormal protein expression and functional changes in the development and progression of colorectal cancer is related to gene expression. The aim of this study was to construct an interference plasmid targeting the Ep-CAM gene and to investigate its effects on the proliferation of colorectal cancer cells. Methods In this study, HT-29 and HCT-116 colorectal cancer cell lines were selected as cell models. The double-stranded micro (mi)RNA oligo was inserted into the pcDNATM6.2-GW/EmGFPmiR vector, which is an expression of miRNA. Lipofectamine™ 2000 was used to transfer plasmid into the empty plasmid group (transfected pcDNATM6.2-GW/EmGFPmiR-neg) and the interference group (transfected pcDNATM6.2-GW/EmGFPmiR-Ep-CAM-1), respectively. Meanwhile, the nontransferred HT-29 and HCT-116 acts as the blank control group. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the transfection efficiency. Western blot was used to detect Ep-CAM protein expression. The cell proliferation in each group was detected by using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Results The results indicated that the Ep-CAM messenger (m)RNA expression in the interference group was lower significantly compared with that of the empty plasmid group and control group (P<0.01). Western blot analysis results showed that Ep-CAM protein expression was significantly lower in interference group compared with that of the empty plasmid group and the control group (P<0.01). MTT assay results demonstrated that the proliferation ability of cells in the interference group was significantly inhibited compared with the two other groups (P<0.05). Conclusion Silencing of Ep-CAM can significantly inhibit the proliferation of colorectal cancer cells. PMID:26028961

  10. Construction of a compatible Gateway-based co-expression vector set for expressing multiprotein complexes in E. coli.

    PubMed

    Salim, Loubna; Feger, Claire; Busso, Didier

    2016-11-01

    We report the construction of a versatile Gateway-based co-expression vector set for producing multiprotein complexes in Escherichia coli. The set consists of two groups of three vectors (pCoGW and pCo0GW), each having a specific antibiotic resistance gene, a compatible origin of replication and allowing cloning of up to two genes, each under control of its own T7 promoter. To validate the set, 33 (co-)expression plasmids encoding fluorescent protein (GFP, DsRed and ECFP) have been generated. Protein expression levels were quantified and (co-)expression visualized by fluorescent microscopy. The results illustrate the applicability of these vectors in co-expression studies. PMID:27558914

  11. Construction of a cyanobacterium synthesizing cyclopropane fatty acids.

    PubMed

    Machida, Shuntaro; Shiraiwa, Yoshihiro; Suzuki, Iwane

    2016-09-01

    Microalgae have received much attention as a next-generation source of biomass energy. However, most of the fatty acids (FAs) from microalgae are multiply unsaturated; thus, the biofuels derived from them are fluid, but vulnerable to oxidation. In this study, we attempted to synthesize cyclopropane FAs in the cyanobacterium Synechocystis sp. PCC 6803 by expressing the cfa gene for cyclopropane FA synthase from Escherichia coli with the aim of producing FAs that are fluid and stable in response to oxidization. We successfully synthesized cyclopropane FAs in Synechocystis with a yield of ~30% of total FAs. Growth of the transformants was altered, particularly at low temperatures, but photosynthesis and respiration were not significantly affected. C16:1(∆9) synthesis in the desA(-)/desD(-) strain by expression of the desC2 gene for sn-2 specific ∆9 desaturase positively affected growth at low temperatures via promotion of various cellular processes, with the exceptions of photosynthesis and respiration. Estimation of the apparent activities of desaturases suggested that some acyl-lipid desaturases might recognize the lipid side chain. PMID:27263419

  12. Characterization of a minimal pKW2124 replicon from Weissella cibaria KLC140 and its application for the construction of the Weissella expression vector pKUCm1.

    PubMed

    Ku, Hye-Jin; Park, Myeong Soo; Lee, Ju-Hoon

    2015-01-01

    A 2.1-kb plasmid was previously isolated from Weissella cibaria KLC140 in kimchi and cloned into pUC19 along with the slpA and gfp genes, resulting in an 8.6-kb pKWCSLGFP construct for use as a novel surface display vector. To reduce the size of the vector, the minimal replicon of pKW2124 was determined. The pKW2124 plasmid contains a putative origin of replication (ori), a potential ribosomal binding site (RBS), and the repA gene encoding a plasmid replication protein. To conduct the minimal replicon experiment, four different PCR products (MR1, ori+RBS+repA; MR2, RBS+repA; MR2', repA; MR3, fragment of repA) were obtained and cloned into pUC19 (pKUCm1, pKUCm2, pKUCm2', and pKUCm3, respectively) containing the chloramphenicol acetyltransferase (CAT) gene. These constructed vectors were electroporated into W. confusa ATCC 10881 with different transformation efficiencies of 1.5 × 10(5) CFU/μg, 1.3 × 10(1) CFU/μg, and no transformation, respectively, suggesting that the putative ori, RBS, and repA gene are essential for optimum plasmid replication. Subsequent segregational plasmid stability testing of pKUCm1 and pKUCm2 showed that the vector pKUCm1 is highly stable up to 100 generations but pKUCm2 was completely lost after 60 generations, suggesting that the putative ori may be important for plasmid stability in the host strain. In addition, a host range test of pKUCm1 revealed that it has a broad host range spectrum including Weissella, Lactococcus, Leuconostoc, and even Lactobacillus. To verify the application of pKUCm1, the β-galactosidase gene and its promoter region from W. cibaria KSD1 were cloned in the vector, resulting in pKUGal. Expression of the β-galactosidase gene was confirmed using blue-white screening after IPTG induction. The small and stable pKUGal vector will be useful for gene transfer, expression, and manipulation in the Weissella genome and in other lactic acid bacteria. PMID:25691882

  13. Characterization of a minimal pKW2124 replicon from Weissella cibaria KLC140 and its application for the construction of the Weissella expression vector pKUCm1

    PubMed Central

    Ku, Hye-Jin; Park, Myeong Soo; Lee, Ju-Hoon

    2015-01-01

    A 2.1-kb plasmid was previously isolated from Weissella cibaria KLC140 in kimchi and cloned into pUC19 along with the slpA and gfp genes, resulting in an 8.6-kb pKWCSLGFP construct for use as a novel surface display vector. To reduce the size of the vector, the minimal replicon of pKW2124 was determined. The pKW2124 plasmid contains a putative origin of replication (ori), a potential ribosomal binding site (RBS), and the repA gene encoding a plasmid replication protein. To conduct the minimal replicon experiment, four different PCR products (MR1, ori+RBS+repA; MR2, RBS+repA; MR2’, repA; MR3, fragment of repA) were obtained and cloned into pUC19 (pKUCm1, pKUCm2, pKUCm2’, and pKUCm3, respectively) containing the chloramphenicol acetyltransferase (CAT) gene. These constructed vectors were electroporated into W. confusa ATCC 10881 with different transformation efficiencies of 1.5 × 105 CFU/μg, 1.3 × 101 CFU/μg, and no transformation, respectively, suggesting that the putative ori, RBS, and repA gene are essential for optimum plasmid replication. Subsequent segregational plasmid stability testing of pKUCm1 and pKUCm2 showed that the vector pKUCm1 is highly stable up to 100 generations but pKUCm2 was completely lost after 60 generations, suggesting that the putative ori may be important for plasmid stability in the host strain. In addition, a host range test of pKUCm1 revealed that it has a broad host range spectrum including Weissella, Lactococcus, Leuconostoc, and even Lactobacillus. To verify the application of pKUCm1, the β-galactosidase gene and its promoter region from W. cibaria KSD1 were cloned in the vector, resulting in pKUGal. Expression of the β-galactosidase gene was confirmed using blue-white screening after IPTG induction. The small and stable pKUGal vector will be useful for gene transfer, expression, and manipulation in the Weissella genome and in other lactic acid bacteria. PMID:25691882

  14. Vector-Mediated Delivery of a Polyamide ("Peptide") Nucleic Acid Analogue through the Blood-Brain Barrier in vivo

    NASA Astrophysics Data System (ADS)

    Pardridge, William M.; Boado, Ruben J.; Kang, Young-Sook

    1995-06-01

    Polyamide ("peptide") nucleic acids (PNAs) are molecules with antigene and antisense effects that may prove to be effective neuropharmaceuticals if these molecules are enabled to undergo transport through the brain capillary endothelial wall, which makes up the blood-brain barrier in vivo. The model PNA used in the present studies is an 18-mer that is antisense to the rev gene of human immunodeficiency virus type 1 and is biotinylated at the amino terminus and iodinated at a tyrosine residue near the carboxyl terminus. The biotinylated PNA was linked to a conjugate of streptavidin (SA) and the OX26 murine monoclonal antibody to the rat transferrin receptor. The blood-brain barrier is endowed with high transferrin receptor concentrations, enabling the OX26-SA conjugate to deliver the biotinylated PNA to the brain. Although the brain uptake of the free PNA was negligible following intravenous administration, the brain uptake of the PNA was increased at least 28-fold when the PNA was bound to the OX26-SA vector. The brain uptake of the PNA bound to the OX26-SA vector was 0.1% of the injected dose per gram of brain at 60 min after an intravenous injection, approximating the brain uptake of intravenously injected morphine. The PNA bound to the OX26-SA vector retained the ability to bind to synthetic rev mRNA as shown by RNase protection assays. In summary, the present studies show that while the transport of PNAs across the blood-brain barrier is negligible, delivery of these potential neuropharmaceutical drugs to the brain may be achieved by coupling them to vector-mediated peptide-drug delivery systems.

  15. Toxicity of aristolochic acids isolated from Aristolochia indica Linn (Aristolochiaceae) against the malarial vector Anopheles stephensi Liston (Diptera: Culicidae).

    PubMed

    Pradeepa, Venkatraman; Sathish-Narayanan, Subbiah; Kirubakaran, Suyambulingam Arunachalam; Thanigaivel, Annamalai; Senthil-Nathan, Sengottayan

    2015-06-01

    With the growth of resistance to overused insecticides, vector management has become highly problematic. Hence more concentration has been focused on botanicals. Therefore our present study was aimed to evaluate the toxicity of compounds, aristolochic acid I and aristolochic acid II from the methanol extract of Aristolochia indica L. (Aristolochiaceae) leaves on larvae of Anopheles stephensi L. (Diptera: Culicidae) employing World Health Organization standard larvicide testing procedures. The soxhlet extraction was carried out using polar solvent, methanol. The isolated toxic compounds were purified through RP-HPLC. The FTIR spectroscopic studies revealed different peak values with functional groups in the mixed compounds (AA-I and AA- II). These two aristolochic acids were further studied through (13)C and (1)HNMR analysis with confirmed by structures. Bioassay-guided fractionation through flash chromatography lead to the isolation of two larvicidal compounds namely aristolochic acid I and II. In these bioassays, the larvae were exposed to concentrations of 100, 250, 500,750 and 1000 ppm for each compound. Between the two, AA-I exerted no significant toxicity difference (P < 0.05) on mosquito larvae with LC₅₀- 171.3, 209.8, 269.1, 502.3 ppm and LC₉₀-751.6, 963.8, 972.7, 990.8 ppm compared to AA-II with LC₅₀-134.8, 166.7, 240.4,543.2 ppm and LC₉₀- 636.7, 792.5, 990.8, 986.2 ppm against first, second, third and fourth instars, respectively. Further, the isolated compounds were severely affecting the mosquito gut. From the results, A. indica toxic compounds could be considered as one of the influential applicant to bring about useful botanicals so as to prevent the resurrection of mosquito vectors. PMID:25660198

  16. Determination of citric acid of lemon vinegar using visible/near infrared spectroscopy and least squares-support vector machine

    NASA Astrophysics Data System (ADS)

    Liu, Fei; Wang, Li; He, Yong

    2008-11-01

    The determination of citric acid of lemon vinegar was processed using visible and near infrared (Vis/NIR) spectroscopy combined with least squares-support vector machine (LS-SVM). Five concentration levels (100%, 80%, 60%, 40% and 20%) of lemon vinegar were studied. The calibration set was consisted of 225 samples (45 samples for each level) and the remaining 75 samples for the validation set. Partial least squares (PLS) analysis was employed for the calibration models as well as extraction of certain latent variables (LVs) and effective wavelengths (EWs). Different preprocessing methods were compared in PLS models including smoothing, standard normal variate (SNV), the first and second derivative. The selected LVs and EWs were employed as the inputs to develop least square-support vector machine (LSSVM) models. The optimal prediction results were achieved by LV-LS-SVM model, and the correlation coefficient (r), root mean square error of prediction (RMSEP) and bias for validation set were 0.9990, 0.1972 and -0.0334, respectively. Moreover, the EW-LS-SVM model was also acceptable and slightly better than all PLS models. The results indicated that Vis/NIR spectroscopy could be utilized as a parsimonious and efficient way for the determination of citric acid of lemon vinegar based on LS-SVM method.

  17. [Construction and expression of a eukaryotic vector co-expressing immunodominant antigens of CFP10, ESAT6, Ag85A and Ag85B of Mycobacterium tuberculosis].

    PubMed

    Li, Wu; Deng, Guangcun; Liu, Xiaoming; Wang, Yujiong

    2014-02-01

    CFP10, ESAT6, Antigen 85A (Ag85A) and antigen 85B (Ag85B) are the key immunodominant antigens of Mycobacterium tuberculosis. In order to construct a eukaryotic vector able to co-express the four genes in one vector, we amplified the target gene fragments encoding the CFP10, ESAT6, Ag85A and Ag85B antigens and inserted them into the multicloning site of the shuttle plasmid vector pcDNA3.1 (+), of which the CFP10 and ESAT6 encoding genes were in frame fused with a linker encoding (Gly4Ser)3 residue, before the fused gene was inserted downstream of CMV promoter with a bovine growth hormone poly A(BGH pA) sequence at the 3'-end; Ag85A and Ag85B encoding genes were fused with a separation of internal ribosome entry site (IRES) sequence before the fused gene cassette was inserted downstream of RSV promoter with a BGH pA sequence at the 3'-end. The final plasmid containing all four genes was confirmed by sequence analysis and designated as pcDNA-CFP10-ESAT6-Ag85A-Ag85B (pcDNA-CEAB). In order to verify the ability of this construct to express target proteins, we then transfected the recombinant plasmid into Human embryonic kidney (HEK) 293T cells and harvested the cell lysates, and the cell lysates were then separated by SDS-PAGE and subjected to Western blot analysis 48 h after transfection. All four of the target proteins were detected in the cell lysates against the respective specific antibodies, suggesting that we have successfully constructed a eukaryotic vector co-expressing the four immunodominant antigens of Mycobacterium tuberculosis, which lay a foundation for the further study of the immunogenicity and protective activity of the four antigens. PMID:24941747

  18. A chiral phosphoric acid catalyst for asymmetric construction of 1,3-dioxanes.

    PubMed

    Matsumoto, Akira; Asano, Keisuke; Matsubara, Seijiro

    2015-07-25

    A novel method of enantioselective 1,3-dioxane construction via a hemiacetalization/intramolecular oxy-Michael addition cascade by a chiral phosphoric acid catalyst was developed. The product was successfully transformed into an optically active 1,3-polyol motif, indicating that the proposed reaction can provide useful chiral building blocks for the de novo synthesis of polyketides. PMID:26103581

  19. Stormwater ponds, constructed wetlands, and other best management practices as potential breeding sites for West Nile virus vectors in Delaware during 2004.

    PubMed

    Gingrich, Jack B; Anderson, Robert D; Williams, Gregory M; O'Connor, Linda; Harkins, Kevin

    2006-06-01

    We performed longitudinal surveys of mosquito larval abundance (mean mosquito larvae per dip) in 87 stormwater ponds and constructed wetland in Delaware from June to September 2004. We analyzed selected water quality factors, water depth, types of vegetation, degree of shade, and level of insect predation in relation to mosquito abundance. The 2004 season was atypical, with most ponds remaining wet for the entire summer. In terms of West Nile virus (WNV) vectors, wetlands predominantly produce Aedes vexans, culex pipiens pipiens, and Culex restuans. Retention ponds generally produced the same species as wetlands, except that Cx. p. pipiens was more abundant than Cx. restuans in retention ponds. Aedes vexans and Culex salinarius were the most abundant species to Conservation Restoration Enhancement Program ponds. Sand filters uniquely produced high numbers of Cx. restuans, Cx. p. pipiens, and Aedes japonicus japonicus, a newly invasive vector species. Site that alternately dried and flooded, mostly detention ponds, forebays of retention ponds, and some wetlands often produced Ae. vexans, an occasional WNV bridge vector species. Overall, seasonal distribution of vectors was bimodal, with peaks occurring during early and late summer. Ponds with shallow sides and heavy shade generally produced an abundance of mosquitoes, unless insect predators were abundant. Bright, sunny ponds with steep sides and little vegetation generally produced the fewest mosquitoes. The associations among mosquito species and selected vegetation types are discussed. PMID:17019774

  20. Formic and Acetic Acids in Degradation Products of Plant Volatiles Elicit Olfactory and Behavioral Responses from an Insect Vector.

    PubMed

    George, Justin; Robbins, Paul S; Alessandro, Rocco T; Stelinski, Lukasz L; Lapointe, Stephen L

    2016-05-01

    Volatile phytochemicals play a role in orientation by phytophagous insects. We studied antennal and behavioral responses of the Asian citrus psyllid, Diaphorina citri, vector of the citrus greening disease pathogen. Little or no response to citrus leaf volatiles was detected by electroantennography. Glass cartridges prepared with β-ocimene or citral produced no response initially but became stimulatory after several days. Both compounds degraded completely in air to a number of smaller molecules. Two peaks elicited large antennal responses and were identified as acetic and formic acids. Probing by D. citri of a wax substrate containing odorants was significantly increased by a blend of formic and acetic acids compared with either compound separately or blends containing β-ocimene and/or citral. Response surface modeling based on a 4-component mixture design and a 2-component mixture-amount design predicted an optimal probing response on wax substrate containing a blend of formic and acetic acids. Our study suggests that formic and acetic acids play a role in host selection by D. citri and perhaps by phytophagous insects in general even when parent compounds from which they are derived are not active. These results have implications for the investigation of arthropod olfaction and may lead to elaboration of attract-and-kill formulations to reduce nontarget effects of chemical control in agriculture. PMID:26857741

  1. Detection of Glutamic Acid in Oilseed Rape Leaves Using Near Infrared Spectroscopy and the Least Squares-Support Vector Machine

    PubMed Central

    Bao, Yidan; Kong, Wenwen; Liu, Fei; Qiu, Zhengjun; He, Yong

    2012-01-01

    Amino acids are quite important indices to indicate the growth status of oilseed rape under herbicide stress. Near infrared (NIR) spectroscopy combined with chemometrics was applied for fast determination of glutamic acid in oilseed rape leaves. The optimal spectral preprocessing method was obtained after comparing Savitzky-Golay smoothing, standard normal variate, multiplicative scatter correction, first and second derivatives, detrending and direct orthogonal signal correction. Linear and nonlinear calibration methods were developed, including partial least squares (PLS) and least squares-support vector machine (LS-SVM). The most effective wavelengths (EWs) were determined by the successive projections algorithm (SPA), and these wavelengths were used as the inputs of PLS and LS-SVM model. The best prediction results were achieved by SPA-LS-SVM (Raw) model with correlation coefficient r = 0.9943 and root mean squares error of prediction (RMSEP) = 0.0569 for prediction set. These results indicated that NIR spectroscopy combined with SPA-LS-SVM was feasible for the fast and effective detection of glutamic acid in oilseed rape leaves. The selected EWs could be used to develop spectral sensors, and the important and basic amino acid data were helpful to study the function mechanism of herbicide. PMID:23203052

  2. Foamy virus vectors.

    PubMed Central

    Russell, D W; Miller, A D

    1996-01-01

    Human foamy virus (HFV) is a retrovirus of the spumavirus family. We have constructed vectors based on HFV that encode neomycin phosphotransferase and alkaline phosphatase. These vectors are able to transduce a wide variety of vertebrate cells by integration of the vector genome. Unlike vectors based on murine leukemia virus, HFV vectors are not inactivated by human serum, and they transduce stationary-phase cultures more efficiently than murine leukemia virus vectors. These properties, as well as their large packaging capacity, make HFV vectors promising gene transfer vehicles. PMID:8523528

  3. Amphiphilic calixresorcinarene associates as effective solubilizing agents for hydrophobic organic acids: construction of nano-aggregates.

    PubMed

    Morozova, Ju E; Syakaev, V V; Kazakova, E Kh; Shalaeva, Ya V; Nizameev, I R; Kadirov, M K; Voloshina, A D; Zobov, V V; Konovalov, A I

    2016-07-01

    Here we represent the first example of the formation of mixed nanoscale associates, constructed from amphiphilic calixresorcinarenes and hydrophobic carboxylic acids including drugs. The amidoamino-calixresorcinarene self-associates effectively solubilize hydrophobic carboxylic acids - drugs such as naproxen, ibuprofen, ursodeoxycholic acid and aliphatic dodecanoic acid - with the formation of the mixed aggregates with the macrocycle/substrate stoichiometry from 1/1 to 1/7. The ionization of organic acids and the peripheral nitrogen atoms of the macrocycles with the subsequent inclusion of hydrophobic acids into the macrocycle self-associates is the driving force of solubilization. In some cases, this leads to the co-assembly of the macrocycle polydisperse associates into supramolecular monodisperse nanoparticles with the diameter of about 100 nm. The efficiency of drug loading into the nanoparticles is up to 45% and depends on the structure of organic acid. The dissociation of the mixed aggregates and release of organic acid are attained by decreasing pH. PMID:27252123

  4. The use of the replication region of plasmid pRS7 from Oenococcus oeni as a putative tool to generate cloning vectors for lactic acid bacteria.

    PubMed

    Rodríguez, M Carmen; Alegre, M Teresa; Martín, M Cruz; Mesas, Juan M

    2015-01-01

    A chimeric plasmid, pRS7Rep (6.1 kb), was constructed using the replication region of pRS7, a large plasmid from Oenococcus oeni, and pEM64, a plasmid derived from pIJ2925 and containing a gene for resistance to chloramphenicol. pRS7Rep is a shuttle vector that replicates in Escherichia coli using its pIJ2925 component and in lactic acid bacteria (LAB) using the replication region of pRS7. High levels of transformants per µg of DNA were obtained by electroporation of pRS7Rep into Pediococcus acidilactici (1.5 × 10(7)), Lactobacillus plantarum (5.7 × 10(5)), Lactobacillus casei (2.3 × 10(5)), Leuconostoc citreum (2.7 × 10(5)), and Enterococcus faecalis (2.4 × 10(5)). A preliminary optimisation of the technical conditions of electrotransformation showed that P. acidilactici and L. plantarum are better transformed at a later exponential phase of growth, whereas L. casei requires the early exponential phase for better electrotransformation efficiency. pRS7Rep contains single restriction sites useful for cloning purposes, BamHI, XbaI, SalI, HincII, SphI and PstI, and was maintained at an acceptable rate (>50%) over 100 generations without selective pressure in L. plantarum, but was less stable in L. casei and P. acidilactici. The ability of pRS7Rep to accept and express other genes was assessed. To the best of our knowledge, this is the first time that the replication region of a plasmid from O. oeni has been used to generate a cloning vector. PMID:25479060

  5. 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

  6. Construction of a recombinant human insulin expression vector for mammary gland-specific expression in buffalo (Bubalus bubalis) mammary epithelial cell line.

    PubMed

    Kaushik, Ramakant; Singh, Karn Pratap; Kumari, Archana; Rameshbabu, K; Singh, Manoj Kumar; Manik, Radhey Shyam; Palta, Prabhat; Singla, Suresh Kumar; Chauhan, Manmohan Singh

    2014-09-01

    The aim of the present study was construction of mammary gland specific expression vector for high level of human insulin (hINS) expression in transgenic buffalo for therapeutic use. We have constructed mammary gland specific vector containing human insulin gene and there expression efficiency was checked into in vitro cultured buffalo mammary epithelial cells (BuMECs). Human pro-insulin coding region was isolated from human genomic DNA by intron skipping PCR primer and furin cleavage site was inserted between B-C and C-A chain of human insulin by overlap extension PCR. A mammary gland-specific buffalo beta-lactoglobulin promoter was isolated from buffalo DNA and used for human insulin expression in BuMEC cells. The construct was transfected into BuMECs by lipofection method and positive transgene cell clones were obtained by G418 selection after 3 weeks. Expression of hINS in transfected cells were confirmed by RT-PCR, Immunocytochemistry, Western Blotting and ELISA. The pAcISUBC insulin-expressing clones secreted insulin at varying levels between 0.18 - 1.43 ng/ml/24 h/2.0 × 10(6) cells. PMID:24969480

  7. Sialic Acid Expression in the Mosquito Aedes aegypti and Its Possible Role in Dengue Virus-Vector Interactions

    PubMed Central

    Cime-Castillo, Jorge; Delannoy, Philippe; Mendoza-Hernández, Guillermo; Monroy-Martínez, Verónica; Lanz-Mendoza, Humberto; Hernández-Hernández, Fidel de la Cruz; Cabello-Gutiérrez, Carlos; Ruiz-Ordaz, Blanca H.

    2015-01-01

    Dengue fever (DF) is the most prevalent arthropod-borne viral disease which affects humans. DF is caused by the four dengue virus (DENV) serotypes, which are transmitted to the host by the mosquito Aedes aegypti that has key roles in DENV infection, replication, and viral transmission (vector competence). Mosquito saliva also plays an important role during DENV transmission. In this study, we detected the presence of sialic acid (Sia) in Aedes aegypti tissues, which may have an important role during DENV-vector competence. We also identified genome sequences encoding enzymes involved in Sia pathways. The cDNA for Aedes aegypti CMP-Sia synthase (CSAS) was amplified, cloned, and functionally evaluated via the complementation of LEC29.Lec32 CSAS-deficient CHO cells. AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface. Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome. Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues. In addition, we evaluated the possible role of sialylated molecules in a salivary gland extract during DENV internalization in mammalian cells. The knowledge of early DENV-host interactions could facilitate a better understanding of viral tropism and pathogenesis to allow the development of new strategies for controlling DENV transmission. PMID:25874215

  8. Sialic acid expression in the mosquito Aedes aegypti and its possible role in dengue virus-vector interactions.

    PubMed

    Cime-Castillo, Jorge; Delannoy, Philippe; Mendoza-Hernández, Guillermo; Monroy-Martínez, Verónica; Harduin-Lepers, Anne; Lanz-Mendoza, Humberto; Hernández-Hernández, Fidel de la Cruz; Zenteno, Edgar; Cabello-Gutiérrez, Carlos; Ruiz-Ordaz, Blanca H

    2015-01-01

    Dengue fever (DF) is the most prevalent arthropod-borne viral disease which affects humans. DF is caused by the four dengue virus (DENV) serotypes, which are transmitted to the host by the mosquito Aedes aegypti that has key roles in DENV infection, replication, and viral transmission (vector competence). Mosquito saliva also plays an important role during DENV transmission. In this study, we detected the presence of sialic acid (Sia) in Aedes aegypti tissues, which may have an important role during DENV-vector competence. We also identified genome sequences encoding enzymes involved in Sia pathways. The cDNA for Aedes aegypti CMP-Sia synthase (CSAS) was amplified, cloned, and functionally evaluated via the complementation of LEC29.Lec32 CSAS-deficient CHO cells. AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface. Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome. Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues. In addition, we evaluated the possible role of sialylated molecules in a salivary gland extract during DENV internalization in mammalian cells. The knowledge of early DENV-host interactions could facilitate a better understanding of viral tropism and pathogenesis to allow the development of new strategies for controlling DENV transmission. PMID:25874215

  9. Application of retinoic acid improves form and function of tissue engineered corneal construct

    PubMed Central

    Abidin, Fadhilah Z; Gouveia, Ricardo M; Connon, Che J

    2015-01-01

    ABSTRACT Retinoic acid has recently been shown to control the phenotype and extracellular matrix composition of corneal stromal cells cultured in vitro as monolayers. This study set out to investigate the effects of retinoic acid on human corneal keratocytes within a 3D environment. Human corneal keratocytes were encapsulated in collagen gels, which were subsequently compressed under load, and cultured in serum-free media supplemented with 10 µM retinoic acid or DMSO vehicle for 30 days. Cell proliferation was quantified on selected days, while the expression of several important keratocytes markers was evaluated at day 30 using RT-PCR and immunoblotting. The weight and size of the collagen constructs were measured before and after hydration and contraction analyses. Retinoic acid enhanced keratocyte proliferation until day 30, whereas cells in control culture conditions showed reduced numbers after day 21. Both gene and protein expressions of keratocyte-characteristic proteoglycans (keratocan, lumican and decorin), corneal crystallins and collagen type I and V were significantly increased following retinoic acid supplementation. Retinoic acid also significantly reduced the expression of matrix metalloproteases 1, 3 and 9 while not increasing α-smooth muscle actin and fibronectin expression. Furthermore, these effects were also correlated with the ability of retinoic acid to significantly inhibit the contractility of keratocytes while allowing the build-up of corneal stromal extracellular matrix within the 3D constructs. Thus, retinoic acid supplementation represents a promising strategy to improve the phenotype of 3D-cultured keratocytes, and their usefulness as a model of corneal stroma for corneal biology and regenerative medicine applications. PMID:26496651

  10. Application of retinoic acid improves form and function of tissue engineered corneal construct.

    PubMed

    Abidin, Fadhilah Z; Gouveia, Ricardo M; Connon, Che J

    2015-01-01

    Retinoic acid has recently been shown to control the phenotype and extracellular matrix composition of corneal stromal cells cultured in vitro as monolayers. This study set out to investigate the effects of retinoic acid on human corneal keratocytes within a 3D environment. Human corneal keratocytes were encapsulated in collagen gels, which were subsequently compressed under load, and cultured in serum-free media supplemented with 10 µM retinoic acid or DMSO vehicle for 30 days. Cell proliferation was quantified on selected days, while the expression of several important keratocytes markers was evaluated at day 30 using RT-PCR and immunoblotting. The weight and size of the collagen constructs were measured before and after hydration and contraction analyses. Retinoic acid enhanced keratocyte proliferation until day 30, whereas cells in control culture conditions showed reduced numbers after day 21. Both gene and protein expressions of keratocyte-characteristic proteoglycans (keratocan, lumican and decorin), corneal crystallins and collagen type I and V were significantly increased following retinoic acid supplementation. Retinoic acid also significantly reduced the expression of matrix metalloproteases 1, 3 and 9 while not increasing α-smooth muscle actin and fibronectin expression. Furthermore, these effects were also correlated with the ability of retinoic acid to significantly inhibit the contractility of keratocytes while allowing the build-up of corneal stromal extracellular matrix within the 3D constructs. Thus, retinoic acid supplementation represents a promising strategy to improve the phenotype of 3D-cultured keratocytes, and their usefulness as a model of corneal stroma for corneal biology and regenerative medicine applications. PMID:26496651

  11. Construction of Expression Vector for Anti-Alpha-Fetoprotein Gene and Its Inhibition Effects on Alpha-Fetoprotein Positive Hepg2 Cells

    NASA Astrophysics Data System (ADS)

    Wang, Ze; Zhang, Hui

    As research previously demonstrated, suppression of AFP expression or its biological activities might inhibit the proliferation of AFP positive human hepatocellular carcinoma cells. In this study, we constructed an anti-AFP gene vector and transfected it to HepG2 cells. RT-PCR showed AFP gene expression in the transfected cells was reduced. MTT assay suggested the proliferation of the transfected cells was also inhibited comparing with the untransfected cells. This result provides a new insight into AFP as the target for preventing and treating hepatocellular carcinoma.

  12. Formulation of a Peptide Nucleic Acid Based Nucleic Acid Delivery Construct

    PubMed Central

    Millili, Peter G.; Yin, Daniel H.; Fan, Haihong; Naik, Ulhas P.; Sullivan, Millicent O.

    2010-01-01

    Gene delivery biomaterials need to be designed to efficiently achieve nuclear delivery of plasmid DNA. Polycations have been used to package DNA and other nucleic acids within sub-micron sized particles, offering protection from shear-induced or enzymatic degradation. However, cytotoxicity issues coupled with limited in vivo transfection efficiencies minimize the effectiveness of this approach. In an effort to improve upon existing technologies aimed at delivering nucleic acids, an alternative approach to DNA packaging was explored. Peptide nucleic acids (PNAs) were used to directly functionalize DNA with poly(ethylene glycol) (PEG) chains that provide a steric layer and inhibit multimolecular aggregation during complexation. DNA prePEGylation by this strategy was predicted to enable the formation of more homogeneous and efficiently packaged polyplexes. In this work, DNA-PNA-peptide-PEG (DP3) conjugates were synthesized and self-assembled with 25 kDa poly(ethylenimine) (PEI). Complexes with small standard deviations and average diameters ranging from 30 – 50 nm were created, with minimal dependence of complex size on N:P ratio (PEI amines to DNA phosphates). Furthermore, PEI-DNA interactions were altered by the derivitization strategy, resulting in tighter compaction of the PEI-DP3 complexes in comparison with PEI-DNA complexes. Transfection experiments in Chinese Hamster Ovary (CHO) cells revealed comparable transfection efficiencies but reduced cytotoxicities of the PEI-DP3 complexes relative to PEI-DNA complexes. The enhanced cellular activities of the PEI-DP3 complexes were maintained following the removal of free PEI from the PEI-DP3 formulations, whereas the cellular activity of the conventional PEI-DNA formulations was reduced by free PEI removal. These findings suggest that DNA prePEGylation by the PNA-based strategy might provide a way to circumvent cytotoxicity and formulation issues related to the use of PEI for in vivo gene delivery. PMID:20131756

  13. MATRIX AND VECTOR SERVICES

    2001-10-18

    PETRA V2 provides matrix and vector services and the ability construct, query, and use matrix and vector objects that are used and computed by TRILINOS solvers. It provides all basic matr5ix and vector operations for solvers in TRILINOS.

  14. Elucidation of Insertion Elements Carried on Plasmids and In Vitro Construction of Shuttle Vectors from the Toxic Cyanobacterium Planktothrix

    PubMed Central

    Christiansen, Guntram; Goesmann, Alexander

    2014-01-01

    Several gene clusters that are responsible for toxin synthesis in bloom-forming cyanobacteria have been found to be associated with transposable elements (TEs). In particular, insertion sequence (IS) elements were shown to play a role in the inactivation or recombination of the genes responsible for cyanotoxin synthesis. Plasmids have been considered important vectors of IS element distribution to the host. In this study, we aimed to elucidate the IS elements propagated on the plasmids and the chromosome of the toxic cyanobacterium Planktothrix agardhii NIVA-CYA126/8 by means of high-throughput sequencing. In total, five plasmids (pPA5.5, pPA14, pPA50, pPA79, and pPA115, of 5, 6, 50, 79, and 120 kbp, respectively) were elucidated, and two plasmids (pPA5.5, pPA115) were found to propagate full IS element copies. Large stretches of shared DNA information between plasmids were constituted of TEs. Two plasmids (pPA5.5, pPA14) were used as candidates to engineer shuttle vectors (named pPA5.5SV and pPA14SV, respectively) in vitro by PCR amplification and the subsequent transposition of the Tn5 cat transposon containing the R6Kγ origin of replication of Escherichia coli. While pPA5.5SV was found to be fully segregated, pPA14SV consistently co-occurred with its wild-type plasmid even under the highest selective pressure. Interestingly, the Tn5 cat transposon became transferred by homologous recombination into another plasmid, pPA50. The availability of shuttle vectors is considered to be of relevance in investigating genome plasticity as a consequence of homologous recombination events. Combining the potential of high-throughput sequencing and in vitro production of shuttle vectors makes it simple to produce species-specific shuttle vectors for many cultivable prokaryotes. PMID:24907328

  15. Construction of Escherichia Coli Cell Factories for Production of Organic Acids and Alcohols.

    PubMed

    Liu, Pingping; Zhu, Xinna; Tan, Zaigao; Zhang, Xueli; Ma, Yanhe

    2016-01-01

    Production of bulk chemicals from renewable biomass has been proved to be sustainable and environmentally friendly. Escherichia coli is the most commonly used host strain for constructing cell factories for production of bulk chemicals since it has clear physiological and genetic characteristics, grows fast in minimal salts medium, uses a wide range of substrates, and can be genetically modified easily. With the development of metabolic engineering, systems biology, and synthetic biology, a technology platform has been established to construct E. coli cell factories for bulk chemicals production. In this chapter, we will introduce this technology platform, as well as E. coli cell factories successfully constructed for production of organic acids and alcohols. PMID:25577396

  16. A benzoic acid derivative and flavokawains from Piper species as schistosomiasis vector controls.

    PubMed

    Rapado, Ludmila N; Freitas, Giovana C; Polpo, Adriano; Rojas-Cardozo, Maritza; Rincón, Javier V; Scotti, Marcus T; Kato, Massuo J; Nakano, Eliana; Yamaguchi, Lydia F

    2014-01-01

    The search of alternative compounds to control tropical diseases such as schistosomiasis has pointed to secondary metabolites derived from natural sources. Piper species are candidates in strategies to control the transmission of schistosomiasis due to their production of molluscicidal compounds. A new benzoic acid derivative and three flavokawains from Piper diospyrifolium, P. cumanense and P. gaudichaudianum displayed significant activities against Biomphalaria glabrata snails. Additionally, "in silico" studies were performed using docking assays and Molecular Interaction Fields to evaluate the physical-chemical differences among the compounds in order to characterize the observed activities of the test compounds against Biomphalaria glabrata snails. PMID:24762961

  17. Construction and detection of the tissue-specific pINV-HPV16 E6/7 vector

    PubMed Central

    GAO, HUI; HUANG, ZHENGFANG; SHI, CHENLONG; LI, HOUDA

    2015-01-01

    A tissue-specific promoter can control downstream gene expression in tissues or organs. The human involucrin (hINV) promoter (pINV) that contains 2474 bp of hINV upstream sequence is able to regulate tissue-specific gene expression. This tissue specificity may be important for the prevention and treatment of human papilloma virus infections. pINV was cloned by polymerase chain reaction and the human papillomavirus (HPV)16 E6/7 gene was obtained from the cancer tissue samples of patients with cervical carcinoma at the Yangzhou Maternal and China Health-Care Center of Jinagsu Province (Yangzhou, China). First, specific primers were designed according to the genomic DNA sequence of the HPV16-type standard strain that has been reported and the E6/7 gene was acquired by PCR. The carcinogenic fraction of the E6/7 gene was removed and the remaining section was cloned into T vectors, sequenced correctly and then cloned into the eukaryotic expression vector pCEP4, which was lacking the CMV promoter. The positive recombinants were identified using blue-white screening and endonuclease digestion, subsequent to sequencing and analysis, and the tissue-specific recombinant pINV-HPV16E6/7 plasmids was detected. PMID:25621060

  18. T-scale as a novel vector of topological descriptors for amino acids and its application in QSARs of peptides

    NASA Astrophysics Data System (ADS)

    Tian, Feifei; Zhou, Peng; Li, Zhiliang

    2007-03-01

    In this paper, a new topological descriptor T-scale is derived from principal component analysis (PCA) on the collected 67 kinds of structural and topological variables of 135 amino acids. Applying T-scale to three peptide panels as 58 angiotensin-converting enzyme (ACE) inhibitors, 20 thromboplastin inhibitors (TI) and 28 bovine lactoferricin-(17-31)-pentadecapeptides (LFB), the resulting QSAR models, constructed by partial least squares (PLS), are all superior to reference reports, with correlative coefficient r2 and cross-validated q2 of 0.845, 0.786; 0.996, 0.782 (0.988, 0.961); 0.760, 0.627, respectively.

  19. [Construction and transfection of eucaryotic expression recombinant vector containing truncated region of UL83 gene of human cytomegalovirus and it's sheltered effect as DNA vaccine].

    PubMed

    Gao, Rong-Bao; Li, Yan-Qiu; Wang, Ming-Li

    2006-06-01

    To construct eucaryotic expression recombinant vector containing vivo truncated region of UL83 gene of human cytomegalovirus, realize its steady expression in Hep-2 cell, and study sheltered effect of the eucaryotic expression recombinant vector as DNA vaccine. A vivo truncated UL83 gene fragment encoding for truncated HCMV pp65 was obtained by PCR from human cytomegalovirus AD169 stock genome. By gene recombinant ways, the truncated UL83 gene fragment was cloned into eucaryotic expression vector pEGFP-C1 with reported gene coding GFP to construct recombinant vector pEGFP-C1-UL83. The recombinant vector pEGFP-C1-UL83 was tested by different methods including PCR, restriction digestion and gene sequencing. Test results showed the recombinant vector was constructed successfully. After pEGFP-C1-UL83 was transfected into Hep-2 cell by lipofectin mediation, expression of GFP and truncated pp65 fusion protein in Hep-2 cell was observed at different time points by fluorescence microscope. Results showed that quantity of fusion protein expression was the highest at 36h point. Then, Hep-2 cell was cultured selectively by RPMI-1640 containing G418 (200 microg/mL) to obtain a new cell stock of expressing truncated UL83 Gene fragment steadily. RT-PCR and Western blot results showed the truncated fragment of UL83 gene could be expressed steadily in Hep-2 cell. The result showed a new cell stock of expressing Tpp65 was established. This cell stock could be useful in some HCMV research fields, for example, it could be a tool in study of pp65 and HCMV infection, and it could provide a platform for the research into the therapy of HCMV infection. Immune sheltered effect of pEGFP-C1-UL83 as DNA vaccine was studied in vivo of HCMV congenital infection mouse model. The mouse model was immunized solely by pEGFP-C1-UL83, and was immunized jointly by pEGFP-C1-UL83 and its expression product. When the mouse was pregnant and brought to bed, differential antibody of anti-HCMV pp65 was

  20. Embedding methods for poly(L-lactic acid) microfiber mesh/human mesenchymal stem cell constructs.

    PubMed

    D'Alessandro, D; Battolla, B; Trombi, L; Barachini, S; Cascone, M G; Bernardini, N; Petrini, M; Mattii, L

    2009-01-01

    Fiber mesh scaffolds were recently investigated in tissue engineering as possible support for stem cell growth and differentiation, in order to repair lesion areas in clinical practice. In particular, the literature is focused on fiber mesh scaffolds constituted of biocompatible and resorbable polymeric structures, like poly(L-lactic acid) (PLLA). However, as regards the study of constructs constituted of PLLA microfibers and cells, only quantitative and SEM analyses were reported, lacking histological analysis. Histological evaluation of these constructs could give important information about cellular distribution in the scaffold, cell-scaffold interactions and extracellular matrix production. The purpose of our study was to find a valid method to analyze PLLA microfiber/cell constructs from both histological and histochemical angles. Biodegradable non-woven fiber meshes were prepared using hollow microfibers, based on PLLA. We first evaluated different embedding methods useable for histological analysis and the results showed that among the paraffin, Killik, and acrylic resin the only suitable medium was the latter. Then we employed the acrylic resin to embed the constructs made up of PLLA microfibers and bone marrow-derived human mesenchymal stromal cells, which we then analyzed with Toluidine Blue, PAS and Alcian Blue staining. These constructs, previously analyzed for cell viability by MTT and CCK-8 tests, showed viable/proliferating cells until 6 weeks of culture. The stainings performed on constructs confirmed viability data obtained with SEM and MTT/CCK-8 and supplied other information on the cell behaviors such as the distribution and organization onto the scaffold and the production of extracellular matrix molecules. In conclusion, this methodological study mainly suggests a suitable method to analyze PLLA microfiber/cell constructs, at the same time confirming and enriching the literature data on the compatibility between PLLA microfibers and hMSCs. PMID

  1. Construction of a Stable Replicating Shuttle Vector for Caldicellulosiruptor Species: Use for Extending Genetic Methodologies to Other Members of This Genus

    PubMed Central

    Chung, Daehwan; Cha, Minseok; Farkas, Joel; Westpheling, Janet

    2013-01-01

    The recalcitrance of plant biomass is the most important barrier to its economic conversion by microbes to products of interest. Thermophiles have special advantages for biomass conversion and members of the genus Caldicellulosiruptor are the most thermophilic cellulolytic microbes known. In this study, we report the construction of a replicating shuttle vector for Caldicellulosiruptor species based on pBAS2, the smaller of two native C. bescii plasmids. The entire plasmid was cloned into an E. coli cloning vector containing a pSC101 origin of replication and an apramycin resistance cassette for selection in E. coli. The wild-type C. bescii pyrF locus was cloned under the transcriptional control of the regulatory region of the ribosomal protein S30EA (Cbes2105), and the resulting vector was transformed into a new spontaneous deletion mutant in the pyrFA locus of C. bescii that allowed complementation with the pyrF gene alone. Plasmid DNA was methylated in vitro with a recently described cognate methyltransferase, M.CbeI, and transformants were selected for uracil prototrophy. The plasmid was stably maintained in low copy with selection but rapidly lost without selection. There was no evidence of DNA rearrangement during transformation and replication in C. bescii. A similar approach was used to screen for transformability of other members of this genus using M.CbeI to overcome restriction as a barrier and was successful for transformation of C. hydrothermalis, an attractive species for many applications. Plasmids containing a carbohydrate binding domain (CBM) and linker region from the C. bescii celA gene were maintained with selection and were structurally stable through transformation and replication in C. bescii and E. coli. PMID:23658781

  2. Modeling and predicting the tensile strength of poly (lactic acid)/graphene nanocomposites by using support vector regression

    NASA Astrophysics Data System (ADS)

    Cheng, W. D.; Cai, C. Z.; Luo, Y.; Li, Y. H.; Zhao, C. J.

    2016-04-01

    According to an experimental dataset under different process parameters, support vector regression (SVR) combined with particle swarm optimization (PSO) for its parameter optimization was employed to establish a mathematical model for prediction of the tensile strength of poly (lactic acid) (PLA)/graphene nanocomposites. Four variables, while graphene loading, temperature, time and speed, were employed as input variables, while tensile strength acted as output variable. Using leave-one-out cross validation test of 30 samples, the maximum absolute percentage error does not exceed 1.5%, the mean absolute percentage error (MAPE) is only 0.295% and the correlation coefficient (R2) is as high as 0.99. Compared with the results of response surface methodology (RSM) model, it is shown that the estimated errors by SVR are smaller than those achieved by RSM. It revealed that the generalization ability of SVR is superior to that of RSM model. Meanwhile, multifactor analysis is adopted for investigation on significances of each experimental factor and their influences on the tensile strength of PLA/graphene nanocomposites. This study suggests that the SVR model can provide important theoretical and practical guide to design the experiment, and control the intensity of the tensile strength of PLA/graphene nanocomposites via rational process parameters.

  3. Water-soluble N-[(2-hydroxy-3-trimethylammonium)propyl]chitosan chloride as a nucleic acids vector for cell transfection.

    PubMed

    Faizuloev, Evgeny; Marova, Anna; Nikonova, Alexandra; Volkova, Irina; Gorshkova, Marina; Izumrudov, Vladimir

    2012-08-01

    To endow the cationic polysaccharides with solubility in the whole pH-range without loss of functionality of the amino groups, different chitosan samples were treated with glycidyltrimethylammonium chloride. Each modified unit of the exhaustively alkylated quaternized chitosan (QCht) contained both quaternary and secondary amino groups. The intercalated dye displacement assay and ζ-potential measurements implied stability of QCht polyplexes at physiological conditions and protonation of the secondary amino groups in slightly acidic media which is favorable for transfection according to proton sponge mechanism. The cytotoxicity and transfection efficacy increased with the chain lengthening. Nevertheless, the longest chains of QCht, 250 kDa were less toxic than PEI for COS-1 cells and revealed comparable and even significantly higher transfection activity of siRNA and plasmid DNA, respectively. Thus, highly polymerized QCht (250 kDa) provided the highest level of the plasmid DNA transfection being 5 and 80 times more active than QCht (100 kDa) and QCht (50 kDa), respectively, and 4-fold more effective than PEI, 25 kDa. The established influence of QCht molecular weight on toxicity and transfection efficacy allows elaborating polysaccharide vectors that possess rational balance of these characteristics. PMID:24750918

  4. Advanced separator construction for long life valve-regulated lead-acid batteries

    NASA Astrophysics Data System (ADS)

    Stevenson, P. R.

    The performance of absorptive glass mat separators in valve-regulated lead-acid (VRLA) batteries is strongly influenced by the diameter of the fibres from which they are made. Coarser diameter fibres are beneficial for the compressive properties of separators while finer fibres maintain the uniform distribution of the electrolyte. Studies of cell compression and electrolyte stratification are reported using separators manufactured with segregated layers of fine and coarse fibres incorporated into a single sheet. This construction locates the different classes of fibre at their location of maximum effectiveness. Improvements in battery life in both cyclic and float charge applications are recorded, and compared with single layer separators.

  5. Construction of infectious cDNA clone derived from a classical swine fever virus field isolate in BAC vector using in vitro overlap extension PCR and recombination.

    PubMed

    Kamboj, Aman; Saini, Mohini; Rajan, Lekshmi S; Patel, Chhabi Lal; Chaturvedi, V K; Gupta, Praveen K

    2015-12-15

    To develop reverse genetics system of RNA viruses, cloning of full-length viral genome is required which is often challenging due to many steps involved. In this study, we report cloning of full-length cDNA from an Indian field isolate (CSFV/IVRI/VB-131) of classical swine fever virus (CSFV) using in vitro overlap extension PCR and recombination which drastically reduced the number of cloning steps. The genome of CSFV was amplified in six overlapping cDNA fragments, linked by overlap extension PCR and cloned in a bacterial artificial chromosome (BAC) vector using in vitro recombination method to generate full-length cDNA clone. The full-length CSFV cDNA clone was found stable in E. coli Stellar and DH10B cells. The full-length RNA was transcribed in vitro using T7 RNA polymerase and transfected in PK15 cells using Neon-tip electroporator to rescue infectious CSFV. The progeny CSFV was propagated in PK15 cells and found indistinguishable from the parent virus. The expression of CSFV proteins were detected in cytoplasm of PK15 cells infected with progeny CSFV at 72 h post-infection. We concluded that the in vitro overlap extension PCR and recombination method is useful to construct stable full-length cDNA clone of RNA virus in BAC vector. PMID:26478540

  6. Assessment of the microbial community in a constructed wetland that receives acid coal mine drainage

    SciTech Connect

    Nicomrat, D.; Dick, W.A.; Tuovinen, O.H.

    2006-01-15

    Constructed wetlands are used to treat acid drainage from surface or underground coal mines. However, little is known about the microbial communities in the receiving wetland cells. The purpose of this work was to characterize the microbial population present in a wetland that was receiving acid coal mine drainage (AMD). Samples were collected from the oxic sediment zone of a constructed wetland cell in southeastern Ohio that was treating acid drainage from an underground coal mine seep. Samples comprised Fe(Ill) precipitates and were pretreated with ammonium oxalate to remove interfering iron, and the DNA was extracted and purified by agarose gel electrophoresis prior to amplification of portions of the 16S rRNA gene. Amplified products were separated by denaturing gradient gel electrophoresis and DNA from seven distinct bands was excised from the gel and sequenced. The sequences were matched to sequences in the GenBank bacterial 16S rDNA database. The DNA in two of the bands yielded matches with Acidithiobacillus ferrooxidans and the DNA in each of the remaining five bands was consistent with one of the following microorganisms: Acidithiobacillus thiooxidans, strain TRA3-20 (a eubacterium), strain BEN-4 (an arsenite-oxidizing bacterium), an Alcaligenes sp., and a Bordetella sp. Low bacterial diversity in these samples reflects the highly inorganic nature of the oxic sediment layer where high abundance of iron- and sulfur-oxidizing bacteria would be expected. The results we obtained by molecular methods supported our findings, obtained using culture methods, that the dominant microbial species in an acid receiving, oxic wetland are A. thiooxidans and A. ferrooxidans.

  7. Production of fired construction brick from high sulfate-containing fly ash with boric acid addition.

    PubMed

    Başpinar, M Serhat; Kahraman, Erhan; Görhan, Gökhan; Demir, Ismail

    2010-01-01

    The increase of power plant capacity has led to the production of an increasing amount of fly ash that causes high environmental impact in Turkey. Some of the fly ash is utilized within the fired brick industry but high sulfate-containing fly ash creates severe problems during sintering of the fired brick. This study attempted to investigate the potential for converting high sulfate-containing fly ash into useful material for the construction industry by the addition of boric acid. The chemical and mineralogical composition of fly ash and clay were investigated. Boric acid (H(3)BO(3)) was added to fly ash-clay mixtures with up to 5 wt.%. Six different series of test samples were produced by uniaxial pressing. The samples were fired at the industrial clay-brick firing temperatures of 800, 900 and 1000 degrees C. The microstructures of the fired samples were investigated by scanning electron microscopy and some physical and mechanical properties were measured. It was concluded that the firing at conventional brick firing temperature of high sulfate fly ash without any addition of boric acid resulted in very weak strength bricks. The addition of boric acid and clay simultaneously to the high sulfate- containing fly ash brick dramatically increased the compressive strength of the samples at a firing temperature of 1000 degrees C by modifying the sintering behaviour of high sulfate fly ash. PMID:19423597

  8. Construction of novel chloroplast expression vector and development of an efficient transformation system for the diatom Phaeodactylum tricornutum.

    PubMed

    Xie, Wei-Hong; Zhu, Cong-Cong; Zhang, Nai-Sheng; Li, Da-Wei; Yang, Wei-Dong; Liu, Jie-Sheng; Sathishkumar, Ramalingam; Li, Hong-Ye

    2014-10-01

    Plastids are ideal subcellular hosts for the expression of transgenes and have been successfully used for the production of different biopolymers, therapeutic proteins and industrial enzymes. Phaeodactylum tricornutum is a widely used aquatic feed species. In this study, we focused on developing a high-efficiency plastid expression system for P. tricornutum. In the plastid transformation vector, the site selected for integration was the transcriptionally active intergenic region present between the trnI and trnA genes, located in the IR (inverted repeat) regions of the plastid genome. Initially, a CAT reporter gene (encoding chloramphenicol acetyltransferase) was integrated at this site in the plastid genome. The expression of CAT in the transformed microalgae conferred resistance to the antibiotic chloramphenicol, which enabled growth in the selection media. Overall, the plastid transformation efficiency was found to be approximately one transplastomic colony per 1,000 microalgae cells. Subsequently, a heterologous gene expression cassette for high-level expression of the target gene was created and cloned between the homologous recombination elements. A TA cloning strategy based on the designed XcmI-XcmI sites could conveniently clone the heterologous gene. An eGFP (green fluorescent protein) reporter gene was used to test the expression level in the plastid system. The relatively high-level expression of eGFP without codon optimisation in stably transformed microalgae was determined to account for 0.12 % of the total soluble protein. Thus, this study presents the first and convenient plastid gene expression system for diatoms and represents an interesting tool to study diatom plastids. PMID:24763817

  9. Geometry of trigonal boron coordination sphere in boronic acids derivatives - a bond-valence vector model approach.

    PubMed

    Czerwińska, Karolina; Madura, Izabela D; Zachara, Janusz

    2016-04-01

    The systematic analysis of the geometry of three-coordinate boron in boronic acid derivatives with a common [CBO2] skeleton is presented. The study is based on the bond-valence vector (BVV) model [Zachara (2007). Inorg. Chem. 46, 9760-9767], a simple tool for the identification and quantitative estimation of both steric and electronic factors causing deformations of the coordination sphere. The empirical bond-valence (BV) parameters in the exponential equation [Brown & Altermatt (1985). Acta Cryst. B41, 244-247] rij and b, for B-O and B-C bonds were determined using data deposited in the Cambridge Structural Database. The values obtained amount to rBO = 1.364 Å, bBO = 0.37 Å, rBC = 1.569 Å, bBC = 0.28 Å, and they were further used in the calculation of BVV lengths. The values of the resultant BVV were less than 0.10 v.u. for 95% of the set comprising 897 [CBO2] fragments. Analysis of the distribution of BVV components allowed for the description of subtle in- and out-of plane deviations from the `ideal' (sp(2)) geometry of boron coordination sphere. The distortions specific for distinct groups of compounds such as boronic acids, cyclic and acyclic esters, benzoxaboroles and hemiesters were revealed. In cyclic esters the direction of strains was found to be controlled by the ring size effect. It was shown that the syn or anti location of substituents on O atoms is decisive for the deformations direction for both acids and acyclic esters. The greatest strains were observed in the case of benzoxaboroles which showed the highest deviation from the zero value of the resultant BVV. The out-of-plane distortions, described by the vz component of the resultant BVV, were ascertained to be useful in the identification of weak secondary interactions on the fourth coordination site of the boron centre. PMID:27048726

  10. Synthesis and evaluation of N-(2,3-dihydroxypropyl)-PEIs as efficient vectors for nucleic acids.

    PubMed

    Tripathi, Sushil K; Yadav, Santosh; Gupta, Kailash C; Kumar, Pradeep

    2012-04-01

    Branched polyethylenimine (bPEI, 25 kDa) has been widely used as an efficient delivery vector for nucleic acids in vitro. However, its charge-associated toxicity has limited its in vivo applications. In an attempt to control its toxicity, it was reacted with varying amounts of glycidol (2,3-epoxy-1-propanol) to obtain a small series of hydrophilic polymers, 2,3-dihydroxypropyl-grafted-polyethylenimines (DHP-g-P). The resulting polymers were characterized by (1)H-NMR and subjected to interaction with negatively charged pDNA, which yielded complexes in the size range of ~171-190 nm with a zeta potential of ∼+33-39 mV. Acid-base titration revealed no effect of substitution on the buffering capacity of the modified polymers. Grafting of 2,3-dihydroxypropyl groups on bPEI significantly improved the cell viability (i.e. almost non-toxic) as well as the DNA release properties of these modified polymers compared to native bPEI. Formation of a relatively loose DHP-g-P25/pDNA complex (the best working system in terms of transfection efficiency) resulted in the efficient nuclear release of pDNA for transcription, a prerequisite for efficient transfection. Subsequently, upon evaluation of their ability to transfer nucleic acids in vitro, the DHP-g-P/pDNA complexes exhibited higher gene transfection efficiency with one of the formulations, DHP-g-P25/DNA complex, displaying ~2.7 folds higher GFP expression than bPEI and ~2.3-3.5 folds higher than the selected commercial transfection reagents used in this study. Further to quantify the extent of GFP positive cells, FACS analysis was performed, which revealed DHP-g-P25/DNA mediated gene expression in ~51% cells outcompeting bPEI, Superfect™, Fugene™ and Lipofectamine™. Sequential delivery of GFP-specific siRNA resulted in ~78% suppression of the target gene compared to ~49% achieved by Fugene™. All these results demonstrate the potential of these polymers for in vivo gene delivery. PMID:22419101

  11. Construction of overexpression vectors of Magnaporthe oryzae genes BAS1 and BAS4 fusion to mCherry and screening of overexpression strains.

    PubMed

    Liang, M L; Yan, J L; Yang, Y Q; Liu, L; Li, C Y; Yang, J

    2015-01-01

    The aim of this study was to construct overexpression vectors and selecting strains of the Magnaporthe oryzae effectors BAS1 and BAS4. Primer pairs of BAS1, BAS4, and mCherry were designed based on their known nucleotide sequences. The coding sequences of BAS1 and BAS4 were amplified, and the pXY201 plasmid was selected as a template to amplify the mCherry sequence. Fragments of BAS1 and mCherry, and BAS4 and mCherry were ligated into the pCAMBIA1302 vector. The recombinant pCAMBIA-BAS1-mCherry and pCAMBIA-BAS4-mCherry plasmids were transformed into E. coli DH5α competent cells. Transformants were screened by PCR, and plasmids from the positive transformants were extracted by enzymatic digestion to obtain pCAMBIA-BAS1-mCherry and pCAMBIA-BAS4-mCherry. The pCAMBIA-BAS1-mCherry and pCAMBIA-BAS4-mCherry plasmids were transformed into protoplasts of rice blast strains and the transformed strains were screened by PCR using primer pairs against the hygromycin gene. The result showed that the PCR products corresponded with the theoretical sizes. RT-PCR was used to analyze the expression of BAS1 and BAS4 in five transformed strains of BAS1 and BAS4, and the result showed that the higher expression level of the two genes was occurred in five transformant strains comparing to wild-type strain A3467-40 (the strain containing BAS1 and BAS4), but there was no difference among the five overexpression strains. The sporulation and spore germination of transformed strains was higher than in wild type strain, and there was no difference in the germination time. Construction of overexpression vectors and strains of M. oryzae effectors BAS1 and BAS4 provide reference material for other new effectors. PMID:26125917

  12. The distribution of common construction materials at risk to acid deposition in the United States

    NASA Astrophysics Data System (ADS)

    Lipfert, Frederick W.; Daum, Mary L.

    Information on the geographic distribution of various types of exposed materials is required to estimate the economic costs of damage to construction materials from acid deposition. This paper focuses on the identification, evaluation and interpretation of data describing the distributions of exterior construction materials, primarily in the United States. This information could provide guidance on how data needed for future economic assessments might be acquired in the most cost-effective ways. Materials distribution surveys from 16 cities in the U.S. and Canada and five related databases from government agencies and trade organizations were examined. Data on residential buildings are more commonly available than on nonresidential buildings; little geographically resolved information on distributions of materials in infrastructure was found. Survey results generally agree with the appropriate ancillary databases, but the usefulness of the databases is often limited by their coarse spatial resolution. Information on those materials which are most sensitive to acid deposition is especially scarce. Since a comprehensive error analysis has never been performed on the data required for an economic assessment, it is not possible to specify the corresponding detailed requirements for data on the distributions of materials.

  13. Introducing Vectors.

    ERIC Educational Resources Information Center

    Roche, John

    1997-01-01

    Suggests an approach to teaching vectors that promotes active learning through challenging questions addressed to the class, as opposed to subtle explanations. Promotes introducing vector graphics with concrete examples, beginning with an explanation of the displacement vector. Also discusses artificial vectors, vector algebra, and unit vectors.…

  14. Enhanced Efficacy of an AAV Vector Encoding Chimeric, Highly-Secreted Acid α-glucosidase in Glycogen Storage Disease Type II

    PubMed Central

    Sun, Baodong; Zhang, Haoyue; Benjamin, Daniel K.; Brown, Talmage; Bird, Andrew; Young, Sarah P.; McVie-Wylie, Alison; Chen, Y-T; Koeberl, Dwight D.

    2009-01-01

    Glycogen storage disease type II (GSD-II; Pompe disease; MIM 232300) is an inherited muscular dystrophy caused by deficiency in the activity of the lysosomal enzyme acid α-glucosidase (GAA). We hypothesized that chimeric GAA containing an alternative signal peptide could increase the secretion of GAA from transduced cells and enhance the receptor-mediated uptake of GAA in striated muscle. The relative secretion of chimeric GAA from transfected 293 cells increased up to 26-fold. Receptor-mediated uptake of secreted, chimeric GAA corrected cultured GSD-II patient cells. High-level hGAA was sustained in the plasma of GSD-II mice for 24 weeks following administration of an AAV2/8 vector encoding chimeric GAA; furthermore, GAA activity was increased and glycogen content was significantly reduced in striated muscle and in the brain. Administration of only 1×1010 vector particles increased GAA activity in the heart and diaphragm for >18 weeks, whereas 3×1010 vector particles increased GAA activity and reduced glycogen content in the heart, diaphragm, and quadriceps. Furthermore, an AAV2/2 vector encoding chimeric GAA produced secreted hGAA for >12 weeks in the majority of treated GSD-II mice. Thus, chimeric, highly secreted GAA enhanced the efficacy of AAV vector-mediated gene therapy in GSD-II mice. PMID:16987711

  15. Synthesis, crystal structure and optical property of three coordination polymer constructed from m-phenylenediacrylate acid

    SciTech Connect

    Xiao, YiFan; Wang, TingTing; Zeng, HePing

    2015-01-15

    Three new complexes of the wholly deprotonated flexible ligand m-phenylenediacrylate acid (H{sub 2}mpda), namely, [Zn (mpda) (H{sub 2}O)] [1], [Pr{sub 2}(mpda){sub 2}(H{sub 2}O){sub 2}(CHOO){sub 2}] [2], and [Mn{sub 2}(mpda){sub 2} (H{sub 2}O){sub 4}] [3], were synthesized by the solvothermal reaction. The complexes were characterized by IR spectroscopy, thermogravimetric analysis, and X-ray single-crystal diffraction. A mixed sample [Nd(mpda)(H{sub 2}O)(CHOO)·Pr(mpda)(H{sub 2}O)(CHOO)] [2{sup ]} was also synthesized and studied. Compounds [1] and [2] exhibit a selective sensing function with respect to acetone, and they are a potential luminescent sensory material for the selective detection of Zn{sup 2+} and Ni{sup 2+} ions, respectively. Furthermore, the fluorescent changes of compound [1] upon the addition of cations (Fe{sup 3+} and Zn{sup 2+}, Bi{sup 2+} and Zn{sup 2+}) are utilized to construct two logic gates at the molecular level. - Graphical abstract: Three new complexes of flexible ligand m-phenylenedicarboxylic acid were synthesized by solvothermal reaction. Three metal−organic frameworks exhibiting 1D, 2D, 3D structures, respectively. - Highlights: • Three new complexes of m-phenylenedicarboxylic acid were synthesized. • Three metal−organic frameworks exhibiting 1D, 2D, 3D structures, respectively. • [1] and [2] exhibits a very high quenching effect with acetone. • [1] and [2] are a potential luminescent sensory material for Zn{sup 2+}, Ni{sup 2+} ions. • Two logic gates have been constructed using compound [1].

  16. Glutamatergic or GABAergic neuron-specific, long-term expression in neocortical neurons from helper virus-free HSV-1 vectors containing the phosphate-activated glutaminase, vesicular glutamate transporter-1, or glutamic acid decarboxylase promoter

    PubMed Central

    Rasmussen, Morten; Kong, Lingxin; Zhang, Guo-rong; Liu, Meng; Wang, Xiaodan; Szabo, Gabor; Curthoys, Norman P.; Geller, Alfred I.

    2009-01-01

    Many potential uses of direct gene transfer into neurons require restricting expression to one of the two major types of forebrain neurons, glutamatergic or GABAergic neurons. Thus, it is desirable to develop virus vectors that contain either a glutamatergic or GABAergic neuron-specific promoter. The brain/kidney phosphate-activated glutaminase (PAG), the product of the GLS1 gene, produces the majority of the glutamate for release as neurotransmitter, and is a marker for glutamatergic neurons. A PAG promoter was partially characterized using a cultured kidney cell line. The three vesicular glutamate transporters (VGLUTs) are expressed in distinct populations of neurons, and VGLUT1 is the predominant VGLUT in the neocortex, hippocampus, and cerebellar cortex. Glutamic acid decarboxylase (GAD) produces GABA; the two molecular forms of the enzyme, GAD65 and GAD67, are expressed in distinct, but largely overlapping, groups of neurons, and GAD67 is the predominant form in the neocortex. In transgenic mice, an ∼9 kb fragment of the GAD67 promoter supports expression in most classes of GABAergic neurons. Here, we constructed plasmid (amplicon) Herpes Simplex Virus (HSV-1) vectors that placed the Lac Z gene under the regulation of putative PAG, VGLUT1, or GAD67 promoters. Helper virus-free vector stocks were delivered into postrhinal cortex, and the rats were sacrificed 4 days or 2 months later. The PAG or VGLUT1 promoters supported ∼90 % glutamatergic neuron-specific expression. The GAD67 promoter supported ∼90 % GABAergic neuron-specific expression. Long-term expression was observed using each promoter. Principles for obtaining long-term expression from HSV-1 vectors, based on these and other results, are discussed. Long-term glutamatergic or GABAergic neuron-specific expression may benefit specific experiments on learning or specific gene therapy approaches. Of note, promoter analyses might identify regulatory elements that determine a glutamatergic or GABAergic

  17. Ethanol production from acid hydrolysates based on the construction and demolition wood waste using Pichia stipitis.

    PubMed

    Cho, Dae Haeng; Shin, Soo-Jeong; Bae, Yangwon; Park, Chulhwan; Kim, Yong Hwan

    2011-03-01

    The feasibility of ethanol production from the construction and demolition (C&D) wood waste acid hydrolysates was investigated. The chemical compositions of the classified C&D wood waste were analyzed. Concentrated sulfuric acid hydrolysis was used to obtain the saccharide hydrolysates and the inhibitors in the hydrolysates were also analyzed. The C&D wood waste composed of lumber, plywood, particleboard, and medium density fiberboard (MDF) had polysaccharide (cellulose, xylan, and glucomannan) fractions of 60.7-67.9%. The sugar composition (glucose, xylose, and mannose) of the C&D wood wastes varied according to the type of wood. The additives used in the wood processing did not appear to be released into the saccharide solution under acid hydrolysis. Although some fermentation inhibitors were detected in the hydrolysates, they did not affect the ethanol production by Pichia stipitis. The hexose sugar-based ethanol yield and ethanol yield efficiency were 0.42-0.46 g ethanol/g substrate and 84.7-90.7%, respectively. Therefore, the C&D wood wastes dumped in landfill sites could be used as a raw material feedstock for the production of bioethanol. PMID:21251816

  18. Construction of tetranuclear metallacycles based on half-sandwich Ir, Rh fragments and pyridyl-substituted ligands with different coordinate vectors.

    PubMed

    Fan, Qi-Jia; Zhang, Wen-Ying; Lin, Yue-Jian; Jin, Guo-Xin

    2016-03-21

    A series of organometallic macrocycles have been constructed by two-step reactions of [Cp*M(μ-Cl)Cl]2 (M = Ir, Rh), firstly with AgOTf to abstract chloride ions and then with simple pyridyl-substituted ligands-pyridyldipyrromethene (HL(1)), pyridin-4-yl (1H-pyrrol-2-yl)methanone (HL(2)) and pyridine-4-carbohydrazide (HL(3))-resulting in the formation of the tetranuclear 32-membered metallacycles [(Cp*Ir)(L(1))]4(OTf)4 (2a) and [(Cp*Rh)(L(1))]4(OTf)4 (2b), and the 28-membered metallacycles [(Cp*Ir)(L(2))]4 (OTf)4 (3a), [(Cp*Rh)(L(2))]4(OTf)4 (3b), [(Cp*Ir)(L(3))]4(OTf)4 (4a) and [(Cp*Rh)(L(3))]4(OTf)4 (4b). Four target complexes were characterised by single crystal X-ray analyses, revealing that these metallacycles, constructed from half-sandwich metal corners and pyridyl-substituted linkers, form large ring structures. The observed variation in the metallacyclic geometries was explained on the basis of the structural flexibility of the corner fragments, subtle changes in coordination geometries, and changes in the orientation of the coordinate vectors in the given ligands, as well as different dihedral angles between the two binding fragments in the nonplanar ligands. PMID:26845526

  19. Hyaluronic Acid Enhances the Mechanical Properties of Tissue-Engineered Cartilage Constructs

    PubMed Central

    Levett, Peter A.; Hutmacher, Dietmar W.; Malda, Jos; Klein, Travis J.

    2014-01-01

    There is a need for materials that are well suited for cartilage tissue engineering. Hydrogels have emerged as promising biomaterials for cartilage repair, since, like cartilage, they have high water content, and they allow cells to be encapsulated within the material in a genuinely three-dimensional microenvironment. In this study, we investigated the mechanical properties of tissue-engineered cartilage constructs using in vitro culture models incorporating human chondrocytes from osteoarthritis patients. We evaluated hydrogels formed from mixtures of photocrosslinkable gelatin-methacrylamide (Gel-MA) and varying concentrations (0–2%) of hyaluronic acid methacrylate (HA-MA). Initially, only small differences in the stiffness of each hydrogel existed. After 4 weeks of culture, and to a greater extent 8 weeks of culture, HA-MA had striking and concentration dependent impact on the changes in mechanical properties. For example, the initial compressive moduli of cell-laden constructs with 0 and 1% HA-MA were 29 and 41 kPa, respectively. After 8 weeks of culture, the moduli of these constructs had increased to 66 and 147 kPa respectively, representing a net improvement of 69 kPa for gels with 1% HA-MA. Similarly the equilibrium modulus, dynamic modulus, failure strength and failure strain were all improved in constructs containing HA-MA. Differences in mechanical properties did not correlate with glycosaminoglycan content, which did not vary greatly between groups, yet there were clear differences in aggrecan intensity and distribution as assessed using immunostaining. Based on the functional development with time in culture using human chondrocytes, mixtures of Gel-MA and HA-MA are promising candidates for cartilage tissue-engineering applications. PMID:25438040

  20. The construction and proliferative effects of a lentiviral vector capable of stably overexpressing SPINK1 gene in human pancreatic cancer AsPC-1 cell line.

    PubMed

    Zhang, Jing; Wang, Dongmei; Hu, Na; Wang, Qian; Yu, Shanice; Wang, Jun

    2016-05-01

    This study aims to design and generate recombinant lentiviral vector containing the complete coding sequence (CDS) region of human serine protease inhibitor Kazal type 1 gene (SPINK1) and establish a human pancreatic cancer cell line (AsPC-1) stably overexpressing SPINK1. Then, to assess the proliferative and oncogenic effects of upregulated SPINK1 gene on pancreatic cancer AsPC-1 cells using different methods. Initially, the target coding sequence (CDS) of SPINK1 was amplified by polymerase chain reaction (PCR) and the synthesized product was subsequently subcloned into the lentiviral vector. The construction of recombinant SPINK1 gene was verified by the restriction digestion and sequencing analysis. The lentiviral particles carrying SPINK1 gene were produced by co-transfection of the recombination lentiviral vector and the packaging mix plasmids into 293 T cells and filtered and concentrated before AsPC-1 cells were infected by the virus particles. The cells transduced were differentially selected with puromycin, and the clones that highly expressed SPINK1 were chosen by real-time PCR and confirmed by Western blot after 7 weeks. The stably transduced AsPC-1 cell line showed significantly increased metabolic and proliferative capability using CCK-8 and Trypan Blue assays (P < 0.001). Cell cycle and DNA content analysis by flow cytometry showed that upregulated SPINK1 elicited significant increase in the percentage of AsPC-1 cells in the S and G2/M phase (P < 0.001). Clone formation assay demonstrated that the number of the colonies formed in the experimental group was greater than that in the control parental cells (P < 0.001). It was concluded that a stable AsPC-1 cell line capable of overexpressing SPINK1 had been successfully created, and that the proliferative capacity of AsPC-1 pancreatic cancer cells was significantly raised by SPINK1 upregulation as well as the ability of a single AsPC-1 cell to grow into a colony. PMID:26586397

  1. Evaluation of Furan Photooxygenation as a Device for Construction of the Zaragozic Acid (Squalestatin) Core.

    PubMed

    Maezaki, Naoyoshi; Gijsen, Harrie J. M.; Sun, Li-Qiang; Paquette, Leo A.

    1996-09-20

    A practical application of the photooxygenation chemistry of 3-substituted furans to construction of the zaragozic acid/squalestatin backbone is described. Although addition of 3-lithiofuran to the tartrate-derived aldehyde 7 proceeds without chelation control to give a 1:1 mixture of diastereomeric alcohols 8, it is demonstrated initially that ready conversion to the polyfunctional intermediate 10 is possible by sequential treatment of 9 with singlet oxygen, sodium borohydride, and triisopropylsilyl triflate. The actual enantiocontrolled route consisted of oxidation of 8 to the ketone and Wittig olefination of the latter in advance of asymmetric dihydroxylation with AD-mix-beta. Once this series of transformations had been accomplished, formation of the target product 30 was realized by an entirely comparable photooxygenation. PMID:11667541

  2. Construction of hyaluronic acid noisome as functional transdermal nanocarrier for tumor therapy.

    PubMed

    Kong, Ming; Park, Hyunjin; Feng, Chao; Hou, Lin; Cheng, Xiaojie; Chen, Xiguang

    2013-04-15

    To develop a functional nanosized transdermal drug delivery system for tumor therapy, amphiphilic hyaluronic acid (HA) based niosome was constructed combining transdermal and tumor targeting ability in one entity. HA esterified with monostearin, the conjugate labeled as HA-GMS self-assembled onto niosome surface and formed HA-niosome. The multilayer vesicle had small size (around 40 nm), good stability and desirable drug encapsulating efficacy, and well compatible with blood. It exhibited better endocytosis to mouse breast tumor cell (4T1) than the control chitosan nanoparticle, which was verified qualitatively and quantitatively. Skin permeation of HA-niosome was proven to be efficient using in vitro stratum corneum model and in vivo fluorescence observation. Histological section study confirmed the security and efficiency of transdermal permeation. The results evidence HA-niosome to be exciting and promising for tumor therapy through trandermal administration. PMID:23544584

  3. A New Multidentate Hexacarboxylic Acid for the Construction of Porous Metal−Organic Frameworks of Diverse Structures and Porosities

    SciTech Connect

    Chen, Zhenxia; Xiang, Shengchang; Liao, Tengbiao; Yang, Yongtai; Chen, Yu-Sheng; Zhou, Yaming; Zhao, Dongyuan; Chen, Banglin

    2010-08-27

    Three porous metal-organic frameworks have been constructed from the new developed C3-symmetric hexacarboxylic acid 3,3{prime},3{double_prime},5,5{prime},5{double_prime}-benzene-1,3,5-triyl-hexabenzoic acid (H{sub 6}BHB) of different NaCl- and Al{sub 2}O{sub 3}-type topologies. The activated metal-organic frameworks (MOFs) exhibit permanent porosities as revealed in their gas and vapor adsorption isotherms, highlighting the promise of this new ligand for the construction of porous MOFs for gas storage and separation.

  4. Two multi-dimensional frameworks constructed from zinc coordination polymers with pyridine carboxylic acids

    SciTech Connect

    Guo Yuanyuan; Ma Pengtao; Wang Jingping; Niu Jingyang

    2011-11-15

    Two novel zinc coordination polymers [Zn{sub 2}(H{sub 2}O)L(MoO{sub 4})]{sub n} (1) and [Zn{sub 4}(PO{sub 4}){sub 2}L'(H{sub 2}O)]{sub n} (2) (H{sub 2}L=2,2'-bipyridine-6.6'-dicarboxylic acid, H{sub 2}L'=2,2'-bipyridine-4,4'-dicarboxylic acid) have been hydrothermally synthesized and characterized by elemental analyses, IR spectra, UV spectra, single-crystal X-ray diffraction and thermogravimetric analyses. Structural analyses indicate that 1 represents a 2-D sheet structure built by dimeric [Zn{sub 2}L(H{sub 2}O)]{sup 2+} units and MoO{sub 4}{sup 2-} groups whereas 2 displays an interesting 3-D framework constructed by tetranuclear zinc clusters, L'{sup 2-} ligands and PO{sub 4}{sup 3-} groups. Examination of UV spectra suggests that both 1 and 2 can stably exist in the pH range of 2.45-5.45 and 3.01-8.55 in aqueous solution, respectively. The room-temperature solid-state photoluminescence of 1 and 2 are derived from the intra-ligands {pi}-{pi}* transitions of H{sub 2}L and H{sub 2}L' ligands and the ligand-to-metal-charge-transfer transitions. - Graphical Abstract: Two new transition metal coordination polymers, namely, [Zn{sub 2}(H{sub 2}O)L{sub 1}(MoO{sub 4})]{sub n} (1), [Zn{sub 4}(PO{sub 4}){sub 2}L{sub 2}(H{sub 2}O)]{sub n} (2) (H{sub 2}L{sub 1}=2,2'-bipyridine-6,6'-dicarboxylic acid, H{sub 2}L{sub 2}=2,2'-bipyridine-4,4'-dicarboxylic acid) have been hydrothermally synthesized. 1 represents a 2-D sheet structure while 2 represents 3-D network. Highlights: {yields}Two new transition metal coordination polymers have been hydrothermally synthesized. > The two compounds have been characterized by elemental analyses, IR, UV spectra, single-crystal X-ray diffraction, thermogravimetric analyses and photoluminescence. > Compound 1 represents a 2-D sheet structure while 2 represents 3-D network.

  5. Chemical vectors for gene delivery: a current review on polymers, peptides and lipids containing histidine or imidazole as nucleic acids carriers

    PubMed Central

    Midoux, Patrick; Pichon, Chantal; Yaouanc, Jean-Jacques; Jaffrès, Paul-Alain

    2009-01-01

    DNA/cationic lipid (lipoplexes), DNA/cationic polymer (polyplexes) and DNA/cationic polymer/cationic lipid (lipopolyplexes) electrostatic complexes are proposed as non-viral nucleic acids delivery systems. These DNA-nanoparticles are taken up by the cells through endocytosis processes, but the low capacity of DNA to escape from endosomes is regarded as the major limitations of their transfection efficiency. Here, we present a current report on a particular class of carriers including the polymers, peptides and lipids, which is based on the exploitation of the imidazole ring as an endosome destabilization device to favour the nucleic acids delivery in the cytosol. The imidazole ring of histidine is a weak base that has the ability to acquire a cationic charge when the pH of the environment drops bellow 6. As it has been demonstrated for poly(histidine), this phenomena can induce membrane fusion and/or membrane permeation in an acidic medium. Moreover, the accumulation of histidine residues inside acidic vesicles can induce a proton sponge effect, which increases their osmolarity and their swelling. The proof of concept has been shown with polylysine partially substituted with histidine residues that has caused a dramatic increase by 3–4.5 orders of magnitude of the transfection efficiency of DNA/polylysine polyplexes. Then, several histidine-rich polymers and peptides as well as lipids with imidazole, imidazolinium or imidazolium polar head have been reported to be efficient carriers to deliver nucleic acids including genes, mRNA or SiRNA in vitro and in vivo. More remarkable, histidylated carriers are often weakly cytotoxic, making them promising chemical vectors for nucleic acids delivery. This article is part of a themed section on Vector Design and Drug Delivery. For a list of all articles in this section see the end of this paper, or visit: http://www3.interscience.wiley.com/journal/121548564/issueyear?year=2009 PMID:19459843

  6. Construction of a novel peptide nucleic acid piezoelectric gene sensor microarray detection system.

    PubMed

    Chen, Ming; Liu, Minghua; Yu, Lili; Cai, Guoru; Chen, Qinghai; Wu, Rong; Wang, Feng; Zhang, Bo; Jiang, Tianlun; Fu, Welling

    2005-08-01

    A novel 2 x 5 clamped style piezoelectric gene sensor microarray has been successfully constructed. Every crystal unit of the fabricated gene sensor can oscillate independently without interfering with each other. The bis-peptide nucleic acid (bis-PNA) probe, which can combine with target DNA or RNA sequences more effectively and specifically than a DNA probe, was designed and immobilized on the surface of the gene sensor microarray to substitute the conventional DNA probe for direct detection of the hepatitis B virus (HBV) genomic DNA. Detection conditions were then explored and optimized. Results showed that PBS buffer of pH 6.8, an ion concentration of 20 mmol/liter, and a probe concentration of 1.5 micromol/liter were optimal for the detection system. Under such optimized experimental conditions, the specificity of bis-PNA was proved much higher than that of DNA probe. The relationship between quantity of target and decrease of frequency showed a typical saturation curve when concentrations of target HBV DNA varied from 10 pg/liter to 100 microg/liter, and 10 microg/liter was the watershed, with a statistic linear regression equation of I gC = -2.7455 + 0.0691 deltaF and the correlating coefficient of 0.9923. Fortunately, this is exactly the most ordinary variant range of the HBV virus concentration in clinical hepatitis samples. So, a good technical platform is successfully constructed and it will be applied to detect HBV quantitatively in clinical samples. PMID:16193990

  7. Construction of a Recombinant Leuconostoc mesenteroides CJNU 0147 Producing 1,4-Dihydroxy-2-Naphthoic Acid, a Bifidogenic Growth Factor

    PubMed Central

    2015-01-01

    1,4-Dihydroxy-2-naphthoic acid (DHNA), a precursor of menaquinone (vitamin K2), has an effect on growth stimulation of bifidobacteria and prevention of osteoporosis, making it a promising functional food material. Therefore, we tried to clone the menB gene encoding DHNA synthase from Leuconostoc mesenteroides CJNU 0147. Based on the genome sequence of Leu. mesenteroides ATCC 8293 (GenBank accession no., CP000414), a primer set (Leu_menBfull_F and Leu_menBfull_R) was designed for the PCR amplification of menB gene of CJNU 0147. A DNA fragment (1,190 bp), including the menB gene, was amplified, cloned into pGEM-T Easy vector, and sequenced. The deduced amino acid sequence of MenB (DHNA synthase) protein of CJNU 0147 had a 98% similarity to the corresponding protein of ATCC 8293. The menB gene was subcloned into pCW4, a lactic acid bacteria - E. coli shuttle vector, and transferred to CJNU 0147. The transcription of menB gene of CJNU 0147 (pCW4::menB) was increased, when compared with those of CJNU 0147 (pCW4) and CJNU 0147 (−). The DHNA was produced from it at a detectable level, indicating that the cloned menB gene of CJNU 0147 encoded a DHNA synthase which is responsible for the production of DHNA, resulting in an increase of bifidogenic growth stimulation activity. PMID:26877648

  8. Broad-Host-Range ProUSER Vectors Enable Fast Characterization of Inducible Promoters and Optimization of p-Coumaric Acid Production in Pseudomonas putida KT2440.

    PubMed

    Calero, Patricia; Jensen, Sheila I; Nielsen, Alex T

    2016-07-15

    Pseudomonas putida KT2440 has gained increasing interest as a host for the production of biochemicals. Because of the lack of a systematic characterization of inducible promoters in this strain, we generated ProUSER broad-host-expression plasmids that facilitate fast uracil-based cloning. A set of ProUSER-reporter vectors was further created to characterize different inducible promoters. The PrhaB and Pm promoters were orthogonal and showed titratable, high, and homogeneous expression. To optimize the production of p-coumaric acid, P. putida was engineered to prevent degradation of tyrosine and p-coumaric acid. Pm and PrhaB were used to control the expression of a tyrosine ammonia lyase or AroG* and TyrA* involved in tyrosine production, respectively. Pathway expression was optimized by modulating inductions, resulting in small-scale p-coumaric acid production of 1.2 mM, the highest achieved in Pseudomonads under comparable conditions. With broad-host-range compatibility, the ProUSER vectors will serve as useful tools for optimizing gene expression in a variety of bacteria. PMID:27092814

  9. Novel one-dimensional lanthanide acrylic acid complexes: an alternative chain constructed by hydrogen bonding

    SciTech Connect

    Li Hui . E-mail: lihui@bit.edu.cn; Hu Changwen

    2004-12-01

    Novel one-dimensional (1D) chains of three lanthanide complexes La(L{sup 1}){sub 3}(CH{sub 3}OH)].CH{sub 3}OH (L{sup 1}=(E)-3-(2-hydroxyl-phenyl)-acrylic acid) 1, La(L{sup 2}){sub 3}(H{sub 2}O){sub 2}].2.75H{sub 2}O (L{sup 2}=(E)-3-(3-hydroxyl-phenyl)-acrylic acid) 2, and La(L{sup 3}){sub 3}(CH{sub 3}OH){sub 2}(H{sub 2}O)].CH{sub 3}OH (L{sup 3}=(E)-3-(4-hydroxyl-phenyl)-acrylic acid) 3 are reported. The crystal structure data are as follows for 1: C{sub 29}H{sub 29}LaO{sub 11}, monoclinic, P2{sub 1}/n, a=15.4289(12)A, b=7.9585(6)A, c=23.041(2)A, {beta}=99.657(2){sup o}, Z=4, R{sub 1}=0.0637, wR{sub 2}=0.0919; for 2: C{sub 27}H{sub 30.50}LaO{sub 13.75}, triclinic, P-1, a=8.4719(17)A, b=13.719(3)A, c=14.570(3)A, {alpha}=62.19(3){sup o}, {beta}=99.657(2){sup o}, {gamma}=78.22(3){sup o}, Z=2, R{sub 1}=0.0384, wR{sub 2}=0.0820; and for 3: C{sub 30}H{sub 35}LaO{sub 13}, monoclinic, P2(1)/c, a=9.5667(6)A, b=24.3911(15)A, c=14.0448(9)A, {beta}=109.245(2){sup o}, Z=4, R{sub 1}=0.0374, wR{sub 2}=0.0630. All the three structure data were collected using graphite monochromated molybdenum K{alpha} radiation and refined using full-matrix least-squares techniques on F{sup 2}. These structures show that four kinds of the carboxylato bridge modes are included in these chains to link the La(III) ions. It is the first time that it has been found that the intra-chain hydrogen bonding can construct an alternative chain even, when the coordination bridge mode is the same along the chain (complex 2). There are 2D and 3D hydrogen bonding in the crystal lattices of complexes 1-3.

  10. Formic and acetic acids in degradation products of plant volatiles elicit olfactory and behavorial responses from an insect vector

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae) vectors the bacterial pathogen presumed to be the etiological agent of citrus greening disease, Huanglongbing (HLB), a major threat to citrus industry worldwide. We studied antennal and behavioral responses of Diaphorina citri...

  11. Predicting Three-Dimensional Conformations of Peptides Constructed of Only Glycine, Alanine, Aspartic Acid, and Valine

    NASA Astrophysics Data System (ADS)

    Oda, Akifumi; Fukuyoshi, Shuichi

    2015-06-01

    The GADV hypothesis is a form of the protein world hypothesis, which suggests that life originated from proteins (Lacey et al. 1999; Ikehara 2002; Andras 2006). In the GADV hypothesis, life is thought to have originated from primitive proteins constructed of only glycine, alanine, aspartic acid, and valine ([GADV]-proteins). In this study, the three-dimensional (3D) conformations of randomly generated short [GADV]-peptides were computationally investigated using replica-exchange molecular dynamics (REMD) simulations (Sugita and Okamoto 1999). Because the peptides used in this study consisted of only 20 residues each, they could not form certain 3D structures. However, the conformational tendencies of the peptides were elucidated by analyzing the conformational ensembles generated by REMD simulations. The results indicate that secondary structures can be formed in several randomly generated [GADV]-peptides. A long helical structure was found in one of the hydrophobic peptides, supporting the conjecture of the GADV hypothesis that many peptides aggregated to form peptide multimers with enzymatic activity in the primordial soup. In addition, these results indicate that REMD simulations can be used for the structural investigation of short peptides.

  12. An analysis of the effectiveness of a constructed wetland treating acid mine drainage

    SciTech Connect

    Huddleston, G.M. III; Grant, A.J.; Ramey, B.A.

    1994-12-31

    Acid mine drainage (AMD) from an abandoned coal mine in southcentral Kentucky had pH levels as low as 2.3 and iron concentrations as high as 641 mg/L. In the summer of 1992, the US Soil Conservation Service constructed a wetland system to treat the AMD that incorporated both physical and biological treatments. The AMD was initially fed into three anoxic limestone beds followed by an aeration pond and four cattail cells. A polishing pond served as the final stage of treatment. Flow of AMD was initiated in the fall of 1992, and treatment effectiveness was monitored for the next year. Chemical analysis and the cladoceran (Ceriodaphnia dubia) survival and reproduction test were performed on water samples collected along the flow path. Water chemistry analysis and determination of toxic levels indicated a substantial increase in pH and removal of metals prior to entering the cattail cells. Water quality in the cattail cells and polishing pond varied throughout the seasons, but had improved substantially by the end of the one-year monitoring period. The use of the wetland system by macroinvertebrates also was evaluated. Results indicated that a limited number of species were found in the cattail cells, while larger numbers were recovered from the polishing pond.

  13. Woven-grid sealed quasi-bipolar lead-acid battery construction and fabricating method

    NASA Technical Reports Server (NTRS)

    Rippel, Wally E. (Inventor)

    1989-01-01

    A quasi-bipolar lead-acid battery construction includes a plurality of bipolar cells disposed in side-by-side relation to form a stack, and a pair of monoplanar plates at opposite ends of the stack, the cell stack and monopolar plates being contained within a housing of the battery. Each bipolar cell is loaded with an electrolyte and composed of a bipolar electrode plate and a pair of separator plates disposed on opposite sides of the electrode plate and peripherally sealed thereto. Each bipolar electrode plate is composed of a partition sheet and two bipolar electrode elements folded into a hairpin configuration and applied over opposite edges of the partition sheet so as to cover the opposite surfaces of the opposite halves thereof. Each bipolar electrode element is comprised of a woven grid with a hot-melt strip applied to a central longitudinal region of the grid along which the grid is folded into the hairpin configuration, and layers of negative and positive active material pastes applied to opposite halves of the grid on opposite sides of the central hot-melt strip. The grid is made up of strands of conductive and non-conductive yarns composing the respective transverse and longitudinal weaves of the grid. The conductive yarn has a multi-stranded glass core surrounded and covered by a lead sheath, whereas the non-conductive yarn has a multi-stranded glass core surrounded and covered by a thermally activated sizing.

  14. Synthesis of nucleoside and nucleotide conjugates of bile acids, and polymerase construction of bile acid-functionalized DNA.

    PubMed

    Ikonen, Satu; Macícková-Cahová, Hana; Pohl, Radek; Sanda, Miloslav; Hocek, Michal

    2010-03-01

    Aqueous Sonogashira cross-coupling reactions of 5-iodopyrimidine or 7-iodo-7-deazaadenine nucleosides with bile acid-derived terminal acetylenes linked via an ester or amide tether gave the corresponding bile acid-nucleoside conjugates. Analogous reactions of halogenated nucleoside triphosphates gave directly bile acid-modified dNTPs. Enzymatic incorporation of these modified nucleotides to DNA was successfully performed using Phusion polymerase for primer extension. One of the dNTPs (dCTP bearing cholic acid) was also efficient for PCR amplification. PMID:20165813

  15. Carbohydrate-interactive pDNA and siRNA gene vectors based on boronic acid functionalized poly(amido amine)s.

    PubMed

    Piest, Martin; Ankoné, Marc; Engbersen, Johan F J

    2013-08-10

    In order to evaluate the influence of incorporation of boronic acid groups on the properties of poly(amido amine)s as gene vectors, a novel poly(amido amine) copolymer p(CBA-ABOL/2AMPBA) containing ortho-aminomethylphenylboronic acid (2AMPBA) moieties was prepared by Michael-type polyaddition of a mixture of 1,4-aminobutanol (ABOL) and 2-((4-aminobutylamino)methyl)phenyl boronic acid to N,N'-cystamine bisacrylamide (CBA). It appeared that the presence of the boronic acid moieties as side groups along the polymer chain strongly enhances the stability of the self-assembled nanoparticles and nanosized polyplexes formed from this polymer; no aggregation was observed after storage for 6days at 37°C. This strong stabilization can be attributed to intermolecular Lewis acid-base interactions between the 2AMPBA groups and the alcohol and amine groups present in the polymer, leading to dynamical (reversible) crosslinking in the nanoparticles. Moreover, since the boronic acids can reversibly form boronic esters with vicinal diol groups, the presence of the 2AMPBA groups add carbohydrate-interactive properties to these polymers that strongly influence their behavior as gene delivery vectors. DNA transfection with p(CBA-ABOL/2AMPBA) polyplexes gave transfection efficiencies that were approximately similar to commercial PEI in different cell lines (COS-7, HUH-6 and H1299-Fluc), but lower than those obtained with reference polyplexes from p(CBA-ABOL). It is hypothesized that the uptake of the boronated polyplexes is suppressed by binding to the glycocalyx of the cells. This is supported by the observation that addition of sorbitol or dextran to the transfection medium significantly enhances the transfection efficiency, which can be attributed to increased cellular uptake of the polyplexes due to boronic ester formation with these agents. AFM, SEM and confocal microscopy showed that polyplexes of p(CBA-ABOL/2AMPBA) become decorated with a dextran layer in the presence of 0.9% (w

  16. Incorporation of nisI-mediated nisin immunity improves vector-based nisin-controlled gene expression in lactic acid bacteria.

    PubMed

    Oddone, Gian M; Mills, David A; Block, David E

    2009-05-01

    Lactic acid bacteria (LAB) have been used successfully to express a wide variety of recombinant proteins, ranging from flavor-active proteins to antibiotic peptides and oral vaccines. The nisin-controlled expression (NICE) system is the most prevalent of the systems for production of heterologous proteins in LAB. Previous optimization of the NICE system has revealed a strong limit on the concentration of the inducer nisin that can be tolerated by the culture of host cells. In this work, the nisin immunity gene, nisI, has been inserted into the recently reported pMSP3535H2 vector that contains the complete NICE system on a high-copy Escherichia coli-LAB shuttle vector. Fed-batch fermentation data show that Lactococcus lactis IL1403 cells transformed with the new vector, pMSP3535H3, tolerate a 5-fold increase in the concentration of the inducer nisin, and, at this elevated concentration, produce a 1.8-fold increased level of green fluorescent protein (GFP), a model recombinant protein. Therefore, the incorporation of nisI in the pMSP3535H3 NICE system described here unveils new ranges of induction parameters to be studied in the course of optimizing recombinant protein expression in LAB. PMID:19141301

  17. Use of submitochondrial particle (SMP) assays for assessing wetlands constructed for sequestering acid mine runoff

    SciTech Connect

    Bettermann, A.D.; Haahr, J.E.; Lazorchak, J.M.

    1995-12-31

    Use of constructed wetlands to sequester metals from acid mine drainage is part of a USEPA SITE demonstration at Burleigh Tunnel near Silverplume, Colorado. Samples are collected on a seasonal basis for toxicity evaluation of two different pilot treatment systems. Water samples were obtained from the outflow of two experimental wetland cells utilizing either upflow and downflow treatment, as well as upstream and downstream of the discharge of Burleigh Tunnel to Clear Creek. Submitochondrial Particle (SMP), Ceriodaphnia dubia and Pimephales promelas acute bioassays were used to evaluate the water quality. The SMP bioassay is based on the electron transfer complex derived from mitochondria. Toxic responses result from subcellular perturbations of various subsets of enzyme systems contained in the complex. In prior work, a 0.79 r{sup 2} was reported between the SMP bioassay and P. promelas for 11 inorganics on the EPA Priority Pollutant list. The SMP bioassay provided data consistent with the whole organism results. The two most toxic samples: the Burleigh outflow, and the Clear Creek Upstream sample, gave C. dubia LC50s of 1.01% and 8.41%, respectively. The Burleigh outflow P. promelas LC50 was 1.55%. SMP EC50s for the Burleigh outflow and the Clear Creek Upstream sample were 0.63% and 1.63%, respectively. As the SMP bioassay requires 1 hour to run and costs approximately 1/10th of whole organism assays, it was feasible to determine EC50 values for 7 samples vs. the two sample LC50s determined using whole organism assays. The SMP bioassays can provide sufficient sampling density, at low cost, allowing effective delineation of wetland performance over time.

  18. [Research on prediction method of fatty acid content in edible oil based on Raman spectroscopy and multi-output least squares support vector regression machine].

    PubMed

    Deng, Zhi-Yin; Zhang, Bing; Dong, Wei; Wang, Xiao-Ping

    2013-11-01

    For the purpose of the rapid prediction of saturated fatty acid, oleic acid, linoleic acid content in edible vegetable oil, the Raman spectra of a batch of edible vegetable oils and their one-one mixtures with different ratios were measured in the range of 800-2 000 cm(-1), 91 samples were measured totally in this research, the obtained Raman spectra data were preprocessed by a new method proposed in this paper called auto-set fulcrums baseline fitting method based on peak-seeking algorithm, and 8 characteristic peak values (872 cm(-1) [v(C-C)], 972 cm(-1) [delta(C=C) trans], 1 082 cm(-1) [v(C-C)], 1 267 cm(-1) [delta(=C-H) cis], 1 303 cm(-1) [delta(CH2) twisting], 1 442 cm(-1) [delta(CH2) scissoring], 1 658 cm(-1) [v(C=C) cis], 1 748 cm(-1) [v(C=O)]) were extracted to be the eigenvalues for the whole spectra, among the 8 peaks there are three peaks (972, 1 267, 1 658 cm(-1)) that play an important role in the establishment of mathematical model, they are closely concerned with C=C band which distinguishes the three fatty acid types. By using these eigenvalues as inputs, and actual saturated fatty acid, oleic acid, linoleic acid contents of sample oils as outputs, a prediction mathematical model that predicts simultaneously the three fatty acid contents was established using multiple regression analysis: multi-output least squares support vector regression machine (MLS-SVR) and partial least squares (PLS). Results show that the MLS-SVR has better effects. The predicting results are compared with results of gas chromatography(GC), and the obtained root mean square error of prediction(RMSEP) for saturated fatty acid, oleic acid, linoleic acid are 0.496 7%, 0.840 0% and 1.019 9%, and the correlation coefficients (r) are 0.813 3, 0.999 2 and 0.998 1, respectively. When this model is applied in the detection of new unknown oil samples, the prediction error does not exceed 5%. Results show that the Raman spectra analysis technology based on MLS-SVR can be a convenient

  19. Analysis of Streptococcus pyogenes promoters by using novel Tn916-based shuttle vectors for the construction of transcriptional fusions to chloramphenicol acetyltransferase.

    PubMed Central

    Geist, R T; Okada, N; Caparon, M G

    1993-01-01

    We have developed a series of shuttle vectors based on the conjugative transposon Tn916 that have been designed for the analysis of transcriptional regulation in Streptococcus pyogenes and other gram-positive bacteria. Designated the pVIT vectors (vectors for integration into Tn916), the vectors are small, stable plasmids in Escherichia coli to facilitate the fusion of promoters from cloned S. pyogenes genes to a promoterless gene which encodes chloramphenicol acetyltransferase. The vectors each contain one or more small regions of Tn916 to direct the integration of the transcriptional fusion into the transposon via homologous recombination following transformation of S. pyogenes or other suitable gram-positive hosts. Integration can be monitored by the inactivation or replacement of an antibiotic resistance determinant in modified derivatives of Tn916. Promoter activity can then be quantitated by the determination of chloramphenicol acetyltransferase-specific activity. In addition, since integration is into loci that do not disrupt the conjugative transpositional functions of Tn916, the vectors are useful for analysis of regulation in strains that are difficult or impossible to transform and can be introduced into these strains by conjugation following transformation of an intermediate host. The promoters for the genes which encode both the M protein and protein F of S. pyogenes were active in pVIT vectors, as was the region which controls transcription of mry, a trans-acting positive regulator of M protein expression. However, neither of the two characterized promoters for mry demonstrated activity when independently analyzed in pVIT-generated partial diploid strains, suggesting that regulation of mry is more complex than predicted by current models. The broad host range of Tn916 should make the pVIT vectors useful for analysis of regulation in numerous other bacterial species. PMID:8244925

  20. Method for construction of bacterial strains with increased succinic acid production

    DOEpatents

    Donnelly, Mark I.; Sanville-Millard, Cynthia; Chatterjee, Ranjini

    2000-01-01

    A fermentation process for producing succinic acid is provided comprising selecting a bacterial strain that does not produce succinic acid in high yield, disrupting the normal regulation of sugar metabolism of said bacterial strain, and combining the mutant bacterial strain and selected sugar in anaerobic conditions to facilitate production of succinic acid. Also provided is a method for changing low yield succinic acid producing bacteria to high yield succinic acid producing bacteria comprising selecting a bacterial strain having a phosphotransferase system and altering the phosphotransferase system so as to allow the bacterial strain to simultaneously metabolize different sugars.

  1. Transferability of different classical force fields for right and left handed α-helices constructed from enantiomeric amino acids.

    PubMed

    Biswas, Santu; Sarkar, Sujit; Pandey, Prithvi Raj; Roy, Sudip

    2016-02-21

    Amino acids can form d and l enantiomers, of which the l enantiomer is abundant in nature. The naturally occurring l enantiomer has a greater preference for a right handed helical conformation, and the d enantiomer for a left handed helical conformation. The other conformations, that is, left handed helical conformations of the l enantiomers and right handed helical conformations of the d enantiomers, are not common. The energetic differences between left and right handed alpha helical peptide chains constructed from enantiomeric amino acids are investigated using quantum chemical calculations (using the M06/6-311g(d,p) level of theory). Further, the performances of commonly used biomolecular force fields (OPLS/AA, CHARMM27/CMAP and AMBER) to represent the different helical conformations (left and right handed) constructed from enantiomeric (D and L) amino acids are evaluated. 5- and 10-mer chains from d and l enantiomers of alanine, leucine, lysine, and glutamic acid, in right and left handed helical conformations, are considered in the study. Thus, in total, 32 α-helical polypeptides (4 amino acids × 4 conformations of 5-mer and 10-mer) are studied. Conclusions, with regards to the performance of the force fields, are derived keeping the quantum optimized geometry as the benchmark, and on the basis of phi and psi angle calculations, hydrogen bond analysis, and different long range helical order parameters. PMID:26863595

  2. Nucleic acid encoding TGF-. beta. and its uses

    SciTech Connect

    Derynck, R.M.A.; Goeddel, D.V.

    1989-12-12

    This patent describes a method. It comprises: constructing a vector which includes nucleic acid encoding biologically active TGF-{beta}, transforming a host eukaryotic cell with the vector, culturing the transformed cell and recovering mature TGF-{beta} from the culture medium.

  3. Selection of acid compositions in well construction in difficult geological conditions

    NASA Astrophysics Data System (ADS)

    Mishchenko, M. V.; Kamartdinov, M. R.

    2016-03-01

    Within the scope of the current work we have presented an approach towards selecting and substantiation of acid composition in accordance with petrophysical characteristics of formations for acid treatment of bottom-hole formation zones. Article presents the results of lab tests of selected acid compositions, in conditions, which model thermobaric conditions of a payzone, combined with an evaluation of hydraulic permeability change, and this, in its turn, should allow us to evaluate the quality of impact of the acid composition recipe on the reservoir formation.

  4. Dose-dependent neuroprotective effect of ciliary neurotrophic factor delivered via tetracycline-regulated lentiviral vectors in the quinolinic acid rat model of Huntington's disease.

    PubMed

    Régulier, E; Pereira de Almeida, L; Sommer, B; Aebischer, P; Déglon, N

    2002-11-01

    The ability to regulate gene expression constitutes a prerequisite for the development of gene therapy strategies aimed at the treatment of neurologic disorders. In the present work, we used tetracycline (Tet)-regulated lentiviral vectors to investigate the dose-dependent neuroprotective effect of human ciliary neurotrophic factor (CNTF) in the quinolinic acid (QA) model of Huntington's disease (HD). The Tet system was split in two lentiviruses, the first one containing the CNTF or green fluorescent protein (GFP) cDNAs under the control of the Tet-response element (TRE) and a second vector encoding the transactivator (tTA). Preliminary coinfection study demonstrated that 63.8% +/- 2.0% of infected cells contain at least two viral copies. Adult rats were then injected with CNTF- and GFP-expressing viral vectors followed 3 weeks later by an intrastriatal administration of QA. A significant reduction of apomorphine-induced rotations was observed in the CNTF-on group. In contrast, GFP-treated animals or CNTF-off rats displayed an ipsilateral turning behavior in response to apomorphine. A selective sparing of DARPP-32-, choline acetyltransferase (ChAT)-, and NADPH-d-positive neurons was observed in the striatum of CNTF-on rats compared to GFP animals and CNTF-off group. Enzyme-linked immunosorbent assay (ELISA) performed on striatal samples of rats sacrificed at the same time point indicated that this neuroprotective effect was associated with the production of 15.5 +/- 4.7 ng CNTF per milligram of protein whereas the residual CNTF expression in the off state (0.54 +/- 0.02 ng/mg of protein) was not sufficient to protect against QA toxicity. These results establish the proof of principle of neurotrophic factor dosing for neurodegenerative diseases and demonstrate the feasibility of lentiviral-mediated tetracycline-regulated gene transfer in the brain. PMID:12427308

  5. A replication-competent retrovirus arising from a split-function packaging cell line was generated by recombination events between the vector, one of the packaging constructs, and endogenous retroviral sequences.

    PubMed

    Chong, H; Starkey, W; Vile, R G

    1998-04-01

    Previously we reported the presence of a replication-competent retrovirus in supernatant from a vector-producing line derived from a widely used split-function amphotropic packaging cell line. Rigorous routine screening of all retroviral stocks produced in our laboratory has not, previously or since, indicated the presence of such a virus. Replication-competent retroviruses have never previously been used in our laboratory, and stringent screening of all routinely used cell lines has not revealed the presence of any helper viruses. Therefore, it is highly unlikely that this virus represents an adventitious cross-contaminant or had been imported unknowingly with our cell line stocks. PCR studies with DNA from infected cell lines and Northern blot analysis and reverse transcriptase PCR with RNA from infected cells suggest that the helper virus arose by recombination events, at sites of partial homology, between sequences in the vector, one of the packaging constructs, and endogenous retroviral elements. These recombinations were not present in stocks of the packaging cell line or in an initial stock of the vector-producing line, indicating that these events occurred while the vector-producing line was being passaged for harvest of supernatant stocks. PMID:9525583

  6. Different types of degradable vectors from low-molecular-weight polycation-functionalized poly(aspartic acid) for efficient gene delivery.

    PubMed

    Dou, X B; Hu, Y; Zhao, N N; Xu, F J

    2014-03-01

    Poly(aspartic acid) (PAsp) has been employed as the potential backbone for the preparation of efficient gene carriers, due to its low cytotoxicity, good biodegradability and excellent biocompatibility. In this work, the degradable linear or star-shaped PBLA was first prepared via ring-opining polymerization of β-benzyl-L-aspartate N-carboxy anhydride (BLA-NCA) initiated by ethylenediamine (ED) or ED-functionalized cyclodextrin cores. Then, PBLA was functionalized via aminolysis reaction with low-molecular-weight poly(2-(dimethylamino)ethyl methacrylate) with one terminal primary amine group (PDMAEMA-NH2), followed by addition of excess ED or ethanolamine (EA) to complete the aminolysis process. The obtained different types of cationic PAsp-based vectors including linear or star PAsp-PDM-NH2 and PAsp-PDM-OH exhibited good condensation capability and degradability, benefiting gene delivery process. In comparison with gold standard polyethylenimine (PEI, ∼ 25 kDa), the cationic PAsp-based vectors, particularly star-shaped ones, exhibited much better transfection performances. PMID:24388816

  7. Lipophilic nucleic acids--a flexible construction kit for organization and functionalization of surfaces.

    PubMed

    Schade, Matthias; Berti, Debora; Huster, Daniel; Herrmann, Andreas; Arbuzova, Anna

    2014-06-01

    Lipophilic nucleic acids have become a versatile tool for structuring and functionalization of lipid bilayers and biological membranes as well as cargo vehicles to transport and deliver bioactive compounds, like interference RNA, into cells by taking advantage of reversible hybridization with complementary strands. This contribution reviews the different types of conjugates of lipophilic nucleic acids, and their physicochemical and self-assembly properties. Strategies for choosing a nucleic acid, lipophilic modification, and linker are discussed. Interaction with lipid membranes and its stability, dynamic structure and assembly of lipophilic nucleic acids upon embedding into biological membranes are specific points of the review. A large diversity of conjugates including lipophilic peptide nucleic acid and siRNA provides tailored solutions for specific applications in bio- and nanotechnology as well as in cell biology and medicine, as illustrated through some selected examples. PMID:24650567

  8. Construction and characterization of thermo-inducible vectors derived from heat-sensitive lacI genes in combination with the T7 A1 promoter.

    PubMed

    Chao, Yun-Peng; Chern, Jong-Tzer; Wen, Chin-Sheng; Fu, Hongyong

    2002-07-01

    The lack of stringency and the cost of induction are two major disadvantages of using lac-derived vectors for recombinant protein productions. To compensate for these drawbacks, a series of thermo-inducible vectors was developed by coupling heat-sensitive lacI (lacIts) with the T7 A1 promoter on a multiple-copynumber plasmid. The lacIts genes were created by the introduction of Gly187-->Ser substitution along with three alternative mutation sites, Leu233-->Lys, Ala241-->Thr, and Gly265-->Asp, generated by site-directed mutagenesis into the wild-type lacI gene. With the LacZ production as a model, the induction profiles for various vectors containing distinct lacIts exhibited a positive trend as the temperature increased. The fully induced level was achieved by applying the temperature shift from 30 degrees C to 42, 40, or 37 degrees C to the cells harboring the plasmid with the Gly187-->Ser, Ala241-->Thr, or Gly265-->Asp substitution in lacI, respectively. As a result, it produced the maximal LacZ production ranging between 46,000 and 54,000 Miller units, corresponding to a 100- to 400-fold amplification over the uninduced level. As a whole, these novel expression vectors are characterized as having tight regulation and facile inducibility, and their practical usefulness in industrial production of recombinant proteins appears promising. PMID:17590925

  9. Impact assessment and remediation strategies for roadway construction in acid-bearing media: case study from Mid-Appalachia

    SciTech Connect

    Viadero, R.C.; Fortney, R.H.; Creel, A.T.

    2008-09-15

    The likelihood of encountering land impacted by current and/or historic coal mining activities is high when constructing roadways in the Mid-Appalachian region. Through additional disturbance of these lands, environmental impacts such as acid and dissolved metals loading and subsequent impacts to aquatic flora and fauna will ensue. Consequently, it is necessary to affect a paradigm shift in roadway design and construction to account for the presence of factors that compound the already difficult task of working in a region characterized by steep topography and aggressive geochemistry. In this study, assessments of the water chemistry and biological impacts of a waste pile containing spoils from previous mining and the presence of an exposed coal mine bench were made as representative microcosmic examples of typical conditions found in the region. Based on quantitative measurements of water quality and biological conditions, recommendations are presented for the assessment and avoidance of impacts prior to construction through acid-bearing materials and suggestions are offered for postconstruction remediation at previously impacted sites.

  10. Constructing a recombinant hyaluronic acid biosynthesis operon and producing food-grade hyaluronic acid in Lactococcus lactis.

    PubMed

    Sheng, Juzheng; Ling, Peixue; Wang, Fengshan

    2015-02-01

    Hyaluronic acid (HA), a natural high molecular weight polysaccharide, is produced by Streptococcus zooepidemicus. However, Streptococcus has several drawbacks including its potential to produce exotoxins, so there is demand for an alternative HA source. Here, a recombinant HA biosynthesis operon, as well as the HA biosynthesis operon of S. zooepidemicus were introduced into L. lactis using the nisin-controlled expression system, respectively. HA was successfully synthesized by recombinant L. lactis. Furthermore, overexpression of the endogenous enzymes directing the synthesis of precursor sugars was effective at increasing HA production, and increasing the supply of UDP-activated monosaccharide donors aided synthesis of monodisperse HA polysaccharides. Besides GRAS host strain (L. lactis) and NICE system, the selecting marker (lacF gene) of the recombinant strain is also food grade. Therefore, HA produced by recombinant L. lactis overcomes the problems associated with Streptococcus and provides a source of food-grading HA appropriate for widespread biotechnological applications. PMID:25447786

  11. Bubble vector in automatic merging

    NASA Technical Reports Server (NTRS)

    Pamidi, P. R.; Butler, T. G.

    1987-01-01

    It is shown that it is within the capability of the DMAP language to build a set of vectors that can grow incrementally to be applied automatically and economically within a DMAP loop that serves to append sub-matrices that are generated within a loop to a core matrix. The method of constructing such vectors is explained.

  12. Rapid construction of multiple sgRNA vectors and knockout of the Arabidopsis IAA2 gene using the CRISPR/Cas9 genomic editing technology.

    PubMed

    Dingyuan, Liu; Ting, Qiu; Xiaohui, Ding; Miaomiao, Li; Muyuan, Zhu; Junhui, Wang

    2016-08-01

    IAA2 is a member of the Aux/IAA auxin responsive gene family in Arabidopsis thaliana. No iaa2 mutant has been reported until now, thus hindering its further mechanistic investigations. The normal genomic editing technology of CRISPR/Cas9 (Clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) uses only a single guide RNA (sgRNA) to target one site in a specific gene, and the gene knockout efficiency is not high. Instead, multiple sgRNAs can target multiple sites; therefore, the efficiency may be improved. In the present investigation, we used the golden-gate cloning strategy and two rounds of PCR reactions to combine three sgRNAs in the same entry vector. The final expression vector was obtained by LR reactions with the destination vector containing the Cas9 expression cassette. Four out of the six sgRNAs were effective, and we also obtained a lot of insertion and deletion mutations. Compared with one sgRNA approach, multiple sgRNAs displayed higher gene-knockout efficiency and produced more germ-line mutants. Thus, we established a more rapid and efficient method and generated five mutants for further studies of IAA2 functions. PMID:27531614

  13. Vector Video

    NASA Astrophysics Data System (ADS)

    Taylor, David P.

    2001-01-01

    Vector addition is an important skill for introductory physics students to master. For years, I have used a fun example to introduce vector addition in my introductory physics classes based on one with which my high school physics teacher piqued my interest many years ago.

  14. Palladium-Catalyzed Construction of Amidines from Arylboronic Acids under Oxidative Conditions.

    PubMed

    Zhu, Fengxiang; Li, Yahui; Wang, Zechao; Orru, Romano V A; Maes, Bert U W; Wu, Xiao-Feng

    2016-06-01

    A valuable palladium-catalyzed three-component coupling reaction for the synthesis of amidines has been developed. Using arylboronic acids, isocyanides, and anilines as the reactants under oxidative conditions, various amidines were isolated in good yields with good functional group tolerances. PMID:27061735

  15. CONSTRUCTION OF MODULAR FIELD-BIOREACTOR FOR ACID MINE DRAINAGE TREATMENT

    EPA Science Inventory

    The paper focuses on the improvements to engineered features of a passive technology that has been used for remediation of acid rock drainage (ARD). This passive remedial technology, a sulfate-reducing bacteria (SRB) bioreactor, takes advantage of the ability of SRB that, if sup...

  16. Development of a small gantry robotic workcell for deoxyribonucleic acid (DNA) filter array construction

    SciTech Connect

    Beugelsdijk, T.J.; Hollen, R.M.; Snider, K.T.

    1990-01-01

    At Los Alamos National Laboratory, we have constructed a primary cosmid library of human chromosome 16. This library consists of an 11-fold representation of the chromosome and is arrayed in microtiter plate format. A need has arisen in the large scale physical mapping of this chromosome, to array spots of DNA from each of these colonies onto filter media for hybridization studies. We are currently developing a small gantry robot-based workcell to array small spots of DNA in an interleaved format. This allows for the construction of a high spot density format filter array. This paper will discuss the features incorporated into this workcell for the handling of thousands of colonies and their automatic tracking and positioning onto the filter. 7 refs., 3 figs., 1 tab.

  17. An efficient method for recovering Lyapunov vectors from singular vectors

    NASA Astrophysics Data System (ADS)

    Wolfe, Christopher L.; Samelson, Roger M.

    2007-05-01

    Lyapunov vectors are natural generalizations of normal modes for linear disturbances to aperiodic deterministic flows and offer insights into the physical mechanisms of aperiodic flow and the maintenance of chaos. Most standard techniques for computing Lyapunov vectors produce results which are norm-dependent and lack invariance under the linearized flow (except for the leading Lyapunov vector) and these features can make computation and physical interpretation problematic. An efficient, norm-independent method for constructing the n most rapidly growing Lyapunov vectors from n - 1 leading forward and n leading backward asymptotic singular vectors is proposed. The Lyapunov vectors so constructed are invariant under the linearized flow in the sense that, once computed at one time, they are defined, in principle, for all time through the tangent linear propagator. An analogous method allows the construction of the n most rapidly decaying Lyapunov vectors from n decaying forward and n - 1 decaying backward singular vectors. This method is demonstrated using two low-order geophysical models.

  18. Scalar-vector bootstrap

    NASA Astrophysics Data System (ADS)

    Rejon-Barrera, Fernando; Robbins, Daniel

    2016-01-01

    We work out all of the details required for implementation of the conformal bootstrap program applied to the four-point function of two scalars and two vectors in an abstract conformal field theory in arbitrary dimension. This includes a review of which tensor structures make appearances, a construction of the projectors onto the required mixed symmetry representations, and a computation of the conformal blocks for all possible operators which can be exchanged. These blocks are presented as differential operators acting upon the previously known scalar conformal blocks. Finally, we set up the bootstrap equations which implement crossing symmetry. Special attention is given to the case of conserved vectors, where several simplifications occur.

  19. Cloning vector

    DOEpatents

    Guilfoyle, R.A.; Smith, L.M.

    1994-12-27

    A vector comprising a filamentous phage sequence containing a first copy of filamentous phage gene X and other sequences necessary for the phage to propagate is disclosed. The vector also contains a second copy of filamentous phage gene X downstream from a promoter capable of promoting transcription in a bacterial host. In a preferred form of the present invention, the filamentous phage is M13 and the vector additionally includes a restriction endonuclease site located in such a manner as to substantially inactivate the second gene X when a DNA sequence is inserted into the restriction site. 2 figures.

  20. Cloning vector

    DOEpatents

    Guilfoyle, Richard A.; Smith, Lloyd M.

    1994-01-01

    A vector comprising a filamentous phage sequence containing a first copy of filamentous phage gene X and other sequences necessary for the phage to propagate is disclosed. The vector also contains a second copy of filamentous phage gene X downstream from a promoter capable of promoting transcription in a bacterial host. In a preferred form of the present invention, the filamentous phage is M13 and the vector additionally includes a restriction endonuclease site located in such a manner as to substantially inactivate the second gene X when a DNA sequence is inserted into the restriction site.

  1. Adsorption compared with sulfide precipitation as metal removal processes from acid mine drainage in a constructed wetland

    NASA Astrophysics Data System (ADS)

    Machemer, Steven D.; Wildeman, Thomas R.

    1992-01-01

    Metal removal processes from acid mine drainage were studied in an experimental constructed wetland in the Idaho Springs-Central City mining district of Colorado. The wetland was designed to passively remove heavy metals from the mine drainage flowing from the Big Five Tunnel. Concurrent studies were performed in the field on the waters flowing from the wetland and in the laboratory on the wetland substrate. Both studies suggest that there is competition for organic adsorption sites among Fe, Cu, Zn and Mn. Iron and Cu appear to be more strongly adsorbed than Zn and Mn. The adsorption of metals varies with the fluctuation of pH in the outflow water. Also indicated by field and laboratory studies is the microbial reduction of sulfate with a corresponding increase in the sulfide concentration of the water. As sulfide is generated. Cu and Zn are completely removed. The field results suggest that upon start up of a constructed wetland, the adsorption of dissolved metals onto organic sites in the substrate material will be an important process. Over time, sulfide precipitation becomes the dominant process for metal removal from acid mine drainage.

  2. Synthesis, structures and properties of two new chiral rare earth-organic frameworks constructed by L/D-tartaric acid

    SciTech Connect

    Hu, Gonghao; Zhang, Haitao; Miao, Hao; Wang, Jiahong; Xu, Yan

    2015-09-15

    Hydrothermal reactions of rare earth cerium with L- or D- tartaric acid afford a pair of novel chiral enantiomer coordination polymers, namely, [Ce(L-tart)(CH{sub 2}OHCH{sub 2}OH)(H{sub 2}O)]Cl (L-1) and [Ce(D-tart)(CH{sub 2}OHCH{sub 2}OH)(H{sub 2}O)]Cl (D-1). Their structures were determined by single crystal X-ray diffraction analyses and further characterized by elemental analyses, XRD, IR spectra, and TG analyses. The circular dichroism (CD) spectra and second-harmonic generation (SHG) efficiency measurements proved that they are of structural chirality in the bulk samples. To the best of our knowledge, the enantiomers of L-1 and D-1 are the first 2D chiral dilayer frameworks constructed from L/D-tartrate ligands, ancillary ligand ethanediol and lanthanide ion Ce. - Graphical abstract: Hydrothermal reactions of rare earth cerium with L- or D- tartaric acid afford a pair of novel chiral enantiomer coordination polymers, namely, [Ce(L-tart)(CH{sub 2}OHCH{sub 2}OH)(H{sub 2}O)]Cl (L-1) and [Ce(D-tart)(CH{sub 2}OHCH{sub 2}OH)(H{sub 2}O)]Cl (D-1). Structural analysis indicates that the enantiomers of L-1 and D-1 are the first 2D chiral dilayer frameworks constructed from L/D-tartrate ligands and ancillary ligands ethanediol reacted with lanthanide ions Ce.

  3. Collagen synthesis promoting pullulan-PEI-ascorbic acid conjugate as an efficient anti-cancer gene delivery vector.

    PubMed

    Ambattu, Lizebona August; Rekha, M R

    2015-08-01

    Cationized pullulan (pullulan-PEI; PP) was synthesized and further modified with an anti-oxidant molecule, ascorbic acid (PPAA) at various ratios. The nanoplexes formed at an optimum ratio of 4:1 was within a size of 150nm and had a zeta potential of 9-14mV. The nanoplexes at this ratio was used for further investigations. The cell internalization and transfection efficiency of these nanoplexes were determined in presence of serum. The internalization and transfection efficiency were found to be unaffected by the presence of fetal bovine serum. Another interesting observation was that this polymer was found to have collagen synthesis promoting property. The collagen synthesis effect of these polymers was quantified and observed that PPAA3 promoted the highest. Transfection efficiency was evaluated by assessing the p53 gene expression in C6 rat glioma cells and cell death was quantified to be 96% by flow cytometry, thus establishing the high efficacy of this polymer. PMID:25933522

  4. Construction of a stable plasmid vector for industrial production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) by a recombinant Cupriavidus necator H16 strain.

    PubMed

    Sato, Shunsuke; Fujiki, Tetsuya; Matsumoto, Keiji

    2013-12-01

    A new stable plasmid vector (pCUP3) was developed for high and stable production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBH) using Cupriavidus necator H16 as the host strain. In pCUP3, it was found that the plasmid partition and replication region of the megaplasmid pMOL28 in the Cupriavidus metallidurans CH34 strain plays an important role in plasmid stability in C. necator H16. Moreover, the partition locus (comprising parA28 and parB28 and the parS28 region) is essential for plasmid maintenance under high-PHBH-accumulation. PHBH productivity by the C. necator H16/ds strain (phaC1 deactivated mutant strain) harboring a phaCAc NSDG within pCUP3 was identical to the productivity of poly(3-hydroxybutyrate) by the C. necator H16 strain when palm kernel oil was used as the sole carbon source without any antibiotics. This new vector is important for industrial mass production of polyhydroxyalkanoates using the C. necator H16 strain as the host, dispensing the necessity of the application of selective pressure such as antibiotics. PMID:23816763

  5. Metal-organic frameworks constructed from crown ether-based 1,4-benzenedicarboxylic acid derivatives.

    PubMed

    Chen, Teng-Hao; Schneemann, Andreas; Fischer, Roland A; Cohen, Seth M

    2016-02-21

    A series of unprecedented crown ether- and thiacrown ether-derivatized benzene dicarboxylic acid (H2bdc) ligands has been synthesized and incorporated into the prototypical isoreticular metal-organic framework (IRMOF) and UiO-66 materials. In the case of UiO-66, only MOFs comprised from a mixed-ligand composition, requiring both unsubstituted bdc and crown ether containing ligands, could be prepared. These are among the few ligand derivatives, and resulting MOFs, that incorporate a macrocyclic group directly on the bdc ligand, providing a new, modular platform for exploring new supramolecular and coordination chemistry within MOFs. PMID:26765588

  6. 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…

  7. Vector quantization

    NASA Technical Reports Server (NTRS)

    Gray, Robert M.

    1989-01-01

    During the past ten years Vector Quantization (VQ) has developed from a theoretical possibility promised by Shannon's source coding theorems into a powerful and competitive technique for speech and image coding and compression at medium to low bit rates. In this survey, the basic ideas behind the design of vector quantizers are sketched and some comments made on the state-of-the-art and current research efforts.

  8. Physical Characterisation as an Insight into a Gene Delivery System Containing Cyclodextrins with Pluronic®-F127 and Folic acid as Non-Viral Vectors.

    PubMed

    Eng, Matthew; Elkordy, Amal A; McCarron, Paul A; Elkordy, Eman A; Faheem, Ahmed

    2014-01-01

    Gene delivery into cells offers opportunities to treat various human genetic diseases. Effective gene delivery is dependent on its stability and ability to transfect across cells. DNA is susceptible to enzymatic degradation and its negatively charge are barriers towards successful transfection. DNA has to be protected from degradation and neutralised. Non-viral vectors are preferred carrier systems, therefore, the use of cyclodextrins with Pluronic(®)-F127 and folic acid at different concentrations to stabilise the formulation was investigated. Formulations were characterised in fresh and freeze dried forms. DNA stability in formulations was tested by determining the stability of DNA against enzymatic degradation. Degree of DNA inclusion into cyclodextrins was investigated using fluorescence spectroscopy. Thermal behaviour was studied using Differential Scanning Calorimetry (DSC). Incorporation of Pluronic(®)-F127 produced most stable formulations regarding enzymatic degradation. These formulations show high percentage inclusion. Shift of peaks in FTIR data, appearance of uniform particulate as detected by SEM and changing in the denaturation temperature as demonstrated by DSC data for Pluronic(®)-F127 containing formulations confirm clear interaction between Pluronic(®)-F127 and cyclodextrin/ DNA complex. It was noted that γ-cyclodextrin provide better protection and inclusion compared to β-cyclodextrin. Pluronic(®)-F127 with cyclodextrins is a promising combination to improve stability. PMID:25158973

  9. New three-fold interpenetrated uranyl organic framework constructed by terephthalic acid and imidazole derivative.

    PubMed

    Chen, Fei; Wang, Cong-zhi; Li, Zi-jie; Lan, Jian-hui; Ji, Yan-qin; Chai, Zhi-fang

    2015-04-20

    A new 3-fold interpenetrated uranyl organic framework, UO2(bdc)(dmpi), was hydrothermally synthesized using 1,4-benzenedicarboxylic acid (H2bdc) and 1-(4-(1H-imidazol-1-yl)-2,5-dimethylphenyl)-1H-imidazole (dmpi). This framework, which was determined by synchrotron radiation X-ray, exhibited a new 3-fold interpenetrated (2,4)-connected topology with the Schläfli symbol of (12(6))(12)2. Additionally, large incurvation happened to the bond angle of [O=U=O](2+), which was always arranged in a rigorous line. Computational results based on density functional theory (DFT) indicated that the bent geometry of uranyl in UO2(bdc)(dmpi) was mainly due to the higher charge populations in the valence 6d shells of uranium, rendered by the electronegative imidazoles. PMID:25835754

  10. A novel nanofiber Cur-loaded polylactic acid constructed by electrospinning

    NASA Astrophysics Data System (ADS)

    Thu Trang Mai, Thi; Thu Thuy Nguyen, Thi; Duong Le, Quang; Ngoan Nguyen, Thi; Cham Ba, Thi; Binh Nguyen, Hai; Bich Hoa Phan, Thi; Tran, Dai Lam; Phuc Nguyen, Xuan; Park, Jun Seo

    2012-06-01

    Curcumin (Cur), extracted from the Curcuma longa L. plant, is well known for its anti-tumor, anti-oxidant, anti-inflammatory and anti-bacterial properties. Nanofiber mats of polylactic acid (PLA) loading Cur (5 wt%) were fabricated by electrospinning (e-spinning). Morphology and structure of the fibers were characterized by field emission scanning electron microscopy (FE-SEM) and Fourier transform infrared (FTIR) spectroscopy, respectively. The diameters of the obtained fibers varied from 200 to 300 nm. The release capacity of curcumin from curcumin-loaded PLA fibers was investigated in phosphate buffer saline (PBS) containing ethanol. After 24 h, 50% of the curcumin was released from curcumin-loaded PLA fibers. These results of electrospun (e-spun) fibers exhibit the potential for biomedical application.

  11. Two metal-organic frameworks with different configurations constructed from a flexible tripodal triaromatic acid

    NASA Astrophysics Data System (ADS)

    Liang, Li-Li; Xu, Lei; Xue, Hong-Bao; Tao, Zhao-Lin; Chen, Fei-Jian

    2016-12-01

    Two metal-organic frameworks [Ce2(L)2(DMF)4] (1), and [Ni3(HL2-)4(bpe)3(H2O)4]·[H2N(CH3)2+]2(DMF)6(H2O)8 (2) (H3L = 4,4‧,4″-{[(2,4,6-trimethylbenzene-1,3,5-triyl)tris-(methylene)] tris(oxy)}tribenzoic acid, bpe = trans-4,4‧-bipyridylethylene) have been solvothermally synthesized from a flexible tripodal ligand. Single-crystal X-ray diffraction shows that compound 1 features a 3,6-connected double-layer network which is further connected by weak π-π interactions between benzene rings to form a 3D porous network. Compound 2 exhibits an unprecedented (4,5)-connected (33.42.5.62.72)2(6.84.10) topology based on zigzag chains and straight chains. In addition, thermal stabilities and luminescent properties of 1 and 2 were investigated in the solid state.

  12. Insight into the structural construction of a perfluorosulfonic acid membrane derived from a polymeric dispersion

    NASA Astrophysics Data System (ADS)

    Wang, Zhao; Tang, Haolin; Li, Junrui; Zeng, Yan; Chen, Lutang; Pan, Mu

    2014-06-01

    The effects of the nature of the solvent on the morphology of perfluorosulfonate ionomers in dispersions and the microstructures of the corresponding formed membranes are investigated. The subsequent electrochemical performances of the formed proton exchange membranes are also studied in detail. It is found that the diameters of the Nafion molecular aggregates in variable solutions decrease with an increase in the solvent dielectric constant (ε) and a decrease in the gap of the solubility parameters (δ) between the resin and the solvent. The micromorphology of Nafion is further examined by means of transmission electron microscopy, small-angle X-ray scattering, and X-ray diffraction. It is found that the membrane cast from a Nafion-2-propanol/water dispersion with ε value of 42.38 and a δ gap of 0.01 (cal cm-3)1/2 has a better ion cluster arrangement, smaller ion cluster size (approximately 13 Å), and higher crystallinity (16.7%) than the other samples. The electrochemical properties of the formed membranes are further investigated as a function of temperature, relative humidity, and the solubility parameter of the applied solvents. The results demonstrate that the electrochemical performance is strongly influenced by the solvent-induced microstructure of the backbone and the ionic clusters in the perfluorosulfonic acid membrane.

  13. Boosting with Averaged Weight Vectors

    NASA Technical Reports Server (NTRS)

    Oza, Nikunj C.; Clancy, Daniel (Technical Monitor)

    2002-01-01

    AdaBoost is a well-known ensemble learning algorithm that constructs its constituent or base models in sequence. A key step in AdaBoost is constructing a distribution over the training examples to create each base model. This distribution, represented as a vector, is constructed to be orthogonal to the vector of mistakes made by the previous base model in the sequence. The idea is to make the next base model's errors uncorrelated with those of the previous model. Some researchers have pointed out the intuition that it is probably better to construct a distribution that is orthogonal to the mistake vectors of all the previous base models, but that this is not always possible. We present an algorithm that attempts to come as close as possible to this goal in an efficient manner. We present experimental results demonstrating significant improvement over AdaBoost and the Totally Corrective boosting algorithm, which also attempts to satisfy this goal.

  14. Two types of rare earth-organic frameworks constructed by racemic tartaric acid

    SciTech Connect

    Jiang Zhanguo; Lv Yaokang; Cheng Jianwen; Feng Yunlong

    2012-01-15

    Hydrothermal reactions of rare earth oxides with racemic tartaric acid (H{sub 2}tar) yielded 7 rare earth(III) MOFs with general formulas [R{sub 2}(tar){sub 3}(H{sub 2}O){sub 2}]{sub n} (R=Y (1), Sm (4), Eu (5), Tb (6), Dy (7)) and [R{sub 2}(tar){sub 2}(C{sub 2}O{sub 4})(H{sub 2}O){sub 2}]{sub n}{center_dot}4nH{sub 2}O (R=La (2), Nd (3)). X-ray powder diffraction analysis and single-crystal X-ray diffraction analysis reveal that they present two different structural types. MOFs 1, 4, 5, 6 and 7 are isostructural and crystallize in the orthorhombic non-centrosymmetric space group Iba2, and feature unusual fsc-3,4-Iba2 topology. MOFs 2 and 3 are isostructural and crystallize in monoclinic P2{sub 1}/c space group and display rare fsx-4,5-P2{sub 1}/c topology containing hydrophilic channels bounded by triple helical chains along a axis. MOFs 3, 4, 5, 6 and 7 exhibit intense lanthanide characteristic photoluminescence at room temperature. - Graphical Abstract: [R{sub 2}(tar){sub 2}(C{sub 2}O{sub 4})(H{sub 2}O){sub 2}]{sub n}{center_dot}4nH{sub 2}O (R=La (2), Nd (3)) display rare fsx-4,5-P2{sub 1}/c topology containing hydrophilic channels bounded by triple helical chains along a axis. Highlights: Black-Right-Pointing-Pointer Two types of rare earth MOFs were synthesized by hydrothermal conditions. Black-Right-Pointing-Pointer MOFs feature unusual fsc-3,4-Iba2 and rare fsx-4,5-P2{sub 1}/c topology structures. Black-Right-Pointing-Pointer MOFs exhibit intense lanthanide characteristic photoluminescence.

  15. Molecular beacon mediated circular strand displacement strategy for constructing a ratiometric electrochemical deoxyribonucleic acid sensor.

    PubMed

    Gao, Fenglei; Du, Lili; Zhang, Yu; Tang, Daoquan; Du, Yan

    2015-07-01

    A novel ratiometric electrochemical sensor for sensitive and selective determination of deoxyribonucleic acid (DNA) had been developed based on signal-on and signal-off strategy. The target DNA hybridized with the loop portion of ferrocene (Fc) labeled hairpin probe immobilized on the gold electrode (GE), the Fc away from the surface of GE and the methylene blue (MB) was attached to an electrode surface by hybridization between hairpin probe and MB labeled primer. Such conformational changes resulted in the oxidation peak current of Fc decreased and that of MB increased, and the changes of dual signals are linear with the concentration of DNA. Furthermore, with the help of strand-displacement polymerization, polymerase catalyzed the extension of the primer and the sequential displacement of the target DNA, which led to the release of target and another polymerization cycle. Thus the circular strand displacement produced the multiplication of the MB confined near the GE surface and Fc got away from the GE surface. Therefore, the recognition of target DNA resulted in both the "signal-off" of Fc and the "signal-on" of MB for dual-signal electrochemical ratiometric readout. The dual signal strategy offered a dramatic enhancement of the stripping response. The dynamic range of the target DNA detection was from 10(-13) to 10(-8) mol L(-1) with a detection limit down to 28 fM level. Compared with the single signaling electrochemical sensor, the dual-signaling electrochemical sensing strategy developed in this paper was more selective. It would have important applications in the sensitive and selective electrochemical determination of other small molecules and proteins. PMID:26088778

  16. Multicistronic lentiviral vector-mediated striatal gene transfer of aromatic L-amino acid decarboxylase, tyrosine hydroxylase, and GTP cyclohydrolase I induces sustained transgene expression, dopamine production, and functional improvement in a rat model of Parkinson's disease.

    PubMed

    Azzouz, Mimoun; Martin-Rendon, Enca; Barber, Robert D; Mitrophanous, Kyriacos A; Carter, Emma E; Rohll, Jonathan B; Kingsman, Susan M; Kingsman, Alan J; Mazarakis, Nicholas D

    2002-12-01

    Parkinson's disease (PD) is a neurodegenerative disorder characterized by the selective loss of dopaminergic neurons in the substantia nigra. This loss leads to complete dopamine depletion in the striatum and severe motor impairment. It has been demonstrated previously that a lentiviral vector system based on equine infectious anemia virus (EIAV) gives rise to highly efficient and sustained transduction of neurons in the rat brain. Therefore, a dopamine replacement strategy using EIAV has been investigated as a treatment in the 6-hydroxydopamine (6-OHDA) animal model of PD. A self-inactivating EIAV minimal lentiviral vector that expresses tyrosine hydroxylase (TH), aromatic amino acid dopa decarboxylase (AADC), and GTP cyclohydrolase 1 (CH1) in a single transcription unit has been generated. In cultured striatal neurons transduced with this vector, TH, AADC, and CH1 proteins can all be detected. After stereotactic delivery into the dopamine-denervated striatum of the 6-OHDA-lesioned rat, sustained expression of each enzyme and effective production of catecholamines were detected, resulting in significant reduction of apomorphine-induced motor asymmetry compared with control animals (p < 0.003). Expression of each enzyme in the striatum was observed for up to 5 months after injection. These data indicate that the delivery of three catecholaminergic synthetic enzymes by a single lentiviral vector can achieve functional improvement and thus open the potential for the use of this vector for gene therapy of late-stage PD patients. PMID:12451130

  17. Enhancement of azo dye Acid Orange 7 removal in newly developed horizontal subsurface-flow constructed wetland.

    PubMed

    Tee, Heng-Chong; Lim, Poh-Eng; Seng, Chye-Eng; Mohd Nawi, Mohd Asri; Adnan, Rohana

    2015-01-01

    Horizontal subsurface-flow (HSF) constructed wetland incorporating baffles was developed to facilitate upflow and downflow conditions so that the treatment of pollutants could be achieved under multiple aerobic, anoxic and anaerobic conditions sequentially in the same wetland bed. The performances of the baffled and conventional HSF constructed wetlands, planted and unplanted, in the removal of azo dye Acid Orange 7 (AO7) were compared at the hydraulic retention times (HRT) of 5, 3 and 2 days when treating domestic wastewater spiked with AO7 concentration of 300 mg/L. The planted baffled unit was found to achieve 100%, 83% and 69% AO7 removal against 73%, 46% and 30% for the conventional unit at HRT of 5, 3 and 2 days, respectively. Longer flow path provided by baffled wetland units allowed more contact of the wastewater with the rhizomes, microbes and micro-aerobic zones resulting in relatively higher oxidation reduction potential (ORP) and enhanced performance as kinetic studies revealed faster AO7 biodegradation rate under aerobic condition. In addition, complete mineralization of AO7 was achieved in planted baffled wetland unit due to the availability of a combination of aerobic, anoxic and anaerobic conditions. PMID:25284799

  18. Microbial populations identified by fluorescence in situ hybridization in a constructed wetland treating acid coal mine drainage

    SciTech Connect

    Nicomrat, D.; Dick, W.A.; Tuovinen, O.H.

    2006-07-15

    Microorganisms are an integral part of the biogeochemical processes in wetlands, yet microbial communities in sediments within constructed wetlands receiving acid mine drainage (AMD) are only poorly understood. The purpose of this study was to characterize the microbial diversity and abundance in a wetland receiving AMD using fluorescence in situ hybridization (FISH) analysis. Seasonal samples of oxic surface sediments, comprised of Fe(III) precipitates, were collected from two treatment cells of the constructed wetland system. The pH of the bulk samples ranged between pH 2.1 and 3.9. Viable counts of acidophilic Fe and S oxidizers and heterotrophs were determined with a most probable number (MPN) method. The MPN counts were only a fraction of the corresponding FISH counts. The sediment samples contained microorganisms in the Bacteria (including the subgroups of acidophilic Fe- and S-oxidizing bacteria and Acidiphilium spp.) and Eukarya domains. Archaea were present in the sediment surface samples at < 0.01% of the total microbial community. The most numerous bacterial species in this wetland system was Acidithiobacillus ferrooxidans, comprising up to 37% of the bacterial population. Acidithiobacillus thiooxidans was also abundant.

  19. Prediction of apoptosis protein locations with genetic algorithms and support vector machines through a new mode of pseudo amino acid composition.

    PubMed

    Kandaswamy, Krishna Kumar; Pugalenthi, Ganesan; Möller, Steffen; Hartmann, Enno; Kalies, Kai-Uwe; Suganthan, P N; Martinetz, Thomas

    2010-12-01

    Apoptosis is an essential process for controlling tissue homeostasis by regulating a physiological balance between cell proliferation and cell death. The subcellular locations of proteins performing the cell death are determined by mostly independent cellular mechanisms. The regular bioinformatics tools to predict the subcellular locations of such apoptotic proteins do often fail. This work proposes a model for the sorting of proteins that are involved in apoptosis, allowing us to both the prediction of their subcellular locations as well as the molecular properties that contributed to it. We report a novel hybrid Genetic Algorithm (GA)/Support Vector Machine (SVM) approach to predict apoptotic protein sequences using 119 sequence derived properties like frequency of amino acid groups, secondary structure, and physicochemical properties. GA is used for selecting a near-optimal subset of informative features that is most relevant for the classification. Jackknife cross-validation is applied to test the predictive capability of the proposed method on 317 apoptosis proteins. Our method achieved 85.80% accuracy using all 119 features and 89.91% accuracy for 25 features selected by GA. Our models were examined by a test dataset of 98 apoptosis proteins and obtained an overall accuracy of 90.34%. The results show that the proposed approach is promising; it is able to select small subsets of features and still improves the classification accuracy. Our model can contribute to the understanding of programmed cell death and drug discovery. The software and dataset are available at http://www.inb.uni-luebeck.de/tools-demos/apoptosis/GASVM. PMID:20666727

  20. Construction of an oral vaccine for transmissible gastroenteritis virus based on the TGEV N gene expressed in an attenuated Salmonella typhimurium vector.

    PubMed

    Zhang, Dan; Huang, Xiaobo; Zhang, Xiaohui; Cao, Sanjie; Wen, Xintian; Wen, Yiping; Wu, Rui; Liang, Entao

    2016-01-01

    This research aimed to develop an oral vaccine for transmissible gastroenteritis virus (TGEV) based on the TGEV N gene expressed in an attenuated Salmonella typhimurium vector and aimed to evaluate the vaccine's immune response in piglets. Recombinant plasmid pVAX-N was transformed into competent cells of attenuated S. typhimurium SL7207 via electroporation. After it was identified via RT-PCR and double digestion, the screened recombinant bacteria presenting pVAX-N were named SL7207 (pVAX-N). To evaluate the safety and stability of the developed vaccine, different dosages (5 × 10(8), 1 × 10(9), and 2 × 10(9) CFU/mice) of SL7207 (pVAX-N) were inoculated to 6-week-old mice. Piglets below 20 days of age were dosed with 1 × 10(12) CFU. Humoral (neutralization titer and specific IgG), cellular (interleukin-4, γ-interferon, and peripheral lymphocyte proliferation), and mucosal (sIgA) immune responses were detected and evaluated. The three immunizing dosages were determined to be safe for mice and were completely eliminated 8 weeks after the first inoculation. Results of antibody and cytokine detection indicated that SL7207 (pVAX-N) could significantly induce antibody-IgG, antibody-IgA, interleukin-4, and γ-interferon, whose value was maximized on the 6th week. Results confirmed that the recombinant vaccine increased the proliferation of peripheral T lymphocyte. In conclusion, the oral vaccine was developed successfully, and the vaccine could significantly induce humoral, cellular, and mucosal immune responses in piglets. PMID:26297958

  1. Construction of Poxviruses as Cloning Vectors: Insertion of the Thymidine Kinase Gene from Herpes Simplex Virus into the DNA of Infectious Vaccinia Virus

    NASA Astrophysics Data System (ADS)

    Panicali, Dennis; Paoletti, Enzo

    1982-08-01

    We have constructed recombinant vaccinia viruses containing the thymidine kinase gene from herpes simplex virus. The gene was inserted into the genome of a variant of vaccinia virus that had undergone spontaneous deletion as well as into the 120-megadalton genome of the large prototypic vaccinia variant. This was accomplished via in vivo recombination by contransfection of eukaryotic tissue culture cells with cloned BamHI-digested thymidine kinase gene from herpes simplex virus containing flanking vaccinia virus DNA sequences and infectious rescuing vaccinia virus. Pure populations of the recombinant viruses were obtained by replica filter techniques or by growth of the recombinant virus in biochemically selective medium. The herpes simplex virus thymidine kinase gene, as an insert in vaccinia virus, is transcribed in vivo and in vitro, and the fidelity of in vivo transcription into a functional gene product was detected by the phosphorylation of 5-[125I]iodo-2'-deoxycytidine.

  2. Microbial populations identified by fluorescence in situ hybridization in a constructed wetland treating acid coal mine drainage.

    PubMed

    Nicomrat, Duongruitai; Dick, Warren A; Tuovinen, Olli H

    2006-01-01

    Microorganisms are an integral part of the biogeochemical processes in wetlands, yet microbial communities in sediments within constructed wetlands receiving acid mine drainage (AMD) are only poorly understood. The purpose of this study was to characterize the microbial diversity and abundance in a wetland receiving AMD using fluorescence in situ hybridization (FISH) analysis. Seasonal samples of oxic surface sediments, comprised of Fe(III) precipitates, were collected from two treatment cells of the constructed wetland system. The pH of the bulk samples ranged between pH 2.1 and 3.9. Viable counts of acidophilic Fe and S oxidizers and heterotrophs were determined with a most probable number (MPN) method. The MPN counts were only a fraction of the corresponding FISH counts. The sediment samples contained microorganisms in the Bacteria (including the subgroups of acidophilic Fe- and S-oxidizing bacteria and Acidiphilium spp.) and Eukarya domains. Archaea were present in the sediment surface samples at < 0.01% of the total microbial community. The most numerous bacterial species in this wetland system was Acidithiobacillus ferrooxidans, comprising up to 37% of the bacterial population. Acidithiobacillus thiooxidans was also abundant. Heterotrophs in the Acidiphilium genus totaled 20% of the bacterial population. Leptospirillum ferrooxidans was below the level of detection in the bacterial community. The results from the FISH technique from this field study are consistent with results from other experiments involving enumeration by most probable number, dot-blot hybridization, and denaturing gradient gel electrophoresis analyses and with the geochemistry of the site. PMID:16825452

  3. Polyelectrolyte Complexes of Low Molecular Weight PEI and Citric Acid as Efficient and Nontoxic Vectors for in Vitro and in Vivo Gene Delivery.

    PubMed

    Giron-Gonzalez, M Dolores; Salto-Gonzalez, Rafael; Lopez-Jaramillo, F Javier; Salinas-Castillo, Alfonso; Jodar-Reyes, Ana Belen; Ortega-Muñoz, Mariano; Hernandez-Mateo, Fernando; Santoyo-Gonzalez, Francisco

    2016-03-16

    Gene transfection mediated by the cationic polymer polyethylenimine (PEI) is considered a standard methodology. However, while highly branched PEIs form smaller polyplexes with DNA that exhibit high transfection efficiencies, they have significant cell toxicity. Conversely, low molecular weight PEIs (LMW-PEIs) with favorable cytotoxicity profiles display minimum transfection activities as a result of inadequate DNA complexation and protection. To solve this paradox, a novel polyelectrolyte complex was prepared by the ionic cross-linking of branched 1.8 kDa PEI with citric acid (CA). This system synergistically exploits the good cytotoxicity profile exhibited by LMW-PEI with the high transfection efficiencies shown by highly branched and high molecular weight PEIs. The polyectrolyte complex (1.8 kDa-PEI@CA) was obtained by a simple synthetic protocol based on the microwave irradiation of a solution of 1.8 kDa PEI and CA. Upon complexation with DNA, intrinsic properties of the resulting particles (size and surface charge) were measured and their ability to form stable polyplexes was determined. Compared with unmodified PEIs the new complexes behave as efficient gene vectors and showed enhanced DNA binding capability associated with facilitated intracellular DNA release and enhanced DNA protection from endonuclease degradation. In addition, while transfection values for LMW-PEIs are almost null, transfection efficiencies of the new reagent range from 2.5- to 3.8-fold to those of Lipofectamine 2000 and 25 kDa PEI in several cell lines in culture such as CHO-k1, FTO2B hepatomas, L6 myoblasts, or NRK cells, simultaneously showing a negligible toxicity. Furthermore, the 1.8 kDa-PEI@CA polyelectrolyte complexes retained the capability to transfect eukaryotic cells in the presence of serum and exhibited the capability to promote in vivo transfection in mouse (as an animal model) with an enhanced efficiency compared to 25 kDa PEI. Results support the polyelectrolyte complex

  4. Corrosion resistance and behavior of construction materials exposed to dilute sulfuric acid at elevated temperatures under static conditions

    SciTech Connect

    Nguyen, D.T.

    1994-10-01

    Laboratory investigation has been undertaken to determine the electrochemical behavior and corrosion resistance of various construction materials in a simulated hydrolysis environment (5 wt % sulfuric acid) at temperatures ranging from 90 to 220C. Tests were performed in an autoclave-type electrochemical cell. The corrosion behavior of the test materials was determined using computer-controlled DC potentiodynamic polarization. Corrosion rates of the test materials were determined using AC impedance techniques. Among the stainless steels tested, only alloy N08026 (Carpenter 20Mo-6) performed satisfactory up to a temperature of 100C. The alloy passivated spontaneously in the environment and corroded at a rate of less than 2 mpy. None of the stainless steels tested could be used at 120{degrees}C or above. A number of nickel-based alloys tested had good corrosion resistance up to 100C, but their corrosion rate exceeded 2 mpy at higher temperatures. Zirconium alloys were satisfactory up to 180C. Only tantalum and a tantalum-niobium alloy were satisfactory up to 220C.

  5. Construction of transplastomic lettuce (Lactuca sativa) dominantly producing astaxanthin fatty acid esters and detailed chemical analysis of generated carotenoids.

    PubMed

    Harada, Hisashi; Maoka, Takashi; Osawa, Ayako; Hattan, Jun-Ichiro; Kanamoto, Hirosuke; Shindo, Kazutoshi; Otomatsu, Toshihiko; Misawa, Norihiko

    2014-04-01

    The plastid genome of lettuce (Lactuca sativa L.) cv. Berkeley was site-specifically modified with the addition of three transgenes, which encoded β,β-carotenoid 3,3'-hydroxylase (CrtZ) and β,β-carotenoid 4,4'-ketolase (4,4'-oxygenase; CrtW) from a marine bacterium Brevundimonas sp. strain SD212, and isopentenyl diphosphate isomerase from a marine bacterium Paracoccus sp. strain N81106. Constructed transplastomic lettuce plants were able to grow on soil at a growth rate similar to that of non-transformed lettuce cv. Berkeley and generate flowers and seeds. The germination ratio of the lettuce transformants (T0) (98.8%) was higher than that of non-transformed lettuce (93.1 %). The transplastomic lettuce (T1) leaves produced the astaxanthin fatty acid (myristate or palmitate) diester (49.2% of total carotenoids), astaxanthin monoester (18.2%), and the free forms of astaxanthin (10.0%) and the other ketocarotenoids (17.5%), which indicated that artificial ketocarotenoids corresponded to 94.9% of total carotenoids (230 μg/g fresh weight). Native carotenoids were there lactucaxanthin (3.8%) and lutein (1.3 %) only. This is the first report to structurally identify the astaxanthin esters biosynthesized in transgenic or transplastomic plants producing astaxanthin. The singlet oxygen-quenching activity of the total carotenoids extracted from the transplastomic leaves was similar to that of astaxanthin (mostly esterified) from the green algae Haematococcus pluvialis. PMID:24287848

  6. Sequence analysis and characterization of pOM1, a small cryptic plasmid from Butyrivibrio fibrisolvens, and its use in construction of a new family of cloning vectors for Butyrivibrios.

    PubMed Central

    Hefford, M A; Kobayashi, Y; Allard, S E; Forster, R J; Teather, R M

    1997-01-01

    As a preliminary step in the development of vector systems, we have isolated and begun to characterize small, cryptic plasmids from several strains of the rumen bacterium Butyrivibrio fibrisolvens. We present here the complete nucleotide sequence of Butyrivibrio plasmid pOM1, which was isolated from B. fibrisolvens Bu49. While it is very similar in size to the previously characterized Butyrivibrio plasmids pRJF1 and pRJF2, pOM1 exhibits a restriction pattern which is quite distinct. Analysis of sequence data reveals that pOM1 contains only two open reading frames of significant length (ORF1 and ORF2), both of which are required for self-replication and maintenance. The protein encoded in ORF1 shows homologies with Pre (plasmid recombination enzyme) proteins encoded in plasmids from gram-positive organisms such as Staphylococcus aureus, Streptococcus agalactiae, Lactobacillus plantarum, and Bacillus thuringiensis. The putative translation product of ORF2, on the other hand, resembles Rep (replication) proteins of a different group of gram-positive plasmids, for which the Staphylococcus plasmid pSN2 is a prototype. Unlike the other characterized-Butyrivibrio plasmids, pOM1 appears to replicate via a rolling-circle mechanism. Experimental evidence showing the presence of a single-stranded replication intermediate consistent with this mechanism is presented. pOM1 has been used in the construction of a new Escherichia coli-B. fibrisolvens shuttle vector, pSMerm1, which has been successfully used to introduce a cloned gene into B. fibrisolvens harboring the pRJF1 plasmid. PMID:9143105

  7. Recombinant adeno-associated virus-mediated high-efficiency, transient expression of the murine cationic amino acid transporter (ecotropic retroviral receptor) permits stable transduction of human HeLa cells by ecotropic retroviral vectors.

    PubMed Central

    Bertran, J; Miller, J L; Yang, Y; Fenimore-Justman, A; Rueda, F; Vanin, E F; Nienhuis, A W

    1996-01-01

    Adeno-associated virus has a broad host range, is nonpathogenic, and integrates into a preferred location on chromosome 19, features that have fostered development of recombinant adeno-associated viruses (rAAV) as gene transfer vectors for therapeutic applications. We have used an rAAV to transfer and express the murine cationic amino acid transporter which functions as the ecotropic retroviral receptor, thereby rendering human cells conditionally susceptible to infection by an ecotropic retroviral vector. The proportion of human HeLa cells expressing the receptor at 60 h varied as a function of the multiplicity of infection (MOI) with the rAAV. Cells expressing the ecotropic receptor were efficiently transduced with an ecotropic retroviral vector encoding a nucleus-localized form of beta-galactosidase. Cells coexpressing the ecotropic receptor and nucleus-localized beta-galactosidase were isolated by fluorescence-activated cell sorting, and cell lines were recovered by cloning at limiting dilution. After growth in culture, all clones contained the retroviral vector genome, but fewer than 10% (3 of 47) contained the rAAV genome and continued to express the ecotropic receptor. The ecotropic receptor coding sequences in the rAAV genome were under the control of a tetracycline-modulated promoter. In the presence of tetracycline, receptor expression was low and the proportion of cells transduced by the ecotropic retroviral vector was decreased. Modulation of receptor expression was achieved with both an episomal and an integrated form of the rAAV genome. These data establish that functional gene expression from an rAAV genome can occur transiently without genome integration. PMID:8794313

  8. A proof of van Aubel's theorem using orthogonal vectors

    NASA Astrophysics Data System (ADS)

    Glaister, P.

    2016-04-01

    We show how two linearly independent vectors can be used to construct two orthogonal vectors of equal magnitude in a simple way. The proof that the constructed vectors are orthogonal and of equal magnitude is a good exercise for students studying properties of scalar and vector triple products. We then show how this result can be used to prove van Aubel's theorem that relates the two line segments joining the centres of squares on opposite sides of a plane quadrilateral.

  9. Design, Construction and Cloning of Truncated ORF2 and tPAsp-PADRE-Truncated ORF2 Gene Cassette From Hepatitis E Virus in the pVAX1 Expression Vector

    PubMed Central

    Farshadpour, Fatemeh; Makvandi, Manoochehr; Taherkhani, Reza

    2015-01-01

    Background: Hepatitis E Virus (HEV) is the causative agent of enterically transmitted acute hepatitis and has high mortality rate of up to 30% among pregnant women. Therefore, development of a novel vaccine is a desirable goal. Objectives: The aim of this study was to construct tPAsp-PADRE-truncated open reading frame 2 (ORF2) and truncated ORF2 DNA plasmid, which can assist future studies with the preparation of an effective vaccine against Hepatitis E Virus. Materials and Methods: A synthetic codon-optimized gene cassette encoding tPAsp-PADRE-truncated ORF2 protein was designed, constructed and analyzed by some bioinformatics software. Furthermore, a codon-optimized truncated ORF2 gene was amplified by the polymerase chain reaction (PCR), with a specific primer from the previous construct. The constructs were sub-cloned in the pVAX1 expression vector and finally expressed in eukaryotic cells. Results: Sequence analysis and bioinformatics studies of the codon-optimized gene cassette revealed that codon adaptation index (CAI), GC content, and frequency of optimal codon usage (Fop) value were improved, and performance of the secretory signal was confirmed. Cloning and sub-cloning of the tPAsp-PADRE-truncated ORF2 gene cassette and truncated ORF2 gene were confirmed by colony PCR, restriction enzymes digestion and DNA sequencing of the recombinant plasmids pVAX-tPAsp-PADRE-truncated ORF2 (aa 112-660) and pVAX-truncated ORF2 (aa 112-660). The expression of truncated ORF2 protein in eukaryotic cells was approved by an Immunofluorescence assay (IFA) and the reverse transcriptase polymerase chain reaction (RT-PCR) method. Conclusions: The results of this study demonstrated that the tPAsp-PADRE-truncated ORF2 gene cassette and the truncated ORF2 gene in recombinant plasmids are successfully expressed in eukaryotic cells. The immunogenicity of the two recombinant plasmids with different formulations will be evaluated as a novel DNA vaccine in future investigations. PMID

  10. Recreating the tumor microenvironment in a bilayer, hyaluronic acid hydrogel construct for the growth of prostate cancer spheroids.

    PubMed

    Xu, Xian; Gurski, Lisa A; Zhang, Chu; Harrington, Daniel A; Farach-Carson, Mary C; Jia, Xinqiao

    2012-12-01

    Cancer cells cultured in physiologically relevant, three-dimensional (3D) matrices can recapture many essential features of native tumor tissues. In this study, a hyaluronic acid (HA)-based bilayer hydrogel system that not only supports the tumoroid formation from LNCaP prostate cancer (PCa) cells, but also simulates their reciprocal interactions with the tumor-associated stroma was developed and characterized. HA hydrogels were prepared by mixing solutions of HA precursors functionalized with acrylate groups (HA-AC) and reactive thiols (HA-SH) under physiological conditions. The resultant viscoelastic gels have an average elastic modulus of 234 ± 30 Pa and can be degraded readily by hyaluronidase. The orthogonal and cytocompatible nature of the crosslinking chemistry permits facile incorporation of cytokine-releasing particles and PCa cells. In our bilayer hydrogel construct, the top layer contains heparin (HP)-decorated, HA-based hydrogel particles (HGPs) capable of releasing heparin-binding epidermal growth factor-like growth factor (HB-EGF) in a sustained manner at a rate of 2.5 wt%/day cumulatively. LNCaP cells embedded in the bottom layer receive the growth factor signals from the top, and in response form enlarging tumoroids with an average diameter of 85 μm by day 7. Cells in 3D hydrogels assemble into spherical tumoroids, form close cellular contacts through E-cadherin, and show cortical organization of F-actin, whereas those plated as 2D monolayers adopt a spread-out morphology. Compared to cells cultured on 2D, the engineered tumoroids significantly increased the expression of two pro-angiogenic factors, vascular endothelial growth factor-165 (VEGF(165)) and interleukin-8 (IL-8), both at mRNA and protein levels. Overall, the HA model system provides a useful platform for the study of tumor cell responses to growth factors and for screening of anticancer drugs targeting these pathways. PMID:22999468

  11. Recreating the Tumor Microenvironment in a Bilayer, Hyaluronic Acid Hydrogel Construct for the Growth of Prostate Cancer Spheroids

    PubMed Central

    Xu, Xian; Gurski, Lisa A.; Zhang, Chu; Harrington, Daniel A.; Farach-Carson, Mary C.; Jia, Xinqiao

    2012-01-01

    Cancer cells cultured in physiologically relevant, three-dimensional (3D) matrices can recapture many essential features of native tumor tissues. In this study, a hyaluronic acid (HA)-based bilayer hydrogel system that not only supports the tumoroid formation from LNCaP prostate cancer (PCa) cells, but also simulates their reciprocal interactions with the tumor-associated stroma was developed and characterized. HA hydrogels were prepared by mixing solutions of HA precursors functionalized with acrylate groups (HA-AC) and reactive thiols (HA-SH) under physiological conditions. The resultant viscoelastic gels have an average elastic modulus of 234 ± 30 Pa and can be degraded readily by hyaluronidase. The orthogonal and cytocompatible nature of the crosslinking chemistry permits facile incorporation of cytokine-releasing particles and PCa cells. In our bilayer hydrogel construct, the top layer contains heparin (HP)-decorated, HA-based hydrogel particles (HGPs) capable of releasing heparin-binding epidermal growth factor-like growth factor (HB-EGF) in a sustained manner at a rate of 2.5wt%/day cumulatively. LNCaP cells embedded in the bottom layer receive the growth factor signals from the top, and in response form enlarging tumoroids with an average diameter of 85 μm by day 7. Cells in 3D hydrogels assemble into spherical tumoroids, form close cellular contacts through E-cadherin, and show cortical organization of F-actin, whereas those plated as 2D monolayers adopt a spread-out morphology. Compared to cells cultured on 2D, the engineered tumoroids significantly increased the expression of two pro-angiogenic factors, vascular endothelial growth factor-165 (VEGF165) and interleukin-8 (IL-8), both at mRNA and protein levels. Overall, the HA model system provides a useful platform for the study of tumor cell responses to growth factors and for screening of anticancer drugs targeting these pathways. PMID:22999468

  12. Copper(II)-lanthanide(III) coordination polymers constructed from pyridine-2,5-dicarboxylic acid: Preparation, crystal structure and photoluminescence

    SciTech Connect

    Xia Zhengqiang; Wei Qing; Chen Sanping; Feng Xinming; Xie Gang; Qiao Chengfang; Zhang Guochun; Gao Shengli

    2013-01-15

    A series of 3d-4f heterometallic coordination polymers, formulated as {l_brace} [Cu{sub 3}Ln{sub 2}(pydc){sub 6}(H{sub 2}O){sub 12}]{center_dot}4H{sub 2}O{r_brace} {sub n} [Ln=Tb (1), Eu (2), Dy (3), Ho (4), Lu (5)], {l_brace} [CuNd{sub 2}(pydc){sub 4}(H{sub 2}O){sub 3}]{center_dot}H{sub 2}O{r_brace} {sub n} (6) and {l_brace} [Cu{sub 3}Pr{sub 2}(pydc){sub 6}(H{sub 2}O){sub 13}]{center_dot}4H{sub 2}O{r_brace} {sub n} (7) (where H{sub 2}pydc=pyridine-2,5-dicarboxylic acid), have been hydrothermally prepared by reactions of H{sub 2}pydc ligand with lanthanide ions in the presence of Cu(II) ion. X-ray crystal structure analysis reveals that these compounds exhibit rich structural chemistry. 1-5 are isomorphous and present a two-dimensional network constructed from Ln{sub 2}Cu{sub 2}L{sub 2}(H{sub 2}O){sub 2} SBU rings and CuL{sub 2}(H{sub 2}O) building blocks. In 6, two-dimensional ladder-like layers based on Nd(III) belts and CuL{sub 2}O{sub 2} units are assembled by H{sub 2}pydc ligands into a three-dimensional open framework. Polymer 7 displays a two-dimensional wave-like layer structure containing two distinct ring units, in which a new coordination mode of the pydc{sup 2-} ligand is observed. The results indicate that the coordination flexibility of the pydc{sup 2-} ligand and lanthanide contraction effect play cooperative roles in the formation of coordination polymers with different polymeric architectures. Compounds 1-2 exhibit intense green and red luminescence emission characteristics of Tb(III) and Eu(III), respectively. Furthermore, elemental analyses (EA), infrared spectra (IR) and thermogravimetric analyses (TGA) of these compounds were also studied. - Graphical abstract: Seven 3d-4f heterometallic coordination polymers were synthesized by reactions of H{sub 2}pydc with lanthanide metal ions in the presence of Cu{sup 2+}, the effects of Cu{sup 2+} on the structures and photoluminescent properties of Ln-pydc{sup 2-} systems were investigated. Highlights

  13. Construction of a colicin E1-R factor composite plasmid in vitro: means for amplification of deoxyribonucleic acid.

    PubMed Central

    Tanaka, T; Weisblum, B

    1975-01-01

    A composite plasmid has been constructed in vitro from colicin E1 factor (mass of 4.2 megadaltons [Md]) and nontransmissible resistance factor RSF 1010 (mass, 5.5. Md) deoxyribonucleic acids (DNAs) by the sequential action of Escherichia coli endonuclease (RI (Eco RI) and T4 phage DNA ligase on the covalently closed circular forms of the constituents. The composite plasmid was selected and amplified in vivo by sequential transformation of E. coli C600 with the ligated mixture and selection of transformants in medium containing streptomycin plus colicin E1, followed by amplification in the presence of chloramphenicol and purification of the extracted plasmid by dye-buoyant density gradient centrifugation in ethidium bromide-cesium chloride solution. Treatment of the composite plasmid with Eco RI yielded two fragments with mobilities corresponding to the linear forms of the parental plasmids, whereas Serratia marscesens endonuclease R (SmaR), which introduces a single scission in the colicin E1 factor but not in RSF 1010, convErted the composite plasmid to a single linear molecule (mass, 9.7 Md). Sequential degradation of colicin E1 factor with Sma R and Eco RI produced two fragments with masses of 3.5 and 0.7 Md; sequential degradation of RSF 1010 produced only one fragment (due to the cleavage with Eco RI), and sequential degradation of the composite plasmid produced the expected three fragments--an RSF 1010 Eco RI linear and the two expected products from the colicin E1 factor moiety. The composite plasmid conferred on the host cell resistance to streptomycin, sulfonamides, and colicin E1, but colicin E1 itself was not synthesized. In contrast, colicin E1 was synthesized by cells containing simultaneously both colicin E1 factor and RSF 1010 as separate entities. In the presence of chloramphenicol, the composite plasmid continued to replicate for 6 h. whereas replication of RSF 1010 and chromosomal DNA stopped within 2 h. Continued replication in the presence of

  14. Integration of narrow-host-range vectors from Escherichia coli into the genomes of amino acid-producing corynebacteria after intergeneric conjugation.

    PubMed Central

    Mateos, L M; Schäfer, A; Kalinowski, J; Martin, J F; Pühler, A

    1996-01-01

    Conjugative transfer of mobilizable derivatives of the Escherichia coli narrow-host-range plasmids pBR322, pBR325, pACYC177, and pACYC184 from E. coli to species of the gram-positive genera Corynebacterium and Brevibacterium resulted in the integration of the plasmids into the genomes of the recipient bacteria. Transconjugants appeared at low frequencies and reproducibly with a delay of 2 to 3 days compared with matings with replicative vectors. Southern analysis of corynebacterial transconjugants and nucleotide sequences from insertion sites revealed that integration occurs at different locations and that different parts of the vector are involved in the process. Integration is not dependent on indigenous insertion sequence elements but results from recombination between very short homologous DNA segments (8 to 12 bp) present in the vector and in the host DNA. In the majority of the cases (90%), integration led to cointegrate formation, and in some cases, deletions or rearrangements occurred during the recombination event. Insertions were found to be quite stable even in the absence of selective pressure. PMID:8824624

  15. Eight supramolecular assemblies constructed from bis(benzimidazole) and organic acids through strong classical hydrogen bonding and weak noncovalent interactions

    NASA Astrophysics Data System (ADS)

    Jin, Shouwen; Wang, Daqi

    2014-05-01

    Eight crystalline organic acid-base adducts derived from alkane bridged bis(N-benzimidazole) and organic acids (2,4,6-trinitrophenol, p-nitrobenzoic acid, m-nitrobenzoic acid, 3,5-dinitrobenzoic acid, 5-sulfosalicylic acid and oxalic acid) were prepared and characterized by X-ray diffraction analysis, IR, mp, and elemental analysis. Of the eight compounds five are organic salts (1, 4, 6, 7 and 8) and the other three (2, 3, and 5) are cocrystals. In all of the adducts except 1 and 8, the ratio of the acid and the base is 2:1. All eight supramolecular assemblies involve extensive intermolecular classical hydrogen bonds as well as other noncovalent interactions. The role of weak and strong noncovalent interactions in the crystal packing is ascertained. These weak interactions combined, all the complexes displayed 3D framework structure. The results presented herein indicate that the strength and directionality of the classical N+-H⋯O-, O-H⋯O, and O-H⋯N hydrogen bonds (ionic or neutral) and other nonbonding associations between acids and ditopic benzimidazoles are sufficient to bring about the formation of cocrystals or organic salts.

  16. Relativistic Gamow vectors: State vectors for unstable particles

    NASA Astrophysics Data System (ADS)

    Kaldas, Hany Kamel Halim

    The relativistic Gamow vectors are derived from the analytic continuation of the angular momentum velocity kets to the resonance pole of the S- matrix. This construction is justifiable within a Rigged Hilbert Space of Hardy class functions. The kets obtained | p j3[ sRjR ]-> are characterized by a spin jR and a complex mass square sR = (MR - iΓ R/2)2. Our use of the velocity kets renders the Gamow vectors | p j3[ sRjR ]-> ``minimally complex'', as the 4-velocities p̂μ = p μ/ s are taken real and they remain real under Lorentz transformations. When the symmetry transformations of the Gamow vectors are considered, it is found that they obey a semigroup time evolution in the forward light cone for the subgroup of P with causal space- time translations, i.e., for space-time translations with 4-vectors x such that x2 >= 0. This semigroup evolution, which is a consequence of the characterization obtained for the Gamow vectors as functionals in a Rigged Hilbert Space, is in conformity with the time directedness associated with decay phenomena. The Gamow vectors, with a Breit-Wigner distribution and exponential decay law, provide a description of decaying particles with a wide range of Γ/ M. Moreover, the Gamow vectors, being members of a complex basis vector expansion, allow the Wigner-Weisskopf's based effective theories, such as the Lee-Oehme-Yang theory for the neutral K-mesons, to be obtained as an approximation in an exact formalism.

  17. Polypeptides having cellulolytic enhancing activity and nucleic acids encoding same

    DOEpatents

    Brown, Kimberly; Harris, Paul; Zaretsky, Elizabeth; Re, Edward; Vlasenko, Elena; McFarland, Keith; Lopez de Leon, Alfredo

    2016-08-09

    The present invention relates to isolated polypeptides having cellulolytic enhancing activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods for producing and using the polypeptides.

  18. Polypeptides having cellulolytic enhancing activity and nucleic acids encoding same

    SciTech Connect

    Brown, Kimberly; Harris, Paul; Zaretsky, Elizabeth; Re, Edward; Vlasenko, Elena; McFarland, Keith; Lopez de Leon, Alfredo

    2014-09-30

    The present invention relates to isolated polypeptides having cellulolytic enhancing activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods for producing and using the polypeptides.

  19. Polypeptides having cellulolytic enhancing activity and nucleic acids encoding same

    SciTech Connect

    Brown, Kimberly; Harris, Paul; Zaretsky, Elizabeth; Re, Edward; Vlasenko, Elena; McFarland, Keith; Lopez de Leon, Alfredo

    2012-10-16

    The present invention relates to isolated polypeptides having cellulolytic enhancing activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods for producing and using the polypeptides.

  20. Multicomponent hydrogen-bonding organic solids constructed from 6-hydroxy-2-naphthoic acid and N-heterocycles: Synthesis, structural characterization and synthon discussion

    NASA Astrophysics Data System (ADS)

    Zong, Yingxia; Shao, Hui; Pang, Yanyan; Wang, Debao; Liu, Kang; Wang, Lei

    2016-07-01

    Seven novel multicomponent crystals involving various substituted organic amine molecules and 6-hydroxy-2-naphthoic acid were prepared and characterized by using single crystal X-ray diffraction, infrared and thermogravimetric analyses (TGA). Crystal structures with 1,4-bis(imidazol) butane (L1) 1, 1,4-bis(imidazol-1-ylmethyl)benzene (L2) 2, 1-phenyl piperazine 3, 2-amino-4-hydroxy-6-methyl pyrimidine 4, 4,4'-bipyridine 5, 5,5'-dimethyl-2,2'-dipyridine 6, 2-amino-4,6-dimethyl pyrimidine 7 were determined. Among the seven molecular complexes, total proton transfer from 6-hydroxy-2-naphthoic acid to coformer has occurred in crystals 1-4, while the remaining were cocrystals. X-ray single-crystal structures of these complexes reveal that strong hydrogen bonding O-H···O/N-H···O/O-H···N and weak C-H···O/C-H···π/π···π intermolecular interactions direct the packing modes of molecular crystals together. The analysis of supramolecular synthons in the present structures shows that some classical supramolecular synthons like pyridine-carboxylic acid heterosynthon R22 (7) and aminopyridine-carboxylic acid heterosynthon R22 (8), are again observed in constructing the hydrogen-bonding networks in this paper. Besides, we noticed that water molecules act as a significant hydrogen-bonding connector in constructing supramolecular architectures of 3, 4, 6, and 7.

  1. Mutation in fiber of adenovirus serotype 5 gene therapy vector decreases liver tropism

    PubMed Central

    Wang, Zhen; Wang, Baoming; Lou, Junfang; Yan, Jingyi; Gao, Lei; Geng, Ranshen; Yu, Bin

    2014-01-01

    Recombinant adenovirus (Ad) vectors are widely used for both in vitro and in vivo gene transfer. However, intravenous administration of Ad vectors results mainly in hepatocyte transduction and subsequent hepatotoxicity. Coxsackie-adenovirus receptor (CAR) and αvβ integrins, which are functional receptors for the fiber and penton proteins, respectively, are the tropism determinants of Ad type 5 (Ad5). We previously developed a system for rapid construction of fiber-modified Ad5 vectors. We also constructed a fiber-modified Ad5 containing an Arg-Gly-Asp (RGD) motif in the HI-loop and showed that it could enhance anti-tumor effects in vitro and in vivo. Here, we constructed a novel Ad5 vector containing two amino acid mutations in the AB loop of the fiber-modified Ad5 fiber knob and showed that it could significantly reduce liver tropism and increase gene transfer in low-CAR or CAR-deficient cancer cells following intravascular delivery. However, anti-tumor effects of the fiber-mutated Ad5 expressing HSV-TK under control of the hTERT promoter was not found when compared with an unmodified Ad5 vector in cancer lines expressing different levels of CAR, likely due to the activity of the hTERT promoter being lower than that of the CMV promoter. Nevertheless, this study describes an enhanced Ad5 vector for intravascular gene delivery, and further modifications such as changes in the promoter may facilitate the development of this vector for cancer treatment. PMID:25663991

  2. Rotations with Rodrigues' Vector

    ERIC Educational Resources Information Center

    Pina, E.

    2011-01-01

    The rotational dynamics was studied from the point of view of Rodrigues' vector. This vector is defined here by its connection with other forms of parametrization of the rotation matrix. The rotation matrix was expressed in terms of this vector. The angular velocity was computed using the components of Rodrigues' vector as coordinates. It appears…

  3. Photoreceptor-targeted gene delivery using intravitreally administered AAV vectors in dogs

    PubMed Central

    Boyd, RF; Sledge, DG; Boye, SL; Boye, SE; Hauswirth, WW; Komáromy, AM; Petersen-Jones, SM; Bartoe, JT

    2016-01-01

    Delivery of therapeutic transgenes to retinal photoreceptors using adeno-associated virus (AAV) vectors has traditionally required subretinal injection. Recently, photoreceptor transduction efficiency following intravitreal injection (IVT) has improved in rodent models through use of capsid-mutant AAV vectors; but remains limited in large animal models. Thickness of the inner limiting membrane (ILM) in large animals is thought to impair retinal penetration by AAV. Our study compared two newly developed AAV vectors containing multiple capsid amino acid substitutions following IVT in dogs. The ability of two promoter constructs to restrict reporter transgene expression to photoreceptors was also evaluated. AAV vectors containing the interphotoreceptor-binding protein (IRBP) promoter drove expression exclusively in rod and cone photoreceptors, with transduction efficiencies of ~ 4% of cones and 2% of rods. Notably, in the central region containing the cone-rich visual streak, 15.6% of cones were transduced. Significant regional variation existed, with lower transduction efficiencies in the temporal regions of all eyes. This variation did not correlate with ILM thickness. Vectors carrying a cone-specific promoter failed to transduce a quantifiable percentage of cone photoreceptors. The newly developed AAV vectors containing the IRBP promoter were capable of producing photoreceptor-specific transgene expression following IVT in the dog. PMID:26467396

  4. Visible-light induced oxidant-free oxidative cross-coupling for constructing allylic sulfones from olefins and sulfinic acids.

    PubMed

    Zhang, Guoting; Zhang, Lingling; Yi, Hong; Luo, Yi; Qi, Xiaotian; Tung, Chen-Ho; Wu, Li-Zhu; Lei, Aiwen

    2016-08-16

    An oxidant-free dehydrogenative sulfonylation of α-methyl-styrene derivatives was developed for the construction of allylic sulfones by using eosin Y as a photosensitizer in conjunction with a cobaloxime catalyst. The process features a low-cost metal catalyst and atom economy, which provides an appealing strategy for future synthetic chemistry. PMID:27481529

  5. Are Bred Vectors The Same As Lyapunov Vectors?

    NASA Astrophysics Data System (ADS)

    Kalnay, E.; Corazza, M.; Cai, M.

    Regional loss of predictability is an indication of the instability of the underlying flow, where small errors in the initial conditions (or imperfections in the model) grow to large amplitudes in finite times. The stability properties of evolving flows have been studied using Lyapunov vectors (e.g., Alligood et al, 1996, Ott, 1993, Kalnay, 2002), singular vectors (e.g., Lorenz, 1965, Farrell, 1988, Molteni and Palmer, 1993), and, more recently, with bred vectors (e.g., Szunyogh et al, 1997, Cai et al, 2001). Bred vectors (BVs) are, by construction, closely related to Lyapunov vectors (LVs). In fact, after an infinitely long breeding time, and with the use of infinitesimal ampli- tudes, bred vectors are identical to leading Lyapunov vectors. In practical applications, however, bred vectors are different from Lyapunov vectors in two important ways: a) bred vectors are never globally orthogonalized and are intrinsically local in space and time, and b) they are finite-amplitude, finite-time vectors. These two differences are very significant in a dynamical system whose size is very large. For example, the at- mosphere is large enough to have "room" for several synoptic scale instabilities (e.g., storms) to develop independently in different regions (say, North America and Aus- tralia), and it is complex enough to have several different possible types of instabilities (such as barotropic, baroclinic, convective, and even Brownian motion). Bred vectors share some of their properties with leading LVs (Corazza et al, 2001a, 2001b, Toth and Kalnay, 1993, 1997, Cai et al, 2001). For example, 1) Bred vectors are independent of the norm used to define the size of the perturba- tion. Corazza et al. (2001) showed that bred vectors obtained using a potential enstro- phy norm were indistinguishable from bred vectors obtained using a streamfunction squared norm, in contrast with singular vectors. 2) Bred vectors are independent of the length of the rescaling period as long as the

  6. Supramolecular architectures constructed by lanthanum, amino acids and 1,10-phenanthroline via non-covalent bond interactions

    NASA Astrophysics Data System (ADS)

    Zheng, Xiang-Jun; Jin, Lin-Pei

    2003-07-01

    Three supramolecular lanthanum coordination compounds of amino acids, with 1,10-phenanthroline (phen), [La 2(APA) 6(phen) 2(H 2O) 2](ClO 4) 6(phen) 4·2H 2O ( 1), [La 2(ABA) 6(phen) 2(H 2O) 2](ClO 4) 6 (phen) 6·4H 2O ( 2), and [La 2(AHA) 4(phen) 4](ClO 4) 6(phen) 4·2H 2O ( 3) (APA=3-aminopropionic acid; ABA=4-aminobutanoic acid; AHA=6-aminohexanoic acid) were synthesized and characterized by single crystal X-ray diffraction. The results show that the three coordination compounds are all composed of binuclear coordination cations built by metal-ligand coordination. Through hydrogen bonding and π-π stacking interactions, complex 1 forms a two-dimensional supramolecular sheet structure extending in the (001) plane, complex 2 forms a three-dimensional supramolecular network with many cavities occupied by ClO 4- and lattice H 2O molecules, and complex 3 forms a two-dimensional supramolecular lamellar structure in the (100) plane.

  7. “Molecular trinity” for soft nanomaterials: Integrating nucleobases, amino acids, and glycosides to construct multifunctional hydrogelators

    PubMed Central

    Li, Xinming; Kuang, Yi

    2013-01-01

    This highlight introduces the development of hydrogelators consisting of nucleobases, amino acids, and glycosides (i.e., molecular trinity), or nucleobases and amino acids (i.e., nucleopeptides). These novel small molecule hydrogelators self-assemble in water to form stable supramolecular nanofibers/hydrogels and exhibit useful biological properties (e.g., biocompatibility, biostability, and the ability to bind and transport DNA into live cells). The approach discussed here not only provides a new strategy to develop soft biomaterials as a form of nanomedicines, but also contributes to the understanding of molecular self-assembly in water by modulating the non-covalent interactions derived from the three basic building blocks used in living organisms. PMID:24368929

  8. Conjugal Strategy for Construction of Fast Acid-Producing, Bacteriophage-Resistant Lactic Streptococci for Use in Dairy Fermentations

    PubMed Central

    Sanders, M. E.; Leonhard, P. J.; Sing, W. D.; Klaenhammer, T. R.

    1986-01-01

    Bacteriophage-resistant dairy streptococci were obtained following conjugal transfer of pTR2030 from a lactose-negative donor, Streptococcus lactis TEK12, to lactose-positive recipient strains, Streptococcus cremoris LMA13 and 924 and S. lactis LMA12. Fast acid-producing, phage-resistant transconjugants were selected by challenge with homologous phage on fast-slow differential agar or lactose indicator agar. Acquisition of pTR2030 by the transconjugants was confirmed by DNA-DNA hybridization. Resistance of transconjugants to homologous phage was complete. Curing or deletion of pTR2030 in the transconjugants confirmed that phage resistance was due to pTR2030 acquisition and not to coincident background mutation. Phage-sensitive pTR2030 deletion derivatives of LMA12 transconjugants were isolated in vivo. The HindIII fragment B of pTR2030 was subcloned into pBR322 to yield a recombinant plasmid, pMET2, useful as a source of pTR2030 DNA. A specific, chemically synthesized oligomer useful as a pTR2030 probe was derived from the sequence of a small portion of pTR2030. The conjugal strategy presented here was effective in yielding fast acid-producing, phage-resistant S. cremoris and S. lactis strains without the use of antibiotic resistance markers and without interfering with the acid-producing ability of the recipient strain. Images PMID:16347196

  9. The Role of DmCatD, a Cathepsin D-Like Peptidase, and Acid Phosphatase in the Process of Follicular Atresia in Dipetalogaster maxima (Hemiptera: Reduviidae), a Vector of Chagas' Disease

    PubMed Central

    Leyria, Jimena; Fruttero, Leonardo L.; Nazar, Magalí; Canavoso, Lilián E.

    2015-01-01

    In this work, we have investigated the involvement of DmCatD, a cathepsin D-like peptidase, and acid phosphatase in the process of follicular atresia of Dipetalogaster maxima, a hematophagous insect vector of Chagas’ disease. For the studies, fat bodies, ovaries and hemolymph were sampled from anautogenous females at representative days of the reproductive cycle: pre-vitellogenesis, vitellogenesis as well as early and late atresia. Real time PCR (qPCR) and western blot assays showed that DmCatD was expressed in fat bodies and ovaries at all reproductive stages, being the expression of its active form significantly higher at the atretic stages. In hemolymph samples, only the immunoreactive band compatible with pro-DmCatD was observed by western blot. Acid phosphatase activity in ovarian tissues significantly increased during follicular atresia in comparison to pre-vitellogenesis and vitellogenesis. A further enzyme characterization with inhibitors showed that the high levels of acid phosphatase activity in atretic ovaries corresponded mainly to a tyrosine phosphatase. Immunofluorescence assays demonstrated that DmCatD and tyrosine phosphatase were associated with yolk bodies in vitellogenic follicles, while in atretic stages they displayed a different cellular distribution. DmCatD and tyrosine phosphatase partially co-localized with vitellin. Moreover, their interaction was supported by FRET analysis. In vitro assays using homogenates of atretic ovaries as the enzyme source and enzyme inhibitors demonstrated that DmCatD, together with a tyrosine phosphatase, were necessary to promote the degradation of vitellin. Taken together, the results strongly suggested that both acid hydrolases play a central role in early vitellin proteolysis during the process of follicular atresia. PMID:26091289

  10. Reduced Vector Preisach Model

    NASA Technical Reports Server (NTRS)

    Patel, Umesh D.; Torre, Edward Della; Day, John H. (Technical Monitor)

    2002-01-01

    A new vector Preisach model, called the Reduced Vector Preisach model (RVPM), was developed for fast computations. This model, derived from the Simplified Vector Preisach model (SVPM), has individual components that like the SVPM are calculated independently using coupled selection rules for the state vector computation. However, the RVPM does not require the rotational correction. Therefore, it provides a practical alternative for computing the magnetic susceptibility using a differential approach. A vector version, using the framework of the DOK model, is implemented. Simulation results for the reduced vector Preisach model are also presented.

  11. Segmentation and texture representation with vector quantizers

    NASA Astrophysics Data System (ADS)

    Yuan, Li; Barba, Joseph

    1990-11-01

    An algorithm for segmentation of cell images and the extraction of texture textons based on vector quantization is presented. Initially a few low dimensional code vectors are employed in a standard vector quantization algorithm to generate a coarse code book a procedure which is equivalent to histogram sharpening. Representative gray level value from each coarse code vector are used to construct a larger fine code book. Coding the original image with the fine code book produces a less distorted image and facilitates cell and nuclear extraction. Texture textons are extracted by application of the same algorithm to the cell area using a larger number of initial code vectors and fine code book. Applications of the algorithm to cytological specimen are presented.

  12. Conserved quantities from piecewise Killing vectors

    NASA Astrophysics Data System (ADS)

    Dray, Tevian; Padmanabhan, T.

    1989-07-01

    In the presence of symmetries, conserved quantities can be obtained by contracting the stress-energy tensor with a Killing vector. We generalize this result to piecewise Killing vectors by giving sufficient conditions for the construction of an associated conserved quantity. A typical example, namely, two stationary space-times joined together in such a way that the resulting space-time is not stationary, is treated in detail.

  13. New silver(I) coordination polymers constructed from pyrazine derivatives and aromatic carboxylic acids: Syntheses, structures and photoluminescence

    NASA Astrophysics Data System (ADS)

    Zhang, Ting; Huang, Hua-Qi; Mei, Hong-Xin; Wang, Dan-Feng; Wang, Xiao-Xiang; Huang, Rong-Bin; Zheng, Lan-Sun

    2015-11-01

    Five one-dimensional to three-dimensional coordination polymers have been synthesized by 2-chlorobenzoic acid (HL1), 2-nitrobenzoic acid (HL2), o-toluic acid (HL3), 2,3,5-trimethylpyrazine (tpyz) and 2,3,5,6-tetramethylpyrazine (mpyz) in the presence of NH3·H2O in mixed solvents systems, namely, {Ag4(tpyz)2(L1)4}n (1), {Ag2(tpyz) (L2)2}n (2), {Ag2(tpyz) (L3)2}n (3), {Ag2(mpyz) (L1)2}n (4), {Ag(mpyz) (L2) (H2O)}n (5). All the complexes have been characterized by elemental analyses, IR spectra and X-ray diffraction. Compound 1 shows a 3D framework. The tpyz ligand links 1D chain which was connected by silver atom and L1 anion into 3D framework. Compounds 2 and 4 possess a similar 2D network with (4, 4) topology. Complex 3 also exhibits a two-dimensional structure. There is a 1D silver chain in 3, which is the main difference from 2 and 4. So, 3 shows three-connected (4 8, 3) topology. For 5, only one oxygen of L2 coordinated to Ag(I) ions. The L2 anions were arranged in both sides of the chain, which was connected by silver atoms and mpyz ligands. Then, the uncoordinated carboxylate oxygen with coordinated water 1molecule oxygen through the hydrogen bond made the resultant structure to a 3D framework. Complexes 1-5 spanning from one-dimensional chains to three-dimensional framework suggest that carboxylates and the kinds of pyrazine derivatives play significant roles in the formation of such coordination architectures. The photoluminescence and thermogravimetric analysis (TGA) of the complexes were also investigated.

  14. A New Hyaluronic Acid Derivative Obtained from Atom Transfer Radical Polymerization as a siRNA Vector for CD44 Receptor Tumor Targeting.

    PubMed

    Palumbo, Fabio Salvatore; Bavuso Volpe, Antonella; Bongiovì, Flavia; Pitarresi, Giovanna; Giammona, Gaetano

    2015-11-01

    Two derivatives of hyaluronic acid (HA) have been synthesized by atom transfer radical polymerization (ATRP), starting from an ethylenediamino HA derivative (HA-EDA) and by using diethylaminoethyl methacrylate (DEAEMA) as a monomer for polymerization. Both samples, indicated as HA-EDA-pDEAEMA a and b, are able to condense siRNA, as determined by gel retardation assay and resulting complexes show a size and a zeta potential value dependent on polymerization number, as determined by dynamic light scattering measurements. In vitro studies performed on HCT 116 cell line, that over express CD44 receptor, demonstrate a receptor mediated uptake of complexes, regardless of their surface charge. PMID:26136372

  15. A pH-responsive drug nanovehicle constructed by reversible attachment of cholesterol to PEGylated poly(l-lysine) via catechol-boronic acid ester formation.

    PubMed

    Yang, Bin; Lv, Yin; Zhu, Jing-Yi; Han, Yun-Tao; Jia, Hui-Zhen; Chen, Wei-Hai; Feng, Jun; Zhang, Xian-Zheng; Zhuo, Ren-Xi

    2014-08-01

    The present work reports the construction of a drug delivery nanovehicle via a pH-sensitive assembly strategy for improved cellular internalization and intracellular drug liberation. Through spontaneous formation of boronate linkage in physiological conditions, phenylboronic acid-modified cholesterol was able to attach onto catechol-pending methoxypoly(ethylene glycol)-block-poly(l-lysine). This comb-type polymer can self-organize into a micellar nanoconstruction that is able to effectively encapsulate poorly water-soluble agents. The blank micelles exhibited negligible in vitro cytotoxicity, yet doxorubicin (DOX)-loaded micelles could effectively induce cell death at a level comparable to free DOX. Owing to the acid-labile feature of the boronate linkage, a reduction in environmental pH from pH 7.4 to 5.0 could trigger the dissociation of the nanoconstruction, which in turn could accelerate the liberation of entrapped drugs. Importantly, the blockage of endosomal acidification in HeLa cells by NH4Cl treatment significantly decreased the nuclear uptake efficiency and cell-killing effect mediated by the DOX-loaded nanoassembly, suggesting that acid-triggered destruction of the nanoconstruction is of significant importance in enhanced drug efficacy. Moreover, confocal fluorescence microscopy and flow cytometry assay revealed the effective internalization of the nanoassemblies, and their cellular uptake exhibited a cholesterol dose-dependent profile, indicating the contribution of introduced cholesterol functionality to the transmembrane process of the nanoassembly. PMID:24879311

  16. Understanding Singular Vectors

    ERIC Educational Resources Information Center

    James, David; Botteron, Cynthia

    2013-01-01

    matrix yields a surprisingly simple, heuristical approximation to its singular vectors. There are correspondingly good approximations to the singular values. Such rules of thumb provide an intuitive interpretation of the singular vectors that helps explain why the SVD is so…

  17. The vector ruling protractor

    NASA Technical Reports Server (NTRS)

    Zahm, A F

    1924-01-01

    The theory, structure and working of a vector slide rule is presented in this report. This instrument is used for determining a vector in magnitude and position when given its components and its moment about a point in their plane.

  18. Validity of manganese as a surrogate of heavy metals removal in constructed wetlands treating acidic mine water

    SciTech Connect

    Royer, E.; Unz, R.F.; Hellier, W.W.

    1998-12-31

    The evaluation of manganese as a surrogate for heavy metal behavior in two wetland treatment systems receiving acidic coal mine drainage in central Pennsylvania was investigated. The use of manganese as an indicator is based on physical/chemical treatment processes quite different from wetland treatment. The treatment systems represented one anoxic, subsurface flow system and one oxic surface flow system. Water quality parameters measured included pH, alkalinity, acidity, and a suite of metals. Correlation and linear regression analysis were used to evaluate the ability of a candidate predictor variable (indicator) to predict heavy metal concentrations and removal. The use of manganese as a predictor of effluent quality proved to be poor in both wetland treatment systems, as evidenced by low linear R{sup 2} values and negative correlations. Zinc emerged as the best predictor of the detectable heavy metals at the anoxic wetland. Zinc exhibited positive strong linear correlations with copper, cobalt, and nickel (R{sup 2} values of 0.843, 0.881, and 0.970, respectively). Effluent pH was a slightly better predictor of effluent copper levels in the anoxic wetland. Iron and cobalt effluent concentrations showed the only strong relationship (R{sup 2} value = 0.778) in the oxic system. The lack of good correlations with manganese strongly challenges its appropriateness as a surrogate for heavy metals in these systems.

  19. Crosslinked self-assemblies of lipoid acid-substituted low molecular weight (1800 Da) polyethylenimine as reductive-sensitive non-viral gene vectors

    NASA Astrophysics Data System (ADS)

    Chen, Xiaojiao; Yuan, Zhefan; Yi, Xiaoqing; Zhuo, Renxi; Li, Feng

    2012-10-01

    In this study, amphiphilic polyethylenimine-graft-thioctic acid (PEI-TA) and polyethylenimine-graft-lauric acid (PEI-LA) were synthesized. Both PEI-TA and PEI-LA could self-assemble into micelles. Due to the existence of disulfide-linked rings at the end of hydrophobic moieties, PEI-TA could form stable micelles with disulfide crosslinked cores (PEI-TA-SS). In comparison with the PEI-LA micelle, PEI-TA-SS possessed higher DNA binding ability according to the gel retardation assay and heparin replacement assay. In vitro transfection experiments indicated that PEI-TA-SS showed comparably high transfection efficiency as compared to 25 kDa PEI. More interestingly, the luciferase expression of PEI-TA-SS was superior to that of PEI-LA at low N/P ratio, which might be ascribed to the stronger binding capacity of PEI-TA-SS facilitating the entering of PEI-TA-SS/pDNA complexes into cells.

  20. Construction and Characterization of New piggyBac Vectors for Constitutive or Inducible Expression of Heterologous Gene Pairs and the Identification of a Previously Unrecognized Activator Sequence in piggyBac

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The piggyBac transposable element was first identified when it inserted itself into baculovirus genes during infection, causing a mutant plaque morphology phenotype. Since then, piggyBac has been developed into a germline tranformation vector usable with a wide variety of organisms. In this study...

  1. Biosafety Features of Lentiviral Vectors

    PubMed Central

    Schambach, Axel; Zychlinski, Daniela; Ehrnstroem, Birgitta

    2013-01-01

    Abstract Over the past decades, lentiviral vectors have evolved as a benchmark tool for stable gene transfer into cells with a high replicative potential. Their relatively flexible genome and ability to transduce many forms of nondividing cells, combined with the potential for cell-specific pseudotyping, provides a rich resource for numerous applications in experimental platforms and therapeutic settings. Here, we give an overview of important biosafety features of lentiviral vectors, with detailed discussion of (i) the principles of the lentiviral split-genome design used for the construction of packaging cells; (ii) the relevance of modifications introduced into the lentiviral long terminal repeat (deletion of enhancer/promoter sequences and introduction of insulators); (iii) the basic features of mRNA processing, including the Rev/Rev-responsive element (RRE) interaction and the modifications of the 3′ untranslated region of lentiviral vectors with various post-transcriptional regulatory elements affecting transcriptional termination, polyadenylation, and differentiation-specific degradation of mRNA; and (iv) the characteristic integration pattern with the associated risk of transcriptional interference with cellular genes. We conclude with considerations regarding the importance of cell targeting via envelope modifications. Along this course, we address canonical biosafety issues encountered with any type of viral vector: the risks of shedding, mobilization, germline transmission, immunogenicity, and insertional mutagenesis. PMID:23311447

  2. Development of a new vector using Soybean yellow common mosaic virus for gene function study or heterologous protein expression in soybeans.

    PubMed

    Lim, Seungmo; Nam, Moon; Kim, Kil Hyun; Lee, Su-Heon; Moon, Jung-Kyung; Lim, Hyoun-Sub; Choung, Myoung-Gun; Kim, Sang-Mok; Moon, Jae Sun

    2016-02-01

    A new vector using Soybean yellow common mosaic virus (SYCMV) was constructed for gene function study or heterologous protein expression in soybeans. The in vitro transcript with a 5' cap analog m7GpppG from an SYCMV full-length infectious vector driven by a T7 promoter infected soybeans (pSYCMVT7-full). The symptoms observed in the soybeans infected with either the sap from SYCMV-infected leaves or pSYCMVT7-full were indistinguishable, suggesting that the vector exhibits equivalent biological activity as the virus itself. To utilize the vector further, a DNA-based vector driven by the Cauliflower mosaic virus (CaMV) 35S promoter was constructed. The complete sequence of the SYCMV genome was inserted into a binary vector flanked by a CaMV 35S promoter at the 5' terminus of the SYCMV genome and a cis-cleaving ribozyme sequence followed by a nopaline synthase terminator at the 3' terminus of the SYCMV genome (pSYCMV-full). The SYCMV-derived vector was tested for use as a virus-induced gene silencing (VIGS) vector for the functional analysis of soybean genes. VIGS constructs containing either a fragment of the Phytoene desaturase (PDS) gene (pSYCMV-PDS1) or a fragment of the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RbcS) gene (pSYCMV-RbcS2) were constructed. Plants infiltrated with each vector using the Agrobacterium-mediated inoculation method exhibited distinct symptoms, such as photo-bleaching in plants infiltrated with pSYCMV-PDS1 and yellow or pale green coloring in plants infiltrated with pSYCMV-RbcS2. In addition, down-regulation of the transcripts of the two target genes was confirmed via northern blot analysis. Particle bombardment and direct plasmid DNA rubbing were also confirmed as alternative inoculation methods. To determine if the SYCMV vector can be used for the expression of heterologous proteins in soybean plants, the vector encoding amino acids 135-160 of VP1 of Foot-and-mouth disease virus (FMDV) serotype O1 Campos (O1C

  3. Coexpression of Tyrosine Hydroxylase, GTP Cyclohydrolase I, Aromatic Amino Acid Decarboxylase, and Vesicular Monoamine Transporter 2 from a Helper Virus-Free Herpes Simplex Virus Type 1 Vector Supports High-Level, Long-Term Biochemical and Behavioral Correction of a Rat Model of Parkinson’s Disease

    PubMed Central

    SUN, MEI; KONG, LINGXIN; WANG, XIAODAN; HOLMES, COURTNEY; GAO, QINGSHENG; ZHANG, GUO-RONG; PFEILSCHIFTER, JOSEF; GOLDSTEIN, DAVID S.; GELLER, ALFRED I.

    2006-01-01

    Parkinson’s disease is due to the selective loss of nigrostriatal dopaminergic neurons. Consequently, many therapeutic strategies have focused on restoring striatal dopamine levels, including direct gene transfer to striatal cells, using viral vectors that express specific dopamine biosynthetic enzymes. The central hypothesis of this study is that coexpression of four dopamine biosynthetic and transporter genes in striatal neurons can support the efficient production and regulated, vesicular release of dopamine: tyrosine hydroxylase (TH) converts tyrosine to l-3,4-dihydroxyphenylalanine (l -DOPA), GTP cyclohydrolase I (GTP CH I) is the rate-limiting enzyme in the biosynthesis of the cofactor for TH, aromatic amino acid decarboxylase (AADC) converts l -DOPA to dopamine, and a vesicular monoamine transporter (VMAT-2) transports dopamine into synaptic vesicles, thereby supporting regulated, vesicular release of dopamine and relieving feedback inhibition of TH by dopamine. Helper virus-free herpes simplex virus type 1 vectors that coexpress the three dopamine biosynthetic enzymes (TH, GTP CH I, and AADC; 3-gene-vector) or these three dopamine biosynthetic enzymes and the vesicular monoamine transporter (TH, GTP CH I, AADC, and VMAT-2; 4-gene-vector) were compared. Both vectors supported production of dopamine in cultured fibroblasts. These vectors were microinjected into the striatum of 6-hydroxydopamine-lesioned rats. These vectors carry a modified neurofilament gene promoter, and γ-aminobutyric acid (GABA)-ergic neuron-specific gene expression was maintained for 14 months after gene transfer. The 4-gene-vector supported higher levels of correction of apomorphine-induced rotational behavior than did the 3-gene-vector, and this correction was maintained for 6 months. Proximal to the injection sites, the 4-gene-vector, but not the 3-gene-vector, supported extracellular levels of dopamine and dihydroxyphenylacetic acid (DOPAC) that were similar to those observed in

  4. Restart 68000 vector remapping

    SciTech Connect

    Gustin, J.

    1984-05-03

    The circuit described allows power-on-reset (POR) vector fetch from ROM for a 68000 microprocessor. It offers programmability of exception vectors, including the restart vector. This method eliminates the need for high-resolution, address-decoder peripheral circuitry.

  5. Rhotrix Vector Spaces

    ERIC Educational Resources Information Center

    Aminu, Abdulhadi

    2010-01-01

    By rhotrix we understand an object that lies in some way between (n x n)-dimensional matrices and (2n - 1) x (2n - 1)-dimensional matrices. Representation of vectors in rhotrices is different from the representation of vectors in matrices. A number of vector spaces in matrices and their properties are known. On the other hand, little seems to be…

  6. Insulated Foamy Viral Vectors.

    PubMed

    Browning, Diana L; Collins, Casey P; Hocum, Jonah D; Leap, David J; Rae, Dustin T; Trobridge, Grant D

    2016-03-01

    Retroviral vector-mediated gene therapy is promising, but genotoxicity has limited its use in the clinic. Genotoxicity is highly dependent on the retroviral vector used, and foamy viral (FV) vectors appear relatively safe. However, internal promoters may still potentially activate nearby genes. We developed insulated FV vectors, using four previously described insulators: a version of the well-studied chicken hypersensitivity site 4 insulator (650cHS4), two synthetic CCCTC-binding factor (CTCF)-based insulators, and an insulator based on the CCAAT box-binding transcription factor/nuclear factor I (7xCTF/NF1). We directly compared these insulators for enhancer-blocking activity, effect on FV vector titer, and fidelity of transfer to both proviral long terminal repeats. The synthetic CTCF-based insulators had the strongest insulating activity, but reduced titers significantly. The 7xCTF/NF1 insulator did not reduce titers but had weak insulating activity. The 650cHS4-insulated FV vector was identified as the overall most promising vector. Uninsulated and 650cHS4-insulated FV vectors were both significantly less genotoxic than gammaretroviral vectors. Integration sites were evaluated in cord blood CD34(+) cells and the 650cHS4-insulated FV vector had fewer hotspots compared with an uninsulated FV vector. These data suggest that insulated FV vectors are promising for hematopoietic stem cell gene therapy. PMID:26715244

  7. Development of non-defective recombinant densovirus vectors for microRNA delivery in the invasive vector mosquito, Aedes albopictus

    PubMed Central

    Liu, Peiwen; Li, Xiaocong; Gu, Jinbao; Dong, Yunqiao; Liu, Yan; Santhosh, Puthiyakunnon; Chen, Xiaoguang

    2016-01-01

    We previously reported that mosquito densoviruses (MDVs) are potential vectors for delivering foreign nucleic acids into mosquito cells. However, considering existing expression strategies, recombinant viruses would inevitably become replication-defective viruses and lose their ability for secondary transmission. The packaging limitations of the virion represent a barrier for the development of MDVs for viral paratransgenesis or as high-efficiency bioinsecticides. Herein, we report the development of a non-defective recombinant Aedes aegypti densovirus (AaeDV) miRNA expression system, mediated by an artificial intron, using an intronic miRNA expression strategy. We demonstrated that this recombinant vector could be used to overexpress endogenous miRNAs or to decrease endogenous miRNAs by generating antisense sponges to explore the biological functions of miRNAs. In addition, the vector could express antisense-miRNAs to induce efficient gene silencing in vivo and in vitro. The recombinant virus effectively self-replicated and retained its secondary transmission ability, similar to the wild-type virus. The recombinant virus was also genetically stable. This study demonstrated the first construction of a non-defective recombinant MDV miRNA expression system, which represents a tool for the functional analysis of mosquito genes and lays the foundation for the application of viral paratransgenesis for dengue virus control. PMID:26879823

  8. Construction of antibacterial poly(ethylene terephthalate) films via layer by layer assembly of chitosan and hyaluronic acid.

    PubMed

    Del Hoyo-Gallego, Sara; Pérez-Álvarez, Leyre; Gómez-Galván, Flor; Lizundia, Erlantz; Kuritka, Ivo; Sedlarik, Vladimir; Laza, Jose Manuel; Vila-Vilela, Jose Luis

    2016-06-01

    Polyelectrolytic multilayers (PEMs) with enhanced antibacterial properties were built up onto commercial poly(ethylene terephthalate) (PET) films based on the layer by layer assembling of bacterial contact killing chitosan and bacterial repelling highly hydrated hyaluronic acid. The optimization of the aminolysis modification reaction of PET was carried out by the study of the mechanical properties and the surface characterization of the modified polymers. The layer by layer assembly was successfully monitored by TEM microscopy, surface zeta-potential, contact angle measurements and, after labeling with fluorescein isothiocyanate (FTIC) by absorption spectroscopy and confocal fluorescent microscopy. Beside, the stability of the PEMs was studied at physiological conditions in absence and in the presence of lysozyme and hyaluronidase enzymes. Antibacterial properties of the obtained PEMs against Escherichia coli were compared with original commercial PET. PMID:27083341

  9. Gene targeting with retroviral vectors

    SciTech Connect

    Ellis, J.; Bernstein, A. )

    1989-04-01

    The authors have designed and constructed integration-defective retroviral vectors to explore their potential for gene targeting in mammalian cells. Two nonoverlapping deletion mutants of the bacterial neomycin resistance (neo) gene were used to detect homologous recombination events between viral and chromosomal sequences. Stable neo gene correction events were selected at a frequency of approximately 1 G418/sup r/ cell per 3 x 10/sup 6/ infected cells. Analysis of the functional neo gene in independent targeted cell clones indicated that unintegrated retroviral linear DNA recombined with the target by gene conversion for variable distances into regions of nonhomology. In addition, transient neo gene correction events which were associated with the complete loss of the chromosomal target sequences were observed. These results demonstrated that retroviral vectors can recombine with homologous chromosomal sequences in rodent and human cells.

  10. Covariantized vector Galileons

    NASA Astrophysics Data System (ADS)

    Hull, Matthew; Koyama, Kazuya; Tasinato, Gianmassimo

    2016-03-01

    Vector Galileons are ghost-free systems containing higher derivative interactions of vector fields. They break the vector gauge symmetry, and the dynamics of the longitudinal vector polarizations acquire a Galileon symmetry in an appropriate decoupling limit in Minkowski space. Using an Arnowitt-Deser-Misner approach, we carefully reconsider the coupling with gravity of vector Galileons, with the aim of studying the necessary conditions to avoid the propagation of ghosts. We develop arguments that put on a more solid footing the results previously obtained in the literature. Moreover, working in analogy with the scalar counterpart, we find indications for the existence of a "beyond Horndeski" theory involving vector degrees of freedom that avoids the propagation of ghosts thanks to secondary constraints. In addition, we analyze a Higgs mechanism for generating vector Galileons through spontaneous symmetry breaking, and we present its consistent covariantization.

  11. Immobilized Lentivirus Vector on Chondroitin Sulfate-Hyaluronate Acid-Silk Fibroin Hybrid Scaffold for Tissue-Engineered Ligament-Bone Junction

    PubMed Central

    Sun, Liguo; Li, Hongguo; Qu, Ling; Zhu, Rui; Fan, Xiangli; Xue, Yingsen; Xie, Zhenghong; Fan, Hongbin

    2014-01-01

    The lack of a fibrocartilage layer between graft and bone remains the leading cause of graft failure after anterior cruciate ligament (ACL) reconstruction. The objective of this study was to develop a gene-modified silk cable-reinforced chondroitin sulfate-hyaluronate acid-silk fibroin (CHS) hybrid scaffold for reconstructing the fibrocartilage layer. The scaffold was fabricated by lyophilizing the CHS mixture with braided silk cables. The scanning electronic microscopy (SEM) showed that microporous CHS sponges were formed around silk cables. Each end of scaffold was modified with lentiviral-mediated transforming growth factor-β3 (TGF-β3) gene. The cells on scaffold were transfected by bonded lentivirus. In vitro culture demonstrated that mesenchymal stem cells (MSCs) on scaffolds proliferated vigorously and produced abundant collagen. The transcription levels of cartilage-specific genes also increased with culture time. After 2 weeks, the MSCs were distributed uniformly throughout scaffold. Deposited collagen was also found to increase. The chondral differentiation of MSCs was verified by expressions of collagen II and TGF-β3 genes in mRNA and protein level. Histology also confirmed the production of cartilage extracellular matrix (ECM) components. The results demonstrated that gene-modified silk cable-reinforced CHS scaffold was capable of supporting cell proliferation and differentiation to reconstruct the cartilage layer of interface. PMID:25019087

  12. DNA shuttling between plasmid vectors and a genome vector: systematic conversion and preservation of DNA libraries using the Bacillus subtilis genome (BGM) vector.

    PubMed

    Kaneko, Shinya; Akioka, Manami; Tsuge, Kenji; Itaya, Mitsuhiro

    2005-06-24

    The combined use of the contemporary vector systems, the bacterial artificial chromosome (BAC) vector and the Bacillus subtilis genome (BGM) vector, makes possible the handling of giant-length DNA (above 100 kb). Our newly constructed BGM vector efficiently integrated DNA prepared in the BAC vector. A BAC library comprised of 18 independent clones prepared from mitochondrial DNA (mtDNA) of Arabidopsis thaliana was converted to a parallel BGM library using the new BGM vector. The effectiveness of the combined use of the vector systems was confirmed by the stable recovery of all 18 DNAs as BAC clones from the respective BGM clones. We show that DNA in BGM was stably preserved at room temperature after spore formation of the host B.subtilis. Rapid and stable shuttling between Escherichiacoli and the B. subtilis host, combined with spore-mediated DNA storage, may facilitate the long-term and low-cost preservation and the transportation of DNA resources. PMID:15913652

  13. Structural variations and photoluminescent properties of a series of metal-organic frameworks constructed from 5-(4-carboxybenzoylamino)-isophthalic acid

    SciTech Connect

    Zhao, Wen; Zhang, Li-Juan; Zhao, Xiao-Li

    2013-06-01

    Five new metal-organic frameworks (MOFs) with 5-(4-carboxybenzoylamino)-isophthalic acid (H₃L), namely, [Cd₉L₆(DMA)₆]·4DMA (1), [Cd₃L₂(H₂O)₉]·4H₂O (2), [LaL(H₂O)₄]·2H₂O (3), [CeL(H₂O)₄]·H₂O (4) and [Tb(HL)(H₂L)(H₂O)₃]·5H₂O (5) (DMA=N,N-dimethylacetamide), have been synthesized. Complex 1 shows a three-dimensional architecture generated from linkage of Cd–O chains via L³⁻ ligands. Minor variations in synthetic conditions of 1 afforded 2, which features an interesting 2D→3D catenation architecture containing helical chains. Complexes 3 and 4 are isostructural and each feature a two-dimensional architecture constructed from the linkage of L³⁻ with Ln³⁺. Complex 5 displays a chain-like structure, of which the most interesting feature is the existence of free carboxylic acid (–COOH) group which may confer unique functionality. Moreover, the investigations of the thermal stability, powder X-ray diffractions and solid-state photoluminescent properties for these crystalline materials have been carried out. - Graphical Abstract: Solvothermal reactions of tricarboxylate ligand H₃L with Cd²⁺/Ln³⁺ has yielded a series of new MOFs containing interesting structural motifs. Highlights: • A tricarboxylate ligand whose coordinating functionalities are not symmetry equivalent is employed to construct MOFs. • Complex 2 features an interesting 2D→3D catenation architecture containing helical chains. • Complex 3 feature chain-like structure containing free – COOH group, which may confer unique functionality. • Photoluminescent properties and thermal behaviors for 1–5 have been reported.

  14. Solar imaging vector magnetograph

    NASA Technical Reports Server (NTRS)

    Canfield, Richard C.

    1993-01-01

    This report describes an instrument which has been constructed at the University of Hawaii to make observations of the magnetic field in solar active regions. Detailed knowledge of active region magnetic structures is crucial to understanding many solar phenomena, because the magnetic field both defines the morphology of structures seen in the solar atmosphere and is the apparent energy source for solar flares. The new vector magnetograph was conceived in response to a perceived discrepancy between the capabilities of X ray imaging telescopes to be operating during the current solar maximum and those of existing magnetographs. There were no space-based magnetographs planned for this period; the existing ground-based instruments variously suffered from lack of sensitivity, poor time resolution, inadequate spatial resolution or unreliable sites. Yet the studies of flares and their relationship to the solar corona planned for the 1991-1994 maximum absolutely required high quality vector magnetic field measurements. By 'vector' measurements we mean that the observation attempts to deduce the complete strength and direction of the field at the measurement site, rather than just the line of sight component as obtained by a traditional longitudinal magnetograph. Knowledge of the vector field permits one to calculate photospheric electric currents, which might play a part in heating the corona, and to calculate energy stored in coronal magnetic fields as the result of such currents. Information about the strength and direction of magnetic fields in the solar atmosphere can be obtained in a number of ways, but quantitative data is best obtained by observing Zeeman-effect polarization in solar spectral lines. The technique requires measuring the complete state of polarization at one or more wavelengths within a magnetically sensitive line of the solar spectrum. This measurement must be done for each independent spatial point for which one wants magnetic field data. All the

  15. The occurrence of domoic acid linked to a toxic diatom bloom in a new potential vector: the tunicate Pyura chilensis (piure).

    PubMed

    López-Rivera, Américo; Pinto, Maricela; Insinilla, Andrea; Suárez Isla, Benjamín; Uribe, Eduardo; Alvarez, Gonzalo; Lehane, Mary; Furey, Ambrose; James, Kevin J

    2009-11-01

    The tunicate Pyura chilensis (Molina, 1782); Phylum Chordata; Subphylum Urochordata; Class Ascidiacea, common local name "piure" or sea squirt; a filter-feeder (plankton and suspended particles) sessile species; may play an important role in monitoring domoic acid (DA) the principal toxic component of Amnesic Shellfish Poisoning (ASP). Significant DA concentrations have been determined in tunicate samples, collected during a recent ASP outbreak in Bahía Inglesa, an important scallop (Argopecten purpuratus) farming area. Several infaunal species were tested for the presence of DA, in addition to the usual scallop monitoring programme. DA was found at sub-toxic levels in filtering bivalves such as mussels (Mytilus chilensis), large mussels (Aulacomya ater) and clams (Protothaca thaca) (6.4, 5.4 and 4.7 microg DA/g tissue respectively). Of particular interest was the observation of significant accumulations of toxic Pseudo-nitzschia sp. diatoms in the internal siphon and atrium spaces of the tunicate. Toxin distribution within major tunicate organs was heterogeneous with 8.7-15.5 microg DA/g in edible tissues, 14.9-17.9 microg DA/g in the fecal material and 13.6-32.7 microg DA/g in the gut content. DA was determined by HPLC-UV and confirmed by diode-array detection and LC-MS/MS analysis. This is the first report of the presence of DA in a tunicate that is regularly consumed by coastal populations. These results confirm the need to include these organisms in sanitation programs for marine toxins. PMID:19505493

  16. Neural network consistent empirical physical formula construction for density functional theory based nonlinear vibrational absorbance and intensity of 6-choloronicotinic acid molecule

    NASA Astrophysics Data System (ADS)

    Yildiz, Nihat; Karabacak, Mehmet; Kurt, Mustafa; Akkoyun, Serkan

    2012-05-01

    Being directly related to the electric charge distributions in a molecule, the vibrational spectra intensities are both experimentally and theoretically important physical quantities. However, these intensities are inherently highly nonlinear and of complex pattern. Therefore, in particular for unknown detailed spatial molecular structures, it is difficult to make ab initio intensity calculations to compare with new experimental data. In this respect, we very recently initiated entirely novel layered feedforward neural network (LFNN) approach to construct empirical physical formulas (EPFs) for density functional theory (DFT) vibrational spectra of some molecules. In this paper, as a new and far improved contribution to our novel molecular vibrational spectra LFNN-EPF approach, we constructed LFFN-EPFs for absorbances and intensities of 6-choloronicotinic acid (6-CNA) molecule. The 6-CNA data, borrowed from our previous study, was entirely different and much larger than the vibrational intensity data of our formerly used LFNN-EPF molecules. In line with our another previous work which theoretically proved the LFNN relevance to EPFs, although the 6-CNA DFT absorbance and intensity were inherently highly nonlinear and sharply fluctuating in character, still the optimally constructed train set LFFN-EPFs very successfully fitted the absorbances and intensities. Moreover, test set (i.e. yet-to-be measured experimental data) LFNN-EPFs consistently and successfully predicted the absorbance and intensity data. This simply means that the physical law embedded in the 6-CNA vibrational data was successfully extracted by the LFNN-EPFs. In conclusion, these vibrational LFNN-EPFs are of explicit form. Therefore, by various suitable operations of mathematical analysis, they can be used to estimate the electronic charge distributions of the unknown molecule of the significant complexity. Additionally, these estimations can be combined with those of theoretical DFT atomic polar

  17. Index Sets and Vectorization

    SciTech Connect

    Keasler, J A

    2012-03-27

    Vectorization is data parallelism (SIMD, SIMT, etc.) - extension of ISA enabling the same instruction to be performed on multiple data items simultaeously. Many/most CPUs support vectorization in some form. Vectorization is difficult to enable, but can yield large efficiency gains. Extra programmer effort is required because: (1) not all algorithms can be vectorized (regular algorithm structure and fine-grain parallelism must be used); (2) most CPUs have data alignment restrictions for load/store operations (obey or risk incorrect code); (3) special directives are often needed to enable vectorization; and (4) vector instructions are architecture-specific. Vectorization is the best way to optimize for power and performance due to reduced clock cycles. When data is organized properly, a vector load instruction (i.e. movaps) can replace 'normal' load instructions (i.e. movsd). Vector operations can potentially have a smaller footprint in the instruction cache when fewer instructions need to be executed. Hybrid index sets insulate users from architecture specific details. We have applied hybrid index sets to achieve optimal vectorization. We can extend this concept to handle other programming models.

  18. Heterotic String Compactification and New Vector Bundles

    NASA Astrophysics Data System (ADS)

    Lin, Hai; Wu, Baosen; Yau, Shing-Tung

    2016-07-01

    We propose a construction of Kähler and non-Kähler Calabi-Yau manifolds by branched double covers of twistor spaces. In this construction we use the twistor spaces of four-manifolds with self-dual conformal structures, with the examples of connected sum of n {mathbb{P}2}s. We also construct K3-fibered Calabi-Yau manifolds from the branched double covers of the blow-ups of the twistor spaces. These manifolds can be used in heterotic string compactifications to four dimensions. We also construct stable and polystable vector bundles. Some classes of these vector bundles can give rise to supersymmetric grand unified models with three generations of quarks and leptons in four dimensions.

  19. Heterotic String Compactification and New Vector Bundles

    NASA Astrophysics Data System (ADS)

    Lin, Hai; Wu, Baosen; Yau, Shing-Tung

    2016-06-01

    We propose a construction of Kähler and non-Kähler Calabi-Yau manifolds by branched double covers of twistor spaces. In this construction we use the twistor spaces of four-manifolds with self-dual conformal structures, with the examples of connected sum of n {P2} s. We also construct K3-fibered Calabi-Yau manifolds from the branched double covers of the blow-ups of the twistor spaces. These manifolds can be used in heterotic string compactifications to four dimensions. We also construct stable and polystable vector bundles. Some classes of these vector bundles can give rise to supersymmetric grand unified models with three generations of quarks and leptons in four dimensions.

  20. Construction of a novel expression system for use in Corynebacterium glutamicum.

    PubMed

    Hu, Jinyu; Li, Yanyan; Zhang, Hailing; Tan, Yanzhen; Wang, Xiaoyuan

    2014-09-01

    Corynebacterium glutamicum is an important microorganism for production of amino acids in industrial fermentation. Suitable vectors are needed for metabolic engineering in C. glutamicum. Most available vectors used in C. glutamicum carry antibiotic resistant genes as a genetic labeling for rapid identification of recombinant strains, and antibiotics have to be added to maintain the vector when growing the cells. These vectors, though excellent for laboratory use, are not preferable choices for industry-scale fermentation. In this work, we developed a novel expression system for use in C. glutamicum, which do not require antibiotics when used for industrial fermentation. This system includes two vectors: the shuttle vector pJYW-4 for expression of genes and the vector pJYW-6 for deletion of the essential gene alr in C. glutamicum. The vector pJYW-4 contains a large multiple cloning site for cloning multiple genes and two selective markers: one is the kanamycin-resistant gene kan and the other is an essential gene alr. The selective marker kan facilitates molecular manipulation or fermentations in the laboratory, and the selection marker alr is good for use in industry-scale fermentation, allowing in vivo maintenance of the expression vector through auxotrophic complementation; therefore, the two selection markers in pJYW-4 make it useful for both laboratory research and industrial fermentation, and convenient to transfer valuable laboratory-developed strains into industrial production. This newly-constructed expression system was successfully used to increase L-valine production in C. glutamicum ATCC 14067, indicating its potential on developing amino acid-producing C. glutamicum strains. PMID:25108235

  1. TWSVR: Regression via Twin Support Vector Machine.

    PubMed

    Khemchandani, Reshma; Goyal, Keshav; Chandra, Suresh

    2016-02-01

    Taking motivation from Twin Support Vector Machine (TWSVM) formulation, Peng (2010) attempted to propose Twin Support Vector Regression (TSVR) where the regressor is obtained via solving a pair of quadratic programming problems (QPPs). In this paper we argue that TSVR formulation is not in the true spirit of TWSVM. Further, taking motivation from Bi and Bennett (2003), we propose an alternative approach to find a formulation for Twin Support Vector Regression (TWSVR) which is in the true spirit of TWSVM. We show that our proposed TWSVR can be derived from TWSVM for an appropriately constructed classification problem. To check the efficacy of our proposed TWSVR we compare its performance with TSVR and classical Support Vector Regression(SVR) on various regression datasets. PMID:26624223

  2. Symbolic computer vector analysis

    NASA Technical Reports Server (NTRS)

    Stoutemyer, D. R.

    1977-01-01

    A MACSYMA program is described which performs symbolic vector algebra and vector calculus. The program can combine and simplify symbolic expressions including dot products and cross products, together with the gradient, divergence, curl, and Laplacian operators. The distribution of these operators over sums or products is under user control, as are various other expansions, including expansion into components in any specific orthogonal coordinate system. There is also a capability for deriving the scalar or vector potential of a vector field. Examples include derivation of the partial differential equations describing fluid flow and magnetohydrodynamics, for 12 different classic orthogonal curvilinear coordinate systems.

  3. Vector Product and an Integrable Dynamical System

    NASA Astrophysics Data System (ADS)

    Willi-Hans, Steeb; Yorick, Hardy; Igor, Tanski

    2011-12-01

    We study an autonomous system of first order ordinary differential equations based on the vector product. We show that the system is completely integrable by constructing the first integrals. The connection with Nambu mechanics is established. The extension to higher dimensions is also discussed.

  4. Short communication. Characterization of chloroplast region rrn16-rrn23S from the tropical timber tree Cedrela odorata L. and de novo construction of a transplastomic expression vector suitable for Meliaceae trees and other economically important crops.

    PubMed

    López-Ochoa, L A; Apolinar-Hernández, M M; Peña-Ramírez, Y J

    2015-01-01

    The forest tree Spanish cedar (Cedrela odorata L.) is well-known for its high-value timber; however, this species is attacked by the shoot borer (Hypsipyla grandella) during its early years of development, resulting in branched stems and making the plants useless for high-quality wood production. The generation of resistant varieties expressing entomotoxic proteins may be an alternative to pesticide treatments. The use of plastid transformation rather than nuclear transformation should be used because it reduces the risk of transgene dissemination by pollen. Chloroplast transformation vectors require an expression cassette flanked by homologous plastid sequences to drive plastome recombination. Thus, C. odorata plastome sequences are a prerequisite. The rrn16-rrn23 plastome region was selected, cloned, and characterized. When the sequence identity among the rrn16-rrn23 regions from C. odorata and Nicotiana tabacum was compared, 3 inDels of 240, 104, and 39 bp were found that might severely affect transformation efficiency. Using this region, a new transformation vector was developed using pUC19 as a backbone by inserting the rrn16-trnI and trnA-rrn23 sequences from C. odorata and adding 2 independent expression cassettes into the trnI-trnA intergenic region, conferring spectinomycin resistance, the ability to express the gfp reporter gene, and a site that can be used to express any other gene of interest. PMID:25730086

  5. Vectors in Use in a 3D Juggling Game Simulation

    ERIC Educational Resources Information Center

    Kynigos, Chronis; Latsi, Maria

    2006-01-01

    The new representations enabled by the educational computer game the "Juggler" can place vectors in a central role both for controlling and measuring the behaviours of objects in a virtual environment simulating motion in three-dimensional spaces. The mathematical meanings constructed by 13 year-old students in relation to vectors as objects, as a…

  6. Mineralized three-dimensional bone constructs

    NASA Technical Reports Server (NTRS)

    Clarke, Mark S. F. (Inventor); Sundaresan, Alamelu (Inventor); Pellis, Neal R. (Inventor)

    2011-01-01

    The present disclosure provides ex vivo-derived mineralized three-dimensional bone constructs. The bone constructs are obtained by culturing osteoblasts and osteoclast precursors under randomized gravity vector conditions. Preferably, the randomized gravity vector conditions are obtained using a low shear stress rotating bioreactor, such as a High Aspect Ratio Vessel (HARV) culture system. The bone constructs of the disclosure have utility in physiological studies of bone formation and bone function, in drug discovery, and in orthopedics.

  7. Mineralized Three-Dimensional Bone Constructs

    NASA Technical Reports Server (NTRS)

    Clarke, Mark S. F. (Inventor); Sundaresan, Alamelu (Inventor); Pellis, Neal R. (Inventor)

    2013-01-01

    The present disclosure provides ex vivo-derived mineralized three-dimensional bone constructs. The bone constructs are obtained by culturing osteoblasts and osteoclast precursors under randomized gravity vector conditions. Preferably, the randomized gravity vector conditions are obtained using a low shear stress rotating bioreactor, such as a High Aspect Ratio Vessel (HARV) culture system. The bone constructs of the disclosure have utility in physiological studies of bone formation and bone function, in drug discovery, and in orthopedics.

  8. Vector theories in cosmology

    SciTech Connect

    Esposito-Farese, Gilles; Pitrou, Cyril; Uzan, Jean-Philippe

    2010-03-15

    This article provides a general study of the Hamiltonian stability and the hyperbolicity of vector field models involving both a general function of the Faraday tensor and its dual, f(F{sup 2},FF-tilde), as well as a Proca potential for the vector field, V(A{sup 2}). In particular it is demonstrated that theories involving only f(F{sup 2}) do not satisfy the hyperbolicity conditions. It is then shown that in this class of models, the cosmological dynamics always dilutes the vector field. In the case of a nonminimal coupling to gravity, it is established that theories involving Rf(A{sup 2}) or Rf(F{sup 2}) are generically pathologic. To finish, we exhibit a model where the vector field is not diluted during the cosmological evolution, because of a nonminimal vector field-curvature coupling which maintains second-order field equations. The relevance of such models for cosmology is discussed.

  9. Vector generator scan converter

    DOEpatents

    Moore, J.M.; Leighton, J.F.

    1988-02-05

    High printing speeds for graphics data are achieved with a laser printer by transmitting compressed graphics data from a main processor over an I/O channel to a vector generator scan converter which reconstructs a full graphics image for input to the laser printer through a raster data input port. The vector generator scan converter includes a microprocessor with associated microcode memory containing a microcode instruction set, a working memory for storing compressed data, vector generator hardware for drawing a full graphic image from vector parameters calculated by the microprocessor, image buffer memory for storing the reconstructed graphics image and an output scanner for reading the graphics image data and inputting the data to the printer. The vector generator scan converter eliminates the bottleneck created by the I/O channel for transmitting graphics data from the main processor to the laser printer, and increases printer speed up to thirty fold. 7 figs.

  10. Vector generator scan converter

    DOEpatents

    Moore, James M.; Leighton, James F.

    1990-01-01

    High printing speeds for graphics data are achieved with a laser printer by transmitting compressed graphics data from a main processor over an I/O (input/output) channel to a vector generator scan converter which reconstructs a full graphics image for input to the laser printer through a raster data input port. The vector generator scan converter includes a microprocessor with associated microcode memory containing a microcode instruction set, a working memory for storing compressed data, vector generator hardward for drawing a full graphic image from vector parameters calculated by the microprocessor, image buffer memory for storing the reconstructed graphics image and an output scanner for reading the graphics image data and inputting the data to the printer. The vector generator scan converter eliminates the bottleneck created by the I/O channel for transmitting graphics data from the main processor to the laser printer, and increases printer speed up to thirty fold.

  11. Line Integral of a Vector.

    ERIC Educational Resources Information Center

    Balabanian, Norman

    This programed booklet is designed for the engineering student who understands and can use vector and unit vector notation, components of a vector, parallel law of vector addition, and the dot product of two vectors. Content begins with work done by a force in moving a body a certain distance along some path. For each of the examples and problem…

  12. Construction of a Multiplex Promoter Reporter Platform to Monitor Staphylococcus aureus Virulence Gene Expression and the Identification of Usnic Acid as a Potent Suppressor of psm Gene Expression

    PubMed Central

    Gao, Peng; Wang, Yanli; Villanueva, Iván; Ho, Pak Leung; Davies, Julian; Kao, Richard Yi Tsun

    2016-01-01

    As antibiotic resistance becomes phenomenal, alternative therapeutic strategies for bacterial infections such as anti-virulence treatments have been advocated. We have constructed a total of 20 gfp-luxABCDE dual-reporter plasmids with selected promoters from S. aureus virulence-associated genes. The plasmids were introduced into various S. aureus strains to establish a gfp-lux based multiplex promoter reporter platform for monitoring S. aureus virulence gene expressions in real time to identify factors or compounds that may perturb virulence of S. aureus. The gene expression profiles monitored by luminescence correlated well with qRT-PCR results and extrinsic factors including carbon dioxide and some antibiotics were shown to suppress or induce the expression of virulence factors in this platform. Using this platform, sub-inhibitory ampicillin was shown to be a potent inducer for the expression of many virulence factors in S. aureus. Bacterial adherence and invasion assays using mammalian cells were employed to measure S. aureus virulence induced by ampicillin. The platform was used for screening of natural extracts that perturb the virulence of S. aureus and usnic acid was identified to be a potent repressor for the expression of psm. PMID:27625639

  13. Construction of a Multiplex Promoter Reporter Platform to Monitor Staphylococcus aureus Virulence Gene Expression and the Identification of Usnic Acid as a Potent Suppressor of psm Gene Expression.

    PubMed

    Gao, Peng; Wang, Yanli; Villanueva, Iván; Ho, Pak Leung; Davies, Julian; Kao, Richard Yi Tsun

    2016-01-01

    As antibiotic resistance becomes phenomenal, alternative therapeutic strategies for bacterial infections such as anti-virulence treatments have been advocated. We have constructed a total of 20 gfp-luxABCDE dual-reporter plasmids with selected promoters from S. aureus virulence-associated genes. The plasmids were introduced into various S. aureus strains to establish a gfp-lux based multiplex promoter reporter platform for monitoring S. aureus virulence gene expressions in real time to identify factors or compounds that may perturb virulence of S. aureus. The gene expression profiles monitored by luminescence correlated well with qRT-PCR results and extrinsic factors including carbon dioxide and some antibiotics were shown to suppress or induce the expression of virulence factors in this platform. Using this platform, sub-inhibitory ampicillin was shown to be a potent inducer for the expression of many virulence factors in S. aureus. Bacterial adherence and invasion assays using mammalian cells were employed to measure S. aureus virulence induced by ampicillin. The platform was used for screening of natural extracts that perturb the virulence of S. aureus and usnic acid was identified to be a potent repressor for the expression of psm. PMID:27625639

  14. Construction and immunological evaluation of truncated hepatitis B core particles carrying HBsAg amino acids 119-152 in the major immunodominant region (MIR).

    PubMed

    Su, Qiudong; Yi, Yao; Guo, Minzhuo; Qiu, Feng; Jia, Zhiyuan; Lu, Xuexin; Meng, Qingling; Bi, Shengli

    2013-09-13

    Hepatitis B capsid protein expressed in Escherichia coli can reassemble into icosahedral particles, which could strongly enhance the immunogenicity of foreign epitopes, especially those inserted into its major immunodominant region. Herein, we inserted the entire 'α' antigenic determinant amino acids (aa) 119-152 of HBsAg into the truncated HBc (aa 1-144), between Asp(78) and Pro(79). Prokaryotic expression showed that the mosaic HBc was mainly in the form of inclusion bodies. After denaturation with urea, it was dialyzed progressively for protein renaturation. We observed that before and after renaturation, mosaic HBc was antigenic as determined by HBsAg ELISA and a lot of viruslike particles were observed after renaturation. Thus, we further purified the mosaic viruslike particles by (NH4)2SO4 precipitation, DEAE chromatography, and Sepharose 4FF chromatography. Negative staining electron microscopy demonstrated the morphology of the viruslike particles. Immunization of Balb/c mice with mosaic particles induced the production of anti-HBs antibody and Th1 cell immune response supported by ELISPOT and CD4/CD8 proportions assay. In conclusion, we constructed mosaic hepatitis core particles displaying the entire 'α' antigenic determinant on the surface and laid a foundation for researching therapeutic hepatits B vaccines. PMID:23969156

  15. Synthesis, structures, and fluorescence properties of two d-d heterometallic cluster-based complexes constructed by N-(phosphonomethyl) iminodiacetic acid

    NASA Astrophysics Data System (ADS)

    Li, Sa-Ying

    2016-12-01

    Two novel d-d heterometallic cluster-based complexes constructed by N-(Phosphonomethyl)imino-diacetic acid (H4PMIDA) multifunctional ligand, [Cu2Zn2(PMIDA)2(H2O)3]·3H2O(1), [ZnNi14(PMIDA)6(H2O)18]·(NO3)6·15H2O(2), have been synthesized under hydrothermal conditions and characterized by elemental analyses, IR spectra, thermal analyses, and single-crystal X-ray diffraction. The complex 1 is one-dimensional heteronuclear molecular chain, which is further extends into a 3D supramolecular network through very extensive Osbnd H⋯O hydrogen bonds. The compound 2 is a rare novel zero-dimensional heteronuclear molecular cluster, which is further extends into a 3D supramolecular network through very extensive Osbnd H⋯O hydrogen bonds. Moreover, the solid-state fluorescence properties of the two complexes have also been investigated at room temperature.

  16. Two new coordination polymers constructed by naphthalene-1,4-dicarboxylic acid and 2,4-diamino-6-methyl-triazine

    NASA Astrophysics Data System (ADS)

    Li, Yamin; Xiao, Changyu; Zhang, Xudong; Xu, Yanhui; Li, Junrui; Lun, Huijie; Chen, Qi

    2013-08-01

    Two new transition metal coordination complexes, {[MnO(nda)](H2dmt)(H2O)}n (1), [Ag5(nda)2.5(dmt)]n (2), (H2nda=naphthalene-1,4-dicarboxylic acid, dmt=2,4-diamine-6-methyl-1,3,5-triazine) have been hydrothermally synthesized by the reactions of H2nda and dmt with the homologous MnCl2·4H2O and AgNO3, respectively, and characterized by single-crystal X-ray diffraction, IR spectra, elemental analysis, thermogravimetric analysis (TGA). The compound 1 exhibits a 3D network comprising 1D metal chain {MnO(CO2)2}n connected by the ligand nda2-, featuring a four-connected uninodal diamond -like topology. In compound 2, it is firstly observed that decanuclear silver units as secondary building units to construct 3D network by the ligands dmt and nda2-, with a rare 2-nodal (3,8)-connected tfz-d topology ({43}2{46.618.84}). The interactions within each Mn(II)-Mn(II) pair of compound 1 are antiferromagnetic (g=2.07, J=-1.42(1) cm-1, zj‧=-0.73(2) cm-1). In addition, compound 2 exhibits photoluminescent property at about 472 nm (λex=394 nm).

  17. Multiclass Reduced-Set Support Vector Machines

    NASA Technical Reports Server (NTRS)

    Tang, Benyang; Mazzoni, Dominic

    2006-01-01

    There are well-established methods for reducing the number of support vectors in a trained binary support vector machine, often with minimal impact on accuracy. We show how reduced-set methods can be applied to multiclass SVMs made up of several binary SVMs, with significantly better results than reducing each binary SVM independently. Our approach is based on Burges' approach that constructs each reduced-set vector as the pre-image of a vector in kernel space, but we extend this by recomputing the SVM weights and bias optimally using the original SVM objective function. This leads to greater accuracy for a binary reduced-set SVM, and also allows vectors to be 'shared' between multiple binary SVMs for greater multiclass accuracy with fewer reduced-set vectors. We also propose computing pre-images using differential evolution, which we have found to be more robust than gradient descent alone. We show experimental results on a variety of problems and find that this new approach is consistently better than previous multiclass reduced-set methods, sometimes with a dramatic difference.

  18. The vector-like twin Higgs

    NASA Astrophysics Data System (ADS)

    Craig, Nathaniel; Knapen, Simon; Longhi, Pietro; Strassler, Matthew

    2016-07-01

    We present a version of the twin Higgs mechanism with vector-like top partners. In this setup all gauge anomalies automatically cancel, even without twin leptons. The matter content of the most minimal twin sector is therefore just two twin tops and one twin bottom. The LHC phenomenology, illustrated with two example models, is dominated by twin glueball decays, possibly in association with Higgs bosons. We further construct an explicit four-dimensional UV completion and discuss a variety of UV completions relevant for both vector-like and fraternal twin Higgs models.

  19. Fractal vector optical fields.

    PubMed

    Pan, Yue; Gao, Xu-Zhen; Cai, Meng-Qiang; Zhang, Guan-Lin; Li, Yongnan; Tu, Chenghou; Wang, Hui-Tian

    2016-07-15

    We introduce the concept of a fractal, which provides an alternative approach for flexibly engineering the optical fields and their focal fields. We propose, design, and create a new family of optical fields-fractal vector optical fields, which build a bridge between the fractal and vector optical fields. The fractal vector optical fields have polarization states exhibiting fractal geometry, and may also involve the phase and/or amplitude simultaneously. The results reveal that the focal fields exhibit self-similarity, and the hierarchy of the fractal has the "weeding" role. The fractal can be used to engineer the focal field. PMID:27420485

  20. Lentiviral vector gene transfer to porcine airways.

    PubMed

    Sinn, Patrick L; Cooney, Ashley L; Oakland, Mayumi; Dylla, Douglas E; Wallen, Tanner J; Pezzulo, Alejandro A; Chang, Eugene H; McCray, Paul B

    2012-01-01

    In this study, we investigated lentiviral vector development and transduction efficiencies in well-differentiated primary cultures of pig airway epithelia (PAE) and wild-type pigs in vivo. We noted gene transfer efficiencies similar to that observed for human airway epithelia (HAE). Interestingly, feline immunodeficiency virus (FIV)-based vectors transduced immortalized pig cells as well as pig primary cells more efficiently than HIV-1-based vectors. PAE express TRIM5α, a well-characterized species-specific lentiviral restriction factor. We contrasted the restrictive properties of porcine TRIM5α against FIV- and HIV-based vectors using gain and loss of function approaches. We observed no effect on HIV-1 or FIV conferred transgene expression in response to porcine TRIM5α overexpression or knockdown. To evaluate the ability of GP64-FIV to transduce porcine airways in vivo, we delivered vector expressing mCherry to the tracheal lobe of the lung and the ethmoid sinus of 4-week-old pigs. One week later, epithelial cells expressing mCherry were readily detected. Our findings indicate that pseudotyped FIV vectors confer similar tropisms in porcine epithelia as observed in human HAE and provide further support for the selection of GP64 as an appropriate envelope pseudotype for future preclinical gene therapy studies in the porcine model of cystic fibrosis (CF).Molecular Therapy - Nucleic Acids (2012) 1, e56; doi:10.1038/mtna.2012.47; published online 27 November 2012. PMID:23187455

  1. Understanding Vector Fields.

    ERIC Educational Resources Information Center

    Curjel, C. R.

    1990-01-01

    Presented are activities that help students understand the idea of a vector field. Included are definitions, flow lines, tangential and normal components along curves, flux and work, field conservation, and differential equations. (KR)

  2. Gammaretroviral vectors: biology, technology and application.

    PubMed

    Maetzig, Tobias; Galla, Melanie; Baum, Christopher; Schambach, Axel

    2011-06-01

    Retroviruses are evolutionary optimized gene carriers that have naturally adapted to their hosts to efficiently deliver their nucleic acids into the target cell chromatin, thereby overcoming natural cellular barriers. Here we will review-starting with a deeper look into retroviral biology-how Murine Leukemia Virus (MLV), a simple gammaretrovirus, can be converted into an efficient vehicle of genetic therapeutics. Furthermore, we will describe how more rational vector backbones can be designed and how these so-called self-inactivating vectors can be pseudotyped and produced. Finally, we will provide an overview on existing clinical trials and how biosafety can be improved. PMID:21994751

  3. Bloch vector projection noise

    NASA Technical Reports Server (NTRS)

    Wang, Li-Jun; Bacon, A. M.; Zhao, H.-Z.; Thomas, J. E.

    1994-01-01

    In the optical measurement of the Bloch vector components describing a system of N two-level atoms, the quantum fluctuations in these components are coupled into the measuring optical field. This paper develops the quantum theory of optical measurement of Bloch vector projection noise. The preparation and probing of coherence in an effective two-level system consisting of the two ground states in an atomic three-level lambda-scheme are analyzed.

  4. Poynting-vector filter

    SciTech Connect

    Carrigan, Charles R.

    2011-08-02

    A determination is made of frequency components associated with a particular bearing or location resulting from sources emitting electromagnetic-wave energy for which a Poynting-Vector can be defined. The broadband frequency components associated with a specific direction or location of interest are isolated from other components in the power spectrum that are not associated with the direction or location of interest. The collection of pointing vectors can be used to characterize the source.

  5. Conceptualizing Vectors in College Geometry: A New Framework for Analysis of Student Approaches and Difficulties

    ERIC Educational Resources Information Center

    Kwon, Oh Hoon

    2012-01-01

    This dissertation documents a new way of conceptualizing vectors in college mathematics, especially in geometry. First, I will introduce three problems to show the complexity and subtlety of the construct of vectors with the classical vector representations. These highlight the need for a new framework that: (1) differentiates abstraction from a…

  6. Construction and immunological evaluation of truncated hepatitis B core particles carrying HBsAg amino acids 119–152 in the major immunodominant region (MIR)

    SciTech Connect

    Su, Qiudong; Yi, Yao; Guo, Minzhuo; Qiu, Feng; Jia, Zhiyuan; Lu, Xuexin; Meng, Qingling; Bi, Shengli

    2013-09-13

    Highlights: •The conformational HBV neutralization antigen domain was successfully displayed on the surface of truncated HBc particles. •Appropriate dialysis procedures to support the renaturing environment for the protein refolding. •Efficient purification procedures to obtain high purity and icosahedral particles of mosaic HBV antigen. •Strong immune responses not only including neutralization antibody response but also Th1 cell response were induced in mice. -- Abstract: Hepatitis B capsid protein expressed in Escherichia coli can reassemble into icosahedral particles, which could strongly enhance the immunogenicity of foreign epitopes, especially those inserted into its major immunodominant region. Herein, we inserted the entire ‘α’ antigenic determinant amino acids (aa) 119–152 of HBsAg into the truncated HBc (aa 1–144), between Asp{sup 78} and Pro{sup 79}. Prokaryotic expression showed that the mosaic HBc was mainly in the form of inclusion bodies. After denaturation with urea, it was dialyzed progressively for protein renaturation. We observed that before and after renaturation, mosaic HBc was antigenic as determined by HBsAg ELISA and a lot of viruslike particles were observed after renaturation. Thus, we further purified the mosaic viruslike particles by (NH{sub 4}){sub 2}SO{sub 4} precipitation, DEAE chromatography, and Sepharose 4FF chromatography. Negative staining electron microscopy demonstrated the morphology of the viruslike particles. Immunization of Balb/c mice with mosaic particles induced the production of anti-HBs antibody and Th1 cell immune response supported by ELISPOT and CD4/CD8 proportions assay. In conclusion, we constructed mosaic hepatitis core particles displaying the entire ‘α’ antigenic determinant on the surface and laid a foundation for researching therapeutic hepatits B vaccines.

  7. Resonance vector soliton of the Rayleigh wave.

    PubMed

    Adamashvili, G T

    2016-02-01

    A theory of acoustic vector solitons of self-induced transparency of the Rayleigh wave is constructed. A thin resonance transition layer on an elastic surface is considered using a model of a two-dimensional gas of impurity paramagnetic atoms or quantum dots. Explicit analytical expressions for the profile and parameters of the Rayleigh vector soliton with two different oscillation frequencies is obtained, as well as simulations of this nonlinear surface acoustic wave with realistic parameters, which can be used in acoustic experiments. It is shown that the properties of a surface vector soliton of the Rayleigh wave depend on the parameters of the resonance layer, the elastic medium, and the transverse structure of the surface acoustic wave. PMID:26986400

  8. Construction of a food-grade cell surface display system for Lactobacillus casei.

    PubMed

    Qin, Jiayang; Wang, Xiuwen; Kong, Jian; Ma, Cuiqing; Xu, Ping

    2014-01-01

    In this study, a food-grade cell surface display host/vector system for Lactobacillus casei was constructed. The food-grade host L. casei Q-5 was a lactose-deficient derivative of L. casei ATCC 334 obtained by plasmid elimination. The food-grade cell surface display vector was constructed based on safe DNA elements from lactic acid bacteria containing the following: pSH71 replicon from Lactococcus lactis, lactose metabolism genes from L. casei ATCC 334 as complementation markers, and surface layer protein gene from Lactobacillus acidophilus ATCC 4356 for cell surface display. The feasibility of the new host/vector system was verified by the expression of green fluorescent protein (GFP) on L. casei. Laser scanning confocal microscopy and immunofluorescence analysis using anti-GFP antibody confirmed that GFP was anchored on the surface of the recombinant cells. The stability of recombinant L. casei cells in artificial gastrointestinal conditions was verified, which is beneficial for oral vaccination applications. These results indicate that the food-grade host/vector system can be an excellent antigen delivery vehicle in oral vaccine construction. PMID:24598012

  9. Multiple Vector Preserving Interpolation Mappings in Algebraic Multigrid

    SciTech Connect

    Vassilevski, P S; Zikatanov, L T

    2004-11-03

    We propose algorithms for the construction of AMG (algebraic multigrid) interpolation mappings such that the resulting coarse space to span (locally and globally) any number of a priori given set of vectors. Specific constructions in the case of element agglomeration AMG methods are given. Some numerical illustration is also provided.

  10. Vector WIMP miracle

    NASA Astrophysics Data System (ADS)

    Abe, Tomohiro; Kakizaki, Mitsuru; Matsumoto, Shigeki; Seto, Osamu

    2012-07-01

    Weakly interacting massive particle (WIMP) is well known to be a good candidate for dark matter, and it is also predicted by many new physics models beyond the standard model at the TeV scale. We found that, if the WIMP is a vector particle (spin-one particle) which is associated with some gauge symmetry broken at the TeV scale, the Higgs mass is often predicted to be 120-125 GeV, which is very consistent with the result of Higgs searches recently reported by ATLAS and CMS Collaborations at the Large Hadron Collider experiment. In this Letter, we consider the vector WIMP using a non-linear sigma model in order to confirm this result as general as possible in a bottom-up approach. Near-future prospects to detect the vector WIMP at both direct and indirect detection experiments of dark matter are also discussed.

  11. Vector financial rogue waves

    NASA Astrophysics Data System (ADS)

    Yan, Zhenya

    2011-11-01

    The coupled nonlinear volatility and option pricing model presented recently by Ivancevic is investigated, which generates a leverage effect, i.e., stock volatility is (negatively) correlated to stock returns, and can be regarded as a coupled nonlinear wave alternative of the Black-Scholes option pricing model. In this Letter, we analytically propose vector financial rogue waves of the coupled nonlinear volatility and option pricing model without an embedded w-learning. Moreover, we exhibit their dynamical behaviors for chosen different parameters. The vector financial rogue wave (rogon) solutions may be used to describe the possible physical mechanisms for the rogue wave phenomena and to further excite the possibility of relative researches and potential applications of vector rogue waves in the financial markets and other related fields.

  12. Vectorized garbage collection

    SciTech Connect

    Appel, A.W.; Bendiksen, A.

    1988-01-01

    Garbage collection can be done in vector mode on supercomputers like the Cray-2 and the Cyber 205. Both copying collection and mark-and-sweep can be expressed as breadth-first searches in which the queue can be processed in parallel. The authors have designed a copying garbage collector whose inner loop works entirely in vector mode. The only significant limitation of the algorithm is that if the size of the records is not constant, the implementation becomes much more complicated. The authors give performance measurements of the algorithm as implemented for Lisp CONS cells on the Cyber 205. Vector-mode garbage collection performs up to 9 times faster than scalar-mode collection.

  13. Nonviral Vectors for Gene Delivery

    NASA Astrophysics Data System (ADS)

    Baoum, Abdulgader Ahmed

    2011-12-01

    The development of nonviral vectors for safe and efficient gene delivery has been gaining considerable attention recently. An ideal nonviral vector must protect the gene against degradation by nuclease in the extracellular matrix, internalize the plasma membrane, escape from the endosomal compartment, unpackage the gene at some point and have no detrimental effects. In comparison to viruses, nonviral vectors are relatively easy to synthesize, less immunogenic, low in cost, and have no limitation in the size of a gene that can be delivered. Significant progress has been made in the basic science and applications of various nonviral gene delivery vectors; however, the majority of nonviral approaches are still inefficient and often toxic. To this end, two nonviral gene delivery systems using either biodegradable poly(D,L-lactide- co-glycolide) (PLG) nanoparticles or cell penetrating peptide (CPP) complexes have been designed and studied using A549 human lung epithelial cells. PLG nanoparticles were optimized for gene delivery by varying particle surface chemistry using different coating materials that adsorb to the particle surface during formation. A variety of cationic coating materials were studied and compared to more conventional surfactants used for PLG nanoparticle fabrication. Nanoparticles (˜200 nm) efficiently encapsulated plasmids encoding for luciferase (80-90%) and slowly released the same for two weeks. After a delay, moderate levels of gene expression appeared at day 5 for certain positively charged PLG particles and gene expression was maintained for at least two weeks. In contrast, gene expression mediated by polyethyleneimine (PEI) ended at day 5. PLG particles were also significantly less cytotoxic than PEI suggesting the use of these vehicles for localized, sustained gene delivery to the pulmonary epithelium. On the other hand, a more simple method to synthesize 50-200 nm complexes capable of high transfection efficiency or high gene knockdown was

  14. Bunyavirus-Vector Interactions

    PubMed Central

    Horne, Kate McElroy; Vanlandingham, Dana L.

    2014-01-01

    The Bunyaviridae family is comprised of more than 350 viruses, of which many within the Hantavirus, Orthobunyavirus, Nairovirus, Tospovirus, and Phlebovirus genera are significant human or agricultural pathogens. The viruses within the Orthobunyavirus, Nairovirus, and Phlebovirus genera are transmitted by hematophagous arthropods, such as mosquitoes, midges, flies, and ticks, and their associated arthropods not only serve as vectors but also as virus reservoirs in many cases. This review presents an overview of several important emerging or re-emerging bunyaviruses and describes what is known about bunyavirus-vector interactions based on epidemiological, ultrastructural, and genetic studies of members of this virus family. PMID:25402172

  15. Two new coordination polymers constructed by naphthalene-1,4-dicarboxylic acid and 2,4-diamino-6-methyl-triazine

    SciTech Connect

    Li, Yamin; Xiao, Changyu; Zhang, Xudong; Xu, Yanhui; Li, Junrui; Lun, Huijie; Chen, Qi

    2013-08-15

    Two new transition metal coordination complexes, ([MnO(nda)](H{sub 2}dmt)(H{sub 2}O)){sub n} (1), [Ag{sub 5}(nda){sub 2.5}(dmt)]{sub n} (2), (H{sub 2}nda=naphthalene-1,4-dicarboxylic acid, dmt=2,4-diamine-6-methyl-1,3,5-triazine) have been hydrothermally synthesized by the reactions of H{sub 2}nda and dmt with the homologous MnCl{sub 2}·4H{sub 2}O and AgNO{sub 3}, respectively, and characterized by single-crystal X-ray diffraction, IR spectra, elemental analysis, thermogravimetric analysis (TGA). The compound 1 exhibits a 3D network comprising 1D metal chain (MnO(CO{sub 2}){sub 2}){sub n} connected by the ligand nda{sup 2−}, featuring a four-connected uninodal diamond -like topology. In compound 2, it is firstly observed that decanuclear silver units as secondary building units to construct 3D network by the ligands dmt and nda{sup 2−}, with a rare 2-nodal (3,8)-connected tfz-d topology ((4{sup 3}){sub 2}(4{sup 6}.6{sup 18}.8{sup 4})). The interactions within each Mn(II)—Mn(II) pair of compound 1 are antiferromagnetic (g=2.07, J=−1.42(1) cm{sup −1}, zj′=−0.73(2) cm{sup −1}). In addition, compound 2 exhibits photoluminescent property at about 472 nm (λ{sub ex}=394 nm). - Graphical abstract: Two new transition metal coordination complexes 1–2 have been hydrothermally synthesized and characterized by single-crystal X-ray diffraction, IR spectra, elemental analysis thermogravimetric analysis (TGA). Highlights: • The compound 1 exhibits a 3D network with four-connected uninodal diamond-like topology. • The first 3D network of 2 with a rare tfz-d topology consists of decanuclear silver clusters as secondary building units. • The magnetic measurement indicates the compound 1 shows antiferromagnetic interactions. • The photoluminescent property of 2 has been measured.

  16. Production of γ-linolenic acid and stearidonic acid by Synechococcus sp. PCC7002 containing cyanobacterial fatty acid desaturase genes

    NASA Astrophysics Data System (ADS)

    Dong, Xuewei; He, Qingfang; Peng, Zhenying; Yu, Jinhui; Bian, Fei; Li, Youzhi; Bi, Yuping

    2015-11-01

    Genetic modification is useful for improving the nutritional qualities of cyanobacteria. To increase the total unsaturated fatty acid content, along with the ratio of ω-3/ω-6 fatty acids, genetic engineering can be used to modify fatty acid metabolism. Synechococcus sp. PCC7002, a fast-growing cyanobacterium, does not contain a Δ6 desaturase gene and is therefore unable to synthesize γ-linolenic acid (GLA) and stearidonic acid (SDA), which are important in human health. In this work, we constructed recombinant vectors Syd6D, Syd15D and Syd6Dd15D to express the Δ15 desaturase and Δ6 desaturase genes from Synechocystis PCC6803 in Synechococcus sp. PCC7002, with the aim of expressing polyunsaturated fatty acids. Overexpression of the Δ15 desaturase gene in Synechococcus resulted in 5.4 times greater accumulation of α-linolenic acid compared with the wild-type while Δ6 desaturase gene expression produced both GLA and SDA. Co-expression of the two genes resulted in low-level accumulation of GLA but much larger amounts of SDA, accounting for as much to 11.64% of the total fatty acid content.

  17. Production of γ-linolenic acid and stearidonic acid by Synechococcus sp. PCC7002 containing cyanobacterial fatty acid desaturase genes

    NASA Astrophysics Data System (ADS)

    Dong, Xuewei; He, Qingfang; Peng, Zhenying; Yu, Jinhui; Bian, Fei; Li, Youzhi; Bi, Yuping

    2016-07-01

    Genetic modification is useful for improving the nutritional qualities of cyanobacteria. To increase the total unsaturated fatty acid content, along with the ratio of ω-3/ω-6 fatty acids, genetic engineering can be used to modify fatty acid metabolism. Synechococcus sp. PCC7002, a fast-growing cyanobacterium, does not contain a Δ6 desaturase gene and is therefore unable to synthesize γ-linolenic acid (GLA) and stearidonic acid (SDA), which are important in human health. In this work, we constructed recombinant vectors Syd6D, Syd15D and Syd6Dd15D to express the Δ15 desaturase and Δ6 desaturase genes from Synechocystis PCC6803 in Synechococcus sp. PCC7002, with the aim of expressing polyunsaturated fatty acids. Overexpression of the Δ15 desaturase gene in Synechococcus resulted in 5.4 times greater accumulation of α-linolenic acid compared with the wild-type while Δ6 desaturase gene expression produced both GLA and SDA. Co-expression of the two genes resulted in low-level accumulation of GLA but much larger amounts of SDA, accounting for as much to 11.64% of the total fatty acid content.

  18. Creation and validation of a widely applicable multiple gene transfer vector system for stable transformation in plant.

    PubMed

    Sun, Quanxi; Liu, Jiang; Li, Yaxiao; Zhang, Qin; Shan, Shihua; Li, Xinzheng; Qi, Baoxiu

    2013-11-01

    Multiple gene transfer (MGT) technology has become a powerful tool for basic and applied plant biology research in recent years. Despite some notable successes in obtaining plant lines harbouring multiple transgenes, these methods are still generally unwieldy and costly. We report here a straightforward and cost effective strategy, utilizing commonly available restriction enzymes for the transfer of multiple genes into plants, hence greatly widening the accessibility of MGT. This methodology exploits the specific 'nested' arrangement of a pair of isocaudomer restriction enzymes (for example XbaI-AvrII-XbaI) so that through the alternate use of these two enzymes in a reiterative fashion multiple genes/constructs (up to five in this study) could be 'stacked' together with ease. In a proof-of-concept experiment, we constructed a plant transformation vector containing three reporter gene expression cassettes flanked by two matrix attachment region sequences. The expression of all three genes was confirmed in transgenic Arabidopsis thaliana. The usefulness of this technology was further validated by the construction of a plant transformation vector containing five transgenes for the production of eicosapentaenoic acid (EPA, C20∆⁵,⁸,¹¹,¹⁴,¹⁷), a polyunsaturated essential fatty acid found in fish oils that is beneficial for health. In addition, we constructed four more vectors, incorporating one seed specific and three promoters conferring constitutive expression. These expression cassettes are flanked by a different isocaudomer pair (AvrII-SpeI-AvrII) and four other unique restriction sites, allowing the exchange of promoters and terminators of choice. PMID:23839253

  19. 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

  20. Support vector machines

    NASA Technical Reports Server (NTRS)

    Garay, Michael J.; Mazzoni, Dominic; Davies, Roger; Wagstaff, Kiri

    2004-01-01

    Support Vector Machines (SVMs) are a type of supervised learning algorith,, other examples of which are Artificial Neural Networks (ANNs), Decision Trees, and Naive Bayesian Classifiers. Supervised learning algorithms are used to classify objects labled by a 'supervisor' - typically a human 'expert.'.

  1. Singular Vectors' Subtle Secrets

    ERIC Educational Resources Information Center

    James, David; Lachance, Michael; Remski, Joan

    2011-01-01

    Social scientists use adjacency tables to discover influence networks within and among groups. Building on work by Moler and Morrison, we use ordered pairs from the components of the first and second singular vectors of adjacency matrices as tools to distinguish these groups and to identify particularly strong or weak individuals.

  2. Killing vectors and anisotropy

    SciTech Connect

    Krisch, J. P.; Glass, E. N.

    2009-08-15

    We consider an action that can generate fluids with three unequal stresses for metrics with a spacelike Killing vector. The parameters in the action are directly related to the stress anisotropies. The field equations following from the action are applied to an anisotropic cosmological expansion and an extension of the Gott-Hiscock cosmic string.

  3. Vector-borne diseases.

    PubMed

    Gubler, D J

    2009-08-01

    Vector-borne diseases have been the scourge of man and animals since the beginning of time. Historically, these are the diseases that caused the great plagues such as the 'Black Death' in Europe in the 14th Century and the epidemics of yellow fever that plagued the development of the New World. Others, such as Nagana, contributed to the lack of development in Africa for many years. At the turn of the 20th Century, vector-borne diseases were among the most serious public and animal health problems in the world. For the most part, these diseases were controlled by the middle of the 20th Century through the application of knowledge about their natural history along with the judicious use of DDT (dichlorodiphenyltrichloroethane) and other residual insecticides to interrupt the transmission cycle between arthropod and vertebrate host. However, this success initiated a period of complacency in the 1960s and 1970s, which resulted in the redirection of resources away from prevention and control of vector-borne diseases. The 1970s was also a time in which there were major changes to public health policy. Global trends, combined with changes in animal husbandry, urbanisation, modern transportation and globalisation, have resulted in a global re-emergence of epidemic vector-borne diseases affecting both humans and animals over the past 30 years. PMID:20128467

  4. Research in vector control

    PubMed Central

    Quarterman, K. D.

    1963-01-01

    Current research on vector control is directed mainly at finding answers to the problem of resistance. Despite considerable advances in knowledge of the genetics, biochemistry, physiology, and ecology of resistant vectors, the only practical answer found so far has been the development of new, substitute insecticides. At present the operational needs of existing large-scale control or eradication programmes swallow up much of the funds, personnel and facilities that might otherwise be devoted to basic research. Moreover, to back up these programmes, there is a continuing need for applied research on such questions as the packaging of pesticides, improvements in equipment and the development of new formulations. The author gives examples of applied research already carried out or in progress and indicates some areas of both basic and applied research demanding urgent attention. Like other participants in the seminar, he stresses the fundamental importance of ecological studies. He also examines the concept of integrated vector control and points out that the realization of this concept presupposes close co-ordination between basic and applied research, laboratory and field studies, and investigations on chemical and non-chemical vector control measures. PMID:20604177

  5. Seven supramolecular frameworks constructed from combination of hydrogen-bonds and other non-covalent associations between organic acids and bis-imidazoles

    NASA Astrophysics Data System (ADS)

    Jin, Shouwen; Zhang, Huan; Zhao, Ying; Jin, Li; Ye, Xianghang; Liu, Hui; Wang, Daqi

    2015-11-01

    Seven crystalline organic acid-base adducts derived from bis(N-imidazolyl) and organic acids (2,4,6-trinitrophenol, p-nitrobenzoic acid, 3,5-dinitrobenzoic acid, oxalic acid, m-phthalic acid, and 1,5-naphthalenedisulfonic acid) were prepared and characterized by X-ray diffraction analysis, IR, mp, and elemental analysis. The seven compounds are all organic salts. In salts 1, and 3, the L1 are monoprotonated, while in 4 and 6 the L1 are diprotonated. All supramolecular architectures involve extensive classical hydrogen bonds and C-H⋯O interactions. The role of weak and strong non-covalent interactions in the crystal packing is analyzed. The complexes displayed 2D-3D framework structures for the synergistic effect of the various non-covalent interactions. The results presented herein indicate that the strength and directionality of the N-H⋯N, N-H⋯O, O-H⋯O, O-H⋯N, N-H⋯S, and O-H⋯S hydrogen bonds between the organic acids and the ditopic imidazoles are sufficient to bring about the formation of binary organic salts.

  6. Monad constructions of omalous bundles

    NASA Astrophysics Data System (ADS)

    Henni, Abdelmoubine Amar; Jardim, Marcos

    2013-12-01

    We consider a particular class of holomorphic vector bundles relevant for supersymmetric string theory, called omalous, over nonsingular projective varieties. We use monads to construct examples of such bundles over 3-fold hypersurfaces in P4, complete intersection Calabi-Yau manifolds in Pk, blow-ups of P2 at n distinct points, and products Pm×Pn.

  7. Correlation properties of the vector signal representation for speckle pattern.

    PubMed

    Wang, Wei; Zhang, Shun; Ma, Ning

    2014-04-01

    In one-dimensional (1D) signal analysis, the complex analytic signal built from a real-valued signal and its Hilbert transform is an important tool providing a mathematical foundation for 1D statistical analysis. For a natural extension beyond 1D signal, Riesz transform has been applied to high-dimensional signal processing as a generalized Hilbert transform to construct a vector signal representation and therefore, to enlarge the traditional analytic signal concept. In this paper, we introduce the vector correlations as new mathematical tools for vector calculus for statistical speckle analysis. Based on vector correlations of a real-valued speckle pattern, we present the associated correlation properties, which can be regarded as mathematical foundation for the vector analysis in speckle metrology. PMID:24787197

  8. New Highly Dynamic Approach for Thrust Vector Control

    NASA Astrophysics Data System (ADS)

    Hecht, M.; Ettl, J.; Grothe, D.; Hrbud, I.

    2015-09-01

    For a new launcher system a thrust vector control system is needed. This launch vehicle system consists of two rockets which are namely the VS-50 (two-stage suborbital vehicle) and the VLM-1 (three-stage microsatellite launch vehicle). VLM-1 and VS-50 are developed in a cooperation between the German Aerospace Center (DLR) and the Brazilian Aeronautics and Space Institute (IAE). To keep these two rockets on its trajectory during flight a highly dynamic thrust vector control system is required. For the purpose of developing such a highly dynamic thrust vector control system a master thesis was written by the author. The development includes all mechanical constructions as well as control algorithms and electronics design. Moreover an optimization of control algorithms was made to increase the dynamic capabilities of the thrust vector control system. The composition of the right components plus the sophisticated control algorithm make the thrust vector control system highly dynamic.

  9. Construction aggregates

    USGS Publications Warehouse

    Tepordei, V.V.

    1995-01-01

    Part of the 1994 Industrial Minerals Review. The production, consumption, and applications of construction aggregates are reviewed. In 1994, the production of construction aggregates, which includes crushed stone and construction sand and gravel combined, increased 7.7 percent to 2.14 Gt compared with the previous year. These record production levels are mostly a result of funding for highway construction work provided by the Intermodal Surface Transportation Efficiency Act of 1991. Demand is expected to increase for construction aggregates in 1995.

  10. The Interaction between Vector Life History and Short Vector Life in Vector-Borne Disease Transmission and Control.

    PubMed

    Brand, Samuel P C; Rock, Kat S; Keeling, Matt J

    2016-04-01

    Epidemiological modelling has a vital role to play in policy planning and prediction for the control of vectors, and hence the subsequent control of vector-borne diseases. To decide between competing policies requires models that can generate accurate predictions, which in turn requires accurate knowledge of vector natural histories. Here we highlight the importance of the distribution of times between life-history events, using short-lived midge species as an example. In particular we focus on the distribution of the extrinsic incubation period (EIP) which determines the time between infection and becoming infectious, and the distribution of the length of the gonotrophic cycle which determines the time between successful bites. We show how different assumptions for these periods can radically change the basic reproductive ratio (R0) of an infection and additionally the impact of vector control on the infection. These findings highlight the need for detailed entomological data, based on laboratory experiments and field data, to correctly construct the next-generation of policy-informing models. PMID:27128163

  11. The Interaction between Vector Life History and Short Vector Life in Vector-Borne Disease Transmission and Control

    PubMed Central

    Brand, Samuel P. C.; Keeling, Matt J.

    2016-01-01

    Epidemiological modelling has a vital role to play in policy planning and prediction for the control of vectors, and hence the subsequent control of vector-borne diseases. To decide between competing policies requires models that can generate accurate predictions, which in turn requires accurate knowledge of vector natural histories. Here we highlight the importance of the distribution of times between life-history events, using short-lived midge species as an example. In particular we focus on the distribution of the extrinsic incubation period (EIP) which determines the time between infection and becoming infectious, and the distribution of the length of the gonotrophic cycle which determines the time between successful bites. We show how different assumptions for these periods can radically change the basic reproductive ratio (R0) of an infection and additionally the impact of vector control on the infection. These findings highlight the need for detailed entomological data, based on laboratory experiments and field data, to correctly construct the next-generation of policy-informing models. PMID:27128163

  12. Rotation vectors for homeomorphisms of non-positively curved manifolds

    NASA Astrophysics Data System (ADS)

    Lessa, Pablo

    2011-11-01

    Rotation vectors, as defined for homeomorphisms of the torus that are isotopic to the identity, are generalized to such homeomorphisms of any complete Riemannian manifold with non-positive sectional curvature. These generalized rotation vectors are shown to exist for almost every orbit of such a dynamical system with respect to any invariant measure with compact support. The concept is then extended to flows and, as an application, it is shown how non-null rotation vectors can be used to construct a measurable semi-conjugacy between a given flow and the geodesic flow of a manifold.

  13. Biological characterization of liver fatty acid binding gene from miniature pig liver cDNA library.

    PubMed

    Gao, Y H; Wang, K F; Zhang, S; Fan, Y N; Guan, W J; Ma, Y H

    2015-01-01

    Liver fatty acid binding proteins (L-FABP) are a family of small, highly conserved, cytoplasmic proteins that bind to long-chain fatty acids and other hydrophobic ligands. In this study, a full-length enriched cDNA library was successfully constructed from Wuzhishan miniature pig, and then the L-FABP gene was cloned from this cDNA library and an expression vector (pEGFP-N3-L-FABP) was constructed in vitro. This vector was transfected into hepatocytes to test its function. The results of western blotting analysis demonstrated that the L-FABP gene from our full-length enriched cDNA library regulated downstream genes, including the peroxisome proliferator-activated receptor family in hepatocytes. This study provides a theoretical basis and experimental evidence for the application of L-FABP for the treatment of liver injury. PMID:26345909

  14. Solving the inverse problem of high numerical aperture focusing using vector Slepian harmonics and vector Slepian multipole fields

    NASA Astrophysics Data System (ADS)

    Jahn, Kornél; Bokor, Nándor

    2013-02-01

    A technique using vector Slepian harmonics and vector Slepian multipole fields is presented for a general treatment of the inverse problem of high numerical aperture focusing. A prescribed intensity distribution or electric field distribution in the focal volume is approximated using numerical optimization and the corresponding illuminating field at the entrance pupil is constructed. Three examples from the recent literature are chosen to illustrate the method.

  15. Simulation of a vector hysteresis measurement system taking hysteresis into account by the vector Preisach model

    NASA Astrophysics Data System (ADS)

    Kuczmann, Miklós

    2008-02-01

    The paper deals with the numerical analysis of a rotational single sheet tester with round-shaped specimen (RRSST) which is now under construction. The measurement setup consists of an induction motor the rotor of which has been removed, and its windings have been replaced to a special two phase one which can generate homogeneous magnetic field inside the motor. The two orthogonal components of the magnetic field intensity and of the magnetic flux density vectors can be measured by H-coils and B-coils, respectively. The Finite Element Method (FEM) with the T, Φ-Φ potential formulation has been applied in the simulations. The vector hysteresis property of the specimen has been approximated by the vector Preisach model. Finally, the nonlinear problem has been solved by the fixed-point technique. The aim of the present work is to focus on the design aspects of this kind of measurement system.

  16. Preloadable vector sensitive latch

    NASA Technical Reports Server (NTRS)

    Acres, William R. (Inventor)

    1987-01-01

    A preloadable vector-sensitive latch which automatically releases when the force vector from a latch memebr reaches a specified release angle is presented. In addition, it contains means to remove clearance between the latched members and to preload the latch to prevent separation at angles less than the specified release angle. The latch comprises a triangular main link, a free link connected between a first corner of the main link and a yoke member, a housing, and an actuator connected between the yoke member and the housing. A return spring bias means connects the main link to a portion of the housing. A second corner of the main link is slidably and pivotally connected to the housing via a slot in a web portion of the housing. The latch housing has a rigid docking ring alignable with a mating locking ring which is engageable by a locking roller journalled on the third corner of the triangular main link.

  17. Vector Magnetograph Design

    NASA Technical Reports Server (NTRS)

    Chipman, Russell A.

    1996-01-01

    This report covers work performed during the period of November 1994 through March 1996 on the design of a Space-borne Solar Vector Magnetograph. This work has been performed as part of a design team under the supervision of Dr. Mona Hagyard and Dr. Alan Gary of the Space Science Laboratory. Many tasks were performed and this report documents the results from some of those tasks, each contained in the corresponding appendix. Appendices are organized in chronological order.

  18. Development of an Escherichia coli-Lactobacillus casei shuttle vector for heterologous protein expression in Lactobacillus casei.

    PubMed

    Suebwongsa, Namfon; Lulitanond, Viraphong; Mayo, Baltasar; Yotpanya, Panjamaporn; Panya, Marutpong

    2016-01-01

    There is an increasing interest to develop various lactic acid bacteria (LAB) species as mucosal delivery vehicles, for which the development of a variety of cloning and expression systems for these bacteria is of primary importance. This study reports the complete nucleotide sequence of the cryptic plasmid pRCEID7.6 derived from the chicken probiotic LAB strain Lactobacillus casei TISTR1341. Sequence analysis and comparison showed that pRCEID7.6 is composed of nine putative open reading frames. The replicon origin of pRCEID7.6 consisted of untranslated origin of replication and translated replication protein B sequences. This region was used to construct Escherichia coli/L. casei shuttle vectors carrying erythromycin and chloramphenicol resistance genes as selective markers. Segregation and structural stability of the vectors in L. casei was sufficient for most genetic applications. The feasibility of this vector for heterologous protein expression in L. casei was determined by cloning in pRCEID-LC7.6, the gene encoding the nucleocapsid protein (NP), from the influenza A virus under the control of the homologous promoter from the lactate dehydrogenase gene. L. casei carrying this recombinant plasmid was shown to successfully express the NP protein. Therefore, this shuttle vector can be used for further study in the development of mucosal delivery vehicles. PMID:27026866

  19. Some experiences with Krylov vectors and Lanczos vectors

    NASA Technical Reports Server (NTRS)

    Craig, Roy R., Jr.; Su, Tzu-Jeng; Kim, Hyoung M.

    1993-01-01

    This paper illustrates the use of Krylov vectors and Lanczos vectors for reduced-order modeling in structural dynamics and for control of flexible structures. Krylov vectors and Lanczos vectors are defined and illustrated, and several applications that have been under study at The University of Texas at Austin are reviewed: model reduction for undamped structural dynamics systems, component mode synthesis using Krylov vectors, model reduction of damped structural dynamics systems, and one-sided and two-sided unsymmetric block-Lanczos model-reduction algorithms.

  20. Isomap based supporting vector machine

    NASA Astrophysics Data System (ADS)

    Liang, W. N.

    2015-12-01

    This research presents a new isomap based supporting vector machine method. Isomap is a dimension reduction method which is able to analyze nonlinear relationship of data on manifolds. Accordingly, support vector machine is established on the isomap manifold to classify given and predict unknown data. A case study of the isomap based supporting vector machine for environmental planning problems is conducted.

  1. Application of Bred Vectors To Data Assimilation

    NASA Astrophysics Data System (ADS)

    Corazza, M.; Kalnay, E.; Patil, Dj

    We introduced a statistic, the BV-dimension, to measure the effective local finite-time dimensionality of the atmosphere. We show that this dimension is often quite low, and suggest that this finding has important implications for data assimilation and the accuracy of weather forecasting (Patil et al, 2001). The original database for this study was the forecasts of the NCEP global ensemble forecasting system. The initial differences between the control forecast and the per- turbed forecasts are called bred vectors. The control and perturbed initial conditions valid at time t=n(t are evolved using the forecast model until time t=(n+1) (t. The differences between the perturbed and the control forecasts are scaled down to their initial amplitude, and constitute the bred vectors valid at (n+1) (t. Their growth rate is typically about 1.5/day. The bred vectors are similar by construction to leading Lya- punov vectors except that they have small but finite amplitude, and they are valid at finite times. The original NCEP ensemble data set has 5 independent bred vectors. We define a local bred vector at each grid point by choosing the 5 by 5 grid points centered at the grid point (a region of about 1100km by 1100km), and using the north-south and east- west velocity components at 500mb pressure level to form a 50 dimensional column vector. Since we have k=5 global bred vectors, we also have k local bred vectors at each grid point. We estimate the effective dimensionality of the subspace spanned by the local bred vectors by performing a singular value decomposition (EOF analysis). The k local bred vector columns form a 50xk matrix M. The singular values s(i) of M measure the extent to which the k column unit vectors making up the matrix M point in the direction of v(i). We define the bred vector dimension as BVDIM={Sum[s(i)]}^2/{Sum[s(i)]^2} For example, if 4 out of the 5 vectors lie along v, and one lies along v, the BV- dimension would be BVDIM[sqrt(4), 1, 0

  2. A second branched-chain alpha-keto acid dehydrogenase gene cluster (bkdFGH) from Streptomyces avermitilis: its relationship to avermectin biosynthesis and the construction of a bkdF mutant suitable for the production of novel antiparasitic avermectins.

    PubMed Central

    Denoya, C D; Fedechko, R W; Hafner, E W; McArthur, H A; Morgenstern, M R; Skinner, D D; Stutzman-Engwall, K; Wax, R G; Wernau, W C

    1995-01-01

    A second cluster of genes encoding the E1 alpha, E1 beta, and E2 subunits of branched-chain alpha-keto acid dehydrogenase (BCDH), bkdFGH, has been cloned and characterized from Streptomyces avermitilis, the soil microorganism which produces anthelmintic avermectins. Open reading frame 1 (ORF1) (bkdF, encoding E1 alpha), would encode a polypeptide of 44,394 Da (406 amino acids). The putative start codon of the incompletely sequenced ORF2 (bkdG, encoding E1 beta) is located 83 bp downstream from the end of ORF1. The deduced amino acid sequence of bkdF resembled the corresponding E1 alpha subunit of several prokaryotic and eukaryotic BCDH complexes. An S. avermitilis bkd mutant constructed by deletion of a genomic region comprising the 5' end of bkdF is also described. The mutant exhibited a typical Bkd- phenotype: it lacked E1 BCDH activity and had lost the ability to grow on solid minimal medium containing isoleucine, leucine, and valine as sole carbon sources. Since BCDH provides an alpha-branched-chain fatty acid starter unit, either S(+)-alpha-methylbutyryl coenzyme A or isobutyryl coenzyme A, which is essential to initiate the synthesis of the avermectin polyketide backbone in S. avermitilis, the disrupted mutant cannot make the natural avermectins in a medium lacking both S(+)-alpha-methylbutyrate and isobutyrate. Supplementation with either one of these compounds restores production of the corresponding natural avermectins, while supplementation of the medium with alternative fatty acids results in the formation of novel avermectins. These results verify that the BCDH-catalyzed reaction of branched-chain amino acid catabolism constitutes a crucial step to provide fatty acid precursors for antibiotic biosynthesis in S. avermitilis. PMID:7768860

  3. Vector representation of tourmaline compositions

    NASA Technical Reports Server (NTRS)

    Burt, Donald M.

    1989-01-01

    The vector method for representing mineral compositions of amphibole and mica groups is applied to the tourmaline group. Consideration is given to the methods for drawing the relevant vector diagrams, relating the exchange vectors to one another, and contouring the diagrams for constant values of Na, Ca, Li, Fe, Mg, Al, Si, and OH. The method is used to depict a wide range of possible tourmaline end-member compositions and solid solutions, starting from a single point. In addition to vector depictions of multicomponent natural tourmalines, vectors are presented for simpler systems such as (Na,Al)-tourmalines, alkali-free tourmalines, and elbaites.

  4. Vector Theory of Ultrasonic Imaging

    NASA Astrophysics Data System (ADS)

    Gan, W. S.

    So far, works on ultrasonic diffraction imaging are based on scalar theory of sound wave. This is not correct as sound has vector nature. But when sound propagates in solids, its vector nature has to be considered as polarization occurs and transverse wave as well as longitudinal wave will appear. Vector theory is especially needed when the obstacle size is smaller than the wavelength. We use the Smythe-Kirchhoff approach for the vector theory of diffraction. We derive the image formation theory based on the vector diffraction theory. The effect of polarization on acoustical imaging is discussed.

  5. Construction Technology.

    ERIC Educational Resources Information Center

    Barnes, James L.

    This curriculum guide is designed to assist junior high school industrial arts teachers in planning new courses and revising existing courses in construction technology. Addressed in the individual units of the guide are the following topics: basic types of construction and the impact of construction on society, preconstruction, personnel…

  6. Electrochemical construction

    DOEpatents

    Einstein, Harry; Grimes, Patrick G.

    1983-08-23

    An electrochemical cell construction features a novel co-extruded plastic electrode in an interleaved construction with a novel integral separator-spacer. Also featured is a leak and impact resistant construction for preventing the spill of corrosive materials in the event of rupture.

  7. A Ring Construction Using Finite Directed Graphs

    ERIC Educational Resources Information Center

    Bardzell, Michael

    2012-01-01

    In this paper we discuss an interesting class of noncommutative rings which can be constructed using finite directed graphs. This construction also creates a vector space. These structures provide undergraduate students connections between ring theory and graph theory and, among other things, allow them to see a ring unity element that looks quite…

  8. DNA-based methods to prepare helper virus-free herpes amplicon vectors and versatile design of amplicon vector plasmids.

    PubMed

    Kasai, Kazue; Saeki, Yoshinaga

    2006-06-01

    The herpes simplex virus (HSV) amplicon vector is a versatile plasmid-based gene delivery vehicle with a large transgene capacity (up to 150 kb) and the ability to infect a broad range of cell types. The vector system was originally developed by Frenkel and her colleagues in 1980. Ever since, a great deal of effort by various investigators has been directed at minimizing the toxicity associated with the inevitable contamination by helper virus. In 1996, Fraefel and his colleagues successfully devised a cosmid-based packaging system that was free of contamination by helper virus (so-called helper virus-free packaging), which utilized as helper a set of 5 overlapping cosmid clones that covered the entire HSV genome, which lacked the DNA packaging/cleavage signals. With the helper virus-free system, broader applications of the vector became possible. Cloning of the entire HSV genome in bacteria artificial chromosome (BAC) plasmids enabled stable maintenance and propagation of the helper HSV genome in bacteria. It also allowed for the development of BAC-based helper virus-free packaging systems. In this article, we review various versions of DNA-based methods to prepare HSV amplicon vectors free of helper virus contamination. We also examine recent advances in vector design, including methods of vector construction, hybrid amplicon vectors, and the infectious BAC system. Future directions in improving packaging systems and vector designs are discussed. PMID:16787182

  9. EasyClone-MarkerFree: A vector toolkit for marker-less integration of genes into Saccharomyces cerevisiae via CRISPR-Cas9.

    PubMed

    Jessop-Fabre, Mathew M; Jakočiūnas, Tadas; Stovicek, Vratislav; Dai, Zongjie; Jensen, Michael K; Keasling, Jay D; Borodina, Irina

    2016-08-01

    Saccharomyces cerevisiae is an established industrial host for production of recombinant proteins, fuels and chemicals. To enable stable integration of multiple marker-free overexpression cassettes in the genome of S. cerevisiae, we have developed a vector toolkit EasyClone-MarkerFree. The integration of linearized expression cassettes into defined genomic loci is facilitated by CRISPR/Cas9. Cas9 is recruited to the chromosomal location by specific guide RNAs (gRNAs) expressed from a set of gRNA helper vectors. Using our genome engineering vector suite, single and triple insertions are obtained with 90-100% and 60-70% targeting efficiency, respectively. We demonstrate application of the vector toolkit by constructing a haploid laboratory strain (CEN.PK113-7D) and a diploid industrial strain (Ethanol Red) for production of 3-hydroxypropionic acid, where we tested three different acetyl-CoA supply strategies, requiring overexpression of three to six genes each. Among the tested strategies was a bacterial cytosolic pyruvate dehydrogenase complex, which was integrated into the genome in a single transformation. The publicly available EasyClone-MarkerFree vector suite allows for facile and highly standardized genome engineering, and should be of particular interest to researchers working on yeast chassis with limited markers available. PMID:27166612

  10. Acceleration of convergence of vector sequences

    NASA Technical Reports Server (NTRS)

    Sidi, A.; Ford, W. F.; Smith, D. A.

    1986-01-01

    A general approach to the construction of convergence acceleration methods for vector sequence is proposed. Using this approach, one can generate some known methods, such as the minimal polynomial extrapolation, the reduced rank extrapolation, and the topological epsilon algorithm, and also some new ones. Some of the new methods are easier to implement than the known methods and are observed to have similar numerical properties. The convergence analysis of these new methods is carried out, and it is shown that they are especially suitable for accelerating the convergence of vector sequences that are obtained when one solves linear systems of equations iterative. A stability analysis is also given, and numerical examples are provided. The convergence and stability properties of the topological epsilon algorithm are likewise given.

  11. Acceleration of convergence of vector sequences

    NASA Technical Reports Server (NTRS)

    Sidi, A.; Ford, W. F.; Smith, D. A.

    1983-01-01

    A general approach to the construction of convergence acceleration methods for vector sequence is proposed. Using this approach, one can generate some known methods, such as the minimal polynomial extrapolation, the reduced rank extrapolation, and the topological epsilon algorithm, and also some new ones. Some of the new methods are easier to implement than the known methods and are observed to have similar numerical properties. The convergence analysis of these new methods is carried out, and it is shown that they are especially suitable for accelerating the convergence of vector sequences that are obtained when one solves linear systems of equations iteratively. A stability analysis is also given, and numerical examples are provided. The convergence and stability properties of the topological epsilon algorithm are likewise given.

  12. Thrust Vector Control using movable probes

    NASA Technical Reports Server (NTRS)

    Cavalleri, Robert; Tiarn, Weihnurng; Readey, Harvey

    1990-01-01

    A study was undertaken to determine if movable probes or struts positioned in the nozzle can be used to provide Thrust Vector Control of the Space Shuttle Solid Rocket Booster. The study employed CFD to determine estimates of the shock standoff distance from the probe. An empirical correlation was used to construct the shock shape and the pressure distribution generated by the probe. The TVC performance for a single and multiple number of probes was then used to determine requirements for a maximum thrust angle offset of 7.5 degrees. Consideration was given to what materials would be suitable for the probe and if active cooling is required. Based on the performance analysis and thermal requirements, a Probe Thrust Vector Control (PTVC) system was sized. Indications are that a PTVC system weight is in the 1500 1bm weight range, compared to the existing weight of 7500 1bm for the SRB nozzle gimble system.

  13. Dual Vector Spaces and Physical Singularities

    NASA Astrophysics Data System (ADS)

    Rowlands, Peter

    Though we often refer to 3-D vector space as constructed from points, there is no mechanism from within its definition for doing this. In particular, space, on its own, cannot accommodate the singularities that we call fundamental particles. This requires a commutative combination of space as we know it with another 3-D vector space, which is dual to the first (in a physical sense). The combination of the two spaces generates a nilpotent quantum mechanics/quantum field theory, which incorporates exact supersymmetry and ultimately removes the anomalies due to self-interaction. Among the many natural consequences of the dual space formalism are half-integral spin for fermions, zitterbewegung, Berry phase and a zero norm Berwald-Moor metric for fermionic states.

  14. Vector ecology of equine piroplasmosis.

    PubMed

    Scoles, Glen A; Ueti, Massaro W

    2015-01-01

    Equine piroplasmosis is a disease of Equidae, including horses, donkeys, mules, and zebras, caused by either of two protozoan parasites, Theileria equi or Babesia caballi. These parasites are biologically transmitted between hosts via tick vectors, and although they have inherent differences they are categorized together because they cause similar pathology and have similar morphologies, life cycles, and vector relationships. To complete their life cycle, these parasites must undergo a complex series of developmental events, including sexual-stage development in their tick vectors. Consequently, ticks are the definitive hosts as well as vectors for these parasites, and the vector relationship is restricted to a few competent tick species. Because the vector relationship is critical to the epidemiology of these parasites, we highlight current knowledge of the vector ecology of these tick-borne equine pathogens, emphasizing tick transmissibility and potential control strategies to prevent their spread. PMID:25564746

  15. Nonlinear Growth of Singular Vector Based Perturbations

    NASA Astrophysics Data System (ADS)

    Reynolds, C. A.

    2002-12-01

    The nonlinearity of singular vector-based perturbation growth is examined within the context of a global atmospheric forecast model. The characteristics of these nonlinearities and their impact on the utility of SV-based diagnostics are assessed both qualitatively and quantitatively. Nonlinearities are quantified by examining the symmetry of evolving positive and negative "twin" perturbations. Perturbations initially scaled to be consistent with estimates of analysis uncertainty become significantly nonlinear by 12 hours. However, the relative magnitude of the nonlinearities is a strong function of scale and metric. Small scales become nonlinear very quickly while synoptic scales can remain significantly linear out to three day. Small shifts between positive and negative perturbations can result in significant nonlinearities even when the basic anomaly patterns are quite similar. Thus, singular vectors may be qualitatively useful even when nonlinearities are large. Post-time pseudo-inverse experiments show that despite significant nonlinear perturbation growth, the nonlinear forecast corrections are similar to the expected linear corrections, even at 72 hours. When the nonlinear correction does differ significantly from the expected linear correction, the nonlinear correction is usually better, indicating that in some cases the pseudo-inverse correction effectively suppresses error growth outside the subspace defined by the leading (dry) singular vectors. Because a significant portion of the nonlinear growth occurs outside of the dry singular vector subspace, an a priori nonlinearity index based on the full perturbations is not a good predictor of when pseudo-inverse based corrections will be ineffective. However, one can construct a reasonable predictor of pseudo-inverse ineffectiveness by focusing on nonlinearities in the synoptic scales or in the singular vector subspace only.

  16. Aerodynamics of thrust vectoring

    NASA Technical Reports Server (NTRS)

    Tseng, J. B.; Lan, C. Edward

    1989-01-01

    Thrust vectoring as a means to enhance maneuverability and aerodynamic performane of a tactical aircraft is discussed. This concept usually involves the installation of a multifunction nozzle. With the nozzle, the engine thrust can be changed in direction without changing the attitude of the aircraft. Change in the direction of thrust induces a significant change in the aerodynamic forces on the aircraft. Therefore, this device can be used for lift-augmenting as well as stability and control purposes. When the thrust is deflected in the longitudinal direction, the lift force and the pitching stability can be manipulated, while the yawing stability can be controlled by directing the thrust in the lateral direction.

  17. Catalytic asymmetric construction of spiro[pyrrolidine-2,3'-oxindole] scaffolds through chiral phosphoric acid-catalyzed 1,3-dipolar cycloaddition involving 3-amino oxindoles.

    PubMed

    Zhu, Guodong; Wang, Baomin; Bao, Xiaoze; Zhang, Huanrui; Wei, Qian; Qu, Jingping

    2015-11-01

    The catalytic asymmetric three-component 1,3-dipolar cycloaddition of 3-amino oxindoles with aldehydes and nitroolefins under the catalysis of a chiral phosphoric acid is reported. The reaction provides a facile approach to synthesize a diverse array of spiro[pyrrolidine-2,3'-oxindoles] in high yields with excellent diastereo- and enantioselectivities under mild conditions. PMID:26345264

  18. Chromatography purification of canine adenoviral vectors.

    PubMed

    Segura, María Mercedes; Puig, Meritxell; Monfar, Mercè; Chillón, Miguel

    2012-06-01

    Canine adenovirus vectors (CAV2) are currently being evaluated for gene therapy, oncolytic virotherapy, and as vectors for recombinant vaccines. Despite the need for increasing volumes of purified CAV2 preparations for preclinical and clinical testing, their purification still relies on the use of conventional, scale-limited CsCl ultracentrifugation techniques. A complete downstream processing strategy for CAV2 vectors based on membrane filtration and chromatography is reported here. Microfiltration and ultra/diafiltration are selected for clarification and concentration of crude viral stocks containing both intracellular and extracellular CAV2 particles. A DNase digestion step is introduced between ultrafiltration and diafiltration operations. At these early stages, concentration of vector stocks with good recovery of viral particles (above 80%) and removal of a substantial amount of protein and nucleic acid contaminants is achieved. The ability of various chromatography techniques to isolate CAV2 particles was evaluated. Hydrophobic interaction chromatography using a Fractogel propyl tentacle resin was selected as a first chromatography step, because it allows removal of the bulk of contaminating proteins with high CAV2 yields (88%). An anion-exchange chromatography step using monolithic supports is further introduced to remove the remaining contaminants with good recovery of CAV2 particles (58-69%). The main CAV2 viral structural components are visualized in purified preparations by electrophoresis analyses. Purified vector stocks contained intact icosahedral viral particles, low contamination with empty viral capsids (10%), and an acceptable total-to-infectious particle ratio (below 30). The downstream processing strategy that was developed allows preparation of large volumes of high-quality CAV2 stocks. PMID:22799886

  19. Vector-vector production in photon-photon interactions

    NASA Astrophysics Data System (ADS)

    Ronan, Micheal T.

    1989-04-01

    Measurements of exclusive untagged ρ0ρ0,ρφ,K*K¯*, and ρω production and tagged ρ0ρ0 production in photon-photon interactions by the TPC/Two-Gamma experiment are reviewed. Comparisons to the results of other experiments and to models of vector-vector production are made. Fits to the data following a four quark model prescription for vector meson pair production are also presented.

  20. A stable RNA virus-based vector for citrus trees

    SciTech Connect

    Folimonov, Alexey S.; Folimonova, Svetlana Y.; Bar-Joseph, Moshe; Dawson, William O.

    2007-11-10

    Virus-based vectors are important tools in plant molecular biology and plant genomics. A number of vectors based on viruses that infect herbaceous plants are in use for expression or silencing of genes in plants as well as screening unknown sequences for function. Yet there is a need for useful virus-based vectors for woody plants, which demand much greater stability because of the longer time required for systemic infection and analysis. We examined several strategies to develop a Citrus tristeza virus (CTV)-based vector for transient expression of foreign genes in citrus trees using a green fluorescent protein (GFP) as a reporter. These strategies included substitution of the p13 open reading frame (ORF) by the ORF of GFP, construction of a self-processing fusion of GFP in-frame with the major coat protein (CP), or expression of the GFP ORF as an extra gene from a subgenomic (sg) mRNA controlled either by a duplicated CTV CP sgRNA controller element (CE) or an introduced heterologous CE of Beet yellows virus. Engineered vector constructs were examined for replication, encapsidation, GFP expression during multiple passages in protoplasts, and for their ability to infect, move, express GFP, and be maintained in citrus plants. The most successful vectors based on the 'add-a-gene' strategy have been unusually stable, continuing to produce GFP fluorescence after more than 4 years in citrus trees.

  1. Equivariant algebraic vector bundles over representations of reductive groups: theory.

    PubMed Central

    Masuda, M; Petrie, T

    1991-01-01

    Let G be a reductive algebraic group and let B be an affine variety with an algebraic action of G. Everything is defined over the field C of complex numbers. Consider the trivial G-vector bundle B x S = S over B where S is a G-module. From the endomorphism ring R of the G-vector bundle S a construction of G-vector bundles over B is given. The bundles constructed this way have the property that when added to S they are isomorphic to F + S for a fixed G-module F. For such a bundle E an invariant rho(E) is defined that lies in a quotient of R. This invariant allows us to distinguish nonisomorphic G-vector bundles. This is applied to the case where B is a G-module and, in that case, an invariant of the underlying equivariant variety is given too. These constructions and invariants are used to produce families of inequivalent G-vector bundles over G-modules and families of inequivalent G actions on affine spaces for some finite and some connected semisimple groups. PMID:11607220

  2. Construction of crystalline 2D covalent organic frameworks with remarkable chemical (acid/base) stability via a combined reversible and irreversible route.

    PubMed

    Kandambeth, Sharath; Mallick, Arijit; Lukose, Binit; Mane, Manoj V; Heine, Thomas; Banerjee, Rahul

    2012-12-01

    Two new chemically stable [acid and base] 2D crystalline covalent organic frameworks (COFs) (TpPa-1 and TpPa-2) were synthesized using combined reversible and irreversible organic reactions. Syntheses of these COFs were done by the Schiff base reactions of 1,3,5-triformylphloroglucinol (Tp) with p-phenylenediamine (Pa-1) and 2,5-dimethyl-p-phenylenediamine (Pa-2), respectively, in 1:1 mesitylene/dioxane. The expected enol-imine (OH) form underwent irreversible proton tautomerism, and only the keto-enamine form was observed. Because of the irreversible nature of the total reaction and the absence of an imine bond in the system, TpPa-1 and TpPa-2 showed strong resistance toward acid (9 N HCl) and boiling water. Moreover, TpPa-2 showed exceptional stability in base (9 N NaOH) as well. PMID:23153356

  3. Construction aggregates

    USGS Publications Warehouse

    Tepordei, V.V.

    1996-01-01

    Part of the Annual Commodities Review 1995. Production of construction aggregates such as crushed stone and construction sand and gravel showed a marginal increase in 1995. Most of the 1995 increases were due to funding for highway construction work. The major areas of concern to the industry included issues relating to wetlands classification and the classification of crystalline silica as a probable human carcinogen. Despite this, an increase in demand is anticipated for 1996.

  4. Construction aggregates

    USGS Publications Warehouse

    Langer, W.H.; Tepordei, V.V.; Bolen, W.P.

    2000-01-01

    Construction aggregates consist primarily of crushed stone and construction sand and gravel. Total estimated production of construction aggregates increased in 1999 by about 2% to 2.39 Gt (2.64 billion st) compared with 1998. This record production level continued an expansion that began in 1992. By commodities, crushed stone production increased 3.3%, while sand and gravel production increased by about 0.5%.

  5. Construction aggregates

    USGS Publications Warehouse

    Tepordei, V.V.

    1994-01-01

    Part of a special section on industrial minerals in 1993. The 1993 production of construction aggregates increased 6.3 percent over the 1992 figure, to reach 2.01 Gt. This represents the highest estimated annual production of combined crushed stone and construction sand and gravel ever recorded in the U.S. The outlook for construction aggregates and the issues facing the industry are discussed.

  6. Solar vector magnetograph for Max 1991 programs

    NASA Technical Reports Server (NTRS)

    Rust, D. M.; Obyrne, J. W.; Harris, T. J.

    1988-01-01

    An instrument for measuring solar magnetic fields is under construction. Key requirements for any solar vector magnetograph are high spatial resolution, high optical throughput, fine spectral selectivity, and ultralow instrumental polarization. An available 25 cm Cassegrain telescope will provide 0.5 arcsec spatial resolution. Spectral selection will be accomplished with a 150 mA filter based on electrically tunable solid Fabry-Perot etalon. Filter and polarization analyzer design concepts for the magnetograph are described in detail. The instrument will be tested at JHU/APL, and then moved to the National Solar Observatory in late 1988. It will be available to support the Max 1991 program.

  7. Three silver-based complexes constructed from organic carboxylic acid and 4,4‧-bipyridine-like ligands: Syntheses, structures and photocatalytic properties

    NASA Astrophysics Data System (ADS)

    Wang, Chong-chen; Jing, Huan-ping; Wang, Peng

    2014-09-01

    The reactions of AgNO3 with combinations of 4,4‧-bipyridine (bpy)/1,2-di(4-pyridyl)ethylene (dpe), and 5-aminophthalic acid (H2ap)/2,6-Naphthalenedicarboxylic acid (H2npdc)/4,4‧-stilbenedicarboxylic acid (H2sbdc) in aqueous alcohol/ammonia at room temperature produce crystals of [Ag4(bpy)4](ap)2·11H2O (1), [Ag2(bpy)2](npdc)·2H2O (2) and [Ag2(dpe)1.5(sbdc)0.5](sbdc)0.5·7H2O (3). All complexes 1, 2 and 3 consist of 1D infinite silver-ligand cationic chains, interspersed with organic carboxylate anions that provide charge compensation in the crystal structures. The lattice water molecules are situated among the framework of the crystal structure and show rich hydrogen-bonding interactions, which serve to orientate of the organic carboxylate anions in the crystal packing, while the presence of Ag⋯N and Ag⋯Ag contacts strengthens the frameworks. In addition, complexes 1-3 exhibit good photocatalytic activities for dye decolorization under UV light.

  8. Constructive Fun.

    ERIC Educational Resources Information Center

    Simanek, Donald E.

    1994-01-01

    Compares and reviews currently available brands of steel construction sets that are useful to physics teachers for building demonstrations, prototypes of mechanisms, robotics, and remote control devices. (ZWH)

  9. Urbanization, land tenure security and vector-borne Chagas disease.

    PubMed

    Levy, Michael Z; Barbu, Corentin M; Castillo-Neyra, Ricardo; Quispe-Machaca, Victor R; Ancca-Juarez, Jenny; Escalante-Mejia, Patricia; Borrini-Mayori, Katty; Niemierko, Malwina; Mabud, Tarub S; Behrman, Jere R; Naquira-Velarde, Cesar

    2014-08-22

    Modern cities represent one of the fastest growing ecosystems on the planet. Urbanization occurs in stages; each stage characterized by a distinct habitat that may be more or less susceptible to the establishment of disease vector populations and the transmission of vector-borne pathogens. We performed longitudinal entomological and epidemiological surveys in households along a 1900 × 125 m transect of Arequipa, Peru, a major city of nearly one million inhabitants, in which the transmission of Trypanosoma cruzi, the aetiological agent of Chagas disease, by the insect vector Triatoma infestans, is an ongoing problem. The transect spans a cline of urban development from established communities to land invasions. We find that the vector is tracking the development of the city, and the parasite, in turn, is tracking the dispersal of the vector. New urbanizations are free of vector infestation for decades. T. cruzi transmission is very recent and concentrated in more established communities. The increase in land tenure security during the course of urbanization, if not accompanied by reasonable and enforceable zoning codes, initiates an influx of construction materials, people and animals that creates fertile conditions for epidemics of some vector-borne diseases. PMID:24990681

  10. Urbanization, land tenure security and vector-borne Chagas disease

    PubMed Central

    Levy, Michael Z.; Barbu, Corentin M.; Castillo-Neyra, Ricardo; Quispe-Machaca, Victor R.; Ancca-Juarez, Jenny; Escalante-Mejia, Patricia; Borrini-Mayori, Katty; Niemierko, Malwina; Mabud, Tarub S.; Behrman, Jere R.; Naquira-Velarde, Cesar

    2014-01-01

    Modern cities represent one of the fastest growing ecosystems on the planet. Urbanization occurs in stages; each stage characterized by a distinct habitat that may be more or less susceptible to the establishment of disease vector populations and the transmission of vector-borne pathogens. We performed longitudinal entomological and epidemiological surveys in households along a 1900 × 125 m transect of Arequipa, Peru, a major city of nearly one million inhabitants, in which the transmission of Trypanosoma cruzi, the aetiological agent of Chagas disease, by the insect vector Triatoma infestans, is an ongoing problem. The transect spans a cline of urban development from established communities to land invasions. We find that the vector is tracking the development of the city, and the parasite, in turn, is tracking the dispersal of the vector. New urbanizations are free of vector infestation for decades. T. cruzi transmission is very recent and concentrated in more established communities. The increase in land tenure security during the course of urbanization, if not accompanied by reasonable and enforceable zoning codes, initiates an influx of construction materials, people and animals that creates fertile conditions for epidemics of some vector-borne diseases. PMID:24990681

  11. Multiscale hierarchical support vector clustering

    NASA Astrophysics Data System (ADS)

    Hansen, Michael Saas; Holm, David Alberg; Sjöstrand, Karl; Ley, Carsten Dan; Rowland, Ian John; Larsen, Rasmus

    2008-03-01

    Clustering is the preferred choice of method in many applications, and support vector clustering (SVC) has proven efficient for clustering noisy and high-dimensional data sets. A method for multiscale support vector clustering is demonstrated, using the recently emerged method for fast calculation of the entire regularization path of the support vector domain description. The method is illustrated on artificially generated examples, and applied for detecting blood vessels from high resolution time series of magnetic resonance imaging data. The obtained results are robust while the need for parameter estimation is reduced, compared to support vector clustering.

  12. VLSI Processor For Vector Quantization

    NASA Technical Reports Server (NTRS)

    Tawel, Raoul

    1995-01-01

    Pixel intensities in each kernel compared simultaneously with all code vectors. Prototype high-performance, low-power, very-large-scale integrated (VLSI) circuit designed to perform compression of image data by vector-quantization method. Contains relatively simple analog computational cells operating on direct or buffered outputs of photodetectors grouped into blocks in imaging array, yielding vector-quantization code word for each such block in sequence. Scheme exploits parallel-processing nature of vector-quantization architecture, with consequent increase in speed.

  13. Localization and vector spherical harmonics

    NASA Astrophysics Data System (ADS)

    von Brecht, James H.

    2016-01-01

    This paper establishes the following localization property for vector spherical harmonics: a wide class of non-local, vector-valued operators reduce to local, multiplication-type operations when applied to a vector spherical harmonic. As localization occurs in a very precise, quantifiable and explicitly computable fashion, the localization property provides a set of useful formulae for analyzing vector-valued fractional diffusion and non-local differential equations defined on S d - 1. As such analyses require a detailed understanding of operators for which localization occurs, we provide several applications of the result in the context of non-local differential equations.

  14. The MSFC Vector Magnetograph

    NASA Astrophysics Data System (ADS)

    Hagyard, M. J.; Cumings, N. P.; West, E. A.; Smith, J. E.

    1982-09-01

    The NASA/Marshall Space Flight Center's solar vector magnetograph system is described; this system allows measurements of all components of the Sun's photospheric magnetic field over a 5 × 5 or 2.0 × 2.0 arc min square field-of-view with an optimum time resolution of ˜ 100 s and an optimum signal-to-noise of ˜1600. The basic system components are described, including the optics, detector, digital system and associated electronics. Automatic sequencing and control functions are outlined as well as manual selections of system parameters which afford unique system flexibility. Results of system calibration and performance are presented, including linearity, dynamic range, uniformity, spatial and spectral resolutions, signal-to-noise, electro-optical retardation and polarization calibration. Scientific investigations which utilize the unique characteristics of the instrument are described and typical results are presented.

  15. The MSFC vector magnetograph

    NASA Astrophysics Data System (ADS)

    Hagyard, M. J.; Cumings, N. P.; West, E. A.

    1981-02-01

    The NASA/Marshall Space Flight Center's solar vector magnetograph system allows measurements of all components of the Sun's photospheric magnetic field over a 5 x 5 or 2.5 x 2.5 arc min square field of view with an optimum time resolution of approximately 100 sec and an optimum signal-to-noise of approximately 1000. The basic system components are described, including the optics, detector, digital system, and associated electronics. Automatic sequencing and control functions are outlined as well as manual selections of system parameters which afford unique system flexibility. Results of system calibration and performance are presented, including linearity, dynamic range, uniformity, spatial and spectral resolutions, signal-to-noise, electro-optical retardation and polarization calibration.

  16. Multistage vector (MSV) therapeutics.

    PubMed

    Wolfram, Joy; Shen, Haifa; Ferrari, Mauro

    2015-12-10

    One of the greatest challenges in the field of medicine is obtaining controlled distribution of systemically administered therapeutic agents within the body. Indeed, biological barriers such as physical compartmentalization, pressure gradients, and excretion pathways adversely affect localized delivery of drugs to pathological tissue. The diverse nature of these barriers requires the use of multifunctional drug delivery vehicles that can overcome a wide range of sequential obstacles. In this review, we explore the role of multifunctionality in nanomedicine by primarily focusing on multistage vectors (MSVs). The MSV is an example of a promising therapeutic platform that incorporates several components, including a microparticle, nanoparticles, and small molecules. In particular, these components are activated in a sequential manner in order to successively address transport barriers. PMID:26264836

  17. Construction Technologies.

    ERIC Educational Resources Information Center

    Columbus State Community Coll., OH.

    This document contains materials developed for and about the construction technologies tech prep program of the South-Western City Schools in Ohio. Part 1 begins with a map of the program, which begins with a construction technologies program in grades 11 and 12 that leads to entry-level employment or one of five 2-year programs at a community…

  18. Construction Management.

    ERIC Educational Resources Information Center

    Abbott, James F.

    This article provides a detailed discussion of a team approach to building that involves a construction manager, an architect, and a contractor. Bidding methods are outlined; the major components in construction management -- value engineering and fast track scheduling -- and the use of performance specifications are discussed. The construction…

  19. Gene Composer: database software for protein construct design, codon engineering, and gene synthesis

    PubMed Central

    Lorimer, Don; Raymond, Amy; Walchli, John; Mixon, Mark; Barrow, Adrienne; Wallace, Ellen; Grice, Rena; Burgin, Alex; Stewart, Lance

    2009-01-01

    Background To improve efficiency in high throughput protein structure determination, we have developed a database software package, Gene Composer, which facilitates the information-rich design of protein constructs and their codon engineered synthetic gene sequences. With its modular workflow design and numerous graphical user interfaces, Gene Composer enables researchers to perform all common bio-informatics steps used in modern structure guided protein engineering and synthetic gene engineering. Results An interactive Alignment Viewer allows the researcher to simultaneously visualize sequence conservation in the context of known protein secondary structure, ligand contacts, water contacts, crystal contacts, B-factors, solvent accessible area, residue property type and several other useful property views. The Construct Design Module enables the facile design of novel protein constructs with altered N- and C-termini, internal insertions or deletions, point mutations, and desired affinity tags. The modifications can be combined and permuted into multiple protein constructs, and then virtually cloned in silico into defined expression vectors. The Gene Design Module uses a protein-to-gene algorithm that automates the back-translation of a protein amino acid sequence into a codon engineered nucleic acid gene sequence according to a selected codon usage table with minimal codon usage threshold, defined G:C% content, and desired sequence features achieved through synonymous codon selection that is optimized for the intended expression system. The gene-to-oligo algorithm of the Gene Design Module plans out all of the required overlapping oligonucleotides and mutagenic primers needed to synthesize the desired gene constructs by PCR, and for physically cloning them into selected vectors by the most popular subcloning strategies. Conclusion We present a complete description of Gene Composer functionality, and an efficient PCR-based synthetic gene assembly procedure with mis

  20. Traditional and robust vector selection methods for use with similarity based models

    SciTech Connect

    Hines, J. W.; Garvey, D. R.

    2006-07-01

    Vector selection, or instance selection as it is often called in the data mining literature, performs a critical task in the development of nonparametric, similarity based models. Nonparametric, similarity based modeling (SBM) is a form of 'lazy learning' which constructs a local model 'on the fly' by comparing a query vector to historical, training vectors. For large training sets the creation of local models may become cumbersome, since each training vector must be compared to the query vector. To alleviate this computational burden, varying forms of training vector sampling may be employed with the goal of selecting a subset of the training data such that the samples are representative of the underlying process. This paper describes one such SBM, namely auto-associative kernel regression (AAKR), and presents five traditional vector selection methods and one robust vector selection method that may be used to select prototype vectors from a larger data set in model training. The five traditional vector selection methods considered are min-max, vector ordering, combination min-max and vector ordering, fuzzy c-means clustering, and Adeli-Hung clustering. Each method is described in detail and compared using artificially generated data and data collected from the steam system of an operating nuclear power plant. (authors)

  1. Pattern vectors from algebraic graph theory.

    PubMed

    Wilson, Richard C; Hancock, Edwin R; Luo, Bin

    2005-07-01

    Graph structures have proven computationally cumbersome for pattern analysis. The reason for this is that, before graphs can be converted to pattern vectors, correspondences must be established between the nodes of structures which are potentially of different size. To overcome this problem, in this paper, we turn to the spectral decomposition of the Laplacian matrix. We show how the elements of the spectral matrix for the Laplacian can be used to construct symmetric polynomials that are permutation invariants. The coefficients of these polynomials can be used as graph features which can be encoded in a vectorial manner. We extend this representation to graphs in which there are unary attributes on the nodes and binary attributes on the edges by using the spectral decomposition of a Hermitian property matrix that can be viewed as a complex analogue of the Laplacian. To embed the graphs in a pattern space, we explore whether the vectors of invariants can be embedded in a low-dimensional space using a number of alternative strategies, including principal components analysis (PCA), multidimensional scaling (MDS), and locality preserving projection (LPP). Experimentally, we demonstrate that the embeddings result in well-defined graph clusters. Our experiments with the spectral representation involve both synthetic and real-world data. The experiments with synthetic data demonstrate that the distances between spectral feature vectors can be used to discriminate between graphs on the basis of their structure. The real-world experiments show that the method can be used to locate clusters of graphs. PMID:16013758

  2. Partial transfer entropy on rank vectors

    NASA Astrophysics Data System (ADS)

    Kugiumtzis, D.

    2013-06-01

    For the evaluation of information flow in bivariate time series, information measures have been employed, such as the transfer entropy (TE), the symbolic transfer entropy (STE), defined similarly to TE but on the ranks of the components of the reconstructed vectors, and the transfer entropy on rank vectors (TERV), similar to STE but forming the ranks for the future samples of the response system with regard to the current reconstructed vector. Here we extend TERV for multivariate time series, and account for the presence of confounding variables, called partial transfer entropy on ranks (PTERV). We investigate the asymptotic properties of PTERV, and also partial STE (PSTE), construct parametric significance tests under approximations with Gaussian and gamma null distributions, and show that the parametric tests cannot achieve the power of the randomization test using time-shifted surrogates. Using simulations on known coupled dynamical systems and applying parametric and randomization significance tests, we show that PTERV performs better than PSTE but worse than the partial transfer entropy (PTE). However, PTERV, unlike PTE, is robust to the presence of drifts in the time series and it is also not affected by the level of detrending.

  3. Hydrogen-Bond Directed Regioselective Pd-Catalyzed Asymmetric Allylic Alkylation: The Construction of Chiral α-Amino Acids with Vicinal Tertiary and Quaternary Stereocenters.

    PubMed

    Wei, Xuan; Liu, Delong; An, Qianjin; Zhang, Wanbin

    2015-12-01

    A Pd-catalyzed asymmetric allylic alkylation of azlactones with 4-arylvinyl-1,3-dioxolan-2-ones was developed, providing "branched" chiral α-amino acids with vicinal tertiary and quaternary stereocenters, in high yields and with excellent selectivities. Mechanistic studies revealed that the formation of a hydrogen bond between the Pd-allylic complex and azlactone isomer is responsible for the excellent regioselectivities. This asymmetric alkylation can be carried out on a gram scale without a loss of catalytic efficiency, and the resulting product can be further transformed to a chiral azetidine in two simple steps. PMID:26575242

  4. BioVector, a flexible system for gene specific-expression in plants

    PubMed Central

    2013-01-01

    Background Functional genomic research always needs to assemble different DNA fragments into a binary vector, so as to express genes with different tags from various promoters with different levels. The cloning systems available bear similar disadvantages, such as promoters/tags are fixed on a binary vector, which is generally with low cloning efficiency and limited for cloning sites if a novel promoter/tag is in need. Therefore, it is difficult both to assemble a gene and a promoter together and to modify the vectors in hand. Another disadvantage is that a long spacer from recombination sites, which may be detrimental to the protein function, exists between a gene and a tag. Multiple GATEWAY system only resolves former problem at the expense of very low efficiency and expensive for multiple LR reaction. Results To improve efficiency and flexibility for constructing expression vectors, we developed a platform, BioVector, by combining classical restriction enzyme/ligase strategy with modern Gateway DNA recombination system. This system included a series of vectors for gene cloning, promoter cloning, and binary vector construction to meet various needs for plant functional genomic study. Conclusion This BioVector platform makes it easy to construct any vectors to express a target gene from a specific promoter with desired intensity, and it is also waiting to be freely modified by researchers themselves for ongoing demands. This idea can also be transferred to the different fields including animal or yeast study. PMID:24304941

  5. Progressive Classification Using Support Vector Machines

    NASA Technical Reports Server (NTRS)

    Wagstaff, Kiri; Kocurek, Michael

    2009-01-01

    An algorithm for progressive classification of data, analogous to progressive rendering of images, makes it possible to compromise between speed and accuracy. This algorithm uses support vector machines (SVMs) to classify data. An SVM is a machine learning algorithm that builds a mathematical model of the desired classification concept by identifying the critical data points, called support vectors. Coarse approximations to the concept require only a few support vectors, while precise, highly accurate models require far more support vectors. Once the model has been constructed, the SVM can be applied to new observations. The cost of classifying a new observation is proportional to the number of support vectors in the model. When computational resources are limited, an SVM of the appropriate complexity can be produced. However, if the constraints are not known when the model is constructed, or if they can change over time, a method for adaptively responding to the current resource constraints is required. This capability is particularly relevant for spacecraft (or any other real-time systems) that perform onboard data analysis. The new algorithm enables the fast, interactive application of an SVM classifier to a new set of data. The classification process achieved by this algorithm is characterized as progressive because a coarse approximation to the true classification is generated rapidly and thereafter iteratively refined. The algorithm uses two SVMs: (1) a fast, approximate one and (2) slow, highly accurate one. New data are initially classified by the fast SVM, producing a baseline approximate classification. For each classified data point, the algorithm calculates a confidence index that indicates the likelihood that it was classified correctly in the first pass. Next, the data points are sorted by their confidence indices and progressively reclassified by the slower, more accurate SVM, starting with the items most likely to be incorrectly classified. The user

  6. pH-Dependent two novel heteronuclear Cu(II)/Sr(II) coordination polymers constructed from 1,3,5-benzenetricarboxylic acid: Synthesis, crystal structures and properties

    NASA Astrophysics Data System (ADS)

    Sun, Qiao-Zhen; Yin, Yi-Biao; Pan, Jun-Qiao; Chai, Li-Yuan; Su, Nan; Liu, Hui; Zhao, Yi-Lin; Liu, Xing-Tao

    2016-02-01

    Two novel heteronuclear coordination polymers, namely, [CuSr2(BTC)2]·10H2O (1) and [Cu2Sr(H4TMA)2]·4H2O (2) (H3BTC = 1,3,5-benzenetricarboxylic acid, H4TMA = 2-hydroxytrimesic acid) were hydrothermally synthesized as pH-dependent products and characterized by elemental analysis (EA), infrared spectroscopy (IR) and single crystal X-ray diffraction. For compound 1, it displays a 3D structure with (2,5,6)-connected net topology. For 2, the H3BTC ligand is oxidized into H4TMA and compound 2 features a 2D layer structure, which is further linked by Cu⋯Cu and Cu⋯O supramolecular interactions into a 3D structure. The results show that the pH plays a crucial role in determining the structure of the compounds. In addition, thermalgravimetric analysis of compounds 1-2 and luminescence property of 1 are also investigated.

  7. PQChPT with Staggered Sea and Valence Ginsparg-Wilson Quarks: Vector Meson Masses

    SciTech Connect

    Hovhannes R. Grigoryan; Anthony W. Thomas

    2005-09-16

    We consider partially quenched, mixed chiral perturbation theory with staggered sea and Ginsparg-Wilson valence quarks in order to extract a chiral-continuum extrapolation expression for the vector meson mass up to order O(a{sup 2}), at one-loop level. Based on general principles, we accomplish the task without explicitly constructing a sophisticated, heavy vector meson chiral Lagrangian.

  8. Insect cell transformation vectors that support high level expression and promoter assessment in insect cell culture

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A somatic transformation vector, pDP9, was constructed that provides a simplified means of producing permanently transformed cultured insect cells that support high levels of protein expression of foreign genes. The pDP9 plasmid vector incorporates DNA sequences from the Junonia coenia densovirus th...

  9. Bipolar battery construction

    NASA Technical Reports Server (NTRS)

    Rippel, Wally E. (Inventor); Edwards, Dean B. (Inventor)

    1981-01-01

    A lightweight, bipolar battery construction for lead acid batteries in which a plurality of thin, rigid, biplates each comprise a graphite fiber thermoplastic composition in conductive relation to lead stripes plated on opposite flat surfaces of the plates, and wherein a plurality of nonconductive thermoplastic separator plates support resilient yieldable porous glass mats in which active material is carried, the biplates and separator plates with active material being contained and maintained in stacked assembly by axial compression of the stacked assembly. A method of assembling such a bipolar battery construction.

  10. Defective herpes simplex virus type 1 vectors harboring gag, pol, and env genes can be used to rescue defective retrovirus vectors.

    PubMed Central

    Savard, N; Cosset, F L; Epstein, A L

    1997-01-01

    A retroviral packaging transcription unit was constructed in which the Moloney murine leukemia virus (MoMLV) gag-pol and env genes are expressed under the control of herpesvirus regulatory sequences. This transcription unit, lacking long terminal repeats, primer binding sites, and most of the retrovirus packaging signal but retaining both retroviral donor and acceptor splice sites, was cloned into a herpes simplex virus type 1 (HSV-1) amplicon plasmid, and amplicon vectors (the gag-pol-env [GPE] vectors) were generated by using a defective HSV-1 vector as helper virus. The GPE vector population was used to infect human TE671 cells (ATCC CRL 8805), harboring a lacZ provirus (TE-lac2 cells), and supernatants of infected cells were collected and filtered at different times after infection. These supernatants were found to contain infectious ecotropic lacZ retroviral particles, as shown both by reverse transcription-PCR and by their ability to transduce a beta-galactosidase activity to murine NIH 3T3 cells but not to human TE671 cells. The titer of retroviral vectors released by GPE vector-infected TE-lac2 cells increased with the dose of infectious amplicon particles. Retrovirus vector production was inhibited by superinfection with helper virus, indicating that helper virus coinfection negatively interfered with retrovirus production. Induction of retrovirus vectors by GPE vectors was neutralized by anti-HSV-1 but not by anti-MoMLV antiserum, while transduction of beta-galactosidase activity to NIH 3T3 cells by supernatants of GPE vector-infected TE-lac2 cells was neutralized by anti-MoMLV antiserum. These results demonstrate that HSV-1 GPE amplicon vectors can rescue defective lacZ retrovirus vectors and suggest that they could be used as a sort of launching ramp to fire defective retrovirus vectors from within virtually any in vitro or in vivo cell type containing defective retroviral vectors. PMID:9094692

  11. Two coordination polymers constructed from a multidentate carboxylic acid ligand with a tertiary amine serve as acid-base catalysts for the synthesis of chloropropene carbonate from CO2 under atmospheric pressure.

    PubMed

    Chen, Chao; Zhang, Jun; Li, Guanghua; Shen, Pan; Jin, Haichao; Zhang, Ning

    2014-10-01

    Two new coordination polymers, [Ni(H2O)(Hpdcd)(H2O)2]·DMF (1) and [Co(H2O)(Hpdcd)(H2O)2]·DMF (2) (H3pdcd = 1-(4-carboxyphenyl)-2,5-dimethyl, 1H-pyrrole-3,4-dicarboxylic acid), which were designed based on a tertiary amine ligand, were synthesized and characterized using multiple spectroscopy techniques, including single-crystal X-ray diffraction. These two 1D linear chains possess the properties of both a Lewis acid and organic base, which was confirmed by temperature programmed desorption of ammonia and on-line mass spectrometry (NH3-TPD-MS), and selective sorption for carbon dioxide. Due to their acid-base properties, the compounds exhibited high catalytic activity, in the absence of co-catalysts, for solvent-free synthesis of chloropropene carbonate from CO2 and epichlorohydrin under atmospheric CO2 pressure. The yields of chloropropene carbonate were 88% and 87% for 1 and 2, respectively, under the optimized conditions. PMID:25113602

  12. Efficient morse decompositions of vector fields.

    PubMed

    Chen, Guoning; Mischaikow, Konstantin; Laramee, Robert S; Zhang, Eugene

    2008-01-01

    Existing topology-based vector field analysis techniques rely on the ability to extract the individual trajectories such as fixed points, periodic orbits, and separatrices that are sensitive to noise and errors introduced by simulation and interpolation. This can make such vector field analysis unsuitable for rigorous interpretations. We advocate the use of Morse decompositions, which are robust with respect to perturbations, to encode the topological structures of a vector field in the form of a directed graph, called a Morse connection graph (MCG). While an MCG exists for every vector field, it need not be unique. Previous techniques for computing MCG's, while fast, are overly conservative and usually results in MCG's that are too coarse to be useful for the applications. To address this issue, we present a new technique for performing Morse decomposition based on the concept of tau-maps, which typically provides finer MCG's than existing techniques. Furthermore, the choice of tau provides a natural tradeoff between the fineness of the MCG's and the computational costs. We provide efficient implementations of Morse decomposition based on tau-maps, which include the use of forward and backward mapping techniques and an adaptive approach in constructing better approximations of the images of the triangles in the meshes used for simulation.. Furthermore, we propose the use of spatial tau-maps in addition to the original temporal tau-maps. These techniques provide additional trade-offs between the quality of the MCGs and the speed of computation. We demonstrate the utility of our technique with various examples in the plane and on surfaces including engine simulation data sets. PMID:18467759

  13. Vectors on the Basketball Court

    ERIC Educational Resources Information Center

    Bergman, Daniel

    2010-01-01

    An Idea Bank published in the April/May 2009 issue of "The Science Teacher" describes an experiential physics lesson on vectors and vector addition (Brown 2009). Like its football predecessor, the basketball-based investigation presented in this Idea Bank addresses National Science Education Standards Content B, Physical Science, 9-12 (NRC 1996)…

  14. GPU Accelerated Vector Median Filter

    NASA Technical Reports Server (NTRS)

    Aras, Rifat; Shen, Yuzhong

    2011-01-01

    Noise reduction is an important step for most image processing tasks. For three channel color images, a widely used technique is vector median filter in which color values of pixels are treated as 3-component vectors. Vector median filters are computationally expensive; for a window size of n x n, each of the n(sup 2) vectors has to be compared with other n(sup 2) - 1 vectors in distances. General purpose computation on graphics processing units (GPUs) is the paradigm of utilizing high-performance many-core GPU architectures for computation tasks that are normally handled by CPUs. In this work. NVIDIA's Compute Unified Device Architecture (CUDA) paradigm is used to accelerate vector median filtering. which has to the best of our knowledge never been done before. The performance of GPU accelerated vector median filter is compared to that of the CPU and MPI-based versions for different image and window sizes, Initial findings of the study showed 100x improvement of performance of vector median filter implementation on GPUs over CPU implementations and further speed-up is expected after more extensive optimizations of the GPU algorithm .

  15. Insect E-probe Diagnostic Nucleic acid Analysis (EDNA): the application of a novel bioinformatic tool to detection of vectors and pathogens in individual insect and simulated insect trap metagenomes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plant pathogen detection takes many forms. In simple cases, researchers are attempting to detect a known pathogen from a known host utilizing targeted nucleic acid or antigenic assays. However, in more complex scenarios researchers may not know the identity of a pathogen, or they may need to screen ...

  16. Method of increasing conversion of a fatty acid to its corresponding dicarboxylic acid

    DOEpatents

    Craft, David L.; Wilson, C. Ron; Eirich, Dudley; Zhang, Yeyan

    2004-09-14

    A nucleic acid sequence including a CYP promoter operably linked to nucleic acid encoding a heterologous protein is provided to increase transcription of the nucleic acid. Expression vectors and host cells containing the nucleic acid sequence are also provided. The methods and compositions described herein are especially useful in the production of polycarboxylic acids by yeast cells.

  17. Construction aggregates

    USGS Publications Warehouse

    Bolen, W.P.; Tepordei, V.V.

    2001-01-01

    The estimated production during 2000 of construction aggregates, crushed stone, and construction sand and gravel increased by about 2.6% to 2.7 Gt (3 billion st), compared with 1999. The expansion that started in 1992 continued with record production levels for the ninth consecutive year. By commodity, construction sand and gravel production increased by 4.5% to 1.16 Gt (1.28 billion st), while crushed stone production increased by 1.3% to 1.56 Gt (1.72 billion st).

  18. Construction of two Cd(II) complexes by flexible adipic acid plus 2-((benzoimidazol-yl)methyl)-1H-tetrazole ligand

    NASA Astrophysics Data System (ADS)

    Duan, Wanlu; Zhang, Yuhong; Wang, Xiuxiu; Meng, Xiangru

    2015-10-01

    Two new complexes with the formulas [Cd(bimt)(adi)]n (1) and {[Cd(bimt)(adi)0.5Br]·H2O}n (2) were synthesized through reactions of 2-((benzoimidazol-yl)methyl)-1H-tetrazole (bimt) with Cd(II) salts in the presence of adipic acid (H2adi). Single crystal X-ray analysis reveals that complex 1 shows a 1D chain structure in which adipate ligand coordinates to Cd(II) ions with μ3-bridging mode. Complex 2 displays a 2D layer structure with 4-connected (44·62) topology in which adipate ligand coordinates to Cd(II) ions with μ2-bridging mode. These results reveal that the versatile coordination modes of adipate ligands play an important role in controlling the structures of the complexes. In addition, their IR spectra, element analyses, PXRD patterns and luminescent properties are investigated.

  19. Divergence-based vector quantization.

    PubMed

    Villmann, Thomas; Haase, Sven

    2011-05-01

    Supervised and unsupervised vector quantization methods for classification and clustering traditionally use dissimilarities, frequently taken as Euclidean distances. In this article, we investigate the applicability of divergences instead, focusing on online learning. We deduce the mathematical fundamentals for its utilization in gradient-based online vector quantization algorithms. It bears on the generalized derivatives of the divergences known as Fréchet derivatives in functional analysis, which reduces in finite-dimensional problems to partial derivatives in a natural way. We demonstrate the application of this methodology for widely applied supervised and unsupervised online vector quantization schemes, including self-organizing maps, neural gas, and learning vector quantization. Additionally, principles for hyperparameter optimization and relevance learning for parameterized divergences in the case of supervised vector quantization are given to achieve improved classification accuracy. PMID:21299418

  20. Rice Reoviruses in Insect Vectors.

    PubMed

    Wei, Taiyun; Li, Yi

    2016-08-01

    Rice reoviruses, transmitted by leafhopper or planthopper vectors in a persistent propagative manner, seriously threaten the stability of rice production in Asia. Understanding the mechanisms that enable viral transmission by insect vectors is a key to controlling these viral diseases. This review describes current understanding of replication cycles of rice reoviruses in vector cell lines, transmission barriers, and molecular determinants of vector competence and persistent infection. Despite recent breakthroughs, such as the discoveries of actin-based tubule motility exploited by viruses to overcome transmission barriers and mutually beneficial relationships between viruses and bacterial symbionts, there are still many gaps in our knowledge of transmission mechanisms. Advances in genome sequencing, reverse genetics systems, and molecular technologies will help to address these problems. Investigating the multiple interaction systems among the virus, insect vector, insect symbiont, and plant during natural infection in the field is a central topic for future research on rice reoviruses. PMID:27296147

  1. A neural support vector machine.

    PubMed

    Jändel, Magnus

    2010-06-01

    Support vector machines are state-of-the-art pattern recognition algorithms that are well founded in optimization and generalization theory but not obviously applicable to the brain. This paper presents Bio-SVM, a biologically feasible support vector machine. An unstable associative memory oscillates between support vectors and interacts with a feed-forward classification pathway. Kernel neurons blend support vectors and sensory input. Downstream temporal integration generates the classification. Instant learning of surprising events and off-line tuning of support vector weights trains the system. Emotion-based learning, forgetting trivia, sleep and brain oscillations are phenomena that agree with the Bio-SVM model. A mapping to the olfactory system is suggested. PMID:20092978

  2. Strategies for targeting lentiviral vectors.

    PubMed

    Frecha, Cecilia; Szécsi, Judit; Cosset, Francois-Loîc; Verhoeyen, Els

    2008-12-01

    Vectors derived from retroviruses such as lentiviruses and onco-retroviruses are probably among the most suitable tools to achieve a long-term gene transfer since they allow stable integration of a transgene and its propagation in daughter cells. Lentiviral vectors should be preferred gene delivery vehicles over vectors derived from onco-retroviruses (MLV) since in contrast to the latter they can transduce non-proliferating target cells. Moreover, lentiviral vectors that have the capacity to deliver transgenes into specific tissues are expected to be of great value for various gene transfer approaches in vivo. Here we provide an overview of innovative approaches to upgrade lentiviral vectors for tissue or cell targeting and which have potential for in vivo gene delivery. In this overview we distinguish between three types of lentiviral vector targeting strategies (Fig 1): 1) targeting of vectors at the level of vector-cell entry through lentiviral vector surface modifications; 2) targeting at the level of transgene transcription by insertion of tissue specific promoters into lentiviral vectors; 3) a novel microRNA technology that rather than targeting the 'right' cells will 'detarget' transgene expression from non-target cells while achieving high expression in the target-cell. It is clear that each strategy is of enormous value for several gene therapy approaches but combining these three layers of transgene expression control will offer tools to really overcome several drawbacks in the field such as side-effect of off-target expression, clearance of transgene modified cells by immune response to the transgene and lack of biosecurity and efficiency in in vivo approaches. PMID:19075628

  3. Constructing Phylogenies.

    ERIC Educational Resources Information Center

    Bilardello, Nicholas; Valdes, Linda

    1998-01-01

    Introduces a method for constructing phylogenies using molecular traits and elementary graph theory. Discusses analyzing molecular data and using weighted graphs, minimum-weight spanning trees, and rooted cube phylogenies to display the data. (DDR)

  4. Construction Administration.

    ERIC Educational Resources Information Center

    Barley, John McKim, II

    1986-01-01

    Successful completion of a construction project requires the efforts of a team composed of the owner, architect, and contractor. A preconstruction conference can clarify the roles of the team as specified in the design contract. (MLF)

  5. Worldwide construction

    SciTech Connect

    Williamson, M.

    1994-10-17

    The paper lists major construction projects in worldwide processing and pipelining, showing capacities, contractors, estimated costs, and time of construction. The lists are divided into refineries, petrochemical plants, sulfur recovery units, gas processing plants, pipelines, and related fuel facilities. This last classification includes cogeneration plants, coal liquefaction and gasification plants, biomass power plants, geothermal power plants, integrated coal gasification combined-cycle power plants, and a coal briquetting plant.

  6. Construction aggregates

    USGS Publications Warehouse

    Tepordei, V.V.

    1993-01-01

    Part of a special section on the market performance of industrial minerals in 1992. Production of construction aggregates increased by 4.6 percent in 1992. This increase was due, in part, to the increased funding for transportation and infrastructure projects. The U.S. produced about 1.05 Gt of crushed stone and an estimated 734 Mt of construction sand and gravel in 1992. Demand is expected to increase by about 5 percent in 1993.

  7. Construction aggregates

    USGS Publications Warehouse

    Nelson, T.I.; Bolen, W.P.

    2007-01-01

    Construction aggregates, primarily stone, sand and gravel, are recovered from widespread naturally occurring mineral deposits and processed for use primarily in the construction industry. They are mined, crushed, sorted by size and sold loose or combined with portland cement or asphaltic cement to make concrete products to build roads, houses, buildings, and other structures. Much smaller quantities are used in agriculture, cement manufacture, chemical and metallurgical processes, glass production and many other products.

  8. Space Construction

    NASA Technical Reports Server (NTRS)

    Hagaman, Jane A. (Editor)

    1987-01-01

    The purpose was to present to the aerospace community an in-depth review of Experimental Assembly of Structures on EVA (EASE)/Assembly Concept for Construction of Erectable Space Structures (ACCESS) space flight experiments and to present the status of activities regarding future space flight experiments and accompanying technology developments that will demonstrate the capability of on-orbit construction required for the Space Station.

  9. Vector Network Analysis

    1997-10-20

    Vector network analyzers are a convenient way to measure scattering parameters of a variety of microwave devices. However, these instruments, unlike oscilloscopes for example, require a relatively high degree of user knowledge and expertise. Due to the complexity of the instrument and of the calibration process, there are many ways in which an incorrect measurement may be produced. The Microwave Project, which is part of Sandia National Laboratories Primary Standards Laboratory, routinely uses check standardsmore » to verify that the network analyzer is operating properly. In the past, these measurements were recorded manually and, sometimes, interpretation of the results was problematic. To aid our measurement assurance process, a software program was developed to automatically measure a check standard and compare the new measurements with an historical database of measurements of the same device. The program acquires new measurement data from selected check standards, plots the new data against the mean and standard deviation of prior data for the same check standard, and updates the database files for the check standard. The program is entirely menu-driven requiring little additional work by the user.« less

  10. Vector-vector production in photon-photon interactions

    SciTech Connect

    Ronan, M.T.

    1988-12-09

    Measurements of exclusive untagged /rho//sup 0//rho//sup 0/, /rho//phi/, K/sup *//bar K//sup */, and /rho/..omega.. production and tagged /rho//sup 0//rho//sup 0/ production in photon-photon interactions by the TPC/Two-Gamma experiment are reviewed. Comparisons to the results of other experiments and to models of vector-vector production are made. Fits to the data following a four quark model prescription for vector meson pair production are also presented. 10 refs., 9 figs.

  11. Vector-vector production in photon-photon interactions

    SciTech Connect

    Ronan, M. T.

    1989-04-25

    Measurements of exclusive untagged /rho//sup 0//rho0/,/rho//phi/,/ital K//sup *//ital K/bar /*/, and /rho/..omega.. production and tagged /rho//sup 0//rho0/ production in photon-photon interactions by the TPC/Two-Gamma experiment are reviewed. Comparisons to the results of other experiments and to models of vector-vector production are made. Fits to the data following a four quark model prescription for vector meson pair production are also presented.

  12. Selection of Novel Vesicular Stomatitis Virus Glycoprotein Variants from a Peptide Insertion Library for Enhanced Purification of Retroviral and Lentiviral Vectors

    PubMed Central

    Yu, Julie H.; Schaffer, David V.

    2006-01-01

    The introduction of new features or functions that are not present in an original protein is a significant challenge in protein engineering. For example, modifications to vesicular stomatitis virus glycoprotein (VSV-G), which is commonly used to pseudotype retroviral and lentiviral vectors for gene delivery, have been hindered by a lack of structural knowledge of the protein. We have developed a transposon-based approach that randomly incorporates designed polypeptides throughout a protein to generate saturated insertion libraries and a subsequent high-throughput selection process in mammalian cells that enables the identification of optimal insertion sites for a novel designed functionality. This method was applied to VSV-G in order to construct a comprehensive library of mutants whose combined members have a His6 tag inserted at likely every site in the original protein sequence. Selecting the library via iterative retroviral infections of mammalian cells led to the identification of several VSV-G-His6 variants that were able to package high-titer viral vectors and could be purified by Ni-nitrilotriacetic acid affinity chromatography. Column purification of vectors reduced protein and DNA impurities more than 5,000-fold and 14,000-fold, respectively, from the viral supernatant. This substantially improved purity elicited a weaker immune response in the brain, without altering the infectivity or tropism from wild-type VSV-G-pseudotyped vectors. This work applies a powerful new tool for protein engineering to construct novel viral envelope variants that can greatly improve the safety and use of retroviral and lentiviral vectors for clinical gene therapy. Furthermore, this approach of library generation and selection can readily be extended to other challenges in protein engineering. PMID:16537595

  13. AAV Vectors Expressing LDLR Gain-of-Function Variants Demonstrate Increased Efficacy in Mouse Models of Familial Hypercholesterolemia

    PubMed Central

    Somanathan, Suryanarayan; Jacobs, Frank; Wang, Qiang; Hanlon, Alexandra L; Wilson, James M; Rader, Daniel J

    2014-01-01

    Rationale Familial hypercholesterolemia (FH) is a genetic disorder that arises due to loss-of-function mutations in the low-density lipoprotein receptor (LDLR) and homozygous FH (hoFH) is a candidate for gene therapy using adeno-associated viral (AAV) vectors. Proprotein convertase subtilisin/kexin type 9 (PCSK9) and inducible degrader of LDLR (IDOL) negatively regulate LDLR protein and could dampen AAV encoded LDLR expression. Objective We sought to create vectors expressing gain-of-function human LDLR variants that are resistant to degradation by human PCSK9 and IDOL and thereby enhance hepatic LDLR protein abundance and plasma LDL cholesterol reduction. Methods and Results Amino acid substitutions were introduced into the coding sequence of human LDLR cDNA to reduce interaction with hPCSK9 and hIDOL. A panel of mutant hLDLRs was initially screened in vitro for escape from PCSK9. The variant hLDLR-L318D was further evaluated using a mouse model of hoFH lacking endogenous LDLR and apolipoprotein B mRNA editing enzyme, APOBEC-1 (DKO). Administration of wild type hLDLR to DKO mice, expressing hPCSK9, led to diminished LDLR activity. However, LDLR-L318D was resistant to hPCSK9 mediated degradation and effectively reduced cholesterol levels. Similarly, the LDLR-K809R\\C818A construct avoided hIDOL regulation and achieved stable reductions in serum cholesterol. An AAV8.LDLR-L318D\\K809R\\C818A vector that carried all three amino acid substitutions conferred partial resistance to both hPCSK9 and hIDOL mediated degradation. Conclusion Amino acid substitutions in the human LDLR confer partial resistance to PCSK9 and IDOL regulatory pathways with improved reduction in cholesterol levels and improve upon a potential gene therapeutic approach to treat homozygous FH subjects. PMID:25023731

  14. 3,4,9,10-perylenetetracarboxylic acid/hemin nanocomposites act as redox probes and electrocatalysts for constructing a pseudobienzyme-channeling amplified electrochemical aptasensor.

    PubMed

    Yuan, Yali; Yuan, Ruo; Chai, Yaqin; Zhuo, Ying; Gan, Xianxue; Bai, Lijuan

    2012-10-29

    A simple wet-chemical strategy for the synthesis of 3,4,9,10-perylenetetracarboxylic acid (PTCA)/hemin nanocomposites through π-π interactions is demonstrated. Significantly, the hemin successfully conciliates PTCA redox activity with a pair of well-defined redox peaks and intrinsic peroxidase-like activity, which provides potential application of the PTCA self-derived redox activity as redox probes. Additionally, PTCA/hemin nanocomposites exhibit a good membrane-forming property, which not only avoids the conventional fussy process for redox probe immobilization, but also reduces the participation of the membrane materials that act as a barrier of electron transfer. On the basis of these unique properties, a pseudobienzyme-channeling amplified electrochemical aptasensor is developed that is coupled with glucose oxidase (GOx) for thrombin detection by using PTCA/hemin nanocomposites as redox probes and electrocatalysts. With the addition of glucose to the electrolytic cell, the GOx on the aptasensor surface bioelectrocatalyzed the reduction of glucose to produce H(2)O(2), which in turn was electrocatalyzed by the PTCA/hemin nanocomposites. Cascade schemes, in which an enzyme is catalytically linked to another enzyme, can produce signal amplification and therefore increase the biosensor sensitivity. As a result, a linear relationship for thrombin from 0.005 to 20 nM and a detection limit of 0.001 nM were obtained. PMID:23001991

  15. Construction a magneto carbon paste electrode using synthesized molecularly imprinted magnetic nanospheres for selective and sensitive determination of mefenamic acid in some real samples.

    PubMed

    Madrakian, Tayyebeh; Haghshenas, Esmaeel; Ahmadi, Mazaher; Afkhami, Abbas

    2015-06-15

    A novel magneto multiwalled carbon nanotube/carbon paste electrode (MMW/CPE) for the determination of mefenamic acid (MFA) was introduced. Magnetic molecularly imprinted polymer nanoparticles (MMIPNPs) were synthesized and then added to the solution of MFA. After stirring for 20 min, the MMW/CPE was immersed in the solution of MFA (contain MMIPNPs) and the MMIPNPs were captured by it. Then oxidation of MFA was analyzed by differential pulse voltammetry (DPV). Electrochemical impedance spectroscopy, cyclic voltammetry, and DPV were employed to characterize the MMW/CPE. The MMIPNPs exhibited a high selectivity and sensitivity toward MFA. The effect of various experimental parameters including pH, MMIPNPs dosage, stirring time, accumulation potential and time on the voltammetric response of MFA were investigated. Under the optimal conditions, selective detection of MFA in a linear concentration range of 2.0-1000.0 nmol L(-1) was performed with the detection limit of 1.2 nmol L(-1) (3S/N). To further study the practical applicability of this method, it was applied to the analysis of some real samples and the obtained results were satisfactory. PMID:25679118

  16. Development of nitrilase promoter-derived inducible vectors for Streptomyces.

    PubMed

    Matsumoto, Masako; Hashimoto, Yoshiteru; Saitoh, Yuki; Kumano, Takuto; Kobayashi, Michihiko

    2016-06-01

    An inducible expression vector, pSH19, which harbors regulatory expression system PnitA-NitR, for streptomycetes was constructed previously. Here, we have modified pSH19 to obtain shuttle vectors for Streptomyces-E. coli by introducing the replication origin of a plasmid for E. coli (ColE1) and an antibiotic-resistant gene. Six inducible shuttle vectors, pESH19cF, pESH19cR, pESH19kF, pESH19kR, pESH19aF, and pESH19aR, for Streptomyces-E. coli, were successfully developed. The stability of these vectors was examined in five different E. coli strains and Streptomyces lividans TK24. The stability test showed that the pSH19-derived shuttle vectors were stable in E. coli Stbl2 and S. lividans TK24. Heterologous expression experiments involving each of the catechol 2,3-dioxygenase, nitrilase, and N-substituted formamide deformylase genes as a reporter gene showed that pESH19cF, pESH19kF, and pESH19aF possess inducible expression ability in S. lividans TK24. Thus, these vectors were found to be useful expression tools for experiments on both Gram-negative and Gram-positive bacterial genes. PMID:26923287

  17. Can MONDian vector theories explain the cosmic speed up?

    SciTech Connect

    Cardone, Vincenzo F.; Radicella, Ninfa

    2009-09-15

    Generalized Einstein-Aether vector field models have been shown to provide, in the weak field regime, modifications to gravity which can be reconciled with the successful modified Newtonian dynamics (MOND) proposal. Very little is known, however, on the function F(K) defining the vector field Lagrangian so that an analysis of the viability of such theories at the cosmological scales has never been performed. As a first step along this route, we rely on the relation between F(K) and the MOND interpolating function {mu}(a/a{sub 0}) to assign the vector field Lagrangian thus obtaining what we refer to as MONDian vector models. Since they are able by construction to recover the MOND successes on galaxy scales, we investigate whether they can also drive the observed accelerated expansion by fitting the models to the type Ia supernovae data. Should this be the case, we have a unified framework where both dark energy and dark matter can be seen as different manifestations of a single vector field. It turns out that both MONDian vector models are able to well fit the low redshift data on type Ia supernovae, while some tension could be present in the high z regime.

  18. Development of lentiviral vectors for antiangiogenic gene delivery.

    PubMed

    Shichinohe, T; Bochner, B H; Mizutani, K; Nishida, M; Hegerich-Gilliam, S; Naldini, L; Kasahara, N

    2001-11-01

    Growth and metastasis of malignant tumors requires angiogenesis. Inhibition of tumor-induced angiogenesis may represent an effective cytostatic strategy. We have constructed recombinant self-inactivating lentiviral vectors expressing angiostatin and endostatin, and have tested their antiangiogenic activities. As VSV-G-pseudotyped lentiviral vectors showed low relative transduction titers on bovine aortic and human umbilical vein endothelial cells, it was difficult to achieve significant inhibition of endothelial cell growth by lentivirus-mediated antiangiogenic gene transfer directly to endothelial cells without concomitant vector-associated cytotoxicity. However, lentivirus vectors could efficiently and stably transduce T24 human bladder cancer cells that are relatively resistant to adenovirus infection due to loss of coxsackievirus-adenovirus receptor expression. Long-term expression and secretion of angiostatin and endostatin from lentivirus-transduced T24 cells resulted in significant inhibition of cellular proliferation on coculture with endothelial cells. This report represents the first use of lentivirus-based vectors to deliver the antiangiogenic factors, angiostatin and endostatin, and suggests the potential utility of antiangiogenic gene therapy with lentiviral vectors for the treatment of cancer. PMID:11773978

  19. Chikungunya Virus–Vector Interactions

    PubMed Central

    Coffey, Lark L.; Failloux, Anna-Bella; Weaver, Scott C.

    2014-01-01

    Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes chikungunya fever, a severe, debilitating disease that often produces chronic arthralgia. Since 2004, CHIKV has emerged in Africa, Indian Ocean islands, Asia, Europe, and the Americas, causing millions of human infections. Central to understanding CHIKV emergence is knowledge of the natural ecology of transmission and vector infection dynamics. This review presents current understanding of CHIKV infection dynamics in mosquito vectors and its relationship to human disease emergence. The following topics are reviewed: CHIKV infection and vector life history traits including transmission cycles, genetic origins, distribution, emergence and spread, dispersal, vector competence, vector immunity and microbial interactions, and co-infection by CHIKV and other arboviruses. The genetics of vector susceptibility and host range changes, population heterogeneity and selection for the fittest viral genomes, dual host cycling and its impact on CHIKV adaptation, viral bottlenecks and intrahost diversity, and adaptive constraints on CHIKV evolution are also discussed. The potential for CHIKV re-emergence and expansion into new areas and prospects for prevention via vector control are also briefly reviewed. PMID:25421891

  20. Enhancing poxvirus vectors vaccine immunogenicity

    PubMed Central

    García-Arriaza, Juan; Esteban, Mariano

    2014-01-01

    Attenuated recombinant poxvirus vectors expressing heterologous antigens from pathogens are currently at various stages in clinical trials with the aim to establish their efficacy. This is because these vectors have shown excellent safety profiles, significant immunogenicity against foreign expressed antigens and are able to induce protective immune responses. In view of the limited efficacy triggered by some poxvirus strains used in clinical trials (i.e, ALVAC in the RV144 phase III clinical trial for HIV), and of the restrictive replication capacity of the highly attenuated vectors like MVA and NYVAC, there is a consensus that further improvements of these vectors should be pursuit. In this review we considered several strategies that are currently being implemented, as well as new approaches, to improve the immunogenicity of the poxvirus vectors. This includes heterologous prime/boost protocols, use of co-stimulatory molecules, deletion of viral immunomodulatory genes still present in the poxvirus genome, enhancing virus promoter strength, enhancing vector replication capacity, optimizing expression of foreign heterologous sequences, and the combined use of adjuvants. An optimized poxvirus vector triggering long-lasting immunity with a high protective efficacy against a selective disease should be sought. PMID:25424927

  1. Generation of RCAS vectors useful for functional genomic analyses.

    PubMed

    Loftus, S K; Larson, D M; Watkins-Chow, D; Church, D M; Pavan, W J

    2001-10-31

    Avian leukosis type A virus-derived retroviral vectors have been used to introduce genes into cells expressing the corresponding avian receptor tv-a. This includes the use of Replication-Competent Avian sarcoma-leukosis virus (ASLV) long terminal repeat (LTR) with Splice acceptor (RCAS) vectors in the analysis of avian development, human and murine cell cultures, murine cell lineage studies and cancer biology. Previously, cloning of genes into this virus was difficult due to the large size of the vector and sparse cloning sites. To overcome some of the disadvantages of traditional cloning using the RCASBP-Y vector, we have modified the RCASBP-Y to incorporate "Gateway" site-specific recombination cloning of genes into the construct, either with or without HA epitope tags. We have found the repetitive "att" sequences, which are the targets for site-specific recombination, do not impair the production of infectious viral particles or the expression of the gene of interest. This is the first instance of site-specific recombination being used to generate retroviral gene constructs. These viral constructs will allow for the efficient transfer and expression of cDNAs needed for functional genomic analyses. PMID:11759842

  2. Rapid identification of an antibody DNA construct rearrangement sequence variant by mass spectrometry.

    PubMed

    Scott, Rebecca A; Rogers, Rich; Balland, Alain; Brady, Lowell J

    2014-01-01

    During cell line development for an IgG1 antibody candidate (mAb1), a C-terminal extension was identified in 2 product candidate clones expressed in CHO-K1 cell line. The extension was initially observed as the presence of anomalous new peaks in these clones after analysis by cation exchange chromatography (CEX-HPLC) and reduced capillary electrophoresis (rCE-SDS). Reduced mass analysis of these CHO-K1 clones revealed that a larger than expected mass was present on a sub-population of the heavy chain species, which could not be explained by any known chemical or post-translational modifications. It was suspected that this additional mass on the heavy chain was due to the presence of an additional amino acid sequence. To identify the suspected additional sequence, de novo sequencing in combination with proteomic searching was performed against translated DNA vectors for the heavy chain and light chain. Peptides unique to the clones containing the extension were identified matching short sequences (corresponding to 9 and 35 amino acids, respectively) from 2 non-coding sections of the light chain vector construct. After investigation, this extension was observed to be due to the re-arrangement of the DNA construct, with the addition of amino acids derived from the light chain vector non-translated sequence to the C-terminus of the heavy chain. This observation showed the power of proteomic mass spectrometric techniques to identify an unexpected antibody sequence variant using de novo sequencing combined with database searching, and allowed for rapid identification of the root cause for new peaks in the cation exchange and rCE-SDS assays. PMID:25484040

  3. Emerging Vector-Borne Diseases - Incidence through Vectors.

    PubMed

    Savić, Sara; Vidić, Branka; Grgić, Zivoslav; Potkonjak, Aleksandar; Spasojevic, Ljubica

    2014-01-01

    Vector-borne diseases use to be a major public health concern only in tropical and subtropical areas, but today they are an emerging threat for the continental and developed countries also. Nowadays, in intercontinental countries, there is a struggle with emerging diseases, which have found their way to appear through vectors. Vector-borne zoonotic diseases occur when vectors, animal hosts, climate conditions, pathogens, and susceptible human population exist at the same time, at the same place. Global climate change is predicted to lead to an increase in vector-borne infectious diseases and disease outbreaks. It could affect the range and population of pathogens, host and vectors, transmission season, etc. Reliable surveillance for diseases that are most likely to emerge is required. Canine vector-borne diseases represent a complex group of diseases including anaplasmosis, babesiosis, bartonellosis, borreliosis, dirofilariosis, ehrlichiosis, and leishmaniosis. Some of these diseases cause serious clinical symptoms in dogs and some of them have a zoonotic potential with an effect to public health. It is expected from veterinarians in coordination with medical doctors to play a fundamental role at primarily prevention and then treatment of vector-borne diseases in dogs. The One Health concept has to be integrated into the struggle against emerging diseases. During a 4-year period, from 2009 to 2013, a total number of 551 dog samples were analyzed for vector-borne diseases (borreliosis, babesiosis, ehrlichiosis, anaplasmosis, dirofilariosis, and leishmaniasis) in routine laboratory work. The analysis was done by serological tests - ELISA for borreliosis, dirofilariosis, and leishmaniasis, modified Knott test for dirofilariosis, and blood smear for babesiosis, ehrlichiosis, and anaplasmosis. This number of samples represented 75% of total number of samples that were sent for analysis for different diseases in dogs. Annually, on average more then half of the samples

  4. CONSTRUCTION OF PLASMIDS FOR USE IN RISK ASSESSMENT RESEARCH

    EPA Science Inventory

    The report describes a series of selftransmissible and nonselftransmissible (cloning vector) plasmids constructed to compare results from different laboratory tests and plasmid systems. Plasmids were designed to overcome problems of reproducibility, confusion due to use of differ...

  5. Vector statistics of LANDSAT imagery

    NASA Technical Reports Server (NTRS)

    Jayroe, R. R., Jr.; Underwood, D.

    1977-01-01

    A digitized multispectral image, such as LANDSAT data, is composed of numerous four dimensional vectors, which quantitatively describe the ground scene from which the data are acquired. The statistics of unique vectors that occur in LANDSAT imagery are studied to determine if that information can provide some guidance on reducing image processing costs. A second purpose of this report is to investigate how the vector statistics are changed by various types of image processing techniques and determine if that information can be useful in choosing one processing approach over another.

  6. Baculovirus as a vaccine vector

    PubMed Central

    Lu, Hsin-Yu; Chen, Yi-Hsuan; Liu, Hung-Jen

    2012-01-01

    Baculovirus is extensively utilized as an excellent tool for production of recombinant protein in insect cells. Baculovirus infects insects in nature and is non-pathogenic to humans. In addition to insect cells, baculovirus is capable of transducing a broad range of animal cells. Due to its biosafety, large cloning capacity, low cytotoxicity, and non-replication nature in the transduced cells as well as the ease of manipulation and production, baculovirus has been utilized as RNA interference mediators, gene delivery vectors, and vaccine vectors for a wide variety of applications. This article focuses on the utilization of baculoviruses as vaccine vectors to prepare antigen or subunit vaccines. PMID:22705893

  7. Generation of Transgenic Porcine Fibroblast Cell Lines Using Nanomagnetic Gene Delivery Vectors.

    PubMed

    Grześkowiak, Bartosz F; Hryhorowicz, Magdalena; Tuśnio, Karol; Grzeszkowiak, Mikołaj; Załęski, Karol; Lipiński, Daniel; Zeyland, Joanna; Mykhaylyk, Olga; Słomski, Ryszard; Jurga, Stefan; Woźniak, Anna

    2016-05-01

    The transgenic process allows for obtaining genetically modified animals for divers biomedical applications. A number of transgenic animals for xenotransplantation have been generated with the somatic cell nuclear transfer (SCNT) method. Thereby, efficient nucleic acid delivery to donor cells such as fibroblasts is of particular importance. The objective of this study was to establish stable transgene expressing porcine fetal fibroblast cell lines using magnetic nanoparticle-based gene delivery vectors under a gradient magnetic field. Magnetic transfection complexes prepared by self-assembly of suitable magnetic nanoparticles, plasmid DNA, and an enhancer under an inhomogeneous magnetic field enabled the rapid and efficient delivery of a gene construct (pCD59-GFPBsd) into porcine fetal fibroblasts. The applied vector dose was magnetically sedimented on the cell surface within 30 min as visualized by fluorescence microscopy. The PCR and RT-PCR analysis confirmed not only the presence but also the expression of transgene in all magnetofected transgenic fibroblast cell lines which survived antibiotic selection. The cells were characterized by high survival rates and proliferative activities as well as correct chromosome number. The developed nanomagnetic gene delivery formulation proved to be an effective tool for the production of genetically engineered fibroblasts and may be used in future in SCNT techniques for breeding new transgenic animals for the purpose of xenotransplantation. PMID:27048425

  8. An approach to 1,3,4-dioxaphospholane complexes through an acid-induced ring expansion of an oxaphosphirane complex: the problem of construction and deconstruction of O,P-heterocycles.

    PubMed

    Pérez, Janaina Marinas; Helten, Holger; Schnakenburg, Gregor; Streubel, Rainer

    2011-06-01

    Treatment of oxaphosphirane complex 1, triflic acid (TfOH), and various aldehydes yielded 1,3,4-dioxaphospholane complexes 5a,b-7a,b after deprotonation with NEt(3). In addition to NMR spectroscopy, IR spectroscopy, and MS data, the X-ray structures of complexes 5a and 7a were determined. (31)P NMR spectroscopic monitoring and DFT calculations provided insight into the reaction course and revealed the transient TfOH 1,3,4-dioxaphospholanium association complex TfOH-5a,b and/or TfOH-5a,b' as key reactive intermediates. Furthermore, it was observed that the five-membered ring system was cleaved upon warming and yielded side-on (E,Z)-methylenephosphonium complexes 8a,b if deprotonation did not occur at low temperature. Overall, a novel temperature- and acid-dependent construction and deconstruction process of the 1,3,4-dioxaphospholane ring system is described. PMID:21433300

  9. Solid rocket thrust vector control

    NASA Technical Reports Server (NTRS)

    1974-01-01

    Thrust vector control systems that superimpose a side force on the motor thrust, steering being achieved by the side force causing a moment about the vehicle center of gravity are described. A brief review of thrust vector control systems is presented, and two systems, flexible joint and liquid injection, are treated in detail. Treatment of the flexible-joint thrust vector control system is limited to the design of the flexible joint and its insulation against hot motor gases. Treatment of the liquid injection thrust vector control system is limited to discussion of the injectant, valves, piping, storage tanks, and pressurization system; no evaluation is presented of the nozzle except for (1) the effect of the injectant and erosion at the injection port and (2) the effect of injection on pressure distribution within the nozzle.

  10. Experiments With Magnetic Vector Potential

    ERIC Educational Resources Information Center

    Skinner, J. W.

    1975-01-01

    Describes the experimental apparatus and method for the study of magnetic vector potential (MVP). Includes a discussion of inherent errors in the calculations involved, precision of the results, and further applications of MVP. (GS)

  11. Effective Masses of Vector Polarons

    NASA Astrophysics Data System (ADS)

    Foell, Charles; Clougherty, Dennis

    2006-03-01

    We consider the vector polarons of a one-dimensional model of an electron in a doubly (or nearly) degenerate band that couples to two elastic distortions, as described previously by Clougherty and Foell [1]. A variational approach is used to analytically and numerically calculate effective masses of the three types of vector polarons. [1] D. P. Clougherty and C. A. Foell, Phys. Rev. B 70, 052301 (2004).

  12. Coulomb problem for vector bosons

    SciTech Connect

    Kuchiev, M.Yu.; Flambaum, V.V.

    2006-05-01

    The Coulomb problem for vector bosons W{sup {+-}} incorporates a well-known difficulty; the charge of the boson localized in a close vicinity of the attractive Coulomb center proves to be infinite. The paradox is shown to be resolved by the QED vacuum polarization, which brings in a strong effective repulsion that eradicates the infinite charge of the boson on the Coulomb center. This property allows one to define the Coulomb problem for vector bosons properly.

  13. Molecular dynamics on vector computers

    NASA Astrophysics Data System (ADS)

    Sullivan, F.; Mountain, R. D.; Oconnell, J.

    1985-10-01

    An algorithm called the method of lights (MOL) has been developed for the computerized simulation of molecular dynamics. The MOL, implemented on the CYBER 205 computer, is based on sorting and reformulating the manner in which neighbor lists are compiled, and it uses data structures compatible with specialized vector statements that perform parallel computations. The MOL is found to reduce running time over standard methods in scalar form, and vectorization is shown to produce an order-of-magnitude reduction in execution time.

  14. Worldwide construction

    SciTech Connect

    Radler, M.

    1998-04-13

    Tables list major construction projects for refineries, petrochemical plants, sulfur plants, natural gas processing plants, and gas and oil pipelines. Data are compiled by country, company name, project type, added capacity, status of the project, expected completion date, contractor and contract type. Gas processes include LPG recovery, cryogenic separation, turboexpanders, LNG, liquefaction, desulfurization, NGL recovery, dehydration, hydrogen plants, and fractionators.

  15. Metal Construction

    NASA Technical Reports Server (NTRS)

    Verduzio, Rodolfo

    1922-01-01

    The future development of aerial navigation is closely connected with the condition of obtaining airplanes of great stability and sufficient strength. Different construction materials such as wood, aluminum, iron, and alloys are examined to determine which materials or combination of materials provides a greater coefficient of safety.

  16. Constructive Criticism.

    ERIC Educational Resources Information Center

    Lieberfeld, Lawrence

    1982-01-01

    Many crucial questions need to be answered before a college embarks on a construction project and makes a substantial financial commitment. Computer modeling techniques can be used to make even complex project feasibility analyses. Available from Peat, Marwick, Mitchell & Co., 345 Park Avenue, New York, NY 10154. (MSE)

  17. Abstract Constructions.

    ERIC Educational Resources Information Center

    Pietropola, Anne

    1998-01-01

    Describes a lesson designed to culminate a year of eighth-grade art classes in which students explore elements of design and space by creating 3-D abstract constructions. Outlines the process of using foam board and markers to create various shapes and optical effects. (DSK)

  18. Axisymmetric Coanda-assisted vectoring

    NASA Astrophysics Data System (ADS)

    Allen, Dustin; Smith, Barton L.

    2009-01-01

    An experimental demonstration of a jet vectoring technique used in our novel spray method called Coanda-assisted Spray Manipulation (CSM) is presented. CSM makes use of the Coanda effect on axisymmetric geometries through the interaction of two jets: a primary jet and a control jet. The primary jet has larger volume flow rate but generally a smaller momentum flux than the control jet. The primary jet flows through the center of a rounded collar. The control jet is parallel to the primary and is adjacent to the convex collar. The Reynolds number range for the primary jet at the exit plane was between 20,000 and 80,000. The flow was in the incompressible Mach number range (Mach < 0.3). The control jet attaches to the convex wall and vectors according to known Coanda effect principles, entraining and vectoring the primary jet, resulting in controllable r - θ directional spraying. Several annular control slots and collar radii were tested over a range of momentum flux ratios to determine the effects of these variables on the vectored jet angle and spreading. Two and Three-component Particle Image Velocimetry systems were used to determine the vectoring angle and the profile of the combined jet in each experiment. The experiments show that the control slot and expansion radius, along with the momentum ratios of the two jets predominantly affected the vectoring angle and profile of the combined jets.

  19. Vectoring of parallel synthetic jets

    NASA Astrophysics Data System (ADS)

    Berk, Tim; Ganapathisubramani, Bharathram; Gomit, Guillaume

    2015-11-01

    A pair of parallel synthetic jets can be vectored by applying a phase difference between the two driving signals. The resulting jet can be merged or bifurcated and either vectored towards the actuator leading in phase or the actuator lagging in phase. In the present study, the influence of phase difference and Strouhal number on the vectoring behaviour is examined experimentally. Phase-locked vorticity fields, measured using Particle Image Velocimetry (PIV), are used to track vortex pairs. The physical mechanisms that explain the diversity in vectoring behaviour are observed based on the vortex trajectories. For a fixed phase difference, the vectoring behaviour is shown to be primarily influenced by pinch-off time of vortex rings generated by the synthetic jets. Beyond a certain formation number, the pinch-off timescale becomes invariant. In this region, the vectoring behaviour is determined by the distance between subsequent vortex rings. We acknowledge the financial support from the European Research Council (ERC grant agreement no. 277472).

  20. Lactococci and lactobacilli as mucosal delivery vectors for therapeutic proteins and DNA vaccines

    PubMed Central

    2011-01-01

    Food-grade Lactic Acid Bacteria (LAB) have been safely consumed for centuries by humans in fermented foods. Thus, they are good candidates to develop novel oral vectors, constituting attractive alternatives to attenuated pathogens, for mucosal delivery strategies. Herein, this review summarizes our research, up until now, on the use of LAB as mucosal delivery vectors for therapeutic proteins and DNA vaccines. Most of our work has been based on the model LAB Lactococcus lactis, for which we have developed efficient genetic tools, including expression signals and host strains, for the heterologous expression of therapeutic proteins such as antigens, cytokines and enzymes. Resulting recombinant lactococci strains have been tested successfully for their prophylactic and therapeutic effects in different animal models: i) against human papillomavirus type 16 (HPV-16)-induced tumors in mice, ii) to partially prevent a bovine β-lactoglobulin (BLG)-allergic reaction in mice and iii) to regulate body weight and food consumption in obese mice. Strikingly, all of these tools have been successfully transposed to the Lactobacillus genus, in recent years, within our laboratory. Notably, anti-oxidative Lactobacillus casei strains were constructed and tested in two chemically-induced colitis models. In parallel, we also developed a strategy based on the use of L. lactis to deliver DNA at the mucosal level, and were able to show that L. lactis is able to modulate the host response through DNA delivery. Today, we consider that all of our consistent data, together with those obtained by other groups, demonstrate and reinforce the interest of using LAB, particularly lactococci and lactobacilli strains, to develop novel therapeutic protein mucosal delivery vectors which should be tested now in human clinical trials. PMID:21995317

  1. Efficacy of an adeno-associated virus 8-pseudotyped vector in glycogen storage disease type II.

    PubMed

    Sun, Baodong; Zhang, Haoyue; Franco, Luis M; Young, Sarah P; Schneider, Ayn; Bird, Andrew; Amalfitano, Andrea; Chen, Y-T; Koeberl, Dwight D

    2005-01-01

    Glycogen storage disease type II (GSD-II; Pompe disease) causes death in infancy from cardiorespiratory failure. The underlying deficiency of acid alpha-glucosidase (GAA; acid maltase) can be corrected by liver-targeted gene therapy in GSD-II, if secretion of GAA is accompanied by receptor-mediated uptake in cardiac and skeletal muscle. An adeno-associated virus (AAV) vector encoding human (h) GAA was pseudotyped as AAV8 (AAV2/8) and injected intravenously into immunodeficient GSD-II mice. High levels of hGAA were maintained in plasma for 24 weeks following AAV2/8 vector administration. A marked increase in vector copy number in the liver was demonstrated for the AAV2/8 vector compared to the analogous AAV2/2 vector. GAA deficiency in the heart and skeletal muscle was corrected with the AAV2/8 vector in male GSD-II mice, consistent with receptor-mediated uptake of hGAA. Male GSD-II mice demonstrated complete correction of glycogen storage in heart and diaphragm with the AAV2/8 vector, while female GSD-II mice had correction only in the heart. A biomarker for GSD-II was reduced in both sexes following AAV2/8 vector administration. Therefore, GAA production with an AAV2/8 vector in a depot organ, the liver, generated evidence for efficacious gene therapy in a mouse model for GSD-II. PMID:15585406

  2. Efficient production of germline transgenic chickens using lentiviral vectors.

    PubMed

    McGrew, Michael J; Sherman, Adrian; Ellard, Fiona M; Lillico, Simon G; Gilhooley, Hazel J; Kingsman, Alan J; Mitrophanous, Kyriacos A; Sang, Helen

    2004-07-01

    An effective method for genetic modification of chickens has yet to be developed. An efficient technology, enabling production of transgenic birds at high frequency and with reliable expression of transgenes, will have many applications, both in basic research and in biotechnology. We investigated the efficiency with which lentiviral vectors could transduce the chicken germ line and examined the expression of introduced reporter transgenes. Ten founder cockerels transmitted the vector to between 4% and 45% of their offspring and stable transmission to the G2 generation was demonstrated. Analysis of expression of reporter gene constructs in several transgenic lines showed a conserved expression profile between individuals that was maintained after transmission through the germ line. These data demonstrate that lentiviral vectors can be used to generate transgenic lines with an efficiency in the order of 100-fold higher than any previously published method, with no detectable silencing of transgene expression between generations. PMID:15192698

  3. Efficient production of germline transgenic chickens using lentiviral vectors

    PubMed Central

    McGrew, Michael J; Sherman, Adrian; Ellard, Fiona M; Lillico, Simon G; Gilhooley, Hazel J; Kingsman, Alan J; Mitrophanous, Kyriacos A; Sang, Helen

    2004-01-01

    An effective method for genetic modification of chickens has yet to be developed. An efficient technology, enabling production of transgenic birds at high frequency and with reliable expression of transgenes, will have many applications, both in basic research and in biotechnology. We investigated the efficiency with which lentiviral vectors could transduce the chicken germ line and examined the expression of introduced reporter transgenes. Ten founder cockerels transmitted the vector to between 4% and 45% of their offspring and stable transmission to the G2 generation was demonstrated. Analysis of expression of reporter gene constructs in several transgenic lines showed a conserved expression profile between individuals that was maintained after transmission through the germ line. These data demonstrate that lentiviral vectors can be used to generate transgenic lines with an efficiency in the order of 100-fold higher than any previously published method, with no detectable silencing of transgene expression between generations. PMID:15192698

  4. Honey Bee Mating Optimization Vector Quantization Scheme in Image Compression

    NASA Astrophysics Data System (ADS)

    Horng, Ming-Huwi

    The vector quantization is a powerful technique in the applications of digital image compression. The traditionally widely used method such as the Linde-Buzo-Gray (LBG) algorithm always generated local optimal codebook. Recently, particle swarm optimization (PSO) is adapted to obtain the near-global optimal codebook of vector quantization. In this paper, we applied a new swarm algorithm, honey bee mating optimization, to construct the codebook of vector quantization. The proposed method is called the honey bee mating optimization based LBG (HBMO-LBG) algorithm. The results were compared with the other two methods that are LBG and PSO-LBG algorithms. Experimental results showed that the proposed HBMO-LBG algorithm is more reliable and the reconstructed images get higher quality than those generated form the other three methods.

  5. Lifshitz effects on vector condensate induced by a magnetic field

    NASA Astrophysics Data System (ADS)

    Wu, Ya-Bo; Lu, Jun-Wang; Liu, Mo-Lin; Lu, Jian-Bo; Zhang, Cheng-Yuan; Yang, Zhuo-Qun

    2014-05-01

    By numerical and analytical methods, we study in detail the effects of the Lifshitz dynamical exponent z on the vector condensate induced by an applied magnetic field in the probe limit. Concretely, in the presence of the magnetic field, we obtain the Landau level independent of z, and we also find the critical value by coupling a Maxwell complex vector field and an SU(2) field into a (3+1)-dimensional Lifshitz black hole, respectively. The research results show that for the two models with the lowest Landau level, the increasing z improves the response of the critical temperature to the applied magnetic field even without the charge density, and the analytical results uphold the numerical results. In addition, we find that, even in the Lifshitz black hole, the Maxwell complex vector model is still a generalization of the SU(2) Yang-Mills model. Furthermore, we construct the square vortex lattice and discuss the implications of these results.

  6. Vector control and surveillance operations in the republic of singapore.

    PubMed

    Yoshikawa, Minako Jen

    2013-06-01

    Singapore is known for its comprehensive vector control methods that keep mosquito populations at low levels in the urban, tropical, and green city-state. This report describes the measures taken by the National Environment Agency on the basis of observations of vector control and surveillance activities in residential areas, construction sites, and foreign worker quarters. The government-led active operations dealt not only with mosquito control but also social issues in urban residential buildings where people with varying preferences live, the responsibilities of the business sector, and the education of multi-cultural/lingual residents and foreign workers. The public health measures implemented in Singapore offer useful ideas to countries/cities that have not yet established vector control programs against mosquito-borne infectious diseases. PMID:23874140

  7. Surrogate-based Reliability Analysis Using Support Vector Machine

    NASA Astrophysics Data System (ADS)

    Li, Gang; Liu, Zhiqiang

    2010-05-01

    An approach of surrogate-based reliability analysis by support vector machine with Monte-Carlo simulation is proposed. The efficient sampling techniques, such as uniform design and Latin Hypercube sampling, are used, and the SVM is trained with the sample pairs of input and output data obtained by the finite element analysis. The trained SVM model, as a solver-surrogate model, is intended to approximate the real performance function. Considering the selection of parameters for SVM affects the learning performance of SVM strongly, the Genetic Algorithm (GA) is integrated to the construction of the SVM, by optimizing the relevant parameters. The influence of the parameters on SVM is discussed and a methodology is proposed for selecting the SVM model. Support Vector Classification (SVC) based and Support Vector Regression (SVR) based reliability analyses are studied. Some numerical examples demonstrate the efficiency and applicability of the proposed method.

  8. Vector Control and Surveillance Operations in the Republic of Singapore

    PubMed Central

    Yoshikawa, Minako Jen

    2013-01-01

    Singapore is known for its comprehensive vector control methods that keep mosquito populations at low levels in the urban, tropical, and green city-state. This report describes the measures taken by the National Environment Agency on the basis of observations of vector control and surveillance activities in residential areas, construction sites, and foreign worker quarters. The government-led active operations dealt not only with mosquito control but also social issues in urban residential buildings where people with varying preferences live, the responsibilities of the business sector, and the education of multi-cultural/lingual residents and foreign workers. The public health measures implemented in Singapore offer useful ideas to countries/cities that have not yet established vector control programs against mosquito-borne infectious diseases. PMID:23874140

  9. A versatile system for USER cloning-based assembly of expression vectors for mammalian cell engineering.

    PubMed

    Lund, Anne Mathilde; Kildegaard, Helene Faustrup; Petersen, Maja Borup Kjær; Rank, Julie; Hansen, Bjarne Gram; Andersen, Mikael Rørdam; Mortensen, Uffe Hasbro

    2014-01-01

    A new versatile mammalian vector system for protein production, cell biology analyses, and cell factory engineering was developed. The vector system applies the ligation-free uracil-excision based technique--USER cloning--to rapidly construct mammalian expression vectors of multiple DNA fragments and with maximum flexibility, both for choice of vector backbone and cargo. The vector system includes a set of basic vectors and a toolbox containing a multitude of DNA building blocks including promoters, terminators, selectable marker- and reporter genes, and sequences encoding an internal ribosome entry site, cellular localization signals and epitope- and purification tags. Building blocks in the toolbox can be easily combined as they contain defined and tested Flexible Assembly Sequence Tags, FASTs. USER cloning with FASTs allows rapid swaps of gene, promoter or selection marker in existing plasmids and simple construction of vectors encoding proteins, which are fused to fluorescence-, purification-, localization-, or epitope tags. The mammalian expression vector assembly platform currently allows for the assembly of up to seven fragments in a single cloning step with correct directionality and with a cloning efficiency above 90%. The functionality of basic vectors for FAST assembly was tested and validated by transient expression of fluorescent model proteins in CHO, U-2-OS and HEK293 cell lines. In this test, we included many of the most common vector elements for heterologous gene expression in mammalian cells, in addition the system is fully extendable by other users. The vector system is designed to facilitate high-throughput genome-scale studies of mammalian cells, such as the newly sequenced CHO cell lines, through the ability to rapidly generate high-fidelity assembly of customizable gene expression vectors. PMID:24879460

  10. A Versatile System for USER Cloning-Based Assembly of Expression Vectors for Mammalian Cell Engineering

    PubMed Central

    Lund, Anne Mathilde; Kildegaard, Helene Faustrup; Petersen, Maja Borup Kjær; Rank, Julie; Hansen, Bjarne Gram; Andersen, Mikael Rørdam; Mortensen, Uffe Hasbro

    2014-01-01

    A new versatile mammalian vector system for protein production, cell biology analyses, and cell factory engineering was developed. The vector system applies the ligation-free uracil-excision based technique – USER cloning – to rapidly construct mammalian expression vectors of multiple DNA fragments and with maximum flexibility, both for choice of vector backbone and cargo. The vector system includes a set of basic vectors and a toolbox containing a multitude of DNA building blocks including promoters, terminators, selectable marker- and reporter genes, and sequences encoding an internal ribosome entry site, cellular localization signals and epitope- and purification tags. Building blocks in the toolbox can be easily combined as they contain defined and tested Flexible Assembly Sequence Tags, FASTs. USER cloning with FASTs allows rapid swaps of gene, promoter or selection marker in existing plasmids and simple construction of vectors encoding proteins, which are fused to fluorescence-, purification-, localization-, or epitope tags. The mammalian expression vector assembly platform currently allows for the assembly of up to seven fragments in a single cloning step with correct directionality and with a cloning efficiency above 90%. The functionality of basic vectors for FAST assembly was tested and validated by transient expression of fluorescent model proteins in CHO, U-2-OS and HEK293 cell lines. In this test, we included many of the most common vector elements for heterologous gene expression in mammalian cells, in addition the system is fully extendable by other users. The vector system is designed to facilitate high-throughput genome-scale studies of mammalian cells, such as the newly sequenced CHO cell lines, through the ability to rapidly generate high-fidelity assembly of customizable gene expression vectors. PMID:24879460

  11. Construction measurements

    SciTech Connect

    Barry, B.A.

    1988-01-01

    This text/reference on construction measurements contains material concerning electronic surveying and remote sensing. New to this edition is coverage of the GPS satellite positioning system, electronic distance measurement (EDM), laser sweep, calculator techniques, radial surveying and tracking, Loran-C, inertial navigation surveying, 3-point resection, computer software, and electronic fieldbooks. It covers the difference of elevation, angle measurements and directions, coordinate surveying and layout, offshore measurements, and random field and office techniques.

  12. Residual stresses and vector hysteresis modeling

    NASA Astrophysics Data System (ADS)

    Ktena, Aphrodite

    2016-04-01

    Residual stresses in magnetic materials, whether the result of processing or intentional loading, leave their footprint on macroscopic data, such hysteresis loops and differential permeability measurements. A Preisach-type vector model is used to reproduce the phenomenology observed based on assumptions deduced from the data: internal stresses lead to smaller and misaligned grains, hence increased domain wall pinning and angular dispersion of local easy axes, favouring rotation as a magnetization reversal mechanism; misaligned grains contribute to magnetostatic fields opposing the direction of the applied field. The model is using a vector operator which accounts for both reversible and irreversible processes; the Preisach concept for interactions for the role of stress related demagnetizing fields; and a characteristic probability density function which is constructed as a weighed sum of constituent functions: the material is modeled as consisting of various subsystems, e.g. reversal mechanisms or areas subject to strong/weak long range interactions and each subsystem is represented by a constituent probability density function. Our assumptions are validated since the model reproduces the hysteresis loops and differential permeability curves observed experimentally and calculations involving rotating inputs at various residual stress levels are consistent and in agreement with experimental evidence.

  13. Construction, characterization and expression of full length cDNA clone of sheep YAP1 gene.

    PubMed

    Sun, Wei; Li, Da; Su, Rui; Musa, Hassan H; Chen, Ling; Zhou, Hong

    2014-02-01

    RT-PCR, 5'RACE, 3'RACE were used to clone sheep full length cDNA sequence of YAP1 (Yes-associated protein 1), eukaryotic expression plasmid and a mutant that cannot be phosphorylated at Ser42 was successfully constructed. The amino acid sequence analysis revealed that sheep YAP1 gene encoded water-soluble protein and its relative molecular weight and isoelectric point was 44,079.0 Da and 4.91, respectively. Sub-cellular localization of YAP1 was in the nucleus, it is hydrophilic non-transmembrane and non-secreted protein. YAP1 protein contained 33 phosphorylation sites, seven glycosylation sites and two WW domains. The secondary structure of YAP1 was mainly composed of random coil, while the tertiary structure of domain area showed a forniciform helix structure. YAP1 gene was expressed in different tissues, the highest expression was in kidney and the lowest was in hypothalamus. The CDS of sheep YAP1was amplified by RT-PCR from healthy sheep longissimus dorsi muscle, cloned into pMD19-T simple vector by T/A ligation. YAP1 coding region was further sub-cloned into pEGFP-C1 vector by T4 Ligase to construct a eukaryotic expression plasmid and then make the eukaryotic expression vector as the template to construct the phosphorylation site mutant. PCR, restriction enzyme and sequencing were used to confirm the recombinant plasmid. The sheep full-length YAP1 cDNA sequence is 1712 in length encoding 403 amino acids. It was confirmed that the sheep YAP1 CDS was correctly inserted into eukaryotic expression vector and serine had been mutated to alanine by PCR, restriction digestion and sequencing. The result showed that the recombinant plasmid pEGFP-C1-YAP1 and pEGFP-C1-YAP1 S42A was constructed correctly, this will help for further studies on the YAP1 protein expression and its biological activities. PMID:24381103

  14. Image Compression Using Vector Quantization with Variable Block Size Division

    NASA Astrophysics Data System (ADS)

    Matsumoto, Hiroki; Kichikawa, Fumito; Sasazaki, Kazuya; Maeda, Junji; Suzuki, Yukinori

    In this paper, we propose a method for compressing a still image using vector quantization (VQ). Local fractal dimension (LFD) is computed to divided an image into variable block size. The LFD shows the complexity of local regions of an image, so that a region of an image that shows higher LFD values than those of other regions is partitioned into small blocks of pixels, while a region of an image that shows lower LFD values than those of other regions is partitioned into large blocks. Furthermore, we developed a division and merging algorithm to decrease the number of blocks to encode. This results in improvement of compression rate. We construct code books for respective blocks sizes. To encode an image, a block of pixels is transformed by discrete cosine transform (DCT) and the closest vector is chosen from the code book (CB). In decoding, the code vector corresponding to the index is selected from the CB and then the code vector is transformed by inverse DCT to reconstruct a block of pixels. Computational experiments were carried out to show the effectiveness of the proposed method. Performance of the proposed method is slightly better than that of JPEG. In the case of learning images to construct a CB being different from test images, the compression rate is comparable to compression rates of methods proposed so far, while image quality evaluated by NPIQM (normalized perceptual image quality measure) is almost the highest step. The results show that the proposed method is effective for still image compression.

  15. Vectors for cancer gene therapy.

    PubMed

    Zhang, J; Russell, S J

    1996-09-01

    Many viral and non-viral vector systems have now been developed for gene therapy applications. In this article, the pros and cons of these vector systems are discussed in relation to the different cancer gene therapy strategies. The protocols used in cancer gene therapy can be broadly divided into six categories including gene transfer to explanted cells for use as cell-based cancer vaccines; gene transfer to a small number of tumour cells in situ to achieve a vaccine effect; gene transfer to vascular endothelial cells (VECs) lining the blood vessels of the tumour to interfere with tumour angiogenesis; gene transfer to T lymphocytes to enhance their antitumour effector capability; gene transfer to haemopoietic stem cells (HSCs) to enhance their resistance to cytotoxic drugs and gene transfer to a large number of tumour cells in situ to achieve nonimmune tumour reduction with or without bystander effect. Each of the six strategies makes unique demands on the vector system and these are discussed with reference to currently available vectors. Aspects of vector biology that are in need of further development are discussed in some detail. The final section points to the potential use of replicating viruses as delivery vehicles for efficient in vivo gene transfer to disseminated cancers. PMID:9034598

  16. A generalized nonlocal vector calculus

    NASA Astrophysics Data System (ADS)

    Alali, Bacim; Liu, Kuo; Gunzburger, Max

    2015-10-01

    A nonlocal vector calculus was introduced in Du et al. (Math Model Meth Appl Sci 23:493-540, 2013) that has proved useful for the analysis of the peridynamics model of nonlocal mechanics and nonlocal diffusion models. A formulation is developed that provides a more general setting for the nonlocal vector calculus that is independent of particular nonlocal models. It is shown that general nonlocal calculus operators are integral operators with specific integral kernels. General nonlocal calculus properties are developed, including nonlocal integration by parts formula and Green's identities. The nonlocal vector calculus introduced in Du et al. (Math Model Meth Appl Sci 23:493-540, 2013) is shown to be recoverable from the general formulation as a special example. This special nonlocal vector calculus is used to reformulate the peridynamics equation of motion in terms of the nonlocal gradient operator and its adjoint. A new example of nonlocal vector calculus operators is introduced, which shows the potential use of the general formulation for general nonlocal models.

  17. Vector Encoding in Biochemical Networks

    NASA Astrophysics Data System (ADS)

    Potter, Garrett; Sun, Bo

    Encoding of environmental cues via biochemical signaling pathways is of vital importance in the transmission of information for cells in a network. The current literature assumes a single cell state is used to encode information, however, recent research suggests the optimal strategy utilizes a vector of cell states sampled at various time points. To elucidate the optimal sampling strategy for vector encoding, we take an information theoretic approach and determine the mutual information of the calcium signaling dynamics obtained from fibroblast cells perturbed with different concentrations of ATP. Specifically, we analyze the sampling strategies under the cases of fixed and non-fixed vector dimension as well as the efficiency of these strategies. Our results show that sampling with greater frequency is optimal in the case of non-fixed vector dimension but that, in general, a lower sampling frequency is best from both a fixed vector dimension and efficiency standpoint. Further, we find the use of a simple modified Ornstein-Uhlenbeck process as a model qualitatively captures many of our experimental results suggesting that sampling in biochemical networks is based on a few basic components.

  18. Adenovirus vectors targeting distinct cell types in the retina.

    PubMed

    Sweigard, J Harry; Cashman, Siobhan M; Kumar-Singh, Rajendra

    2010-04-01

    Purpose. Gene therapy for a number of retinal diseases necessitates efficient transduction of photoreceptor cells. Whereas adenovirus (Ad) serotype 5 (Ad5) does not transduce photoreceptors efficiently, previous studies have demonstrated improved photoreceptor transduction by Ad5 pseudotyped with Ad35 (Ad5/F35) or Ad37 (Ad5/F37) fiber or by the deletion of the RGD domain in the Ad5 penton base (Ad5DeltaRGD). However, each of these constructs contained a different transgene cassette, preventing the evaluation of the relative performance of these vectors, an important consideration before the use of these vectors in the clinic. The aim of this study was to evaluate these vectors in the retina and to attempt photoreceptor-specific transgene expression. Methods. Three Ad5-based vectors containing the same expression cassette were generated and injected into the subretinal space of adult mice. Eyes were analyzed for green fluorescence protein expression in flat-mounts, cross-sections, quantitative RT-PCR, and a modified stereological technique. A 257-bp fragment derived from the mouse opsin promoter was analyzed in the context of photoreceptor-specific transgene expression. Results. Each virus tested efficiently transduced the retinal pigment epithelium. The authors found no evidence that Ad5/F35 or Ad5/F37 transduced photoreceptors. Instead, they found that Ad5/F37 transduced Müller cells. Robust photoreceptor transduction by Ad5DeltaRGD was detected. Photoreceptor-specific transgene expression from the 257-bp mouse opsin promoter in the context of Ad5DeltaRGD vectors was found. Conclusions. Adenovirus vectors may be designed with tropism to distinct cell populations. Robust photoreceptor-specific transgene expression can be achieved in the context of Ad5DeltaRGD vectors. PMID:19892875

  19. Symbolic vector analysis in plasma physics

    NASA Astrophysics Data System (ADS)

    Qin, H.; Tang, W. M.; Rewoldt, G.

    1999-01-01

    Many problems in plasma physics involve substantial amounts of analytical vector calculation. The complexity usually originates from both the vector operations themselves and the underlying coordinate systems. A computer algebra package for symbolic vector analysis in general coordinate systems, GeneralVectorAnalysis (GVA), is developed using Mathematica. The modern viewpoint for 3D vector calculus, differential forms on 3-manifolds, is adopted to unify and systematize the vector calculus operations in general coordinate systems. Besides the basic vector analysis functions, the package provides asymptotic capabilities, 2D vector analysis notation, and a simple interface for users to define their own coordinate systems. These features will benefit physicists and applied mathematicians in their research where complicated vector analysis in complicated coordinate systems is required. Several applications of this symbolic vector analysis package to plasma physics are also given.

  20. Viral vector-based models of Parkinson's disease.

    PubMed

    Van der Perren, Anke; Van den Haute, Chris; Baekelandt, Veerle

    2015-01-01

    In order to study the molecular pathways of Parkinson's disease (PD) and to develop novel therapeutic strategies, scientific investigators rely on animal models. The identification of PD-associated genes has led to the development of genetic PD models as an alternative to toxin-based models. Viral vector-mediated loco-regional gene delivery provides an attractive way to express transgenes in the central nervous system. Several vector systems based on various viruses have been developed. In this chapter, we give an overview of the different viral vector systems used for targeting the CNS. Further, we describe the different viral vector-based PD models currently available based on overexpression strategies for autosomal dominant genes such as α-synuclein and LRRK2, and knockout or knockdown strategies for autosomal recessive genes, such as parkin, DJ-1, and PINK1. Models based on overexpression of α-synuclein are the most prevalent and extensively studied, and therefore the main focus of this chapter. Many efforts have been made to increase the expression levels of α-synuclein in the dopaminergic neurons. The best α-synuclein models currently available have been developed from a combined approach using newer AAV serotypes and optimized vector constructs, production, and purification methods. These third-generation α-synuclein models show improved face and predictive validity, and therefore offer the possibility to reliably test novel therapeutics. PMID:24839101

  1. Probability-theoretical analog of the vector Lyapunov function method

    SciTech Connect

    Nakonechnyi, A.N.

    1995-01-01

    The main ideas of the vector Lyapunov function (VLF) method were advanced in 1962 by Bellman and Matrosov. In this method, a Lyapunov function and a comparison equation are constructed for each subsystem. Then the dependences between the subsystems and the effect of external noise are allowed for by constructing a vector Lyapunov function (as a collection of the scalar Lyapunov functions of the subsystems) and an aggregate comparison function for the entire complex system. A probability-theoretical analog of this method for convergence analysis of stochastic approximation processes has been developed. The abstract approach proposed elsewhere eliminates all restrictions on the system phase space, the system trajectories, the class of Lyapunov functions, etc. The analysis focuses only on the conditions that relate sequences of Lyapunov function values with the derivative and ensure a particular type (mode, character) of stability. In our article, we extend this approach to the VLF method for discrete stochastic dynamic systems.

  2. Murine leukemia virus-based Tat-inducible long terminal repeat replacement vectors: a new system for anti-human immunodeficiency virus gene therapy.

    PubMed

    Cannon, P M; Kim, N; Kingsman, S M; Kingsman, A J

    1996-11-01

    We have constructed new murine leukemia virus (MLV)-based vectors (TIN vectors) which, following integration, contain human immunodeficiency virus (HIV) type 1 U3 and R sequences in place of the MLV U3 and R regions. This provides, for the first time, single transcriptional unit retroviral vectors under the control of Tat. TIN vectors have several advantages for anti-HIV gene therapy applications. PMID:8892960

  3. Murine leukemia virus-based Tat-inducible long terminal repeat replacement vectors: a new system for anti-human immunodeficiency virus gene therapy.

    PubMed Central

    Cannon, P M; Kim, N; Kingsman, S M; Kingsman, A J

    1996-01-01

    We have constructed new murine leukemia virus (MLV)-based vectors (TIN vectors) which, following integration, contain human immunodeficiency virus (HIV) type 1 U3 and R sequences in place of the MLV U3 and R regions. This provides, for the first time, single transcriptional unit retroviral vectors under the control of Tat. TIN vectors have several advantages for anti-HIV gene therapy applications. PMID:8892960

  4. The molecular basis of multiple vector insertion by gene targeting in mammalian cells.

    PubMed Central

    Ng, P; Baker, M D

    1999-01-01

    Gene targeting using sequence insertion vectors generally results in integration of one copy of the targeting vector generating a tandem duplication of the cognate chromosomal region of homology. However, occasionally the target locus is found to contain >1 copy of the integrated vector. The mechanism by which the latter recombinants arise is not known. In the present study, we investigated the molecular basis by which multiple vectors become integrated at the chromosomal immunoglobulin mu locus in a murine hybridoma. To accomplish this, specially designed insertion vectors were constructed that included six diagnostic restriction enzyme markers in the Cmu region of homology to the target chromosomal mu locus. This enabled contributions by the vector-borne and chromosomal Cmu sequences at the recombinant locus to be ascertained. Targeted recombinants were isolated and analyzed to determine the number of vector copies integrated at the chromosomal immunoglobulin mu locus. Targeted recombinants identified as bearing >1 copy of the integrated vector resulted from a Cmu triplication formed by two vector copies in tandem. Examination of the fate of the Cmu region markers suggested that this class of recombinant was generated predominantly, if not exclusively, by two targeted vector integration events, each involving insertion of a single copy of the vector. Both vector insertion events into the chromosomal mu locus were consistent with the double-strand-break repair mechanism of homologous recombination. We interpret our results, taken together, to mean that a proportion of recipient cells is in a predetermined state that is amenable to targeted but not random vector integration. PMID:10049930

  5. Extrapolation methods for vector sequences

    NASA Technical Reports Server (NTRS)

    Smith, David A.; Ford, William F.; Sidi, Avram

    1987-01-01

    This paper derives, describes, and compares five extrapolation methods for accelerating convergence of vector sequences or transforming divergent vector sequences to convergent ones. These methods are the scalar epsilon algorithm (SEA), vector epsilon algorithm (VEA), topological epsilon algorithm (TEA), minimal polynomial extrapolation (MPE), and reduced rank extrapolation (RRE). MPE and RRE are first derived and proven to give the exact solution for the right 'essential degree' k. Then, Brezinski's (1975) generalization of the Shanks-Schmidt transform is presented; the generalized form leads from systems of equations to TEA. The necessary connections are then made with SEA and VEA. The algorithms are extended to the nonlinear case by cycling, the error analysis for MPE and VEA is sketched, and the theoretical support for quadratic convergence is discussed. Strategies for practical implementation of the methods are considered.

  6. Gauge Theories of Vector Particles

    DOE R&D Accomplishments Database

    Glashow, S. L.; Gell-Mann, M.

    1961-04-24

    The possibility of generalizing the Yang-Mills trick is examined. Thus we seek theories of vector bosons invariant under continuous groups of coordinate-dependent linear transformations. All such theories may be expressed as superpositions of certain "simple" theories; we show that each "simple theory is associated with a simple Lie algebra. We may introduce mass terms for the vector bosons at the price of destroying the gauge-invariance for coordinate-dependent gauge functions. The theories corresponding to three particular simple Lie algebras - those which admit precisely two commuting quantum numbers - are examined in some detail as examples. One of them might play a role in the physics of the strong interactions if there is an underlying super-symmetry, transcending charge independence, that is badly broken. The intermediate vector boson theory of weak interactions is discussed also. The so-called "schizon" model cannot be made to conform to the requirements of partial gauge-invariance.

  7. Toward lattice fractional vector calculus

    NASA Astrophysics Data System (ADS)

    Tarasov, Vasily E.

    2014-09-01

    An analog of fractional vector calculus for physical lattice models is suggested. We use an approach based on the models of three-dimensional lattices with long-range inter-particle interactions. The lattice analogs of fractional partial derivatives are represented by kernels of lattice long-range interactions, where the Fourier series transformations of these kernels have a power-law form with respect to wave vector components. In the continuum limit, these lattice partial derivatives give derivatives of non-integer order with respect to coordinates. In the three-dimensional description of the non-local continuum, the fractional differential operators have the form of fractional partial derivatives of the Riesz type. As examples of the applications of the suggested lattice fractional vector calculus, we give lattice models with long-range interactions for the fractional Maxwell equations of non-local continuous media and for the fractional generalization of the Mindlin and Aifantis continuum models of gradient elasticity.

  8. Vector potential and metric perturbations of a rotating black hole

    NASA Technical Reports Server (NTRS)

    Chrzanowski, P. L.

    1975-01-01

    The assumption of factorized Green's functions together with the inhomogeneous Teukolsky equations are used to derive analytic expressions for homogeneous metric (and vector potential) perturbations of a Kerr black hole. These homogeneous solutions are used to construct solutions to the perturbation equations when sources are present. What one finds are particularly simple formulas for the energy and angular momentum flux in the asymptotic regions at plus or minus infinity.-

  9. Curved manifolds with conserved Runge-Lenz vectors

    SciTech Connect

    Ngome, J.-P.

    2009-12-15

    van Holten's algorithm is used to construct Runge-Lenz-type conserved quantities, induced by Killing tensors, on curved manifolds. For the generalized Taub-Newman-Unti-Tamburino metric, the most general external potential such that the combined system admits a conserved Runge-Lenz-type vector is found. In the multicenter case, the subclass of two-center metric exhibits a conserved Runge-Lenz-type scalar.

  10. Producing a Mammalian GFP Expression Vector Containing Neomycin Resistance Gene.

    PubMed

    Izadi, Manizheh; Abiri, Maryam; Keramatipour, Mohammad

    2009-04-01

    The green fluorescent protein (GFP) was originally isolated from the Jellyfish Aequorea Victoria that fluoresces green when exposed to blue light. GFP protein is composed of 238 amino acids with the molecular mass of 26.9 kD. The GFP gene is frequently used in cellular and molecular biology as a reporter gene. To date, many bacterial, yeast, fungal, plants, fly and mammalian cells, including human, have been created which express GFP. Martin Chalfie, Osamu Shimomura, and Roger Tsien were awarded the 2008 noble prize in chemistry for their discovery and development of GFP. In many studies on mammalian cells, GFP gene is introduced into cells using vector-based systems or a recombinant virus to track the location of a target protein or to study the expression level of the gene of interest, but in these studies there is no selection marker to normalize transfection. According to the importance of neomycin gene as a selection marker in mammalian cells, we aimed to produce a GFP expression vector that contains neomycin gene. GFP gene was separated from pEGFP-N1 vector and was inserted in the back-bone of pCDNA3.1/His/LacZ vector that contained the neomycin gene. The resulted vector contained GFP beside neomycin gene. PMID:23407141

  11. Incompatibility of Lactobacillus Vectors with Replicons Derived from Small Cryptic Lactobacillus Plasmids and Segregational Instability of the Introduced Vectors

    PubMed Central

    Posno, M.; Leer, R. J.; van Luijk, N.; van Giezen, M. J. F.; Heuvelmans, P. T. H. M.; Lokman, B. C.; Pouwels, P. H.

    1991-01-01

    Three new Lactobacillus vectors based on cryptic Lactobacillus plasmids were constructed. The shuttle vector pLP3537 consists of a 2.3-kb plasmid from Lactobacillus pentosus MD353, an erythromycin resistance gene from Staphylococcus aureus plasmid pE194, and pUC19 as a replicon for Escherichia coli. The vectors pLPE317 and pLPE323, which do not contain E. coli sequences, were generated by introducing the erythromycin resistance gene of pE194 into a 1.7- and a 2.3-kb plasmid from L. pentosus MD353, respectively. These vectors and the shuttle vector pLP825 (M. Posno, R. J. Leer, J. M. M. van Rijn, B. C. Lokman, and P. H. Pouwels, p. 397-401, in A. T. Ganesan and J. A. Hoch, ed., Genetics and biotechnology of bacilli, vol. 2, 1988) could be introduced by electroporation into Lactobacillus casei, L. pentosus, L. plantarum, L. acidophilus, L. fermentum, and L. brevis strains with similar efficiencies. Transformation efficiencies were strain dependent and varied from 102 to 107 transformants per μg of DNA. Plasmid DNA analysis of L. pentosus MD353 transformants revealed that the introduction of pLP3537 or pLPE323 was invariably accompanied by loss of the endogenous 2.3-kb plasmid. Remarkably, pLPE317 could only be introduced into an L. pentosus MD353 strain that had been previously cured of its endogenous 1.7-kb plasmid. The curing phenomena are most likely to be explained by the incompatibility of the vectors and resident plasmids. Lactobacillus vectors are generally rapidly lost when cells are cultivated in the absence of selective pressure. However, pLPE323 is stable in three of four Lactobacillus strains tested so far. Images PMID:16348515

  12. Bred vectors, singular vectors, and Lyapunov vectors in simple and complex models

    NASA Astrophysics Data System (ADS)

    Norwood, Adrienne

    We compute and compare three types of vectors frequently used to explore the instability properties of dynamical models, Lyapunov vectors (LVs), singular vectors (SVs), and bred vectors (BVs). The first model is the Lorenz (1963) three-variable model. We find BVs align with the locally fastest growing LV, which is often the second fastest growing global LV. The growth rates of the three types of vectors reveal all predict regime changes and durations of new regimes, as shown for BVs by Evans et al. (2004). The second model is the toy 'atmosphere-ocean model' developed by Pena and Kalnay (2004) coupling three Lorenz (1963) models with different time scales to test the effects of fast and slow modes of growth on the dynamical vectors. A fast 'extratropical atmosphere' is weakly coupled to a fast 'tropical atmosphere' which is strongly coupled to a slow 'ocean' system, the latter coupling imitating the tropical El Nino--Southern Oscillation. BVs separate the fast and slow modes of growth through appropriate selection of the breeding parameters. LVs successfully separate the fast 'extratropics' but cannot completely decouple the 'tropics' from the 'ocean,' leading to 'coupled' LVs that are affected by both systems but mainly dominated by one. SVs identify the fast modes but cannot capture the slow modes until the fast 'extratropics' are replaced with faster 'convection.' The dissimilar behavior of the three types of vectors degrades the similarities of the subspaces they inhabit (Norwood et al. 2013). The third model is a quasi-geostrophic channel model (Rotunno and Bao 1996) that is a simplification of extratropical synoptic-scale motions with baroclinic instabilities only. We were unable to successfully compute LVs for it. However, randomly initialized BVs quickly converge to a single vector that is the leading LV. The last model is the SPEEDY model created by Molteni (2003). It is a simplified general atmospheric circulation model with several types of instabilities

  13. Helper-Dependent Adenoviral Vectors

    PubMed Central

    Rosewell, Amanda; Vetrini, Francesco; Ng, Philip

    2012-01-01

    Helper-dependent adenoviral vectors are devoid of all viral coding sequences, possess a large cloning capacity, and can efficiently transduce a wide variety of cell types from various species independent of the cell cycle to mediate long-term transgene expression without chronic toxicity. These non-integrating vectors hold tremendous potential for a variety of gene transfer and gene therapy applications. Here, we review the production technologies, applications, obstacles to clinical translation and their potential resolutions, and the future challenges and unanswered questions regarding this promising gene transfer technology. PMID:24533227

  14. Construction material

    DOEpatents

    Wagh, Arun S.; Antink, Allison L.

    2008-07-22

    A structural material of a polystyrene base and the reaction product of the polystyrene base and a solid phosphate ceramic is applied as a slurry which includes one or more of a metal oxide or a metal hydroxide with a source of phosphate to produce a phosphate ceramic and a poly (acrylic acid or acrylate) or combinations or salts thereof and polystyrene or MgO applied to the polystyrene base and allowed to cure so that the dried aqueous slurry chemically bonds to the polystyrene base. A method is also disclosed of applying the slurry to the polystyrene base.

  15. Developing Adenoviral Vectors Encoding Therapeutic Genes Toxic to Host Cells: Comparing Binary and Single Inducible Vectors Expressing Truncated E2F-1

    PubMed Central

    Gomez-Gutierrez, Jorge G.; Rao, Xiao-Mei; Garcia-Garcia, Aracely; Hao, Hongying; McMasters, Kelly M.; Zhou, H. Sam

    2010-01-01

    Adenoviral vectors are highly efficient at transferring genes into cells and are broadly used in cancer gene therapy. However, many therapeutic genes are toxic to vector host cells and thus inhibit vector production. The truncated form of E2F-1 (E2Ftr), which lacks the transactivation domain, can significantly induce cancer cell apoptosis, but is also toxic to HEK-293 cells and inhibits adenovirus replication. To overcome this, we have developed binary- and single-vector systems with a modified tetracycline-off inducible promoter to control E2Ftr expression. We compared several vectors and found that the structure of expression cassettes in vectors significantly affects E2Ftr expression. One construct expresses high levels of inducible E2Ftr and efficiently causes apoptotic cancer cell death by activation of caspase-3. The approach developed in this study may be applied in other viral vectors for encoding therapeutic genes that are toxic to their host cells and/or inhibit vector propagation. PMID:20003994

  16. Vector Acoustics, Vector Sensors, and 3D Underwater Imaging

    NASA Astrophysics Data System (ADS)

    Lindwall, D.

    2007-12-01

    Vector acoustic data has two more dimensions of information than pressure data and may allow for 3D underwater imaging with much less data than with hydrophone data. The vector acoustic sensors measures the particle motions due to passing sound waves and, in conjunction with a collocated hydrophone, the direction of travel of the sound waves. When using a controlled source with known source and sensor locations, the reflection points of the sound field can be determined with a simple trigonometric calculation. I demonstrate this concept with an experiment that used an accelerometer based vector acoustic sensor in a water tank with a short-pulse source and passive scattering targets. The sensor consists of a three-axis accelerometer and a matched hydrophone. The sound source was a standard transducer driven by a short 7 kHz pulse. The sensor was suspended in a fixed location and the hydrophone was moved about the tank by a robotic arm to insonify the tank from many locations. Several floats were placed in the tank as acoustic targets at diagonal ranges of approximately one meter. The accelerometer data show the direct source wave as well as the target scattered waves and reflections from the nearby water surface, tank bottom and sides. Without resorting to the usual methods of seismic imaging, which in this case is only two dimensional and relied entirely on the use of a synthetic source aperture, the two targets, the tank walls, the tank bottom, and the water surface were imaged. A directional ambiguity inherent to vector sensors is removed by using collocated hydrophone data. Although this experiment was in a very simple environment, it suggests that 3-D seismic surveys may be achieved with vector sensors using the same logistics as a 2-D survey that uses conventional hydrophones. This work was supported by the Office of Naval Research, program element 61153N.

  17. A reagentless amperometric electrode based on carbon paste, chemically modified with D-lactate dehydrogenase, NAD(+), and mediator containing polymer for D-lactic acid analysis. I. Construction, composition, and characterization.

    PubMed

    Shu, H C; Mattiasson, B; Persson, B; Nagy, G; Gorton, L; Sahni, S; Geng, L; Boguslavsky, L; Skotheim, T

    1995-05-01

    A reagentless carbon paste electrode was designed for D-lactic acid analysis in a flow injection system for the monitoring of the production of D-lactate in a batch fermentation. D-Lactate dehydrogenase, nicotinamide adenine dinucleotide (NAD(+)), a synthetic redox polymer containing covalently attached toluidine blue O as mediator, graphite powder, and paraffin oil were used for the construction of the modified carbon paste electrode. D-Lactate selectivity was indicated by insignificant responses from a variety of possible interfernces including L-lactate. The electrodes gave a linear response in the range between 0.05 and 5 mM D-lactate, with a detecting limit of 30 muM, allowing a sample throughput of 20 h(-1). Preliminary investigations were made by covering the electrode surface with electropolymerized membranes. Satisfactory stability was observed, indicated by a reproducibility of 3.3% relative standard deviation (RSD, n = 31), with a non-membrane-covered electrode for the analysis of D-lactate in fermentation broth. A long-term stability (230 broth samples) was proven, suggesting the electrodes to have a good potential for use in on-line monitoring of fermentation processes. (c) 1995 John Wiley & Sons, Inc. PMID:18623311

  18. Syntheses, crystal structures and fluorescent properties of Cd(II), Hg(II) and Ag(I) coordination polymers constructed from 1H-1,2,4-triazole-1-acetic acid

    SciTech Connect

    Ding Degang; Xie Lixia; Fan Yaoting; Hou Hongwei; Xu Yan

    2009-06-15

    Three new d{sup 10} coordination polymers, namely [Cd(taa)Cl]{sub n}1, [Hg(taa)Cl]{sub n}2, and [Ag{sub 1.5}(taa)(NO{sub 3}){sub 0.5}]{sub n}3 (taa=1H-1,2,4-triazole-1-acatate anion) have been prepared and characterized by elemental analysis, IR, and single crystal X-ray diffraction. Compound 1 consists of two-dimensional layers constructed by carboxyl-linked helical chains, which are further linked through carboxyl group to generate a unique 3D open framework. Topological analysis reveals that the structure of 1 can be classified as an unprecedented (3,8)-connected network with the Schlaefli symbol (4.5{sup 2}){sub 2}(4{sup 2}.5{sup 8}.6{sup 14}.7{sup 3}.8). Compound 2 manifests a doubly interpenetrated decorated alpha-polonium cubic network with the Schlaefli symbol of (4{sup 10}.6{sup 2}.8{sup 3}). Compound 3 consists of 2D puckered layers made up of Ag centers and taa{sup -} bridges. In addition, all of these compounds are photoluminescent in the solid state with spectra that closely resemble those of the ligand precursor. - Graphical abstract: Three new compounds based on 1H-1,2,4-triazole-1-acetic acid and Cd(II), Hg(II) and Ag(I) salts display luminescent properties and may be potential candidates for luminescent materials.

  19. Methods of treating Parkinson's disease using viral vectors

    DOEpatents

    Bankiewicz, Krys; Cunningham, Janet

    2012-11-13

    Methods of delivering viral vectors, particularly recombinant AAV virions, to the central nervous system (CNS) are provided for the treatment of CNS disorders, particularly those disorders which involve the neurotransmitter dopamine. The methods entail providing rAAV virions that comprise a transgene encoding aromatic amino acid decarboxylase (AADC) and administering the virions to the brain of a mammal using a non-manual pump.

  20. Transposon-containing DNA cloning vector and uses thereof

    DOEpatents

    Berg, C.M.; Berg, D.E.; Wang, G.

    1997-07-08

    The present invention discloses a rapid method of restriction mapping, sequencing or localizing genetic features in a segment of deoxyribonucleic acid (DNA) that is up to 42 kb in size. The method in part comprises cloning of the DNA segment in a specialized cloning vector and then isolating nested deletions in either direction in vivo by intramolecular transposition into the cloned DNA. A plasmid has been prepared and disclosed. 4 figs.