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Sample records for hierarchical oligonucleotide primer

  1. Application of hierarchical oligonucleotide primer extension (HOPE) to assess relative abundances of ammonia- and nitrite-oxidizing bacteria.

    Science.gov (United States)

    Scarascia, Giantommaso; Cheng, Hong; Harb, Moustapha; Hong, Pei-Ying

    2017-04-04

    Establishing an optimal proportion of nitrifying microbial populations, including ammonia-oxidizing bacteria (AOB), nitrite-oxidizing bacteria (NOB), complete nitrite oxidizers (comammox) and ammonia-oxidizing archaea (AOA), is important for ensuring the efficiency of nitrification in water treatment systems. Hierarchical oligonucleotide primer extension (HOPE), previously developed to rapidly quantify relative abundances of specific microbial groups of interest, was applied in this study to track the abundances of the important nitrifying bacterial populations. The method was tested against biomass obtained from a laboratory-scale biofilm-based trickling reactor, and the findings were validated against those obtained by 16S rRNA gene-based amplicon sequencing. Our findings indicated a good correlation between the relative abundance of nitrifying bacterial populations obtained using both HOPE and amplicon sequencing. HOPE showed a significant increase in the relative abundance of AOB, specifically Nitrosomonas, with increasing ammonium content and shock loading (p < 0.001). In contrast, Nitrosospira remained stable in its relative abundance against the total community throughout the operational phases. There was a corresponding significant decrease in the relative abundance of NOB, specifically Nitrospira and those affiliated to comammox, during the shock loading. Based on the relative abundance of AOB and NOB (including commamox) obtained from HOPE, it was determined that the optimal ratio of AOB against NOB ranged from 0.2 to 2.5 during stable reactor performance. Overall, the HOPE method was developed and validated against 16S rRNA gene-based amplicon sequencing for the purpose of performing simultaneous monitoring of relative abundance of nitrifying populations. Quantitative measurements of these nitrifying populations obtained via HOPE would be indicative of reactor performance and nitrification functionality.

  2. Application of hierarchical oligonucleotide primer extension (HOPE) to assess relative abundances of ammonia- and nitrite-oxidizing bacteria

    KAUST Repository

    Scarascia, Giantommaso

    2017-04-04

    Background: Establishing an optimal proportion of nitrifying microbial populations, including ammonia-oxidizing bacteria (AOB), nitrite-oxidizing bacteria (NOB), complete nitrite oxidizers (comammox) and ammonia-oxidizing archaea (AOA), is important for ensuring the efficiency of nitrification in water treatment systems. Hierarchical oligonucleotide primer extension (HOPE), previously developed to rapidly quantify relative abundances of specific microbial groups of interest, was applied in this study to track the abundances of the important nitrifying bacterial populations. Results: The method was tested against biomass obtained from a laboratory-scale biofilm-based trickling reactor, and the findings were validated against those obtained by 16S rRNA gene-based amplicon sequencing. Our findings indicated a good correlation between the relative abundance of nitrifying bacterial populations obtained using both HOPE and amplicon sequencing. HOPE showed a significant increase in the relative abundance of AOB, specifically Nitrosomonas, with increasing ammonium content and shock loading (p < 0.001). In contrast, Nitrosospira remained stable in its relative abundance against the total community throughout the operational phases. There was a corresponding significant decrease in the relative abundance of NOB, specifically Nitrospira and those affiliated to comammox, during the shock loading. Based on the relative abundance of AOB and NOB (including commamox) obtained from HOPE, it was determined that the optimal ratio of AOB against NOB ranged from 0.2 to 2.5 during stable reactor performance. Conclusions: Overall, the HOPE method was developed and validated against 16S rRNA gene-based amplicon sequencing for the purpose of performing simultaneous monitoring of relative abundance of nitrifying populations. Quantitative measurements of these nitrifying populations obtained via HOPE would be indicative of reactor performance and nitrification functionality.

  3. Family-Specific Degenerate Primer Design: A Tool to Design Consensus Degenerated Oligonucleotides

    Science.gov (United States)

    Goñi, Sandra Elizabeth; Lozano, Mario Enrique

    2013-01-01

    Designing degenerate PCR primers for templates of unknown nucleotide sequence may be a very difficult task. In this paper, we present a new method to design degenerate primers, implemented in family-specific degenerate primer design (FAS-DPD) computer software, for which the starting point is a multiple alignment of related amino acids or nucleotide sequences. To assess their efficiency, four different genome collections were used, covering a wide range of genomic lengths: Arenavirus (10 × 104 nucleotides), Baculovirus (0.9 × 105 to 1.8 × 105 bp), Lactobacillus sp. (1 × 106 to 2 × 106 bp), and Pseudomonas sp. (4 × 106 to 7 × 106 bp). In each case, FAS-DPD designed primers were tested computationally to measure specificity. Designed primers for Arenavirus and Baculovirus were tested experimentally. The method presented here is useful for designing degenerate primers on collections of related protein sequences, allowing detection of new family members. PMID:23533783

  4. An Improved Barcoded Oligonucleotide Primers-based Next-generation Sequencing Approach for Direct Identification of Viral Pathogens in Clinical Specimens.

    Science.gov (United States)

    Wang, Chun Hua; Nie, Kai; Zhang, Yi; Wang, Ji; Zhou, Shuai Feng; Li, Xin Na; Zhou, Hang Yu; Qi, Shun Xiang; Ma, Xue Jun

    2017-01-01

    To provide a feasible and cost-effective next-generation sequencing (NGS) method for accurate identification of viral pathogens in clinical specimens, because enormous limitations impede the clinical use of common NGS, such as high cost, complicated procedures, tremendous data analysis, and high background noise in clinical samples. Viruses from cell culture materials or clinical specimens were identified following an improved NGS procedure: reduction of background noise by sample preprocessing, viral enrichment by barcoded oligonucleotide (random hexamer or non-ribosomal hexanucleotide) primer-based amplification, fragmentation-free library construction and sequencing of one-tube mixtures, as well as rapid data analysis using an in-house pipeline. NGS data demonstrated that both barcoded primer sets were useful to simultaneously capture multiple viral pathogens in cell culture materials or clinical specimens and verified that hexanucleotide primers captured as many viral sequences as hexamers did. Moreover, direct testing of clinical specimens using this improved hexanucleotide primer-based NGS approach provided further detailed genotypes of enteroviruses causing hand, foot, and mouth disease (HFMD) and identified other potential viruses or differentiated misdiagnosis events. The improved barcoded oligonucleotide primer-based NGS approach is simplified, time saving, cost effective, and appropriate for direct identification of viral pathogens in clinical practice. Copyright © 2017 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  5. Application of Locked Nucleic Acid (LNA) Primer and PCR Clamping by LNA Oligonucleotide to Enhance the Amplification of Internal Transcribed Spacer (ITS) Regions in Investigating the Community Structures of Plant-Associated Fungi.

    Science.gov (United States)

    Ikenaga, Makoto; Tabuchi, Masakazu; Kawauchi, Tomohiro; Sakai, Masao

    2016-09-29

    The simultaneous extraction of host plant DNA severely limits investigations of the community structures of plant-associated fungi due to the similar homologies of sequences in primer-annealing positions between fungi and host plants. Although fungal-specific primers have been designed, plant DNA continues to be excessively amplified by PCR, resulting in the underestimation of community structures. In order to overcome this limitation, locked nucleic acid (LNA) primers and PCR clamping by LNA oligonucleotides have been applied to enhance the amplification of fungal internal transcribed spacer (ITS) regions. LNA primers were designed by converting DNA into LNA, which is specific to fungi, at the forward primer side. LNA oligonucleotides, the sequences of which are complementary to the host plants, were designed by overlapping a few bases with the annealing position of the reverse primer. Plant-specific DNA was then converted into LNA at the shifted position from the 3' end of the primer-binding position. PCR using the LNA technique enhanced the amplification of fungal ITS regions, whereas those of the host plants were more likely to be amplified without the LNA technique. A denaturing gradient gel electrophoresis (DGGE) analysis displayed patterns that reached an acceptable level for investigating the community structures of plant-associated fungi using the LNA technique. The sequences of the bands detected using the LNA technique were mostly affiliated with known isolates. However, some sequences showed low similarities, indicating the potential to identify novel fungi. Thus, the application of the LNA technique is considered effective for widening the scope of community analyses of plant-associated fungi.

  6. Selection strategy and the design of hybrid oligonucleotide primers for RACE-PCR: cloning a family of toxin-like sequences from Agelena orientalis

    Directory of Open Access Journals (Sweden)

    Lipkin Alexey

    2007-05-01

    Full Text Available Abstract Background the use of specific but partially degenerate primers for nucleic acid hybridisations and PCRs amplification of known or unknown gene families was first reported well over a decade ago and the technique has been used widely since then. Results here we report a novel and successful selection strategy for the design of hybrid partially degenerate primers for use with RT-PCR and RACE-PCR for the identification of unknown gene families. The technique (named PaBaLiS has proven very effective as it allowed us to identify and clone a large group of mRNAs encoding neurotoxin-like polypeptide pools from the venom of Agelena orientalis species of spider. Our approach differs radically from the generally accepted CODEHOP principle first reported in 1998. Most importantly, our method has proven very efficient by performing better than an independently generated high throughput EST cloning programme. Our method yielded nearly 130 non-identical sequences from Agelena orientalis, whilst the EST cloning technique yielded only 48 non-identical sequences from 2100 clones obtained from the same Agelena material. In addition to the primer design approach reported here, which is almost universally applicable to any PCR cloning application, our results also indicate that venom of Agelena orientalis spider contains a much larger family of related toxin-like sequences than previously thought. Conclusion with upwards of 100,000 species of spider thought to exist, and a propensity for producing diverse peptide pools, many more peptides of pharmacological importance await discovery. We envisage that some of these peptides and their recombinant derivatives will provide a new range of tools for neuroscience research and could also facilitate the development of a new generation of analgesic drugs and insecticides.

  7. Gene Assembly from Chip-Synthesized Oligonucleotides

    Science.gov (United States)

    Eroshenko, Nikolai; Kosuri, Sriram; Marblestone, Adam H; Conway, Nicholas; Church, George M.

    2012-01-01

    De novo synthesis of long double-stranded DNA constructs has a myriad of applications in biology and biological engineering. However, its widespread adoption has been hindered by high costs. Cost can be significantly reduced by using oligonucleotides synthesized on high-density DNA chips. However, most methods for using off-chip DNA for gene synthesis have failed to scale due to the high error rates, low yields, and high chemical complexity of the chip-synthesized oligonucleotides. We have recently demonstrated that some commercial DNA chip manufacturers have improved error rates, and that the issues of chemical complexity and low yields can be solved by using barcoded primers to accurately and efficiently amplify subpools of oligonucleotides. This article includes protocols for computationally designing the DNA chip, amplifying the oligonucleotide subpools, and assembling 500-800 basepair (bp) constructs. PMID:25077042

  8. Hybridization with synthetic oligonucleotides

    Energy Technology Data Exchange (ETDEWEB)

    Szostak, J.W.; Stiles, J.I.; Tye, B.K.; Sherman, F.; Wu, R.

    1978-01-01

    Procedures are described for the use of synthetic oligonucleotides for Southern blot experiments and gene bank screening, and the effect of various mismatches on the efficiency of hybridization is demonstrated. The following topics are discussed: sensitivity vs. specificity, hybridization of a 12-mer to the lambda endolysin gene; hybridization of oligonucleotide probes to the E. coli lac operator; hybridization of synthetic probes to the CYC1 gene of yeast; and cloning eucaryotic genes. (HLW)

  9. SBE primer : multiplexing minisequencing-based genotyping

    Energy Technology Data Exchange (ETDEWEB)

    Kaderali, L. (Lars); Deshpande, A. (Alina); Uribe-Romeo, F. J. (Francisco J.); Schliep, A.; Torney, D. C. (David C.)

    2002-01-01

    Single-nucleotide polymorphism (SNP) analysis is a powerful tool for mapping and diagnosing disease-related alleles. Most of the known genetic diseases are caused by point mutations, and a growing number of SNPs will be routinely analyzed to diagnose genetic disorders. Mutation analysis by polymerase mediated single-base primer extension (minisequencing) can be massively parallelized using for example DNA microchips or flow cytometry with microspheres as solid support. By adding a unique oligonucleotide tag to the 5-inch end of the minisequencing primer and attaching the complementary anti-tag to the array or bead surface, the assay can be 'demultiplexed'. However, such high-throughput scoring of SNPs requires a high level of primer multiplexing in order to analyze multiple loci in one assay, thus enabling inexpensive and fast polymorphism scoring. Primers can be chosen from either the plus or the minus strand, and primers used in the same experiment must not bind to one another. To genotype a given number of polymorphic sites, the question is which primer to use for each SNP, and which primers to group into the same experiment. Furthermore, a crosshybridization-free tag/anti-tag code is required in order to sort the extended primers to the corresponding microspheres or chip spots. These problems pose challenging algorithmic questions. We present a computer program lo automate the design process for the assay. Oligonucleotide primers for the reaction are automatically selected by the software, a unique DNA tag/anti-tag system is generated, and the pairing of primers and DNA-Tags is automatically done in a way to avoid any crossreactivity. We report first results on a 45-plex genotyping assay, indicating that minisequencing can be adapted to be a powerful tool for high-throughput, massively parallel genotyping.

  10. EvOligo: A Novel Software to Design and Group Libraries of Oligonucleotides Applicable for Nucleic Acid-Based Experiments.

    Science.gov (United States)

    Milewski, Marek C; Kamel, Karol; Kurzynska-Kokorniak, Anna; Chmielewski, Marcin K; Figlerowicz, Marek

    2017-10-01

    Experimental methods based on DNA and RNA hybridization, such as multiplex polymerase chain reaction, multiplex ligation-dependent probe amplification, or microarray analysis, require the use of mixtures of multiple oligonucleotides (primers or probes) in a single test tube. To provide an optimal reaction environment, minimal self- and cross-hybridization must be achieved among these oligonucleotides. To address this problem, we developed EvOligo, which is a software package that provides the means to design and group DNA and RNA molecules with defined lengths. EvOligo combines two modules. The first module performs oligonucleotide design, and the second module performs oligonucleotide grouping. The software applies a nearest-neighbor model of nucleic acid interactions coupled with a parallel evolutionary algorithm to construct individual oligonucleotides, and to group the molecules that are characterized by the weakest possible cross-interactions. To provide optimal solutions, the evolutionary algorithm sorts oligonucleotides into sets, preserves preselected parts of the oligonucleotides, and shapes their remaining parts. In addition, the oligonucleotide sets can be designed and grouped based on their melting temperatures. For the user's convenience, EvOligo is provided with a user-friendly graphical interface. EvOligo was used to design individual oligonucleotides, oligonucleotide pairs, and groups of oligonucleotide pairs that are characterized by the following parameters: (1) weaker cross-interactions between the non-complementary oligonucleotides and (2) more uniform ranges of the oligonucleotide pair melting temperatures than other available software products. In addition, in contrast to other grouping algorithms, EvOligo offers time-efficient sorting of paired and unpaired oligonucleotides based on various parameters defined by the user.

  11. Peptide-LNA oligonucleotide conjugates

    DEFF Research Database (Denmark)

    Astakhova, I Kira; Hansen, Lykke Haastrup; Vester, Birte

    2013-01-01

    Although peptide-oligonucleotide conjugates (POCs) are well-known for nucleic acids delivery and therapy, reports on internal attachment of peptides to oligonucleotides are limited in number. To develop a convenient route for preparation of internally labeled POCs with improved biomedical...... properties, peptides were introduced into oligonucleotides via a 2'-alkyne-2'-amino-LNA scaffold. Derivatives of methionine- and leucine-enkephalins were chosen as model peptides of mixed amino acid content, which were singly and doubly incorporated into LNA/DNA strands using highly efficient copper......(i)-catalyzed azide-alkyne cycloaddition (CuAAC) "click" chemistry. DNA/RNA target binding affinity and selectivity of the resulting POCs were improved in comparison to LNA/DNA mixmers and unmodified DNA controls. This clearly demonstrates that internal attachment of peptides to oligonucleotides can significantly...

  12. Oligonucleotide conjugates for therapeutic applications.

    Science.gov (United States)

    Winkler, Johannes

    2013-07-01

    Insufficient pharmacokinetic properties and poor cellular uptake are the main hurdles for successful therapeutic development of oligonucleotide agents. The covalent attachment of various ligands designed to influence the biodistribution and cellular uptake or for targeting specific tissues is an attractive possibility to advance therapeutic applications and to expand development options. In contrast to advanced formulations, which often consist of multiple reagents and are sensitive to a variety of preparation conditions, oligonucleotide conjugates are defined molecules, enabling structure-based analytics and quality control techniques. This review gives an overview of current developments of oligonucleotide conjugates for therapeutic applications. Attached ligands comprise peptides, proteins, carbohydrates, aptamers and small molecules, including cholesterol, tocopherol and folic acid. Important linkage types and conjugation methods are summarized. The distinct ligands directly influence biochemical parameters, uptake mechanisms and pharmacokinetic properties.

  13. Oligonucleotide fingerprinting to individualize ungulates.

    Science.gov (United States)

    Schwaiger, F W; Gomolka, M; Geldermann, H; Zischler, H; Buitkamp, J; Epplen, J T; Ammer, H

    1992-01-01

    The optimal combination of restriction enzyme and oligonucleotide probe has been determined for the individualization of hoofed animal species (cattle, pig, goat, sheep, horse and camel). Four different restriction endonucleases were used as well as five synthetic oligonucleotide probes hybridizing to different simple tandem repeats for fingerprint analyses in unrelated cattle (Swiss and German Simmental of unknown relationship and three families): 4 x 10(15) cows and oxes would reveal different banding patterns after HinfI digestion using the probe (CAC)5/(GTG)5. The other species were investigated using HinfI (and HaeIII) and five different oligonucleotide probes specific for simple tandem repeats. Using (CAC)5/(GTG)5 the discrimination potential in sheep was about one order of magnitude lower than in cattle while in goats 6 x 10(10) specimen are easily differentiated with (CA)8/(GT)8. From an evolutionary standpoint it may be of interest that also in all other ungulate species tested, (CAC)5/(GTG)5 and (CA)8/(GT)8 exhibited the highest potential for individualization. Advantages of oligonucleotide fingerprinting are discussed.

  14. GENOMEMASKER package for designing unique genomic PCR primers

    Directory of Open Access Journals (Sweden)

    Kaplinski Lauris

    2006-03-01

    Full Text Available Abstract Background The design of oligonucleotides and PCR primers for studying large genomes is complicated by the redundancy of sequences. The eukaryotic genomes are particularly difficult to study due to abundant repeats. The speed of most existing primer evaluation programs is not sufficient for large-scale experiments. Results In order to improve the efficiency and success rate of automatic primer/oligo design, we created a novel method which allows rapid masking of repeats in large sequence files, for example in eukaryotic genomes. It also allows the detection of all alternative binding sites of PCR primers and the prediction of PCR products. The new method was implemented in a collection of efficient programs, the GENOMEMASKER package. The performance of the programs was compared to other similar programs. We also modified the PRIMER3 program, to be able to design primers from lowercase-masked sequences. Conclusion The GENOMEMASKER package is able to mask the entire human genome for non-unique primers within 6 hours and find locations of all binding sites for 10 000 designed primer pairs within 10 minutes. Additionally, it predicts all alternative PCR products from large genomes for given primer pairs.

  15. Hierarchical photocatalysts.

    Science.gov (United States)

    Li, Xin; Yu, Jiaguo; Jaroniec, Mietek

    2016-05-07

    As a green and sustainable technology, semiconductor-based heterogeneous photocatalysis has received much attention in the last few decades because it has potential to solve both energy and environmental problems. To achieve efficient photocatalysts, various hierarchical semiconductors have been designed and fabricated at the micro/nanometer scale in recent years. This review presents a critical appraisal of fabrication methods, growth mechanisms and applications of advanced hierarchical photocatalysts. Especially, the different synthesis strategies such as two-step templating, in situ template-sacrificial dissolution, self-templating method, in situ template-free assembly, chemically induced self-transformation and post-synthesis treatment are highlighted. Finally, some important applications including photocatalytic degradation of pollutants, photocatalytic H2 production and photocatalytic CO2 reduction are reviewed. A thorough assessment of the progress made in photocatalysis may open new opportunities in designing highly effective hierarchical photocatalysts for advanced applications ranging from thermal catalysis, separation and purification processes to solar cells.

  16. Simultaneous Detection of Escherichia coli, Salmonella enterica, Listeria monocytegenes and Bacillus cereus by Oligonucleotide Microarray

    Directory of Open Access Journals (Sweden)

    Meysam Sarshar

    2015-11-01

    Full Text Available Background: Traditional laboratory methods to detect pathogenic bacteria are time consuming and laborious. Therefore, it is essential to use powerful and reliable molecular methods for quick and simultaneous detection of microbial pathogens. Objectives: The current study aimed to evaluate the capability and efficiency of 23S rDNA sequence for rapid and simultaneous detection of four important food-borne pathogens by an oligonucleotide microarray technique. Materials and Methods: The 23S rDNA sequences of Escherichia coli, Salmonella enterica, Listeria monocytogenes and Bacillus cereus were obtained from GenBank databases and used to design the oligonucleotide probes and primers by Vector NTI software. Oligonucleotide probes were placed on a nylon membrane and hybridization was performed between probes and 23S rDNA digoxigenin-labeled polymerase chain reaction (PCR products. Hybridization signals were visualized by NBT/BCIP color development. Results: Positive hybridization color was produced for Escherichia coli, Salmonella enterica, Listeria monocytogenes and Bacillus cereus. The oligonucleotide microarray detected all bacterial strains in a single reaction in less than five hours. The sensitivity of the performed microarray assay was 103 cfu/mL for each species of pathogen. No cross reaction was found between the tested bacterial species. Conclusions: The obtained results indicated that amplification of 23S rDNA gene followed by oligonucleotide microarray hybridization is a rapid and reliable method to identify and discriminate foodborne pathogens tested under the study.

  17. Oligonucleotide microarrays for the detection and identification of viable beer spoilage bacteria.

    Science.gov (United States)

    Weber, D G; Sahm, K; Polen, T; Wendisch, V F; Antranikian, G

    2008-10-01

    The design and evaluation of an oligonucleotide microarray in order to detect and identify viable bacterial species that play a significant role in beer spoilage. These belong to the species of the genera Lactobacillus, Megasphaera, Pediococcus and Pectinatus. Oligonucleotide probes specific to beer spoilage bacteria were designed. In order to detect viable bacteria, the probes were designed to target the intergenic spacer regions (ISR) between 16S and 23S rRNA. Prior to hybridization the ISR were amplified by combining reverse transcriptase and polymerase chain reactions using a designed consenus primer. The developed oligonucleotide microarrays allows the detection of viable beer spoilage bacteria. This method allows the detection and discrimination of single bacterial species in a sample containing complex microbial community. Furthermore, microarrays using oligonucleotide probes targeting the ISR allow the distinction between viable bacteria with the potential to grow and non growing bacteria. The results demonstrate the feasibility of oligonucleotide microarrays as a contamination control in food industry for the detection and identification of spoilage micro-organisms within a mixed population.

  18. Chemoselective Coupling Preserves the Substrate Integrity of Surface-Immobilized Oligonucleotides for Emulsion PCR-Based Gene Library Construction.

    Science.gov (United States)

    Malone, Marie L; Cavett, Valerie J; Paegel, Brian M

    2017-01-09

    Combinatorial bead libraries figure prominently in next-generation sequencing and are also important tools for in vitro evolution. The most common methodology for generating such bead libraries, emulsion PCR (emPCR), enzymatically extends bead-immobilized oligonucleotide PCR primers in emulsion droplets containing a single progenitor library member. Primers are almost always immobilized on beads via noncovalent biotin-streptavidin binding. Here, we describe covalent bead functionalization with primers (∼106 primers/2.8-μm-diameter bead) via either azide-alkyne click chemistry or Michael addition. The primers are viable polymerase substrates (4-7% bead-immobilized enzymatic extension product yield from one thermal cycle). Carbodiimide-activated carboxylic acid beads only react with oligonucleotides under conditions that promote nonspecific interactions (low salt, low pH, no detergent), comparably immobilizing primers on beads, but yielding no detectable enzymatic extension product. Click-functionalized beads perform satisfactorily in emPCR of a site-saturation mutagenesis library, generating monoclonal templated beads (104-105 copies/bead, 1.4-kb amplicons). This simpler, chemical approach to primer immobilization may spur more economical library preparation for high-throughput sequencing and enable more complex surface elaboration for in vitro evolution.

  19. LNA-modified isothermal oligonucleotide microarray for ...

    Indian Academy of Sciences (India)

    2014-10-20

    Oct 20, 2014 ... Bacillus licheniformis, Bacillus pumilus, Bacillus megaterium and Bacillus circulans), we unified the melting temperatures of .... Sequences and calculated melting temperatures of DNA oligonucleotide probes and LNA-modified oligonucleotide probes for. Bacillus .... quick chilling on ice for 5 min. 13.5 μL of ...

  20. RNase H sequence preferences influence antisense oligonucleotide efficiency.

    Science.gov (United States)

    Kielpinski, Lukasz J; Hagedorn, Peter H; Lindow, Morten; Vinther, Jeppe

    2017-11-08

    RNase H cleaves RNA in RNA-DNA duplexes. It is present in all domains of life as well as in multiple viruses and is essential for mammalian development and for human immunodeficiency virus replication. Here, we developed a sequencing-based method to measure the cleavage of thousands of different RNA-DNA duplexes and thereby comprehensively characterized the sequence preferences of HIV-1, human and Escherichia coli RNase H enzymes. We find that the catalytic domains of E. coli and human RNase H have nearly identical sequence preferences, which correlate with the efficiency of RNase H-recruiting antisense oligonucleotides. The sequences preferred by HIV-1 RNase H are distributed in the HIV genome in a way suggesting selection for efficient RNA cleavage during replication. Our findings can be used to improve the design of RNase H-recruiting antisense oligonucleotides and show that sequence preferences of HIV-1 RNase H may have shaped evolution of the viral genome and contributed to the use of tRNA-Lys3 as primer during viral replication. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  1. Hierarchical XP

    OpenAIRE

    Jacobi, Carsten; Rumpe, Bernhard

    2014-01-01

    XP is a light-weight methodology suited particularly for small-sized teams that develop software which has only vague or rapidly changing requirements. The discipline of systems engineering knows it as approach of incremental system change or also of "muddling through". In this paper, we introduce three well known methods of reorganizing companies, namely, the holistic approach, the incremental approach, and the hierarchical approach. We show similarities between software engineering methods ...

  2. Identification of sex in Cetaceans by multiplexing with three ZFX and ZFY specific primers

    NARCIS (Netherlands)

    Berube, M; Palsboll, P

    We sequenced 540 nucleotides of the last exon in the ZFY/ZFX gene in two males and two females for eight cetacean species; four odontocetes (toothed whales) and four mysticetes (baleen whales). Based upon the obtained nucleotide sequences, we designed two sets of oligonucleotide primers for specific

  3. Primer3Plus, an enhanced web interface to Primer3

    NARCIS (Netherlands)

    Untergasser, A.; Nijveen, H.; Rao, X.; Bisseling, T.; Geurts, R.; Leunissen, J.A.M.

    2007-01-01

    Here we present Primer3Plus, a new web interface to the popular Primer3 primer design program as an enhanced alternative for the CGI- scripts that come with Primer3. Primer3 consists of a command line program and a web interface. The web interface is one large form showing all of the possible

  4. Electronic Structures of LNA Phosphorothioate Oligonucleotides

    DEFF Research Database (Denmark)

    Bohr, Henrik G.; Shim, Irene; Stein, Cy

    2017-01-01

    Important oligonucleotides in anti-sense research have been investigated in silico and experimentally. This involves quantum mechanical (QM) calculations and chromatography experiments on locked nucleic acid (LNA) phosphorothioate (PS) oligonucleotides. iso-potential electrostatic surfaces...... or differentiate between the individual PS diastereoisomers determined by the position of sulfur atoms. Rules are derived from the electronic calculations of these molecules and include the effects of the phosphorothioate chirality and formation of electrostatic potential surfaces. Physical and electrochemical...

  5. Methods for the preparation of large quantities of complex single-stranded oligonucleotide libraries.

    Science.gov (United States)

    Murgha, Yusuf E; Rouillard, Jean-Marie; Gulari, Erdogan

    2014-01-01

    Custom-defined oligonucleotide collections have a broad range of applications in fields of synthetic biology, targeted sequencing, and cytogenetics. Also, they are used to encode information for technologies like RNA interference, protein engineering and DNA-encoded libraries. High-throughput parallel DNA synthesis technologies developed for the manufacture of DNA microarrays can produce libraries of large numbers of different oligonucleotides, but in very limited amounts. Here, we compare three approaches to prepare large quantities of single-stranded oligonucleotide libraries derived from microarray synthesized collections. The first approach, alkaline melting of double-stranded PCR amplified libraries with a biotinylated strand captured on streptavidin coated magnetic beads results in little or no non-biotinylated ssDNA. The second method wherein the phosphorylated strand of PCR amplified libraries is nucleolyticaly hydrolyzed is recommended when small amounts of libraries are needed. The third method combining in vitro transcription of PCR amplified libraries to reverse transcription of the RNA product into single-stranded cDNA is our recommended method to produce large amounts of oligonucleotide libraries. Finally, we propose a method to remove any primer binding sequences introduced during library amplification.

  6. The Use of Degenerate Primers in qPCR Analysis of Functional Genes Can Cause Dramatic Quantification Bias as Revealed by Investigation of nifH Primer Performance.

    Science.gov (United States)

    Gaby, John Christian; Buckley, Daniel H

    2017-10-01

    The measurement of functional gene abundance in diverse microbial communities often employs quantitative PCR (qPCR) with highly degenerate oligonucleotide primers. While degenerate PCR primers have been demonstrated to cause template-specific bias in PCR applications, the effect of such bias on qPCR has been less well explored. We used a set of diverse, full-length nifH gene standards to test the performance of several universal nifH primer sets in qPCR. We found significant template-specific bias in all but the PolF/PolR primer set. Template-specific bias caused more than 1000-fold mis-estimation of nifH gene copy number for three of the primer sets and one primer set resulted in more than 10,000-fold mis-estimation. Furthermore, such template-specific bias will cause qPCR estimates to vary in response to beta-diversity, thereby causing mis-estimation of changes in gene copy number. A reduction in bias was achieved by increasing the primer concentration. We conclude that degenerate primers should be evaluated across a range of templates, annealing temperatures, and primer concentrations to evaluate the potential for template-specific bias prior to their use in qPCR.

  7. Mean and variance of the Gibbs free energy of oligonucleotides in the nearest neighbor model under varying conditions.

    Science.gov (United States)

    Rahmann, Sven; Gräfe, Christine

    2004-11-22

    In order to assess the stability of DNA-DNA hybridizations-for example during PCR primer design or oligonucleotide selection for microarrays-one needs to predict the change in Gibbs free energy DeltaG during hybridization. The nearest neighbor model provides a good compromise between accuracy and computational simplicity for this task. To determine optimal combinations of reaction parameters (temperature, salt concentration, oligonucleotide length and GC-content), one would like to understand how DeltaG depends on all of these parameters simultaneously. We derive analytic results about the distribution of nearest neighbor DeltaG values for a Bernoulli random sequence model (specified by oligonucleotide length and average GC-content) under given experimental conditions. We find that the distribution of DeltaG values is approximately Gaussian and provide exact formulas for expectation and variance.

  8. Asset management primer

    Science.gov (United States)

    1999-12-01

    According to this primer, asset management is a systematic process of maintaining, upgrading, and operating physical assets cost-effectively. It combines engineering principles with sound business practices and economic theory, and it provides tools ...

  9. Electron detachment dissociation (EDD) pathways in oligonucleotides

    Science.gov (United States)

    Kinet, Catherine; Gabelica, Valérie; Balbeur, Dorothée; de Pauw, Edwin

    2009-06-01

    Electron detachment dissociation (EDD) and electron photodetachment dissociation (EPD) are two novel fragmentation methods yielding radicals from negatively charged ions. With the goal of comparing EDD, EPD and the more traditional collision-induced dissociation (CID) and infrared multiphoton dissociation (IRMPD) fragmentation processes in oligonucleotides, we studied here the EDD fragmentation pathways of oligonucleotides of varying length. We chose polythymine oligonucleotides because these are the least prone to secondary structure formation, and found complete sequence coverage by EDD for up to dT20. We also found that the fragmentation pathways change with oligonucleotide length: electron detachment is a mandatory step in the fragmentation of larger sequences, while shorter oligonucleotides can also fragment via direct electronic or vibrational excitation by the electrons. This is supported by (1) the fact that continuous ejection of the charge-reduced species does not totally prevent fragmentation of short oligonucleotides dT5 and dT6, (2) the fact that CID and EDD fragments are more similar for small oligonucleotides (although double resonance experiments show that they are not all issued from the same mechanisms), and (3) the fact that electron-induced dissociation (EID) of singly charged dT3 and dT4 gives similar fragments as EDD of doubly charged dT5 and dT6. Finally, the detachment efficiency as a function of the nature of the nucleobase was studied. The effect of base on electron detachment in EDD (G > T > A > C) is different than in EPD (G > A > C > T), indicating different electron loss mechanisms.

  10. China Energy Primer

    Energy Technology Data Exchange (ETDEWEB)

    Ni, Chun Chun

    2009-11-16

    Based on extensive analysis of the 'China Energy Databook Version 7' (October 2008) this Primer for China's Energy Industry draws a broad picture of China's energy industry with the two goals of helping users read and interpret the data presented in the 'China Energy Databook' and understand the historical evolution of China's energy inustry. Primer provides comprehensive historical reviews of China's energy industry including its supply and demand, exports and imports, investments, environment, and most importantly, its complicated pricing system, a key element in the analysis of China's energy sector.

  11. Merlin: Computer-Aided Oligonucleotide Design for Large Scale Genome Engineering with MAGE.

    Science.gov (United States)

    Quintin, Michael; Ma, Natalie J; Ahmed, Samir; Bhatia, Swapnil; Lewis, Aaron; Isaacs, Farren J; Densmore, Douglas

    2016-06-17

    Genome engineering technologies now enable precise manipulation of organism genotype, but can be limited in scalability by their design requirements. Here we describe Merlin ( http://merlincad.org ), an open-source web-based tool to assist biologists in designing experiments using multiplex automated genome engineering (MAGE). Merlin provides methods to generate pools of single-stranded DNA oligonucleotides (oligos) for MAGE experiments by performing free energy calculation and BLAST scoring on a sliding window spanning the targeted site. These oligos are designed not only to improve recombination efficiency, but also to minimize off-target interactions. The application further assists experiment planning by reporting predicted allelic replacement rates after multiple MAGE cycles, and enables rapid result validation by generating primer sequences for multiplexed allele-specific colony PCR. Here we describe the Merlin oligo and primer design procedures and validate their functionality compared to OptMAGE by eliminating seven AvrII restriction sites from the Escherichia coli genome.

  12. Photochemical immobilization of anthraquinone conjugated oligonucleotides and PCR amplicons on solid surfaces

    DEFF Research Database (Denmark)

    Koch, T; Jacobsen, N; Fensholdt, J

    2000-01-01

    facilitate AQ conjugation during routine DNA synthesis, thus enabling the AQ-oligonucleotides to be immobilized in a very convenient and efficient manner. AQ-conjugated PCR primers can be used directly in PCR. When the PCR is performed in solution, the amplicons can be immobilized after the PCR. Moreover...... advantages as they are able to generate highly reactive species in an orientation specific manner. As presented here, anthraquinone (AQ) mediated covalent DNA immobilization appears to be superior to currently known procedures. A synthetic procedure providing AQ-phosphoramidites is presented. These reagents......, when the primers are immobilized prior to the PCR, a solid-phase PCR can be performed and the amplicons are thus produced directly on the solid support....

  13. An oligonucleotide-tagged microarray for routine diagnostics of colon cancer by genotyping KRAS mutations

    DEFF Research Database (Denmark)

    Liu, Yuliang; Guðnason, Haukur; Li, Yiping

    2014-01-01

    and therapeutic importance. In this study, KRAS gene fragments containing mutations in codon 12 were amplified by multiplex PCR using a 5'-Cy5-labeled reverse primer in combination with 3'-mutation-specific forward primers that were linked with four unique nucleotide-sequence tags at the 5'-end. The Cy5-labeled...... or spiked fecal samples. The immobilized tag-probes were stable under multiple thermal cycling treatments, allowing re-use of the tag-microarray and further optimization to solid PCR. Our results demonstrated that a novel oligonucleotide-tagged microarray system has been developed which would be suitable...... to be used for detection of KRAS mutations and clinical diagnosis of CRC....

  14. Semantic Web Primer

    NARCIS (Netherlands)

    Antoniou, Grigoris; Harmelen, Frank van

    2004-01-01

    The development of the Semantic Web, with machine-readable content, has the potential to revolutionize the World Wide Web and its use. A Semantic Web Primer provides an introduction and guide to this still emerging field, describing its key ideas, languages, and technologies. Suitable for use as a

  15. Identification of ISSR Primers

    African Journals Online (AJOL)

    leaf preferred by consumers. These genotypes ... plants, primers that work for one may not work for another. Hence .... Naidoo, Madhupa and Dr Bayo Shittu for access to laboratory facilities, training in different techniques and editing of manuscript. This work was made possible by a fellowship for Female. Academic Staff ...

  16. An SAT® Validity Primer

    Science.gov (United States)

    Shaw, Emily J.

    2015-01-01

    This primer should provide the reader with a deeper understanding of the concept of test validity and will present the recent available validity evidence on the relationship between SAT® scores and important college outcomes. In addition, the content examined on the SAT will be discussed as well as the fundamental attention paid to the fairness of…

  17. Synthetic oligonucleotides : Useful molecules? A review

    NARCIS (Netherlands)

    Calogero, A; Hospers, GAP; Mulder, NH

    1997-01-01

    Specific inhibition of mammalian genes is possible through the use of antisense oligonucleotides (AS ODNs) or ribozymes. These strategies have led to a better understanding of several cellular and molecular mechanisms, among which cancer development. Recently, these strategies have been applied also

  18. Associating Oligonucleotides with Positively Charged Liposomes

    Czech Academy of Sciences Publication Activity Database

    Jurkiewicz, P.; Okruszek, A.; Hof, Martin; Langner, M.

    2003-01-01

    Roč. 8, č. 1 (2003), s. 77-84 ISSN 1425-8153 R&D Projects: GA MŠk LN00A032 Institutional research plan: CEZ:AV0Z4040901 Keywords : oligonucleotides * fluorescence correlation spectroscopy * DOTAP Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 0.455, year: 2003

  19. A comparative analysis of existing oligonucleotides selection ...

    African Journals Online (AJOL)

    SERVER

    2007-07-04

    Jul 4, 2007 ... rence of poly-morphisms in genomic DNA (Schena,. 1996). Two DNA chips format currently in wide use are, the cDNA array format and high density synthetic oligo- nucleotide array format (oligos). The oligonucleotide app- roach has some advantages because it allows the user to design oligos for each ...

  20. Metallo-Phthalocyanine Near-IR Fluorophores: Oligonucleotide Conjugates and Their Applications in PCR Assays

    Science.gov (United States)

    Nesterova, Irina V.; Verdree, Vera T.; Pakhomov, Serhii; Strickler, Karen L.; Allen, Michael W.; Hammer, Robert P.; Soper, Steven A.

    2011-01-01

    Water soluble, metallo-pthalocyanine (MPc) near-IR fluorophores were designed, synthesized, and evaluated as highly stable and sensitive reporters for fluorescence assays. Their conjugation to oligonucleotides was achieved via succinimidyl ester-amino coupling chemistry with the conditions for conjugation extensively examined and optimized. In addition, various conjugate purification and isolation techniques were evaluated as well. Results showed that under proper conditions and following purification using reverse-phase ion-pair chromatography, labeling efficiencies near 80% could be achieved using ZnPc (Zn phthalocyanine) as the labeling fluorophore. Absorption and fluorescence spectra accumulated for the conjugates indicated that the intrinsic fluorescence properties of the MPc’s were not significantly altered by covalent attachment to oligonucleotides. As an example of the utility of MPc reporters, we used the MPc–oligonucleotide conjugates as primers for PCR (polymerase chain reaction) amplifications with the products sorted via electrophoresis and detected using near-IR fluorescence (λex = 680 nm). The MPc dyes were found to be more chemically stable under typical thermal cycling conditions used for PCR compared to the carbocyanine-based near-IR reporter systems typically used and produced single and narrow bands in the electrophoretic traces, indicative of producing a single PCR product during amplification. PMID:18030995

  1. Primer on molecular genetics

    Energy Technology Data Exchange (ETDEWEB)

    1992-04-01

    This report is taken from the April 1992 draft of the DOE Human Genome 1991--1992 Program Report, which is expected to be published in May 1992. The primer is intended to be an introduction to basic principles of molecular genetics pertaining to the genome project. The material contained herein is not final and may be incomplete. Techniques of genetic mapping and DNA sequencing are described.

  2. Coal Bed Methane Primer

    Energy Technology Data Exchange (ETDEWEB)

    Dan Arthur; Bruce Langhus; Jon Seekins

    2005-05-25

    During the second half of the 1990's Coal Bed Methane (CBM) production increased dramatically nationwide to represent a significant new source of income and natural gas for many independent and established producers. Matching these soaring production rates during this period was a heightened public awareness of environmental concerns. These concerns left unexplained and under-addressed have created a significant growth in public involvement generating literally thousands of unfocused project comments for various regional NEPA efforts resulting in the delayed development of public and fee lands. The accelerating interest in CBM development coupled to the growth in public involvement has prompted the conceptualization of this project for the development of a CBM Primer. The Primer is designed to serve as a summary document, which introduces and encapsulates information pertinent to the development of Coal Bed Methane (CBM), including focused discussions of coal deposits, methane as a natural formed gas, split mineral estates, development techniques, operational issues, producing methods, applicable regulatory frameworks, land and resource management, mitigation measures, preparation of project plans, data availability, Indian Trust issues and relevant environmental technologies. An important aspect of gaining access to federal, state, tribal, or fee lands involves education of a broad array of stakeholders, including land and mineral owners, regulators, conservationists, tribal governments, special interest groups, and numerous others that could be impacted by the development of coal bed methane. Perhaps the most crucial aspect of successfully developing CBM resources is stakeholder education. Currently, an inconsistent picture of CBM exists. There is a significant lack of understanding on the parts of nearly all stakeholders, including industry, government, special interest groups, and land owners. It is envisioned the Primer would being used by a variety of

  3. Application of Oligonucleotide Microarrays for Bacterial Source Tracking of Environmental Enterococcus sp. Isolates

    Directory of Open Access Journals (Sweden)

    John S. Furey

    2005-04-01

    Full Text Available In an effort towards adapting new and defensible methods for assessing and managing the risk posed by microbial pollution, we evaluated the utility of oligonucleotide microarrays for bacterial source tracking (BST of environmental Enterococcus sp. isolates derived from various host sources. Current bacterial source tracking approaches rely on various phenotypic and genotypic methods to identify sources of bacterial contamination resulting from point or non-point pollution. For this study Enterococcus sp. isolates originating from deer, bovine, gull, and human sources were examined using microarrays. Isolates were subjected to Box PCR amplification and the resulting amplification products labeled with Cy5. Fluorescent-labeled templates were hybridized to in-house constructed nonamer oligonucleotide microarrays consisting of 198 probes. Microarray hybridization profiles were obtained using the ArrayPro image analysis software. Principal Components Analysis (PCA and Hierarchical Cluster Analysis (HCA were compared for their ability to visually cluster microarray hybridization profiles based on the environmental source from which the Enterococcus sp. isolates originated. The PCA was visually superior at separating origin-specific clusters, even for as few as 3 factors. A Soft Independent Modeling (SIM classification confirmed the PCA, resulting in zero misclassifications using 5 factors for each class. The implication of these results for the application of random oligonucleotide microarrays for BST is that, given the reproducibility issues, factor-based variable selection such as in PCA and SIM greatly outperforms dendrogram-based similarity measures such as in HCA and K-Nearest Neighbor KNN.

  4. Assembly of Designed Oligonucleotides: a useful tool in synthetic biology for creating high-quality combinatorial DNA libraries.

    Science.gov (United States)

    Acevedo-Rocha, Carlos G; Reetz, Manfred T

    2014-01-01

    The method dubbed Assembly of Designed Oligonucleotides (ADO) is a powerful tool in synthetic biology to create combinatorial DNA libraries for gene, protein, metabolic, and genome engineering. In directed evolution of proteins, ADO benefits from using reduced amino acid alphabets for saturation mutagenesis and/or DNA shuffling, but all 20 canonical amino acids can be also used as building blocks. ADO is performed in a two-step reaction. The first involves a primer-free, polymerase cycling assembly or overlap extension PCR step using carefully designed overlapping oligonucleotides. The second step is a PCR amplification using the outer primers, resulting in a high-quality and bias-free double-stranded DNA library that can be assembled with other gene fragments and/or cloned into a suitable plasmid subsequently. The protocol can be performed in a few hours. In theory, neither the length of the DNA library nor the number of DNA changes has any limits. Furthermore, with the costs of synthetic DNA dropping every year, after an initial investment is made in the oligonucleotides, these can be exchanged for alternative ones with different sequences at any point in the process, fully exploiting the potential of creating highly diverse combinatorial libraries. In the example chosen here, we show the construction of a high-quality combinatorial ADO library targeting sixteen different codons simultaneously with nonredundant degenerate codons encoding various reduced alphabets of four amino acids along the heme region of the monooxygenase P450-BM3.

  5. Nanocrystalline silicon-based oligonucleotide chips.

    Science.gov (United States)

    Zhu, Z Q; Zhu, B; Zhang, J; Zhu, J Z; Fan, C H

    2007-04-15

    A novel oligonucleotide array sensor has been developed with nanocrystalline Si (ncSi) substrates. The ncSi was prepared by electrochemical etching technique. Our study indicated that both the binding capacity and the hybridization efficiency are dependent upon the particle size of ncSi. In contrary, the chips developed with Si substrates exhibit the lower binding capacity and hybridization efficiency. The improved performances of the sensor chips are attributed to the large specific surface area of ncSi compared to the existing conventional techniques. The sensor chips with the ncSi substrate of 13 nm-sized particle can be regenerated and reused for at least 12 times. The oligonucleotide array sensor also shows high stability, which can bear relatively the stringent conditions (e.g. 80 degrees C, 75% of relative humidity and 3.6 klx of irradiation).

  6. Math primer for engineers

    CERN Document Server

    Cryer, CW

    2014-01-01

    Mathematics and engineering are inevitably interrelated, and this interaction will steadily increase as the use of mathematical modelling grows. Although mathematicians and engineers often misunderstand one another, their basic approach is quite similar, as is the historical development of their respective disciplines. The purpose of this Math Primer is to provide a brief introduction to those parts of mathematics which are, or could be, useful in engineering, especially bioengineering. The aim is to summarize the ideas covered in each subject area without going into exhaustive detail. Formula

  7. Primer on Health Surveys

    Directory of Open Access Journals (Sweden)

    David L Nordstrom

    2012-06-01

    Full Text Available The aim of this paper is to introduce novice researchers to surveys as a method of data collection. It starts with the definition of a survey, its major purposes and types as well as changes in the goals surveys have helped to achieve over time. Advantages and disadvantages of surveys over population censuses and medical examinations are discussed. Approaches to questionnaire construction are introduced along with properties that questionnaires are evaluated for. Modes of administration, sample size issues, and data analysis approaches are also introduced. The primer is illustrated with examples of surveys conducted in different countries with various public health purposes.

  8. El primer virreinato americano

    Directory of Open Access Journals (Sweden)

    Cassá, Roberto

    2006-12-01

    Full Text Available This article explores the government of viceroy Christopher Columbus in the American territories. We return to the first Spanish settlement in Santo Domingo and the contradictions inherent to this expansionist proyect. The contradictions were part of the logic of the absolutist state and Columbus’ reaction against the controls imposed by the monarchs. Secondly, we look into the dificulties that the Admiral encountered to develop a mercantilist model. In this context, we examine the rationale behind the first government of the Indies and the features that defined the new West Indian society.

    El artículo trata sobre el gobierno de Cristóbal Colón en tierras americanas. Retomamos el tema del primer emplazamiento español en Santo Domingo y las contradicciones que tuvo aquel proyecto debido a la lógica del estado absolutista, a la ambición desmedida del descubridor y a su reacción ante los controles que desde un principio impusieron los monarcas. En un segundo momento analizamos las dificultades que encontró el Almirante para desarrollar un modelo mercantilista acorde a sus ideas y a los acuerdos a que llegó con la Corona. En ese contexto analizamos la lógica del primer gobierno colombinista en las Indias y los rasgos que definieron la nueva sociedad antillana.

  9. Universal and blocking primer mismatches limit the use of high-throughput DNA sequencing for the quantitative metabarcoding of arthropods.

    Science.gov (United States)

    Piñol, J; Mir, G; Gomez-Polo, P; Agustí, N

    2015-07-01

    The quantification of the biological diversity in environmental samples using high-throughput DNA sequencing is hindered by the PCR bias caused by variable primer-template mismatches of the individual species. In some dietary studies, there is the added problem that samples are enriched with predator DNA, so often a predator-specific blocking oligonucleotide is used to alleviate the problem. However, specific blocking oligonucleotides could coblock nontarget species to some degree. Here, we accurately estimate the extent of the PCR biases induced by universal and blocking primers on a mock community prepared with DNA of twelve species of terrestrial arthropods. We also compare universal and blocking primer biases with those induced by variable annealing temperature and number of PCR cycles. The results show that reads of all species were recovered after PCR enrichment at our control conditions (no blocking oligonucleotide, 45 °C annealing temperature and 40 cycles) and high-throughput sequencing. They also show that the four factors considered biased the final proportions of the species to some degree. Among these factors, the number of primer-template mismatches of each species had a disproportionate effect (up to five orders of magnitude) on the amplification efficiency. In particular, the number of primer-template mismatches explained most of the variation (~3/4) in the amplification efficiency of the species. The effect of blocking oligonucleotide concentration on nontarget species relative abundance was also significant, but less important (below one order of magnitude). Considering the results reported here, the quantitative potential of the technique is limited, and only qualitative results (the species list) are reliable, at least when targeting the barcoding COI region. © 2014 John Wiley & Sons Ltd.

  10. Introduction on Using the FastPCR Software and the Related Java Web Tools for PCR and Oligonucleotide Assembly and Analysis.

    Science.gov (United States)

    Kalendar, Ruslan; Tselykh, Timofey V; Khassenov, Bekbolat; Ramanculov, Erlan M

    2017-01-01

    This chapter introduces the FastPCR software as an integrated tool environment for PCR primer and probe design, which predicts properties of oligonucleotides based on experimental studies of the PCR efficiency. The software provides comprehensive facilities for designing primers for most PCR applications and their combinations. These include the standard PCR as well as the multiplex, long-distance, inverse, real-time, group-specific, unique, overlap extension PCR for multi-fragments assembling cloning and loop-mediated isothermal amplification (LAMP). It also contains a built-in program to design oligonucleotide sets both for long sequence assembly by ligase chain reaction and for design of amplicons that tile across a region(s) of interest. The software calculates the melting temperature for the standard and degenerate oligonucleotides including locked nucleic acid (LNA) and other modifications. It also provides analyses for a set of primers with the prediction of oligonucleotide properties, dimer and G/C-quadruplex detection, linguistic complexity as well as a primer dilution and resuspension calculator. The program consists of various bioinformatical tools for analysis of sequences with the GC or AT skew, CG% and GA% content, and the purine-pyrimidine skew. It also analyzes the linguistic sequence complexity and performs generation of random DNA sequence as well as restriction endonucleases analysis. The program allows to find or create restriction enzyme recognition sites for coding sequences and supports the clustering of sequences. It performs efficient and complete detection of various repeat types with visual display. The FastPCR software allows the sequence file batch processing that is essential for automation. The program is available for download at http://primerdigital.com/fastpcr.html , and its online version is located at http://primerdigital.com/tools/pcr.html .

  11. Application of nonsense-mediated primer exclusion (NOPE) for preparation of unique molecular barcoded libraries.

    Science.gov (United States)

    Shagin, Dmitriy A; Turchaninova, Maria A; Shagina, Irina A; Shugay, Mikhail; Zaretsky, Andrew R; Zueva, Olga I; Bolotin, Dmitriy A; Lukyanov, Sergey; Chudakov, Dmitriy M

    2017-06-05

    Recently we proposed efficient method to exclude undesirable primers at any stage of amplification reaction, here termed NOPE (NOnsense-mediated Primer Exclusion). According to this method, added oligonucleotide overlapping with the 3'-end of unwanted amplification primer (NOPE oligo) simultaneously provides a template for its elongation. This elongation disrupts specificity of unwanted primer, preventing its further participation in PCR. The suggested approach allows to rationally manage the course of PCR reactions in order to facilitate analysis of complex DNA mixtures as well as to perform multistage PCR bypassing intermediate purification steps. Here we apply NOPE method to DNA library preparation for the high-throughput sequencing (HTS) with the PCR-based introduction of unique molecular identifiers (UMI). We show that NOPE oligo efficiently neutralizes UMI-containing oligonucleotides after introduction of UMI into sample DNA molecules, thus allowing to proceed with further amplification steps without purification and associated loss of starting material. At the same time, NOPE oligo does not affect the efficiency of target PCR amplification. We describe a simple, robust and cheap modification of UMI-labeled HTS libraries preparation procedure, that allows to bypass purification step and thus to preserve starting material which may be limited, e.g. circulating tumor DNA, circulating fetal DNA, or small amounts of isolated cells of interest. Furthermore, demonstrated simplicity and robustness of NOPE method should make it popular in various PCR protocols.

  12. A primer on water

    Science.gov (United States)

    Leopold, Luna Bergere; Langbein, Walter Basil

    1960-01-01

    When you open the faucet you expect water to flow. And you expect it to flow night or day, summer or winter, whether you want to fill a glass or water the lawn. It should be clean and pure, without any odor.You have seen or read about places where the water doesn't have these qualities. You may have lived in a city where you were allowed to water the lawn only during a few hours of certain days. We know a large town where the water turns brown after every big rainstorm.Beginning shortly after World War II, large areas in the Southwestern United States had a 10-year drought, and newspapers published a lot of information about its effects. Some people say that the growing demand for water will cause serious shortages over much of the country in the next 10 to 40 years. But it has always been true that while water wells and springs dry up in some places, floods may be occurring in other places at the same time.Nearly every month news stories are published describing floods somewhere in the country. In fact, every year, on the average, 75,000 persons are forced from their homes by floods. In some years, as in 1951 when the lower Kansas River experienced a great flood, half a million people are affected. To understand the reasons for such recurring distress, it is necessary to know something about rivers and about the flat land or flood plain that borders the river.Interest in water and related problems is growing as our population increases and as the use of water becomes steadily greater. To help meet this heightened interest in general information about water and its use and control is the reason this primer was written. The primer is in two parts. The first part tells about hydrology, or the science that concerns the relation of water to our earth, and the second part describes the development of water supplies and the use of water. The Geological Survey is publishing this primer in nontechnical language in the hope that it will enable the general reader to

  13. Comparative analysis of measles virus RNA by oligonucleotide fingerprinting

    Energy Technology Data Exchange (ETDEWEB)

    Stephenson, J.R.; Meulen, V. ter (Wuerzburg Univ. (Germany, F.R.))

    1982-01-01

    Isolates from two cases of acute measles, one case of acute measles encephalitis and three patients with subacute sclerosing panencephalitis were compared. This comparison was based upon the electrophoretic analysis of T/sub 1/ oligonucleotides from single-stranded, full-length RNA isolated from cytoplasmic nucleocapsids. Although all viruses have oligonucleotides in common, each isolate generated a unique pattern of oligonucleotides. However, no group of oligonucleotides was observed which would allow differentiation between viruses isolated from acute infections and those isolated from CNS diseases; indicating that probably all measles viruses differ in their nucleotide sequence, regardless of origin.

  14. Metabolomics: A Primer.

    Science.gov (United States)

    Liu, Xiaojing; Locasale, Jason W

    2017-04-01

    Metabolomics generates a profile of small molecules that are derived from cellular metabolism and can directly reflect the outcome of complex networks of biochemical reactions, thus providing insights into multiple aspects of cellular physiology. Technological advances have enabled rapid and increasingly expansive data acquisition with samples as small as single cells; however, substantial challenges in the field remain. In this primer we provide an overview of metabolomics, especially mass spectrometry (MS)-based metabolomics, which uses liquid chromatography (LC) for separation, and discuss its utilities and limitations. We identify and discuss several areas at the frontier of metabolomics. Our goal is to give the reader a sense of what might be accomplished when conducting a metabolomics experiment, now and in the near future. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. A primer in combinatorics

    CERN Document Server

    Kheyfits, Alexander

    2010-01-01

    This textbook is devoted to Combinatorics and Graph Theory, which are cornerstones of Discrete Mathematics. Every section begins with simple model problems. Following their detailed analysis, the reader is led through the derivation of definitions, concepts and methods for solving typical problems. Theorems then are formulated, proved and illustrated by more problems of increasing difficulty. Topics covered include elementary combinatorial constructions, application to probability theory, introduction to graphs and trees with application to hierarchical clustering algorithms, more advanced counting techniques, and existence theorems in combinatorial analysis. The text systematically employs the basic language of set theory. This approach is often useful for solving combinatorial problems, especially problems where one has to identify some objects, and significantly reduces the number of the students' errors; it is demonstrated in the text on many examples. The textbook is suitable for undergraduate and entry-...

  16. Oligonucleotide probes for DNA fingerprinting in dogs.

    Science.gov (United States)

    Jung, M; Wilke, K; Geldermann, H

    1994-01-12

    Ten different oligonucleotide probes were evaluated for DNA fingerprinting in dogs. Seven probes are able to detect polymorphic bands. Probes (GT)(8) , (GTG)(5) and (GGAT)(4) are most informative for individual identification. The probabilities that two individuals from different breeds have the same DNA fingerprint pattern are 1.7 × 10(-7) , 5.5 × 10(-8) and 4.5 × 10(-6) , respectively. Using a combination of the three probes, paternity tests were performed with exclusion probabilities between 0.006% and 3%. ZUSAMMENFASSUNG: Oligonucleotid Sonden für DNA fingerprinting bei Hunden Zur Darstellung von DNA-Fingerprints beim Hund wurden zehn verschiedene Oligonukleotid-Sonden verglichen. Mit sieben Sonden konnten polymorphe Banden nachgewiesen werden. Die Sonden (GT)(8) , (GTG)(5) und (GGAT)(4) besaßen die größte Informativität für den Indentitätsnachweis. Die Wahrscheinlichkeit, daß zwei Individuen dieselben Fingerprint-Muster aufweisen, liegt bei 1,7 × 10(-7) , 5,5 × 10(-8) bzw. 4,5 × 10(-6) . Bei Verwendung einer Kombination der drei Sonden wurden Vaterschaftskontrollen mit Ausschlußwahrscheinlichkeiten zwischen 0,006% und 3 % erreicht. 1994 Blackwell Verlag GmbH.

  17. Catalysis with hierarchical zeolites

    DEFF Research Database (Denmark)

    Holm, Martin Spangsberg; Taarning, Esben; Egeblad, Kresten

    2011-01-01

    topic. Until now, the main reason for developing hierarchical zeolites has been to achieve heterogeneous catalysts with improved performance but this particular facet has not yet been reviewed in detail. Thus, the present paper summaries and categorizes the catalytic studies utilizing hierarchical...

  18. Investigations of oligonucleotide usage variance within and between prokaryotes

    DEFF Research Database (Denmark)

    Bohlin, J.; Skjerve, E.; Ussery, David

    2008-01-01

    Oligonucleotide usage in archaeal and bacterial genomes can be linked to a number of properties, including codon usage (trinucleotides), DNA base-stacking energy (dinucleotides), and DNA structural conformation (di-to tetranucleotides). We wanted to assess the statistical information potential...... of different DNA 'word-sizes' and explore how oligonucleotide frequencies differ in coding and non-coding regions. In addition, we used oligonucleotide frequencies to investigate DNA composition and how DNA sequence patterns change within and between prokaryotic organisms. Among the results found...... was that prokaryotic chromosomes can be described by hexanucleotide frequencies, suggesting that prokaryotic DNA is predominantly short range correlated, i. e., information in prokaryotic genomes is encoded in short oligonucleotides. Oligonucleotide usage varied more within AT-rich and host-associated genomes than...

  19. Photochemical immobilization of anthraquinone conjugated oligonucleotides and PCR amplicons on solid surfaces

    DEFF Research Database (Denmark)

    Koch, T; Jacobsen, N; Fensholdt, J

    2000-01-01

    Ligand immobilization on solid surfaces is an essential step in fields such as diagnostics, bio sensor manufacturing, and new material sciences in general. In this paper a photochemical approach based on anthraquinone as the chromophore is presented. Photochemical procedures offer special...... advantages as they are able to generate highly reactive species in an orientation specific manner. As presented here, anthraquinone (AQ) mediated covalent DNA immobilization appears to be superior to currently known procedures. A synthetic procedure providing AQ-phosphoramidites is presented. These reagents...... facilitate AQ conjugation during routine DNA synthesis, thus enabling the AQ-oligonucleotides to be immobilized in a very convenient and efficient manner. AQ-conjugated PCR primers can be used directly in PCR. When the PCR is performed in solution, the amplicons can be immobilized after the PCR. Moreover...

  20. Generation of Aptamers from A Primer-Free Randomized ssDNA Library Using Magnetic-Assisted Rapid Aptamer Selection

    Science.gov (United States)

    Tsao, Shih-Ming; Lai, Ji-Ching; Horng, Horng-Er; Liu, Tu-Chen; Hong, Chin-Yih

    2017-04-01

    Aptamers are oligonucleotides that can bind to specific target molecules. Most aptamers are generated using random libraries in the standard systematic evolution of ligands by exponential enrichment (SELEX). Each random library contains oligonucleotides with a randomized central region and two fixed primer regions at both ends. The fixed primer regions are necessary for amplifying target-bound sequences by PCR. However, these extra-sequences may cause non-specific bindings, which potentially interfere with good binding for random sequences. The Magnetic-Assisted Rapid Aptamer Selection (MARAS) is a newly developed protocol for generating single-strand DNA aptamers. No repeat selection cycle is required in the protocol. This study proposes and demonstrates a method to isolate aptamers for C-reactive proteins (CRP) from a randomized ssDNA library containing no fixed sequences at 5‧ and 3‧ termini using the MARAS platform. Furthermore, the isolated primer-free aptamer was sequenced and binding affinity for CRP was analyzed. The specificity of the obtained aptamer was validated using blind serum samples. The result was consistent with monoclonal antibody-based nephelometry analysis, which indicated that a primer-free aptamer has high specificity toward targets. MARAS is a feasible platform for efficiently generating primer-free aptamers for clinical diagnoses.

  1. DNA fingerprinting in cattle using oligonucleotide probes.

    Science.gov (United States)

    Buitkamp, J; Zischler, H; Epplen, J T; Geldermann, H

    1991-01-01

    Oligonucleotide probes specific for simple tandem repeat sequences produce individual specific DNA fingerprints in man and all animal species tested so far. Here 11 different synthetic probes were hybridized to bovine genomic DNAs which had been digested with the restriction endonucleases HinfI, AluI and HaeIII. Two of these probes gave DNA fingerprint patterns which were analysed for three German breeds. Different parameters were calculated, such as the average number of bands per individual or the probability of finding identical fingerprints in two unrelated individuals. The number of polymorphic bands varies from 11 to 23 in the different breeds and the probability of finding the same banding pattern in two unrelated individuals ranges from 1.5 x 10(-7) to 2.4 x 10(-7). Hence this DNA fingerprinting procedure allows precise identification of individuals. It is also a useful additional method for paternity testing in cattle.

  2. Antisense oligonucleotide therapeutics for inherited neurodegenerative diseases.

    Science.gov (United States)

    Southwell, Amber L; Skotte, Niels H; Bennett, C Frank; Hayden, Michael R

    2012-11-01

    The rising median age of our population and the age-dependent risk of neurodegeneration translate to exponentially increasing numbers of afflicted individuals in the coming years. Although symptomatic treatments are available for some neurodegenerative diseases, most are only moderately efficacious and are often associated with significant side effects. The development of small molecule, disease-modifying drugs has been hindered by complex pathogenesis and a failure to clearly define the rate-limiting steps in disease progression. An alternative approach is to directly target the mutant gene product or a defined causative protein. Antisense oligonucleotides (ASOs) - with their diverse functionality, high target specificity, and relative ease of central nervous system (CNS) delivery - are uniquely positioned as potential therapies for neurological diseases. Here we review the development of ASOs for the treatment of inherited neurodegenerative diseases. Copyright © 2012 Elsevier Ltd. All rights reserved.

  3. [The detection of plasmid pCAMBIA1301 in transgenic rice by arrayed primer extension].

    Science.gov (United States)

    Gao, Xiu-Li; Yang, Jian-Bo; Jing, Feng-Xiang; Zhao, Jian-Long

    2005-03-01

    Biochip technology which had emerged from the fusion of biotechnology and micro/nanofabrication technology at the end of 1980s has been widely used in life science, medicine, clinical diagnosis, drug development, agriculture, environmental protection and strategies. DNA microarray (also call gene chip, DNA chip), one kind of biochips, is small chip containing many oligonucleotide probes. It can hybridize with labelled sample, making it possible to detect large numbers of oligonucleotides at one time. So DNA microarray can overcome the disadvantage of traditional hybridization technology such as complexity, low automation, poor efficiency and quantity of molecules detected. This paper describes a new method to detect transgenic plant with gene chip. We have developed a novel arrayed-primer extension technique. It combines hybridization and PCR in one step, while ordinary DNA microarray needs two separate steps. Therefore our method provide a feasibility to detect long DNA fragment .

  4. Design and analysis of mismatch probes for long oligonucleotide microarrays

    Energy Technology Data Exchange (ETDEWEB)

    Deng, Ye; He, Zhili; Van Nostrand, Joy D.; Zhou, Jizhong

    2008-08-15

    Nonspecific hybridization is currently a major concern with microarray technology. One of most effective approaches to estimating nonspecific hybridizations in oligonucleotide microarrays is the utilization of mismatch probes; however, this approach has not been used for longer oligonucleotide probes. Here, an oligonucleotide microarray was constructed to evaluate and optimize parameters for 50-mer mismatch probe design. A perfect match (PM) and 28 mismatch (MM) probes were designed for each of ten target genes selected from three microorganisms. The microarrays were hybridized with synthesized complementary oligonucleotide targets at different temperatures (e.g., 42, 45 and 50 C). In general, the probes with evenly distributed mismatches were more distinguishable than those with randomly distributed mismatches. MM probes with 3, 4 and 5 mismatched nucleotides were differentiated for 50-mer oligonucleotide probes hybridized at 50, 45 and 42 C, respectively. Based on the experimental data generated from this study, a modified positional dependent nearest neighbor (MPDNN) model was constructed to adjust the thermodynamic parameters of matched and mismatched dimer nucleotides in the microarray environment. The MM probes with four flexible positional mismatches were designed using the newly established MPDNN model and the experimental results demonstrated that the redesigned MM probes could yield more consistent hybridizations. Conclusions: This study provides guidance on the design of MM probes for long oligonucleotides (e.g., 50 mers). The novel MPDNN model has improved the consistency for long MM probes, and this modeling method can potentially be used for the prediction of oligonucleotide microarray hybridizations.

  5. Antisense Oligonucleotide-Based Therapy for Neuromuscular Disease

    Directory of Open Access Journals (Sweden)

    Valentina Sardone

    2017-04-01

    Full Text Available Neuromuscular disorders such as Duchenne Muscular Dystrophy and Spinal Muscular Atrophy are neurodegenerative genetic diseases characterized primarily by muscle weakness and wasting. Until recently there were no effective therapies for these conditions, but antisense oligonucleotides, a new class of synthetic single stranded molecules of nucleic acids, have demonstrated promising experimental results and are at different stages of regulatory approval. The antisense oligonucleotides can modulate the protein expression via targeting hnRNAs or mRNAs and inducing interference with splicing, mRNA degradation, or arrest of translation, finally, resulting in rescue or reduction of the target protein expression. Different classes of antisense oligonucleotides are being tested in several clinical trials, and limitations of their clinical efficacy and toxicity have been reported for some of these compounds, while more encouraging results have supported the development of others. New generation antisense oligonucleotides are also being tested in preclinical models together with specific delivery systems that could allow some of the limitations of current antisense oligonucleotides to be overcome, to improve the cell penetration, to achieve more robust target engagement, and hopefully also be associated with acceptable toxicity. This review article describes the chemical properties and molecular mechanisms of action of the antisense oligonucleotides and the therapeutic implications these compounds have in neuromuscular diseases. Current strategies and carrier systems available for the oligonucleotides delivery will be also described to provide an overview on the past, present and future of these appealing molecules.

  6. Antisense oligonucleotide induction of progerin in human myogenic cells.

    Directory of Open Access Journals (Sweden)

    Yue-Bei Luo

    Full Text Available We sought to use splice-switching antisense oligonucleotides to produce a model of accelerated ageing by enhancing expression of progerin, translated from a mis-spliced lamin A gene (LMNA transcript in human myogenic cells. The progerin transcript (LMNA Δ150 lacks the last 150 bases of exon 11, and is translated into a truncated protein associated with the severe premature ageing disease, Hutchinson-Gilford progeria syndrome (HGPS. HGPS arises from de novo mutations that activate a cryptic splice site in exon 11 of LMNA and result in progerin accumulation in tissues of mesodermal origin. Progerin has also been proposed to play a role in the 'natural' ageing process in tissues. We sought to test this hypothesis by producing a model of accelerated muscle ageing in human myogenic cells. A panel of splice-switching antisense oligonucleotides were designed to anneal across exon 11 of the LMNA pre-mRNA, and these compounds were transfected into primary human myogenic cells. RT-PCR showed that the majority of oligonucleotides were able to modify LMNA transcript processing. Oligonucleotides that annealed within the 150 base region of exon 11 that is missing in the progerin transcript, as well as those that targeted the normal exon 11 donor site induced the LMNA Δ150 transcript, but most oligonucleotides also generated variable levels of LMNA transcript missing the entire exon 11. Upon evaluation of different oligomer chemistries, the morpholino phosphorodiamidate oligonucleotides were found to be more efficient than the equivalent sequences prepared as oligonucleotides with 2'-O-methyl modified bases on a phosphorothioate backbone. The morpholino oligonucleotides induced nuclear localised progerin, demonstrated by immunostaining, and morphological nuclear changes typical of HGPS cells. We show that it is possible to induce progerin expression in myogenic cells using splice-switching oligonucleotides to redirect splicing of LMNA. This may offer a model

  7. Introduction into Hierarchical Matrices

    KAUST Repository

    Litvinenko, Alexander

    2013-12-05

    Hierarchical matrices allow us to reduce computational storage and cost from cubic to almost linear. This technique can be applied for solving PDEs, integral equations, matrix equations and approximation of large covariance and precision matrices.

  8. Programming with Hierarchical Maps

    DEFF Research Database (Denmark)

    Ørbæk, Peter

    This report desribes the hierarchical maps used as a central data structure in the Corundum framework. We describe its most prominent features, ague for its usefulness and briefly describe some of the software prototypes implemented using the technology....

  9. Micromechanics of hierarchical materials

    DEFF Research Database (Denmark)

    Mishnaevsky, Leon, Jr.

    2012-01-01

    A short overview of micromechanical models of hierarchical materials (hybrid composites, biomaterials, fractal materials, etc.) is given. Several examples of the modeling of strength and damage in hierarchical materials are summarized, among them, 3D FE model of hybrid composites...... with nanoengineered matrix, fiber bundle model of UD composites with hierarchically clustered fibers and 3D multilevel model of wood considered as a gradient, cellular material with layered composite cell walls. The main areas of research in micromechanics of hierarchical materials are identified, among them......, the investigations of the effects of load redistribution between reinforcing elements at different scale levels, of the possibilities to control different material properties and to ensure synergy of strengthening effects at different scale levels and using the nanoreinforcement effects. The main future directions...

  10. Oligonucleotides Containing Aminated 2′-Amino-LNA Nucleotides

    DEFF Research Database (Denmark)

    Lou, Chenguang; Samuelsen, Simone V.; Christensen, Niels Johan

    2017-01-01

    Mono- and diaminated 2′-amino-LNA monomers were synthesized and introduced into oligonucleotides. Each modification imparts significant stabilization of nucleic acid duplexes and triplexes, excellent sequence selectivity, and significant nuclease resistance. Molecular modeling suggested that stru...... that structural stabilization occurs via intrastrand electrostatic attraction between the protonated amino groups of the aminated 2′-amino-LNA monomers and the host oligonucleotide backbone.......Mono- and diaminated 2′-amino-LNA monomers were synthesized and introduced into oligonucleotides. Each modification imparts significant stabilization of nucleic acid duplexes and triplexes, excellent sequence selectivity, and significant nuclease resistance. Molecular modeling suggested...

  11. Modern methods for the synthesis of peptide-oligonucleotide conjugates

    Science.gov (United States)

    Zubin, Evgenii M.; Romanova, Elena A.; Oretskaya, Tat'yana S.

    2002-03-01

    The published data on the methods of chemical solution and solid-phase synthesis of peptide-oligonucleotide conjugates are reviewed. The known methods are systematised and their advantages and disadvantages are considered. The approaches to the solution synthesis of peptide-oligonucleotide conjugates are systematised according to the type of chemical bonds between the fragments, whereas those to the solid-phase synthesis are classified according to the procedure used for the preparation of conjugates, viz., stepwise elongation of oligonucleotide and peptide chains on the same polymeric support or solid-phase condensation of two presynthesised fragments. The bibliography includes 141 references.

  12. Optical Characterization of Oligonucleotide DNA Influenced by Magnetic Fields

    Directory of Open Access Journals (Sweden)

    Seyedeh Maryam Banihashemian

    2013-09-01

    Full Text Available UV-VIS spectroscopic analysis of oligonucleotide DNA exposed to different magnetic fields was performed in order to investigate the relationship between DNA extinction coefficients and optical parameters according to magnetic-field strength. The results with the oligonucleotides adenine-thymine 100 mer (AT-100 DNA and cytosine-guanine 100 mer (CG-100 DNA indicate that the magnetic field influences DNA molar extinction coefficients and refractive indexes. The imaginary parts of the refractive index and molar extinction coefficients of the AT-100 and CG-100 DNA decreased after exposure to a magnetic field of 750 mT due to cleavage of the DNA oligonucleotides into smaller segments.

  13. RTPrimerDB: the real-time PCR primer and probe database, major update 2006.

    Science.gov (United States)

    Pattyn, Filip; Robbrecht, Piet; De Paepe, Anne; Speleman, Frank; Vandesompele, Jo

    2006-01-01

    The RTPrimerDB (http://medgen.ugent.be/rtprimerdb) project provides a freely accessible data retrieval system and an in silico assay evaluation pipeline for real-time quantitative PCR assays. Over the last year the number of user submitted assays has grown to 3500. Data conveyance from Entrez Gene by establishing an assay-to-gene relationship enables the addition of new primer assays for one of the 1.5 million different genes from 2300 species stored in the system. Easy access to the primer and probe data is possible by using multiple search criteria. Assay reports contain gene information, assay details (such as oligonucleotide sequences, detection chemistry and reaction conditions), publication information, users' experimental evaluation feedback and submitter's contact details. Gene expression assays are extended with a scalable assay viewer that provides detailed information on the alignment of primer and probe sequences on the known transcript variants of a gene, along with Single Nucleotide Polymorphisms (SNP) positions and peptide domain information. Furthermore, an mfold module is implemented to predict the secondary structure of the amplicon sequence, as this has been reported to impact the efficiency of the PCR. RTPrimerDB is also extended with an in silico analysis pipeline to streamline the evaluation of custom designed primer and probe sequences prior to ordering and experimental evaluation. In a secured environment, the pipeline performs automated BLAST specificity searches, mfold secondary structure prediction, SNP or plain sequence error identification, and graphical visualization of the aligned primer and probe sequences on the target gene.

  14. Sequence-dependent theory of oligonucleotide hybridization kinetics

    Energy Technology Data Exchange (ETDEWEB)

    Marimuthu, Karthikeyan [Department of Chemical Engineering and Center for Advanced Process Decision-Making, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213 (United States); Chakrabarti, Raj, E-mail: raj@pmc-group.com, E-mail: rajc@andrew.cmu.edu [Department of Chemical Engineering and Center for Advanced Process Decision-Making, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213 (United States); Division of Fundamental Research, PMC Advanced Technology, Mount Laurel, New Jersey 08054 (United States)

    2014-05-07

    A theoretical approach to the prediction of the sequence and temperature-dependent rate constants for oligonucleotide hybridization reactions has been developed based on the theory of relaxation kinetics. One-sided and two-sided melting reaction mechanisms for oligonucleotide hybridization reactions have been considered, analyzed, modified, and compared to select a physically consistent as well as robust model for prediction of the relaxation times of DNA hybridization reactions that agrees with the experimental evidence. The temperature- and sequence-dependent parameters of the proposed model have been estimated using available experimental data. The relaxation time model that we developed has been combined with the nearest neighbor model of hybridization thermodynamics to estimate the temperature- and sequence-dependent rate constants of an oligonucleotide hybridization reaction. The model-predicted rate constants are compared to experimentally determined rate constants for the same oligonucleotide hybridization reactions. Finally, we consider a few important applications of kinetically controlled DNA hybridization reactions.

  15. Oligonucleotide-based theranostic nanoparticles in cancer therapy.

    Science.gov (United States)

    Shahbazi, Reza; Ozpolat, Bulent; Ulubayram, Kezban

    2016-05-01

    Theranostic approaches, combining the functionality of both therapy and imaging, have shown potential in cancer nanomedicine. Oligonucleotides such as small interfering RNA and microRNA, which are powerful therapeutic agents, have been effectively employed in theranostic systems against various cancers. Nanoparticles are used to deliver oligonucleotides into tumors by passive or active targeting while protecting the oligonucleotides from nucleases in the extracellular environment. The use of quantum dots, iron oxide nanoparticles and gold nanoparticles and tagging with contrast agents, like fluorescent dyes, optical or magnetic agents and various radioisotopes, has facilitated early detection of tumors and evaluation of therapeutic efficacy. In this article, we review the advantages of theranostic applications in cancer therapy and imaging, with special attention to oligonucleotide-based therapeutics.

  16. Sequence-dependent theory of oligonucleotide hybridization kinetics.

    Science.gov (United States)

    Marimuthu, Karthikeyan; Chakrabarti, Raj

    2014-05-07

    A theoretical approach to the prediction of the sequence and temperature-dependent rate constants for oligonucleotide hybridization reactions has been developed based on the theory of relaxation kinetics. One-sided and two-sided melting reaction mechanisms for oligonucleotide hybridization reactions have been considered, analyzed, modified, and compared to select a physically consistent as well as robust model for prediction of the relaxation times of DNA hybridization reactions that agrees with the experimental evidence. The temperature- and sequence-dependent parameters of the proposed model have been estimated using available experimental data. The relaxation time model that we developed has been combined with the nearest neighbor model of hybridization thermodynamics to estimate the temperature- and sequence-dependent rate constants of an oligonucleotide hybridization reaction. The model-predicted rate constants are compared to experimentally determined rate constants for the same oligonucleotide hybridization reactions. Finally, we consider a few important applications of kinetically controlled DNA hybridization reactions.

  17. Oligonucleotide probes for DNA fingerprinting in horses.

    Science.gov (United States)

    Wilke, K; Weimann, M; Jung, M; Geldermann, H

    1993-01-12

    10 different oligonucleotide probes were evaluated for DNA fingerprinting in horses. Five probes were able to detect polymorphic bands. The probes (GT)(8) , (GTG)(5) and (GGAT)(4) are most informative for individual identification and were used to analyze a population of Hannoveranian horses. The probability that two individuals have the same DNA fingerprint pattern is 1.2 × 10(-8) , 5.2 × 10(-10) and 1.5 × 10(-7) respectively. Using a combination of the three probes, paternity tests were performed with exclusion probabilities between 0.08% and 4%. ZUSAMMENFASSUNG: Oligonukleotide-Sonden für DNS-Fingerprints von Pferden Zur Darstellung von DNA-Fingerprints beim Pferd wurden zehn verschiedene Oligonukleotid-Sonden verglichen. Mit fünf Sonden konnten polymorphe Banden nachgewiesen werden. Die Sonden (GT)(8) , (GTG)(5) und (GGAT)(4) besaßen die größte Informativität für den Identitätsnachweis und wurden für die Analyse einer Population von Hannoverschen Pferden benutzt. Die Wahrscheinlichkeit, daß zwei Individuen dieselben Fingerprint-Muster aufweisen, liegt bei 1,2 × 10(-8) , 5,2 × 10(-10) bzw. 1,5 × 10(-7) . Bei Verwendung einer Kombination der drei Sonden wurden Vaterschaftskontrollen mit Ausschlußwahrscheinlichkeiten zwischen 0,08% und 4% erreicht. 1993 Blackwell Verlag GmbH.

  18. Hole hopping rates in single strand oligonucleotides

    Energy Technology Data Exchange (ETDEWEB)

    Borrelli, Raffaele [Dipartimento di Scienze Agrarie, Forestali e Alimentari, Università di Torino, Largo Paolo Braccini 2, I-10095 Grugliasco, TO (Italy); Capobianco, Amedeo [Dipartimento di Chimica e Biologia, Università di Salerno, Via Giovanni Paolo II, I-84084 Fisciano, SA (Italy); Peluso, Andrea, E-mail: apeluso@unisa.it [Dipartimento di Chimica e Biologia, Università di Salerno, Via Giovanni Paolo II, I-84084 Fisciano, SA (Italy)

    2014-08-31

    Highlights: • DNA hole transfer rates have been computed. • Delocalized adenine domains significantly affect hole transfer rates in DNA. • Franck–Condon weighted density of state from DFT normal modes. • DNA application in molecular electronics. - Abstract: The rates of hole transfer between guanine and adenine in single strand DNA have been evaluated by using Fermi’s golden rule and Kubo’s generating function approach for the Franck–Condon weighted density of states. The whole sets of the normal modes and vibrational frequencies of the two nucleobases, obtained at DFT/B3LYP level of calculation, have been considered in computations. The results show that in single strand the pyramidalization/planarization mode of the amino groups of both nucleobases plays the major role. At room temperature, the Franck–Condon density of states extends over a wide range of hole site energy difference, 0–1 eV, giving some hints about the design of oligonucleotides of potential technological interest.

  19. Ferrocene conjugated oligonucleotide for electrochemical detection of DNA base mismatch.

    Science.gov (United States)

    Hasegawa, Yusuke; Takada, Tadao; Nakamura, Mitsunobu; Yamana, Kazushige

    2017-08-01

    We describe the synthesis, binding, and electrochemical properties of ferrocene-conjugated oligonucleotides (Fc-oligos). The key step for the preparation of Fc-oligos contains the coupling of vinylferrocene to 5-iododeoxyuridine via Heck reaction. The Fc-conjugated deoxyuridine phosphoramidite was used in the Fc-oligonucleotide synthesis. We show that thiol-modified Fc-oligos deposited onto gold electrodes possess potential ability in electrochemical detection of DNA base mismatch. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Spotted cotton oligonucleotide microarrays for gene expression analysis

    Directory of Open Access Journals (Sweden)

    Nettleton Dan

    2007-03-01

    Full Text Available Abstract Background Microarrays offer a powerful tool for diverse applications plant biology and crop improvement. Recently, two comprehensive assemblies of cotton ESTs were constructed based on three Gossypium species. Using these assemblies as templates, we describe the design and creation and of a publicly available oligonucleotide array for cotton, useful for all four of the cultivated species. Results Synthetic oligonucleotide probes were generated from exemplar sequences of a global assembly of 211,397 cotton ESTs derived from >50 different cDNA libraries representing many different tissue types and tissue treatments. A total of 22,787 oligonucleotide probes are included on the arrays, optimized to target the diversity of the transcriptome and previously studied cotton genes, transcription factors, and genes with homology to Arabidopsis. A small portion of the oligonucleotides target unidentified protein coding sequences, thereby providing an element of gene discovery. Because many oligonucleotides were based on ESTs from fiber-specific cDNA libraries, the microarray has direct application for analysis of the fiber transcriptome. To illustrate the utility of the microarray, we hybridized labeled bud and leaf cDNAs from G. hirsutum and demonstrate technical consistency of results. Conclusion The cotton oligonucleotide microarray provides a reproducible platform for transcription profiling in cotton, and is made publicly available through http://cottonevolution.info.

  1. Parallel hierarchical radiosity rendering

    Energy Technology Data Exchange (ETDEWEB)

    Carter, Michael [Iowa State Univ., Ames, IA (United States)

    1993-07-01

    In this dissertation, the step-by-step development of a scalable parallel hierarchical radiosity renderer is documented. First, a new look is taken at the traditional radiosity equation, and a new form is presented in which the matrix of linear system coefficients is transformed into a symmetric matrix, thereby simplifying the problem and enabling a new solution technique to be applied. Next, the state-of-the-art hierarchical radiosity methods are examined for their suitability to parallel implementation, and scalability. Significant enhancements are also discovered which both improve their theoretical foundations and improve the images they generate. The resultant hierarchical radiosity algorithm is then examined for sources of parallelism, and for an architectural mapping. Several architectural mappings are discussed. A few key algorithmic changes are suggested during the process of making the algorithm parallel. Next, the performance, efficiency, and scalability of the algorithm are analyzed. The dissertation closes with a discussion of several ideas which have the potential to further enhance the hierarchical radiosity method, or provide an entirely new forum for the application of hierarchical methods.

  2. Vygotsky on Education Primer. Peter Lang Primer. Volume 30

    Science.gov (United States)

    Lake, Robert

    2012-01-01

    The "Vygotsky on Education Primer" serves as an introduction to the life and work of the Russian psychologist Lev Vygotsky. Even though he died almost eighty years ago, his life's work remains both relevant and significant to the field of education today. This book examines Vygotsky's emphasis on the role of cultural and historical context in…

  3. Hierarchical Network Design

    DEFF Research Database (Denmark)

    Thomadsen, Tommy

    2005-01-01

    design. The papers have all been submitted for journals, and except for two papers, are awaiting review. The papers are mostly concerned with optimal methods and, in a few cases, heuristics for designing hierarchical and ring networks. All papers develop bounds which are used in the optimal methods...... danne grundlag for et studie af design af hierarkiske netværk. Afhandlings vigtigste bidrag best ar af syv artikler, der er inkluderet i appendiks. Artiklerne handler om design af hierarkisk netværk og ring netværk. Artiklerne er alle indsendt til videnskablige journaler og afventer bedømmelse, bortset......Communication networks are immensely important today, since both companies and individuals use numerous services that rely on them. This thesis considers the design of hierarchical (communication) networks. Hierarchical networks consist of layers of networks and are well-suited for coping...

  4. DNA Extraction and Primer Selection

    DEFF Research Database (Denmark)

    Karst, Søren Michael; Nielsen, Per Halkjær; Albertsen, Mads

    Talk regarding pitfalls in DNA extraction and 16S amplicon primer choice when performing community analysis of complex microbial communities. The talk was a part of Workshop 2 "Principles, Potential, and Limitations of Novel Molecular Methods in Water Engineering; from Amplicon Sequencing to -omics...

  5. A Hearing Aid Primer 1

    Science.gov (United States)

    Yetter, Carol J.

    2009-01-01

    This hearing aid primer is designed to define the differences among the three levels of hearing instrument technology: conventional analog circuit technology (most basic), digitally programmable/analog circuit technology (moderately advanced), and fully digital technology (most advanced). Both moderate and advanced technologies mean that hearing…

  6. Freshwater Wetlands: A Citizen's Primer.

    Science.gov (United States)

    Catskill Center for Conservation and Development, Inc., Hobart, NY.

    The purpose of this "primer" for the general public is to describe the general characteristics of wetlands and how wetland alteration adversely affects the well-being of humans. Particular emphasis is placed on wetlands in New York State and the northeast. Topics discussed include wetland values, destruction of wetlands, the costs of…

  7. Biominetic High Density Lipoproteins for the Delivery of Therapeutic Oligonucleotides

    Science.gov (United States)

    Tripathy, Sushant

    Advances in nanotechnology have brought about novel inorganic and hybrid nanoparticles with unique physico-chemical properties that make them suitable for a broad range of applications---from nano-circuitry to drug delivery. A significant part of those advancements have led to ground-breaking discoveries that have changed the approaches to formulation of therapeutics against diseases, such as cancer. Now-a-days the focus does not lie solely on finding a candidate small-molecule therapeutic with minimal adverse effects, but researchers are looking up to nanoparticles to improve biodistribution and biocompatibility profile of clinically proven therapeutics. The plethora of conjugation chemistries offered by currently extant inorganic nanoparticles have, in recent years, led to great leaps in the field of biomimicry---a modality that promises high biocompatibility. Further, in the pursuit of highly specific therapeutic molecules, researchers have turned to silencing oligonucleotides and some have already brought together the strengths of nanoparticles and silencing oligonucleotides in search of an efficacious therapy for cancer with minimal adverse effects. This dissertation work focuses on such a biomimetic platform---a gold nanoparticle based high density lipoprotein biomimetic (HDL NP), for the delivery of therapeutic oligonucleotides. The first chapter of this body of work introduces the molecular target of the silencing oligonucleotides---VEGFR2, and its role in the progression of solid tumor cancers. The background information also covers important aspects of natural high density lipoproteins (HDL), especially their innate capacity to bind and deliver exogenous and endogenous silencing oligonucleotides to tissues that express their high affinity receptor SRB1. We subsequently describe the synthesis of the biomimetic HDL NP and its oligonucleotide conjugates, and establish their biocompatibility. Further on, experimental data demonstrate the efficacy of silencing

  8. Hierarchical Porous Structures

    Energy Technology Data Exchange (ETDEWEB)

    Grote, Christopher John [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2016-06-07

    Materials Design is often at the forefront of technological innovation. While there has always been a push to generate increasingly low density materials, such as aero or hydrogels, more recently the idea of bicontinuous structures has gone more into play. This review will cover some of the methods and applications for generating both porous, and hierarchically porous structures.

  9. Microparticles with hierarchical porosity

    Science.gov (United States)

    Petsev, Dimiter N; Atanassov, Plamen; Pylypenko, Svitlana; Carroll, Nick; Olson, Tim

    2012-12-18

    The present disclosure provides oxide microparticles with engineered hierarchical porosity and methods of manufacturing the same. Also described are structures that are formed by templating, impregnating, and/or precipitating the oxide microparticles and method for forming the same. Suitable applications include catalysts, electrocatalysts, electrocatalysts support materials, capacitors, drug delivery systems, sensors and chromatography.

  10. Hierarchical models and functional traits

    NARCIS (Netherlands)

    van Loon, E.E.; Shamoun-Baranes, J.; Sierdsema, H.; Bouten, W.; Cramer, W.; Badeck, F.; Krukenberg, B.; Klotz, S.; Kühn, I.; Schweiger, O.; Böhning-Gaese, K.; Schaefer, H.-C.; Kissling, D.; Brandl, R.; Brändle, M.; Fricke, R.; Leuschner, C.; Buschmann, H.; Köckermann, B.; Rose, L.

    2006-01-01

    Hierarchical models for animal abundance prediction are conceptually elegant. They are generally more parsimonous than non-hierarchical models derived from the same data, give relatively robust predictions and automatically provide consistent output at multiple (spatio-temporal) scales. Another

  11. Synthesis of oligonucleotides containing novel G-clamp analogue with C8-tethered group in phenoxazine ring: Implication to qPCR detection of the low-copy Kemerovo virus dsRNA.

    Science.gov (United States)

    Varizhuk, Anna M; Zatsepin, Timofei S; Golovin, Andrey V; Belyaev, Evgeny S; Kostyukevich, Yury I; Dedkov, Vladimir G; Shipulin, German A; Shpakovski, George V; Aralov, Andrey V

    2017-07-15

    Nowadays modified oligonucleotides are widely used in diagnostics and as novel therapeutics. Introduction of modified or unnatural residues into oligonucleotides allows fine tuning of their binding properties to complementary nucleic acids and leads to improved stability both in vitro and in vivo. Previously it was demonstrated that insertion of phenoxazine nucleotides with various groups in C9-position into oligonucleotides leads to a significant increase of duplex stability with complementary DNA and RNA. Here the synthesis of a novel G-clamp nucleoside analogue (G8AE-clamp) bearing 2-aminoethyl tether at C8-atom is presented. Introduction of such modified residues into oligonucleotides lead to enhanced specificity of duplex formation towards complementary DNA and RNA targets with increased thermal and 3'-exonuclease stability. According to CD-spectroscopy studies G8AE-clamp does not substantially disrupt helix geometry. Primers containing G8AE-clamp demonstrated superior sensitivity in qPCR detection of dsRNA of Kemerovo virus in comparison to native oligonucleotides. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Bioinformatic tools for PCR Primer design

    African Journals Online (AJOL)

    ES

    choosing the most efficient way to design a new specific-primer by applying current publicly available links and Web services. Also, the purpose here is to provide general recommendations for the design and use of PCR primers. Key words: Bio-computing, primer design, web-based resources. INTRODUCTION. In the last ...

  13. Delivery of RNAi-Based Oligonucleotides by Electropermeabilization

    Directory of Open Access Journals (Sweden)

    Muriel Golzio

    2013-04-01

    Full Text Available For more than a decade, understanding of RNA interference (RNAi has been a growing field of interest. The potent gene silencing ability that small oligonucleotides have offers new perspectives for cancer therapeutics. One of the present limits is that many biological barriers exist for their efficient delivery into target cells or tissues. Electropermeabilization (EP is one of the physical methods successfully used to transfer small oligonucleotides into cells or tissues. EP consists in the direct application of calibrated electric pulses to cells or tissues that transiently permeabilize the plasma membranes, allowing efficient in vitro and in vivo. cytoplasmic delivery of exogenous molecules. The present review reports on the type of therapeutic RNAi-based oligonucleotides that can be electrotransferred, the mechanism(s of their electrotransfer and the technical settings for pre-clinical purposes.

  14. Current progress on aptamer-targeted oligonucleotide therapeutics

    Science.gov (United States)

    Dassie, Justin P; Giangrande, Paloma H

    2014-01-01

    Exploiting the power of the RNAi pathway through the use of therapeutic siRNA drugs has remarkable potential for treating a vast array of human disease conditions. However, difficulties in delivery of these and similar nucleic acid-based pharmacological agents to appropriate organs or tissues, remains a major impediment to their broad clinical application. Synthetic nucleic acid ligands (aptamers) have emerged as effective delivery vehicles for therapeutic oligonucleotides, including siRNAs. In this review, we summarize recent attractive developments in creatively employing cell-internalizing aptamers to deliver therapeutic oligonucleotides (e.g., siRNAs, miRNAs, anti-miRs and antisense oligos) to target cells. We also discuss advancements in aptamer-siRNA chimera technology, as well as, aptamer-functionalized nanoparticles for siRNA delivery. In addition, the challenges and future prospects of aptamer-targeted oligonucleotide drugs for clinical translation are further highlighted. PMID:24304250

  15. Challenges to oligonucleotides-based therapeutics for Duchenne muscular dystrophy

    Directory of Open Access Journals (Sweden)

    Goyenvalle Aurélie

    2011-02-01

    Full Text Available Abstract Antisense oligonucleotides are short nucleic acids designed to bind to specific messenger RNAs in order to modulate splicing patterns or inhibit protein translation. As such, they represent promising therapeutic tools for many disorders and have been actively developed for more than 20 years as a form of molecular medicine. Although significant progress has been made in developing these agents as drugs, they are yet not recognized as effective therapeutics and several hurdles remain to be overcome. Within the last few years, however, the prospect of successful oligonucleotides-based therapies has moved a step closer, in particular for Duchenne muscular dystrophy. Clinical trials have recently been conducted for this myopathy, where exon skipping is being used to achieve therapeutic outcomes. In this review, the recent developments and clinical trials using antisense oligonucleotides for Duchenne muscular dystrophy are discussed, with emphasis on the challenges ahead for this type of therapy, especially with regards to delivery and regulatory issues.

  16. Oligonucleotide Therapy for Obstructive and Restrictive Respiratory Diseases

    Directory of Open Access Journals (Sweden)

    Wupeng Liao

    2017-01-01

    Full Text Available Inhaled oligonucleotide is an emerging therapeutic modality for various common respiratory diseases, including obstructive airway diseases like asthma and chronic obstructive pulmonary disease (COPD and restrictive airway diseases like idiopathic pulmonary fibrosis (IPF. The advantage of direct accessibility for oligonucleotide molecules to the lung target sites, bypassing systemic administration, makes this therapeutic approach promising with minimized potential systemic side effects. Asthma, COPD, and IPF are common chronic respiratory diseases, characterized by persistent airway inflammation and dysregulated tissue repair and remodeling, although each individual disease has its unique etiology. Corticosteroids have been widely prescribed for the treatment of asthma, COPD, and IPF. However, the effectiveness of corticosteroids as an anti-inflammatory drug is limited by steroid resistance in severe asthma, the majority of COPD cases, and pulmonary fibrosis. There is an urgent medical need to develop target-specific drugs for the treatment of these respiratory conditions. Oligonucleotide therapies, including antisense oligonucleotide (ASO, small interfering RNA (siRNA, and microRNA (miRNA are now being evaluated both pre-clinically and clinically as potential therapeutics. The mechanisms of action of ASO and siRNA are highly target mRNA specific, ultimately leading to target protein knockdown. miRNA has both biomarker and therapeutic values, and its knockdown by a miRNA antagonist (antagomir has a broader but potentially more non-specific biological outcome. This review will compile the current findings of oligonucleotide therapeutic targets, verified in various respiratory disease models and in clinical trials, and evaluate different chemical modification approaches to improve the stability and potency of oligonucleotides for the treatment of respiratory diseases.

  17. Immunomodulation of hematological malignancies using oligonucleotides based-nanomedicines.

    Science.gov (United States)

    Hazan-Halevy, Inbal; Landesman-Milo, Dalit; Rosenblum, Daniel; Mizrahy, Shoshy; Ng, Brandon D; Peer, Dan

    2016-12-28

    Hematological malignancies are a group of diseases characterized by clonal proliferation of blood-forming cells. Malignant blood cells are classified as myeloid or lymphoid cells depending on their stem cell origin. Lymphoid malignancies are characterized by lymphocyte accumulation in the blood stream, in the bone marrow, or in lymphatic nodes and organs. Several of these diseases are associated with chromosomal translocations, which cause gene fusion and amplification of expression, while others are characterized with aberrant expression of oncogenes. Overall, these genes play a major role in development and maintenance of malignant clones. The discovery of antisense oligonucleotides and RNA interference (RNAi) mechanisms offer new tools to specifically manipulate gene expression. Systemic delivery of inhibitory oligonucleotides molecules for manipulation of gene expression in lymphocytes holds a great potential for facilitating the development of an oligonucleotides -based therapy platform for lymphoid blood cancer. However, lymphocytes are among the most difficult targets for oligonucleotides delivery, as they are resistant to conventional transfection reagents and are dispersed throughout the body, making it difficult to successfully localize or deliver oligonucleotides payloads via systemic administration. In this review, we will survey the latest progress in the field of oligonucleotides based nanomedicine in the heterogeneous group of hematological malignancies with special emphasis on RNA based strategies. We will describe the most advanced non-viral nanocarriers for RNA delivery to malignant blood cells. We will also discuss targeted strategies for cell specific delivery of RNA molecules using nanoparticles and the therapeutic benefit of manipulating gene function in hematological malignancies. Finally, we will focus on the ex vivo, in vivo, and clinical trial strategies, that are currently under development in hematological malignancies - strategies that

  18. Hierarchical species distribution models

    Science.gov (United States)

    Hefley, Trevor J.; Hooten, Mevin B.

    2016-01-01

    Determining the distribution pattern of a species is important to increase scientific knowledge, inform management decisions, and conserve biodiversity. To infer spatial and temporal patterns, species distribution models have been developed for use with many sampling designs and types of data. Recently, it has been shown that count, presence-absence, and presence-only data can be conceptualized as arising from a point process distribution. Therefore, it is important to understand properties of the point process distribution. We examine how the hierarchical species distribution modeling framework has been used to incorporate a wide array of regression and theory-based components while accounting for the data collection process and making use of auxiliary information. The hierarchical modeling framework allows us to demonstrate how several commonly used species distribution models can be derived from the point process distribution, highlight areas of potential overlap between different models, and suggest areas where further research is needed.

  19. Inhibition of microRNA with antisense oligonucleotides.

    Science.gov (United States)

    Esau, Christine C

    2008-01-01

    Antisense inhibition of microRNA (miRNA) function has been an important tool for uncovering miRNA biology. Chemical modification of anti-miRNA oligonucleotides (AMOs) is necessary to improve affinity for target miRNA, stabilize the AMO to nuclease degradation, and to promote tissue uptake for in vivo delivery. Here I summarize the work done to evaluate the effectiveness of various chemically modified AMOs for use in cultured cells and rodent models, and outline important issues to consider when inhibiting miRNAs with antisense oligonucleotides.

  20. Hierarchically Structured Electrospun Fibers

    Directory of Open Access Journals (Sweden)

    Nicole E. Zander

    2013-01-01

    Full Text Available Traditional electrospun nanofibers have a myriad of applications ranging from scaffolds for tissue engineering to components of biosensors and energy harvesting devices. The generally smooth one-dimensional structure of the fibers has stood as a limitation to several interesting novel applications. Control of fiber diameter, porosity and collector geometry will be briefly discussed, as will more traditional methods for controlling fiber morphology and fiber mat architecture. The remainder of the review will focus on new techniques to prepare hierarchically structured fibers. Fibers with hierarchical primary structures—including helical, buckled, and beads-on-a-string fibers, as well as fibers with secondary structures, such as nanopores, nanopillars, nanorods, and internally structured fibers and their applications—will be discussed. These new materials with helical/buckled morphology are expected to possess unique optical and mechanical properties with possible applications for negative refractive index materials, highly stretchable/high-tensile-strength materials, and components in microelectromechanical devices. Core-shell type fibers enable a much wider variety of materials to be electrospun and are expected to be widely applied in the sensing, drug delivery/controlled release fields, and in the encapsulation of live cells for biological applications. Materials with a hierarchical secondary structure are expected to provide new superhydrophobic and self-cleaning materials.

  1. Anti sense and sensibility : renal and skin effects of (antisense) oligonucleotides

    NARCIS (Netherlands)

    Meer, van L.

    2017-01-01

    This thesis describes the clinical investigation of a novel treatment strategy for type 2 diabetes mellitus (t2dm) using an antisense oligonucleotide(aon)to inhibit the sglt2 receptor. Furthermore it describes skin effects of oligonucleotides

  2. A flexible and fully integrated system for amplification, detection and genotyping of genomic DNA targets based on microfluidic oligonucleotide arrays.

    Science.gov (United States)

    Summerer, Daniel; Hevroni, Dona; Jain, Amit; Oldenburger, Olga; Parker, Jefferson; Caruso, Anthony; Stähler, Cord F; Stähler, Peer F; Beier, Markus

    2010-05-31

    A strategy allowing for amplification, detection and genotyping of different genomic DNA targets in a single reaction container is described. The method makes use of primer-directed solution-phase amplification with integrated labeling in a closed, microfluidic oligonucleotide array. Selective array probes allow for subsequent detection and genotyping of generated amplicons by hybridization. The array contains up to 15,624 programmable features that can be designed, de novo synthesized and tested within 24 hours using an automated benchtop microarray synthesizer. This enables rapid prototyping and adaptation of the system to newly emerging targets such as pathogenic bacterial or viral subtypes. The system was evaluated by amplifying and detecting different loci of viral (HPV), bacterial (Bacillus sp.) and eukaryotic (human) genomes. Multiplex PCR and semi-quantitative detection with excellent detection limits of automation grade of the system reduces contamination risk and workload and should enhance safety and reproducibility. 2010 Elsevier B.V. All rights reserved.

  3. methBLAST and methPrimerDB: web-tools for PCR based methylation analysis

    Directory of Open Access Journals (Sweden)

    Herzog Robert

    2006-11-01

    Full Text Available Abstract Background DNA methylation plays an important role in development and tumorigenesis by epigenetic modification and silencing of critical genes. The development of PCR-based methylation assays on bisulphite modified DNA heralded a breakthrough in speed and sensitivity for gene methylation analysis. Despite this technological advancement, these approaches require a cumbersome gene by gene primer design and experimental validation. Bisulphite DNA modification results in sequence alterations (all unmethylated cytosines are converted into uracils and a general sequence complexity reduction as cytosines become underrepresented. Consequently, standard BLAST sequence homology searches cannot be applied to search for specific methylation primers. Results To address this problem we developed methBLAST, a sequence similarity search program, based on the original BLAST algorithm but querying in silico bisulphite modified genome sequences to evaluate oligonucleotide sequence similarities. Apart from the primer specificity analysis tool, we have also developed a public database termed methPrimerDB for the storage and retrieval of validated PCR based methylation assays. The web interface allows free public access to perform methBLAST searches or database queries and to submit user based information. Database records can be searched by gene symbol, nucleotide sequence, analytical method used, Entrez Gene or methPrimerDB identifier, and submitter's name. Each record contains a link to Entrez Gene and PubMed to retrieve additional information on the gene, its genomic context and the article in which the methylation assay was described. To assure and maintain data integrity and accuracy, the database is linked to other reference databases. Currently, the database contains primer records for the most popular PCR-based methylation analysis methods to study human, mouse and rat epigenetic modifications. methPrimerDB and methBLAST are available at http

  4. Montmorillonite, oligonucleotides, RNA and origin of life

    Science.gov (United States)

    Ertem, Gozen

    2004-01-01

    Na-montmorillonite prepared from Volclay by the titration method facilitates the self-condensation of ImpA, the 5'-phosphorimidazolide derivative of adenosine. As was shown by AE-HPLC analysis and selective enzymatic hydrolysis of products, oligo(A)s formed in this reaction are 10 monomer units long and contain 67% 3',5'-phosphodiester bonds (Ferris and Ertem, 1992a). Under the same reaction conditions, 5'-phosphorimidazolide derivatives of cytidine, uridine and guanosine also undergo self-condensation producing oligomers containing up to 12-14 monomer units for oligo(C)s to 6 monomer units for oligo(G)s. In oligo(C)s and oligo(U)s, 75-80% of the monomers are linked by 2',5'-phosphodiester bonds. Hexamer and higher oligomers isolated from synthetic oligo(C)s formed by montmorillonite catalysis, which contain both 3',5'- and 2',5'-linkages, serve as catalysts for the non-enzymatic template directed synthesis of oligo(G)s from activated monomer 2-MeImpG, guanosine 5'-phospho-2-methylimidazolide (Ertem and Ferris, 1996). Pentamer and higher oligomers containing exclusively 2',5'-linkages, which were isolated from the synthetic oligo(C)s, also serve as templates and produce oligo(G)s with both 2',5'- and 3',5'-phosphodiester bonds. Kinetic studies on montmorillonite catalyzed elongation rates of oligomers using the computer program SIMFIT demonstrated that the rate constants for the formation of oligo(A)s increased in the order of 2-mer < 3-mer < 4-mer ... < 7-mer (Kawamura and Ferris, 1994). A decameric primer, dA(pdA)8pA bound to montmorillonite was elongated to contain up to 50 monomer units by daily addition of activated monomer ImpA to the reaction mixture (Ferris, Hill and Orgel, 1996). Analysis of dimer fractions formed in the montmorillonite catalyzed reaction of binary and quaternary mixtures of ImpA, ImpC, 2-MeImpG and ImpU suggested that only a limited number of oligomers could have formed on the primitive Earth rather than equal amounts of all possible

  5. Lipid-modified G4-decoy oligonucleotide anchored to nanoparticles

    DEFF Research Database (Denmark)

    Cogoi, S; Jakobsen, U; Pedersen, E B

    2016-01-01

    factor essential for KRAS transcription. It is based on the use of palmitoyl-oleyl-phosphatidylcholine (POPC) liposomes functionalized with lipid-modified G4-decoy oligonucleotides and a lipid-modified cell penetrating TAT peptide. The potency of the strategy in pancreatic cancer cells is demonstrated...

  6. Linear model for fast background subtraction in oligonucleotide microarrays

    NARCIS (Netherlands)

    Kroll, K.M.; Barkema, G.T.; Carlon, E.

    2009-01-01

    Background One important preprocessing step in the analysis of microarray data is background subtraction. In high-density oligonucleotide arrays this is recognized as a crucial step for the global performance of the data analysis from raw intensities to expression values. Results We propose here an

  7. Solid-phase-supported synthesis of morpholinoglycine oligonucleotide mimics

    Directory of Open Access Journals (Sweden)

    Tatyana V. Abramova

    2014-05-01

    Full Text Available An efficient solid-phase-supported peptide synthesis (SPPS of morpholinoglycine oligonucleotide (MorGly mimics has been developed. The proposed strategy includes a novel specially designed labile linker group containing the oxalyl residue and the 2-aminomethylmorpholino nucleoside analogues as first subunits.

  8. Chromosome-specific painting in Cucumis species using bulked oligonucleotides

    Science.gov (United States)

    Chromosome-specific painting is a powerful technique in molecular cytogenetic and genome research. We developed an oligonucleotide (oligo)-based chromosome painting technique in cucumber (Cucumis sativus) that will be applicable in any plant species with a sequenced genome. Oligos specific to a sing...

  9. Tandem Oligonucleotide Probe Annealing and Elongation To Discriminate Viral Sequence

    DEFF Research Database (Denmark)

    Taskova, Maria; Uhd, Jesper; Miotke, Laura

    2017-01-01

    opportunities in transcriptome analysis, virology, and other fields. Herein, we report for the first time a "click" chemistry approach to oligonucleotide probe elongation as a novel approach to specifically detect a viral sequence. We hybridized a library of short, terminally labeled probes to Ebola virus RNA...

  10. Fragmentation of protonated oligonucleotides by energetic photons and Cq+ ions

    NARCIS (Netherlands)

    Gonzalez-Magana, O.; Tiemens, M.; Reitsma, G.; Boschman, L.; Door, M.; Bari, S.; Lahaie, P. O.; Wagner, J. R.; Huels, M. A.; Hoekstra, R.; Schlathölter, Thomas

    2013-01-01

    The ionization and fragmentation of trapped protonated dGCAT oligonucleotides upon interaction with energetic photons (h nu = 10-570 eV) and keV Cq+ ions was investigated by means of time-of-flight mass spectrometry. The observed fragmentation patterns are dominated by protonated and nonprotonated

  11. Chemically modified oligonucleotides with efficient RNase H response

    DEFF Research Database (Denmark)

    Vester, Birte; Boel, Anne Marie; Lobedanz, Sune

    2008-01-01

    Ten different chemically modified nucleosides were incorporated into short DNA strands (chimeric oligonucleotides ON3-ON12 and ON15-ON24) and then tested for their capacity to mediate RNAse H cleavage of the complementary RNA strand. The modifications were placed at two central positions directly...

  12. Oligonucleotide-directed mutagenesis for precision gene editing.

    Science.gov (United States)

    Sauer, Noel J; Mozoruk, Jerry; Miller, Ryan B; Warburg, Zachary J; Walker, Keith A; Beetham, Peter R; Schöpke, Christian R; Gocal, Greg F W

    2016-02-01

    Differences in gene sequences, many of which are single nucleotide polymorphisms, underlie some of the most important traits in plants. With humanity facing significant challenges to increase global agricultural productivity, there is an urgent need to accelerate the development of these traits in plants. oligonucleotide-directed mutagenesis (ODM), one of the many tools of Cibus' Rapid Trait Development System (RTDS(™) ) technology, offers a rapid, precise and non-transgenic breeding alternative for trait improvement in agriculture to address this urgent need. This review explores the application of ODM as a precision genome editing technology, with emphasis on using oligonucleotides to make targeted edits in plasmid, episomal and chromosomal DNA of bacterial, fungal, mammalian and plant systems. The process of employing ODM by way of RTDS technology has been improved in many ways by utilizing a fluorescence conversion system wherein a blue fluorescent protein (BFP) can be changed to a green fluorescent protein (GFP) by editing a single nucleotide of the BFP gene (CAC→TAC; H66 to Y66). For example, dependent on oligonucleotide length, applying oligonucleotide-mediated technology to target the BFP transgene in Arabidopsis thaliana protoplasts resulted in up to 0.05% precisely edited GFP loci. Here, the development of traits in commercially relevant plant varieties to improve crop performance by genome editing technologies such as ODM, and by extension RTDS, is reviewed. © 2015 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  13. A primer of special relativity

    CERN Document Server

    Sardesai, PL

    2004-01-01

    A Primer of Special Relativity1 is an unusually lucid introduction to the subject specifically written for Indian students. It is intended to give the beginner a firm grounding for a more advanced course in relativity. An entire chapter is devoted to applications of the theory to elucidate a large number of topics the students (B.Sc. Physics) come across in Modern Physics. Detailed and well-selected examples are used to illuminate aspects of the theory as well as to show techniques of application. A large number of Illustrative Examples enables the students to gain confidence to solve any problem in relativity normally expected of B.Sc. students.

  14. A primer of multivariate statistics

    CERN Document Server

    Harris, Richard J

    2014-01-01

    Drawing upon more than 30 years of experience in working with statistics, Dr. Richard J. Harris has updated A Primer of Multivariate Statistics to provide a model of balance between how-to and why. This classic text covers multivariate techniques with a taste of latent variable approaches. Throughout the book there is a focus on the importance of describing and testing one's interpretations of the emergent variables that are produced by multivariate analysis. This edition retains its conversational writing style while focusing on classical techniques. The book gives the reader a feel for why

  15. Sample Return Primer and Handbook

    Science.gov (United States)

    Barrow, Kirk; Cheuvront, Allan; Faris, Grant; Hirst, Edward; Mainland, Nora; McGee, Michael; Szalai, Christine; Vellinga, Joseph; Wahl, Thomas; Williams, Kenneth; hide

    2007-01-01

    This three-part Sample Return Primer and Handbook provides a road map for conducting the terminal phase of a sample return mission. The main chapters describe element-by-element analyses and trade studies, as well as required operations plans, procedures, contingencies, interfaces, and corresponding documentation. Based on the experiences of the lead Stardust engineers, the topics include systems engineering (in particular range safety compliance), mission design and navigation, spacecraft hardware and entry, descent, and landing certification, flight and recovery operations, mission assurance and system safety, test and training, and the very important interactions with external support organizations (non-NASA tracking assets, landing site support, and science curation).

  16. A primer of Lebesgue integration

    CERN Document Server

    Bear, H S

    2001-01-01

    The Lebesgue integral is now standard for both applications and advanced mathematics. This books starts with a review of the familiar calculus integral and then constructs the Lebesgue integral from the ground up using the same ideas. A Primer of Lebesgue Integration has been used successfully both in the classroom and for individual study.Bear presents a clear and simple introduction for those intent on further study in higher mathematics. Additionally, this book serves as a refresher providing new insight for those in the field. The author writes with an engaging, commonsense style that appeals to readers at all levels.

  17. UniPrimer: A Web-Based Primer Design Tool for Comparative Analyses of Primate Genomes

    Directory of Open Access Journals (Sweden)

    Nomin Batnyam

    2012-01-01

    Full Text Available Whole genome sequences of various primates have been released due to advanced DNA-sequencing technology. A combination of computational data mining and the polymerase chain reaction (PCR assay to validate the data is an excellent method for conducting comparative genomics. Thus, designing primers for PCR is an essential procedure for a comparative analysis of primate genomes. Here, we developed and introduced UniPrimer for use in those studies. UniPrimer is a web-based tool that designs PCR- and DNA-sequencing primers. It compares the sequences from six different primates (human, chimpanzee, gorilla, orangutan, gibbon, and rhesus macaque and designs primers on the conserved region across species. UniPrimer is linked to RepeatMasker, Primer3Plus, and OligoCalc softwares to produce primers with high accuracy and UCSC In-Silico PCR to confirm whether the designed primers work. To test the performance of UniPrimer, we designed primers on sample sequences using UniPrimer and manually designed primers for the same sequences. The comparison of the two processes showed that UniPrimer was more effective than manual work in terms of saving time and reducing errors.

  18. Context updates are hierarchical

    Directory of Open Access Journals (Sweden)

    Anton Karl Ingason

    2016-10-01

    Full Text Available This squib studies the order in which elements are added to the shared context of interlocutors in a conversation. It focuses on context updates within one hierarchical structure and argues that structurally higher elements are entered into the context before lower elements, even if the structurally higher elements are pronounced after the lower elements. The crucial data are drawn from a comparison of relative clauses in two head-initial languages, English and Icelandic, and two head-final languages, Korean and Japanese. The findings have consequences for any theory of a dynamic semantics.

  19. Detecting Hierarchical Structure in Networks

    DEFF Research Database (Denmark)

    Herlau, Tue; Mørup, Morten; Schmidt, Mikkel Nørgaard

    2012-01-01

    a generative Bayesian model that is able to infer whether hierarchies are present or not from a hypothesis space encompassing all types of hierarchical tree structures. For efficient inference we propose a collapsed Gibbs sampling procedure that jointly infers a partition and its hierarchical structure......Many real-world networks exhibit hierarchical organization. Previous models of hierarchies within relational data has focused on binary trees; however, for many networks it is unknown whether there is hierarchical structure, and if there is, a binary tree might not account well for it. We propose....... On synthetic and real data we demonstrate that our model can detect hierarchical structure leading to better link-prediction than competing models. Our model can be used to detect if a network exhibits hierarchical structure, thereby leading to a better comprehension and statistical account the network....

  20. Nested and Hierarchical Archimax copulas

    KAUST Repository

    Hofert, Marius

    2017-07-03

    The class of Archimax copulas is generalized to nested and hierarchical Archimax copulas in several ways. First, nested extreme-value copulas or nested stable tail dependence functions are introduced to construct nested Archimax copulas based on a single frailty variable. Second, a hierarchical construction of d-norm generators is presented to construct hierarchical stable tail dependence functions and thus hierarchical extreme-value copulas. Moreover, one can, by itself or additionally, introduce nested frailties to extend Archimax copulas to nested Archimax copulas in a similar way as nested Archimedean copulas extend Archimedean copulas. Further results include a general formula for the density of Archimax copulas.

  1. Design and evaluation of 16S rRNA sequence based oligonucleotide probes for the detection and quantification of Comamonas testosteroni in mixed microbial communities

    Directory of Open Access Journals (Sweden)

    Bathe Stephan

    2006-06-01

    Full Text Available Abstract Background The β-proteobacterial species Comamonas testosteroni is capable of biotransformation and also biodegradation of a range of chemical compounds and thus potentially useful in chemical manufacturing and bioremediation. The ability to detect and quantify members of this species in mixed microbial communities thus may be desirable. Results We have designed an oligonucleotide probe for use in fluorescent in situ hybridization (FISH and two pairs of PCR primers targeting a C. testosteroni subgroup. The FISH probe and one of the PCR primer pairs are suitable for quantification of C. testosteroni in mixed microbial communities using FISH followed by quantitative image analysis or real-time quantitative PCR, respectively. This has been shown by analysis of samples from an enrichment of activated sludge on testosterone resulting in an increase in abundance and finally isolation of C. testosteroni. Additionally, we have successfully used quantitative PCR to follow the C. testosteroni abundance during a laboratory scale wastewater bioaugmentation experiment. Conclusion The oligonucleotides presented here provide a useful tool to study C. testosteroni population dynamics in mixed microbial communities.

  2. Synthesis of Oligonucleotides Carrying Thiol Groups Using a Simple Reagent Derived from Threoninol

    Directory of Open Access Journals (Sweden)

    Ramon Eritja

    2012-08-01

    Full Text Available Oligonucleotides carrying thiol groups are useful intermediates for a remarkable number of applications involving nucleic acids. In this study, DNA oligonucleotides carrying tert-butylsulfanyl (t-BuS protected thiol groups have been prepared. A building block derived from threoninol has been developed to introduce a thiol group at any predetemined position of an oligonucleotide. The resulting thiolated oligonucleotides have been used for the preparation of oligonucleotide conjugates and for the functionalization of gold nanoparticles using the reactivity of the thiol groups.

  3. A degenerate pair of primers for simultaneous detection of four alpha- and betanecroviruses.

    Science.gov (United States)

    Varanda, C M R; Cardoso, J M S; Oliveira, M D M; Oliveira, S; Clara, M I E; Félix, M R F

    2014-11-01

    The high infection levels due to Olive latent virus 1 (OLV-1), Olive mild mosaic virus (OMMV) (alphanecrovirus) and Tobacco necrosis virus D (TNV-D) (betanecrovirus) in Portuguese olive orchards prompted us to develop a rapid PCR-based assay for the simultaneous detection of these viruses aimed at the sanitary selection and marketing of plant material in compliance with European Union regulations. A pair of degenerate oligonucleotide primers, parRdRp5' and parCoat3' was designed based on conserved regions located in the RNA-dependent RNA polymerase (RdRp) and coat protein (CP) genes of these viruses and one other alphanecrovirus, Tobacco necrosis virus A. Its use in RT-PCR assays generated a product of ca. 2000 bp for the 4 viral species tested. These primers were compared with virus specific primers in multiplex RT-PCR, and identical results were obtained. Its application to dsRNA extracted from 54 olive field growing trees originated the expected ca. 2000 bp amplicon in 17 trees. The virus identity was determined by sequencing the cloned RT-PCR products. No TNV-A was found. The RT-PCR assay using the degenerate primers described in this study were shown to be reliable in detecting any of the above-mentioned alpha- and betanecroviruses, and it is as sensitive as that which uses virus specific primers in multiplex assays. Therefore, this assay is well suited for the rapid screen of virus-free plant material in selection and improvement crop programmes. Additionally, it has the potential to reveal virus diversity and the presence of new viruses, provided the RT-PCR generated amplicon is further sequenced. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Genome-wide identification of specific oligonucleotides using artificial neural network and computational genomic analysis

    Directory of Open Access Journals (Sweden)

    Chen Jiun-Ching

    2007-05-01

    Full Text Available Abstract Background Genome-wide identification of specific oligonucleotides (oligos is a computationally-intensive task and is a requirement for designing microarray probes, primers, and siRNAs. An artificial neural network (ANN is a machine learning technique that can effectively process complex and high noise data. Here, ANNs are applied to process the unique subsequence distribution for prediction of specific oligos. Results We present a novel and efficient algorithm, named the integration of ANN and BLAST (IAB algorithm, to identify specific oligos. We establish the unique marker database for human and rat gene index databases using the hash table algorithm. We then create the input vectors, via the unique marker database, to train and test the ANN. The trained ANN predicted the specific oligos with high efficiency, and these oligos were subsequently verified by BLAST. To improve the prediction performance, the ANN over-fitting issue was avoided by early stopping with the best observed error and a k-fold validation was also applied. The performance of the IAB algorithm was about 5.2, 7.1, and 6.7 times faster than the BLAST search without ANN for experimental results of 70-mer, 50-mer, and 25-mer specific oligos, respectively. In addition, the results of polymerase chain reactions showed that the primers predicted by the IAB algorithm could specifically amplify the corresponding genes. The IAB algorithm has been integrated into a previously published comprehensive web server to support microarray analysis and genome-wide iterative enrichment analysis, through which users can identify a group of desired genes and then discover the specific oligos of these genes. Conclusion The IAB algorithm has been developed to construct SpecificDB, a web server that provides a specific and valid oligo database of the probe, siRNA, and primer design for the human genome. We also demonstrate the ability of the IAB algorithm to predict specific oligos through

  5. Genome-wide identification of specific oligonucleotides using artificial neural network and computational genomic analysis.

    Science.gov (United States)

    Liu, Chun-Chi; Lin, Chin-Chung; Li, Ker-Chau; Chen, Wen-Shyen E; Chen, Jiun-Ching; Yang, Ming-Te; Yang, Pan-Chyr; Chang, Pei-Chun; Chen, Jeremy J W

    2007-05-22

    Genome-wide identification of specific oligonucleotides (oligos) is a computationally-intensive task and is a requirement for designing microarray probes, primers, and siRNAs. An artificial neural network (ANN) is a machine learning technique that can effectively process complex and high noise data. Here, ANNs are applied to process the unique subsequence distribution for prediction of specific oligos. We present a novel and efficient algorithm, named the integration of ANN and BLAST (IAB) algorithm, to identify specific oligos. We establish the unique marker database for human and rat gene index databases using the hash table algorithm. We then create the input vectors, via the unique marker database, to train and test the ANN. The trained ANN predicted the specific oligos with high efficiency, and these oligos were subsequently verified by BLAST. To improve the prediction performance, the ANN over-fitting issue was avoided by early stopping with the best observed error and a k-fold validation was also applied. The performance of the IAB algorithm was about 5.2, 7.1, and 6.7 times faster than the BLAST search without ANN for experimental results of 70-mer, 50-mer, and 25-mer specific oligos, respectively. In addition, the results of polymerase chain reactions showed that the primers predicted by the IAB algorithm could specifically amplify the corresponding genes. The IAB algorithm has been integrated into a previously published comprehensive web server to support microarray analysis and genome-wide iterative enrichment analysis, through which users can identify a group of desired genes and then discover the specific oligos of these genes. The IAB algorithm has been developed to construct SpecificDB, a web server that provides a specific and valid oligo database of the probe, siRNA, and primer design for the human genome. We also demonstrate the ability of the IAB algorithm to predict specific oligos through polymerase chain reaction experiments. Specific

  6. A primer on quantum fluids

    CERN Document Server

    Barenghi, Carlo

    2016-01-01

    The aim of this primer is to cover the essential theoretical information, quickly and concisely, in order to enable senior undergraduate and beginning graduate students to tackle projects in topical research areas of quantum fluids, for example, solitons, vortices and collective modes. The selection of the material, both regarding the content and level of presentation, draws on the authors analysis of the success of relevant research projects with newcomers to the field, as well as of the students feedback from many taught and self-study courses on the subject matter. Starting with a brief historical overview, this text covers particle statistics, weakly interacting condensates and their dynamics and finally superfluid helium and quantum turbulence. At the end of each chapter (apart from the first) there will be some exercises. Detailed solutions can be made available to instructors upon request to the authors. .

  7. Trees and Hierarchical Structures

    CERN Document Server

    Haeseler, Arndt

    1990-01-01

    The "raison d'etre" of hierarchical dustering theory stems from one basic phe­ nomenon: This is the notorious non-transitivity of similarity relations. In spite of the fact that very often two objects may be quite similar to a third without being that similar to each other, one still wants to dassify objects according to their similarity. This should be achieved by grouping them into a hierarchy of non-overlapping dusters such that any two objects in ~ne duster appear to be more related to each other than they are to objects outside this duster. In everyday life, as well as in essentially every field of scientific investigation, there is an urge to reduce complexity by recognizing and establishing reasonable das­ sification schemes. Unfortunately, this is counterbalanced by the experience of seemingly unavoidable deadlocks caused by the existence of sequences of objects, each comparatively similar to the next, but the last rather different from the first.

  8. URPD: a specific product primer design tool.

    Science.gov (United States)

    Chuang, Li-Yeh; Cheng, Yu-Huei; Yang, Cheng-Hong

    2012-06-19

    Polymerase chain reaction (PCR) plays an important role in molecular biology. Primer design fundamentally determines its results. Here, we present a currently available software that is not located in analyzing large sequence but used for a rather straight-forward way of visualizing the primer design process for infrequent users. URPD (yoUR Primer Design), a web-based specific product primer design tool, combines the NCBI Reference Sequences (RefSeq), UCSC In-Silico PCR, memetic algorithm (MA) and genetic algorithm (GA) primer design methods to obtain specific primer sets. A friendly user interface is accomplished by built-in parameter settings. The incorporated smooth pipeline operations effectively guide both occasional and advanced users. URPD contains an automated process, which produces feasible primer pairs that satisfy the specific needs of the experimental design with practical PCR amplifications. Visual virtual gel electrophoresis and in silico PCR provide a simulated PCR environment. The comparison of Practical gel electrophoresis comparison to virtual gel electrophoresis facilitates and verifies the PCR experiment. Wet-laboratory validation proved that the system provides feasible primers. URPD is a user-friendly tool that provides specific primer design results. The pipeline design path makes it easy to operate for beginners. URPD also provides a high throughput primer design function. Moreover, the advanced parameter settings assist sophisticated researchers in performing experiential PCR. Several novel functions, such as a nucleotide accession number template sequence input, local and global specificity estimation, primer pair redesign, user-interactive sequence scale selection, and virtual and practical PCR gel electrophoresis discrepancies have been developed and integrated into URPD. The URPD program is implemented in JAVA and freely available at http://bio.kuas.edu.tw/urpd/.

  9. URPD: a specific product primer design tool

    Directory of Open Access Journals (Sweden)

    Chuang Li-Yeh

    2012-06-01

    Full Text Available Abstract Background Polymerase chain reaction (PCR plays an important role in molecular biology. Primer design fundamentally determines its results. Here, we present a currently available software that is not located in analyzing large sequence but used for a rather straight-forward way of visualizing the primer design process for infrequent users. Findings URPD (yoUR Primer Design, a web-based specific product primer design tool, combines the NCBI Reference Sequences (RefSeq, UCSC In-Silico PCR, memetic algorithm (MA and genetic algorithm (GA primer design methods to obtain specific primer sets. A friendly user interface is accomplished by built-in parameter settings. The incorporated smooth pipeline operations effectively guide both occasional and advanced users. URPD contains an automated process, which produces feasible primer pairs that satisfy the specific needs of the experimental design with practical PCR amplifications. Visual virtual gel electrophoresis and in silico PCR provide a simulated PCR environment. The comparison of Practical gel electrophoresis comparison to virtual gel electrophoresis facilitates and verifies the PCR experiment. Wet-laboratory validation proved that the system provides feasible primers. Conclusions URPD is a user-friendly tool that provides specific primer design results. The pipeline design path makes it easy to operate for beginners. URPD also provides a high throughput primer design function. Moreover, the advanced parameter settings assist sophisticated researchers in performing experiential PCR. Several novel functions, such as a nucleotide accession number template sequence input, local and global specificity estimation, primer pair redesign, user-interactive sequence scale selection, and virtual and practical PCR gel electrophoresis discrepancies have been developed and integrated into URPD. The URPD program is implemented in JAVA and freely available at http://bio.kuas.edu.tw/urpd/.

  10. Nanomaterial building blocks based on spider silk-oligonucleotide conjugates.

    Science.gov (United States)

    Humenik, Martin; Scheibel, Thomas

    2014-02-25

    Self-assembling protein nanofibrils are promising structures for the "bottom-up" fabrication of bionanomaterials. Here, the recombinant protein eADF4(C16), a variant of Araneus diadematus dragline silk ADF4, which self-assembles into nanofibrils, and short oligonucleotides were modified for site-specific azide-alkyne coupling. Corresponding oligonuleotide-eADF4(C16) "click" conjugates were hybridized in linear or branched fashion according to the designed complementarities of the DNA moieties. Self-assembly properties of higher ordered structures of the spider silk-DNA conjugates were dominated by the silk component. Assembled β-sheet rich conjugate fibrils were similar in appearance to fibrils of unmodified eADF4(C16) but enabled the specific attachment of neutravidin-modified gold nanoparticles on their surface directed by complementary biotin-oligonucleotides, providing the basis for functionalization of such conjugates.

  11. Palladium-Catalyzed Modification of Unprotected Nucleosides, Nucleotides, and Oligonucleotides

    Directory of Open Access Journals (Sweden)

    Kevin H. Shaughnessy

    2015-05-01

    Full Text Available Synthetic modification of nucleoside structures provides access to molecules of interest as pharmaceuticals, biochemical probes, and models to study diseases. Covalent modification of the purine and pyrimidine bases is an important strategy for the synthesis of these adducts. Palladium-catalyzed cross-coupling is a powerful method to attach groups to the base heterocycles through the formation of new carbon-carbon and carbon-heteroatom bonds. In this review, approaches to palladium-catalyzed modification of unprotected nucleosides, nucleotides, and oligonucleotides are reviewed. Polar reaction media, such as water or polar aprotic solvents, allow reactions to be performed directly on the hydrophilic nucleosides and nucleotides without the need to use protecting groups. Homogeneous aqueous-phase coupling reactions catalyzed by palladium complexes of water-soluble ligands provide a general approach to the synthesis of modified nucleosides, nucleotides, and oligonucleotides.

  12. Typing of enteroviruses by use of microwell oligonucleotide arrays.

    Science.gov (United States)

    Susi, P; Hattara, L; Waris, M; Luoma-Aho, T; Siitari, H; Hyypiä, T; Saviranta, P

    2009-06-01

    We have developed a straightforward assay for the rapid typing of enteroviruses using oligonucleotide arrays in microtiter wells. The viral nucleic acids are concomitantly amplified and labeled during reverse transcription-PCR, and unpurified PCR products are used for hybridization. DNA strands are separated by alkaline denaturation, and hybridization is started by neutralization. The microarray hybridization reactions and the subsequent washes are performed in standard 96-well microtiter plates, which makes the method easily adaptable to high-throughput analysis. We describe here the assay principle and its potential in clinical laboratory use by correctly identifying 10 different enterovirus reference strains. Furthermore, we explore the detection of unknown sequence variants using serotype consensus oligonucleotide probes. With just two consensus probes for the coxsackievirus A9 (CVA9) serotype, we detected 23 out of 25 highly diverse CVA9 isolates. Overall, the assay involves several features aiming at ease of performance, robustness, and applicability to large-scale studies.

  13. Typing of Enteroviruses by Use of Microwell Oligonucleotide Arrays▿ †

    Science.gov (United States)

    Susi, P.; Hattara, L.; Waris, M.; Luoma-aho, T.; Siitari, H.; Hyypiä, T.; Saviranta, P.

    2009-01-01

    We have developed a straightforward assay for the rapid typing of enteroviruses using oligonucleotide arrays in microtiter wells. The viral nucleic acids are concomitantly amplified and labeled during reverse transcription-PCR, and unpurified PCR products are used for hybridization. DNA strands are separated by alkaline denaturation, and hybridization is started by neutralization. The microarray hybridization reactions and the subsequent washes are performed in standard 96-well microtiter plates, which makes the method easily adaptable to high-throughput analysis. We describe here the assay principle and its potential in clinical laboratory use by correctly identifying 10 different enterovirus reference strains. Furthermore, we explore the detection of unknown sequence variants using serotype consensus oligonucleotide probes. With just two consensus probes for the coxsackievirus A9 (CVA9) serotype, we detected 23 out of 25 highly diverse CVA9 isolates. Overall, the assay involves several features aiming at ease of performance, robustness, and applicability to large-scale studies. PMID:19357207

  14. Oligonucleotide?directed mutagenesis for precision gene editing

    OpenAIRE

    Sauer, Noel J.; Mozoruk, Jerry; Miller, Ryan B.; Warburg, Zachary J.; Walker, Keith A.; Beetham, Peter R.; Sch?pke, Christian R.; Gocal, Greg F. W.

    2015-01-01

    Summary Differences in gene sequences, many of which are single nucleotide polymorphisms, underlie some of the most important traits in plants. With humanity facing significant challenges to increase global agricultural productivity, there is an urgent need to accelerate the development of these traits in plants. oligonucleotide?directed mutagenesis (ODM), one of the many tools of Cibus? Rapid Trait Development System ( RTDS ?) technology, offers a rapid, precise and non?transgenic breeding a...

  15. Thermoplastic polymers surfaces for Dip-Pen Nanolithography of oligonucleotides

    Energy Technology Data Exchange (ETDEWEB)

    Suriano, Raffaella [Dipartimento di Chimica, Materiali e Ingegneria Chimica “Giulio Natta”, Politecnico di Milano, Piazza Leonardo da Vinci 32, 20133 Milano (Italy); Biella, Serena, E-mail: serena.biella@polimi.it [Dipartimento di Chimica, Materiali e Ingegneria Chimica “Giulio Natta”, Politecnico di Milano, Piazza Leonardo da Vinci 32, 20133 Milano (Italy); Cesura, Federico; Levi, Marinella; Turri, Stefano [Dipartimento di Chimica, Materiali e Ingegneria Chimica “Giulio Natta”, Politecnico di Milano, Piazza Leonardo da Vinci 32, 20133 Milano (Italy)

    2013-05-15

    Different thermoplastic polymers were spin-coated to prepare smooth surfaces for the direct deposition of end-group modified oligonucleotides by Dip-Pen Nanolithography. A study of the diffusion process was done in order to investigate the dependence of calibration coefficient and quality of deposited features on environmental parameters (temperature, relative humidity) and ink's molecular weight and functionality. The optimization of the process parameters led to the realization of high quality and density nanoarrays on plastics.

  16. Differentiation of regions with atypical oligonucleotide composition in bacterial genomes

    Directory of Open Access Journals (Sweden)

    Reva Oleg N

    2005-10-01

    Full Text Available Abstract Background Complete sequencing of bacterial genomes has become a common technique of present day microbiology. Thereafter, data mining in the complete sequence is an essential step. New in silico methods are needed that rapidly identify the major features of genome organization and facilitate the prediction of the functional class of ORFs. We tested the usefulness of local oligonucleotide usage (OU patterns to recognize and differentiate types of atypical oligonucleotide composition in DNA sequences of bacterial genomes. Results A total of 163 bacterial genomes of eubacteria and archaea published in the NCBI database were analyzed. Local OU patterns exhibit substantial intrachromosomal variation in bacteria. Loci with alternative OU patterns were parts of horizontally acquired gene islands or ancient regions such as genes for ribosomal proteins and RNAs. OU statistical parameters, such as local pattern deviation (D, pattern skew (PS and OU variance (OUV enabled the detection and visualization of gene islands of different functional classes. Conclusion A set of approaches has been designed for the statistical analysis of nucleotide sequences of bacterial genomes. These methods are useful for the visualization and differentiation of regions with atypical oligonucleotide composition prior to or accompanying gene annotation.

  17. Site-directed selection of oligonucleotide antagonists by competitive elution.

    Science.gov (United States)

    Bridonneau, P; Chang, Y F; Buvoli, A V; O'Connell, D; Parma, D

    1999-02-01

    Oligonucleotide ligands that bind a protein or a small molecule of interest are readily isolated by in vitro selection and amplification of rare sequences from combinatorial libraries of sequence-randomized oligonucleotides (Gold et al., 1995). Classic systematic evolution of ligands by exponential enrichment (SELEX) protocols are affinity based (Tuerk and Gold, 1990), but because many problems and applications require antagonists, protocols for selecting inhibitors are both desirable and valuable. A widely applicable approach for isolating inhibitors is competitive elution with a molecule that binds the targeted molecule's active or binding site. We have used this approach to isolate antagonists of wheat germ agglutinin (WGA) from a library of 2'NH2-pyrimidine, 2'OH-purine oligonucleotides by elution with N N' N"-triacetylchitotriose, (GlcNAc)3. The highest affinity aptamers have equilibrium dissociation constants of 1 nM-20 nM for WGA, a 10(3)-10(4)-fold improvement relative to (GlcNAc)3, and unlike the carbohydrate, are highly specific. In addition to competing for binding with (GlcNAc)3, aptamers inhibit WGA-mediated agglutination of sheep erythrocytes, demonstrating that they are able to compete with natural ligands presented on the surfaces of cells. These results illustrate the feasibility of isolating high-affinity, high-specificity antagonists by competitive elution with low molecular weight, relatively low-affinity, and low-specificity small molecules.

  18. Recent developments in reversible photoregulation of oligonucleotide structure and function.

    Science.gov (United States)

    Lubbe, Anouk S; Szymanski, Wiktor; Feringa, Ben L

    2017-02-20

    There is a growing interest in the photoregulation of biological functions, due to the high level of spatiotemporal precision achievable with light. Additionally, light is non-invasive and waste-free. In particular, the photoregulation of oligonucleotide structure and function is a rapidly developing study field with relevance to biological, physical and material sciences. Molecular photoswitches have been incorporated in oligonucleotides for 20 years, and the field has currently grown beyond fundamental studies on photochemistry of the switches and DNA duplex stability, and is moving towards applications in chemical biology, nanotechnology and material science. Moreover, the currently emerging field of photopharmacology indicates the relevance of photocontrol in future medicine. In recent years, a large number of publications has appeared on photoregulation of DNA and RNA structure and function. New strategies are evaluated and novel, exciting applications are shown. In this comprehensive review, the key strategies for photoswitch inclusion in oligonucleotides are presented and illustrated with recent examples. Additionally the applications that have emerged in recent years are discussed, including gene regulation, drug delivery and materials design. Finally, we identify the challenges that the field currently faces and look forward to future applications.

  19. Are "universal" DNA primers really universal?

    Science.gov (United States)

    Sharma, Pranay; Kobayashi, Tsuyoshi

    2014-11-01

    "Universal" DNA primers LCO 1490 and HCO 2198 were originally designed from three coding and six anticoding strands by comparing highly conserved regions of mitochondrial cytochrome c oxidase subunit I (COI) genes across 15 taxa. These primers have been successful in amplifying a 710-bp fragment of highly conserved regions of the COI gene for more than 80 invertebrate species from 11 phyla. In the present study, 130,843 variations were reviewed in the primer region of mitochondrial molecular markers by comparing 725 COI sequences from the kingdom Animalia. It was found that, for 177 invertebrate species, the forward primer (LCO 1490) showed only four conserved regions, compared to 12 in the original study. For ascidians, fungi and vertebrates, it showed approximately 50 % conserved regions, dropping to one conserved region for echinoderms. However, the reverse primer (HCO 2198) was highly conserved across 725 COI primer sequences. A similar pattern was observed in amino acid distributions. There was a significant difference in the means of base pair differences from the level of family, genus and species for LCO 1490 [analysis of variance (ANOVA), F 6,188 = 8.193, P primers from reference sequences belonging to the level of order (maximum 5 bp differences), family (maximum 6 bp differences) or genus (maximum 1 bp difference). Reverse primers can be designed from the level of family (maximum 5 bp differences) or genus (maximum 2 bp differences).

  20. Electrostatic Discharge testing of propellants and primers

    Energy Technology Data Exchange (ETDEWEB)

    Berry, R.B.

    1994-02-01

    This report presents the results of testing of selected propellants and primers to Electrostatic Discharge (ESD) characteristic of the human body. It describes the tests and the fixturing built to accommodate loose material (propellants) and the packed energetic material of the primer. The results indicate that all powders passed and some primers, especially the electric primers, failed to pass established requirements which delineate insensitive energetic components. This report details the testing of components and materials to four ESD environments (Standard ESD, Severe ESD, Modified Standard ESD, and Modified Severe ESD). The purpose of this study was to collect data based on the customer requirements as defined in the Sandia Environmental Safety & Health (ES&H) Manual, Chapter 9, and to define static sensitive and insensitive propellants and primers.

  1. [DNA amplification using PCR with abutting primers].

    Science.gov (United States)

    Garafutdinov, R R; Galimova, A A; Sakhabutdinova, A R; Vakhitov, V A; Chemeris, A V

    2015-01-01

    DNA analysis of ñîmplex biological objects (wastewater, soil, archaeological and forensic samples, etc.) is currently of great interest. DNA of these objects is characterized by low suitability for research due to the violation of its integrity and chemical structure; thus, the detection of specific nucleic acid fragments can be achieved by PCR with contiguous primers. In this paper, we present the results that clarify the specific characteristics of PCR with abutting primers. The 3'-ends of these primers are annealed at adjacent nucleotides of complementary chains of DNA target. It has been shown that the proximity of primers enables the formation of specific reaction products with a higher sensitivity and less reaction time. Using artificially damaged DNA and DNA from the soil we demonstrated that the abutting primers provide assured detection of specific DNA fragments. The results of this work may be taken into account in PCR with degraded (fragmented) DNA.

  2. Static magnetic field reduced exogenous oligonucleotide uptake by spermatozoa using magnetic nanoparticle gene delivery system

    Energy Technology Data Exchange (ETDEWEB)

    Katebi, Samira; Esmaeili, Abolghasem, E-mail: aesmaeili@sci.ui.ac.ir; Ghaedi, Kamran

    2016-03-15

    Spermatozoa could introduce exogenous oligonucleotides of interest to the oocyte. The most important reason of low efficiency of sperm mediated gene transfer (SMGT) is low uptake of exogenous DNA by spermatozoa. The aim of this study was to evaluate the effects of static magnetic field on exogenous oligonucleotide uptake of spermatozoa using magnetofection method. Magnetic nanoparticles (MNPs) associated with the labeled oligonucleotides were used to increase the efficiency of exogenous oligonucleotide uptake by rooster spermatozoa. We used high-field/high-gradient magnet (NdFeB) to enhance and accelerate exogenous DNA sedimentation at the spermatozoa surface. Flow cytometry analysis was performed to measure viability and percentage of exogenous oligonucleotide uptake by sperm. Flow cytometry analysis showed a significant increase in exogenous oligonucleotide uptake by rooster spermatozoa (P<0.001) when spermatozoa were incubated in exogenous oligonucleotide solution and MNPs. However, by applying static magnetic field during magnetofection method, a significant decrease in exogenous oligonucleotide uptake was observed (P<0.05). Findings of this study showed that MNPs were effective to increase exogenous oligonucleotide uptake by rooster spermatozoa; however unlike others studies, static magnetic field, was not only ineffective to enhance exogenous oligonucleotide uptake by rooster spermatozoa but also led to reduction in efficiency of magnetic nanoparticles in gene transfer. - Highlights: • Core/shell type Iron oxide nanoparticles were used as a novel and efficient method. • This method increases exogenous DNA uptake by rooster spermatozoa. • Static magnetic field decreased DNA uptake by rooster spermatozoa.

  3. Primer3_masker: integrating masking of template sequence with primer design software.

    Science.gov (United States)

    Kõressaar, Triinu; Lepamets, Maarja; Kaplinski, Lauris; Raime, Kairi; Andreson, Reidar; Remm, Maido

    2018-01-19

    Designing PCR primers for amplifying regions of eukaryotic genomes is a complicated task because the genomes contain a large number of repeat sequences and other regions unsuitable for amplification by PCR. We have developed a novel k-mer based masking method that uses a statistical model to detect and mask failure-prone regions on the DNA template prior to primer design. We implemented the software as a standalone software primer3_masker and integrated it into the primer design program Primer3. The standalone version of primer3_masker is implemented in C. The source code is freely available at https://github.com/bioinfo-ut/primer3_masker/ (standalone version for Linux and macOS) and at https://github.com/primer3-org/primer3/(integrated version). Primer3 web application that allows masking sequences of 196 animal and plant genomes is available at http://primer3.ut.ee/. maido.remm@ut.ee. Supplementary data are available at Bioinformatics online.

  4. Hierarchical multifunctional nanocomposites

    Science.gov (United States)

    Ghasemi-Nejhad, Mehrdad N.

    2014-03-01

    properties of the fibers can also be improved by the growth of nanotubes on the fibers. The combination of the two will produce super-performing materials, not currently available. Since the improvement of fiber starts with carbon nanotube grown on micron-size fibers (and matrix with a nanomaterial) to give the macro-composite, this process is a bottom-up "hierarchical" advanced manufacturing process, and since the resulting nanocomposites will have "multifunctionality" with improve properties in various functional areas such as chemical and fire resistance, damping, stiffness, strength, fracture toughness, EMI shielding, and electrical and thermal conductivity, the resulting nanocomposites are in fact "multifunctional hierarchical nanocomposites." In this paper, the current state of knowledge in processing, performance, and characterization of these materials are addressed.

  5. PHUSER (Primer Help for USER): a novel tool for USER fusion primer design

    DEFF Research Database (Denmark)

    Olsen, Lars Rønn; Hansen, Niels Bjørn; Bonde, Mads

    2011-01-01

    Uracil-Specific Exision Reagent (USER) fusion is a recently developed technique that allows for assembly of multiple DNA fragments in a few simple steps. However, designing primers for USER fusion is both tedious and time consuming. Here, we present the Primer Help for USER (PHUSER) software......, a novel tool for designing primers specifically for USER fusion and USER cloning applications. We also present proof-of-concept experimental validation of its functionality. PHUSER offers quick and easy design of PCR optimized primers ensuring directionally correct fusion of fragments into a plasmid...... containing a customizable USER cassette. Designing primers using PHUSER ensures that the primers have similar annealing temperature (Tm), which is essential for efficient PCR. PHUSER also avoids identical overhangs, thereby ensuring correct order of assembly of DNA fragments. All possible primers...

  6. Hierarchical Discriminant Analysis

    Directory of Open Access Journals (Sweden)

    Di Lu

    2018-01-01

    Full Text Available The Internet of Things (IoT generates lots of high-dimensional sensor intelligent data. The processing of high-dimensional data (e.g., data visualization and data classification is very difficult, so it requires excellent subspace learning algorithms to learn a latent subspace to preserve the intrinsic structure of the high-dimensional data, and abandon the least useful information in the subsequent processing. In this context, many subspace learning algorithms have been presented. However, in the process of transforming the high-dimensional data into the low-dimensional space, the huge difference between the sum of inter-class distance and the sum of intra-class distance for distinct data may cause a bias problem. That means that the impact of intra-class distance is overwhelmed. To address this problem, we propose a novel algorithm called Hierarchical Discriminant Analysis (HDA. It minimizes the sum of intra-class distance first, and then maximizes the sum of inter-class distance. This proposed method balances the bias from the inter-class and that from the intra-class to achieve better performance. Extensive experiments are conducted on several benchmark face datasets. The results reveal that HDA obtains better performance than other dimensionality reduction algorithms.

  7. A novel photoinduced electron transfer (PET) primer technique for rapid real-time PCR detection of Cryptosporidium spp

    Energy Technology Data Exchange (ETDEWEB)

    Jothikumar, N., E-mail: jin2@cdc.gov; Hill, Vincent R.

    2013-06-28

    Highlights: •Uses a single-labeled fluorescent primer for real-time PCR. •The detection sensitivity of PET PCR was comparable to TaqMan PCR. •Melt curve analysis can be performed to confirm target amplicon production. •Conventional PCR primers can be converted to PET PCR primers. -- Abstract: We report the development of a fluorescently labeled oligonucleotide primer that can be used to monitor real-time PCR. The primer has two parts, the 3′-end of the primer is complimentary to the target and a universal 17-mer stem loop at the 5′-end forms a hairpin structure. A fluorescent dye is attached to 5′-end of either the forward or reverse primer. The presence of guanosine residues at the first and second position of the 3′ dangling end effectively quenches the fluorescence due to the photo electron transfer (PET) mechanism. During the synthesis of nucleic acid, the hairpin structure is linearized and the fluorescence of the incorporated primer increases several-fold due to release of the fluorescently labeled tail and the absence of guanosine quenching. As amplicons are synthesized during nucleic acid amplification, the fluorescence increase in the reaction mixture can be measured with commercially available real-time PCR instruments. In addition, a melting procedure can be performed to denature the double-stranded amplicons, thereby generating fluorescence peaks that can differentiate primer dimers and other non-specific amplicons if formed during the reaction. We demonstrated the application of PET-PCR for the rapid detection and quantification of Cryptosporidium parvum DNA. Comparison with a previously published TaqMan® assay demonstrated that the two real-time PCR assays exhibited similar sensitivity for a dynamic range of detection of 6000–0.6 oocysts per reaction. PET PCR primers are simple to design and less-expensive than dual-labeled probe PCR methods, and should be of interest for use by laboratories operating in resource

  8. Primer on spontaneous heating and pyrophoricity

    Energy Technology Data Exchange (ETDEWEB)

    1994-12-01

    This primer was prepared as an information resource for personnel responsible for operation of DOE nuclear facilities. It has sections on combustion principles, spontaneous heating/ignition of hydrocarbons and organics, pyrophoric gases and liquids, pyrophoric nonmetallic solids, pyrophoric metals (including Pu and U), and accident case studies. Although the information in this primer is not all-encompassing, it should provide the reader with a fundamental knowledge level sufficient to recognize most spontaneous combustion hazards and how to prevent ignition and widespread fires. This primer is provided as an information resource only, and is not intended to replace any fire protection or hazardous material training.

  9. Primers-4-Yeast: a comprehensive web tool for planning primers for Saccharomyces cerevisiae.

    Science.gov (United States)

    Yofe, Ido; Schuldiner, Maya

    2014-02-01

    The budding yeast Saccharomyces cerevisiae is a key model organism of functional genomics, due to its ease and speed of genetic manipulations. In fact, in this yeast, the requirement for homologous sequences for recombination purposes is so small that 40 base pairs (bp) are sufficient. Hence, an enormous variety of genetic manipulations can be performed by simply planning primers with the correct homology, using a defined set of transformation plasmids. Although designing primers for yeast transformations and for the verification of their correct insertion is a common task in all yeast laboratories, primer planning is usually done manually and a tool that would enable easy, automated primer planning for the yeast research community is still lacking. Here we introduce Primers-4-Yeast, a web tool that allows primers to be designed in batches for S. cerevisiae gene-targeting transformations, and for the validation of correct insertions. This novel tool enables fast, automated, accurate primer planning for large sets of genes, introduces consistency in primer planning and is therefore suggested to serve as a standard in yeast research. Primers-4-Yeast is available at: http://www.weizmann.ac.il/Primers-4-Yeast Copyright © 2013 John Wiley & Sons, Ltd.

  10. Hierarchical clustering for graph visualization

    CERN Document Server

    Clémençon, Stéphan; Rossi, Fabrice; Tran, Viet Chi

    2012-01-01

    This paper describes a graph visualization methodology based on hierarchical maximal modularity clustering, with interactive and significant coarsening and refining possibilities. An application of this method to HIV epidemic analysis in Cuba is outlined.

  11. Direct hierarchical assembly of nanoparticles

    Science.gov (United States)

    Xu, Ting; Zhao, Yue; Thorkelsson, Kari

    2014-07-22

    The present invention provides hierarchical assemblies of a block copolymer, a bifunctional linking compound and a nanoparticle. The block copolymers form one micro-domain and the nanoparticles another micro-domain.

  12. Hierarchical materials: Background and perspectives

    DEFF Research Database (Denmark)

    2016-01-01

    Hierarchical design draws inspiration from analysis of biological materials and has opened new possibilities for enhancing performance and enabling new functionalities and extraordinary properties. With the development of nanotechnology, the necessary technological requirements for the manufactur...

  13. Hierarchical classification as relational framing.

    Science.gov (United States)

    Slattery, Brian; Stewart, Ian

    2014-01-01

    The purpose of this study was to model hierarchical classification as contextually controlled, generalized relational responding or relational framing. In Experiment 1, a training procedure involving nonarbitrarily related multidimensional stimuli was used to establish two arbitrary shapes as contextual cues for 'member of' and 'includes' relational responding, respectively. Subsequently those cues were used to establish a network of arbitrary stimuli in particular hierarchical relations with each other, and then test for derivation of further untrained hierarchical relations as well as for transformation of functions. Resultant patterns of relational framing showed properties of transitive class containment, asymmetrical class containment, and unilateral property induction, consistent with conceptions of hierarchical classification as described within the cognitive developmental literature. Experiment 2 extended the basic model by using "fuzzy category" stimuli and providing a better controlled test of transformation of functions. Limitations and future research directions are discussed. © Society for the Experimental Analysis of Behavior.

  14. MFEprimer: multiple factor evaluation of the specificity of PCR primers.

    Science.gov (United States)

    Qu, Wubin; Shen, Zhiyong; Zhao, Dongsheng; Yang, Yi; Zhang, Chenggang

    2009-01-15

    We developed a program named MFEprimer for evaluating the specificity of PCR primers based on multiple factors, including sequence similarity, stability at the 3'-end of the primer, melting temperature, GC content and number of binding sites between the primer and DNA templates. MFEprimer can help the user to select more suitable primers before running either standard or multiplex PCR reactions. The cDNA and genomic DNA databases of 10 widely used species, as well as user custom databases, were used as DNA templates for analyzing primers specificity. Furthermore, we maintained a Primer3Plus server with a modified Primer3Manager for one-stop primer design and specificity checking.

  15. Production of cis-syn thymine-thymine cyclobutane dimer oligonucleotide in the presence of acetone photosensitizer.

    Science.gov (United States)

    Mu, Wanmeng; Han, Qingkai; Luo, Zhaofeng; Wang, Yuzhen

    2006-06-01

    cis-syn Cyclobutane pyrimidine dimer (CPD) oligonucleotide was produced by UV irradiation in the presence of acetone photosensitizer. Acetone could enhance the productivity but evidently induced the photocleavage of oligonucleotide under a long time irradiation. A statistical approach of orthogonal design was applied to optimize the preparation condition for the production of the modified oligonucleotide. Optimal conditions for maximal cis-syn CPD oligonucleotide productivity were determined based on three factors: acetone concentration, initial oligonucleotide concentration, and irradiation time at several different levels. The optimal modified oligonucleotide that this optimization could produce was 32.7%. Through analysis of 20% polyacrylamide gel electrophoresis, it was found that modified oligonucleotide migrated slightly more slowly than the parent oligonucleotide. The photoreactivation of cis-syn thymine-thymine dimer oligonucleotide displayed the selectivity of the substrate specificity of DNA photolyase with high-performance liquid chromatography (HPLC) analysis.

  16. Oligonucleotide Antiviral Therapeutics: Antisense and RNA Interference for Highly Pathogenic RNA Viruses

    National Research Council Canada - National Science Library

    Spurgers, Kevin B; Sharkey, C. M; Warfield, Kelly L; Bavari, Sina

    2008-01-01

    .... Important advances in the field include the characterization of RNA interference in mammalian cells and chemical modifications that can dramatically increase the in vivo stability of therapeutic oligonucleotides...

  17. Terahertz response of DNA oligonucleotides on the surface of silicon nanostructures

    Energy Technology Data Exchange (ETDEWEB)

    Bagraev, N. T., E-mail: bagraev@mail.ioffe.ru [Peter the Great Saint-Petersburg Polytechnic University (Russian Federation); Chernev, A. L. [Russian Academy of Sciences, Saint Petersburg Academic University—Nanotechnology Research and Education Center (Russian Federation); Klyachkin, L. E.; Malyarenko, A. M. [Russian Academy of Sciences, Ioffe Physical–Technical Institute (Russian Federation); Emel’yanov, A. K.; Dubina, M. V. [Russian Academy of Sciences, Saint Petersburg Academic University—Nanotechnology Research and Education Center (Russian Federation)

    2016-09-15

    The possibility of identifying DNA oligonucleotides deposited onto the region of the edge channels of silicon nanostructures is considered. The role of various THz (terahertz) radiation harmonics of silicon nanostructures in the resonance response of oligonucleotides is analyzed. In particular, this makes it possible to compare single-stranded 100- and 50-mer DNA oligonucleotides. A technique for the rapid identification of different oligonucleotides by measuring changes in the conductance and transverse potential difference of silicon nanostructures with microcavities, embedded in the edge channels for selecting THz radiation characteristics, is proposed.

  18. A practical primer on geostatistics

    Science.gov (United States)

    Olea, Ricardo A.

    2009-01-01

    has significant methodological implications.Historical Remarks—As a discipline, geostatistics was firmly established in the 1960s by the French engineer Georges Matheron, who was interested in the appraisal of ore reserves in mining. Geostatistics did not develop overnight. Like other disciplines, it has built on previous results, many of which were formulated with different objectives in various fields.Pioneers—Seminal ideas conceptually related to what today we call geostatistics or spatial statistics are found in the work of several pioneers, including: 1940s: A.N. Kolmogorov in turbulent flow and N. Wiener in stochastic processing; 1950s: D. Krige in mining; 1960s: B. Mathern in forestry and L.S. Gandin in meteorologyCalculations—Serious applications of geostatistics require the use of digital computers. Although for most geostatistical techniques rudimentary implementation from scratch is fairly straightforward, coding programs from scratch is recommended only as part of a practice that may help users to gain a better grasp of the formulations.Software—For professional work, the reader should employ software packages that have been thoroughly tested to handle any sampling scheme, that run as efficiently as possible, and that offer graphic capabilities for the analysis and display of results. This primer employs primarily the package Stanford Geomodeling Software (SGeMS) - recently developed at the Energy Resources Engineering Department at Stanford University - as a way to show how to obtain results practically. This applied side of the primer should not be interpreted as the notes being a manual for the use of SGeMS. The main objective of the primer is to help the reader gain an understanding of the fundamental concepts and tools in geostatistics.Organization of the Primer—The chapters of greatest importance are those covering kriging and simulation. All other materials are peripheral and are included for better comprehension of these main

  19. Multi-gene detection and identification of mosquito-borne RNA viruses using an oligonucleotide microarray.

    Directory of Open Access Journals (Sweden)

    Nathan D Grubaugh

    Full Text Available BACKGROUND: Arthropod-borne viruses are important emerging pathogens world-wide. Viruses transmitted by mosquitoes, such as dengue, yellow fever, and Japanese encephalitis viruses, infect hundreds of millions of people and animals each year. Global surveillance of these viruses in mosquito vectors using molecular based assays is critical for prevention and control of the associated diseases. Here, we report an oligonucleotide DNA microarray design, termed ArboChip5.1, for multi-gene detection and identification of mosquito-borne RNA viruses from the genera Flavivirus (family Flaviviridae, Alphavirus (Togaviridae, Orthobunyavirus (Bunyaviridae, and Phlebovirus (Bunyaviridae. METHODOLOGY/PRINCIPAL FINDINGS: The assay utilizes targeted PCR amplification of three genes from each virus genus for electrochemical detection on a portable, field-tested microarray platform. Fifty-two viruses propagated in cell-culture were used to evaluate the specificity of the PCR primer sets and the ArboChip5.1 microarray capture probes. The microarray detected all of the tested viruses and differentiated between many closely related viruses such as members of the dengue, Japanese encephalitis, and Semliki Forest virus clades. Laboratory infected mosquitoes were used to simulate field samples and to determine the limits of detection. Additionally, we identified dengue virus type 3, Japanese encephalitis virus, Tembusu virus, Culex flavivirus, and a Quang Binh-like virus from mosquitoes collected in Thailand in 2011 and 2012. CONCLUSIONS/SIGNIFICANCE: We demonstrated that the described assay can be utilized in a comprehensive field surveillance program by the broad-range amplification and specific identification of arboviruses from infected mosquitoes. Furthermore, the microarray platform can be deployed in the field and viral RNA extraction to data analysis can occur in as little as 12 h. The information derived from the ArboChip5.1 microarray can help to establish

  20. Fluorous methods for the synthesis of peptides and oligonucleotides.

    Science.gov (United States)

    Miriyala, Bruhaspathy

    2012-01-01

    The non-covalent affinity of a perfluoro chain towards similar has been exploited by many to separate fluorous tagged compounds from non-fluorous compounds by F-SPE or F-LLE. This purification strategy found its application across diverse fields including peptide and oligonucleotide synthesis where even slight inefficient couplings result in deletion sequences that are often difficult to remove from the target sequence. Two commonly employed strategies to address this problem involve end-tagging the target sequence or capping the deletion sequences with fluorous tags. Solution phase syntheses using soluble fluorous supports are easier and quicker. These approaches are reviewed here in detail.

  1. Universal detection of phytoplasmas and Xylella spp. by TaqMan singleplex and multiplex real-time PCR with dual priming oligonucleotides.

    Directory of Open Access Journals (Sweden)

    Takao Ito

    Full Text Available Phytoplasmas and Xylella spp. are bacteria that cause many economically important plant diseases worldwide. TaqMan probe-based quantitative real-time polymerase chain reaction (qPCR assays have been utilized to universally detect phytoplasmas or Xylella fastidiosa. To develop a superior universal qPCR method, we used a dual priming oligonucleotide (DPO with two annealing sites as a reverse primer to target the well-conserved bacterial 16S rDNA. The new qPCR assays universally detected various species of phytoplasmas and subspecies of X. fastidiosa as well as Xylella taiwanensis, and generally showed superior threshold cycle values when amplifying specific or non-specific products compared to current universal qPCR assays. The proposed qPCR assays were integrated to develop a multiplex qPCR assay that simultaneously detected phytoplasmas, Xylella spp., and an internal plant DNA positive control within 1 hour. This assay could detect a minimum of ten bacterial cells and was compatible with crude extractions used in the rapid screening of various plants. The amplicons were of sufficient lengths to be directly sequenced for preliminary identification, and the primers could be used in universal conventional PCR assays. Additionally, reverse DPO primers can be utilized to improve other probe-based qPCR assays.

  2. Use of oligonucleotide array for identification of six foodborne pathogens and Pseudomonas aeruginosa grown on selective media.

    Science.gov (United States)

    Lin, Miao Chu; Huang, Ay Huey; Tsen, Hau Yang; Wong, Hin-Chung; Chang, Tsung Chain

    2005-11-01

    Identification of presumptive foodborne pathogens grown on selective media may take one to several days and requires a different battery of biochemical tests for each microorganism. A molecular identification method was developed in which universal primers were used to amplify the 16S to 23S rDNA intergenic spacer of target microorganisms, and PCR products were hybridized to a panel of species-specific oligonucleotides that were immobilized on a nylon membrane. The seven target microorganisms were Bacillus cereus, Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella, Staphylococcus aureus, and Vibrio parahaemolyticus. After testing a large collection of target bacteria (29 to 51 strains) and nontarget bacteria (> 500 strains), the performances (sensitivity and specificity) of the oligonucleotide array were as follows: B. cereus (100 and 77%), E. coli (100 and 100%), L. monocytogenes (100 and 90%), P. aeruginosa (100 and 100%), Salmonella (100 and 100%), S. aureus (100 and 100%), and V. parahaemolyticus (100 and 94.2%). Other species in the B. cereus group cross-hybridized to the probes used for identification of B. cereus, and positive results should be confirmed by additional morphological observation of colonies. Listeria innocua cross-reacted with probes used to identify L. monocytogenes, but a simple hemolysis test was used to differentiate the two species. Some strains of Vibrio harveyi and Vibrio mimicus cross-hybridized with probes used for identification of V. parahaemolyticus and caused false-positive reactions. The advantage of the array is that a common protocol was used to identify the seven target microorganisms and multiple different microorganisms could be simultaneously identified on a single array.

  3. Multiplexing Short Primers for Viral Family PCR

    Energy Technology Data Exchange (ETDEWEB)

    Gardner, S N; Hiddessen, A L; Hara, C A; Williams, P L; Wagner, M; Colston, B W

    2008-06-26

    We describe a Multiplex Primer Prediction (MPP) algorithm to build multiplex compatible primer sets for large, diverse, and unalignable sets of target sequences. The MPP algorithm is scalable to larger target sets than other available software, and it does not require a multiple sequence alignment. We applied it to questions in viral detection, and demonstrated that there are no universally conserved priming sequences among viruses and that it could require an unfeasibly large number of primers ({approx}3700 18-mers or {approx}2000 10-mers) to generate amplicons from all sequenced viruses. We then designed primer sets separately for each viral family, and for several diverse species such as foot-and-mouth disease virus, hemagglutinin and neuraminidase segments of influenza A virus, Norwalk virus, and HIV-1.

  4. Hexavalent Chromium IV-Free Primer Development

    Science.gov (United States)

    Alldredge, Michael J.; Buck, Amy L.

    2015-01-01

    Primer materials provide corrosion protection for metal parts as well as an increased adhesion between metallic substrates and thermal protection systems (TPSs). Current primers for use in cryogenic applications contain hexavalent chromium. This hexavalent chromium provides excellent corrosion protection even in a cryogenic environment, but it is a carcinogen that requires special equipment and waste control procedures to use. The hazardous nature of hexavalent chromium makes it an obsolescence risk in the future. This study included two phases of evaluation. Thirteen primers were initially identified as candidates and twelve of those primers were tested in phase 1. Four of the best performing candidates from phase 1 continued into phase 2 testing. Phase 1 testing consisted mostly of liquid constituent and physical property testing. Cryoflex and salt fog testing were included in phase 1 because of their importance to the overall success of a candidate material. Phase 2 consisted of physical, thermal, and mechanical properties for nominally processed and fabricated specimens.

  5. Primer on transportation and climate change

    Science.gov (United States)

    2008-04-01

    This primer is an introduction to the issue of climate change and its implications for transportation policy in the United States. Its purpose is to outline the current thinking of governmental agencies, researchers, and advocacy groups on the issue ...

  6. ANL supplement to the UNICOS primer

    Energy Technology Data Exchange (ETDEWEB)

    Wiley, M.S. (ed.); Karlovsky, S.R.

    1991-06-01

    The ANL Supplement to the UNICOS Primer (ANL/TM 460) introduces the Cray X-MP interactive and batch services available at Argonne National Laboratory. It serves as a companion to the UNICOS Primer (Cray publication SG-2010 6.0). Whereas the UNICOS Primer discusses standard Unix issues of Cray computing, this manual discusses those issues specific to Cray computing at ANL. If this is your first experience on a Unix-based system, we assume that you have read at least Chapters 1 through 3 of the UNICOS Primer. The Glossary at the back of the UNICOS Primer will also be useful to you. If you are already familiar with a Unix system, it should suffice to keep the UNICOS Primer handy as you use this document. To learn about Unix programming in greater detail, we recommend A Practical Guide to the Unix System, by Mark G. Sobell. This manual and all other sources referred to in this document are available for purchase at the Document Distribution Counter in Building 221, Room A-134. We assume that you have already read the Guide to Computing at ANL (ANL/TM 336) to get an overview of all the computing facilities and services available at Argonne National Laboratory. You should also refer to Recommended Documentation for Computer Users at ANL (ANL/TM 379) for additional guidance in selecting available documentation that will best fill your particular computing needs.

  7. AmoA-Targeted Polymerase Chain Reaction Primers for the Specific Detection and Quantification of Comammox Nitrospira in the Environment

    Directory of Open Access Journals (Sweden)

    Petra Pjevac

    2017-08-01

    Full Text Available Nitrification, the oxidation of ammonia via nitrite to nitrate, has always been considered to be catalyzed by the concerted activity of ammonia- and nitrite-oxidizing microorganisms. Only recently, complete ammonia oxidizers (“comammox”, which oxidize ammonia to nitrate on their own, were identified in the bacterial genus Nitrospira, previously assumed to contain only canonical nitrite oxidizers. Nitrospira are widespread in nature, but for assessments of the distribution and functional importance of comammox Nitrospira in ecosystems, cultivation-independent tools to distinguish comammox from strictly nitrite-oxidizing Nitrospira are required. Here we developed new PCR primer sets that specifically target the amoA genes coding for subunit A of the distinct ammonia monooxygenase of comammox Nitrospira. While existing primers capture only a fraction of the known comammox amoA diversity, the new primer sets cover as much as 95% of the comammox amoA clade A and 92% of the clade B sequences in a reference database containing 326 comammox amoA genes with sequence information at the primer binding sites. Application of the primers to 13 samples from engineered systems (a groundwater well, drinking water treatment and wastewater treatment plants and other habitats (rice paddy and forest soils, rice rhizosphere, brackish lake sediment and freshwater biofilm detected comammox Nitrospira in all samples and revealed a considerable diversity of comammox in most habitats. Excellent primer specificity for comammox amoA was achieved by avoiding the use of highly degenerate primer preparations and by using equimolar mixtures of oligonucleotides that match existing comammox amoA genes. Quantitative PCR with these equimolar primer mixtures was highly sensitive and specific, and enabled the efficient quantification of clade A and clade B comammox amoA gene copy numbers in environmental samples. The measured relative abundances of comammox Nitrospira, compared

  8. AmoA-Targeted Polymerase Chain Reaction Primers for the Specific Detection and Quantification of Comammox Nitrospira in the Environment.

    Science.gov (United States)

    Pjevac, Petra; Schauberger, Clemens; Poghosyan, Lianna; Herbold, Craig W; van Kessel, Maartje A H J; Daebeler, Anne; Steinberger, Michaela; Jetten, Mike S M; Lücker, Sebastian; Wagner, Michael; Daims, Holger

    2017-01-01

    Nitrification, the oxidation of ammonia via nitrite to nitrate, has always been considered to be catalyzed by the concerted activity of ammonia- and nitrite-oxidizing microorganisms. Only recently, complete ammonia oxidizers ("comammox"), which oxidize ammonia to nitrate on their own, were identified in the bacterial genus Nitrospira, previously assumed to contain only canonical nitrite oxidizers. Nitrospira are widespread in nature, but for assessments of the distribution and functional importance of comammox Nitrospira in ecosystems, cultivation-independent tools to distinguish comammox from strictly nitrite-oxidizing Nitrospira are required. Here we developed new PCR primer sets that specifically target the amoA genes coding for subunit A of the distinct ammonia monooxygenase of comammox Nitrospira. While existing primers capture only a fraction of the known comammox amoA diversity, the new primer sets cover as much as 95% of the comammox amoA clade A and 92% of the clade B sequences in a reference database containing 326 comammox amoA genes with sequence information at the primer binding sites. Application of the primers to 13 samples from engineered systems (a groundwater well, drinking water treatment and wastewater treatment plants) and other habitats (rice paddy and forest soils, rice rhizosphere, brackish lake sediment and freshwater biofilm) detected comammox Nitrospira in all samples and revealed a considerable diversity of comammox in most habitats. Excellent primer specificity for comammox amoA was achieved by avoiding the use of highly degenerate primer preparations and by using equimolar mixtures of oligonucleotides that match existing comammox amoA genes. Quantitative PCR with these equimolar primer mixtures was highly sensitive and specific, and enabled the efficient quantification of clade A and clade B comammox amoA gene copy numbers in environmental samples. The measured relative abundances of comammox Nitrospira, compared to canonical ammonia

  9. PHUSER (Primer Help for USER): a novel tool for USER fusion primer design.

    Science.gov (United States)

    Olsen, Lars Rønn; Hansen, Niels Bjørn; Bonde, Mads Tvillinggaard; Genee, Hans Jasper; Holm, Dorte Koefoed; Carlsen, Simon; Hansen, Bjarne Gram; Patil, Kiran Raosaheb; Mortensen, Uffe Hasbro; Wernersson, Rasmus

    2011-07-01

    Uracil-Specific Exision Reagent (USER) fusion is a recently developed technique that allows for assembly of multiple DNA fragments in a few simple steps. However, designing primers for USER fusion is both tedious and time consuming. Here, we present the Primer Help for USER (PHUSER) software, a novel tool for designing primers specifically for USER fusion and USER cloning applications. We also present proof-of-concept experimental validation of its functionality. PHUSER offers quick and easy design of PCR optimized primers ensuring directionally correct fusion of fragments into a plasmid containing a customizable USER cassette. Designing primers using PHUSER ensures that the primers have similar annealing temperature (T(m)), which is essential for efficient PCR. PHUSER also avoids identical overhangs, thereby ensuring correct order of assembly of DNA fragments. All possible primers are individually analysed in terms of GC content, presence of GC clamp at 3'-end, the risk of primer dimer formation, the risk of intra-primer complementarity (secondary structures) and the presence of polyN stretches. Furthermore, PHUSER offers the option to insert linkers between DNA fragments, as well as highly flexible cassette options. PHUSER is publicly available at http://www.cbs.dtu.dk/services/phuser/.

  10. VizPrimer: a web server for visualized PCR primer design based on known gene structure.

    Science.gov (United States)

    Zhou, Yang; Qu, Wubin; Lu, Yiming; Zhang, Yanchun; Wang, Xiaolei; Zhao, Dongsheng; Yang, Yi; Zhang, Chenggang

    2011-12-15

    The visualization of gene structure plays an important role in polymerase chain reaction (PCR) primer design, especially for eukaryotic genes with a number of splice variants that users need to distinguish between via PCR. Here, we describe a visualized web server for primer design named VizPrimer. It utilizes the new information technology (IT) tools, HTML5 to display gene structure and JavaScript to interact with the users. In VizPrimer, the users can focus their attention on the gene structure and primer design strategy, without wasting time calculating the exon positions of splice variants or manually configuring complicated parameters. In addition, VizPrimer is also suitable for the design of PCR primers for amplifying open reading frames and detecting single nucleotide polymorphisms (SNPs). VizPrimer is freely available at http://biocompute.bmi.ac.cn/CZlab/VizPrimer/. The web server supported browsers: Chrome (≥5.0), Firefox (≥3.0), Safari (≥4.0) and Opera (≥10.0). zhangcg@bmi.ac.cn; yangyi528@vip.sina.com.

  11. Ionic strength of electrospray droplets affects charging of DNA oligonucleotides.

    Science.gov (United States)

    Xu, Ning; Chingin, Konstantin; Chen, Huanwen

    2014-01-01

    The fundamental aspects of charging in electrospray ionization (ESI) are hotly debated. In the present study, ESI charging of DNA oligonucleotides was explored in both positive (ESI+) and negative (ESI-) polarity using mass spectrometry detection. Single-stranded 12-mer CCCCAATTCCCC in buffer solution (aqueous NH4Ac, 100 mM) produced similar charge state distribution (CSD) in either ESI+ or ESI-. Similarity of CSD in ESI+ and ESI- was also observed for the double-stranded 12-mer CGCGAATTCGCG. By adding typical low-vapor reagents (e.g. m-nitro benzyl alcohol, m-NBA; sulfolane) into the same buffer solution (sulfolane or m-NBA, the CGCGAATTCGCG duplex dissociated into single strands in ESI-. No SC was observed in both ESI+ and ESI- for thermally denatured CGCGAATTCGCG duplex in NH4 Ac buffer without the reagents. These findings are difficult to reconcile with the earlier model, which attributes SC in aqueous buffer solution to the conformational changes of analytes. Our observations suggest that the ionic strength of ESI droplets strongly affects the CSD of biopolymers such as DNA oligonucleotides and that SC effect is related to the depletion of ionic strength during the ESI process. Copyright © 2014 John Wiley & Sons, Ltd.

  12. Multiplex pairwise assembly of array-derived DNA oligonucleotides.

    Science.gov (United States)

    Klein, Jason C; Lajoie, Marc J; Schwartz, Jerrod J; Strauch, Eva-Maria; Nelson, Jorgen; Baker, David; Shendure, Jay

    2016-03-18

    While the cost of DNA sequencing has dropped by five orders of magnitude in the past decade, DNA synthesis remains expensive for many applications. Although DNA microarrays have decreased the cost of oligonucleotide synthesis, the use of array-synthesized oligos in practice is limited by short synthesis lengths, high synthesis error rates, low yield and the challenges of assembling long constructs from complex pools. Toward addressing these issues, we developed a protocol for multiplex pairwise assembly of oligos from array-synthesized oligonucleotide pools. To evaluate the method, we attempted to assemble up to 2271 targets ranging in length from 192-252 bases using pairs of array-synthesized oligos. Within sets of complexity ranging from 131-250 targets, we observed error-free assemblies for 90.5% of all targets. When all 2271 targets were assembled in one reaction, we observed error-free constructs for 70.6%. While the assembly method intrinsically increased accuracy to a small degree, we further increased accuracy by using a high throughput 'Dial-Out PCR' protocol, which combines Illumina sequencing with an in-house set of unique PCR tags to selectively amplify perfect assemblies from complex synthetic pools. This approach has broad applicability to DNA assembly and high-throughput functional screens. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  13. Synthesis of peptide-oligonucleotide conjugates for chromium coordination.

    Science.gov (United States)

    Civitello, E R; Leniek, R G; Hossler, K A; Haebe, K; Stearns, D M

    2001-01-01

    The synthesis of the first peptide-oligonucleotide conjugate designed to coordinate chromium(III) is reported. The overall goal of this work is to synthesize di-deoxynucleotides tethered with chromium(III)-coordinating appendages to model chromium-DNA-protein cross-links, which are a type of DNA lesion that may be involved in chromium-induced cancers. The conjugate dGp(NHCH(2)CH(2)S-Ac-Gly-Ser-Gly-OH)G was made by coupling the peptide, ClAc-Gly-Ser-Gly-OH, and dinucleotide, dGp(NHCH(2)CH(2)SH)G, through a thioether moiety. The conjugate was characterized by HPLC and mass spectrometry. Previously reported methods for small-scale solid-phase synthesis of peptides and dinucleotide were unsuitable; therefore, gram-scale solution-phase methods were developed. We also report the gram-scale syntheses of two other serine-containing peptides, ClAc-betaAla-Ser-Gly-OH and ClAc-Ser-Gly-OH, and three histidine-containing peptides, ClAc-Gly-His-Gly-OH, ClAc-betaAla-His-Gly-OH, and ClAc-His-Gly-OH. The synthesis and characterization of chromium-containing peptide-oligonucleotide conjugates will ultimately help us to understand chromium-DNA interactions at a molecular level, which is necessary before we can determine how chromium causes cancer.

  14. The use of oligonucleotide probes for meningococcal serotype characterization

    Directory of Open Access Journals (Sweden)

    SACCHI Claudio Tavares

    1998-01-01

    Full Text Available In the present study we examine the potential use of oligonucleotide probes to characterize Neisseria meningitidis serotypes without the use of monoclonal antibodies (MAbs. Antigenic diversity on PorB protein forms the bases of serotyping method. However, the current panel of MAbs underestimated, by at least 50% the PorB variability, presumably because reagents for several PorB variable regions (VRs are lacking, or because a number of VR variants are not recognized by serotype-defining MAbs12. We analyzed the use of oligonucleotide probes to characterize serotype 10 and serotype 19 of N. meningitidis. The porB gene sequence for the prototype strain of serotype 10 was determined, aligned with 7 other porB sequences from different serotypes, and analysis of individual VRs were performed. The results of DNA probes 21U (VR1-A and 615U (VR3-B used against 72 N. meningitidis strains confirm that VR1 type A and VR3 type B encode epitopes for serotype-defined MAbs 19 and 10, respectively. The use of probes for characterizing serotypes possible can type 100% of the PorB VR diversity. It is a simple and rapid method specially useful for analysis of large number of samples.

  15. Sense oligonucleotide competition for gene promoter binding and activation.

    Science.gov (United States)

    Cutroneo, Kenneth R; Chiu, Jen-Fu

    2003-01-01

    Considerable evidence has ensued on the importance of growth factors during regeneration both for cell replication and for stimulation of reparative cells to synthesize and secrete extracellular matrix components. During the healing process if the growth factor concentration is too high because of over-expression, abnormal wound healing and tissue fibrosis will occur. The growth factor concentration at the wound site may be controlled by gene therapy and the titration of gene dosage. However, if there is a narrow window between the beneficial effects and adverse effects of gene therapy, oligonucleotide approaches may be used concurrently with gene therapy to control growth factor concentration(s) at the wound site. Antisense oligos offer a method to control the concentration of growth factors at the level of translation. A novel method using sense oligos to the proalpha1 (I) collagen gene to inhibit gene transcription and collagen synthesis has recently been reported. The exogenous modified oligodeoxynucleotide competes with the cis-element (i.e. the transforming growth factor-beta (TGF-beta) element) in the distal 5'-flanking region of the proalpha1 (I) collagen gene for the trans-acting factor (i.e. the TGF-beta activator protein complex), thereby down regulating promoter activity of the proalpha1 (I) collagen gene and inhibiting type I collagen synthesis. The oligonucleotide approaches, both antisense and sense therapies, may be used to regulate over-expression of growth factors and thereby either eliminate or lessen the potential adverse effects of gene therapy.

  16. Adsorption of DNA oligonucleotides by titanium dioxide nanoparticles.

    Science.gov (United States)

    Zhang, Xu; Wang, Feng; Liu, Biwu; Kelly, Erin Y; Servos, Mark R; Liu, Juewen

    2014-01-28

    Titanium dioxide (TiO2) or titania shows great promise in detoxification and drug delivery. To reach its full potential, it is important to interface TiO2 with biomolecules to harness their molecular recognition function. To this end, DNA attachment is an important topic. Previous work has mainly focused on long double-stranded DNA or single nucleotides. For biosensor development and targeted drug delivery, it is more important to use single-stranded oligonucleotides. Herein, the interaction between fluorescently labeled oligonucleotides and TiO2 nanoparticles is reported. The point of zero charge (PZC) of TiO2 is around 6 in water or acetate buffer; therefore, the particles are positively charged at lower pH. However, if in phosphate or citrate buffer, the particles are negatively charged, even at pH ∼2, suggesting strong adsorption of buffer anions. DNA adsorption takes place mainly via the phosphate backbone, although the bases might also have moderate contributions. Peptide nucleic acids (PNAs) with an amide backbone cannot be adsorbed. DNA adsorption is strongly affected by inorganic anions, where phosphate and citrate can strongly inhibit DNA adsorption. DNA adsorption is promoted by adding salt or lowering pH. DNA adsorption is accompanied with fluorescence quenching, and double-stranded DNA showed reduced quenching, allowing for the detection of DNA using TiO2 nanoparticles.

  17. Fusion primer and nested integrated PCR (FPNI-PCR): a new high-efficiency strategy for rapid chromosome walking or flanking sequence cloning

    Science.gov (United States)

    2011-01-01

    Background The advent of genomics-based technologies has revolutionized many fields of biological enquiry. However, chromosome walking or flanking sequence cloning is still a necessary and important procedure to determining gene structure. Such methods are used to identify T-DNA insertion sites and so are especially relevant for organisms where large T-DNA insertion libraries have been created, such as rice and Arabidopsis. The currently available methods for flanking sequence cloning, including the popular TAIL-PCR technique, are relatively laborious and slow. Results Here, we report a simple and effective fusion primer and nested integrated PCR method (FPNI-PCR) for the identification and cloning of unknown genomic regions flanked known sequences. In brief, a set of universal primers was designed that consisted of various 15-16 base arbitrary degenerate oligonucleotides. These arbitrary degenerate primers were fused to the 3' end of an adaptor oligonucleotide which provided a known sequence without degenerate nucleotides, thereby forming the fusion primers (FPs). These fusion primers are employed in the first step of an integrated nested PCR strategy which defines the overall FPNI-PCR protocol. In order to demonstrate the efficacy of this novel strategy, we have successfully used it to isolate multiple genomic sequences namely, 21 orthologs of genes in various species of Rosaceace, 4 MYB genes of Rosa rugosa, 3 promoters of transcription factors of Petunia hybrida, and 4 flanking sequences of T-DNA insertion sites in transgenic tobacco lines and 6 specific genes from sequenced genome of rice and Arabidopsis. Conclusions The successful amplification of target products through FPNI-PCR verified that this novel strategy is an effective, low cost and simple procedure. Furthermore, FPNI-PCR represents a more sensitive, rapid and accurate technique than the established TAIL-PCR and hiTAIL-PCR procedures. PMID:22093809

  18. Logistic regression: a brief primer.

    Science.gov (United States)

    Stoltzfus, Jill C

    2011-10-01

    Regression techniques are versatile in their application to medical research because they can measure associations, predict outcomes, and control for confounding variable effects. As one such technique, logistic regression is an efficient and powerful way to analyze the effect of a group of independent variables on a binary outcome by quantifying each independent variable's unique contribution. Using components of linear regression reflected in the logit scale, logistic regression iteratively identifies the strongest linear combination of variables with the greatest probability of detecting the observed outcome. Important considerations when conducting logistic regression include selecting independent variables, ensuring that relevant assumptions are met, and choosing an appropriate model building strategy. For independent variable selection, one should be guided by such factors as accepted theory, previous empirical investigations, clinical considerations, and univariate statistical analyses, with acknowledgement of potential confounding variables that should be accounted for. Basic assumptions that must be met for logistic regression include independence of errors, linearity in the logit for continuous variables, absence of multicollinearity, and lack of strongly influential outliers. Additionally, there should be an adequate number of events per independent variable to avoid an overfit model, with commonly recommended minimum "rules of thumb" ranging from 10 to 20 events per covariate. Regarding model building strategies, the three general types are direct/standard, sequential/hierarchical, and stepwise/statistical, with each having a different emphasis and purpose. Before reaching definitive conclusions from the results of any of these methods, one should formally quantify the model's internal validity (i.e., replicability within the same data set) and external validity (i.e., generalizability beyond the current sample). The resulting logistic regression model

  19. RExPrimer: an integrated primer designing tool increases PCR effectiveness by avoiding 3' SNP-in-primer and mis-priming from structural variation

    Directory of Open Access Journals (Sweden)

    Suwannasri Payiarat

    2009-12-01

    Full Text Available Abstract Background Polymerase chain reaction (PCR is very useful in many areas of molecular biology research. It is commonly observed that PCR success is critically dependent on design of an effective primer pair. Current tools for primer design do not adequately address the problem of PCR failure due to mis-priming on target-related sequences and structural variations in the genome. Methods We have developed an integrated graphical web-based application for primer design, called RExPrimer, which was written in Python language. The software uses Primer3 as the primer designing core algorithm. Locally stored sequence information and genomic variant information were hosted on MySQLv5.0 and were incorporated into RExPrimer. Results RExPrimer provides many functionalities for improved PCR primer design. Several databases, namely annotated human SNP databases, insertion/deletion (indel polymorphisms database, pseudogene database, and structural genomic variation databases were integrated into RExPrimer, enabling an effective without-leaving-the-website validation of the resulting primers. By incorporating these databases, the primers reported by RExPrimer avoid mis-priming to related sequences (e.g. pseudogene, segmental duplication as well as possible PCR failure because of structural polymorphisms (SNP, indel, and copy number variation (CNV. To prevent mismatching caused by unexpected SNPs in the designed primers, in particular the 3' end (SNP-in-Primer, several SNP databases covering the broad range of population-specific SNP information are utilized to report SNPs present in the primer sequences. Population-specific SNP information also helps customize primer design for a specific population. Furthermore, RExPrimer offers a graphical user-friendly interface through the use of scalable vector graphic image that intuitively presents resulting primers along with the corresponding gene structure. In this study, we demonstrated the program

  20. RExPrimer: an integrated primer designing tool increases PCR effectiveness by avoiding 3' SNP-in-primer and mis-priming from structural variation.

    Science.gov (United States)

    Piriyapongsa, Jittima; Ngamphiw, Chumpol; Assawamakin, Anunchai; Wangkumhang, Pongsakorn; Suwannasri, Payiarat; Ruangrit, Uttapong; Agavatpanitch, Gallissara; Tongsima, Sissades

    2009-12-03

    Polymerase chain reaction (PCR) is very useful in many areas of molecular biology research. It is commonly observed that PCR success is critically dependent on design of an effective primer pair. Current tools for primer design do not adequately address the problem of PCR failure due to mis-priming on target-related sequences and structural variations in the genome. We have developed an integrated graphical web-based application for primer design, called RExPrimer, which was written in Python language. The software uses Primer3 as the primer designing core algorithm. Locally stored sequence information and genomic variant information were hosted on MySQLv5.0 and were incorporated into RExPrimer. RExPrimer provides many functionalities for improved PCR primer design. Several databases, namely annotated human SNP databases, insertion/deletion (indel) polymorphisms database, pseudogene database, and structural genomic variation databases were integrated into RExPrimer, enabling an effective without-leaving-the-website validation of the resulting primers. By incorporating these databases, the primers reported by RExPrimer avoid mis-priming to related sequences (e.g. pseudogene, segmental duplication) as well as possible PCR failure because of structural polymorphisms (SNP, indel, and copy number variation (CNV)). To prevent mismatching caused by unexpected SNPs in the designed primers, in particular the 3' end (SNP-in-Primer), several SNP databases covering the broad range of population-specific SNP information are utilized to report SNPs present in the primer sequences. Population-specific SNP information also helps customize primer design for a specific population. Furthermore, RExPrimer offers a graphical user-friendly interface through the use of scalable vector graphic image that intuitively presents resulting primers along with the corresponding gene structure. In this study, we demonstrated the program effectiveness in successfully generating primers for strong

  1. Beam shaping for laser initiated optical primers

    Science.gov (United States)

    Lizotte, Todd E.

    2008-08-01

    Remington was one of the first firearm manufacturing companies to file a patent for laser initiated firearms, in 1969. Nearly 40 years later, the development of laser initiated firearms has not become a mainstream technology in the civilian market. Requiring a battery is definitely a short coming, so it is easy to see how such a concept would be problematic. Having a firearm operate reliably and the delivery of laser energy in an efficient manner to ignite the shock-sensitive explosive primer mixtures is a tall task indeed. There has been considerable research on optical element based methods of transferring or compressing laser energy to ignite primer charges, including windows, laser chip primers and various lens shaped windows to focus the laser energy. The focusing of laser light needs to achieve igniting temperatures upwards of >400°C. Many of the patent filings covering this type of technology discuss simple approaches where a single point of light might be sufficient to perform this task. Alternatively a multi-point method might provide better performance, especially for mission critical applications, such as precision military firearms. This paper covers initial design and performance test of the laser beam shaping optics to create simultaneous multiple point ignition locations and a circumferential intense ring for igniting primer charge compounds. A simple initial test of the ring beam shaping technique was evaluated on a standard large caliber primer to determine its effectiveness on igniting the primer material. Several tests were conducted to gauge the feasibility of laser beam shaping, including optic fabrication and mounting on a cartridge, optic durability and functional ignition performance. Initial data will be presented, including testing of optically elements and empirical primer ignition / burn analysis.

  2. and OMeN-modified oligonucleotide probes on the stability and ...

    Indian Academy of Sciences (India)

    Supplementary table 1a. Melting temperatures (°C) of duplexes containing a DNA:DNA mismatch. Base pair ... Sequence. Nature of primers with template. Forward primer 1 ... The four DNA forward primers and the four LNA forward primers were designed to include matching and all possible mismatchingnucleotides at the ...

  3. Removal of PCR error products and unincorporated primers by metal-chelate affinity chromatography.

    Directory of Open Access Journals (Sweden)

    Indhu Kanakaraj

    Full Text Available Immobilized Metal Affinity Chromatography (IMAC has been used for decades to purify proteins on the basis of amino acid content, especially surface-exposed histidines and "histidine tags" genetically added to recombinant proteins. We and others have extended the use of IMAC to purification of nucleic acids via interactions with the nucleotide bases, especially purines, of single-stranded RNA and DNA. We also have demonstrated the purification of plasmid DNA from contaminating genomic DNA by IMAC capture of selectively-denatured genomic DNA. Here we describe an efficient method of purifying PCR products by specifically removing error products, excess primers, and unincorporated dNTPs from PCR product mixtures using flow-through metal-chelate affinity adsorption. By flowing a PCR product mixture through a Cu(2+-iminodiacetic acid (IDA agarose spin column, 94-99% of the dNTPs and nearly all the primers can be removed. Many of the error products commonly formed by Taq polymerase also are removed. Sequencing of the IMAC-processed PCR product gave base-calling accuracy comparable to that obtained with a commercial PCR product purification method. The results show that IMAC matrices (specifically Cu(2+-IDA agarose can be used for the purification of PCR products. Due to the generality of the base-specific mechanism of adsorption, IMAC matrices may also be used in the purification of oligonucleotides, cDNA, mRNA and micro RNAs.

  4. Synthesis of 3'-, or 5'-, or internal methacrylamido-modified oligonucleotides

    Science.gov (United States)

    Golova, Julia B.; Chernov, Boris K.

    2010-04-27

    New modifiers were synthesized for incorporation of a methacrylic function in 3'-, 5'- and internal positions of oligonucleotides during solid phase synthesis. A modifier was used for synthesis of 5'-methacrylated oligonucleotides for preparation of microarrays by a co-polymerization method.

  5. Phosphorothioate oligonucleotides induction into experimental choroidal neovascularization by HVJ-liposome system.

    Science.gov (United States)

    Ogata, N; Otsuji, T; Matsushima, M; Kimoto, T; Yamanaka, R; Takahashi, K; Wada, M; Uyama, M; Kaneda, Y

    1999-04-01

    The purpose of this study was to determine whether the inactivated hemagglutinating virus of Japan (HVJ)-liposome method can induce phosphorothioate oligonucleotides effectively into an experimentally-induced choroidal neovascularization of rats. We also examined whether antisense phosphorothioate oligonucleotides against VEGF could be induced into choroidal neovascularization as a therapeutic agent by the HVJ-liposome method. The experiments were conducted on a rat model of choroidal neovascularization. FITC-labeled phosphorothioate oligonucleotides were coencapsulated in liposomes. The liposomes were coated with the envelope of inactivated HVJ and injected into the vitreous cavity following photocoagulation of pigmented rat eyes. The eyes were removed following injection, fixed, frozen and cut into thin sections. Induction of oligonucleotides was observed under a laser confocal scanning microscope for fluorescence and the development of choroidal neovascularization was evaluated histopathologically. Phosphorothioate oligonucleotides were effectively induced into ganglion cells and into the cells of the choroidal neovascularization induced by laser photocoagulation. Highly effective induction of oligos was observed 3 to 14 days after intravitreal injection of HVJ-liposomes after which the level decreased. Antisense oligonucleotides against VEGF were induced specifically into cells in the choroidal neovascularization, however neovascularization was still observed. Phosphorothioate oligonucleotides can be effectively induced into ganglion cells, and specifically into cells in choroidal neovascularization. Although antisense oligonucleotides against VEGF failed to prevent choroidal neovascularization, the HVJ-liposome method provided a highly effective means of inducing antisense oligos for in vivo antisense therapy.

  6. Thioctic Acid Derivatives as Building Blocks to Incorporate DNA Oligonucleotides onto Gold Nanoparticles

    Directory of Open Access Journals (Sweden)

    Sónia Pérez-Rentero

    2014-07-01

    Full Text Available Oligonucleotide gold nanoparticle conjugates are being used as diagnostic tools and gene silencing experiments. Thiol-chemistry is mostly used to functionalize gold nanoparticles with oligonucleotides and to incorporate DNA or RNA molecules onto gold surfaces. However, the stability of such nucleic acid–gold nanoparticle conjugates in certain conditions may be a limitation due to premature break of the thiol-gold bonds followed by aggregation processes. Here, we describe a straightforward synthesis of oligonucleotides carrying thioctic acid moiety based on the use of several thioctic acid-L-threoninol derivatives containing different spacers, including triglycine, short polyethyleneglycol, or aliphatic spacers. The novel thioctic-oligonucleotides were used for the functionalization of gold nanoparticles and the surface coverage and stability of the resulting thioctic-oligonucleotide gold nanoparticles were assessed. In all cases gold nanoparticles functionalized with thioctic-oligonucleotides had higher loadings and higher stability in the presence of thiols than gold nanoparticles prepared with commercially available thiol-oligonucleotides. Furthermore, the thioctic derivative carrying the triglycine linker is sensitive to cathepsin B present in endosomes. In this way this derivative may be interesting for the cellular delivery of therapeutic oligonucleotides as these results provides the basis for a potential endosomal escape.

  7. Deliberate change without hierarchical influence?

    DEFF Research Database (Denmark)

    Nørskov, Sladjana; Kesting, Peter; Ulhøi, John Parm

    2017-01-01

    Purpose This paper aims to present that deliberate change is strongly associated with formal structures and top-down influence. Hierarchical configurations have been used to structure processes, overcome resistance and get things done. But is deliberate change also possible without formal...... reveals that deliberate change is indeed achievable in a non-hierarchical collaborative OSS community context. However, it presupposes the presence and active involvement of informal change agents. The paper identifies and specifies four key drivers for change agents’ influence. Originality....../value The findings contribute to organisational analysis by providing a deeper understanding of the importance of leadership in making deliberate change possible in non-hierarchical settings. It points to the importance of “change-by-conviction”, essentially based on voluntary behaviour. This can open the door...

  8. Static Correctness of Hierarchical Procedures

    DEFF Research Database (Denmark)

    Schwartzbach, Michael Ignatieff

    1990-01-01

    basis for a general type hierarchy with static type checking, which enables first-order polymorphism combined with multiple inheritance and specialization in a language with assignments. We extend the results to include opaque types. An opaque version of a type is different from the original but has......A system of hierarchical, fully recursive types in a truly imperative language allows program fragments written for small types to be reused for all larger types. To exploit this property to enable type-safe hierarchical procedures, it is necessary to impose a static requirement on procedure calls....... We introduce an example language and prove the existence of a sound requirement which preserves static correctness while allowing hierarchical procedures. This requirement is further shown to be optimal, in the sense that it imposes as few restrictions as possible. This establishes the theoretical...

  9. Nucleobase-modified antisense oligonucleotides containing 5-(phenyltriazol)-2′-deoxyuridine nucleotides induce exon-skipping

    DEFF Research Database (Denmark)

    Le, Bao T.; Hornum, Mick; Sharma, Pawan K.

    2017-01-01

    Chemically-modified antisense oligonucleotide-mediated exon-skipping has been validated as a therapeutic strategy for tackling several disease pathologies, particularly duchenne muscular dystrophy. To date, only sugar-modified and internucleotide linkage-modified oligonucleotide chemistries have...

  10. Interfacing click chemistry with automated oligonucleotide synthesis for the preparation of fluorescent DNA probes containing internal xanthene and cyanine dyes

    DEFF Research Database (Denmark)

    Astakhova, I Kira; Wengel, Jesper

    2013-01-01

    of novel oligonucleotide probes containing internally positioned xanthene and cyanine dye pairs and thus represents a significant step forward for the preparation of advanced fluorescent oligonucleotide probes. Furthermore, we demonstrate that the novel xanthene and cyanine labeled probes display unusual...

  11. Shotgun sequencing small oligonucleotides by nozzle-skimmer dissociation and electrospray ionization mass spectrometry.

    Science.gov (United States)

    Meng, Zhaojing; Limbach, Patrick A

    2005-01-01

    Nozzle-skimmer dissociation in combination with de novo sequencing was investigated as an approach for increasing the throughput of oligonucleotide analysis attainable by electrospray ionization mass spectrometry. An experimental method allowing for the sequential generation of precursor and fragment ion data during direct infusion of sample was developed. These data can then be used with readily available de novo sequencing software to characterize small oligonucleotides. When this approach was applied to mixtures of oligonucleotides, it was found that de novo sequencing becomes limited due to spectral congestion and overlapping oligonucleotide m/z dissociation product values. Self-packed C(18) microspray emitters were investigated as a means of reducing spectral complexity. It was found that such emitters allow for the analysis of oligonucleotide mixtures with minimal component overlap, and these emitters provide additional benefits of pre- concentrating and desalting the sample. These developments can provide a route for the more rapid characterization of ribonucleic acid endonuclease digestion mixtures.

  12. Design of oligonucleotides for microarrays and perspectives for design of multi-transcriptome arrays

    DEFF Research Database (Denmark)

    Nielsen, Henrik Bjørn; Wernersson, Rasmus; Knudsen, Steen

    2003-01-01

    with an overview of these parameters. We present here a flexible tool named OligoWiz for designing oligonucleotides for multiple purposes. OligoWiz presents a set of parameter scores in a graphical interface to facilitate an overview for the user. Additional custom parameter scores can easily be added......Optimal design of oligonucleotides for microarrays involves tedious and laborious work evaluating potential oligonucleotides relative to a series of parameters. The currently available tools for this purpose are limited in their flexibility and do not present the oligonucleotide designer...... to the program to extend the default parameters: homology, DeltaTm, low-complexity, position and GATC-only. Furthermore we present an analysis of the limitations in designing oligonucleotide sets that can detect transcripts from multiple organisms. OligoWiz is available at www.cbs.dtu.dk/services/OligoWiz/....

  13. Validation of a spectrophotometric method to estimate the adsorption on nanoemulsions of an antimalarial oligonucleotide

    Directory of Open Access Journals (Sweden)

    Fernanda Bruxel

    2011-09-01

    Full Text Available This study describes the validation of a spectrophotometric method to estimate oligonucleotides association with cationic nanoemulsions. Phosphodiester and phosphorothioate oligonucleotides targeting Plasmodium falciparum topoisomerase II were analyzed at 262 nm. Linear response (r > 0.998 was observed from 0.4 to 1.0 nmol/mL, the relative standard deviation values for the intra- and inter-days precision were lower than 2.6% and the recovery ranged from 98.8 to 103.6% for both oligonucleotides. The association efficiency was estimated based on an ultrafiltration/centrifugation method. Oligonucleotides recovery through 30 kDa-membranes was higher than 92%. The extent of oligonucleotides association (42 to 98% varied with the composition of nanoemulsions

  14. Structural integrity of hierarchical composites

    Directory of Open Access Journals (Sweden)

    Marco Paggi

    2012-01-01

    Full Text Available Interface mechanical problems are of paramount importance in engineering and materials science. Traditionally, due to the complexity of modelling their mechanical behaviour, interfaces are often treated as defects and their features are not explored. In this study, a different approach is illustrated, where the interfaces play an active role in the design of innovative hierarchical composites and are fundamental for their structural integrity. Numerical examples regarding cutting tools made of hierarchical cellular polycrystalline materials are proposed, showing that tailoring of interface properties at the different scales is the way to achieve superior mechanical responses that cannot be obtained using standard materials

  15. Ferrocene-oligonucleotide conjugates for electrochemical probing of DNA.

    Science.gov (United States)

    Ihara, T; Maruo, Y; Takenaka, S; Takagi, M

    1996-01-01

    Toward the development of a universal, sensitive and convenient method of DNA (or RNA) detection, electrochemically active oligonucleotides were prepared by covalent linkage of a ferrocenyl group to the 5'-aminohexyl-terminated synthetic oligonucleotides. Using these electrochemically active probes, we have been able to demonstrate the detection of DNA and RNA at femtomole levels by HPLC equipped with an ordinary electrochemical detector (ECD) [Takenaka,S., Uto,Y., Kondo,H., Ihara,T. and Takagi,M. (1994) Anal. Biochem., 218, 436-443]. Thermodynamic and electrochemical studies of the interaction between the probes and the targets are presented here. The thermodynamics obtained revealed that the conjugation stabilizes the triple-helix complexes by 2-3 kcal mol-1 (1-2 orders increment in binding constant) at 298 K, which corresponds to the effect of elongation of additional several base triplets. The main cause of this thermodynamic stabilization by the conjugation is likely to be the overall conformational change of whole structure of the conjugate rather than the additional local interaction. The redox potential of the probe was independent of the target structure, which is either single- or double stranded. However, the potential is slightly dependent (with a 10-30 mV negative shift on complexation) on the extra sequence in the target, probably because the individual sequence is capable of contacting or interacting with the ferrocenyl group in a slightly different way from each other. This small potential shift itself, however, does not cause any inconvenience on practical applications in detecting the probes by using ECD. These results lead to the conclusion that the redox-active probes are very useful for the microanalysis of nucleic acids due to the stability of the complexes, high detection sensitivity and wide applicability to the target structures (DNA and RNA; single- and double strands) and the sequences. PMID:8932383

  16. Direct microcontact printing of oligonucleotides for biochip applications

    Directory of Open Access Journals (Sweden)

    Trévisiol E

    2005-07-01

    Full Text Available Abstract Background A critical step in the fabrication of biochips is the controlled placement of probes molecules on solid surfaces. This is currently performed by sequential deposition of probes on a target surface with split or solid pins. In this article, we present a cost-effective procedure namely microcontact printing using stamps, for a parallel deposition of probes applicable for manufacturing biochips. Results Contrary to a previous work, we showed that the stamps tailored with an elastomeric poly(dimethylsiloxane material did not require any surface modification to be able to adsorb oligonucleotides or PCR products. The adsorbed DNA molecules are subsequently printed efficiently on a target surface with high sub-micron resolution. Secondly, we showed that successive stamping is characterized by an exponential decay of the amount of transferred DNA molecules to the surface up the 4th print, then followed by a second regime of transfer that was dependent on the contact time and which resulted in reduced quality of the features. Thus, while consecutive stamping was possible, this procedure turned out to be less reproducible and more time consuming than simply re-inking the stamps between each print. Thirdly, we showed that the hybridization signals on arrays made by microcontact printing were 5 to 10-times higher than those made by conventional spotting methods. Finally, we demonstrated the validity of this microcontact printing method in manufacturing oligonucleotides arrays for mutations recognition in a yeast gene. Conclusion The microcontact printing can be considered as a new potential technology platform to pattern DNA microarrays that may have significant advantages over the conventional spotting technologies as it is easy to implement, it uses low cost material to make the stamp, and the arrays made by this technology are 10-times more sensitive in term of hybridization signals than those manufactured by conventional spotting

  17. Hierarchical Engineered Materials and Structures

    Science.gov (United States)

    2012-11-30

    of lightweight cellular materials (such as honeycombs and foams ) which are commonly used in "hierarchically designed" structural materials, especially...response past the elevated yield stress is modeled as perfectly plastic. The viscoelastic properties are not modeled because it is assumed that the

  18. Microsatellite Primers for Fungus-Growing Ants

    DEFF Research Database (Denmark)

    Villesen Fredsted, Palle; Gertsch, Pia J.; Boomsma, Jacobus Jan (Koos)

    2002-01-01

    We isolated five polymorphic microsatellite loci from a library of two thousand recombinant clones of two fungus-growing ant species, Cyphomyrmex longiscapus and Trachymyrmex cf. zeteki. Amplification and heterozygosity were tested in five species of higher attine ants using both the newly...... developed primers and earlier published primers that were developed for fungus-growing ants. A total of 20 variable microsatellite loci, developed for six different species of fungus-growing ants, are now available for studying the population genetics and colony kin-structure of these ants....

  19. Microsatellite primers for fungus-growing ants

    DEFF Research Database (Denmark)

    Villesen, Palle; Gertsch, P J; Boomsma, JJ

    2002-01-01

    We isolated five polymorphic microsatellite loci from a library of two thousand recombinant clones of two fungus-growing ant species, Cyphomyrmex longiscapus and Trachymyrmex cf. zeteki. Amplification and heterozygosity were tested in five species of higher attine ants using both the newly...... developed primers and earlier published primers that were developed for fungus-growing ants. A total of 20 variable microsatellite loci, developed for six different species of fungus-growing ants, are now available for studying the population genetics and colony kin-structure of these ants....

  20. Comparison between Mt-DNA D-Loop and Cyt B primers for porcine DNA detection in meat products

    Science.gov (United States)

    Hamzah, Azhana; Mutalib, Sahilah Abd.; Babji, Abdul Salam

    2013-11-01

    This study was conducted to detect the presence of porcine DNA in meat products in the market using conventional polymerase chain reaction (PCR) and commercial PCR-southern hybridization analysis. Porcine DNA detection in meat products was tested due to some issues associated with the adulteration of food products in Malaysia. This is an important issue especially for Halal authentication which is required for some religious practices such as in Islam and Hinduisms. Many techniques have been developed for determining the Halal status of food products. In this paper, mt-DNA D-loop primer and cytochrome (cyt) b were used to detect the presence of porcine DNA in meat products. Positive and negative controls were always present for each batch of extraction. DNA of raw pork meat was used as a positive control while nucleus free water is used as negative control. A pair of oligonucleotide primer was used namely Pork1 and Pork2 which produced amplicon of 531 base pair (bp) in size. While, PCR-southern hybridization was conducted using primers readily supplied by commercial PCR-Southern hybridization and produced amplicon with 276 bp in size. In the present study, demonstrated that none of the samples were contaminated with porcine residuals but selected samples with pork meat were positive. The species-specific PCR amplification yielded excellent results for identification of pork derivatives in food products and it is a potentially reliable and suitable technique in routine food analysis for Halal certification.

  1. A new enzymatic route for production of long 5'-phosphorylated oligonucleotides using suicide cassettes and rolling circle DNA synthesis

    Directory of Open Access Journals (Sweden)

    Stougaard Magnus

    2007-08-01

    Full Text Available Abstract Background The quality of chemically synthesized oligonucleotides falls with the length of the oligonucleotide, not least due to depurinations and premature termination during production. This limits the use of long oligonucleotides in assays where long high-quality oligonucleotides are needed (e.g. padlock probes. Another problem with chemically synthesized oligonucleotides is that secondary structures contained within an oligonucleotide reduce the efficiency of HPLC and/or PAGE purification. Additionally, ligation of chemically synthesized oligonucleotides is less efficient than the ligation of enzymatically produced DNA molecules. Results Chemically synthesized oligonucleotides with hairpin structures were acquired from our standard supplier. The stem of the hairpin contained recognition sequences for the Nt. Alw I nicking enzyme and the Mly I restriction enzyme. These double stranded regions were positioned in a way to allow self-templated circularization of the oligonucleotide. Following ligation, tandem repeats of the complementary sequence of the circular oligonucleotide could be produced through rolling circle DNA synthesis. By running successive rounds of ligation, rolling circle DNA synthesis, and nicking, the original oligonucleotide could be amplified as either the (+-strand or the (--strand. Alternatively, the hairpin structure could be removed by cleavage with the Mly I restriction enzyme, thereby releasing the oligonucleotide sequence contained within the hairpin structure from the hairpin. Conclusion We present here a method for the enzymatic production through DNA amplification of oligonucleotides with freely designable 5'-ends and 3'-ends, using hairpin-containing self-templating oligonucleotides. The hairpin comprises recognition sequences for a nicking enzyme and a restriction enzyme. The oligonucleotides are amplified by successive rounds of ligation, rolling circle DNA synthesis and nicking. Furthermore, the

  2. Polymerase-endonuclease amplification reaction (PEAR for large-scale enzymatic production of antisense oligonucleotides.

    Directory of Open Access Journals (Sweden)

    Xiaolong Wang

    Full Text Available Antisense oligonucleotides targeting microRNAs or their mRNA targets prove to be powerful tools for molecular biology research and may eventually emerge as new therapeutic agents. Synthetic oligonucleotides are often contaminated with highly homologous failure sequences. Synthesis of a certain oligonucleotide is difficult to scale up because it requires expensive equipment, hazardous chemicals and a tedious purification process. Here we report a novel thermocyclic reaction, polymerase-endonuclease amplification reaction (PEAR, for the amplification of oligonucleotides. A target oligonucleotide and a tandem repeated antisense probe are subjected to repeated cycles of denaturing, annealing, elongation and cleaving, in which thermostable DNA polymerase elongation and strand slipping generate duplex tandem repeats, and thermostable endonuclease (PspGI cleavage releases monomeric duplex oligonucleotides. Each round of PEAR achieves over 100-fold amplification. The product can be used in one more round of PEAR directly, and the process can be further repeated. In addition to avoiding dangerous materials and improved product purity, this reaction is easy to scale up and amenable to full automation. PEAR has the potential to be a useful tool for large-scale production of antisense oligonucleotide drugs.

  3. Selection of long oligonucleotides for gene expression microarrays using weighted rank-sum strategy

    Directory of Open Access Journals (Sweden)

    Preiser Peter

    2007-09-01

    Full Text Available Abstract Background The design of long oligonucleotides for spotted DNA microarrays requires detailed attention to ensure their optimal performance in the hybridization process. The main challenge is to select an optimal oligonucleotide element that represents each genetic locus/gene in the genome and is unique, devoid of internal structures and repetitive sequences and its Tm is uniform with all other elements on the microarray. Currently, all of the publicly available programs for DNA long oligonucleotide microarray selection utilize various combinations of cutoffs in which each parameter (uniqueness, Tm, and secondary structure is evaluated and filtered individually. The use of the cutoffs can, however, lead to information loss and to selection of suboptimal oligonucleotides, especially for genomes with extreme distribution of the GC content, a large proportion of repetitive sequences or the presence of large gene families with highly homologous members. Results Here we present the program OligoRankPick which is using a weighted rank-based strategy to select microarray oligonucleotide elements via an integer weighted linear function. This approach optimizes the selection criteria (weight score for each gene individually, accommodating variable properties of the DNA sequence along the genome. The designed algorithm was tested using three microbial genomes Escherichia coli, Saccharomyces cerevisiae and the human malaria parasite species Plasmodium falciparum. In comparison to other published algorithms OligoRankPick provides significant improvements in oligonucleotide design for all three genomes with the most significant improvements observed in the microarray design for P. falciparum whose genome is characterized by large fluctuations of GC content, and abundant gene duplications. Conclusion OligoRankPick is an efficient tool for the design of long oligonucleotide DNA microarrays which does not rely on direct oligonucleotide exclusion by

  4. Issues Primer. EEE708 Negotiated Study Program.

    Science.gov (United States)

    Jennings, Leonie

    This issues primer is structured around a series of 20 contemporary concerns in the changing world of work and training in Australia in the early 1990s. It is part of the study materials for the one-semester distance education unit, Negotiated Study Program, in the Open Campus Program at Deakin University (Australia). Information on each issue is…

  5. Scrimer: designing primers from transcriptome data

    Czech Academy of Sciences Publication Activity Database

    Mořkovský, Libor; Pačes, Jan; Rídl, Jakub; Reifová, R.

    2015-01-01

    Roč. 15, č. 6 (2015), s. 1415-1420 ISSN 1755-098X R&D Projects: GA MŠk EE2.3.20.0303 Institutional support: RVO:68081766 ; RVO:68378050 Keywords : next-generation sequencing * primer design * SNaPshot * SNP genotyping * transcriptome Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.298, year: 2015

  6. A Primer on the Functional Equation

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 22; Issue 10. A Primer on the Functional Equation: f(x+y) = f(x) + f(y). Kaushal Verma. General Article Volume 22 Issue 10 October 2017 pp 935-941. Fulltext. Click here to view fulltext PDF. Permanent link:

  7. A Primer on Policies for Jobs

    OpenAIRE

    Bhattacharya, Rwitwika; Griffith, Breda; Chen, Derek H. C.; Nallari, Raj; Andriamananjara, Soamiely; Wang, Yidan

    2012-01-01

    A primer on policies for jobs is based on materials and input provided during the labor market courses conducted during the past 10 years. Its objective is to provide government policy makers, researchers, and labor market practitioners and other specialists with a practical guide on how to strengthen labor market institutions, especially in light of the global financial crisis. This prime...

  8. Reforming Science Instruction in California: A Primer

    Science.gov (United States)

    EdSource, 2017

    2017-01-01

    As schools across the state implement the Next Generation Science Standards, this EdSource primer provides an easy-to-read guide for parents and other community members to understand the rationale for the standards and their potential to affect science instruction in California schools. California adopted the Next Generation Science Standards, or…

  9. Hierarchical Prisoner's Dilemma in Hierarchical Public-Goods Game

    CERN Document Server

    Fujimoto, Yuma; Kaneko, Kunihiko

    2016-01-01

    The dilemma in cooperation is one of the major concerns in game theory. In a public-goods game, each individual pays a cost for cooperation, or to prevent defection, and receives a reward from the collected cost in a group. Thus, defection is beneficial for each individual, while cooperation is beneficial for the group. Now, groups (say, countries) consisting of individual players also play games. To study such a multi-level game, we introduce a hierarchical public-goods (HPG) game in which two groups compete for finite resources by utilizing costs collected from individuals in each group. Analyzing this HPG game, we found a hierarchical prisoner's dilemma, in which groups choose the defection policy (say, armaments) as a Nash strategy to optimize each group's benefit, while cooperation optimizes the total benefit. On the other hand, for each individual within a group, refusing to pay the cost (say, tax) is a Nash strategy, which turns to be a cooperation policy for the group, thus leading to a hierarchical d...

  10. Solid-phase synthesis of 2{sup '}-O-methoxyethyl oligonucleotides using dimeric phosphoramidate blocks

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Gi Weon; Kang, Yong Han [Dept. of Applied Chemistry, Hanyang University, Ansan (Korea, Republic of)

    2016-11-15

    This research focused on the method of using dimeric phosphoramidite blocks to synthesize oligonucleotides for development as oligonucleotide drugs. A 16-mer oligonucleotide with the randomly selected sequence of C*C*T*C*G*C *T*C*T*C*G*C*C* C*G*C was synthesized using CC, GC, and TC dimers, a combination of monomers and dimers, or only monomers as building blocks. Using dimer blocks in this synthetic method provided a significant decrease in critical impurities that had similar properties to the main product, which was confirmed by LC-MS and HPLC analysis.

  11. Synthesis and properties of cholesteryl-modified triple-helix forming oligonucleotides containing a triglycyl linker.

    Science.gov (United States)

    Vu, H; Hill, T S; Jayaraman, K

    1994-01-01

    In order to enhance the nuclear uptake of triple-helix forming oligonucleotides (TFOs), a triglycylcholesterol group was attached to the 3' end. The peptide unit was introduced as a "labile" linker with the aim of releasing the oligonucleotide from the endosomes by the action of peptidases after crossing the cell membrane. Cholesteryl-CPG (8) and -TentaGel (9) supports containing 2-[N-(glycylglycylglycyl)amino]propane-1,3-diol (GAP-3) linker were prepared and used for automated oligonucleotide synthesis. The synthesis, characterization, and stability of these compounds are described.

  12. Antisense c-myb oligonucleotides inhibit intimal arterial smooth muscle cell accumulation in vivo

    Science.gov (United States)

    Simons, Michael; Edelman, Elazer R.; Dekeyser, Jean-Luc; Langer, Robert; Rosenberg, Robert D.

    1992-09-01

    SYNTHETIC antisense oligonucleotides have been used to dissect gene function in vitro. Technical difficulties prevented the use of this approach for investigating the effect of gene products in vivo. Here we report the use of local delivery of antisense c-myb oligonu-cleotide to suppress intimal accumulation of rat carotid arterial smooth muscle cells. Our results suggest that antisense oligonucleotides can be used to define the in vivo biological role of specific macromolecules in the blood vessel wall and could potentially serve as a new class of therapeutic agents for cardiovascular disorders.

  13. Synthetic Method for Oligonucleotide Block by Using Alkyl-Chain-Soluble Support.

    Science.gov (United States)

    Matsuno, Yuki; Shoji, Takao; Kim, Shokaku; Chiba, Kazuhiro

    2016-02-19

    A straightforward method for the synthesis of oligonucleotide blocks using a Cbz-type alkyl-chain-soluble support (Z-ACSS) attached to the 3'-OH group of 3'-terminal nucleosides was developed. The Z-ACSS allowed for the preparation of fully protected deoxyribo- and ribo-oligonucleotides without chromatographic purification and released dimer- to tetramer-size oligonucleotide blocks via hydrogenation using a Pd/C catalyst without significant loss or migration of protective groups such as 5'-end 4,4'-dimethoxtrityl, 2-cyanoethyl on internucleotide bonds, or 2'-TBS.

  14. Universal primers for rapid detection of hytrosaviruses.

    Science.gov (United States)

    Abd-Alla, Adly M M; Salem, Tamer Z; Parker, Andrew G; Wang, Yongjie; Jehle, Johannes A; Vreysen, Marc J B; Boucias, Drion

    2011-01-01

    Hytrosaviridae is a proposed virus family encompassing viruses that cause salivary gland hypertrophy (SGH) syndrome in infected insects and reduce the fertility in their dipteran insect hosts. They contain a large, double stranded DNA genome of 120-190 kbp. To date, these viruses have been detected only in adult Diptera. These include hytrosaviruses detected in various tsetse fly species (Glossina spp.), the narcissus bulb fly Merodon equestris and the house fly Musca domestica. The limited number of hytrosaviruses reported to date may be a reflection of the frequent absence of external symptoms in infected adult flies and the fact that the virus does not cause rapid mortality. Based on the complete genome sequence of Glossinia pallidipes (GpSGHV) and Musca domestica (MdSGHV) salivary gland hypertrophy viruses, a PCR based methodology was developed to detect the viruses in these species. To be able to detect hytrosaviruses in other Diptera, five degenerate primer pairs were designed and tested on GpSGHV and MdSGHV DNA using gradient PCR with annealing temperatures from 37 to 61°C. Two pairs of primers were selected from p74, two pairs from PIF-1 and one pair from ODV-e66 homologous proteins. Four primer pairs generated a virus specific PCR product on both MdSGHV and GpSGHV at all tested annealing temperatures, while the ODV-e66 based primers did not generate a virus specific product with annealing temperatures higher that 47°C. No non-specific PCR product was found when using genomic DNA of infected flies as template DNA. These results offer new sets of primers that could be used to detect hytrosaviruses in other insects. Copyright © 2010 Elsevier B.V. All rights reserved.

  15. Multicast Routing of Hierarchical Data

    Science.gov (United States)

    Shacham, Nachum

    1992-01-01

    The issue of multicast of broadband, real-time data in a heterogeneous environment, in which the data recipients differ in their reception abilities, is considered. Traditional multicast schemes, which are designed to deliver all the source data to all recipients, offer limited performance in such an environment, since they must either force the source to overcompress its signal or restrict the destination population to those who can receive the full signal. We present an approach for resolving this issue by combining hierarchical source coding techniques, which allow recipients to trade off reception bandwidth for signal quality, and sophisticated routing algorithms that deliver to each destination the maximum possible signal quality. The field of hierarchical coding is briefly surveyed and new multicast routing algorithms are presented. The algorithms are compared in terms of network utilization efficiency, lengths of paths, and the required mechanisms for forwarding packets on the resulting paths.

  16. Hierarchical matrices algorithms and analysis

    CERN Document Server

    Hackbusch, Wolfgang

    2015-01-01

    This self-contained monograph presents matrix algorithms and their analysis. The new technique enables not only the solution of linear systems but also the approximation of matrix functions, e.g., the matrix exponential. Other applications include the solution of matrix equations, e.g., the Lyapunov or Riccati equation. The required mathematical background can be found in the appendix. The numerical treatment of fully populated large-scale matrices is usually rather costly. However, the technique of hierarchical matrices makes it possible to store matrices and to perform matrix operations approximately with almost linear cost and a controllable degree of approximation error. For important classes of matrices, the computational cost increases only logarithmically with the approximation error. The operations provided include the matrix inversion and LU decomposition. Since large-scale linear algebra problems are standard in scientific computing, the subject of hierarchical matrices is of interest to scientists ...

  17. High-performance hierarchical fracturing

    Science.gov (United States)

    Cobb, Nicolas B.; Zhang, Weidong

    2002-07-01

    We describe in more detail a mask data preparation (MDP) flow previously proposed. The focus on this paper is a performance comparison of hierarchical fracturing techniques compared to standard fracturing. Our flow uses GDSII data as input, including a GDSII-based job deck description. The output is maximally compacted, trapezoidal mask writer (MW) formatted data. Our flow takes advantage of hierarchy explicit in the GDSII file(s). This allows optimal determination of 'cover cells', which are repeatable groups of patterns within the data. The use of cover cells allows a reduction of fracturing runtime. In one case, a 21 GB MEBES file was fractured in 30 hours using the standard technique and 53 minutes using the hierarchical cover cell technique.

  18. Topological properties of hierarchical networks.

    Science.gov (United States)

    Agliari, Elena; Barra, Adriano; Galluzzi, Andrea; Guerra, Francesco; Tantari, Daniele; Tavani, Flavia

    2015-06-01

    Hierarchical networks are attracting a renewal interest for modeling the organization of a number of biological systems and for tackling the complexity of statistical mechanical models beyond mean-field limitations. Here we consider the Dyson hierarchical construction for ferromagnets, neural networks, and spin glasses, recently analyzed from a statistical-mechanics perspective, and we focus on the topological properties of the underlying structures. In particular, we find that such structures are weighted graphs that exhibit a high degree of clustering and of modularity, with a small spectral gap; the robustness of such features with respect to the presence of thermal noise is also studied. These outcomes are then discussed and related to the statistical-mechanics scenario in full consistency. Last, we look at these weighted graphs as Markov chains and we show that in the limit of infinite size, the emergence of ergodicity breakdown for the stochastic process mirrors the emergence of metastabilities in the corresponding statistical mechanical analysis.

  19. Topological properties of hierarchical networks

    Science.gov (United States)

    Agliari, Elena; Barra, Adriano; Galluzzi, Andrea; Guerra, Francesco; Tantari, Daniele; Tavani, Flavia

    2015-06-01

    Hierarchical networks are attracting a renewal interest for modeling the organization of a number of biological systems and for tackling the complexity of statistical mechanical models beyond mean-field limitations. Here we consider the Dyson hierarchical construction for ferromagnets, neural networks, and spin glasses, recently analyzed from a statistical-mechanics perspective, and we focus on the topological properties of the underlying structures. In particular, we find that such structures are weighted graphs that exhibit a high degree of clustering and of modularity, with a small spectral gap; the robustness of such features with respect to the presence of thermal noise is also studied. These outcomes are then discussed and related to the statistical-mechanics scenario in full consistency. Last, we look at these weighted graphs as Markov chains and we show that in the limit of infinite size, the emergence of ergodicity breakdown for the stochastic process mirrors the emergence of metastabilities in the corresponding statistical mechanical analysis.

  20. Automatic Hierarchical Color Image Classification

    Directory of Open Access Journals (Sweden)

    Jing Huang

    2003-02-01

    Full Text Available Organizing images into semantic categories can be extremely useful for content-based image retrieval and image annotation. Grouping images into semantic classes is a difficult problem, however. Image classification attempts to solve this hard problem by using low-level image features. In this paper, we propose a method for hierarchical classification of images via supervised learning. This scheme relies on using a good low-level feature and subsequently performing feature-space reconfiguration using singular value decomposition to reduce noise and dimensionality. We use the training data to obtain a hierarchical classification tree that can be used to categorize new images. Our experimental results suggest that this scheme not only performs better than standard nearest-neighbor techniques, but also has both storage and computational advantages.

  1. Hybrid and hierarchical composite materials

    CERN Document Server

    Kim, Chang-Soo; Sano, Tomoko

    2015-01-01

    This book addresses a broad spectrum of areas in both hybrid materials and hierarchical composites, including recent development of processing technologies, structural designs, modern computer simulation techniques, and the relationships between the processing-structure-property-performance. Each topic is introduced at length with numerous  and detailed examples and over 150 illustrations.   In addition, the authors present a method of categorizing these materials, so that representative examples of all material classes are discussed.

  2. Statistical significance for hierarchical clustering.

    Science.gov (United States)

    Kimes, Patrick K; Liu, Yufeng; Neil Hayes, David; Marron, James Stephen

    2017-09-01

    Cluster analysis has proved to be an invaluable tool for the exploratory and unsupervised analysis of high-dimensional datasets. Among methods for clustering, hierarchical approaches have enjoyed substantial popularity in genomics and other fields for their ability to simultaneously uncover multiple layers of clustering structure. A critical and challenging question in cluster analysis is whether the identified clusters represent important underlying structure or are artifacts of natural sampling variation. Few approaches have been proposed for addressing this problem in the context of hierarchical clustering, for which the problem is further complicated by the natural tree structure of the partition, and the multiplicity of tests required to parse the layers of nested clusters. In this article, we propose a Monte Carlo based approach for testing statistical significance in hierarchical clustering which addresses these issues. The approach is implemented as a sequential testing procedure guaranteeing control of the family-wise error rate. Theoretical justification is provided for our approach, and its power to detect true clustering structure is illustrated through several simulation studies and applications to two cancer gene expression datasets. © 2017, The International Biometric Society.

  3. Aluminum Rich Epoxy Primer for Ground and Air Vehicles

    Science.gov (United States)

    2017-03-01

    UNCLASSIFIED DOCUMENT Aluminum Rich Epoxy Primer for Ground and Air Vehicles Monthly Technical Report for the Period: January 20, 2017...Objective: To further develop the Aluminum Rich Epoxy Primer systems for Air and Ground Vehicles while addressing the objective requirements

  4. New primers for adhesive bonding of aluminum alloys

    Science.gov (United States)

    Burrell, B. W.; Port, W. S.

    1971-01-01

    Synthetic polypeptide adhesive primers are effective, with high temperature epoxy resins, at temperatures from 100 deg to 300 deg C. Lap-shear failure loads and lap-shear strength of both primers are discussed.

  5. Economics : pricing, demand, and economic efficiency : a primer.

    Science.gov (United States)

    2008-11-01

    The Congestion Pricing Primer Series is part of : FHWAs outreach efforts to introduce the various : aspects of congestion pricing to decision-makers and : transportation professionals in the United States. The : primers are intended to lay out the...

  6. Primer on consumer marketing research : procedures, methods, and tools

    Science.gov (United States)

    1994-03-01

    The Volpe Center developed a marketing research primer which provides a guide to the approach, procedures, and research tools used by private industry in predicting consumer response. The final two chapters of the primer focus on the challenges of do...

  7. Oligonucleotide-based biosensors for in vitro diagnostics and environmental hazard detection.

    Science.gov (United States)

    Jung, Il Young; Lee, Eun Hee; Suh, Ah Young; Lee, Seung Jin; Lee, Hyukjin

    2016-04-01

    Oligonucleotide-based biosensors have drawn much attention because of their broad applications in in vitro diagnostics and environmental hazard detection. They are particularly of interest to many researchers because of their high specificity as well as excellent sensitivity. Recently, oligonucleotide-based biosensors have been used to achieve not only genetic detection of targets but also the detection of small molecules, peptides, and proteins. This has further broadened the applications of these sensors in the medical and health care industry. In this review, we highlight various examples of oligonucleotide-based biosensors for the detection of diseases, drugs, and environmentally hazardous chemicals. Each example is provided with detailed schematics of the detection mechanism in addition to the supporting experimental results. Furthermore, future perspectives and new challenges in oligonucleotide-based biosensors are discussed.

  8. Exploiting protected maleimides to modify oligonucleotides, peptides and peptide nucleic acids.

    Science.gov (United States)

    Paris, Clément; Brun, Omar; Pedroso, Enrique; Grandas, Anna

    2015-04-10

    This manuscript reviews the possibilities offered by 2,5-dimethylfuran-protected maleimides. Suitably derivatized building blocks incorporating the exo Diels-Alder cycloadduct can be introduced at any position of oligonucleotides, peptide nucleic acids, peptides and peptoids, making use of standard solid-phase procedures. Maleimide deprotection takes place upon heating, which can be followed by either Michael-type or Diels-Alder click conjugation reactions. However, the one-pot procedure in which maleimide deprotection and conjugation are simultaneously carried out provides the target conjugate more quickly and, more importantly, in better yield. This procedure is compatible with conjugates involving oligonucleotides, peptides and peptide nucleic acids. A variety of cyclic peptides and oligonucleotides can be obtained from peptide and oligonucleotide precursors incorporating protected maleimides and thiols.

  9. Antiproliferative effects of antisense oligonucleotides directed to the RNA of c-myc oncogene.

    Science.gov (United States)

    Degols, G; Leonetti, J P; Mechti, N; Lebleu, B

    1991-01-01

    Several groups have reported the use of antisense oligonucleotides to inhibit c-myc gene expression and study its biological role. However high concentrations of free oligonucleotides were generally needed. To lower their concentration and stabilize the antisense effect against c-myc, oligonucleotides were covalently linked to poly(L-lysine) and administered in ternary complexes formed with heparin (100 micrograms/ml). A sequence specific growth inhibition was observed at concentrations lower than 1 microM, while oligonucleotide-poly(L-lysine) conjugates alone were inefficient. Similar results occurred with other polyanionic compounds. Inhibition of proliferation was correlated to a reduction of c-myc protein and to a transient decrease in c-myc mRNA level. However, implication of RNase H in this process could not be demonstrated. Images PMID:1708128

  10. Exploiting Protected Maleimides to Modify Oligonucleotides, Peptides and Peptide Nucleic Acids

    Directory of Open Access Journals (Sweden)

    Clément Paris

    2015-04-01

    Full Text Available This manuscript reviews the possibilities offered by 2,5-dimethylfuran-protected maleimides. Suitably derivatized building blocks incorporating the exo Diels-Alder cycloadduct can be introduced at any position of oligonucleotides, peptide nucleic acids, peptides and peptoids, making use of standard solid-phase procedures. Maleimide deprotection takes place upon heating, which can be followed by either Michael-type or Diels-Alder click conjugation reactions. However, the one-pot procedure in which maleimide deprotection and conjugation are simultaneously carried out provides the target conjugate more quickly and, more importantly, in better yield. This procedure is compatible with conjugates involving oligonucleotides, peptides and peptide nucleic acids. A variety of cyclic peptides and oligonucleotides can be obtained from peptide and oligonucleotide precursors incorporating protected maleimides and thiols.

  11. Direct in situ hybridization with oligonucleotide functionalized quantum dot probes.

    Science.gov (United States)

    Bentolila, Laurent A

    2010-01-01

    Coming from the material sciences, fluorescent semiconductor nanocrystals, also known as quantum dots (QDs), have emerged as powerful fluorescent probes for a wide range of biological imaging applications. QDs have several advantages over organic dyes which include higher brightness, better resistance to photobleaching, and simplified multicolor target detection. In this chapter, we describe a rapid assay for the direct imaging of multiple repetitive subnuclear genetic sequences using QD-based FISH probes. Streptavidin-coated QDs (SAvQDs) are functionalized with short biotinylated oligonucleotides and used in a single hybridization/detection step. These QD-FISH probes penetrate both intact interphase nuclei and metaphase chromosomes and show good targeting of dense chromatin domains. Importantly, the broad absorption spectra of QDs allows two sequence specific QD-FISH probes of different colors to be simultaneously imaged with a single laser excitation wavelength. This method, which requires minimal custom conjugation, is easily expandable and offers the experimentalist a new alternative to increase flexibility in multicolor cytogenetic FISH applications of repetitive DNAs.

  12. Linear model for fast background subtraction in oligonucleotide microarrays

    Directory of Open Access Journals (Sweden)

    Barkema Gerard T

    2009-11-01

    Full Text Available Abstract Background One important preprocessing step in the analysis of microarray data is background subtraction. In high-density oligonucleotide arrays this is recognized as a crucial step for the global performance of the data analysis from raw intensities to expression values. Results We propose here an algorithm for background estimation based on a model in which the cost function is quadratic in a set of fitting parameters such that minimization can be performed through linear algebra. The model incorporates two effects: 1 Correlated intensities between neighboring features in the chip and 2 sequence-dependent affinities for non-specific hybridization fitted by an extended nearest-neighbor model. Conclusion The algorithm has been tested on 360 GeneChips from publicly available data of recent expression experiments. The algorithm is fast and accurate. Strong correlations between the fitted values for different experiments as well as between the free-energy parameters and their counterparts in aqueous solution indicate that the model captures a significant part of the underlying physical chemistry.

  13. Chimeric Antisense Oligonucleotide Conjugated to α-Tocopherol

    Directory of Open Access Journals (Sweden)

    Tomoko Nishina

    2015-01-01

    Full Text Available We developed an efficient system for delivering short interfering RNA (siRNA to the liver by using α-tocopherol conjugation. The α-tocopherol–conjugated siRNA was effective and safe for RNA interference–mediated gene silencing in vivo. In contrast, when the 13-mer LNA (locked nucleic acid-DNA gapmer antisense oligonucleotide (ASO was directly conjugated with α-tocopherol it showed markedly reduced silencing activity in mouse liver. Here, therefore, we tried to extend the 5′-end of the ASO sequence by using 5′-α-tocopherol–conjugated 4- to 7-mers of unlocked nucleic acid (UNA as a “second wing.” Intravenous injection of mice with this α-tocopherol–conjugated chimeric ASO achieved more potent silencing than ASO alone in the liver, suggesting increased delivery of the ASO to the liver. Within the cells, the UNA wing was cleaved or degraded and α-tocopherol was released from the 13-mer gapmer ASO, resulting in activation of the gapmer. The α-tocopherol–conjugated chimeric ASO showed high efficacy, with hepatic tropism, and was effective and safe for gene silencing in vivo. We have thus identified a new, effective LNA-DNA gapmer structure in which drug delivery system (DDS molecules are bound to ASO with UNA sequences.

  14. Alternative interpretations of the oligonucleotide transport literature: insights from nature.

    Science.gov (United States)

    Wu-Pong, S

    2000-10-31

    Elucidation of the mechanism of oligonucleotide (ON) cellular internalization has met an impasse at the lipid penetration stage. ON internalization is commonly regarded to involve endocytosis, yet the method by which the ON penetrates the endosome membrane remains a mystery despite more than 10 years of research by multiple laboratories. In addition, the literature regarding this topic is fraught with discrepancies and inconsistencies. Therefore, the goal of this review is to propose and illustrate the feasibility of the notion that the literature discrepancies are perhaps an indication of a complex transport mechanism involving more than one uptake pathway. Accordingly, ON- and cell-differences in uptake may be attributed to differences in the relative importance of these pathways for different cell types and ONs. An example of one such pathway is reviewed and critiqued in this communication with respect to its hypothetical role in ON uptake. Other innovative mechanisms should similarly be considered to stimulate new ideas, discussion and research in this unique and interesting field.

  15. Selective Covalent Conjugation of Phosphorothioate DNA Oligonucleotides with Streptavidin

    Directory of Open Access Journals (Sweden)

    Christof M. Niemeyer

    2011-08-01

    Full Text Available Protein-DNA conjugates have found numerous applications in the field of diagnostics and nanobiotechnology, however, their intrinsic susceptibility to DNA degradation by nucleases represents a major obstacle for many applications. We here report the selective covalent conjugation of the protein streptavidin (STV with phosphorothioate oligonucleotides (psDNA containing a terminal alkylthiolgroup as the chemically addressable linking unit, using a heterobifunctional NHS-/maleimide crosslinker. The psDNA-STV conjugates were synthesized in about 10% isolated yields. We demonstrate that the terminal alkylthiol group selectively reacts with the maleimide while the backbone sulfur atoms are not engaged in chemical conjugation. The novel psDNA-STV conjugates retain their binding capabilities for both biotinylated macromolecules and the complementary nucleic acid. Moreover, the psDNA-STV conjugate retained its binding capacity for complementary oligomers even after a nuclease digestion step, which effectively degrades deoxyribonucleotide oligomers and thus the binding capability of regular DNA-STV conjugates. The psDNA-STV therefore hold particular promise for applications e.g. in proteome research and novel biosensing devices, where interfering endogenous nucleic acids need to be removed from analytes by nuclease digestion.

  16. EvoOligo: oligonucleotide probe design with multiobjective evolutionary algorithms.

    Science.gov (United States)

    Shin, Soo-Yong; Lee, In-Hee; Cho, Young-Min; Yang, Kyung-Ae; Zhang, Byoung-Tak

    2009-12-01

    Probe design is one of the most important tasks in successful deoxyribonucleic acid microarray experiments. We propose a multiobjective evolutionary optimization method for oligonucleotide probe design based on the multiobjective nature of the probe design problem. The proposed multiobjective evolutionary approach has several distinguished features, compared with previous methods. First, the evolutionary approach can find better probe sets than existing simple filtering methods with fixed threshold values. Second, the multiobjective approach can easily incorporate the user's custom criteria or change the existing criteria. Third, our approach tries to optimize the combination of probes for the given set of genes, in contrast to other tools that independently search each gene for qualifying probes. Lastly, the multiobjective optimization method provides various sets of probe combinations, among which the user can choose, depending on the target application. The proposed method is implemented as a platform called EvoOligo and is available for service on the web. We test the performance of EvoOligo by designing probe sets for 19 types of Human Papillomavirus and 52 genes in the Arabidopsis Calmodulin multigene family. The design results from EvoOligo are proven to be superior to those from well-known existing probe design tools, such as OligoArray and OligoWiz.

  17. Stable oligonucleotide-functionalized gold nanosensors for environmental biocontaminant monitoring.

    Science.gov (United States)

    Riquelme, Maria V; Leng, Weinan; Carzolio, Marcos; Pruden, Amy; Vikesland, Peter

    2017-12-01

    The global propagation of environmental biocontaminants such as antibiotic resistant pathogens and their antibiotic resistance genes (ARGs) is a public health concern that highlights the need for improved monitoring strategies. Here, we demonstrate the environmental stability and applicability of an oligonucleotide-functionalized gold nanosensor. The mecA ARG was targeted as model biocontaminant due to its presence in clinically-relevant pathogens and to its emergence as an environmental contaminant. mecA-specific nanosensors were tested for antibiotic resistance gene (ARG) detection in ARG-spiked effluent from four wastewater treatment plants (WWTPs). The mecA-specific nanosensors showed stability in environmental conditions and in high ionic strength ([MgCl2]<50mM), and high selectivity against mismatched targets. Spectrophotometric detection was reproducible with an LOD of 70pM (≈4×107genes/μL), even in the presence of interferences associated with non-target genomic DNA and complex WWTP effluent. This contribution supports the environmental applicability of a new line of cost-effective, field-deployable tools needed for wide-scale biocontaminant monitoring. Copyright © 2017. Published by Elsevier B.V.

  18. Oligonucleotide fishing for STAT6: cross-talk between IL-4 and chemokines

    DEFF Research Database (Denmark)

    Eriksen, K W; Nielsen, M; Kaltoft, K

    2001-01-01

    -stranded oligonucleotide probes containing a STAT6-binding gene-sequence from the promotor of the immunoglobulin heavy chain germline epsilon transcript to study the IL-4-induced DNA binding of STAT6. Using these probes, we show that repeated adjacent STAT6-binding sites result in enhanced STAT6-DNA binding. Moreover...... activation, whereas other chemokines and cytokines do not. In conclusion, our data show that oligonucleotide fishing is a supplementary tool for studying cytokine cross-talk at a genomic level....

  19. Complexes of carbon nanotubes with oligonucleotides in thin Langmuir-Blodgett films to detect electrochemically hybridization

    Science.gov (United States)

    Egorov, A. S.; Egorova, V. P.; Krylova, H. V.; Lipnevich, I. V.; Orekhovskaya, T. I.; Veligura, A. A.; Govorov, M. I.; Shulitsky, B. G.

    2014-10-01

    Self-assembled complexes consisting of thin multi-walled carbon nanotubes (MWCNTs) and DNA-oligonucleotides which are able to a cooperative binding to complementary oligonucleotides have been investigated. It was establised a high-performance charge transport in nanostructured Langmuir-Blodgett complexes thin MWCNTs/DNA. A method to electrochemically detect DNA hybridization on the self-organized structures has been proposed.

  20. Synthesis and Excellent Duplex Stability of Oligonucleotides Containing 2'-Amino-LNA Functionalized with Galactose Units

    DEFF Research Database (Denmark)

    Kumar, Rajesh; Ries, Annika; Wengel, Jesper

    2017-01-01

    A convenient method for the preparation of oligonucleotides containing internally-attached galactose and triantennary galactose units has been developed based on click chemistry between 2'-N-alkyne 2'-amino-LNA nucleosides and azido-functionalized galactosyl building blocks. The synthesized oligo...... oligonucleotides show excellent binding affinity and selectivity towards complementary DNA/RNA strands with an increase in the melting temperature of up to +23.5 °C for triply-modified variants....

  1. Fragment-based solid-phase assembly of oligonucleotide conjugates with peptide and polyethylene glycol ligands.

    Science.gov (United States)

    Dirin, Mehrdad; Urban, Ernst; Noe, Christian R; Winkler, Johannes

    2016-10-04

    Ligand conjugation to oligonucleotides is an attractive strategy for enhancing the therapeutic potential of antisense and siRNA agents by inferring properties such as improved cellular uptake or better pharmacokinetic properties. Disulfide linkages enable dissociation of ligands and oligonucleotides in reducing environments found in endosomal compartments after cellular uptake. Solution-phase fragment coupling procedures for producing oligonucleotide conjugates are often tedious, produce moderate yields and reaction byproducts are frequently difficult to remove. We have developed an improved method for solid-phase coupling of ligands to oligonucleotides via disulfides directly after solid-phase synthesis. A 2'-thiol introduced using a modified nucleotide building block was orthogonally deprotected on the controlled pore glass solid support with N-butylphosphine. Oligolysine peptides and a short monodisperse ethylene glycol chain were successfully coupled to the deprotected thiol. Cleavage from the resin and full removal of oligonucleotide protection groups were achieved using methanolic ammonia. After standard desalting, and without further purification, homogenous conjugates were obtained as demonstrated by HPLC, gel electrophoresis, and mass spectrometry. The attachment of both amphiphilic and cationic ligands proves the versatility of the conjugation procedure. An antisense oligonucleotide conjugate with hexalysine showed pronounced gene silencing in a cell culture tumor model in the absence of a transfection reagent and the corresponding ethylene glycol conjugate resulted in down regulation of the target gene to nearly 50% after naked application. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  2. Sensitive and specific detection of Xanthomonas campestris pv. pelargonii with DNA primers and probes identified by random amplified polymorphic DNA analysis.

    Science.gov (United States)

    Manulis, S; Valinsky, L; Lichter, A; Gabriel, D W

    1994-11-01

    The random amplified polymorphic DNA method was used to distinguish strains of Xanthomonas campestris pv. pelargonii from 21 other Xanthomonas species and/or pathovars. Among the 42 arbitrarily chosen primers evaluated, 3 were found to reveal diagnostic polymorphisms when purified DNAs from compared strains were amplified by the PCR. The three primers revealed DNA amplification patterns which were conserved among all 53 strains tested of X. campestris pv. pelargonii isolated from various locations worldwide. The distinctive X. compestris pv. pelargonii patterns were clearly different from those obtained with any of 46 other Xanthomonas strains tested. An amplified 1.2-kb DNA fragment, apparently unique to X. campestris pv. pelargonii by these random amplified polymorphic DNA tests, was cloned and evaluated as a diagnostic DNA probe. It hybridized with total DNA from all 53 X. campestris pv. pelargonii strains tested and not with any of the 46 other Xanthomonas strains tested. The DNA sequence of the terminal ends of this 1.2-kb fragment was obtained and used to design a pair of 18-mer oligonucleotide primers specific for X. campestris pv. pelargonii. The custom-synthesized primers amplified the same 1.2-kb DNA fragment from all 53 X. campestris pv. pelargonii strains tested and failed to amplify DNA from any of the 46 other Xanthomonas strains tested. DNA isolated from saprophytes associated with the geranium plant also did not produce amplified DNA with these primers. The sensitivity of the PCR assay using the custom-synthesized primers was between 10 and 50 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Universal COI primers for DNA barcoding amphibians.

    Science.gov (United States)

    Che, Jing; Chen, Hong-Man; Yang, Jun-Xiao; Jin, Jie-Qiong; Jiang, Ke; Yuan, Zhi-Yong; Murphy, Robert W; Zhang, Ya-Ping

    2012-03-01

    DNA barcoding is a proven tool for the rapid and unambiguous identification of species, which is essential for many activities including the vouchering tissue samples in the genome 10K initiative, genealogical reconstructions, forensics and biodiversity surveys, among many other applications. A large-scale effort is underway to barcode all amphibian species using the universally sequenced DNA region, a partial fragment of mitochondrial cytochrome oxidase subunit I COI. This fragment is desirable because it appears to be superior to 16S for barcoding, at least for some groups of salamanders. The barcoding of amphibians is essential in part because many species are now endangered. Unfortunately, existing primers for COI often fail to achieve this goal. Herein, we report two new pairs of primers (➀, ➁) that in combination serve to universally amplify and sequence all three orders of Chinese amphibians as represented by 36 genera. This taxonomic diversity, which includes caecilians, salamanders and frogs, suggests that the new primer pairs will universally amplify COI for the vast majority species of amphibians. © 2011 Blackwell Publishing Ltd.

  4. MPprimer: a program for reliable multiplex PCR primer design

    Directory of Open Access Journals (Sweden)

    Wang Xiaolei

    2010-03-01

    Full Text Available Abstract Background Multiplex PCR, defined as the simultaneous amplification of multiple regions of a DNA template or multiple DNA templates using more than one primer set (comprising a forward primer and a reverse primer in one tube, has been widely used in diagnostic applications of clinical and environmental microbiology studies. However, primer design for multiplex PCR is still a challenging problem and several factors need to be considered. These problems include mis-priming due to nonspecific binding to non-target DNA templates, primer dimerization, and the inability to separate and purify DNA amplicons with similar electrophoretic mobility. Results A program named MPprimer was developed to help users for reliable multiplex PCR primer design. It employs the widely used primer design program Primer3 and the primer specificity evaluation program MFEprimer to design and evaluate the candidate primers based on genomic or transcript DNA database, followed by careful examination to avoid primer dimerization. The graph-expanding algorithm derived from the greedy algorithm was used to determine the optimal primer set combinations (PSCs for multiplex PCR assay. In addition, MPprimer provides a virtual electrophotogram to help users choose the best PSC. The experimental validation from 2× to 5× plex PCR demonstrates the reliability of MPprimer. As another example, MPprimer is able to design the multiplex PCR primers for DMD (dystrophin gene which caused Duchenne Muscular Dystrophy, which has 79 exons, for 20×, 20×, 20×, 14×, and 5× plex PCR reactions in five tubes to detect underlying exon deletions. Conclusions MPprimer is a valuable tool for designing specific, non-dimerizing primer set combinations with constrained amplicons size for multiplex PCR assays.

  5. Hierarchical organisation of causal graphs

    Energy Technology Data Exchange (ETDEWEB)

    Dziopa, P. [CEA Centre d`Etudes de la Vallee du Rhone, 30 - Marcoule (France). Dept. des Procedes de Retraitement; Gentil, S. [ENSIEG, 38 - Saint-Martin d`Heres (France)

    1993-12-31

    This paper deals with the design of a supervision system using a hierarchy of models formed by graphs, in which the variables are the nodes and the causal relations between the variables of the arcs. To obtain a representation of the variables evolutions which contains only the relevant features of their real evolutions, the causal relations are completed with qualitative transfer functions (QTFs) which produce roughly the behaviour of the classical transfer functions. Major improvements have been made in the building of the hierarchical organization. First, the basic variables of the uppermost level and the causal relations between them are chosen. The next graph is built by adding intermediary variables to the upper graph. When the undermost graph has been built, the transfer functions parameters corresponding to its causal relations are identified. The second task consists in the upwelling of the information from the undermost graph to the uppermost one. A fusion procedure of the causal relations has been designed to compute the QFTs relevant for each level. This procedure aims to reduce the number of parameters needed to represent an evolution at a high level of abstraction. These techniques have been applied to the hierarchical modelling of nuclear process. (authors). 8 refs., 12 figs.

  6. Hierarchical Structures in Hypertext Learning Environments

    NARCIS (Netherlands)

    Bezdan, Eniko; Kester, Liesbeth; Kirschner, Paul A.

    2011-01-01

    Bezdan, E., Kester, L., & Kirschner, P. A. (2011, 9 September). Hierarchical Structures in Hypertext Learning Environments. Presentation for the visit of KU Leuven, Open University, Heerlen, The Netherlands.

  7. Discovering hierarchical structure in normal relational data

    DEFF Research Database (Denmark)

    Schmidt, Mikkel Nørgaard; Herlau, Tue; Mørup, Morten

    2014-01-01

    Hierarchical clustering is a widely used tool for structuring and visualizing complex data using similarity. Traditionally, hierarchical clustering is based on local heuristics that do not explicitly provide assessment of the statistical saliency of the extracted hierarchy. We propose a non......-parametric generative model for hierarchical clustering of similarity based on multifurcating Gibbs fragmentation trees. This allows us to infer and display the posterior distribution of hierarchical structures that comply with the data. We demonstrate the utility of our method on synthetic data and data of functional...

  8. Hierarchical Robust and Adaptive Nonlinear Control Design

    National Research Council Canada - National Science Library

    Haddad, Wassim

    2003-01-01

    The authors proposed the development of a general multiechelon hierarchical nonlinear switching control design framework that minimizes control law complexity subject to the achievement of control law robustness...

  9. Analyses of point mutation repair and allelic heterogeneity generated by CRISPR/Cas9 and single-stranded DNA oligonucleotides

    OpenAIRE

    Bialk, Pawel; Sansbury, Brett; Rivera-Torres, Natalia; Bloh, Kevin; Man, Dula; Kmiec, Eric B.

    2016-01-01

    The repair of a point mutation can be facilitated by combined activity of a single-stranded oligonucleotide and a CRISPR/Cas9 system. While the mechanism of action of combinatorial gene editing remains to be elucidated, the regulatory circuitry of nucleotide exchange executed by oligonucleotides alone has been largely defined. The presence of the appropriate CRISPR/Cas9 system leads to an enhancement in the frequency of gene editing directed by single-stranded DNA oligonucleotides. While CRIS...

  10. Primer Concilio Provincial del Nuevo Reino

    Directory of Open Access Journals (Sweden)

    Manuel Lucena Salmoral

    1963-01-01

    Full Text Available El acontecimiento más sobresaliente del patriarcado de don Fernando Arias de Ugarte, en el que hubo muchos notables, fue el Primer Concilio Provincial del Nuevo Reino de Granada, celebrado en el año 1623. Cumplió así una vieja aspiración de los arzobispos santafereños y la obligación impuesta en el Concilio de Trento, por lo que resulta incomprensible lo historiado por don José Antonio Plaza quien, al referirse a este hecho, dice lo siguiente...

  11. Signals and systems primer with Matlab

    CERN Document Server

    Poularikas, Alexander D

    2006-01-01

    Signals and Systems Primer with MATLAB® equally emphasizes the fundamentals of both analog and digital signals and systems. To ensure insight into the basic concepts and methods, the text presents a variety of examples that illustrate a wide range of applications, from microelectromechanical to worldwide communication systems. It also provides MATLAB functions and procedures for practice and verification of these concepts.Taking a pedagogical approach, the author builds a solid foundation in signal processing as well as analog and digital systems. The book first introduces orthogonal signals,

  12. Bayesian models a statistical primer for ecologists

    CERN Document Server

    Hobbs, N Thompson

    2015-01-01

    Bayesian modeling has become an indispensable tool for ecological research because it is uniquely suited to deal with complexity in a statistically coherent way. This textbook provides a comprehensive and accessible introduction to the latest Bayesian methods-in language ecologists can understand. Unlike other books on the subject, this one emphasizes the principles behind the computations, giving ecologists a big-picture understanding of how to implement this powerful statistical approach. Bayesian Models is an essential primer for non-statisticians. It begins with a definition of probabili

  13. Emulsion primers, their contribution to bonding

    OpenAIRE

    González Sburlati, Rubén Osmar; Sapei, José

    2014-01-01

    Asphalt irrigation in its various types, performs specific functions in the structure of a road during the construction phase or their service life. In particular, the so-called "Irrigation Primer " is used in underlying layers, in order to generate a transition surface with the new asphalt layer; thus the tack or prime coat will be placed on a surface to ensure a good bond with the overlying layer. For a long time Diluted Asphalts (Cut Back mediums) where used, but have been discontinued for...

  14. Identification of mosquito-borne flavivirus sequences using universal primers and reverse transcription/polymerase chain reaction.

    Science.gov (United States)

    Pierre, V; Drouet, M T; Deubel, V

    1994-01-01

    A reverse transcription/polymerase chain reaction (RT/PCR) protocol for the rapid detection and identification of flaviviruses was developed using a set of universal oligonucleotide primers. These primers correspond to sequences in the 3' non-coding region and in the NS5 gene which are highly conserved among the mosquito-borne flaviviruses. The sequences of the resulting amplified products were analysed for dengue 1, dengue 2, dengue 3, dengue 4, Japanese encephalitis, West Nile, yellow fever and Zika viruses, and compared with the published sequences of other flaviviruses. The 291-297 nucleotides corresponding to the C-terminus of NS5 gene showed 56 to 76% similarity, whereas the 3' non-coding region (190 to 421 nucleotides) showed only 20 to 36% similarity. Genetic classification of the Zika virus supported its traditional serological grouping. Recombinant plasmids containing the flavivirus sequences were used in a nucleic acid hybridization test to identify the RT/PCR products derived from viral RNA extracted from experimentally infected mosquitoes. The plasmids were dotted on a strip of nitrocellulose membrane and incubated with the RT/PCR product labelled with digoxigenin during the PCR step. This is a valuable method for the rapid and specific identification of mosquito-borne flaviviruses in biological specimens and for subsequent sequence analysis.

  15. Novel Efficient Cell-Penetrating, Peptide-Mediated Strategy for Enhancing Telomerase Inhibitor Oligonucleotides.

    Science.gov (United States)

    Muñoz-Alarcón, Andrés; Eriksson, Jonas; Langel, Ülo

    2015-12-01

    At present, there are several therapeutic approaches for targeting telomerase in tumors. One in particular, currently undergoing clinical trials, is based on synthetic lipid-modified oligonucleotide antagonists aimed at inhibiting the ribonucleoprotein subunit of human telomerase. However, while enabling efficient uptake, the lipid modifications reduce the potency of the therapeutic oligonucleotides compared to nonmodified oligonucleotides. Moreover, lipid modification may increase oligonucleotide accumulation in the liver causing undesirable hepatotoxicity. Noncovalent complexation strategies for cell-penetrating peptide (CPP)-mediated delivery present an option to circumvent the need for potency-reducing modifications, while allowing for a highly efficient uptake, and could significantly improve the efficiency of telomerase-targeting cancer therapeutics. Delivery of a nonlipidated locked nucleic acid/2'-O-methyl mixmer significantly inhibits the telomerase activity in treated HeLa cells. The inhibitory effect was further improved through addition of a CPP. Furthermore, calculated IC50-values for the oligonucleotide delivered by CPPs into HeLa cells are more than 20 times lower than telomerase inhibitor Imetelstat, currently undergoing clinical trials. These results emphasize the potential of CPP-mediated delivery of future pharmaceuticals and provide means by which to enhance an already promising therapeutic strategy for cancer treatment.

  16. Development, characterization and experimental validation of a cultivated sunflower (Helianthus annuus L.) gene expression oligonucleotide microarray.

    Science.gov (United States)

    Fernandez, Paula; Soria, Marcelo; Blesa, David; DiRienzo, Julio; Moschen, Sebastian; Rivarola, Maximo; Clavijo, Bernardo Jose; Gonzalez, Sergio; Peluffo, Lucila; Príncipi, Dario; Dosio, Guillermo; Aguirrezabal, Luis; García-García, Francisco; Conesa, Ana; Hopp, Esteban; Dopazo, Joaquín; Heinz, Ruth Amelia; Paniego, Norma

    2012-01-01

    Oligonucleotide-based microarrays with accurate gene coverage represent a key strategy for transcriptional studies in orphan species such as sunflower, H. annuus L., which lacks full genome sequences. The goal of this study was the development and functional annotation of a comprehensive sunflower unigene collection and the design and validation of a custom sunflower oligonucleotide-based microarray. A large scale EST (>130,000 ESTs) curation, assembly and sequence annotation was performed using Blast2GO (www.blast2go.de). The EST assembly comprises 41,013 putative transcripts (12,924 contigs and 28,089 singletons). The resulting Sunflower Unigen Resource (SUR version 1.0) was used to design an oligonucleotide-based Agilent microarray for cultivated sunflower. This microarray includes a total of 42,326 features: 1,417 Agilent controls, 74 control probes for sunflower replicated 10 times (740 controls) and 40,169 different non-control probes. Microarray performance was validated using a model experiment examining the induction of senescence by water deficit. Pre-processing and differential expression analysis of Agilent microarrays was performed using the Bioconductor limma package. The analyses based on p-values calculated by eBayes (psunflower unigene collection, and a custom, validated sunflower oligonucleotide-based microarray using Agilent technology. Both the curated unigene collection and the validated oligonucleotide microarray provide key resources for sunflower genome analysis, transcriptional studies, and molecular breeding for crop improvement.

  17. A novel multifunctional oligonucleotide microarray for Toxoplasma gondii

    Directory of Open Access Journals (Sweden)

    Chen Feng

    2010-10-01

    Full Text Available Abstract Background Microarrays are invaluable tools for genome interrogation, SNP detection, and expression analysis, among other applications. Such broad capabilities would be of value to many pathogen research communities, although the development and use of genome-scale microarrays is often a costly undertaking. Therefore, effective methods for reducing unnecessary probes while maintaining or expanding functionality would be relevant to many investigators. Results Taking advantage of available genome sequences and annotation for Toxoplasma gondii (a pathogenic parasite responsible for illness in immunocompromised individuals and Plasmodium falciparum (a related parasite responsible for severe human malaria, we designed a single oligonucleotide microarray capable of supporting a wide range of applications at relatively low cost, including genome-wide expression profiling for Toxoplasma, and single-nucleotide polymorphism (SNP-based genotyping of both T. gondii and P. falciparum. Expression profiling of the three clonotypic lineages dominating T. gondii populations in North America and Europe provides a first comprehensive view of the parasite transcriptome, revealing that ~49% of all annotated genes are expressed in parasite tachyzoites (the acutely lytic stage responsible for pathogenesis and 26% of genes are differentially expressed among strains. A novel design utilizing few probes provided high confidence genotyping, used here to resolve recombination points in the clonal progeny of sexual crosses. Recent sequencing of additional T. gondii isolates identifies >620 K new SNPs, including ~11 K that intersect with expression profiling probes, yielding additional markers for genotyping studies, and further validating the utility of a combined expression profiling/genotyping array design. Additional applications facilitating SNP and transcript discovery, alternative statistical methods for quantifying gene expression, etc. are also pursued at

  18. Efficient oligonucleotide probe selection for pan-genomic tiling arrays

    Directory of Open Access Journals (Sweden)

    Zhang Wei

    2009-09-01

    Full Text Available Abstract Background Array comparative genomic hybridization is a fast and cost-effective method for detecting, genotyping, and comparing the genomic sequence of unknown bacterial isolates. This method, as with all microarray applications, requires adequate coverage of probes targeting the regions of interest. An unbiased tiling of probes across the entire length of the genome is the most flexible design approach. However, such a whole-genome tiling requires that the genome sequence is known in advance. For the accurate analysis of uncharacterized bacteria, an array must query a fully representative set of sequences from the species' pan-genome. Prior microarrays have included only a single strain per array or the conserved sequences of gene families. These arrays omit potentially important genes and sequence variants from the pan-genome. Results This paper presents a new probe selection algorithm (PanArray that can tile multiple whole genomes using a minimal number of probes. Unlike arrays built on clustered gene families, PanArray uses an unbiased, probe-centric approach that does not rely on annotations, gene clustering, or multi-alignments. Instead, probes are evenly tiled across all sequences of the pan-genome at a consistent level of coverage. To minimize the required number of probes, probes conserved across multiple strains in the pan-genome are selected first, and additional probes are used only where necessary to span polymorphic regions of the genome. The viability of the algorithm is demonstrated by array designs for seven different bacterial pan-genomes and, in particular, the design of a 385,000 probe array that fully tiles the genomes of 20 different Listeria monocytogenes strains with overlapping probes at greater than twofold coverage. Conclusion PanArray is an oligonucleotide probe selection algorithm for tiling multiple genome sequences using a minimal number of probes. It is capable of fully tiling all genomes of a species on

  19. Adaptive Sampling in Hierarchical Simulation

    Energy Technology Data Exchange (ETDEWEB)

    Knap, J; Barton, N R; Hornung, R D; Arsenlis, A; Becker, R; Jefferson, D R

    2007-07-09

    We propose an adaptive sampling methodology for hierarchical multi-scale simulation. The method utilizes a moving kriging interpolation to significantly reduce the number of evaluations of finer-scale response functions to provide essential constitutive information to a coarser-scale simulation model. The underlying interpolation scheme is unstructured and adaptive to handle the transient nature of a simulation. To handle the dynamic construction and searching of a potentially large set of finer-scale response data, we employ a dynamic metric tree database. We study the performance of our adaptive sampling methodology for a two-level multi-scale model involving a coarse-scale finite element simulation and a finer-scale crystal plasticity based constitutive law.

  20. Multicollinearity in hierarchical linear models.

    Science.gov (United States)

    Yu, Han; Jiang, Shanhe; Land, Kenneth C

    2015-09-01

    This study investigates an ill-posed problem (multicollinearity) in Hierarchical Linear Models from both the data and the model perspectives. We propose an intuitive, effective approach to diagnosing the presence of multicollinearity and its remedies in this class of models. A simulation study demonstrates the impacts of multicollinearity on coefficient estimates, associated standard errors, and variance components at various levels of multicollinearity for finite sample sizes typical in social science studies. We further investigate the role multicollinearity plays at each level for estimation of coefficient parameters in terms of shrinkage. Based on these analyses, we recommend a top-down method for assessing multicollinearity in HLMs that first examines the contextual predictors (Level-2 in a two-level model) and then the individual predictors (Level-1) and uses the results for data collection, research problem redefinition, model re-specification, variable selection and estimation of a final model. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. The Case for a Hierarchical Cosmology

    Science.gov (United States)

    Vaucouleurs, G. de

    1970-01-01

    The development of modern theoretical cosmology is presented and some questionable assumptions of orthodox cosmology are pointed out. Suggests that recent observations indicate that hierarchical clustering is a basic factor in cosmology. The implications of hierarchical models of the universe are considered. Bibliography. (LC)

  2. Hierarchical decision making for flood risk reduction

    DEFF Research Database (Denmark)

    Custer, Rocco; Nishijima, Kazuyoshi

    2013-01-01

    River flood events often cause large economic damages and casualties requiring stakeholders to manage flood risk. In flood prone areas, flood risk management can be achieved through a series hierarchically integrated protection structures, which together form a hierarchical flood protection system...

  3. Biased trapping issue on weighted hierarchical networks

    Indian Academy of Sciences (India)

    In this paper, we present trapping issues of weight-dependent walks on weighted hierarchical networks which are based on the classic scale-free hierarchical networks. Assuming that edge's weight is used as local information by a random walker, we introduce a biased walk. The biased walk is that a walker, at each step, ...

  4. Aggregation in hierarchical discrete-event systems

    NARCIS (Netherlands)

    Al-Falou, AA; Van Schuppen, JH

    2003-01-01

    We propose an algorithm for the hierarchical decomposition of a large automaton-based discrete-event system. We also provide an estimation of the numerical cost in terms of the size of the system. Our hierarchical decomposition is illustrated with simple examples from dynamical ST-control and

  5. Primer on CDM programme of activities

    Energy Technology Data Exchange (ETDEWEB)

    Hinostroza, M. (UNEP Risoe Centre, Roskilde (Denmark)); Lescano, A.D. (A2G Carbon Partners (Peru)); Alvarez, J.M. (Ministerio del Ambiente del Peru (Peru)); Avendano, F.M. (EEA Fund Management Ltd. (United Kingdom)

    2009-07-01

    As an advanced modality introduced in 2005, the Programmatic CDM (POA) is expected to address asymmetries of participation, especially of very small-scale project activities in certain areas, key sectors and many countries with considerable potential for greenhouse gas emission reductions, not reached by the traditional single-project-based CDM. Latest experiences with POAs and the recently finalized official guidance governing the Programmatic CDM are the grassroots of this Primer, which has the purpose of supporting the fully understanding of rules and procedures of POAs by interpreting them and analyzing real POA cases. Professional and experts from the public and private entities have contributed to the development of this Primer, produced by the UNEP Risoe Centre, as part of knowledge support activities for the Capacity Development for the CDM (CD4CDM) project. The overall objective of the CD4CDM is to develop the capacities of host countries to identify, design, approve, finance, implement CDM projects and commercialize CERs in participating countries. The CDM4CDM is funded by the Netherlands Ministry of Foreign Affairs. (author)

  6. High Functional Stability of a Low-cost HBV DNA qPCR Primer Pair and Plasmid Standard

    Science.gov (United States)

    García, Gerardo; León, Yamila; Canales, Eduardo; Angel Silva, José; Gell, Omar; Estrada, Regla; Morán, Ivis; Muzio, Verena; Guillén, Gerardo; Pentón, Eduardo; Aguilar, Julio Cesar

    2016-01-01

    ABSTRACT Aim We studied the functional stability of a primer pair and the standard curve based on a plasmid carrying full-length HBV genome, from a novel low-cost real-time quantitative polymerase chain reaction (qPCR) assay. The assay was developed at the Center for Genetic Engineering and Biotechnology (CIGB) in Havana, to quantify the serum hepatitis B virus (HBV) DNA from chronic HBV-infected (CHB) patients. Materials and methods In-house generated oligonucleotides and plasmids were incubated at 37°C during 1 month and compared with the same materials incubated at –20, 4, and 25°C during the same time in qPCR experiments. Results This work shows that the oligonucleotide pair and the plasmid for the quantitative standard curve are functionally stable in severe temperature conditions during 1 month. Polymerase chain reaction amplification with both materials after its incubation 30 days at 37°C produced similar cycle threshold (CT) values and similar degree of sample quantifications compared with the same materials preserved using the conventional storage conditions at –20°C. Conclusion These results are indicative of the robustness of this low-cost qPCR system for HBV DNA quantification. These results also support that this qPCR assay can be used as a low-cost technology in clinical studies to monitor the viral load changes of serum HBV DNA of CHB patients, which could be used by poor people of third world countries, where there are frequent blackouts and temperature changes that can hinder the primer and plasmid stability. How to cite this article Aguiar J, García G, León Y, Canales E, Silva JA, Gell O, Estrada R, Morán I, Muzio V, Guillén G, Pentón E, Aguilar JC. High Functional Stability of a Low-cost HBV DNA qPCR Primer Pair and Plasmid Standard. Euroasian J Hepato-Gastroenterol 2016;6(1):19-24. PMID:29201719

  7. An iterative method for selecting degenerate multiplex PCR primers.

    Science.gov (United States)

    Souvenir, Richard; Buhler, Jeremy; Stormo, Gary; Zhang, Weixiong

    2007-01-01

    Single-nucleotide polymorphism (SNP) genotyping is an important molecular genetics process, which can produce results that will be useful in the medical field. Because of inherent complexities in DNA manipulation and analysis, many different methods have been proposed for a standard assay. One of the proposed techniques for performing SNP genotyping requires amplifying regions of DNA surrounding a large number of SNP loci. To automate a portion of this particular method, it is necessary to select a set of primers for the experiment. Selecting these primers can be formulated as the Multiple Degenerate Primer Design (MDPD) problem. The Multiple, Iterative Primer Selector (MIPS) is an iterative beam-search algorithm for MDPD. Theoretical and experimental analyses show that this algorithm performs well compared with the limits of degenerate primer design. Furthermore, MIPS outperforms an existing algorithm that was designed for a related degenerate primer selection problem.

  8. Screening of rapd primer for teak (Tectona grandis

    Directory of Open Access Journals (Sweden)

    Imas Cintamulya

    2016-04-01

    Full Text Available Identification of DNA polymorphisms in teak is important. It is a first step to determine the presence of genetic varia-tion in teak. The information of genetic variation is needed for teak breeding development. RAPD is one of method which can be used for identification of DNA polymorphism. This study aim to get the RAPD primer which can detect the DNA polymorphism in teak. Benefits of this study are provide information about primer which can detect the DNA polymorphism in teak, DNA polymorphism data can be used for genetic variation analysis which needed for teak breeding development. The primers which used in this study shown the DNA polymorphism in teak. The primer are OPF6 (5'-GGGAATTCGG-3 ', OPF8 (5'-GGGATATCGC-3', and OPF11 (5'-TTGGTACCCC-3 '. The highest DNA poly-morphism is shown in DNA which amplified with OPF-8 primer. Keywords: RAPD, Primer, Polymorphism, DNA, Tectona grandis

  9. Processing of hierarchical syntactic structure in music.

    Science.gov (United States)

    Koelsch, Stefan; Rohrmeier, Martin; Torrecuso, Renzo; Jentschke, Sebastian

    2013-09-17

    Hierarchical structure with nested nonlocal dependencies is a key feature of human language and can be identified theoretically in most pieces of tonal music. However, previous studies have argued against the perception of such structures in music. Here, we show processing of nonlocal dependencies in music. We presented chorales by J. S. Bach and modified versions in which the hierarchical structure was rendered irregular whereas the local structure was kept intact. Brain electric responses differed between regular and irregular hierarchical structures, in both musicians and nonmusicians. This finding indicates that, when listening to music, humans apply cognitive processes that are capable of dealing with long-distance dependencies resulting from hierarchically organized syntactic structures. Our results reveal that a brain mechanism fundamental for syntactic processing is engaged during the perception of music, indicating that processing of hierarchical structure with nested nonlocal dependencies is not just a key component of human language, but a multidomain capacity of human cognition.

  10. Hierarchically Nanostructured Materials for Sustainable Environmental Applications

    Directory of Open Access Journals (Sweden)

    Zheng eRen

    2013-11-01

    Full Text Available This article presents a comprehensive overview of the hierarchical nanostructured materials with either geometry or composition complexity in environmental applications. The hierarchical nanostructures offer advantages of high surface area, synergistic interactions and multiple functionalities towards water remediation, environmental gas sensing and monitoring as well as catalytic gas treatment. Recent advances in synthetic strategies for various hierarchical morphologies such as hollow spheres and urchin-shaped architectures have been reviewed. In addition to the chemical synthesis, the physical mechanisms associated with the materials design and device fabrication have been discussed for each specific application. The development and application of hierarchical complex perovskite oxide nanostructures have also been introduced in photocatalytic water remediation, gas sensing and catalytic converter. Hierarchical nanostructures will open up many possibilities for materials design and device fabrication in environmental chemistry and technology.

  11. Streptavidin-coated gold nanoparticles: critical role of oligonucleotides on stability and fractal aggregation

    Directory of Open Access Journals (Sweden)

    Roberta D'Agata

    2017-01-01

    Full Text Available Gold nanoparticles (AuNPs exhibit unique properties that can be modulated through a tailored surface functionalization, enabling their targeted use in biochemical sensing and medical diagnostics. In particular, streptavidin-modified AuNPs are increasingly used for biosensing purposes. We report here a study of AuNPs surface-functionalized with streptavidin-biotinylated oligonucleotide, focussing on the role played by the oligonucleotide probes in the stabilization/destabilization of the functionalized nanoparticle dispersion. The behaviour of the modified AuNP dispersion as a consequence of the competitive displacement of the biotinylated oligonucleotide has been investigated and the critical role of displaced oligonucletides in triggering the quasi one-dimensional aggregation of nanoparticles is demonstrated for the first time. The thorough understanding of the fundamental properties of bioconjugated AuNPs is of great importance for the design of highly sensitive and reliable functionalized AuNP-based assays.

  12. Toward a new paradigm of DNA writing using a massively parallel sequencing platform and degenerate oligonucleotide.

    Science.gov (United States)

    Hwang, Byungjin; Bang, Duhee

    2016-11-23

    All synthetic DNA materials require prior programming of the building blocks of the oligonucleotide sequences. The development of a programmable microarray platform provides cost-effective and time-efficient solutions in the field of data storage using DNA. However, the scalability of the synthesis is not on par with the accelerating sequencing capacity. Here, we report on a new paradigm of generating genetic material (writing) using a degenerate oligonucleotide and optomechanical retrieval method that leverages sequencing (reading) throughput to generate the desired number of oligonucleotides. As a proof of concept, we demonstrate the feasibility of our concept in digital information storage in DNA. In simulation, the ability to store data is expected to exponentially increase with increase in degenerate space. The present study highlights the major framework change in conventional DNA writing paradigm as a sequencer itself can become a potential source of making genetic materials.

  13. Ultramild protein-mediated click chemistry creates efficient oligonucleotide probes for targeting and detecting nucleic acids

    DEFF Research Database (Denmark)

    Nåbo, Lina J.; Madsen, Charlotte Stahl; Jensen, Knud Jørgen

    2015-01-01

    Functionalized synthetic oligonucleotides are finding growing applications in research, clinical studies, and therapy. However, it is not easy to prepare them in a biocompatible and highly efficient manner. We report a new strategy to synthesize oligonucleotides with promising nucleic acid...... conditions with fluorophore, peptide, and carbohydrate azide derivatives. In thermal denaturation studies, the modified probes showed specific binding to complementary DNA and RNA targets. Finally, we demonstrated the pH sensitivity of the new rhodamine-based fluorescent probes in vitro and rationalize our...... results by electronic structure calculations. Functionalized oligonucleotides were prepared in good yields by protein-mediated CuAAC click reactions for the first time with a human copper-binding chaperon. The carbohydrate, peptide, and fluorescent derivatives display high binding affinity and selectivity...

  14. Hydration-dependent dynamics of human telomeric oligonucleotides in the picosecond timescale: A neutron scattering study

    Science.gov (United States)

    Sebastiani, F.; Longo, M.; Orecchini, A.; Comez, L.; De Francesco, A.; Muthmann, M.; Teixeira, S. C. M.; Petrillo, C.; Sacchetti, F.; Paciaroni, A.

    2015-07-01

    The dynamics of the human oligonucleotide AG3(T2AG3)3 has been investigated by incoherent neutron scattering in the sub-nanosecond timescale. A hydration-dependent dynamical activation of thermal fluctuations in weakly hydrated samples was found, similar to that of protein powders. The amplitudes of such thermal fluctuations were evaluated in two different exchanged wave-vector ranges, so as to single out the different contributions from intra- and inter-nucleotide dynamics. The activation energy was calculated from the temperature-dependent characteristic times of the corresponding dynamical processes. The trends of both amplitudes and activation energies support a picture where oligonucleotides possess a larger conformational flexibility than long DNA sequences. This additional flexibility, which likely results from a significant relative chain-end contribution to the average chain dynamics, could be related to the strong structural polymorphism of the investigated oligonucleotides.

  15. Hydration-dependent dynamics of human telomeric oligonucleotides in the picosecond timescale: A neutron scattering study

    Energy Technology Data Exchange (ETDEWEB)

    Sebastiani, F.; Comez, L.; Sacchetti, F. [Dipartimento di Fisica e Geologia, Università degli Studi di Perugia, Via A. Pascoli, 06123 Perugia (Italy); CNR, Istituto Officina dei Materiali, Unità di Perugia, c/o Dipartimento di Fisica e Geologia, Università di Perugia, 06123 Perugia (Italy); Longo, M. [Dipartimento di Fisica e Geologia, Università degli Studi di Perugia, Via A. Pascoli, 06123 Perugia (Italy); Elettra—Sincrotrone Trieste, 34149 Basovizza, Trieste (Italy); Orecchini, A.; Petrillo, C.; Paciaroni, A., E-mail: alessandro.paciaroni@fisica.unipg.it [Dipartimento di Fisica e Geologia, Università degli Studi di Perugia, Via A. Pascoli, 06123 Perugia (Italy); De Francesco, A. [CNR-IOM OGG c/o Institut Laue-Langevin, 71 Avenue des Martyrs, CS20156, 38042 Grenoble Cedex 9 (France); Muthmann, M. [Jülich Centre for Neutron Science, Forschungszentrum Jülich GmbH, Outstation at Heinz Maier-Leibnitz Zentrum, Lichtenbergstrasse 1, 85747 Garching (Germany); Teixeira, S. C. M. [EPSAM, Keele University, Staffordshire ST5 5BG (United Kingdom); Institut Laue–Langevin, 71 Avenue des Martyrs, CS20156, 38042 Grenoble Cedex 9 (France)

    2015-07-07

    The dynamics of the human oligonucleotide AG{sub 3}(T{sub 2}AG{sub 3}){sub 3} has been investigated by incoherent neutron scattering in the sub-nanosecond timescale. A hydration-dependent dynamical activation of thermal fluctuations in weakly hydrated samples was found, similar to that of protein powders. The amplitudes of such thermal fluctuations were evaluated in two different exchanged wave-vector ranges, so as to single out the different contributions from intra- and inter-nucleotide dynamics. The activation energy was calculated from the temperature-dependent characteristic times of the corresponding dynamical processes. The trends of both amplitudes and activation energies support a picture where oligonucleotides possess a larger conformational flexibility than long DNA sequences. This additional flexibility, which likely results from a significant relative chain-end contribution to the average chain dynamics, could be related to the strong structural polymorphism of the investigated oligonucleotides.

  16. Specific PCR product primer design using memetic algorithm.

    Science.gov (United States)

    Yang, Cheng-Hong; Cheng, Yu-Huei; Chuang, Li-Yeh; Chang, Hsueh-Wei

    2009-01-01

    To provide feasible primer sets for performing a polymerase chain reaction (PCR) experiment, many primer design methods have been proposed. However, the majority of these methods require a relatively long time to obtain an optimal solution since large quantities of template DNA need to be analyzed. Furthermore, the designed primer sets usually do not provide a specific PCR product size. In recent years, evolutionary computation has been applied to PCR primer design and yielded promising results. In this article, a memetic algorithm (MA) is proposed to solve primer design problems associated with providing a specific product size for PCR experiments. The MA is compared with a genetic algorithm (GA) using an accuracy formula to estimate the quality of the primer design and test the running time. Overall, 50 accession nucleotide sequences were sampled for the comparison of the accuracy of the GA and MA for primer design. Five hundred runs of the GA and MA primer design were performed with PCR product lengths of 150-300 bps and 500-800 bps, and two different methods of calculating T(m) for each accession nucleotide sequence were tested. A comparison of the accuracy results for the GA and MA primer design showed that the MA primer design yielded better results than the GA primer design. The results further indicate that the proposed method finds optimal or near-optimal primer sets and effective PCR products in a dry dock experiment. Related materials are available online at http://bio.kuas.edu.tw/ma-pd/. 2009 American Institute of Chemical Engineers

  17. Sequence analysis of peptide:oligonucleotide heteroconjugates by electron capture dissociation and electron transfer dissociation.

    Science.gov (United States)

    Krivos, Kady L; Limbach, Patrick A

    2010-08-01

    Mass spectrometry analysis of protein-nucleic acid cross-links is challenging due to the dramatically different chemical properties of the two components. Identifying specific sites of attachment between proteins and nucleic acids requires methods that enable sequencing of both the peptide and oligonucleotide component of the heteroconjugate cross-link. While collision-induced dissociation (CID) has previously been used for sequencing such heteroconjugates, CID generates fragmentation along the phosphodiester backbone of the oligonucleotide preferentially. The result is a reduction in peptide fragmentation within the heteroconjugate. In this work, we have examined the effectiveness of electron capture dissociation (ECD) and electron-transfer dissociation (ETD) for sequencing heteroconjugates. Both methods were found to yield preferential fragmentation of the peptide component of a peptide:oligonucleotide heteroconjugate, with minimal differences in sequence coverage between these two electron-induced dissociation methods. Sequence coverage was found to increase with increasing charge state of the heteroconjugate, but decreases with increasing size of the oligonucleotide component. To overcome potential intermolecular interactions between the two components of the heteroconjugate, supplemental activation with ETD was explored. The addition of a supplemental activation step was found to increase peptide sequence coverage over ETD alone, suggesting that electrostatic interactions between the peptide and oligonucleotide components are one limiting factor in sequence coverage by these two approaches. These results show that ECD/ETD methods can be used for the tandem mass spectrometry sequencing of peptide:oligonucleotide heteroconjugates, and these methods are complementary to existing CID methods already used for sequencing of protein-nucleic acid cross-links. Copyright 2010. Published by Elsevier Inc.

  18. Characteristics of Primer of High Performance Bismaleimide Resin

    Directory of Open Access Journals (Sweden)

    LI Hong-feng

    2016-06-01

    Full Text Available A primer of the bismaleimide resin toughened with block copolymer was prepared. The properties of the primer, such as adhesion, pencil hardness, impact resistance, environment resistant, ageing resistant performance and etc. meet the requirements of the technical specifications of GJB1388. The thermal property of the primer was characterized by differential scanning calorimeter (DSC, thermal gravity analysis (TG, and dynamic mechanical analysis (DMA. The results show that the primer possesses good heat resistance, the glass transition temperature of the primer reach 238℃, 5% mass heat lost temperature is 384℃ after 200℃ cured, and the glass transition temperature of the primer reach 268℃, 5% mass heat lost temperature is 407℃ after 230℃ heat treatment respectively. Bonding strength will increase when the bismaleimide primer doubled with bismaleimide film adhesive, when the primer doubled with J-188 bismaleimide film adhesive, peel strength increases to 107%, the room temperature shear strength and high temperature shear strength increase by 10%.The primer can also be used with other bismaleimide film structural adhesives, which are suitable for the bonding between metals or bismaleimide composites with metals.

  19. Hierarchically Nanoporous Bioactive Glasses for High Efficiency Immobilization of Enzymes

    DEFF Research Database (Denmark)

    He, W.; Min, D.D.; Zhang, X.D.

    2014-01-01

    Bioactive glasses with hierarchical nanoporosity and structures have been heavily involved in immobilization of enzymes. Because of meticulous design and ingenious hierarchical nanostructuration of porosities from yeast cell biotemplates, hierarchically nanostructured porous bioactive glasses can...

  20. Direct Mutagenesis of Thousands of Genomic Targets using Microarray-derived Oligonucleotides

    DEFF Research Database (Denmark)

    Bonde, Mads; Kosuri, Sriram; Genee, Hans Jasper

    2015-01-01

    Multiplex Automated Genome Engineering (MAGE) allows simultaneous mutagenesis of multiple target sites in bacterial genomes using short oligonucleotides. However, large-scale mutagenesis requires hundreds to thousands of unique oligos, which are costly to synthesize and impossible to scale-up by ...... insertions per cell. MO-MAGE enables cost-effective large-scale targeted genome engineering that should be useful for a variety of applications in synthetic biology and metabolic engineering.......Multiplex Automated Genome Engineering (MAGE) allows simultaneous mutagenesis of multiple target sites in bacterial genomes using short oligonucleotides. However, large-scale mutagenesis requires hundreds to thousands of unique oligos, which are costly to synthesize and impossible to scale...

  1. Synthesis and properties of cationic 2'-O-[N-(4-aminobutyl)carbamoyl] modified oligonucleotides.

    Science.gov (United States)

    Seio, Kohji; Tokugawa, Munefumi; Kanamori, Takashi; Tsunoda, Hirosuke; Ohkubo, Akihiro; Sekine, Mitsuo

    2012-04-01

    2'-O-[N-(4-Aminobutylcarbamoyl)]uridine (U(abcm)) was synthesized and incorporated into oligonucleotides. The oligonucleotides incorporating U(abcm) formed more stable duplexes with their complementary and mismatched RNAs than those containing 2'-O-carbamoyluridine (U(cm)). The stability of duplex with a U(abcm)-rG base pair showed higher thermostability than the duplex having unmodified U-rG base pair. The U(abcm) residue showed enhanced resistance to snake venome phosphodiesterase. Copyright © 2012 Elsevier Ltd. All rights reserved.

  2. A hierarchical artificial retina architecture

    Science.gov (United States)

    Parker, Alice C.; Azar, Adi N.

    2009-05-01

    Connectivity in the human retina is complex. Over one hundred million photoreceptors transduce light into electrical signals. These electrical signals are sent to the ganglion cells through amacrine and bipolar cells. Lateral connections involving horizontal and amacrine cells span throughout the outer plexiform layer and inner plexiform layer respectively. Horizontal cells are important for photoreceptor regulation by depolarizing them after an illumination occurs. Horizontal cells themselves form an electrical network that communicates by gap junctions, and these cells exhibit plasticity (change in behavior and structure) with respect to glycine receptors. The bipolar and amacrine cells transfer electrical signals from photoreceptors to the ganglion cells. Furthermore, amacrine cells are responsible for further processing the retinal image. Finally, the ganglion cells receive electrical signals from the bipolar and amacrine cells and will spike at a faster rate if there is a change in the overall intensity for a group of photoreceptors, sending a signal to the brain. Dramatic progress is being made with respect to retinal prostheses, raising hope for an entire synthetic retina in the future. We propose a bio-inspired 3D hierarchical pyramidal architecture for a synthetic retina that mimics the overall structure of the human retina. We chose to use a 3D architecture to facilitate connectivity among retinal cells, maintaining a hierarchical structure similar to that of the biological retina. The first layer of the architecture contains electronic circuits that model photoreceptors and horizontal cells. The second layer contains amacrine and bipolar electronic cells, and the third layer contains ganglion cells. Layer I has the highest number of cells, and layer III has the lowest number of cells, resulting in a pyramidal architecture. In our proposed architecture we intend to use photodetectors to transduce light into electrical signals. We propose to employ

  3. The thermal stability of oligonucleotide duplexes is sequence independent in tetraalkylammonium salt solutions: application to identifying recombinant DNA clones.

    OpenAIRE

    Jacobs, K. A.; Rudersdorf, R; Neill, S D; Dougherty, J P; Brown, E L; Fritsch, E F

    1988-01-01

    In solutions of tetraalkylammonium salts the melting temperature of oligonucleotide duplexes is independent of nucleotide sequence and thus GC content. Data quantitating the destabilizing effects of various mismatches in these solvents are also presented. The results are in accord with theories on DNA melting and establish conditions under which oligonucleotides can be used as hybridization probes with predictable and controllable specificity.

  4. Synthesis and Biophysical Investigations of Oligonucleotides Containing Galactose-Modified DNA, LNA and 2'-Amino-LNA Monomers

    DEFF Research Database (Denmark)

    Ries, Annika; Kumar, Rajesh; Lou, Chenguang

    2016-01-01

    Galactose-modified thymidine, LNA-T and 2'-amino-LNA-T nucleosides were synthesized, converted into the corresponding phosphoramidite derivatives and introduced into short oligonucleotides. Compared to the unmodified control strands, the galactose-modified oligonucleotides in general, and the N2'...

  5. Antisense oligonucleotide inhibition of Heat Shock Protein (HSP 47 improves bleomycin-induced pulmonary fibrosis in rats

    Directory of Open Access Journals (Sweden)

    Noguchi Takayuki

    2007-05-01

    Full Text Available Abstract Background The most common pathologic form of pulmonary fibrosis arises from excessive deposition of extracellular matrix proteins such as collagen. The 47 kDa heat shock protein 47 (HSP47 is a collagen-specific molecular chaperone that has been shown to play a major role during the processing and/or secretion of procollagen. Objectives To determine whether inhibition of HSP47 could have beneficial effects in mitigating bleomycin-induced pulmonary fibrosis in rats. Methods All experiments were performed with 250–300 g male Wistar rats. Animals were randomly divided into five experimental groups that were administered: 1 saline alone, 2 bleomycin alone, 3 antisense HSP47 oligonucleotides alone, 4 bleomycin + antisense HSP47 oligonucleotides, and 5 bleomycin + sense control oligonucleotides. We investigated lung histopathology and performed immunoblot and immunohistochemistry analyses. Results In rats treated with HSP47 antisense oligonucleotides, pulmonary fibrosis was significantly reduced. In addition, treatment with HSP47 antisense oligonucleotides significantly improved bleomycin-induced morphological changes. Treatment with HSP47 antisense oligonucleotides alone did not produce any significant changes to lung morphology. Immunoblot analyses of lung homogenates confirmed the inhibition of HSP47 protein by antisense oligonucleotides. The bleo + sense group, however, did not exhibit any improvement in lung pathology compared to bleomycin alone groups, and also had no effect on HSP47 expression. Conclusion These findings suggest that HSP47 antisense oligonucleotide inhibition of HSP47 improves bleomycin-induced pulmonary fibrosis pathology in rats.

  6. Complex and adaptive dynamical systems a primer

    CERN Document Server

    Gros, Claudius

    2015-01-01

    This primer offers readers an introduction to the central concepts that form our modern understanding of complex and emergent behavior, together with detailed coverage of accompanying mathematical methods. All calculations are presented step by step and are easy to follow. This new fourth edition has been fully reorganized and includes new chapters, figures and exercises. The core aspects of modern complex system sciences are presented in the first chapters, covering network theory, dynamical systems, bifurcation and catastrophe theory, chaos and adaptive processes, together with the principle of self-organization in reaction-diffusion systems and social animals. Modern information theoretical principles are treated in further chapters, together with the concept of self-organized criticality, gene regulation networks, hypercycles and coevolutionary avalanches, synchronization phenomena, absorbing phase transitions and the cognitive system approach to the brain. Technical course prerequisites are the standard ...

  7. Complex and adaptive dynamical systems a primer

    CERN Document Server

    Gros, Claudius

    2007-01-01

    We are living in an ever more complex world, an epoch where human actions can accordingly acquire far-reaching potentialities. Complex and adaptive dynamical systems are ubiquitous in the world surrounding us and require us to adapt to new realities and the way of dealing with them. This primer has been developed with the aim of conveying a wide range of "commons-sense" knowledge in the field of quantitative complex system science at an introductory level, providing an entry point to this both fascinating and vitally important subject. The approach is modular and phenomenology driven. Examples of emerging phenomena of generic importance treated in this book are: -- The small world phenomenon in social and scale-free networks. -- Phase transitions and self-organized criticality in adaptive systems. -- Life at the edge of chaos and coevolutionary avalanches resulting from the unfolding of all living. -- The concept of living dynamical systems and emotional diffusive control within cognitive system theory. Techn...

  8. Complex and Adaptive Dynamical Systems A Primer

    CERN Document Server

    Gros, Claudius

    2011-01-01

    We are living in an ever more complex world, an epoch where human actions can accordingly acquire far-reaching potentialities. Complex and adaptive dynamical systems are ubiquitous in the world surrounding us and require us to adapt to new realities and the way of dealing with them. This primer has been developed with the aim of conveying a wide range of "commons-sense" knowledge in the field of quantitative complex system science at an introductory level, providing an entry point to this both fascinating and vitally important subject. The approach is modular and phenomenology driven. Examples of emerging phenomena of generic importance treated in this book are: -- The small world phenomenon in social and scale-free networks. -- Phase transitions and self-organized criticality in adaptive systems. -- Life at the edge of chaos and coevolutionary avalanches resulting from the unfolding of all living. -- The concept of living dynamical systems and emotional diffusive control within cognitive system theory. Techn...

  9. Complex and adaptive dynamical systems a primer

    CERN Document Server

    Gros, Claudius

    2013-01-01

    Complex system theory is rapidly developing and gaining importance, providing tools and concepts central to our modern understanding of emergent phenomena. This primer offers an introduction to this area together with detailed coverage of the mathematics involved. All calculations are presented step by step and are straightforward to follow. This new third edition comes with new material, figures and exercises. Network theory, dynamical systems and information theory, the core of modern complex system sciences, are developed in the first three chapters, covering basic concepts and phenomena like small-world networks, bifurcation theory and information entropy. Further chapters use a modular approach to address the most important concepts in complex system sciences, with the emergence and self-organization playing a central role. Prominent examples are self-organized criticality in adaptive systems, life at the edge of chaos, hypercycles and coevolutionary avalanches, synchronization phenomena, absorbing phase...

  10. Scope and Limitations of the Nicking Enzyme Amplification Reaction for the Synthesis of Base-Modified Oligonucleotides and Primers for PCR

    Czech Academy of Sciences Publication Activity Database

    Ménová, Petra; Raindlová, Veronika; Hocek, Michal

    2013-01-01

    Roč. 24, č. 6 (2013), s. 1081-1093 ISSN 1043-1802 R&D Projects: GA ČR GA203/09/0317 Institutional support: RVO:61388963 Keywords : isothermal DNA amplification * cross-coupling reactions * nucleoside triphosphates * polymerase incorporation * functionalized DNA * nucleic- acids Subject RIV: CC - Organic Chemistry Impact factor: 4.821, year: 2013

  11. Comparative analysis of oligonucleotide primers for high-throughput screening of genes encoding adenylation domains of nonribosomal peptide synthetases in actinomycetes

    Czech Academy of Sciences Publication Activity Database

    Vopálenská, I.; Váchová, Libuše; Palková, Z.

    2015-01-01

    Roč. 72, OCT 2015 (2015), s. 160-167 ISSN 0956-5663 R&D Projects: GA TA ČR(CZ) TA01011461; GA MŠk(CZ) ED1.1.00/02.0109; GA MŠk(CZ) EE2.3.30.0003 Institutional support: RVO:61388971 Keywords : Yeast biosensor * Copper ion detection * Purine synthesis pathway Subject RIV: DJ - Water Pollution ; Quality Impact factor: 7.476, year: 2015

  12. Formaldehyde as hypothetical primer of biohomochirality

    Energy Technology Data Exchange (ETDEWEB)

    Goldanskii, V.I. [N. N. Semenov Institute of Chemical Physics of the Russian Academy of Sciences, Kosygin Street 4, Moscow, 117334 (Russia)

    1996-07-01

    One of the most intriguing and crucial problems of the prebiotic evolution and the origin of life is the explanation of the origin of biohomochirality. A scheme of conversions originated by formaldehyde (FA) as hypothetical primer of biohomochirality is proposed. The merit of FA as executor of this function is based -inter alia - on the distinguished role of FA as one of the earliest and simplest molecules in both warm, terrestrial and cold, extraterrestrial scenarios of the origin of life. The confirmation of the role of FA as primer of biohomochirality would support the option of an RNA world as an alternative to the protein world. The suggested hypothesis puts forward for the first time a concrete sequence of chemical reactions which can lead to biohomochirality. The spontaneous breaking of the mirror symmetry is secured by the application of the well-known Frank scheme (combination of autocatalysis and {open_quote}{open_quote}annihilation{close_quote}{close_quote} of L and D enantiomers) to the series of interactions of FA {open_quote}{open_quote}trimers{close_quote}{close_quote} (i.e. C{sub 3}H{sub 6}O{sub 3} compounds) of (aaa), (apa) and (app) types, where the monomeric groups (a) means {open_quote}{open_quote}achirons{close_quote}{close_quote} (a=CH{sub n}, n{ge}2 and C=M, M=C,O) and (p) mean {open_quote}{open_quote}prochirons{close_quote}{close_quote} (p=HC{asterisk}OM, M=H,C). {copyright} {ital 1996 American Institute of Physics.}

  13. Synthesis, properties, and NMR studies of a C8-phenylguanine modified oligonucleotide that preferentially adopts the Z DNA conformation.

    Science.gov (United States)

    Gannett, Peter M; Heavner, Sue; Daft, Jonathan R; Shaughnessy, Kevin H; Epperson, Jon D; Greenbaum, Nancy L

    2003-10-01

    Carcinogenic aryl hydrazines produce C8-arylated purine adducts. The effect of these adducts on DNA conformation and their role in hydrazine carcinogenesis are unknown. Here, we describe a new synthetic route to produce these adducts that is also compatible with the synthesis of the corresponding phosphoramidites needed for oligonucleotide synthesis. Two oligonucleotides were prepared, an unmodified oligonucleotide, d((5)(')CGCGCGCGCG(3)(')), and a C8-phenylguanine modified oligonucleotide, d((5)(')CGCGCGCGCG(3)(')) (G = 8-phenylguanine). These oligonucleotides were compared using thermal denaturation, circular dichroism, NMR, and molecular modeling. The phenyl modification destabilizes the B DNA form and stabilizes the Z DNA form such that the B:Z ratio is near one under physiological conditions. In light of recent studies that show a role for Z DNA in gene expression and cell transformation, Z DNA stabilization by C8-arylguanine formation from aryl hydrazines may be relevant to their role in carcinogenesis.

  14. Hierarchical resolution of power spectra

    Science.gov (United States)

    Badii, R.; Finardi, M.; Broggi, G.; Sepúlveda, M. A.

    1992-09-01

    We identify the basic ingredients determining the structure of the power spectra of non-linear dynamical systems in a hierarchical order of importance. The analysis, performed with the help of symbolic methods, shows that dynamical invariants such as topological and metric properties of the symbolic orbits explain the main qualitative features of the spectra, whereas the coordinate-dependent values of the observable itself represent a less relevant contribution. Consideration of simple dynamical models with increasing number of topological transition rules evidences the formation of coherent structures (peaks) and explains their position and size. By constructing the parse tree of the allowed symbolic itineraries, it is possible to estimate conditional probabilities by considering orbits belonging to adjacent tree levels. Accordingly, a Markov transition matrix is obtained for each level l and is used to generate signals with statistical properties which approximate those of the actual one increasingly better for l → ∞. A considerable improvement is achieved by recoding the original signal in terms of variable-length words and by re-applying the above procedure to the transformed signal, which is equivalent to a renormalization operation of the associated dynamical map. The accuracy of the estimates is directly related to the convergence of the scaling function for the conditional probabilities. Analytic results are presented for the simplest five Markov models arising from piecewise-linear, continuous, one-dimensional maps. Numerical studies have been performed for the logistic and Hénon maps and for the Lorenz system.

  15. Hierarchically nanostructured barium sulfate fibers.

    Science.gov (United States)

    Romero-Ibarra, Issis C; Rodríguez-Gattorno, Geonel; García-Sánchez, Mario F; Sánchez-Solís, Antonio; Manero, Octavio

    2010-05-18

    BaSO(4) nanostructures with controlled morphologies were successfully produced via one-step process through precipitation of BaSO(4) in aqueous and organic media. The synthesis is carried out by mixing solutions of BaCl(2) and Na(2)SO(4) in presence of EDTA (disodium ethylenediaminetetraacetic acid) at room temperature. The influence of the reaction conditions such as initial reactants concentration, pH, EDTA/[Ba(2+)] ratio and aging on the BaSO(4) nanoparticles organization is studied. Using EDTA in aqueous media, spherical secondary particles of 500 nm diameter are obtained, which are formed by 4 nm size primary particles. With dimethyl sulfoxide and small amounts of water (5%) and EDTA, the aging process allows the production of long homogeneous fibers, related to hierarchical organization of BaSO(4) nanoparticles. Direct observation of self-assembling of primary particles by HRTEM allows proposing a mechanism for fiber formation, which is based on multipolar attractions that lead to a brick-by-brick organization along a preferential orientation. Results evidence the role of EDTA as controlling agent of the morphology and primary and secondary mean particle size.

  16. A hierarchical theory of macroecology.

    Science.gov (United States)

    Passy, Sophia I

    2012-09-01

    The relationships of local population density (N) with body size (M) and distribution (D) have been extensively studied because they reveal how ecological and historical factors structure species communities; however, a unifying model explaining their joint behaviour, has not been developed. Here, I propose a theory that explores these relationships hierarchically and predicts that: (1) at a metacommunity level, niche breadth, population density and regional distribution are all related and size-dependent and (2) at a community level, the exponents b and d of the relationships N ~ M (b) and N ~ D (d) are functions (f) of the environment and, consequently, species richness (S), allowing the following reformulation of the power laws: N ~ M (f(S)) and N ~ D (f(S)) . Using this framework and continental data on stream environment, diatoms, invertebrates and fish, I address the following fundamental, but unresolved ecological questions: how do species partition their resources across environments, is energetic equivalence among them possible, are generalists more common than specialists, why are locally abundant species also regionally prevalent, and, do microbes have different biogeography than macroorganisms? The discovery that community scaling behaviour is environmentally constrained calls for better integration of macroecology and environmental science. © 2012 Blackwell Publishing Ltd/CNRS.

  17. Hierarchical analysis of acceptable use policies

    Directory of Open Access Journals (Sweden)

    P. A. Laughton

    2008-01-01

    Full Text Available Acceptable use policies (AUPs are vital tools for organizations to protect themselves and their employees from misuse of computer facilities provided. A well structured, thorough AUP is essential for any organization. It is impossible for an effective AUP to deal with every clause and remain readable. For this reason, some sections of an AUP carry more weight than others, denoting importance. The methodology used to develop the hierarchical analysis is a literature review, where various sources were consulted. This hierarchical approach to AUP analysis attempts to highlight important sections and clauses dealt with in an AUP. The emphasis of the hierarchal analysis is to prioritize the objectives of an AUP.

  18. Classification using Hierarchical Naive Bayes models

    DEFF Research Database (Denmark)

    Langseth, Helge; Dyhre Nielsen, Thomas

    2006-01-01

    Classification problems have a long history in the machine learning literature. One of the simplest, and yet most consistently well-performing set of classifiers is the Naïve Bayes models. However, an inherent problem with these classifiers is the assumption that all attributes used to describe......, termed Hierarchical Naïve Bayes models. Hierarchical Naïve Bayes models extend the modeling flexibility of Naïve Bayes models by introducing latent variables to relax some of the independence statements in these models. We propose a simple algorithm for learning Hierarchical Naïve Bayes models...

  19. Hierarchical modeling and analysis for spatial data

    CERN Document Server

    Banerjee, Sudipto; Gelfand, Alan E

    2003-01-01

    Among the many uses of hierarchical modeling, their application to the statistical analysis of spatial and spatio-temporal data from areas such as epidemiology And environmental science has proven particularly fruitful. Yet to date, the few books that address the subject have been either too narrowly focused on specific aspects of spatial analysis, or written at a level often inaccessible to those lacking a strong background in mathematical statistics.Hierarchical Modeling and Analysis for Spatial Data is the first accessible, self-contained treatment of hierarchical methods, modeling, and dat

  20. Hierarchically structured, nitrogen-doped carbon membranes

    KAUST Repository

    Wang, Hong

    2017-08-03

    The present invention is a structure, method of making and method of use for a novel macroscopic hierarchically structured, nitrogen-doped, nano-porous carbon membrane (HNDCMs) with asymmetric and hierarchical pore architecture that can be produced on a large-scale approach. The unique HNDCM holds great promise as components in separation and advanced carbon devices because they could offer unconventional fluidic transport phenomena on the nanoscale. Overall, the invention set forth herein covers a hierarchically structured, nitrogen-doped carbon membranes and methods of making and using such a membranes.

  1. Oligonucleotide-assisted cleavage and ligation: a novel directional DNA cloning technology to capture cDNAs. Application in the construction of a human immune antibody phage-display library

    Science.gov (United States)

    Schoonbroodt, Sonia; Frans, Nicolas; DeSouza, Mark; Eren, Rachel; Priel, Smadar; Brosh, Naama; Ben-Porath, Judith; Zauberman, Arie; Ilan, Ehud; Dagan, Shlomo; Cohen, Edward H.; Hoogenboom, Hennie R.; Ladner, Robert Charles; Hoet, René M.

    2005-01-01

    The use of oligonucleotide-assisted cleavage and ligation (ONCL), a novel approach to the capture of gene repertoires, in the construction of a phage-display immune antibody library is described. ONCL begins with rapid amplification of cDNA ends to amplify all members equally. A single, specific cut near 5′ and/or 3′ end of each gene fragment (in single stranded form) is facilitated by hybridization with an appropriate oligonucleotide adapter. Directional cloning of targeted DNA is accomplished by ligation of a partially duplex DNA molecule (containing suitable restriction sites) and amplification with primers in constant regions. To demonstrate utility and reliability of ONCL, a human antibody repertoire was cloned from IgG mRNA extracted from human B-lymphocytes engrafted in Trimera mice. These mice were transplanted with peripheral blood lymphocytes from Candida albicans infected individuals and subsequently immunized with C.albicans glyceraldehyde-3-phosphate dehydrogenase (GAPDH). DNA sequencing showed that ONCL resulted in efficient capture of gene repertoires. Indeed, full representation of all VH families/segments was observed showing that ONCL did not introduce cloning biases for or against any VH family. We validated the efficiency of ONCL by creating a functional Fab phage-display library with a size of 3.3 × 1010 and by selecting five unique Fabs against GAPDH antigen. PMID:15905471

  2. Short locked nucleic acid antisense oligonucleotides potently reduce apolipoprotein B mRNA and serum cholesterol in mice and non-human primates

    DEFF Research Database (Denmark)

    Straarup, Ellen Marie; Fisker, Niels; Hedtjärn, Maj

    2010-01-01

    -high-density lipoprotein (non-HDL) cholesterol without increasing serum liver toxicity markers. The data presented here show that oligonucleotide length as a parameter needs to be considered in the design of antisense oligonucleotide and that potent short oligonucleotides with sufficient target affinity can be generated...... using the LNA chemistry. Conclusively, we present a 13-mer LNA oligonucleotide with therapeutic potential that produce beneficial cholesterol lowering effect in non-human primates....

  3. Adhesive primers for bonding cobalt-chromium alloy to resin.

    Science.gov (United States)

    Yoshida, K; Kamada, K; Atsuta, M

    1999-06-01

    This study evaluated the effect of five adhesive primers on the shear bond strength of a self-curing resin to cobalt-chromium (Co-Cr) alloy. The adhesive primers Acryl Bond (AB, Shofu), Cesead Opaque Primer (COP, Kuraray), Metacolor Opaque Bonding Liner (MOBL, Sun-Medical), Metal PrimerII (MPII, GC) and MR. Bond (MRB, Tokuyama) were used. A brass ring which was placed over the casting alloy disk surface non-primed or primed with each primer was filled with the self-curing MMA-PMMA resin. The specimens were stored in water at 37 degrees C for 24 h and then immersed alternately in water baths at 4 C and 60 degrees C for 1 min each for up to 50000 thermal cycles before shear mode testing at a crosshead speed of 0.5 mm/min. All of the primers examined, except MOBL, improved the shear bond strength between the resin and Co-Cr alloy compared with nonprimed specimens prior to thermal cycling. Regardless of which primer was used, the shear bond strength significantly differed between thermal cycles 0 and 50000. However, after 50000 thermal cycles, the bond strengths of resin to Co-Cr alloy primed with COP or MPII were significantly greater than those of specimens primed with AB, MOBL or MRB and non-primed controls. This study indicated that COP and MPII are effective primers to obtain higher bond strength between resin and Co-Cr alloy.

  4. Marketing Information Products and Services: A Primer for Librarians ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    ... and Services: A Primer for Librarians and Information Professionals. Book cover Marketing Information Products and Services: A Primer for Librarians and Information Professionals. Directeur(s) : Abhinandan K. Jain, Ashok Jambhekar, T.P.Rama Rao, and S. Sreenivas Rao. Maison(s) d'édition : Tata McGraw-Hill, IDRC.

  5. Marketing Information Products and Services : A Primer for ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    ... and Services : A Primer for Librarians and Information Professionals. Couverture du livre Marketing Information Products and Services : A Primer for Librarians and Information Professionals. Editor(s):. Abhinandan K. Jain, Ashok Jambhekar, T.P.Rama Rao et S. Sreenivas Rao. Publisher(s):. Tata McGraw-Hill, CRDI.

  6. Mapping of randomly amplified polymorphic DNA primer (RAPD) on ...

    African Journals Online (AJOL)

    Genet & Botany only

    2012-08-14

    Aug 14, 2012 ... RAPD primers GLC-07 and GLB-11. PCR amplification using primer GLC-07 produced single band of ... PCR amplification profile of the two genetic stocks of common wheat,. NT2A2B and NT1D1B using .... Isolation of recombinants involving barley arms 3HL and 6HL. Theor. Appl. Genet. 83:489-494.

  7. Bioinformatic tools and guideline for PCR primer design | Abd ...

    African Journals Online (AJOL)

    This review intends to provide a guide to choosing the most efficient way to design a new specific-primer by applying current publicly available links and Web services. Also, the purpose here is to provide general recommendations for the design and use of PCR primers. (African Journal of Biotechnology: 2003 2(5): 91-95) ...

  8. Detection of mutations in genes by specific LNA primers

    DEFF Research Database (Denmark)

    2001-01-01

    The present invention relates to a method of detecting variant nucleic acid whose nucleotide sequence differs from one another at a single (or more) position(s). The method uses a set of chimeric oligonucleotides containing DNA monomers and monomers of a novel class of DNA analogues, locked nucle...

  9. Pd0-Catalyzed Methyl Transfer on Nucleosides and Oligonucleotides, Envisaged as a PET Tracer

    Directory of Open Access Journals (Sweden)

    Eric Fouquet

    2013-11-01

    Full Text Available The methyl transfer reaction from activated monomethyltin, via a modified Stille coupling reaction, was studied under “ligandless” conditions on fully deprotected 5'-modified nucleosides and one dinucleotide. The reaction was optimized to proceed in a few minutes and quantitative yield, even under dilute conditions, thus affording a rapid and efficient new method for oligonucleotide labelling with carbon-11.

  10. Fatty acid-modified gapmer antisense oligonucleotide and human serum albumin constructs for pharmacokinetic modulation

    DEFF Research Database (Denmark)

    Hvam, Michael Lykke; Cai, Yunpeng; Dagnæs-Hansen, Frederik

    2017-01-01

    oligonucleotides (ASOs)/albumin constructs. We show by an electrophoretic mobility assay that fatty acid-modified gapmer and human serum albumin (HSA) can self-assemble into constructs that offer favorable pharmacokinetics. The interaction was dependent on fatty acid type (either palmitic or myristic acid), number...

  11. Optimizing anti-gene oligonucleotide 'Zorro-LNA' for improved strand invasion into duplex DNA

    DEFF Research Database (Denmark)

    Zaghloul, Eman M; Madsen, Andreas S; Moreno, Pedro M D

    2011-01-01

    Zorro-LNA (Zorro) is a newly developed, oligonucleotide (ON)-based, Z-shaped construct with the potential of specific binding to each strand of duplex DNA. The first-generation Zorros are formed by two hybridized LNA/DNA mixmers (2-ON Zorros) and was hypothesized to strand invade. We have now...

  12. LNA for Optimization of Fluorescent Oligonucleotide Probes: Improved Spectral Properties and Target Binding

    DEFF Research Database (Denmark)

    Astakhova, Irina V; Ustinov, Alexey V; Korshun, Vladimir A

    2011-01-01

    Mixmer LNA/DNA fluorescent probes containing the 1-(phenylethynyl)pyrene fluorophore attached to 2'-arabino-uridine were synthesized and studied. The conjugates displayed significantly higher hybridization affinity to target DNA, increased fluorescence quantum yields of single-stranded oligonucle......-stranded oligonucleotides and their duplexes, and improved ability to form an interstrand excimer compared to analogous non-LNA probes....

  13. Whole-cell hybridization of Methanosarcina cells with two new oligonucleotide probes

    DEFF Research Database (Denmark)

    Sørensen, A.H.; Torsvik, V.L.; Torsvik, T.

    1997-01-01

    Two new oligonucleotide probes targeting the 16S rRNA of the methanogenic genus Methanosarcina were developed. The probes have the following sequences (Escherichia coli numbering): probe SARCI551, 5'-GAC CCAATAATCACGATCAC-3', and probe SARCI645, 5'-TCCCGGTTCCAAGTCTGGC-3'. In situ hybridization...

  14. Comparison of small molecules and oligonucleotides that target a toxic, non-coding RNA.

    Science.gov (United States)

    Costales, Matthew G; Rzuczek, Suzanne G; Disney, Matthew D

    2016-06-01

    Potential RNA targets for chemical probes and therapeutic modalities are pervasive in the transcriptome. Oligonucleotide-based therapeutics are commonly used to target RNA sequence. Small molecules are emerging as a modality to target RNA structures selectively, but their development is still in its infancy. In this work, we compare the activity of oligonucleotides and several classes of small molecules that target the non-coding r(CCUG) repeat expansion (r(CCUG)(exp)) that causes myotonic dystrophy type 2 (DM2), an incurable disease that is the second-most common cause of adult onset muscular dystrophy. Small molecule types investigated include monomers, dimers, and multivalent compounds synthesized on-site by using RNA-templated click chemistry. Oligonucleotides investigated include phosphorothioates that cleave their target and vivo-morpholinos that modulate target RNA activity via binding. We show that compounds assembled on-site that recognize structure have the highest potencies amongst small molecules and are similar in potency to a vivo-morpholino modified oligonucleotide that targets sequence. These studies are likely to impact the design of therapeutic modalities targeting other repeats expansions that cause fragile X syndrome and amyotrophic lateral sclerosis, for example. Copyright © 2016. Published by Elsevier Ltd.

  15. A facile inhibitor screening of SARS coronavirus N protein using nanoparticle-based RNA oligonucleotide.

    Science.gov (United States)

    Roh, Changhyun

    2012-01-01

    Hundreds of million people worldwide have been infected with severe acute respiratory syndrome (SARS), and the rate of global death from SARS has remarkably increased. Hence, the development of efficient drug treatments for the biological effects of SARS is highly needed. We have previously shown that quantum dots (QDs)-conjugated RNA oligonucleotide is sensitive to the specific recognition of the SARS-associated coronavirus (SARS-CoV) nucleocapsid (N) protein. In this study, we found that a designed biochip could analyze inhibitors of the SARS-CoV N protein using nanoparticle-based RNA oligonucleotide. Among the polyphenolic compounds examined, (-)-catechin gallate and (-)-gallocatechin gallate demonstrated a remarkable inhibition activity on SARS-CoV N protein. (-)-catechin gallate and (-)-gallocatechin gallate attenuated the binding affinity in a concentrated manner as evidenced by QDs-conjugated RNA oligonucleotide on a designed biochip. At a concentration of 0.05 μg mL(-1), (-)-catechin gallate and (-)-gallocatechin gallate showed more than 40% inhibition activity on a nanoparticle-based RNA oligonucleotide biochip system.

  16. Efficient targeting of fatty-acid modified oligonucleotides to live cell membranes through stepwise assembly

    OpenAIRE

    Desai, Tejal; Gartner, Zev; Weber, RJ; Liang, Si; Selden, NS; Desai, TA; Gartner, ZJ

    2014-01-01

    © 2014 American Chemical Society.Lipid modifications provide efficient targeting of oligonucleotides to live cell membranes in a range of applications. Targeting efficiency is a function of the rate of lipid DNA insertion into the cell surface and its pers

  17. Properties of amphiphilic oligonucleotide films at the air/water interface and after film transfer

    NARCIS (Netherlands)

    Keller, R.; Kwak, M.; de Vries, J. W.; Sawaryn, C.; Wang, J.; Anaya, M.; Muellen, K.; Butt, H. -J.; Herrmann, A.; Berger, R.; Müllen, K.

    2013-01-01

    The self-assembly of amphiphilic hybrid materials containing an oligonucleotide sequence at the air/water interface was investigated by means of pressure-molecular area (Pi-A) isotherms. In addition, films were transferred onto solid substrates and imaged using scanning force microscopy. We used

  18. Folding Topology of a Short Coiled-Coil Peptide Structure Templated by an Oligonucleotide Triplex

    DEFF Research Database (Denmark)

    Lou, Chenguang; Christensen, Niels Johan; Martos Maldonado, Manuel Cristo

    2017-01-01

    The rational design of a well-defined protein-like tertiary structure formed by small peptide building blocks is still a formidable challenge. By using peptide-oligonucleotide conjugates (POC) as building blocks, we present the self-assembly of miniature coiled-coil α-helical peptides guided...

  19. High-frequency genome editing using ssDNA oligonucleotides with zinc-finger nucleases

    DEFF Research Database (Denmark)

    Chen, Fuqiang; Pruett-Miller, Shondra M; Huang, Yuping

    2011-01-01

    Zinc-finger nucleases (ZFNs) have enabled highly efficient gene targeting in multiple cell types and organisms. Here we describe methods for using simple ssDNA oligonucleotides in tandem with ZFNs to efficiently produce human cell lines with three distinct genetic outcomes: (i) targeted point...

  20. [Use of a synthetic oligonucleotide (CAC)5 in the genomic "dactyloscopy" method].

    Science.gov (United States)

    Korokhov, N P; Popovskiĭ, A V; Sharonova, D A; Novoselov, V P

    1993-01-01

    Potentialities of a chemically synthetized oligonucleotide of a certain structure in the genomic "dactyloscopy" method were under study. A random sample analysis has demonstrated a high resolving power of this variant of the method. Arguments in favor of introducing this approach in practical forensic medical direct identification of biologic material are presented.

  1. Microfluidic bead trap as a visual bar for quantitative detection of oligonucleotides.

    Science.gov (United States)

    Zhao, Zichen; Bao, Yuanye; Chu, Lok Ting; Ho, John Kin Lim; Chieng, Ching-Chang; Chen, Ting-Hsuan

    2017-09-26

    We demonstrate a microfluidic bead trap capable of forming a dipstick-type bar visible to the naked eye for simple and quantitative detection of oligonucleotides. We use magnetic microparticles (MMPs) and polystyrene microparticles (PMPs) that are connected and form MMPs-targets-PMPs when target oligonucleotides are present, leaving free PMPs with a number inversely proportional to the amount of targets. Using a capillary flow-driven microfluidic circuitry consisting of a magnetic separator to remove the MMPs-targets-PMPs, the free PMPs can be trapped at the narrowing nozzle downstream, forming a visual bar quantifiable based on the length of PMP accumulation. Such a power-free and instrument-free platform enables a limit of detection at 13 fmol (0.65 nM in 20 μl, S/N = 3) of oligonucleotides and is compatible with single-nucleotide polymorphisms and operation in a complex bio-fluid. Moreover, using DNAzyme as the target oligonucleotide that catalyzes a specific hydrolytic cleavage in the presence of lead ions, we demonstrate a model application that detects lead ions with a limit of detection of 12.2 nM (2.5 μg l-1), providing quantitative and visual detection of lead contamination at resource-limited sites.

  2. nuID: a universal naming scheme of oligonucleotides for Illumina, Affymetrix, and other microarrays

    Directory of Open Access Journals (Sweden)

    Kibbe Warren A

    2007-05-01

    Full Text Available Abstract Background Oligonucleotide probes that are sequence identical may have different identifiers between manufacturers and even between different versions of the same company's microarray; and sometimes the same identifier is reused and represents a completely different oligonucleotide, resulting in ambiguity and potentially mis-identification of the genes hybridizing to that probe. Results We have devised a unique, non-degenerate encoding scheme that can be used as a universal representation to identify an oligonucleotide across manufacturers. We have named the encoded representation 'nuID', for nucleotide universal identifier. Inspired by the fact that the raw sequence of the oligonucleotide is the true definition of identity for a probe, the encoding algorithm uniquely and non-degenerately transforms the sequence itself into a compact identifier (a lossless compression. In addition, we added a redundancy check (checksum to validate the integrity of the identifier. These two steps, encoding plus checksum, result in an nuID, which is a unique, non-degenerate, permanent, robust and efficient representation of the probe sequence. For commercial applications that require the sequence identity to be confidential, we have an encryption schema for nuID. We demonstrate the utility of nuIDs for the annotation of Illumina microarrays, and we believe it has universal applicability as a source-independent naming convention for oligomers. Reviewers This article was reviewed by Itai Yanai, Rong Chen (nominated by Mark Gerstein, and Gregory Schuler (nominated by David Lipman.

  3. Rapid identification of bacteria in blood cultures by using fluorescently labeled oligonucleotide probes

    NARCIS (Netherlands)

    Jansen, GJ; Mooibroek, M; Idema, J; Harmsen, HJM; Welling, GW; Degener, JE

    The applicability of whole-cell hybridization for the identification of pathogenic bacteria in blood from septic patients was examined. Oligonucleotide probes, fluorescently labeled with fluorescein isothiocyanate, directed against the variable regions of the 16S rRNAs of the following bacterial

  4. Splice-correcting oligonucleotides restore BTK function in X-linked agammaglobulinemia model

    DEFF Research Database (Denmark)

    Bestas, Burcu; Moreno, Pedro M D; Blomberg, K Emelie M

    2014-01-01

    , splice-correcting oligonucleotides (SCOs) targeting mutated BTK transcripts for treating XLA. Both the SCO structural design and chemical properties were optimized using 2'-O-methyl, locked nucleic acid, or phosphorodiamidate morpholino backbones. In order to have access to an animal model of XLA, we...

  5. Precise construction of oligonucleotide-Fab fragment conjugate for homogeneous immunoassay using HaloTag technology.

    Science.gov (United States)

    Päkkilä, Henna; Peltomaa, Riikka; Lamminmäki, Urpo; Soukka, Tero

    2015-03-01

    The use of oligonucleotide-protein conjugates enables the development of novel types of bioanalytical assays. However, convenient methods for producing covalent and stoichiometric oligonucleotide-protein conjugates are still rare. Here we demonstrate, for the first time, covalent conjugation of DNA oligonucleotide to Fab fragments with a 1:1 ratio using HaloTag self-labeling technology. The oligonucleotide coupling was carried out while the Fab was attached to protein G matrix, thereby enabling straightforward production of covalent conjugates. Furthermore, it allowed convenient purification of the product because the unreacted components were easily removed before the elution of the high-purity conjugate. The prepared conjugate was employed in a homogeneous immunoassay where prostate-specific antigen was used as a model analyte. Switchable lanthanide luminescence was used for detection, and the obtained limit of detection was 0.27 ng/ml. In the future, the developed method for covalent conjugation and successive purification in protein G column could also be applied for introducing other kinds of modifications to Fab fragments in a simple and site-specific manner. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Synthetic Nucleic Acid Analogues in Gene Therapy: An Update for Peptide–Oligonucleotide Conjugates

    DEFF Research Database (Denmark)

    Taskova, Maria; Mantsiou, Anna; Astakhova, Kira

    2017-01-01

    The main objective of this work is to provide an update on synthetic nucleic acid analogues and nanoassemblies as tools in gene therapy. In particular, the synthesis and properties of peptide–oligonucleotide conjugates (POCs), which have high potential in research and as therapeutics, are described...

  7. A new achiral reagent for the incorporation of multiple amino groups into oligonucleotides

    DEFF Research Database (Denmark)

    Behrens, Carsten; Petersen, Kenneth H.; Egholm, Michael

    1995-01-01

    The synthesis of a new functionalized achiral linker reagent (10) for the incorporation of multiple primary amino groups into oligonucleotides is described. The linker reagent is compatible with conventional DNA-synthesis following the phosphoramidite methodology, and the linker can be incorporated...

  8. Species‐specific oligonucleotide probes for the detection and identification of Lactobacillus isolated from mouse faeces

    National Research Council Canada - National Science Library

    Park, S.H; Itoh, K

    2005-01-01

    .... Methods and Results:  The specificity of oligonucleotide probes was evaluated by dot blot hybridization to 16S rDNA and 23S rDNA amplified by PCR from 12 Lactobacillus type strains and 100 strains of Lactobacillus isolated from mouse faeces...

  9. High-speed measurement of firearm primer blast waves

    CERN Document Server

    Courtney, Michael; Eng, Jonathan; Courtney, Amy

    2012-01-01

    This article describes a method and results for direct high-speed measurements of firearm primer blast waves employing a high-speed pressure transducer located at the muzzle to record the blast pressure wave produced by primer ignition. Key findings are: 1) Most of the lead styphnate based primer models tested show 5.2-11.3% standard deviation in the magnitudes of their peak pressure. 2) In contrast, lead-free diazodinitrophenol (DDNP) based primers had standard deviations of the peak blast pressure of 8.2-25.0%. 3) Combined with smaller blast waves, these large variations in peak blast pressure of DDNP-based primers led to delayed ignition and failure to fire in brief field tests.

  10. HIERARCHICAL ORGANIZATION OF INFORMATION, IN RELATIONAL DATABASES

    Directory of Open Access Journals (Sweden)

    Demian Horia

    2008-05-01

    Full Text Available In this paper I will present different types of representation, of hierarchical information inside a relational database. I also will compare them to find the best organization for specific scenarios.

  11. Packaging glass with hierarchically nanostructured surface

    KAUST Repository

    He, Jr-Hau

    2017-08-03

    An optical device includes an active region and packaging glass located on top of the active region. A top surface of the packaging glass includes hierarchical nanostructures comprised of honeycombed nanowalls (HNWs) and nanorod (NR) structures extending from the HNWs.

  12. Hierarchical DSE for multi-ASIP platforms

    DEFF Research Database (Denmark)

    Micconi, Laura; Corvino, Rosilde; Gangadharan, Deepak

    2013-01-01

    This work proposes a hierarchical Design Space Exploration (DSE) for the design of multi-processor platforms targeted to specific applications with strict timing and area constraints. In particular, it considers platforms integrating multiple Application Specific Instruction Set Processors (ASIPs...

  13. Hierarchical modularity in human brain functional networks

    CERN Document Server

    Meunier, D; Fornito, A; Ersche, K D; Bullmore, E T; 10.3389/neuro.11.037.2009

    2010-01-01

    The idea that complex systems have a hierarchical modular organization originates in the early 1960s and has recently attracted fresh support from quantitative studies of large scale, real-life networks. Here we investigate the hierarchical modular (or "modules-within-modules") decomposition of human brain functional networks, measured using functional magnetic resonance imaging (fMRI) in 18 healthy volunteers under no-task or resting conditions. We used a customized template to extract networks with more than 1800 regional nodes, and we applied a fast algorithm to identify nested modular structure at several hierarchical levels. We used mutual information, 0 < I < 1, to estimate the similarity of community structure of networks in different subjects, and to identify the individual network that is most representative of the group. Results show that human brain functional networks have a hierarchical modular organization with a fair degree of similarity between subjects, I=0.63. The largest 5 modules at ...

  14. HIERARCHICAL OPTIMIZATION MODEL ON GEONETWORK

    Directory of Open Access Journals (Sweden)

    Z. Zha

    2012-07-01

    Full Text Available In existing construction experience of Spatial Data Infrastructure (SDI, GeoNetwork, as the geographical information integrated solution, is an effective way of building SDI. During GeoNetwork serving as an internet application, several shortcomings are exposed. The first one is that the time consuming of data loading has been considerately increasing with the growth of metadata count. Consequently, the efficiency of query and search service becomes lower. Another problem is that stability and robustness are both ruined since huge amount of metadata. The final flaw is that the requirements of multi-user concurrent accessing based on massive data are not effectively satisfied on the internet. A novel approach, Hierarchical Optimization Model (HOM, is presented to solve the incapability of GeoNetwork working with massive data in this paper. HOM optimizes the GeoNetwork from these aspects: internal procedure, external deployment strategies, etc. This model builds an efficient index for accessing huge metadata and supporting concurrent processes. In this way, the services based on GeoNetwork can maintain stable while running massive metadata. As an experiment, we deployed more than 30 GeoNetwork nodes, and harvest nearly 1.1 million metadata. From the contrast between the HOM-improved software and the original one, the model makes indexing and retrieval processes more quickly and keeps the speed stable on metadata amount increasing. It also shows stable on multi-user concurrent accessing to system services, the experiment achieved good results and proved that our optimization model is efficient and reliable.

  15. Feature Selection in Hierarchical Feature Spaces

    OpenAIRE

    Ristoski, Petar; Paulheim, Heiko

    2014-01-01

    Feature selection is an important preprocessing step in data mining, which has an impact on both the runtime and the result quality of the subsequent processing steps. While there are many cases where hierarchic relations between features exist, most existing feature selection approaches are not capable of exploiting those relations. In this paper, we introduce a method for feature selection in hierarchical feature spaces. The method first eliminates redundant features along paths in the hier...

  16. Hierarchical Solution of Large Markov Decision Processes

    OpenAIRE

    Barry, Jennifer; Kaelbling, Leslie P.; Lozano-Perez, Tomas

    2010-01-01

    This paper presents an algorithm for finding approximately optimal policies in very large Markov decision processes by constructing a hierarchical model and then solving it. This strategy sacrifices optimality for the ability to address a large class of very large problems. Our algorithm works efficiently on enumerated-states and factored MDPs by constructing a hierarchical structure that is no larger than both the reduced model of the MDP and the regression tree for the ...

  17. Metal hierarchical patterning by direct nanoimprint lithography

    OpenAIRE

    Boya Radha; Su Hui Lim; Saifullah, Mohammad S. M.; Kulkarni, Giridhar U.

    2013-01-01

    Three-dimensional hierarchical patterning of metals is of paramount importance in diverse fields involving photonics, controlling surface wettability and wearable electronics. Conventionally, this type of structuring is tedious and usually involves layer-by-layer lithographic patterning. Here, we describe a simple process of direct nanoimprint lithography using palladium benzylthiolate, a versatile metal-organic ink, which not only leads to the formation of hierarchical patterns but also is a...

  18. Hierarchical Model of Container Ports Throughput

    Directory of Open Access Journals (Sweden)

    Monika Rozmarynowska

    2015-12-01

    Full Text Available In this article the attempt has been made to construct hierarchical model of container ports throughput development. The presented hierarchical approach uses the relationships of development of global economy and container flows at different geographical levels: global (container throughput in all seaport on the world, regional (container throughput in the Baltic seaports and national (container throughput in Polish seaports. Model have been evaluated for their fit and usefulness for predictive purposes.

  19. Hierarchical organization versus self-organization

    OpenAIRE

    Busseniers, Evo

    2014-01-01

    In this paper we try to define the difference between hierarchical organization and self-organization. Organization is defined as a structure with a function. So we can define the difference between hierarchical organization and self-organization both on the structure as on the function. In the next two chapters these two definitions are given. For the structure we will use some existing definitions in graph theory, for the function we will use existing theory on (self-)organization. In the t...

  20. Translating Management Practices in Hierarchical Organizations

    DEFF Research Database (Denmark)

    Wæraas, Arild; Nielsen, Jeppe Agger

    structures affect translators’ approaches taken towards management ideas. This paper reports the findings from a longitudinal case study of the translation of Leadership Pipeline in a Danish fire department and how the translators’ approach changed over time from a modifying to a reproducing mode. The study...... hierarchical structures offer translators considerable control over the translation process. This hierarchical control seems to reduce the likelihood of unpredictable translation outcomes....

  1. Learning Hierarchical Feature Extractors for Image Recognition

    Science.gov (United States)

    2012-09-01

    Learning Hierarchical Feature Extractors For Image Recognition by Y-Lan Boureau A dissertation submitted in partial fulfillment of the requirements...DATES COVERED 00-00-2012 to 00-00-2012 4. TITLE AND SUBTITLE Learning Hierarchical Feature Extractors For Image Recognition 5a. CONTRACT...pooling for all weighting schemes. With average pooling, weighting by the square root of the cluster weight performs best. P = 16 configuration space

  2. Hierarchical control architecture for mobile offshore bases

    OpenAIRE

    Girard, A; Karl Hedrick; João Tasso de Figueiredo Borges de Sousa

    2000-01-01

    A hierarchical architecture for Mobile Offshore Bases (MOB) control is presented. By a control architecture we mean a specific way of organizing the motion control and navigation functions performed by the MOB. It is convenient to organize the functions into hierarchical layers. This way, a complex design problem is partitioned into a number of more manageable sub-problems that are addressed in separate layers. The decomposition also allows for modular design and testing and the incorporation...

  3. Development, characterization and experimental validation of a cultivated sunflower (Helianthus annuus L. gene expression oligonucleotide microarray.

    Directory of Open Access Journals (Sweden)

    Paula Fernandez

    Full Text Available Oligonucleotide-based microarrays with accurate gene coverage represent a key strategy for transcriptional studies in orphan species such as sunflower, H. annuus L., which lacks full genome sequences. The goal of this study was the development and functional annotation of a comprehensive sunflower unigene collection and the design and validation of a custom sunflower oligonucleotide-based microarray. A large scale EST (>130,000 ESTs curation, assembly and sequence annotation was performed using Blast2GO (www.blast2go.de. The EST assembly comprises 41,013 putative transcripts (12,924 contigs and 28,089 singletons. The resulting Sunflower Unigen Resource (SUR version 1.0 was used to design an oligonucleotide-based Agilent microarray for cultivated sunflower. This microarray includes a total of 42,326 features: 1,417 Agilent controls, 74 control probes for sunflower replicated 10 times (740 controls and 40,169 different non-control probes. Microarray performance was validated using a model experiment examining the induction of senescence by water deficit. Pre-processing and differential expression analysis of Agilent microarrays was performed using the Bioconductor limma package. The analyses based on p-values calculated by eBayes (p<0.01 allowed the detection of 558 differentially expressed genes between water stress and control conditions; from these, ten genes were further validated by qPCR. Over-represented ontologies were identified using FatiScan in the Babelomics suite. This work generated a curated and trustable sunflower unigene collection, and a custom, validated sunflower oligonucleotide-based microarray using Agilent technology. Both the curated unigene collection and the validated oligonucleotide microarray provide key resources for sunflower genome analysis, transcriptional studies, and molecular breeding for crop improvement.

  4. Drop drying on surfaces determines chemical reactivity - the specific case of immobilization of oligonucleotides on microarrays

    Science.gov (United States)

    2013-01-01

    Background Drop drying is a key factor in a wide range of technical applications, including spotted microarrays. The applied nL liquid volume provides specific reaction conditions for the immobilization of probe molecules to a chemically modified surface. Results We investigated the influence of nL and μL liquid drop volumes on the process of probe immobilization and compare the results obtained to the situation in liquid solution. In our data, we observe a strong relationship between drop drying effects on immobilization and surface chemistry. In this work, we present results on the immobilization of dye labeled 20mer oligonucleotides with and without an activating 5′-aminoheptyl linker onto a 2D epoxysilane and a 3D NHS activated hydrogel surface. Conclusions Our experiments identified two basic processes determining immobilization. First, the rate of drop drying that depends on the drop volume and the ambient relative humidity. Oligonucleotides in a dried spot react unspecifically with the surface and long reaction times are needed. 3D hydrogel surfaces allow for immobilization in a liquid environment under diffusive conditions. Here, oligonucleotide immobilization is much faster and a specific reaction with the reactive linker group is observed. Second, the effect of increasing probe concentration as a result of drop drying. On a 3D hydrogel, the increasing concentration of probe molecules in nL spotting volumes accelerates immobilization dramatically. In case of μL volumes, immobilization depends on whether the drop is allowed to dry completely. At non-drying conditions, very limited immobilization is observed due to the low oligonucleotide concentration used in microarray spotting solutions. The results of our study provide a general guideline for microarray assay development. They allow for the initial definition and further optimization of reaction conditions for the immobilization of oligonucleotides and other probe molecule classes to different

  5. Modulation of 5' splice site selection using tailed oligonucleotides carrying splicing signals

    Directory of Open Access Journals (Sweden)

    Elela Sherif

    2006-01-01

    Full Text Available Abstract Background We previously described the use of tailed oligonucleotides as a means of reprogramming alternative pre-mRNA splicing in vitro and in vivo. The tailed oligonucleotides that were used interfere with splicing because they contain a portion complementary to sequences immediately upstream of the target 5' splice site combined with a non-hybridizing 5' tail carrying binding sites for the hnRNP A1/A2 proteins. In the present study, we have tested the inhibitory activity of RNA oligonucleotides carrying different tail structures. Results We show that an oligonucleotide with a 5' tail containing the human β-globin branch site sequence inhibits the use of the 5' splice site of Bcl-xL, albeit less efficiently than a tail containing binding sites for the hnRNP A1/A2 proteins. A branch site-containing tail positioned at the 3' end of the oligonucleotide also elicited splicing inhibition but not as efficiently as a 5' tail. The interfering activity of a 3' tail was improved by adding a 5' splice site sequence next to the branch site sequence. A 3' tail carrying a Y-shaped branch structure promoted similar splicing interference. The inclusion of branch site or 5' splice site sequences in the Y-shaped 3' tail further improved splicing inhibition. Conclusion Our in vitro results indicate that a variety of tail architectures can be used to elicit splicing interference at low nanomolar concentrations, thereby broadening the scope and the potential impact of this antisense technology.

  6. Gas-phase Reactivity of meta-Benzyne Analogs Toward Small Oligonucleotides of Differing Lengths

    Science.gov (United States)

    Widjaja, Fanny; Max, Joann P.; Jin, Zhicheng; Nash, John J.; Kenttämaa, Hilkka I.

    2017-07-01

    The gas-phase reactivity of two aromatic carbon-centered σ,σ-biradicals ( meta-benzyne analogs) and a related monoradical towards small oligonucleotides of differing lengths was investigated in a Fourier-transform ion cyclotron resonance (FT-ICR) mass spectrometer coupled with laser-induced acoustic desorption (LIAD). The mono- and biradicals were positively charged to allow for manipulation in the mass spectrometer. The oligonucleotides were evaporated into the gas phase as intact neutral molecules by using LIAD. One of the biradicals was found to be unreactive. The reactive biradical reacts with dinucleoside phosphates and trinucleoside diphosphates mainly by addition to a nucleobase moiety followed by cleavage of the glycosidic bond, leading to a nucleobase radical (e.g., base-H) abstraction. In some instances, after the initial cleavage, the unquenched radical site of the biradical abstracts a hydrogen atom from the neutral fragment, which results in a net nucleobase abstraction. In sharp contrast, the related monoradical mainly undergoes facile hydrogen atom abstraction from the sugar moiety. As the size of the oligonucleotides increases, the rate of hydrogen atom abstraction from the sugar moiety by the monoradical was found to increase due to the presence of more hydrogen atom donor sites, and it is the only reaction observed for tetranucleoside triphosphates. Hence, the monoradical only attacks sugar moieties in these substrates. The biradical also shows significant attack at the sugar moiety for tetranucleoside triphosphates. This drastic change in reactivity indicates that the size of the oligonucleotides plays a key role in the outcome of these reactions. This finding is attributed to more compact conformations in the gas phase for the tetranucleoside triphosphates than for the smaller oligonucleotides, which result from stronger stabilizing interactions between the nucleobases.

  7. Coupling Strategies for the Synthesis of Peptide-Oligonucleotide Conjugates for Patterned Synthetic Biomineralization

    Directory of Open Access Journals (Sweden)

    Joshua D. Carter

    2011-01-01

    Full Text Available This work describes preparation strategies for peptide-oligonucleotide conjugates that combine the self-assembling behavior of DNA oligonucleotides with the molecular recognition capabilities of peptides. The syntheses include a solution-phase fragment coupling reaction and a solid-phase fragment coupling strategy where the oligonucleotide has been immobilized on DEAE Sepharose. The yield of four coupling reagents is evaluated, two reagents in water, EDC (1-ethyl-3-(3-dimethylaminopropyl carbodiimide hydrochloride and DMTMM (4-(4,6-dimethoxy[1,3,5]triazin-2-yl-4-methyl-morpholinium chloride, and two in dimethylformamide (DMF, PyBOP ((Benzotriazol-1-yloxy tripyrrolidinophosphonium hexafluorophosphate and HBTU (O-benzotriazole-N,N,N′,N′-tetramethyluronium hexafluorophosphate, while the oligonucleotide fragment is either in solution or immobilized on DEAE. These coupling strategies rely on an unprotected 5′ amino linker on the oligonucleotide reacting with the peptide C-terminus. The peptide, selected from a combinatorial library for its gold-binding behavior, was 12 amino acids long with an N-terminus acetyl cap. Formation of the conjugates was confirmed by gel electrophoresis and mass spectrometry while molecular recognition functionality of the peptide portion was verified using atomic force microscopy. Solution-phase yields were superior to their solid-phase counterparts. EDC resulted in the highest yield for both solution-phase (95% and solid-phase strategies (24%, while the DMF-based reagents, PyBOP and HBTU, resulted in low yields with reduced recovery. All recoverable conjugates demonstrated gold nanoparticle templating capability.

  8. Phage annealing proteins promote oligonucleotide-directed mutagenesis in Escherichia coli and mouse ES cells

    Directory of Open Access Journals (Sweden)

    Muyrers Joep PP

    2003-01-01

    Full Text Available Abstract Background The phage protein pairs, RecE/RecT from Rac or Redα/Redβ from λ, initiate efficient double strand break repair (DSBR in Escherichia coli that has proven very useful for DNA engineering. These phage pairs initiate DSBR either by annealing or by another mechanism that is not defined. Results Here we report that these proteins also mediate single strand oligonucleotide repair (ssOR at high efficiencies. The ssOR activity, unlike DSBR, does not require a phage exonuclease (RecE or Redα but only requires a phage annealing protein (RecT or Redβ. Notably, the P22 phage annealing protein Erf, which does not mediate the same DSBR reactions, also delivers ssOR activity. By altering aspects of the oligonucleotides, we document length and design parameters that affect ssOR efficiency to show a simple relationship to homologies either side of the repair site. Notably, ssOR shows strand bias. Oligonucleotides that can prime lagging strand replication deliver more ssOR than their leading complements. This suggests a model in which the annealing proteins hybridize the oligonucleotides to single stranded regions near the replication fork. We also show that ssOR is a highly efficient way to engineer BACs and can be detected in a eukaryotic cell upon expression of a phage annealing protein. Conclusion Phage annealing proteins can initiate the recombination of single stranded oligonucleotides into endogenous targets in Escherichia coli at very high efficiencies. This expands the repertoire of useful DNA engineering strategies, shows promise for applications in eukaryotic cells, and has implications for the unanswered questions regarding DSBR mediated by RecE/RecT and Redα/Redβ.

  9. Evaluating Hierarchical Structure in Music Annotations

    Directory of Open Access Journals (Sweden)

    Brian McFee

    2017-08-01

    Full Text Available Music exhibits structure at multiple scales, ranging from motifs to large-scale functional components. When inferring the structure of a piece, different listeners may attend to different temporal scales, which can result in disagreements when they describe the same piece. In the field of music informatics research (MIR, it is common to use corpora annotated with structural boundaries at different levels. By quantifying disagreements between multiple annotators, previous research has yielded several insights relevant to the study of music cognition. First, annotators tend to agree when structural boundaries are ambiguous. Second, this ambiguity seems to depend on musical features, time scale, and genre. Furthermore, it is possible to tune current annotation evaluation metrics to better align with these perceptual differences. However, previous work has not directly analyzed the effects of hierarchical structure because the existing methods for comparing structural annotations are designed for “flat” descriptions, and do not readily generalize to hierarchical annotations. In this paper, we extend and generalize previous work on the evaluation of hierarchical descriptions of musical structure. We derive an evaluation metric which can compare hierarchical annotations holistically across multiple levels. sing this metric, we investigate inter-annotator agreement on the multilevel annotations of two different music corpora, investigate the influence of acoustic properties on hierarchical annotations, and evaluate existing hierarchical segmentation algorithms against the distribution of inter-annotator agreement.

  10. Hierarchical screening for multiple mental disorders.

    Science.gov (United States)

    Batterham, Philip J; Calear, Alison L; Sunderland, Matthew; Carragher, Natacha; Christensen, Helen; Mackinnon, Andrew J

    2013-10-01

    There is a need for brief, accurate screening when assessing multiple mental disorders. Two-stage hierarchical screening, consisting of brief pre-screening followed by a battery of disorder-specific scales for those who meet diagnostic criteria, may increase the efficiency of screening without sacrificing precision. This study tested whether more efficient screening could be gained using two-stage hierarchical screening than by administering multiple separate tests. Two Australian adult samples (N=1990) with high rates of psychopathology were recruited using Facebook advertising to examine four methods of hierarchical screening for four mental disorders: major depressive disorder, generalised anxiety disorder, panic disorder and social phobia. Using K6 scores to determine whether full screening was required did not increase screening efficiency. However, pre-screening based on two decision tree approaches or item gating led to considerable reductions in the mean number of items presented per disorder screened, with estimated item reductions of up to 54%. The sensitivity of these hierarchical methods approached 100% relative to the full screening battery. Further testing of the hierarchical screening approach based on clinical criteria and in other samples is warranted. The results demonstrate that a two-phase hierarchical approach to screening multiple mental disorders leads to considerable increases efficiency gains without reducing accuracy. Screening programs should take advantage of prescreeners based on gating items or decision trees to reduce the burden on respondents. © 2013 Elsevier B.V. All rights reserved.

  11. Evaluating Hierarchical Structure in Music Annotations

    Science.gov (United States)

    McFee, Brian; Nieto, Oriol; Farbood, Morwaread M.; Bello, Juan Pablo

    2017-01-01

    Music exhibits structure at multiple scales, ranging from motifs to large-scale functional components. When inferring the structure of a piece, different listeners may attend to different temporal scales, which can result in disagreements when they describe the same piece. In the field of music informatics research (MIR), it is common to use corpora annotated with structural boundaries at different levels. By quantifying disagreements between multiple annotators, previous research has yielded several insights relevant to the study of music cognition. First, annotators tend to agree when structural boundaries are ambiguous. Second, this ambiguity seems to depend on musical features, time scale, and genre. Furthermore, it is possible to tune current annotation evaluation metrics to better align with these perceptual differences. However, previous work has not directly analyzed the effects of hierarchical structure because the existing methods for comparing structural annotations are designed for “flat” descriptions, and do not readily generalize to hierarchical annotations. In this paper, we extend and generalize previous work on the evaluation of hierarchical descriptions of musical structure. We derive an evaluation metric which can compare hierarchical annotations holistically across multiple levels. sing this metric, we investigate inter-annotator agreement on the multilevel annotations of two different music corpora, investigate the influence of acoustic properties on hierarchical annotations, and evaluate existing hierarchical segmentation algorithms against the distribution of inter-annotator agreement. PMID:28824514

  12. Evaluating Hierarchical Structure in Music Annotations.

    Science.gov (United States)

    McFee, Brian; Nieto, Oriol; Farbood, Morwaread M; Bello, Juan Pablo

    2017-01-01

    Music exhibits structure at multiple scales, ranging from motifs to large-scale functional components. When inferring the structure of a piece, different listeners may attend to different temporal scales, which can result in disagreements when they describe the same piece. In the field of music informatics research (MIR), it is common to use corpora annotated with structural boundaries at different levels. By quantifying disagreements between multiple annotators, previous research has yielded several insights relevant to the study of music cognition. First, annotators tend to agree when structural boundaries are ambiguous. Second, this ambiguity seems to depend on musical features, time scale, and genre. Furthermore, it is possible to tune current annotation evaluation metrics to better align with these perceptual differences. However, previous work has not directly analyzed the effects of hierarchical structure because the existing methods for comparing structural annotations are designed for "flat" descriptions, and do not readily generalize to hierarchical annotations. In this paper, we extend and generalize previous work on the evaluation of hierarchical descriptions of musical structure. We derive an evaluation metric which can compare hierarchical annotations holistically across multiple levels. sing this metric, we investigate inter-annotator agreement on the multilevel annotations of two different music corpora, investigate the influence of acoustic properties on hierarchical annotations, and evaluate existing hierarchical segmentation algorithms against the distribution of inter-annotator agreement.

  13. Hierarchical Nanoceramics for Industrial Process Sensors

    Energy Technology Data Exchange (ETDEWEB)

    Ruud, James, A.; Brosnan, Kristen, H.; Striker, Todd; Ramaswamy, Vidya; Aceto, Steven, C.; Gao, Yan; Willson, Patrick, D.; Manoharan, Mohan; Armstrong, Eric, N., Wachsman, Eric, D.; Kao, Chi-Chang

    2011-07-15

    This project developed a robust, tunable, hierarchical nanoceramics materials platform for industrial process sensors in harsh-environments. Control of material structure at multiple length scales from nano to macro increased the sensing response of the materials to combustion gases. These materials operated at relatively high temperatures, enabling detection close to the source of combustion. It is anticipated that these materials can form the basis for a new class of sensors enabling widespread use of efficient combustion processes with closed loop feedback control in the energy-intensive industries. The first phase of the project focused on materials selection and process development, leading to hierarchical nanoceramics that were evaluated for sensing performance. The second phase focused on optimizing the materials processes and microstructures, followed by validation of performance of a prototype sensor in a laboratory combustion environment. The objectives of this project were achieved by: (1) synthesizing and optimizing hierarchical nanostructures; (2) synthesizing and optimizing sensing nanomaterials; (3) integrating sensing functionality into hierarchical nanostructures; (4) demonstrating material performance in a sensing element; and (5) validating material performance in a simulated service environment. The project developed hierarchical nanoceramic electrodes for mixed potential zirconia gas sensors with increased surface area and demonstrated tailored electrocatalytic activity operable at high temperatures enabling detection of products of combustion such as NOx close to the source of combustion. Methods were developed for synthesis of hierarchical nanostructures with high, stable surface area, integrated catalytic functionality within the structures for gas sensing, and demonstrated materials performance in harsh lab and combustion gas environments.

  14. Binding of Dumbbell Oligonucleotides to MoMuLV Reverse Transcriptase: Inhibitory Properties of RNase H Activity

    Directory of Open Access Journals (Sweden)

    Ajay Kumar

    2010-01-01

    Full Text Available Dumbbell oligonucleotides with loops of various chemistry were synthesized. Incubation of dumbbell oligonucleotides containing phosphorothioate bonds or trimethylene phosphate linkages in loops with S1 nuclease did not result in significant cleavage under conditions which led to the degradation of dumbbell oligonucleotide containing phophodiester bonds in the loops. The binding of reverse transcriptase of Moloney Murine Leukemia Virus (MoMuLV was evaluated with all the five oligonucleotides. The protein binds to all the dumbbell oligonucleotides with similar affinity. The dissociation constants evaluated using PAGE band mobility shift assays were of the order of 10-7. The inhibitory properties of the retroviral RNase H activity was evaluated using 3H –UTP-labeled RNA:RNA-DNA hybrid. It was found that the best dumbbell oligonucleotide, inhibitor contained phosphorothioate residues in both the loops. Our value studies demonstrated that this particularly designed oligonucleotide displays an IC50 of 18 nM in its inhibition on the reverse transcriptase RNase H activity, a magnitude lower than that of first nucleotide reverse transcriptase of HIV-1, tenofovir, introduced by Gilead Science in the market.

  15. The Dynamics of Compound, Transcript, and Protein Effects After Treatment With 2OMePS Antisense Oligonucleotides in mdx Mice

    Directory of Open Access Journals (Sweden)

    Ingrid E C Verhaart

    2014-01-01

    Full Text Available Antisense-mediated exon skipping is currently in clinical development for Duchenne muscular dystrophy (DMD to amend the consequences of the underlying genetic defect and restore dystrophin expression. Due to turnover of compound, transcript, and protein, chronic treatment with effector molecules (antisense oligonucleotides will be required. To investigate the dynamics and persistence of antisense 2′-O-methyl phosphorothioate oligonucleotides, exon skipping, and dystrophin expression after dosing was concluded, mdx mice were treated subcutaneously for 8 weeks with 100 mg/kg oligonucleotides twice weekly. Thereafter, mice were sacrificed at different time points after the final injection (36 hours–24 weeks. Oligonucleotide half-life was longer in heart (~65 days compared with that in skeletal muscle, liver, and kidney (~35 days. Exon skipping half-lives varied between 33 and 53 days, whereas dystrophin protein showed a long half-life (>100 days. Oligonucleotide and exon-skipping levels peaked in the first week and declined thereafter. By contrast, dystrophin expression peaked after 3–8 weeks and then slowly declined, remaining detectable after 24 weeks. Concordance between levels of oligonucleotides, exon skipping, and proteins was observed, except in heart, wherein high oligonucleotide levels but low exon skipping and dystrophin expression were seen. Overall, these results enhance our understanding of the pharmacokinetics and pharmacodynamics of 2′-O-methyl phosphorothioate oligos used for the treatment of DMD.

  16. Primers on molecular pathways--caspase pathway.

    Science.gov (United States)

    Lomberk, Gwen; Urrutia, Raul

    2009-01-01

    Apoptosis, or programmed cell death, is a physiological process of cellular autodestruction, or cell suicide. This process is strictly controlled in response to integrity of pro-death signaling and plays critical roles in development, maintenance of homeostasis and host defense in multicellular organisms. As pancreatologists, apoptosis plays a central role in the pancreas and its disease states, from diabetes to pancreatitis to pancreatic cancer. In pancreatic beta-cells, apoptotic cell death is involved in the pathogenesis of diabetes, as signals from death receptors and DNA damage have been widely accepted as being triggers of apoptosis in beta-cells. During acute pancreatitis, this common clinical condition is of variable severity in which some patients experience mild, self-limited attacks while others manifest a severe, highly morbid, and frequently lethal attack. However, recent research in this area has demonstrated the importance of acinar cell death in the form of apoptosis and necrosis as a determinant of pancreatitis severity. In pancreatic cancer, various survival mechanisms have been shown to act in the prevention of cell death to result in promotion of tumor growth and metastasis. Thus, resistance of pancreatic cancer to apoptosis is the key factor preventing responses to therapies. Thus, it is for these reasons that in the current 'Primers on Molecular Pathways,' we take a closer look at the pathway cascade that is triggered during apoptosis. Copyright 2008 S. Karger AG, Basel and IAP.

  17. Primer on electricity futures and other derivatives

    Energy Technology Data Exchange (ETDEWEB)

    Stoft, S.; Belden, T.; Goldman, C.; Pickle, S.

    1998-01-01

    Increased competition in bulk power and retail electricity markets is likely to lower electricity prices, but will also result in greater price volatility as the industry moves away from administratively determined, cost-based rates and encourages market-driven prices. Price volatility introduces new risks for generators, consumers, and marketers. Electricity futures and other derivatives can help each of these market participants manage, or hedge, price risks in a competitive electricity market. Futures contracts are legally binding and negotiable contracts that call for the future delivery of a commodity. In most cases, physical delivery does not take place, and the futures contract is closed by buying or selling a futures contract on or near the delivery date. Other electric rate derivatives include options, price swaps, basis swaps, and forward contracts. This report is intended as a primer for public utility commissioners and their staff on futures and other financial instruments used to manage price risks. The report also explores some of the difficult choices facing regulators as they attempt to develop policies in this area.

  18. Transforming practices: a primer on action research.

    Science.gov (United States)

    Acosta, Sandra; Goltz, Heather Honoré

    2014-07-01

    Action research (AR) is a powerful tool for health education and promotion practitioners who want to focus on improving the quality of their programs and services. In this Tool, we describe the characteristics and controversial aspects of AR, differentiate between traditional and action research, present the benefits of applying AR methods/techniques for investigating problems related to professional practice, and offer a four-phase methodological framework for conducting AR studies. Unlike traditional research, AR is a methodology that links theory, research, and practice; advances new knowledge and understandings via iterative action cycles; employs frontline health practitioners as researchers; and promotes collaborative practitioner-community partnerships. Egalitarian in its approach, AR offers an "insider's perspective" centered on context-specific problems and issues related to health promotion. AR falls into two categories: large-scale community-based research and small-scale practice-based research. Each cycle of the AR framework includes four phases: (a) preplanning/needs assessment, (b) planning/study organization, (c) action and observation/study implementation, and (d) reflection and planning/data analysis and interpretation. Using the AR primer in this Tool has the potential to empower health education/promotion practitioners, encourage collaborative partnerships, enhance practitioners' knowledge base, and promote social change. © 2014 Society for Public Health Education.

  19. Value for money assessment for public-private partnerships : a primer.

    Science.gov (United States)

    2015-01-01

    This primer addresses Value for Money Assessment for public-private partnerships (P3s). Companion : primers on Financial Assessment and Risk Assessment for P3s are also available as part of this series of : primers.

  20. Complementation of a primer binding site-impaired murine leukemia virus-derived retroviral vector by a genetically engineered tRNA-like primer

    DEFF Research Database (Denmark)

    Lund, Anders Henrik; Duch, M; Lovmand, J

    1997-01-01

    , but not with a noncomplementary tRNA-like molecule. The engineered primer was shown to be involved in both the initiation of first-strand synthesis and second-strand transfer. These results provide an in vivo demonstration that the retroviral replication machinery may recognize sequence complementarity rather than actual primer...... binding site and 3' primer sequences. Use of mutated primer binding site vectors replicating via engineered primers may add additional control features to retroviral gene transfer technology....

  1. Synthesis, Dynamic Combinatorial Chemistry, and PCR Amplification of 3'-5' and 3'-6' Disulfide-linked Oligonucleotides

    DEFF Research Database (Denmark)

    Hansen, Dennis Jul; Manuguerra, Ilenia; Kjelstrup, Michael Brøndum

    2014-01-01

    Disulfide dithymidines linked 3'-5' or 3'-6' were synthesized and incorporated into oligonucleotides through a combined phosphotriester and phosphoramidite solid-phase oligonucleotide synthesis approach. The disulfide links are cleaved and formed reversibly in the presence of thiols...... and oligonucleotides. This link was shown to be sequence-adaptive in response to given templates in the presence of mercaptoethanol. The artificial 3'-5' and 3'-6' disulfide link was tolerated by polymerases in the polymerase chain reaction (PCR). By using sequencing analysis, we show that single mutations frequently...

  2. Amino acids attached to 2'-amino-LNA: Synthesis of DNA mixmer oligonucleotides with increased duplex stability

    DEFF Research Database (Denmark)

    Johannsen, Marie Willaing; Wengel, Jesper; Wamberg, Michael Chr.

    2010-01-01

    The synthesis of 2'-amino-LNA (locked nucleic acid) opens up exciting possibilities for modification of nucleic acids by conjugation to the 2'-nitrogen. Incorporation of unmodified and N-functionalized 2'-amino-LNA nucleotides improve duplex stability compared to unmodified DNA. 2'-Amino......-LNA nucleosides derivatized with amino acids have been synthesized and incorporated into DNA oligonucleotides. Following oligonucleotide synthesis, peptides have been added using solid phase peptide coupling chem. Modification of oligonucleotides with pos. charged residues greatly improves thermal stability....

  3. Metabolic systems biology: a brief primer.

    Science.gov (United States)

    Edwards, Lindsay M

    2017-05-01

    In the early to mid-20th century, reductionism as a concept in biology was challenged by key thinkers, including Ludwig von Bertalanffy. He proposed that living organisms were specific examples of complex systems and, as such, they should display characteristics including hierarchical organisation and emergent behaviour. Yet the true study of complete biological systems (for example, metabolism) was not possible until technological advances that occurred 60 years later. Technology now exists that permits the measurement of complete levels of the biological hierarchy, for example the genome and transcriptome. The complexity and scale of these data require computational models for their interpretation. The combination of these - systems thinking, high-dimensional data and computation - defines systems biology, typically accompanied by some notion of iterative model refinement. Only sequencing-based technologies, however, offer full coverage. Other 'omics' platforms trade coverage for sensitivity, although the densely connected nature of biological networks suggests that full coverage may not be necessary. Systems biology models are often characterised as either 'bottom-up' (mechanistic) or 'top-down' (statistical). This distinction can mislead, as all models rely on data and all are, to some degree, 'middle-out'. Systems biology has matured as a discipline, and its methods are commonplace in many laboratories. However, many challenges remain, especially those related to large-scale data integration. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

  4. cutPrimers: A New Tool for Accurate Cutting of Primers from Reads of Targeted Next Generation Sequencing.

    Science.gov (United States)

    Kechin, Andrey; Boyarskikh, Uljana; Kel, Alexander; Filipenko, Maxim

    2017-11-01

    Cutting of primers from reads is an important step of processing targeted amplicon-based next generation sequencing data. Existing tools are adapted for cutting of one or several primer/adapter sequences from reads and removing all of their occurrences. Also most of the existing tools use kmers and may cut only part of primers or primers with studied sequence of gene. Because of this, use of such programs leads to incorrect trimming, reduction of coverage, and increase in the number of false-positive and/or false-negative results. We have developed a new tool named cutPrimers for accurate cutting of any number of primers from reads. Using sequencing reads that were obtained during study of BRCA1/2 genes, we compared it with cutadapt, AlienTrimmer, and BBDuk. All of them trimmed reads in such a way that coverage of at least two amplicons decreased to unacceptable level (cut all occurrences of primer sequences, so the length of the remaining reads was less than prospective.

  5. Iodine-125-labeled DNA-Triplex-forming oligonucleotides reveal increased cyto- and genotoxic effectiveness compared to Phosphorus-32.

    Science.gov (United States)

    Dahmen, Volker; Pomplun, Ekkehard; Kriehuber, Ralf

    2016-11-01

    The efficacy of DNA-targeting radionuclide therapies might be strongly enhanced by employing short range particle-emitters. However, the gain of effectiveness is not yet well substantiated. We compared the Auger electron emitter I-125 to the ß(-)-emitter P-32 in terms of biological effectiveness per decay and radiation dose when located in the close proximity to DNA using DNA Triplex-forming oligonucleotides (TFO). The clonogenicity and the induction of DNA double-strand breaks (DSB) were investigated in SCL-II cells after exposure to P-32- or I-125-labeled TFO targeting the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene and after external homogeneous exposure to gamma-rays as reference radiation. TFO were labeled with P-32 or I-125 using the primer extension method. Cell survival was analyzed by colony-forming assay and DNA damage was assessed by microscopic quantification of protein 53 binding protein 1 (53BP1) foci in SCL-II cells. I-125-TFO induced a pronounced decrease of cell survival (D37 at ∼360 accumulated decays per cell, equivalent to 1.22 Gy cell nucleus dose) and a significant increase of 53BP1 foci with increasing decays. The P-32-labeled TFO induced neither a strong decrease of cell survival nor an increase of 53BP1 foci up to ∼4000 accumulated decays per cell, equivalent to ∼1 Gy cell nucleus dose. The RBE for I-125-TFO was in the range of 3-4 for both biological endpoints. I-125-TFO proved to be much more radiotoxic than P-32-TFO per decay and per unit dose although targeting the same sequence in the GAPDH gene. This might be well explained by the high number of low energy Auger electrons emitted by I-125 per decay, leading to a high ionization density in the immediate vicinity of the decay site, probably producing highly complex DNA lesions overcharging DNA repair mechanisms.

  6. A Psicologia Latinoamericana: El Primer Medio Siglo

    Directory of Open Access Journals (Sweden)

    Rubén Ardila

    2004-01-01

    Full Text Available Se presentan los principales desarrollos de la psicología en América Latina, desde el contexto de la psicología interamericana. La Sociedad Interamericana de Psicología (SIP fue fundada el 17 de diciembre de 1951 en Ciudad de México. El primer Congreso Interamericano de Psicología se llevó a cabo en Santo Domingo (República Dominicana entre el 10 y el 20 de diciembre de 1953. Este medio siglo ha marcado un acelerado proceso de desarrollo en la psicología en América Latina, en todo el continente americano, y en el mundo en general. Se han creado programas de formación profesional a lo largo y ancho del continente, y podemos afirmar que hoy existen psicólogos profesionales en todos los países de las Américas El papel del psicólogo en la sociedad ha logrado tener importancia y relevancia habiendo partido de comienzos muy modestos. Se está llevando a cabo investigación científica de alto nivel, especialmente en algunos países líderes, como son México y Brasil, pero también en Perú, Venezuela, Argentina, Cuba, Chile, Colombia, y en general en América Latina. La falta de originalidad que señalaba Alarcón en su libro Orientaciones Teóricas de la Psicología en América Latina, en 1997, ha comenzado a dar paso a trabajos originales y a investigaciones pioneras. Los siguientes 50 años de la psicología en este contexto cultural probablemente van a continuar con estos desarrollos, y seguramente llevarán a integrar la psicología latinoamericana al contexto internacional

  7. propósito del primer centenario

    Directory of Open Access Journals (Sweden)

    René Martínez Lemoine

    2007-01-01

    Full Text Available Este trabajo, trata de las apreciaciones públicas y urbanas que se dieron en la ciudad de Santiago a instancias de la celebración del primer Centenario de la República, en el año 1910. Es una rememoranza de aquellas visiones que la ciudad sustrajo a los medios periodísticos, personajes y liderazgos de una sociedad autosatisfecha por el progreso notable alcanzado por el país en las últimas décadas del siglo XIX y los primeros años del siglo XX, pero que ignoró los movimientos obreros y expresiones de disconformidad social que ya comenzaban a ebullir en esos años. La ciudad, como una vasta, compleja y heterogénea construcción en el espacio, erigida a través de las edades por innumerables y, la más de las veces, anónimos constructores, representa la mayor suma de obra humana acumulada en el tiempo, en la que cada generación va dejando una muestra de su aporte en vivienda, espacios, instalaciones y monumentos, vale decir, de su particular cultura y modo de vida en su propio tiempo. Ciertamente, cada ciudad es historia y memoria de sí misma, testimonio permanente de la continuidad del hombre y de la sociedad humana con su propio pasado. En ese sentido, como somos herederos de nuestra historia y de los hombres y mujeres que construyeron y legaron las ciudades en las que vivimos, es importante rescatar esos valores culturales, sociales, arquitectónicos y urbanísticos de modo de visualizar el paso del tiempo, que se materializa, se hace objeto y se torna visible en la ciudad, en la medida que nos “cuenta” algo.

  8. Self-assembled biomimetic superhydrophobic hierarchical arrays.

    Science.gov (United States)

    Yang, Hongta; Dou, Xuan; Fang, Yin; Jiang, Peng

    2013-09-01

    Here, we report a simple and inexpensive bottom-up technology for fabricating superhydrophobic coatings with hierarchical micro-/nano-structures, which are inspired by the binary periodic structure found on the superhydrophobic compound eyes of some insects (e.g., mosquitoes and moths). Binary colloidal arrays consisting of exemplary large (4 and 30 μm) and small (300 nm) silica spheres are first assembled by a scalable Langmuir-Blodgett (LB) technology in a layer-by-layer manner. After surface modification with fluorosilanes, the self-assembled hierarchical particle arrays become superhydrophobic with an apparent water contact angle (CA) larger than 150°. The throughput of the resulting superhydrophobic coatings with hierarchical structures can be significantly improved by templating the binary periodic structures of the LB-assembled colloidal arrays into UV-curable fluoropolymers by a soft lithography approach. Superhydrophobic perfluoroether acrylate hierarchical arrays with large CAs and small CA hysteresis can be faithfully replicated onto various substrates. Both experiments and theoretical calculations based on the Cassie's dewetting model demonstrate the importance of the hierarchical structure in achieving the final superhydrophobic surface states. Copyright © 2013 Elsevier Inc. All rights reserved.

  9. Design factors that influence PCR amplification success of cross-species primers among 1147 mammalian primer pairs

    Directory of Open Access Journals (Sweden)

    Beckett Stephanie E

    2006-10-01

    Full Text Available Abstract Background Cross-species primers have been used with moderate success to address a variety of questions concerning genome structure, evolution, and gene function. However, the factors affecting their success have never been adequately addressed, particularly with respect to producing a consistent method to achieve high throughput. Using 1,147 mammalian cross-species primer pairs (1089 not previously reported, we tested several factors to determine their influence on the probability that a given target will amplify in a given species under a single amplification condition. These factors included: number of mismatches between the two species (the index species used to identify conserved regions to which the primers were designed, GC-content of the gene and amplified region, CpG dinucleotides in the primer region, degree of encoded protein conservation, length of the primers, and the degree of evolutionary distance between the target species and the two index species. Results The amplification success rate for the cross-species primers was significantly influenced by the number of mismatches between the two index species (6–8% decrease per mismatch in a primer pair, the GC-content within the amplified region (for the dog, GC ≥ 50%, 56.9% amplified; GC2 = 0.14 and the relatedness of the target species to the index species. For the dog, 598 products of 930 primer pairs (64.3% (excluding primers in which dog was an index species were sequenced and shown to be the expected product, with an additional three percent producing the incorrect sequence. When hamster DNA was used with the single amplification condition in a microtiter plate-based format, 510 of 1087 primer pairs (46.9% produced amplified products. The primer pairs are spaced at an average distance of 2.3 Mb in the human genome and may be used to produce up to several hundred thousand bp of species-specific sequence. Conclusion The most important factors influencing the proportion

  10. New primers for DNA barcoding of digeneans and cestodes (Platyhelminthes).

    Science.gov (United States)

    Van Steenkiste, Niels; Locke, Sean A; Castelin, Magalie; Marcogliese, David J; Abbott, Cathryn L

    2015-07-01

    Digeneans and cestodes are species-rich taxa and can seriously impact human health, fisheries, aqua- and agriculture, and wildlife conservation and management. DNA barcoding using the COI Folmer region could be applied for species detection and identification, but both 'universal' and taxon-specific COI primers fail to amplify in many flatworm taxa. We found that high levels of nucleotide variation at priming sites made it unrealistic to design primers targeting all flatworms. We developed new degenerate primers that enabled acquisition of the COI barcode region from 100% of specimens tested (n = 46), representing 23 families of digeneans and 6 orders of cestodes. This high success rate represents an improvement over existing methods. Primers and methods provided here are critical pieces towards redressing the current paucity of COI barcodes for these taxa in public databases. © 2014 Her Majesty the Queen in Right of Canada Molecular Ecology Resources © 2014 John Wiley & Sons Ltd.

  11. Preparation and Characterization of Acrylic Primer for Concrete Substrate Application

    Directory of Open Access Journals (Sweden)

    El-Sayed Negim

    2016-01-01

    Full Text Available This study dealt with the properties of acrylic primer for concrete substrate using acrylic syrup, made from a methyl methacrylate monomer solution of terpolymers. Terpolymer systems consisting of methyl methacrylate (MMA, 2-ethylhexyl acrylate (2-EHA, and methacrylic acid (MAA with different chemical composition ratios of MMA and 2-EHA were synthesized through bulk polymerization using azobisisobutyronitrile (AIBN as initiator. The terpolymer composition is characterized by FTIR, 1H NMR, DSC, TGA, and SEM. The glass transition temperature and the thermal stability increased with increasing amounts of MMA in the terpolymer backbone. The effect of chemical composition of terpolymers on physicomechanical properties of primer films was investigated. However, increasing the amount of MMA in terpolymer backbone increased tensile and contact angle of primer films while elongation at break, water absorption, and bond strength are decreased. In particular, the primer syrup containing 65% 2-EHA has good bonding strength with concrete substrate around 1.1 MPa.

  12. USER-derived cloning methods and their primer design.

    Science.gov (United States)

    Salomonsen, Bo; Mortensen, Uffe H; Halkier, Barbara A

    2014-01-01

    Uracil excision-based cloning through USER™ (Uracil-Specific Excision Reagent) is an efficient ligase-free cloning technique that comprises USER cloning, USER fusion, and USER cassette-free (UCF) USER fusion. These USER-derived cloning techniques enable seamless assembly of multiple DNA fragments in one construct. Though governed by a few simple rules primer design for USER-based fusion of PCR fragments can prove time-consuming for inexperienced users. The Primer Help for USER (PHUSER) software is an easy-to-use primer design tool for USER-based methods. In this chapter, we present a PHUSER software protocol for designing primers for USER-derived cloning techniques.

  13. Best Development Practices: A Primer for Smart Growth

    Science.gov (United States)

    Best Development Practices: A Primer for Smart Growth lists specific practices to achieve development principles that mix land uses, support transportation options, protect natural systems, and provide housing choices.

  14. PrecisePrimer: an easy-to-use web server for designing PCR primers for DNA library cloning and DNA shuffling

    Science.gov (United States)

    Pauthenier, Cyrille; Faulon, Jean-Loup

    2014-01-01

    PrecisePrimer is a web-based primer design software made to assist experimentalists in any repetitive primer design task such as preparing, cloning and shuffling DNA libraries. Unlike other popular primer design tools, it is conceived to generate primer libraries with popular PCR polymerase buffers proposed as pre-set options. PrecisePrimer is also meant to design primers in batches, such as for DNA libraries creation of DNA shuffling experiments and to have the simplest interface possible. It integrates the most up-to-date melting temperature algorithms validated with experimental data, and cross validated with other computational tools. We generated a library of primers for the extraction and cloning of 61 genes from yeast DNA genomic extract using default parameters. All primer pairs efficiently amplified their target without any optimization of the PCR conditions. PMID:24829457

  15. Hierarchical Micro-Nano Coatings by Painting

    Science.gov (United States)

    Kirveslahti, Anna; Korhonen, Tuulia; Suvanto, Mika; Pakkanen, Tapani A.

    2016-03-01

    In this paper, the wettability properties of coatings with hierarchical surface structures and low surface energy were studied. Hierarchically structured coatings were produced by using hydrophobic fumed silica nanoparticles and polytetrafluoroethylene (PTFE) microparticles as additives in polyester (PES) and polyvinyldifluoride (PVDF). These particles created hierarchical micro-nano structures on the paint surfaces and lowered or supported the already low surface energy of the paint. Two standard application techniques for paint application were employed and the presented coatings are suitable for mass production and use in large surface areas. By regulating the particle concentrations, it was possible to modify wettability properties gradually. Highly hydrophobic surfaces were achieved with the highest contact angle of 165∘. Dynamic contact angle measurements were carried out for a set of selected samples and low hysteresis was obtained. Produced coatings possessed long lasting durability in the air and in underwater conditions.

  16. [Comparative hierarchic structure of the genetic language].

    Science.gov (United States)

    Ratner, V A

    1993-05-01

    The genetical texts and genetic language are built according to hierarchic principle and contain no less than 6 levels of coding sequences, separated by marks of punctuation, separation and indication: codons, cistrons, scriptons, replicons, linkage groups, genomes. Each level has all the attributes of the language. This hierarchic system expresses some general properties and regularities. The rules of genetic language being determined, the variability of genetical texts is generated by block-modular combinatorics on each level. Between levels there are some intermediate sublevels and module types capable of being combined. The genetic language is compared with two different independent linguistic systems: human natural languages and artificial programming languages. Genetic language is a natural one by its origin, but it is a typical technical language of the functioning genetic regulatory system--by its predestination. All three linguistic systems under comparison have evident similarity of the organization principles and hierarchical structures. This argues for similarity of their principles of appearance and evolution.

  17. Towards a sustainable manufacture of hierarchical zeolites.

    Science.gov (United States)

    Verboekend, Danny; Pérez-Ramírez, Javier

    2014-03-01

    Hierarchical zeolites have been established as a superior type of aluminosilicate catalysts compared to their conventional (purely microporous) counterparts. An impressive array of bottom-up and top-down approaches has been developed during the last decade to design and subsequently exploit these exciting materials catalytically. However, the sustainability of the developed synthetic methods has rarely been addressed. This paper highlights important criteria to ensure the ecological and economic viability of the manufacture of hierarchical zeolites. Moreover, by using base leaching as a promising case study, we verify a variety of approaches to increase reactor productivity, recycle waste streams, prevent the combustion of organic compounds, and minimize separation efforts. By reducing their synthetic footprint, hierarchical zeolites are positioned as an integral part of sustainable chemistry. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Hierarchical Neural Network Structures for Phoneme Recognition

    CERN Document Server

    Vasquez, Daniel; Minker, Wolfgang

    2013-01-01

    In this book, hierarchical structures based on neural networks are investigated for automatic speech recognition. These structures are evaluated on the phoneme recognition task where a  Hybrid Hidden Markov Model/Artificial Neural Network paradigm is used. The baseline hierarchical scheme consists of two levels each which is based on a Multilayered Perceptron. Additionally, the output of the first level serves as a second level input. The computational speed of the phoneme recognizer can be substantially increased by removing redundant information still contained at the first level output. Several techniques based on temporal and phonetic criteria have been investigated to remove this redundant information. The computational time could be reduced by 57% whilst keeping the system accuracy comparable to the baseline hierarchical approach.

  19. Ultrahigh molecular recognition specificity of competing DNA oligonucleotide strands in thermal equilibrium

    CERN Document Server

    Schenkelberger, Marc; Mai, Timo; Ott, Albrecht

    2016-01-01

    The specificity of molecular recognition is important to molecular self-organization. A prominent example is the biological cell where, within a highly crowded molecular environment, a myriad of different molecular receptor pairs recognize their binding partner with astonishing accuracy. In thermal equilibrium it is usually admitted that the affinity of recognizer pairs only depends on the nature of the two binding molecules. Accordingly, Boltzmann factors of binding energy differences relate the molecular affinities among different target molecules that compete for the same probe. Here, we consider the molecular recognition of short DNA oligonucleotide single strands. We show that a better matching oligonucleotide strand can prevail against a disproportionally more concentrated competitor that exhibits reduced affinity due to a mismatch. The magnitude of deviation from the simple picture above may reach several orders of magnitude. In our experiments the effective molecular affinity of a given strand remains...

  20. Thermal Stability of Modified i-Motif Oligonucleotides with Naphthalimide Intercalating Nucleic Acids

    DEFF Research Database (Denmark)

    El-Sayed, Ahmed Ali; Pedersen, Erik B.; Khaireldin, Nahid Y.

    2016-01-01

    of naphthalimide (1H-benzo[de]isoquinoline-1,3(2H)-dione) as the intercalating nucleic acid. The stabilities of i-motif structures with inserted naphthalimide intercalating nucleotides were studied using UV melting temperatures (Tm) and circular dichroism spectra at different pH values and conditions (crowding......In continuation of our investigation of characteristics and thermodynamic properties of the i-motif 5′-d[(CCCTAA)3CCCT)] upon insertion of intercalating nucleotides into the cytosine-rich oligonucleotide, this article evaluates the stabilities of i-motif oligonucleotides upon insertion...... and non-crowding). This study indicated a positive effect of the naphthalimide intercalating nucleotides on the stabilities of the i-motif structures compared to the wild-type structure which is in contrast to a previous observation for a pyrene-intercalating nucleotide showing a decrease in Tm values....

  1. Electrochemical study of ellipticine interaction with single and double stranded oligonucleotides.

    Science.gov (United States)

    Tmejova, Katerina; Krejcova, Ludmila; Hynek, David; Adam, Vojtech; Babula, Petr; Trnkova, Libuse; Stiborova, Marie; Eckschlager, Tomas; Kizek, Rene

    2014-02-01

    Ellipticine (5,11-dimethyl-6H-pyrido[4,3-b]carbazole) is an alkaloid that has been isolated from plants of an Apocynaceae family. It is one of the simplest naturally occurring alkaloids with a planar structure. Over the past decades, ellipticine became a very promising antitumor agent. Interaction with DNA is one of the most studied ellipticine effects on cell division. This phenomenon is not clearly explained so far. In our experiments we studied interaction of ellipticine with single-stranded and double-stranded oligonucleotides by electrochemical methods on mercury electrode. Differential pulse voltammetry was applied for ellipticine (Elli) and CA peak detection. Square wave voltammetry was applied for G peak detection. The effect of the interaction time and ellipticine concentrations on interactions of ellipticine with single- and double-stranded oligonucleotides was tested too.

  2. Recognition and sensing of low-epitope targets via ternary complexes with oligonucleotides and synthetic receptors

    Science.gov (United States)

    Yang, Kyung-Ae; Barbu, Mihaela; Halim, Marlin; Pallavi, Payal; Kim, Benjamin; Kolpashchikov, Dmitry M.; Pecic, Stevan; Taylor, Steven; Worgall, Tilla S.; Stojanovic, Milan N.

    2014-11-01

    Oligonucleotide-based receptors or aptamers can interact with small molecules, but the ability to achieve high-affinity and specificity of these interactions depends strongly on functional groups or epitopes displayed by the binding targets. Some classes of targets are particularly challenging: for example, monosaccharides have scarce functionalities and no aptamers have been reported to recognize, let alone distinguish from each other, glucose and other hexoses. Here we report aptamers that differentiate low-epitope targets such as glucose, fructose or galactose by forming ternary complexes with high-epitope organic receptors for monosaccharides. In a follow-up example, we expand this method to isolate high-affinity oligonucleotides against aromatic amino acids complexed in situ with a nonspecific organometallic receptor. The method is general and enables broad clinical use of aptamers for the detection of small molecules in mix-and-measure assays, as demonstrated by monitoring postprandial waves of phenylalanine in human subjects.

  3. Heated oligonucleotide ligation assay (HOLA): an affordable single nucleotide polymorphism assay.

    Science.gov (United States)

    Black, W C; Gorrochotegui-Escalante, N; Duteau, N M

    2006-03-01

    Most single nucleotide polymorphism (SNP) detection requires expensive equipment and reagents. The oligonucleotide ligation assay (OLA) is an inexpensive SNP assay that detects ligation between a biotinylated "allele-specific detector" and a 3' fluorescein-labeled "reporter" oligonucleotide. No ligation occurs unless the 3' detector nucleotide is complementary to the SNP nucleotide. The original OLA used chemical denaturation and neutralization. Heated OLA (HOLA) instead uses a thermal stable ligase and cycles of denaturing and hybridization for ligation and SNP detection. The cost per genotype is approximately US$1.25 with two-allele SNPs or approximately US$1.75 with three-allele SNPs. We illustrate the development of HOLA for SNP detection in the Early Trypsin and Abundant Trypsin loci in the mosquito Aedes aegypti (L.) and at the a-glycerophosphate dehydrogenase locus in the mosquito Anopheles gambiae s.s.

  4. LNA-modified oligonucleotides mediate specific inhibition of microRNA function

    DEFF Research Database (Denmark)

    Ørom, Ulf Andersson; Kauppinen, Sakari; Lund, Anders H

    2006-01-01

    observed in many human cancers. Here we present a method for specific inhibition of miRNA function through interaction with LNA-modified antisense oligonucleotides and report the specificity of this application. We show that LNA-modified oligonucleotides can inhibit exogenously introduced miRNAs with high......microRNAs are short, endogenous non-coding RNAs that act as post-transcriptional modulators of gene expression. Important functions for microRNAs have been found in the regulation of development, cellular proliferation and differentiation, while perturbed miRNA expression patterns have been...... specificity using a heterologous reporter assay, and furthermore demonstrate their ability to inhibit an endogenous miRNA in Drosophila melanogaster cells, leading to up-regulation of the cognate target protein. The method shows stoichiometric and reliable inhibition of the targeted miRNA and can thus...

  5. Single-Labeled Oligonucleotides Showing Fluorescence Changes upon Hybridization with Target Nucleic Acids

    Directory of Open Access Journals (Sweden)

    Gil Tae Hwang

    2018-01-01

    Full Text Available Sequence-specific detection of nucleic acids has been intensively studied in the field of molecular diagnostics. In particular, the detection and analysis of single-nucleotide polymorphisms (SNPs is crucial for the identification of disease-causing genes and diagnosis of diseases. Sequence-specific hybridization probes, such as molecular beacons bearing the fluorophore and quencher at both ends of the stem, have been developed to enable DNA mutation detection. Interestingly, DNA mutations can be detected using fluorescently labeled oligonucleotide probes with only one fluorophore. This review summarizes recent research on single-labeled oligonucleotide probes that exhibit fluorescence changes after encountering target nucleic acids, such as guanine-quenching probes, cyanine-containing probes, probes containing a fluorophore-labeled base, and microenvironment-sensitive probes.

  6. Development and characterization of microsatellite primers in Pogostemon cablin (Lamiaceae).

    Science.gov (United States)

    Sandes, S S; Pinheiro, J B; Zucchi, M I; Monteiro, M; Arrigoni-Blank, M F; Blank, A F

    2013-08-08

    Microsatellite primers were developed and optimized for patchouli (Pogostemon cablin) to characterize the patchouli Active Germplasm Bank of Universidade Federal de Sergipe. Creation of a genomic library for patchouli enabled the design of 12 microsatellite primers. Six of these microsatellites were polymorphic, revealing two well-defined groups of individuals that possess exclusive alleles. The data allowed us to characterize the patchouli active Germplasm Bank, identify its genetic diversity, and provide new information for researching this species.

  7. Karakterisasi Unjuk Kerja Pompa Sistem Pendingin Primer Reaktor Triga 2000

    OpenAIRE

    Soekodijat, B

    2002-01-01

    KARAKTERISASI UNJUK KERJA POMPA SISTEM PENDINGIN PRIMER REAKTOR TRIGA 2000. Pompa sistem pendingin primer adalah salah satu komponen utama pada sistem pendingin Reaktor TRIGA 2000. Karakteristik unjuk kerja pompa sangat penting diketahui agar pompa dapat dioperasikan dengan baik dan aman. Penelitian dilakukan dengan cara memverifikasi kurva kinerja pompa yang diberikan oleh pabrik. Proses verifikasi dilakukan dengan mengubah-ubah laju alir pompa (dengan cara mengatur besar kecilnya pembukaan ...

  8. Chemical Etiology of Nucleic Acid Structure: The α-Threofuranosyl-(3′→2′) Oligonucleotide System

    National Research Council Canada - National Science Library

    K.-U. Schöning; P. Scholz; S. Guntha; X. Wu; R. Krishnamurthy; A. Eschenmoser

    2000-01-01

    TNAs [(L)-α-threofuranosyl oligonucleotides] containing vicinally connected (3′→2′) phosphodiester bridges undergo informational base pairing in antiparallel strand orientation and are capable of cross-pairing with RNA and DNA...

  9. Crystallization of a member of the recFOR DNA repair pathway, RecO, with and without bound oligonucleotide

    Energy Technology Data Exchange (ETDEWEB)

    Aono, Shelly; Hartsch, Thomas; Schulze-Gahmen, Ursula

    2003-01-22

    RecFOR proteins are important for DNA repair by homologous recombination in bacteria. The RecO protein from Thermus thermophilus was cloned, purified and characterized for its binding to oligonucleotides. The protein was crystallized alone and in complex with a 14-mer oligonucleotide. Both crystal forms grow under different crystallization conditions in the same space group, P3121 or P3221, with almost identical unit cell parameters. Complete data sets were collected to 2.8 Angstrom and 2.5 Angstrom for RecO alone and the RecO-oligonucleotide complex, respectively. Visual comparison of the diffraction patterns between the two crystal forms and calculation of an Rmerge of 33.9 percent on F indicate that one of the crystal forms is indeed a complex of RecO with bound oligonucleotide.

  10. Direct profiling of environmental microbial populations by thermal dissociation analysis of native rRNAs hybridized to oligonucleotide microarrays

    NARCIS (Netherlands)

    Fantroussi, El S.; Urakawa, H.; Bernhard, A.E.; Kelly, J.J.; Noble, P.A.; Smidt, H.; Yershov, G.M.; Stahl, D.A.

    2003-01-01

    Oligonucleotide microarrays were used to profile directly extracted rRNA from environmental microbial populations without PCR amplification. In our initial inspection of two distinct estuarine study sites, the hybridization patterns were reproducible and varied between estuarine sediments of

  11. Site-specific synthesis of oligonucleotides containing malondialdehyde adducts of deoxyguanosine and deoxyadenosine via a postsynthetic modification strategy.

    Science.gov (United States)

    Wang, Hao; Kozekov, Ivan D; Kozekova, Albena; Tamura, Pamela J; Marnett, Lawrence J; Harris, Thomas M; Rizzo, Carmelo J

    2006-11-01

    Malondialdehyde (MDA) and its reactive equivalent, base propenal, are products of oxidative damage to lipids and DNA, respectively; they are mutagenic in bacterial and mammalian systems, and MDA is carcinogenic in rats. MDA adducts of deoxyguanosine (M1dG), deoxyadenosine (OPdA), and deoxycytidine (OPdC) have been characterized. We have developed site-specific syntheses of M1dG and OPdA adducted oligonucleotides that rely on a postsynthetic modification strategy. This work provides an alternative route to the M1dG adducted oligonucleotide and, to date, the only viable strategy for the site-specific synthesis of OPdA-modified oligonucleotides. The stability of the modified oligonucleotides was examined by UV thermal melting studies (Tm). In contrast to the M1dG adduct, OPdA caused very little change in the Tm.

  12. Site-specific synthesis of oligonucleotides containing malondialdehyde adducts of deoxyguanosine and deoxyadenosine via a post-synthetic modification strategy

    Science.gov (United States)

    Wang, Hao; Kozekov, Ivan D.; Kozekova, Albena; Tamura, Pamela; Marnett, Lawrence J.; Harris, Thomas M.; Rizzo, Carmelo J.

    2008-01-01

    Malondialdehyde (MDA) and its reactive equivalent, base propenal, are products of oxidative damage to lipids and DNA, respectively; they are mutagenic in bacterial and mammalian systems and MDA is carcinogenic in rats. MDA adducts of deoxyguanosine (M1dG), deoxyadenosine (OPdA) and deoxycytidine (OPdC) have been characterized. We have developed site-specific syntheses of M1dG and OPdA adducted oligonucleotides that rely on a post-synthetic modification strategy. This work provides an alternative route to the M1dG adducted oligonucleotide and to date, the only viable strategy for the site-specific synthesis of OPdA modified oligonucleotides. The stability of the modified oligonucleotides was examined by UV thermal melting studies (Tm). In contrast to the M1dG adduct, OPdA caused very little change in the Tm. PMID:17112234

  13. Genome dynamics of short oligonucleotides: the example of bacterial DNA uptake enhancing sequences.

    Directory of Open Access Journals (Sweden)

    Mohammed Bakkali

    Full Text Available Among the many bacteria naturally competent for transformation by DNA uptake-a phenomenon with significant clinical and financial implications- Pasteurellaceae and Neisseriaceae species preferentially take up DNA containing specific short sequences. The genomic overrepresentation of these DNA uptake enhancing sequences (DUES causes preferential uptake of conspecific DNA, but the function(s behind this overrepresentation and its evolution are still a matter for discovery. Here I analyze DUES genome dynamics and evolution and test the validity of the results to other selectively constrained oligonucleotides. I use statistical methods and computer simulations to examine DUESs accumulation in Haemophilus influenzae and Neisseria gonorrhoeae genomes. I analyze DUESs sequence and nucleotide frequencies, as well as those of all their mismatched forms, and prove the dependence of DUESs genomic overrepresentation on their preferential uptake by quantifying and correlating both characteristics. I then argue that mutation, uptake bias, and weak selection against DUESs in less constrained parts of the genome combined are sufficient enough to cause DUESs accumulation in susceptible parts of the genome with no need for other DUES function. The distribution of overrepresentation values across sequences with different mismatch loads compared to the DUES suggests a gradual yet not linear molecular drive of DNA sequences depending on their similarity to the DUES. Other genomically overrepresented sequences, both pro- and eukaryotic, show similar distribution of frequencies suggesting that the molecular drive reported above applies to other frequent oligonucleotides. Rare oligonucleotides, however, seem to be gradually drawn to genomic underrepresentation, thus, suggesting a molecular drag. To my knowledge this work provides the first clear evidence of the gradual evolution of selectively constrained oligonucleotides, including repeated, palindromic and protein

  14. DNA Damage Response Pathway and Replication Fork Stress During Oligonucleotide Directed Gene Editing

    OpenAIRE

    Bonner, Melissa; Strouse,Bryan; Applegate, Mindy; Livingston, Paula; Kmiec, Eric B.

    2012-01-01

    Single-stranded DNA oligonucleotides (ODNs) can be used to direct the exchange of nucleotides in the genome of mammalian cells in a process known as gene editing. Once refined, gene editing should become a viable option for gene therapy and molecular medicine. Gene editing is regulated by a number of DNA recombination and repair pathways whose natural activities often lead to single- and double-stranded DNA breaks. It has been previously shown that introduction of a phosphorotioated ODN, desi...

  15. Studies on oligonucleotides and analogues: from drug candidates to components for high-ordered supramolecular structures

    OpenAIRE

    Nici, Fabrizia

    2016-01-01

    Last century advances in molecular biology and biotechnologies enabled us to understand the exceptionally broad repertoire of nucleic acids functions. For a long time DNA was regarded as a rigid molecule with the sole purpose to store and transmit genetic information. However, this idea has been completely overtaken with the discovery of the catalytic activity of specific RNA molecules. In effect, the early 1970s and the embryonic works on antisense oligonucleotides paved the way for our c...

  16. Scalable Amplification of Strand Subsets from Chip-Synthesized Oligonucleotide Libraries (Open Access)

    Science.gov (United States)

    2015-11-16

    oligonucleotides can have practically arbitrary sequences, as fixed sequence domains are cleaved off using nicking endonucleases . Our process is more...with two nicking sites (blue), a restriction site (orange) and one or two orthogonal subpool- specific barcodes of 10 nucleotides (nt) each (black... restriction sites are shared by all strands of the library, barcodes are subpool-specific. The ‘intervening region’ (dark blue) comprises all non-target

  17. An empirical study of choosing efficient discriminative seeds for oligonucleotide design.

    Science.gov (United States)

    Chung, Won-Hyoung; Park, Seong-Bae

    2009-12-03

    Oligonucleotide design is known as a time-consuming work in bioinformatics. In order to accelerate and be efficient the oligonucleotide design process, one of widely used approach is the prescreening unreliable regions using a hashing (or seeding) algorithm. Since the seeding algorithm is originally proposed to increase sensitivity for local alignment, the specificity should be considered as well as the sensitivity for the oligonucleotide design problem. However, a measure of evaluating the seeds regarding how adequate and efficient they are in the oligo design is not yet proposed. Here, we propose novel measures of evaluating the seeding algorithms based on the discriminability and the efficiency. To evaluate the proposed measures, we examine five seeding algorithms in oligonucleotide design. We carried out a series of experiments to compare the seeding algorithms. As the result, the spaced seed is recorded as the most efficient discriminative seed for oligo design. The performance of transition-constrained seed is slightly lower than the spaced seed. Because BLAT seeding algorithm and Vector seeding algorithm give poor scores in specificity and efficiency, we conclude that these algorithms are not adequate to design oligos. Consequently, we recommend spaced seeds or transition-constrained seeds with 15 approximately 18 weight in order to design oligos with the length of 50 mer. The empirical experiments in real biological data reveal that the recommended seeds show consequently good performance. We also propose a software package which enables the users to get the adequate seeds under their own experimental conditions. Our study is valuable to the two points. One is that our study can be applied to the oligo design programs in order to improve the performance by suggesting the experiment-specific seeds. The other is that our study is useful to improve the performance of the mapping assembly in the field of Next-Generation Sequencing. Our proposed measures are

  18. Universal PCR primers for ribosomal protein gene introns of fish

    Directory of Open Access Journals (Sweden)

    Seinen Chow

    2016-01-01

    Full Text Available Abstract Human ribosomal protein (RP gene sequences with respect to intron/exon structures and corresponding cDNA or genomic data of fish species were obtained from the GenBank database. Based on conserved exon sequences, 128 primer pairs for 41 genes were designed for exon-primed intron-crossing (EPIC polymerase chain reaction (PCR. In reference to the draft genome sequences of the Pacific bluefin tuna (Thunnus orientalis, 12 primer pairs expected to amplify introns of the bluefin tuna with lengths of 500–1000 bp were selected and applied to six distantly related fish species belonging to the Orders Clupeiformes, Tetraodontiformes, Pleuronectiformes, Perciformes, Scorpaeniformes, and Anguilliformes. PCR amplification was observed for at least four species in each primer pair, and all fragments were larger than those expected for intronless amplification. Single fragment amplification was observed for at least seven primer pairs per species. Fragment sizes of the bluefin tuna for nine primer pairs corresponded to those expected from the genomic data. Thus, our primer pairs are potentially applicable to a wide variety of fish species and serve as an initial step for isolating single-copy nuclear DNA sequences.

  19. Hierarchical Analysis of the Omega Ontology

    Energy Technology Data Exchange (ETDEWEB)

    Joslyn, Cliff A.; Paulson, Patrick R.

    2009-12-01

    Initial delivery for mathematical analysis of the Omega Ontology. We provide an analysis of the hierarchical structure of a version of the Omega Ontology currently in use within the US Government. After providing an initial statistical analysis of the distribution of all link types in the ontology, we then provide a detailed order theoretical analysis of each of the four main hierarchical links present. This order theoretical analysis includes the distribution of components and their properties, their parent/child and multiple inheritance structure, and the distribution of their vertical ranks.

  20. Hierarchical MAS Based Control Strategy for Microgrid

    Directory of Open Access Journals (Sweden)

    Wei Wu

    2010-09-01

    Full Text Available Microgrids have become a hot topic driven by the dual pressures of environmental protection concerns and the energy crisis. In this paper, a challenge for the distributed control of a modern electric grid incorporating clusters of residential microgrids is elaborated and a hierarchical multi-agent system (MAS is proposed as a solution. The issues of how to realize the hierarchical MAS and how to improve coordination and control strategies are discussed. Based on MATLAB and ZEUS platforms, bilateral switching between grid-connected mode and island mode is performed under control of the proposed MAS to enhance and support its effectiveness.

  1. Translating Management Ideas in Hierarchical Organization

    DEFF Research Database (Denmark)

    Wæraas, Arild; Nielsen, Jeppe

    of Leadership Pipeline in a Danish fire department and how the translators’ approach changed over time from a modifying to a reproducing mode. The study finds that translation does not necessarily imply transformation of the management idea at every phase of the process, pointing instead to efforts of exact...... imitation and copying of the ”original” idea. It also suggests that translation is likely to involve multiple and successive translation modes and, furthermore, that strongly hierarchical structures offer translators control over the translation process. This hierarchical control seems to reduce...

  2. Hierarchical control of electron-transfer

    DEFF Research Database (Denmark)

    Westerhoff, Hans V.; Jensen, Peter Ruhdal; Egger, Louis

    1997-01-01

    In this chapter the role of electron transfer in determining the behaviour of the ATP synthesising enzyme in E. coli is analysed. It is concluded that the latter enzyme lacks control because of special properties of the electron transfer components. These properties range from absence of a strong...... back pressure by the protonmotive force on the rate of electron transfer to hierarchical regulation of the expression of the gens that encode the electron transfer proteins as a response to changes in the bioenergetic properties of the cell.The discussion uses Hierarchical Control Analysis...

  3. Hierarchical machining materials and their performance

    DEFF Research Database (Denmark)

    Sidorenko, Daria; Loginov, Pavel; Levashov, Evgeny

    2016-01-01

    Machining is an important technological process in many areas of industry. The efficiency of machining determines the quality of many industrial products. Machining efficiency and cost depend on the properties, strength, and microstructure of the machining materials. One of the promising ways...... to increase the reliability and wear resistance of machining tools is the development and use of hierarchical machining materials. In the area of machining materials, designed typically as binder/reinforcement composites, hierarchical structures are realized as lower-scale secondary reinforcements (such...... machining materials and their performance....

  4. Hierarchical social networks and information flow

    Science.gov (United States)

    López, Luis; F. F. Mendes, Jose; Sanjuán, Miguel A. F.

    2002-12-01

    Using a simple model for the information flow on social networks, we show that the traditional hierarchical topologies frequently used by companies and organizations, are poorly designed in terms of efficiency. Moreover, we prove that this type of structures are the result of the individual aim of monopolizing as much information as possible within the network. As the information is an appropriate measurement of centrality, we conclude that this kind of topology is so attractive for leaders, because the global influence each actor has within the network is completely determined by the hierarchical level occupied.

  5. Solution conformation of an oligonucleotide containing a G.G mismatch determined by nuclear magnetic resonance and molecular mechanics.

    OpenAIRE

    Cognet, J A; Gabarro-Arpa, J; Le Bret, M; van der Marel, G A; van Boom, J H; Fazakerley, G V

    1991-01-01

    We have determined by two-dimensional nuclear magnetic resonance studies and molecular mechanics calculations the three dimensional solution structure of the non-selfcomplementary oligonucleotide, d(GAGGAGGCACG). d(CGTGCGTCCTC) in which the central base pair is G.G. This is the first structural determination of a G.G mismatch in a oligonucleotide. Two dimensional nuclear magnetic resonance spectra show that the bases of the mismatched pair are stacked into the helix and that the helix adopts ...

  6. Proposal for new European pharmaceutical legislation to permit access to custom-made anti-sense oligonucleotide medicinal products.

    Science.gov (United States)

    Johnston, John D; Feldschreiber, Peter

    2014-06-01

    Current European pharmaceutical legislation is not adequate to meet advances in science and technologies that will lead to rapid development of custom-made medicines. Using existing legislation for custom-made medical devices as a template and anti-sense oligonucleotides as model medicinal products, we propose new European pharmaceutical legislation to permit timely access to custom-made anti-sense oligonucleotide medicinal products. The proposals may be more widely applicable to other medicinal products. © 2013 The British Pharmacological Society.

  7. Synthesis and Excellent Duplex Stability of Oligonucleotides Containing 2′-Amino-LNA Functionalized with Galactose Units

    Directory of Open Access Journals (Sweden)

    Rajesh Kumar

    2017-05-01

    Full Text Available A convenient method for the preparation of oligonucleotides containing internally-attached galactose and triantennary galactose units has been developed based on click chemistry between 2′-N-alkyne 2′-amino-LNA nucleosides and azido-functionalized galactosyl building blocks. The synthesized oligonucleotides show excellent binding affinity and selectivity towards complementary DNA/RNA strands with an increase in the melting temperature of up to +23.5 °C for triply-modified variants.

  8. Targeting the r(CGG) repeats that cause FXTAS with modularly assembled small molecules and oligonucleotides.

    Science.gov (United States)

    Tran, Tuan; Childs-Disney, Jessica L; Liu, Biao; Guan, Lirui; Rzuczek, Suzanne; Disney, Matthew D

    2014-04-18

    We designed small molecules that bind the structure of the RNA that causes fragile X-associated tremor ataxia syndrome (FXTAS), an incurable neuromuscular disease. FXTAS is caused by an expanded r(CGG) repeat (r(CGG)(exp)) that inactivates a protein regulator of alternative pre-mRNA splicing. Our designed compounds modulate r(CGG)(exp) toxicity in cellular models of FXTAS, and pull-down experiments confirm that they bind r(CGG)(exp) in vivo. Importantly, compound binding does not affect translation of the downstream open reading frame (ORF). We compared molecular recognition properties of our optimal compound to oligonucleotides. Studies show that r(CGG)(exp)'s self-structure is a significant energetic barrier for oligonucleotide binding. A fully modified 2'-OMethyl phosphorothioate is incapable of completely reversing an FXTAS-associated splicing defect and inhibits translation of the downstream ORF, which could have deleterious effects. Taken together, these studies suggest that a small molecule that recognizes structure may be more well suited for targeting highly structured RNAs that require strand invasion by a complementary oligonucleotide.

  9. Specificity tests of an oligonucleotide probe against food-outbreak salmonella for biosensor detection

    Science.gov (United States)

    Chen, I.-H.; Horikawa, S.; Xi, J.; Wikle, H. C.; Barbaree, J. M.; Chin, B. A.

    2017-05-01

    Phage based magneto-elastic (ME) biosensors have been shown to be able to rapidly detect Salmonella in various food systems to serve food pathogen monitoring purposes. In this ME biosensor platform, the free-standing strip-shaped magneto-elastic sensor is the transducer and the phage probe that recognizes Salmonella in food serves as the bio-recognition element. According to Sorokulova et al. at 2005, a developed oligonucleotide probe E2 was reported to have high specificity to Salmonella enterica Typhimurium. In the report, the specificity tests were focused in most of Enterobacterace groups outside of Salmonella family. Here, to understand the specificity of phage E2 to different Salmonella enterica serotypes within Salmonella Family, we further tested the specificity of the phage probe to thirty-two Salmonella serotypes that were present in the major foodborne outbreaks during the past ten years (according to Centers for Disease Control and Prevention). The tests were conducted through an Enzyme linked Immunosorbent Assay (ELISA) format. This assay can mimic probe immobilized conditions on the magnetoelastic biosensor platform and also enable to study the binding specificity of oligonucleotide probes toward different Salmonella while avoiding phage/ sensor lot variations. Test results confirmed that this oligonucleotide probe E2 was high specific to Salmonella Typhimurium cells but showed cross reactivity to Salmonella Tennessee and four other serotypes among the thirty-two tested Salmonella serotypes.

  10. Site-Selective Artificial Ribonucleases: Oligonucleotide Conjugates Containing Multiple Imidazole Residues in the Catalytic Domain

    Directory of Open Access Journals (Sweden)

    Natalia G. Beloglazova

    2011-01-01

    Full Text Available Design of site-selective artificial ribonucleases (aRNases is one of the most challenging tasks in RNA targeting. Here, we designed and studied oligonucleotide-based aRNases containing multiple imidazole residues in the catalytic part and systematically varied structure of cleaving constructs. We demonstrated that the ribonuclease activity of the conjugates is strongly affected by the number of imidazole residues in the catalytic part, the length of a linker between the catalytic imidazole groups of the construct and the oligonucleotide, and the type of anchor group, connecting linker structure and the oligonucleotide. Molecular modeling of the most active aRNases showed that preferable orientation(s of cleaving constructs strongly depend on the structure of the anchor group and length of the linker. The inclusion of deoxyribothymidine anchor group significantly reduced the probability of cleaving groups to locate near the cleavage site, presumably due to a stacking interaction with the neighbouring nucleotide residue. Altogether the obtained results show that dynamics factors play an important role in site-specific RNA cleavage. Remarkably high cleavage activity was displayed by the conjugates with the most flexible and extended cleaving construct, which presumably provides a better opportunity for imidazole residues to be correctly positioned in the vicinity of scissile phosphodiester bond.

  11. Probe selection algorithm for oligonucleotide array-based medium-resolution genotyping.

    Science.gov (United States)

    Zhou, Y; Peng, S; Gao, H; Cheng, J

    2004-11-01

    Medium-resolution genotyping has the goal of distinguishing different subgroups instead of each element in a group. An oligonucleotide array provides an inexpensive, high-throughput method to identify differences in DNA sequence among individuals, which is fundamental for genotyping. As the cost and difficulty of designing and fabricating the oligonucleotide array dramatically increase with the number of probes used, it is therefore important to have a design with a minimum number of probes meeting the requirement of medium-resolution genotyping. The first algorithm for designing and selecting probes for oligonucleotide array-based medium-resolution typing is reported. The goal in deriving the algorithm was to select a minimum number of probes from a large probe set on the premise of minimum loss of resolution. The algorithm, which was based on entropy, conditional entropy and mutual information theory, was used to select the minimum number of probes from a large probe set. The algorithm was tested on a human leukocyte antigen (HLA) sequence data set Thirty probes were selected from 390 probes for HLA-A, and 60 probes were selected from 767 probes for HLA-B. Although the number of probes was reduced by almost ten times, the distinguishability was reduced only a little, by 0.45% (from 99.90% to 99.45%) for HLA-A and 0.27% (from 99.84% to 99.57%) for HLA-B, respectively. This is a satisfactory and practical result.

  12. The estimation of quantitative parameters of oligonucleotides immobilization on mica surface

    Science.gov (United States)

    Sharipov, T. I.; Bakhtizin, R. Z.

    2017-05-01

    Immobilization of nucleic acids on the surface of various materials is increasingly being used in research and some practical applications. Currently, the DNA chip technology is rapidly developing. The basis of the immobilization process can be both physical adsorption and chemisorption. A useful way to control the immobilization of nucleic acids on a surface is to use atomic force microscopy. It allows you to investigate the topography of the surface by its direct imaging with high resolution. Usually, to fix the DNA on the surface of mica are used cations which mediate the interaction between the mica surface and the DNA molecules. In our work we have developed a method for estimation of quantitative parameter of immobilization of oligonucleotides is their degree of aggregation depending on the fixation conditions on the surface of mica. The results on study of aggregation of oligonucleotides immobilized on mica surface will be presented. The single oligonucleotides molecules have been imaged clearly, whereas their surface areas have been calculated and calibration curve has been plotted.

  13. Long-range order of organized oligonucleotide monolayers on Au(111) electrodes

    DEFF Research Database (Denmark)

    Wackerbarth, Hainer; Grubb, Mikala; Zhang, Jingdong

    2004-01-01

    , with electrochemical potential control of both the sample electrode and the tip. All the data are based on single-crystal, atomically planar Au(111)-electrode surfaces. The high sensitivity of such surfaces provides accurate HS-10A and HS-10AT electrode coverages on the basis of the reductive desorption of the Au......Oligonucleotides modified by a hexamethylene linker group adsorb on gold electrodes via Au-S bond formation. We have obtained novel data for adsorption of thiol-modified (HS) single-strand HS-10A and double-stranded HS-10AT oligonucleotides and for analogous thiol-free 10A (A = adenine) and 10T (T......-S bond. The coverage is high and in keeping with dense monolayers of adsorbed HS-10A and HS-10AT in an upright or tilted orientation, with the oligonucleotide backbone repelled from the strongly negatively charged electrode surface. Adsorbed thiol-free 10A only gives aAu(111)-reconstruction peak, while...

  14. Structure/nuclease activity relationships of DNA cleavers based on cationic metalloporphyrin-oligonucleotide conjugates.

    Science.gov (United States)

    Mestre, B; Jakobs, A; Pratviel, G; Meunier, B

    1996-07-16

    The covalent attachment of a managanese-tris(methylpyridiniumyl)porphyrin entity to an antisense oligonucleotide allowed sequence-selective oxidative cleavage of DNA when the metalloporphyrin was activated by potassium monopersulfate (KHSO5). We prepared several structurally modified metallo-porphyrin-oligonucleotide conjugates in order to find out the most efficient compound for in vitro DNA cleavage. The nature and the length of the tether were modulated, the metalloporphyrin entity was modified (metal, ligand), and different ways of activation of the metalloporphyrin were assayed. We noticed that the location of the peptidic bond within the linker could greatly affect the cleavage efficiency of the different conjugates. We showed that the most efficient conjugate for oxidative DNA cleavage was a manganese tetracationic porphyrin-oligonucleotide compound. When the metalloporphyrin moiety was activated by a reducing agent in the presence of molecular oxygen, DNA cleavage was efficient at suitable concentrations of the reducing agent, in order to avoid the reduction of the activated DNA cleaver, a putative high-valent metal-oxo species, by the excess of reducing agent.

  15. Efficient surface patterning of oligonucleotides inside a glass capillary through oxime bond formation.

    Science.gov (United States)

    Dendane, Nabil; Hoang, Antoine; Guillard, Ludovic; Defrancq, Eric; Vinet, Françoise; Dumy, Pascal

    2007-01-01

    The efficient surface patterning of oligonucleotides was accomplished onto the inner wall of fused-silica capillary tubes as well as on the surface of glass slides through oxime bond formation. The robustness of the method was demonstrated by achieving the surface immobilization of up to three different oligonucleotide sequences inside the same capillary tube. The method involves the preparation of surfaces grafted with reactive aminooxy functionalities masked with the photocleavable protecting group, 2-(2-nitrophenyl) propyloxycarbonyl group (NPPOC). Briefly, NPPOC-aminooxy silane 1 was prepared and used to silanize the glass surfaces. The NPPOC group was cleaved under brief irradiation to unmask the reactive aminooxy group on surfaces. These reactive aminooxy groups were allowed to react with aldehyde-containing oligonucleotides to achieve an efficient surface immobilization. The advantage associated with the present approach is that it combines the high-coupling efficiency of oxime bond formation with the convenience associated with the use of photolabile groups. The present strategy thus offers an alternative approach for the immobilization of biomolecules in the microchannels of "labs on a chip" devices.

  16. Recent Advances in Nucleic Acid Targeting Probes and Supramolecular Constructs Based on Pyrene-Modified Oligonucleotides

    Directory of Open Access Journals (Sweden)

    Olga A. Krasheninina

    2017-11-01

    Full Text Available In this review, we summarize the recent advances in the use of pyrene-modified oligonucleotides as a platform for functional nucleic acid-based constructs. Pyrene is of special interest for the development of nucleic acid-based tools due to its unique fluorescent properties (sensitivity of fluorescence to the microenvironment, ability to form excimers and exciplexes, long fluorescence lifetime, high quantum yield, ability to intercalate into the nucleic acid duplex, to act as a π-π-stacking (including anchoring moiety, and others. These properties of pyrene have been used to construct novel sensitive fluorescent probes for the sequence-specific detection of nucleic acids and the discrimination of single nucleotide polymorphisms (SNPs, aptamer-based biosensors, agents for binding of double-stranded DNAs, and building blocks for supramolecular complexes. Special attention is paid to the influence of the design of pyrene-modified oligonucleotides on their properties, i.e., the structure-function relationships. The perspectives for the applications of pyrene-modified oligonucleotides in biomolecular studies, diagnostics, and nanotechnology are discussed.

  17. Development of Multiexon Skipping Antisense Oligonucleotide Therapy for Duchenne Muscular Dystrophy

    Science.gov (United States)

    Yokota, Toshifumi; Wood, Matthew J. A.

    2013-01-01

    Duchenne muscular dystrophy (DMD) is an incurable, X-linked progressive muscle degenerative disorder that results from the absence of dystrophin protein and leads to premature death in affected individuals due to respiratory and/or cardiac failure typically by age of 30. Very recently the exciting prospect of an effective oligonucleotide therapy has emerged which restores dystrophin protein expression to affected tissues in DMD patients with highly promising data from a series of clinical trials. This therapeutic approach is highly mutation specific and thus is personalised. Therefore DMD has emerged as a model genetic disorder for understanding and overcoming of the challenges of developing personalised genetic medicines. One of the greatest weaknesses of the current oligonucleotide approach is that it is a mutation-specific therapy. To address this limitation, we have recently demonstrated that exons 45–55 skipping therapy has the potential to treat clusters of mutations that cause DMD, which could significantly reduce the number of compounds that would need to be developed in order to successfully treat all DMD patients. Here we discuss and review the latest preclinical work in this area as well as a variety of accompanying issues, including efficacy and potential toxicity of antisense oligonucleotides, prior to human clinical trials. PMID:23984357

  18. Development of Multiexon Skipping Antisense Oligonucleotide Therapy for Duchenne Muscular Dystrophy

    Directory of Open Access Journals (Sweden)

    Yoshitsugu Aoki

    2013-01-01

    Full Text Available Duchenne muscular dystrophy (DMD is an incurable, X-linked progressive muscle degenerative disorder that results from the absence of dystrophin protein and leads to premature death in affected individuals due to respiratory and/or cardiac failure typically by age of 30. Very recently the exciting prospect of an effective oligonucleotide therapy has emerged which restores dystrophin protein expression to affected tissues in DMD patients with highly promising data from a series of clinical trials. This therapeutic approach is highly mutation specific and thus is personalised. Therefore DMD has emerged as a model genetic disorder for understanding and overcoming of the challenges of developing personalised genetic medicines. One of the greatest weaknesses of the current oligonucleotide approach is that it is a mutation-specific therapy. To address this limitation, we have recently demonstrated that exons 45–55 skipping therapy has the potential to treat clusters of mutations that cause DMD, which could significantly reduce the number of compounds that would need to be developed in order to successfully treat all DMD patients. Here we discuss and review the latest preclinical work in this area as well as a variety of accompanying issues, including efficacy and potential toxicity of antisense oligonucleotides, prior to human clinical trials.

  19. Optimal conditions for hybridization with oligonucleotides: a study with myc-oncogene DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Albretsen, C.; Haukanes, B.I.; Aasland, R.; Kleppe, K.

    1988-04-01

    The authors present a study on the refinement of filter-hybridization conditions for a series of synthetic oligonucleotides in the range from 17 to 50 base residues in length. Experimental conditions for hybridization and the subsequent washing steps of the filter were optimized for different lengths of the synthetic oligonucleotides by varying the formamide concentration and washing conditions. Target DNA was immobilized to the nitrocellulose filter with the slot blot technique. The sequences of the synthetic oligonucleotides are derived from the third exon of the human oncogene c-myc and the corresponding viral gene v-myc and the G+C content was between 43 and 47%. Optimal conditions for hybridization with a 82% homologous 30-mer and 100% homologous 17-, 20-, 25-, 30-, and 50-mers were found to be a concentration of formamide of 15, 15, 30, 30, 40, and 50%, respectively. The melting temperature for these optimal hybridization and washing conditions was calculated to be up to 11/sup 0/C below the hybridization temperature actually used. This confirms that the duplexes are more stable than expected. The melting points for 17-, 20-, and 30-mers were measured in the presence of 5x SSC and found to be 43, 58, and 60/sup 0/C, respectively. Competition between double- and single-stranded DNA probes to the target DNA was investigated. The single-stranded DNA probes were about 30- to 40-fold more sensitive than the double-stranded DNA probes.

  20. Detection of Glucose with Atomic Absorption Spectroscopy by Using Oligonucleotide Functionalized Gold Nanoparticle.

    Science.gov (United States)

    Zhang, Hong; Yan, Honglian; Ling, Liansheng

    2016-06-01

    A novel method for the detection of glucose was established with atomic absorption spectroscopy by using the label of gold nanoparticle (AuNP). Silver-coated glass assembled with oligonucleotide 5'-SH-T12-AGA CAA GAG AGG-3' (Oligo 1) was acted as separation probe, oligonucleotide 5'-CAA CAG AGA ACG-T12-SH-3' modified gold nanoparticle (AuNP-Oligo 2) was acted as signal-reporting probe. Oligonucleotide 5'-CGT TCT CTG TTG CCT CTC TTG TCT-3' (Oligo 3) could hybridize with Oligo 1 on the surface of silver-coated glass and AuNP-Oligo 2, and free AuNP-Oligo 2 could be removed by rinsing with buffer. Hence the concentration of Oligo 3 was transformed into the concentration of gold element. In addition, Oligo 3 could be cleaved into DNA fragments by glucose, glucose oxidase and Fe(2+)-EDTA through Fenton reaction. Thereby the concentration of glucose could be transformed to the absorbance of gold element. Under the optimum conditions, the integrated absorbance decreased proportionally to the concentration of glucose over the range from 50.0 μM to 1.0 mM with a detection limit of 40.0 μM. Moreover, satisfactory result was obtained when the assay was used to determinate glucose in human serum.

  1. Oligonucleotide and Parylene Surface Coating of Polystyrene and ePTFE for Improved Endothelial Cell Attachment and Hemocompatibility

    Directory of Open Access Journals (Sweden)

    Martina Schleicher

    2012-01-01

    Full Text Available In vivo self-endothelialization by endothelial cell adhesion on cardiovascular implants is highly desirable. DNA-oligonucleotides are an intriguing coating material with nonimmunogenic characteristics and the feasibility of easy and rapid chemical fabrication. The objective of this study was the creation of cell adhesive DNA-oligonucleotide coatings on vascular implant surfaces. DNA-oligonucleotides immobilized by adsorption on parylene (poly(monoaminomethyl-para-xylene coated polystyrene and ePTFE were resistant to high shear stress (9.5 N/m2 and human blood serum for up to 96 h. Adhesion of murine endothelial progenitor cells, HUVECs and endothelial cells from human adult saphenous veins as well as viability over a period of 14 days of HUVECs on oligonucleotide coated samples under dynamic culture conditions was significantly enhanced (P<0.05. Oligonucleotide-coated surfaces revealed low thrombogenicity and excellent hemocompatibility after incubation with human blood. These properties suggest the suitability of immobilization of DNA-oligonucleotides for biofunctionalization of blood vessel substitutes for improved in vivo endothelialization.

  2. Discovery and development of the G-rich oligonucleotide AS1411 as a novel treatment for cancer.

    Science.gov (United States)

    Bates, Paula J; Laber, Damian A; Miller, Donald M; Thomas, Shelia D; Trent, John O

    2009-06-01

    Certain guanine-rich (G-rich) DNA and RNA molecules can associate intermolecularly or intramolecularly to form four stranded or "quadruplex" structures, which have unusual biophysical and biological properties. Several synthetic G-rich quadruplex-forming oligodeoxynucleotides have recently been investigated as therapeutic agents for various human diseases. We refer to these biologically active G-rich oligonucleotides as aptamers because their activities arise from binding to protein targets via shape-specific recognition (analogous to antibody-antigen binding). As therapeutic agents, the G-rich aptamers may have some advantages over monoclonal antibodies and other oligonucleotide-based approaches. For example, quadruplex oligonucleotides are non-immunogenic, heat stable and they have increased resistance to serum nucleases and enhanced cellular uptake compared to unstructured sequences. In this review, we describe the characteristics and activities of G-rich oligonucleotides. We also give a personal perspective on the discovery and development of AS1411, an antiproliferative G-rich phosphodiester oligonucleotide that is currently being tested as an anticancer agent in Phase II clinical trials. This molecule functions as an aptamer to nucleolin, a multifunctional protein that is highly expressed by cancer cells, both intracellularly and on the cell surface. Thus, the serendipitous discovery of the G-rich oligonucleotides also led to the identification of nucleolin as a new molecular target for cancer therapy.

  3. TSUNAMI Primer: A Primer for Sensitivity/Uncertainty Calculations with SCALE

    Energy Technology Data Exchange (ETDEWEB)

    Rearden, Bradley T [ORNL; Mueller, Don [ORNL; Bowman, Stephen M [ORNL; Busch, Robert D. [University of New Mexico, Albuquerque; Emerson, Scott [University of New Mexico, Albuquerque

    2009-01-01

    This primer presents examples in the application of the SCALE/TSUNAMI tools to generate k{sub eff} sensitivity data for one- and three-dimensional models using TSUNAMI-1D and -3D and to examine uncertainties in the computed k{sub eff} values due to uncertainties in the cross-section data used in their calculation. The proper use of unit cell data and need for confirming the appropriate selection of input parameters through direct perturbations are described. The uses of sensitivity and uncertainty data to identify and rank potential sources of computational bias in an application system and TSUNAMI tools for assessment of system similarity using sensitivity and uncertainty criteria are demonstrated. Uses of these criteria in trending analyses to assess computational biases, bias uncertainties, and gap analyses are also described. Additionally, an application of the data adjustment tool TSURFER is provided, including identification of specific details of sources of computational bias.

  4. Modular networks with hierarchical organization: The dynamical ...

    Indian Academy of Sciences (India)

    another example, this time a hierarchical structure, viz., the Cayley tree with b branches at each vertex. Again, it is easy to see that the clustering vs. degree curve will not show the characteristic scaling seen for the RB model. In fact, in the next section, we show that even for networks where both hierarchy and modularity are.

  5. Modular networks with hierarchical organization: The dynamical ...

    Indian Academy of Sciences (India)

    As both hierarchy and modularity are seen in natural systems, which necessarily have to be robust against environmental fluctuations, we conclude that additional constraints are necessary for the emergence of hierarchical structure, similar to the occurrence of modularity through multi-constraint optimization as shown by us ...

  6. Tanzania: A Hierarchical Cluster Analysis Approach | Ngaruko ...

    African Journals Online (AJOL)

    Using survey data from Kibondo district, west Tanzania, we use hierarchical cluster analysis to classify borrower farmers according to their borrowing behaviour into four distinctive clusters. The appreciation of the existence of heterogeneous farmer clusters is vital in forging credit delivery policies that are not only ...

  7. Transforming Hierarchical Relationships in Student Conduct Administration

    Science.gov (United States)

    Jacobson, Kelly A.

    2013-01-01

    Conflict transformation theory provided a philosophical lens for this critical cultural, constructivist study, wherein four student conduct administrators who engage in leveling hierarchical relationships with students in conduct processes shared ways they make meaning of their professional practice. Through informal, unstructured interviews, a…

  8. Ultrafast Hierarchical OTDM/WDM Network

    Directory of Open Access Journals (Sweden)

    Hideyuki Sotobayashi

    2003-12-01

    Full Text Available Ultrafast hierarchical OTDM/WDM network is proposed for the future core-network. We review its enabling technologies: C- and L-wavelength-band generation, OTDM-WDM mutual multiplexing format conversions, and ultrafast OTDM wavelengthband conversions.

  9. SORM applied to hierarchical parallel system

    DEFF Research Database (Denmark)

    Ditlevsen, Ove Dalager

    2006-01-01

    The old hierarchical stochastic load combination model of Ferry Borges and Castanheta and the corresponding problem of determining the distribution of the extreme random load effect is the inspiration to this paper. The evaluation of the distribution function of the extreme value by use of a part...

  10. A hierarchical classification scheme of psoriasis images

    DEFF Research Database (Denmark)

    Maletti, Gabriela Mariel; Ersbøll, Bjarne Kjær

    2003-01-01

    A two-stage hierarchical classification scheme of psoriasis lesion images is proposed. These images are basically composed of three classes: normal skin, lesion and background. The scheme combines conventional tools to separate the skin from the background in the first stage, and the lesion from...

  11. Parallel Temporal Dynamics in Hierarchical Cognitive Control

    Science.gov (United States)

    Ranti, Carolyn; Chatham, Christopher H.; Badre, David

    2015-01-01

    Cognitive control allows us to follow abstract rules in order to choose appropriate responses given our desired outcomes. Cognitive control is often conceptualized as a hierarchical decision process, wherein decisions made at higher, more abstract levels of control asymmetrically influence lower-level decisions. These influences could evolve sequentially across multiple levels of a hierarchical decision, consistent with much prior evidence for central bottlenecks and seriality in decision-making processes. However, here, we show that multiple levels of hierarchical cognitive control are processed primarily in parallel. Human participants selected responses to stimuli using a complex, multiply contingent (third order) rule structure. A response deadline procedure allowed assessment of the accuracy and timing of decisions made at each level of the hierarchy. In contrast to a serial decision process, error rates across levels of the decision mostly declined simultaneously and at identical rates, with only a slight tendency to complete the highest level decision first. Simulations with a biologically plausible neural network model demonstrate how such parallel processing could emerge from a previously developed hierarchically nested frontostriatal architecture. Our results support a parallel processing model of cognitive control, in which uncertainty on multiple levels of a decision is reduced simultaneously. PMID:26051820

  12. Hierarchical Logistic Regression in Course Placement

    Science.gov (United States)

    Schulz, E. Matthew; Betebenner, Damian; Ahn, Meeyeon

    2004-01-01

    Whether hierarchical logistic regression can reduce the sample size requirement for estimating optimal cutoff scores in a course placement service where predictive validity is measured by a threshold utility function is explored. Data from courses with varying class size were randomly partitioned into two halves per course. Nonhierarchical and…

  13. Analyzing security protocols in hierarchical networks

    DEFF Research Database (Denmark)

    Zhang, Ye; Nielson, Hanne Riis

    2006-01-01

    Validating security protocols is a well-known hard problem even in a simple setting of a single global network. But a real network often consists of, besides the public-accessed part, several sub-networks and thereby forms a hierarchical structure. In this paper we first present a process calculus...

  14. Integrating Linear Programming and Analytical Hierarchical ...

    African Journals Online (AJOL)

    A comprehensive Linear Programming model established, including 106 variables and 43 ecological-socio-economic constraints. Land capability and suitability evaluation accomplished using ecological factors and Comparative Advantages of the uses and the factors, respectively. Analytical Hierarchical Process followed ...

  15. Hierarchical Network Design Using Simulated Annealing

    DEFF Research Database (Denmark)

    Thomadsen, Tommy; Clausen, Jens

    2002-01-01

    networks are described and a mathematical model is proposed for a two level version of the hierarchical network problem. The problem is to determine which edges should connect nodes, and how demand is routed in the network. The problem is solved heuristically using simulated annealing which as a sub...

  16. Hierarchical Scaling in Systems of Natural Cities

    CERN Document Server

    Chen, Yanguang

    2016-01-01

    Hierarchies can be modeled by a set of exponential functions, from which we can derive a set of power laws indicative of scaling. These scaling laws are followed by many natural and social phenomena such as cities, earthquakes, and rivers. This paper is devoted to revealing the scaling patterns in systems of natural cities by reconstructing the hierarchy with cascade structure. The cities of America, Britain, France, and Germany are taken as examples to make empirical analyses. The hierarchical scaling relations can be well fitted to the data points within the scaling ranges of the size and area of the natural cities. The size-number and area-number scaling exponents are close to 1, and the allometric scaling exponent is slightly less than 1. The results suggest that natural cities follow hierarchical scaling laws and hierarchical conservation law. Zipf's law proved to be one of the indications of the hierarchical scaling, and the primate law of city-size distribution represents a local pattern and can be mer...

  17. Sharing the proceeds from a hierarchical venture

    DEFF Research Database (Denmark)

    Hougaard, Jens Leth; Moreno-Ternero, Juan D.; Tvede, Mich

    2017-01-01

    We consider the problem of distributing the proceeds generated from a joint venture in which the participating agents are hierarchically organized. We introduce and characterize a family of allocation rules where revenue ‘bubbles up’ in the hierarchy. The family is flexible enough to accommodate...

  18. Evaluation of a new universal primer for ceramics and alloys.

    Science.gov (United States)

    Azimian, Farhad; Klosa, Karsten; Kern, Matthias

    2012-06-01

    To evaluate in vitro the effect of using a universal primer on the resin bond strength to ceramics (silicate and zirconia) and two noble alloys for metal-ceramic restorations. Disk-like specimens made of four dental restoration materials (silicate ceramic, zirconia ceramic, gold-based alloy and palladium-based alloy) were ground with abrasive paper, prepared for bonding (silicate ceramic: etching with 5% hydrofluoric acid for 60 s; the other materials: air abrasion with 50-µm Al2O3 at 2.5 bar) and ultrasonically cleaned in 96% isopropanol. Eight specimens of each material were primed with a universal primer, 8 specimens were primed with a well-established material-specific method (silicate ceramic: silanization; zirconia ceramic: phosphate monomer-containing primer; alloys: sulfuric monomer-containing primer). Plexiglas tubes filled with a composite resin were bonded to the specimens using an alignment apparatus and a composite luting resin. After storage in 37°C tap water for three days, or 167 days with additional thermocycling, tensile bond strength (TBS) was measured in a universal testing machine at a crosshead speed of 2 mm/min. After 167 days of storage, median TBS values for each material were (well-established method/universal primer) 40 MPa/37 MPa (silicate ceramic), 41 MPa/41 MPa (zirconia ceramic), 20 MPa/17 MPa (gold-based alloy), and 20 MPa/19 MPa (palladium-based alloy). Statistical analysis revealed no statistically significant differences among control groups and test groups. The universal primer seems to be a promising alternative compared to using three different primers for alloys, silicate and zirconia ceramics.

  19. Hierarchically Self-Assembled Nanofiber Films from Amylose-Grafted Carboxymethyl Cellulose

    Directory of Open Access Journals (Sweden)

    Daisuke Hatanaka

    2014-01-01

    Full Text Available In this paper, we report the formation of hierarchically self-assembled nanofiber films from amylose-grafted sodium carboxymethyl celluloses (NaCMCs that were synthesized by a chemoenzymatic approach. First, maltooligosaccharide primer-grafted NaCMCs were prepared by a chemical reaction using two kinds of NaCMCs with different degrees of polymerization (DPs from Avicel and cotton sources. Then, phosphorylase-catalyzed enzymatic polymerization of α-d-glucose 1-phosphate from the nonreducing ends of the primer chains on the products was conducted to produce the prescribed amylose-grafted NaCMCs. The films were obtained by drying aqueous alkaline solutions of the amylose-grafted NaCMCs. The scanning electron microscopy (SEM image of the film fabricated from the material with the higher DP from the cotton source showed a clear, self-assembled, highly condensed tangle of nanofibers. The SEM image of the material with the lower DP from the Avicel source, on the other hand, showed an unclear nanofiber morphology. These results indicate that the DPs of the main chains in the materials strongly affected the hierarchically self-assembled nanofiber formation. The SEM images of the films after washing out the alkali, furthermore, showed that the fibers partially merged with each other at the interfacial area owing to the double helix formation between the amylose-grafted chains. The mechanical properties of the films under tensile mode also depended on the self-assembled morphologies of the amylose-grafted NaCMCs from the different sources.

  20. Hierarchical organisation in perception of orientation.

    Science.gov (United States)

    Spinelli, D; Antonucci, G; Daini, R; Martelli, M L; Zoccolotti, P

    1999-01-01

    According to Rock [1990, in The Legacy of Solomon Asch (Hillsdale, NJ: Lawrence Erlbaum Associates)], hierarchical organisation of perception describes cases in which the orientation of an object is affected by the immediately surrounding elements in the visual field. Various experiments were performed to study the hierarchical organisation of orientation perception. In most of them the rod-and-frame-illusion (RFI: change of the apparent vertical measured on a central rod surrounded by a tilted frame) was measured in the presence/absence of a second inner frame. The first three experiments showed that, when the inner frame is vertical, the direction and size of the illusion are consistent with expectancies based on the hierarchical organisation hypothesis. An analysis of published and unpublished data collected on a large number of subjects showed that orientational hierarchical effects are independent from the absolute size of the RFI. In experiments 4 to 7 we examined the perceptual conditions of the inner stimulus (enclosure, orientation, and presence of luminance borders) critical for obtaining a hierarchical organisation effect. Although an inner vertical square was effective in reducing the illusion (experiment 3), an inner circle enclosing the rod was ineffective (experiment 4). This indicates that definite orientation is necessary to modulate the illusion. However, orientational information provided by a vertical or horizontal rectangle presented near the rod, but not enclosing it, did not modulate the RFI (experiment 5). This suggests that the presence of a figure with oriented contours enclosing the rod is critical. In experiments 6 and 7 we studied whether the presence of luminance borders is important or whether the inner upright square might be effective also if made of subjective contours. When the subjective contour figure was salient and the observers perceived it clearly, its effectiveness in modulating the RFI was comparable to that observed with

  1. Generic hierarchical engine for mask data preparation

    Science.gov (United States)

    Kalus, Christian K.; Roessl, Wolfgang; Schnitker, Uwe; Simecek, Michal

    2002-07-01

    Electronic layouts are usually flattened on their path from the hierarchical source downstream to the wafer. Mask data preparation has certainly been identified as a severe bottleneck since long. Data volumes are not only doubling every year along the ITRS roadmap. With the advent of optical proximity correction and phase-shifting masks data volumes are escalating up to non-manageable heights. Hierarchical treatment is one of the most powerful means to keep memory and CPU consumption in reasonable ranges. Only recently, however, has this technique acquired more public attention. Mask data preparation is the most critical area calling for a sound infrastructure to reduce the handling problem. Gaining more and more attention though, are other applications such as large area simulation and manufacturing rule checking (MRC). They all would profit from a generic engine capable to efficiently treat hierarchical data. In this paper we will present a generic engine for hierarchical treatment which solves the major problem, steady transitions along cell borders. Several alternatives exist how to walk through the hierarchy tree. They have, to date, not been thoroughly investigated. One is a bottom-up attempt to treat cells starting with the most elementary cells. The other one is a top-down approach which lends itself to creating a new hierarchy tree. In addition, since the variety, degree of hierarchy and quality of layouts extends over a wide range a generic engine has to take intelligent decisions when exploding the hierarchy tree. Several applications will be shown, in particular how far the limits can be pushed with the current hierarchical engine.

  2. Mutated primer binding sites interacting with different tRNAs allow efficient murine leukemia virus replication

    DEFF Research Database (Denmark)

    Lund, Anders Henrik; Duch, M; Lovmand, J

    1993-01-01

    Two Akv murine leukemia virus-based retroviral vectors with primer binding sites matching tRNA(Gln-1) and tRNA(Lys-3) were constructed. The transduction efficiency of these mutated vectors was found to be comparable to that of a vector carrying the wild-type primer binding site matching t......RNA(Pro). Polymerase chain reaction amplification and sequence analysis of transduced proviruses confirmed the transfer of vectors with mutated primer binding sites and further showed that tRNA(Gln-2) may act efficiently in conjunction with the tRNA(Gln-1) primer binding site. We conclude that murine leukemia virus...... can replicate by using various tRNA molecules as primers and propose primer binding site-tRNA primer interactions to be of major importance for tRNA primer selection. However, efficient primer selection does not require perfect Watson-Crick base pairing at all 18 positions of the primer binding site....

  3. Self-assembly of hierarchically ordered structures in DNA nanotube systems

    Science.gov (United States)

    Glaser, Martin; Schnauß, Jörg; Tschirner, Teresa; Schmidt, B. U. Sebastian; Moebius-Winkler, Maximilian; Käs, Josef A.; Smith, David M.

    2016-05-01

    The self-assembly of molecular and macromolecular building blocks into organized patterns is a complex process found in diverse systems over a wide range of size and time scales. The formation of star- or aster-like configurations, for example, is a common characteristic in solutions of polymers or other molecules containing multi-scaled, hierarchical assembly processes. This is a recurring phenomenon in numerous pattern-forming systems ranging from cellular constructs to solutions of ferromagnetic colloids or synthetic plastics. To date, however, it has not been possible to systematically parameterize structural properties of the constituent components in order to study their influence on assembled states. Here, we circumvent this limitation by using DNA nanotubes with programmable mechanical properties as our basic building blocks. A small set of DNA oligonucleotides can be chosen to hybridize into micron-length DNA nanotubes with a well-defined circumference and stiffness. The self-assembly of these nanotubes to hierarchically ordered structures is driven by depletion forces caused by the presence of polyethylene glycol. This trait allowed us to investigate self-assembly effects while maintaining a complete decoupling of density, self-association or bundling strength, and stiffness of the nanotubes. Our findings show diverse ranges of emerging structures including heterogeneous networks, aster-like structures, and densely bundled needle-like structures, which compare to configurations found in many other systems. These show a strong dependence not only on concentration and bundling strength, but also on the underlying mechanical properties of the nanotubes. Similar network architectures to those caused by depletion forces in the low-density regime are obtained when an alternative hybridization-based bundling mechanism is employed to induce self-assembly in an isotropic network of pre-formed DNA nanotubes. This emphasizes the universal effect inevitable

  4. PCR primers for metazoan mitochondrial 12S ribosomal DNA sequences.

    Directory of Open Access Journals (Sweden)

    Ryuji J Machida

    Full Text Available BACKGROUND: Assessment of the biodiversity of communities of small organisms is most readily done using PCR-based analysis of environmental samples consisting of mixtures of individuals. Known as metagenetics, this approach has transformed understanding of microbial communities and is beginning to be applied to metazoans as well. Unlike microbial studies, where analysis of the 16S ribosomal DNA sequence is standard, the best gene for metazoan metagenetics is less clear. In this study we designed a set of PCR primers for the mitochondrial 12S ribosomal DNA sequence based on 64 complete mitochondrial genomes and then tested their efficacy. METHODOLOGY/PRINCIPAL FINDINGS: A total of the 64 complete mitochondrial genome sequences representing all metazoan classes available in GenBank were downloaded using the NCBI Taxonomy Browser. Alignment of sequences was performed for the excised mitochondrial 12S ribosomal DNA sequences, and conserved regions were identified for all 64 mitochondrial genomes. These regions were used to design a primer pair that flanks a more variable region in the gene. Then all of the complete metazoan mitochondrial genomes available in NCBI's Organelle Genome Resources database were used to determine the percentage of taxa that would likely be amplified using these primers. Results suggest that these primers will amplify target sequences for many metazoans. CONCLUSIONS/SIGNIFICANCE: Newly designed 12S ribosomal DNA primers have considerable potential for metazoan metagenetic analysis because of their ability to amplify sequences from many metazoans.

  5. A general strategy to determine the congruence between a hierarchical and a non-hierarchical classification

    Directory of Open Access Journals (Sweden)

    Marín Ignacio

    2007-11-01

    Full Text Available Abstract Background Classification procedures are widely used in phylogenetic inference, the analysis of expression profiles, the study of biological networks, etc. Many algorithms have been proposed to establish the similarity between two different classifications of the same elements. However, methods to determine significant coincidences between hierarchical and non-hierarchical partitions are still poorly developed, in spite of the fact that the search for such coincidences is implicit in many analyses of massive data. Results We describe a novel strategy to compare a hierarchical and a dichotomic non-hierarchical classification of elements, in order to find clusters in a hierarchical tree in which elements of a given "flat" partition are overrepresented. The key improvement of our strategy respect to previous methods is using permutation analyses of ranked clusters to determine whether regions of the dendrograms present a significant enrichment. We show that this method is more sensitive than previously developed strategies and how it can be applied to several real cases, including microarray and interactome data. Particularly, we use it to compare a hierarchical representation of the yeast mitochondrial interactome and a catalogue of known mitochondrial protein complexes, demonstrating a high level of congruence between those two classifications. We also discuss extensions of this method to other cases which are conceptually related. Conclusion Our method is highly sensitive and outperforms previously described strategies. A PERL script that implements it is available at http://www.uv.es/~genomica/treetracker.

  6. A general strategy to determine the congruence between a hierarchical and a non-hierarchical classification.

    Science.gov (United States)

    Marco, Antonio; Marín, Ignacio

    2007-11-15

    Classification procedures are widely used in phylogenetic inference, the analysis of expression profiles, the study of biological networks, etc. Many algorithms have been proposed to establish the similarity between two different classifications of the same elements. However, methods to determine significant coincidences between hierarchical and non-hierarchical partitions are still poorly developed, in spite of the fact that the search for such coincidences is implicit in many analyses of massive data. We describe a novel strategy to compare a hierarchical and a dichotomic non-hierarchical classification of elements, in order to find clusters in a hierarchical tree in which elements of a given "flat" partition are overrepresented. The key improvement of our strategy respect to previous methods is using permutation analyses of ranked clusters to determine whether regions of the dendrograms present a significant enrichment. We show that this method is more sensitive than previously developed strategies and how it can be applied to several real cases, including microarray and interactome data. Particularly, we use it to compare a hierarchical representation of the yeast mitochondrial interactome and a catalogue of known mitochondrial protein complexes, demonstrating a high level of congruence between those two classifications. We also discuss extensions of this method to other cases which are conceptually related. Our method is highly sensitive and outperforms previously described strategies. A PERL script that implements it is available at http://www.uv.es/~genomica/treetracker.

  7. Refinements in hierarchical phrase-based translation systems

    OpenAIRE

    Pino, Juan Miguel

    2015-01-01

    The relatively recently proposed hierarchical phrase-based translation model for statistical machine translation (SMT) has achieved state-of-the-art performance in numerous recent translation evaluations. Hierarchical phrase-based systems comprise a pipeline of modules with complex interactions. In this thesis, we propose refinements to the hierarchical phrase-based model as well as improvements and analyses in various modules for hierarchical phrase-based systems. We too...

  8. [Commemorative lecture of receiving Imamura Memorial Prize. II. Mode of action of oligonucleotide fraction extracted from Mycobacterium bovis BCG].

    Science.gov (United States)

    Yamamoto, S

    1994-09-01

    A fraction extracted from Mycobacterium bovis BCG was found to exhibit strong antitumor activity. This fraction, which was designated MY-1, caused some animal tumors to regress and/or prevent metastasis very effectively. MY-1 after digestion with DNase had almost completely reduced activity, while MY-1 digested with RNase did not. MY-1 also augmented natural killer (NK) cell activity of mouse spleen cells in vitro, and produced factors which showed anti-viral activity and rendered macrophages cytotoxic towards tumor cells. The function of the factor to activate macrophages was destroyed by treatment with anti-interferon (IFN)-gamma antibody, while the anti-viral activity was destroyed by treatment with anti-INF alpha/beta antibody. The oligonucleotides contained in MY-1 distributed in a broad range of molecular size, and peaked at 45 nucleotides. We synthesized 13 kinds of 45-mer nucleotides with sequence present in the known cDNA encoding various BCG proteins. Six out of these oligonucleotides, which contained one or more hexameric palindromic structures, showed strong antitumor activity, while the other without palindrome did not. These active oligonucleotides possessed the capability to induce IFN and to augment NK cell activity of mouse spleen cells by coincubation in vitro. When a portion of the sequence of the inactive oligonucleotides was substituted with either palindromic sequence of GACGTC, AGCGCT or AACGTT, the oligonucleotide acquired the ability to augment NK activity. In contrast, the oligonucleotides substituted with another palindromic sequence such as ACCGGT was without effect. Furthermore, exchange of two neighboring mononucleotides within, but not outside, the active palindromic sequence destroyed the ability of the oligonucleotide to augment NK activity.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. Incorporating Usability Criteria into the Development of Animated Hierarchical Maps

    Science.gov (United States)

    Shih, Yu-Cheng; Huang, Pei-Ren; Chen, Sherry Y.

    2013-01-01

    Nowadays, Web-based learning systems have become popular because they can provide multiple tools, among which hierarchical maps are widely used to support teaching and learning. However, traditional hierarchical maps may let learners easily get lost within large information space. This study proposes an animated hierarchical map to address this…

  10. Microsatellite Primers for the Fungi Rhizopogon kretzerae and R. salebrosus (Rhizopogonaceae from 454 Shotgun Pyrosequencing

    Directory of Open Access Journals (Sweden)

    Lisa C. Grubisha

    2014-07-01

    Full Text Available Premise of the study: Rhizopogon kretzerae and R. salebrosus (Rhizopogonaceae are ectomycorrhizal fungi symbiotic with pines and the mycoheterotrophic plant Pterospora andromedea (Ericaceae. Microsatellite loci will allow population genetic study of fungal hosts to P. andromedea. Methods and Results: Shotgun pyrosequencing of R. kretzerae DNA resulted in primer development of 23 perfect microsatellite loci and screened across two populations each for R. kretzerae and R. salebrosus. Twelve loci were polymorphic in R. kretzerae populations, and 11 loci cross-amplified in R. salebrosus populations. For R. kretzerae and R. salebrosus, number of alleles was one to eight and one to nine, respectively, and observed heterozygosity ranged from 0.00–0.57 and 0.00–0.70, respectively. Conclusions: These are the first microsatellite loci developed for any species within Rhizopogon subgenus Amylopogon. These microsatellite loci will be used in conservation genetic studies of rare to endangered eastern populations and to compare plant and fungal population genetic structure at different hierarchical levels.

  11. New cyt b gene universal primer set for forensic analysis.

    Science.gov (United States)

    Lopez-Oceja, A; Gamarra, D; Borragan, S; Jiménez-Moreno, S; de Pancorbo, M M

    2016-07-01

    Analysis of mitochondrial DNA, and in particular the cytochrome b gene (cyt b), has become an essential tool for species identification in routine forensic practice. In cases of degraded samples, where the DNA is fractionated, universal primers that are highly efficient for the amplification of the target region are necessary. Therefore, in the present study a new universal cyt b primer set with high species identification capabilities, even in samples with highly degraded DNA, has been developed. In order to achieve this objective, the primers were designed following the alignment of complete sequences of the cyt b from 751 species from the Class of Mammalia listed in GenBank. A highly variable region of 148bp flanked by highly conserved sequences was chosen for placing the primers. The effectiveness of the new pair of primers was examined in 63 animal species belonging to 38 Families from 14 Orders and 5 Classes (Mammalia, Aves, Reptilia, Actinopterygii, and Malacostraca). Species determination was possible in all cases, which shows that the fragment analyzed provided a high capability for species identification. Furthermore, to ensure the efficiency of the 148bp fragment, the intraspecific variability was analyzed by calculating the concordance between individuals with the BLAST tool from the NCBI (National Center for Biotechnological Information). The intraspecific concordance levels were superior to 97% in all species. Likewise, the phylogenetic information from the selected fragment was confirmed by obtaining the phylogenetic tree from the sequences of the species analyzed. Evidence of the high power of phylogenetic discrimination of the analyzed fragment of the cyt b was obtained, as 93.75% of the species were grouped within their corresponding Orders. Finally, the analysis of 40 degraded samples with small-size DNA fragments showed that the new pair of primers permits identifying the species, even when the DNA is highly degraded as it is very common in

  12. RUCS: rapid identification of PCR primers for unique core sequences.

    Science.gov (United States)

    Thomsen, Martin Christen Frølund; Hasman, Henrik; Westh, Henrik; Kaya, Hülya; Lund, Ole

    2017-12-15

    Designing PCR primers to target a specific selection of whole genome sequenced strains can be a long, arduous and sometimes impractical task. Such tasks would benefit greatly from an automated tool to both identify unique targets, and to validate the vast number of potential primer pairs for the targets in silico. Here we present RUCS, a program that will find PCR primer pairs and probes for the unique core sequences of a positive genome dataset complement to a negative genome dataset. The resulting primer pairs and probes are in addition to simple selection also validated through a complex in silico PCR simulation. We compared our method, which identifies the unique core sequences, against an existing tool called ssGeneFinder, and found that our method was 6.5-20 times more sensitive. We used RUCS to design primer pairs that would target a set of genomes known to contain the mcr-1 colistin resistance gene. Three of the predicted pairs were chosen for experimental validation using PCR and gel electrophoresis. All three pairs successfully produced an amplicon with the target length for the samples containing mcr-1 and no amplification products were produced for the negative samples. The novel methods presented in this manuscript can reduce the time needed to identify target sequences, and provide a quick virtual PCR validation to eliminate time wasted on ambiguously binding primers. Source code is freely available on https://bitbucket.org/genomicepidemiology/rucs. Web service is freely available on https://cge.cbs.dtu.dk/services/RUCS. mcft@cbs.dtu.dk. Supplementary data are available at Bioinformatics online.

  13. Identification of GalNAc-Conjugated Antisense Oligonucleotide Metabolites Using an Untargeted and Generic Approach Based on High Resolution Mass Spectrometry.

    Science.gov (United States)

    Husser, Christophe; Brink, Andreas; Zell, Manfred; Müller, Martina B; Koller, Erich; Schadt, Simone

    2017-06-20

    Antisense oligonucleotides linked by phosphorothioates are an important class of therapeutics under investigation in various pharmaceutical companies. Antisense oligonucleotides may be coupled to high-affinity ligands (triantennary N-acetyl galactosamine = GalNAc) for hepatocyte-specific asialoglycoprotein receptors (ASGPR) to enhance uptake to hepatocytes and to increase potency. Since disposition and biotransformation of GalNAc-conjugated oligonucleotides is different from unconjugated oligonucleotides, appropriate analytical methods are required to identify main cleavage sites and degradation products of GalNAc conjugated and unconjugated oligonucleotides in target cells. A highly sensitive method was developed to identify metabolites of oligonucleotides using capillary flow liquid chromatography with column switching coupled to a high resolution Orbitrap Fusion mass spectrometer. Detection of GalNAc-conjugated oligonucleotides and their metabolites was achieved by combining full scan MS with two parallel MS2 experiments, one data-dependent scan and an untargeted MS2 experiment (all ion fragmentation) applying high collision energy. In the all ion fragmentation scan, a diagnostic fragment originating from the phosphorothioate backbone (O2PS-: m/z 94.936) was formed efficiently upon collisional activation. Based on this fragment an accurate determination of metabolites of oligonucleotides was achieved, independent of their sequence or conjugation in an untargeted but highly selective manner. The method was effectively applied to investigate uptake and metabolism of GalNAc-conjugated oligonucleotides in incubations of primary rat hepatocytes; the elucidation of expected and unexpected degradation products was achieved in subnanomolar range.

  14. Preselection of shotgun clones by oligonucleotide fingerprinting: an efficient and high throughput strategy to reduce redundancy in large-scale sequencing projects

    National Research Council Canada - National Science Library

    Radelof, U; Hennig, S; Seranski, P; Steinfath, M; Ramser, J; Reinhardt, R; Poustka, A; Francis, F; Lehrach, H

    1998-01-01

    .... To reduce the overall effort and cost of those projects and to accelerate the sequencing throughput, we have developed an efficient, high throughput oligonucleotide fingerprinting protocol to select...

  15. Sequence-dependent structural changes in a self-assembling DNA oligonucleotide.

    Science.gov (United States)

    Saoji, Maithili; Paukstelis, Paul J

    2015-12-01

    DNA has proved to be a remarkable molecule for the construction of sophisticated two-dimensional and three-dimensional architectures because of its programmability and structural predictability provided by complementary Watson-Crick base pairing. DNA oligonucleotides can, however, exhibit a great deal of local structural diversity. DNA conformation is strongly linked to both environmental conditions and the nucleobase identities inherent in the oligonucleotide sequence, but the exact relationship between sequence and local structure is not completely understood. This study examines how a single-nucleotide addition to a class of self-assembling DNA 13-mers leads to a significantly different overall structure under identical crystallization conditions. The DNA 13-mers self-assemble in the presence of Mg(2+) through a combination of Watson-Crick and noncanonical base-pairing interactions. The crystal structures described here show that all of the predicted Watson-Crick base pairs are present, with the major difference being a significant rearrangement of noncanonical base pairs. This includes the formation of a sheared A-G base pair, a junction of strands formed from base-triple interactions, and tertiary interactions that generate structural features similar to tandem sheared G-A base pairs. The adoption of this alternate noncanonical structure is dependent in part on the sequence in the Watson-Crick duplex region. These results provide important new insights into the sequence-structure relationship of short DNA oligonucleotides and demonstrate a unique interplay between Watson-Crick and noncanonical base pairs that is responsible for crystallization fate.

  16. Development and production of an oligonucleotide MuscleChip: use for validation of ambiguous ESTs

    Directory of Open Access Journals (Sweden)

    Lanfranchi Gerolamo

    2002-10-01

    Full Text Available Abstract Background We describe the development, validation, and use of a highly redundant 120,000 oligonucleotide microarray (MuscleChip containing 4,601 probe sets representing 1,150 known genes expressed in muscle and 2,075 EST clusters from a non-normalized subtracted muscle EST sequencing project (28,074 EST sequences. This set included 369 novel EST clusters showing no match to previously characterized proteins in any database. Each probe set was designed to contain 20–32 25 mer oligonucleotides (10–16 paired perfect match and mismatch probe pairs per gene, with each probe evaluated for hybridization kinetics (Tm and similarity to other sequences. The 120,000 oligonucleotides were synthesized by photolithography and light-activated chemistry on each microarray. Results Hybridization of human muscle cRNAs to this MuscleChip (33 samples showed a correlation of 0.6 between the number of ESTs sequenced in each cluster and hybridization intensity. Out of 369 novel EST clusters not showing any similarity to previously characterized proteins, we focused on 250 EST clusters that were represented by robust probe sets on the MuscleChip fulfilling all stringent rules. 102 (41% were found to be consistently "present" by analysis of hybridization to human muscle RNA, of which 40 ESTs (39% could be genome anchored to potential transcription units in the human genome sequence. 19 ESTs of the 40 ESTs were furthermore computer-predicted as exons by one or more than three gene identification algorithms. Conclusion Our analysis found 40 transcriptionally validated, genome-anchored novel EST clusters to be expressed in human muscle. As most of these ESTs were low copy clusters (duplex and triplex in the original 28,000 EST project, the identification of these as significantly expressed is a robust validation of the transcript units that permits subsequent focus on the novel proteins encoded by these genes.

  17. Simplified Paper Format for Detecting HIV Drug Resistance in Clinical Specimens by Oligonucleotide Ligation.

    Directory of Open Access Journals (Sweden)

    Nuttada Panpradist

    Full Text Available Human immunodeficiency virus (HIV is a chronic infection that can be managed by antiretroviral treatment (ART. However, periods of suboptimal viral suppression during lifelong ART can select for HIV drug resistant (DR variants. Transmission of drug resistant virus can lessen or abrogate ART efficacy. Therefore, testing of individuals for drug resistance prior to initiation of treatment is recommended to ensure effective ART. Sensitive and inexpensive HIV genotyping methods are needed in low-resource settings where most HIV infections occur. The oligonucleotide ligation assay (OLA is a sensitive point mutation assay for detection of drug resistance mutations in HIV pol. The current OLA involves four main steps from sample to analysis: (1 lysis and/or nucleic acid extraction, (2 amplification of HIV RNA or DNA, (3 ligation of oligonucleotide probes designed to detect single nucleotide mutations that confer HIV drug resistance, and (4 analysis via oligonucleotide surface capture, denaturation, and detection (CDD. The relative complexity of these steps has limited its adoption in resource-limited laboratories. Here we describe a simplification of the 2.5-hour plate-format CDD to a 45-minute paper-format CDD that eliminates the need for a plate reader. Analysis of mutations at four HIV-1 DR codons (K103N, Y181C, M184V, and G190A in 26 blood specimens showed a strong correlation of the ratios of mutant signal to total signal between the paper CDD and the plate CDD. The assay described makes the OLA easier to perform in low resource laboratories.

  18. Effect of Terminal Groups of Dendrimers in the Complexation with Antisense Oligonucleotides and Cell Uptake

    Science.gov (United States)

    Márquez-Miranda, Valeria; Peñaloza, Juan Pablo; Araya-Durán, Ingrid; Reyes, Rodrigo; Vidaurre, Soledad; Romero, Valentina; Fuentes, Juan; Céric, Francisco; Velásquez, Luis; González-Nilo, Fernando D.; Otero, Carolina

    2016-02-01

    Poly(amidoamine) dendrimers are the most recognized class of dendrimer. Amino-terminated (PAMAM-NH2) and hydroxyl-terminated (PAMAM-OH) dendrimers of generation 4 are widely used, since they are commercially available. Both have different properties, mainly based on their different overall charges at physiological pH. Currently, an important function of dendrimers as carriers of short single-stranded DNA has been applied. These molecules, known as antisense oligonucleotides (asODNs), are able to inhibit the expression of a target mRNA. Whereas PAMAM-NH2 dendrimers have shown to be able to transfect plasmid DNA, PAMAM-OH dendrimers have not shown the same successful results. However, little is known about their interaction with shorter and more flexible molecules such as asODNs. Due to several initiatives, the use of these neutral dendrimers as a scaffold to introduce other functional groups has been proposed. Because of its low cytotoxicity, it is relevant to understand the molecular phenomena involving these types of dendrimers. In this work, we studied the behavior of an antisense oligonucleotide in presence of both types of dendrimers using molecular dynamics simulations, in order to elucidate if they are able to form stable complexes. In this manner, we demonstrated at atomic level that PAMAM-NH2, unlike PAMAM-OH, could form a well-compacted complex with asODN, albeit PAMAM-OH can also establish stable interactions with the oligonucleotide. The biological activity of asODN in complex with PAMAM-NH2 dendrimer was also shown. Finally, we revealed that in contact with PAMAM-OH, asODN remains outside the cells as TIRF microscopy results showed, due to its poor interaction with this dendrimer and cell membranes.

  19. Analysis of oligonucleotide array experiments with repeated measures using mixed models

    Directory of Open Access Journals (Sweden)

    Getchell Thomas V

    2004-12-01

    Full Text Available Abstract Background Two or more factor mixed factorial experiments are becoming increasingly common in microarray data analysis. In this case study, the two factors are presence (Patients with Alzheimer's disease or absence (Control of the disease, and brain regions including olfactory bulb (OB or cerebellum (CER. In the design considered in this manuscript, OB and CER are repeated measurements from the same subject and, hence, are correlated. It is critical to identify sources of variability in the analysis of oligonucleotide array experiments with repeated measures and correlations among data points have to be considered. In addition, multiple testing problems are more complicated in experiments with multi-level treatments or treatment combinations. Results In this study we adopted a linear mixed model to analyze oligonucleotide array experiments with repeated measures. We first construct a generalized F test to select differentially expressed genes. The Benjamini and Hochberg (BH procedure of controlling false discovery rate (FDR at 5% was applied to the P values of the generalized F test. For those genes with significant generalized F test, we then categorize them based on whether the interaction terms were significant or not at the α-level (αnew = 0.0033 determined by the FDR procedure. Since simple effects may be examined for the genes with significant interaction effect, we adopt the protected Fisher's least significant difference test (LSD procedure at the level of αnew to control the family-wise error rate (FWER for each gene examined. Conclusions A linear mixed model is appropriate for analysis of oligonucleotide array experiments with repeated measures. We constructed a generalized F test to select differentially expressed genes, and then applied a specific sequence of tests to identify factorial effects. This sequence of tests applied was designed to control for gene based FWER.

  20. Development and validation of an oligonucleotide microarray to characterise ectomycorrhizal fungal communities

    Directory of Open Access Journals (Sweden)

    Reich Marlis

    2009-01-01

    Full Text Available Abstract Background In forest ecosystems, communities of ectomycorrhizal fungi (ECM are influenced by several biotic and abiotic factors. To understand their underlying dynamics, ECM communities have been surveyed with ribosomal DNA-based sequencing methods. However, most identification methods are both time-consuming and limited by the number of samples that can be treated in a realistic time frame. As a result of ongoing implementation, the array technique has gained throughput capacity in terms of the number of samples and the capacity for parallel identification of several species. Thus far, although phylochips (microarrays that are used to detect species have been mostly developed to trace bacterial communities or groups of specific fungi, no phylochip has been developed to carry oligonucleotides for several ectomycorrhizal species that belong to different genera. Results We have constructed a custom ribosomal DNA phylochip to identify ECM fungi. Specific oligonucleotide probes were targeted to the nuclear internal transcribed spacer (ITS regions from 95 fungal species belonging to 21 ECM fungal genera. The phylochip was first validated using PCR amplicons of reference species. Ninety-nine percent of the tested oligonucleotides generated positive hybridisation signals with their corresponding amplicons. Cross-hybridisation was mainly restricted at the genus level, particularly for Cortinarius and Lactarius species. The phylochip was subsequently tested with environmental samples that were composed of ECM fungal DNA from spruce and beech plantation fungal communities. The results were in concordance with the ITS sequencing of morphotypes and the ITS clone library sequencing results that were obtained using the same PCR products. Conclusion For the first time, we developed a custom phylochip that is specific for several ectomycorrhizal fungi. To overcome cross-hybridisation problems, specific filter and evaluation strategies that used spot

  1. Ultrasensitive electrochemical biosensor based on the oligonucleotide self-assembled monolayer-mediated immunosensing interface

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Dengyou; Luo, Qimei [Science College of Hunan Agricultural University, Changsha 410128 (China); Deng, Fawen [The Fourth Hospital of Chansha, Changsha 410006 (China); Li, Zhen [Science College of Hunan Agricultural University, Changsha 410128 (China); Li, Benxiang, E-mail: 172170960@qq.com [Science College of Hunan Agricultural University, Changsha 410128 (China); Shen, Zhifa, E-mail: shenzhifa@wmu.edu.cn [Key Laboratory of Laboratory Medicine, Ministry of Education, Zhejiang Provincial Key Laboratory of Medical Genetics, College of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035 (China)

    2017-06-08

    Highly sensitive and selective quantitation of a variety of proteins over a wide concentration range is highly desirable for increased accuracy of biomarker detection or for multidisease diagnostics. In the present contribution, using human immunoglobulin G (HIgG) as the model target protein, an electrochemical ultrasensitive immunosensing platform was developed based on the oligonucleotide self-assembled monolayer-mediated (OSAM) sensing interface. For this immunosensor, the “signal-on” signaling mechanism and enzymatic signal amplification effect were integrated into one sensing architecture. Moreover, the thiolated flexible single-stranded DNAs immobilized onto gold electrode surface not only performs the wobbling motion to facilitate the electron transfer between the electrode surface and biosensing layer but also fundamentally prohibiting the direct interaction of proteins with gold substrate. Thus, the electrochemical signal could be efficiently enhanced and the unspecific adsorption or cross-reaction might be eliminated. As a result, utilizing the newly-proposed immunosensor, the HIgG can be detected down to 0.5 ng/mL, and the high detection specificity is offered. The successful design of OSAM and the highly desirable detection capability of new immunosensor are expected to provide a perspective for fabricating new robust immunosensing platform and for promising potential of oligonucleotide probe in biological research and biomedical diagnosis. - Highlights: • An electrochemical ultrasensitive immunosensing platform was developed based on the oligonucleotide self-assembled monolayer (OASM). • OASM severs as a flexible monolayer to promote electron transfer and prohibits the direct interaction of proteins with gold substrate. • The electrochemical signal is efficiently enhanced and the unspecific adsorption or cross-reaction is eliminated. • Target protein can be detected down to 0.5 ng/mL, and the high detection specificity can be obtained.

  2. Detection of cyclin D1 mRNA by hybridization sensitive NIC-oligonucleotide probe.

    Science.gov (United States)

    Kovaliov, Marina; Segal, Meirav; Kafri, Pinhas; Yavin, Eylon; Shav-Tal, Yaron; Fischer, Bilha

    2014-05-01

    A large group of fluorescent hybridization probes, includes intercalating dyes for example thiazole orange (TO). Usually TO is coupled to nucleic acids post-synthetically which severely limits its use. Here, we have developed a phosphoramidite monomer, 10, and prepared a 2'-OMe-RNA probe, labeled with 5-(trans-N-hexen-1-yl-)-TO-2'-deoxy-uridine nucleoside, dU(TO), (Nucleoside bearing an Inter-Calating moiety, NIC), for selective mRNA detection. We investigated a series of 15-mer 2'-OMe-RNA probes, targeting the cyclin D1 mRNA, containing one or several dU(TO) at various positions. dU(TO)-2'-OMe-RNA exhibited up to 7-fold enhancement of TO emission intensity upon hybridization with the complementary RNA versus that of the oligomer alone. This NIC-probe was applied for the specific detection of a very small amount of a breast cancer marker, cyclin D1 mRNA, in total RNA extract from cancerous cells (250 ng/μl). Furthermore, this NIC-probe was found to be superior to our related NIF (Nucleoside with Intrinsic Fluorescence)-probe which could detect cyclin D1 mRNA target only at high concentrations (1840 ng/μl). Additionally, dU(T) can be used as a monomer in solid-phase oligonucleotide synthesis, thus avoiding the need for post-synthetic modification of oligonucleotide probes. Hence, we propose dU(TO) oligonucleotides, as hybridization probes for the detection of specific RNA in homogeneous solutions and for the diagnosis of breast cancer. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Object tracking with hierarchical multiview learning

    Science.gov (United States)

    Yang, Jun; Zhang, Shunli; Zhang, Li

    2016-09-01

    Building a robust appearance model is useful to improve tracking performance. We propose a hierarchical multiview learning framework to construct the appearance model, which has two layers for tracking. On the top layer, two different views of features, grayscale value and histogram of oriented gradients, are adopted for representation under the cotraining framework. On the bottom layer, for each view of each feature, three different random subspaces are generated to represent the appearance from multiple views. For each random view submodel, the least squares support vector machine is employed to improve the discriminability for concrete and efficient realization. These two layers are combined to construct the final appearance model for tracking. The proposed hierarchical model assembles two types of multiview learning strategies, in which the appearance can be described more accurately and robustly. Experimental results in the benchmark dataset demonstrate that the proposed method can achieve better performance than many existing state-of-the-art algorithms.

  4. Hierarchical Self-organization of Complex Systems

    Institute of Scientific and Technical Information of China (English)

    CHAI Li-he; WEN Dong-sheng

    2004-01-01

    Researches on organization and structure in complex systems are academic and industrial fronts in modern sciences. Though many theories are tentatively proposed to analyze complex systems, we still lack a rigorous theory on them. Complex systems possess various degrees of freedom, which means that they should exhibit all kinds of structures. However, complex systems often show similar patterns and structures. Then the question arises why such similar structures appear in all kinds of complex systems. The paper outlines a theory on freedom degree compression and the existence of hierarchical self-organization for all complex systems is found. It is freedom degree compression and hierarchical self-organization that are responsible for the existence of these similar patterns or structures observed in the complex systems.

  5. Application of hierarchical matrices for partial inverse

    KAUST Repository

    Litvinenko, Alexander

    2013-11-26

    In this work we combine hierarchical matrix techniques (Hackbusch, 1999) and domain decomposition methods to obtain fast and efficient algorithms for the solution of multiscale problems. This combination results in the hierarchical domain decomposition (HDD) method, which can be applied for solution multi-scale problems. Multiscale problems are problems that require the use of different length scales. Using only the finest scale is very expensive, if not impossible, in computational time and memory. Domain decomposition methods decompose the complete problem into smaller systems of equations corresponding to boundary value problems in subdomains. Then fast solvers can be applied to each subdomain. Subproblems in subdomains are independent, much smaller and require less computational resources as the initial problem.

  6. Hierarchical photo stream segmentation using context

    Science.gov (United States)

    Gong, Bo; Jain, Ramesh

    2008-01-01

    Photo stream segmentation is to segment photo streams into groups, each of which corresponds to an event. Photo stream segmentation can be done with or without prior knowledge of event structure. In this paper, we study the problem by assuming that there is no a priori event model available. Although both context and content information are important for photo stream segmentation, we focus on investigating the usage of context information in this work. We consider different information components of context such as time, location, and optical setting for inexpensive segmentation of photo streams from common users of modern digital camera. As events are hierarchical, we propose to segment photo stream using hierarchical mixture model. We compare the generated hierarchy with that created by users to see how well results can be obtained without knowing the prior event model. We experimented with about 3000 photos from amateur photographers to study the efficacy of the approach for these context information components.

  7. Hierarchical silica particles by dynamic multicomponent assembly

    DEFF Research Database (Denmark)

    Wu, Z. W.; Hu, Q. Y.; Pang, J. B.

    2005-01-01

    Abstract: Aerosol-assisted assembly of mesoporous silica particles with hierarchically controllable pore structure has been prepared using cetyltrimethylammonium bromide (CTAB) and poly(propylene oxide) (PPO, H[OCH(CH3)CH2],OH) as co-templates. Addition of the hydrophobic PPO significantly...... influences the delicate hydrophilic-hydrophobic balance in the well-studied CTAB-silicate co-assembling system, resulting in various mesostructures (such as hexagonal, lamellar, and hierarchical structure). The co-assembly of CTAB, silicate clusters, and a low-molecular-weight PPO (average M-n 425) results......-silicate assembling system was discussed. The mesostructure of these particles was characterized by transmission electron microscope (TEM), scanning electron microscope (SEM), X-ray diffraction (XRD), and N-2 sorption. (c) 2005 Elsevier Inc. All rights reserved....

  8. Information Limited Oligonucleotide Amplification Assay for Affinity-Based, Parallel Detection Studies.

    Directory of Open Access Journals (Sweden)

    Harish Bokkasam

    Full Text Available Molecular communication systems encounter similar constraints as telecommunications. In either case, channel crosstalk at the receiver end will result in information loss that statistical analysis cannot compensate. This is because in any communication channel there is a physical limit to the amount of information that can be transmitted. We present a novel and simple modified end amplification (MEA technique to generate reduced and defined amounts of specific information in form of short fragments from an oligonucleotide source that also contains unrelated and redundant information. Our method can be a valuable tool to investigate information overflow and channel capacity in biomolecular recognition systems.

  9. Pressure refilled polyacrylamide columns for the separation of oligonucleotides by capillary electrophoresis.

    Science.gov (United States)

    Sudor, J; Foret, F; Bocek, P

    1991-12-01

    The separation of oligonucleotides by capillary zone electrophoresis (CZE) was studied in fused silica separation capillaries filled by linear (noncrosslinked) polyacrylamide (PAA) solutions, introduced into the capillary from the stock by pressure after each analysis. The time-consuming in-capillary polymerization step could thus be avoided, and fast and reproducible repetition of the analyses was assured. The PAA concentrations varied within the range of 3-10% and both the reproducibility of the analyses and the stability of the solution in the capillary, with and without a chemically treated inner wall, were tested. Ferguson plots were used to assess the size selectivity of the separation.

  10. Chemosensitization of Human Renal Cell Cancer Using Antisense Oligonucleotides Targeting the Antiapoptotic Gene Clusterin

    Directory of Open Access Journals (Sweden)

    Tobias Zellweger

    2001-01-01

    Full Text Available BACKGROUND: Renal cell cancer (RCC is a chemoresistant disease with no active chemotherapeutic agent achieving objective response rates higher than 15%. Clusterin is a cell survival gene that increases in human renal tubular epithelial cells after various states of injury and disease. Downregulation of clusterin, using antisense oligonucleotides (ASO, has recently been shown to increase chemosensitivity in several prostate cancer models. The objectives in this study were to evaluate clusterin expression levels in human RCC and normal kidney tissue, and to test whether clusterin ASO could also enhance chemosensitivity in human RCC Caki-2 cells both in vitro and in vivo. METHODS: Immunohistochemical staining was used to characterize clusterin expression in 67 RCC and normal kidney tissues obtained from radical nephrectomy specimens. Northern blot analysis was used to assess changes in clusterin mRNA expression after ASO and paclitaxel treatment. The effects of combined clusterin ASO and paclitaxel treatment on Caki-2 cell growth was examined using an MTT assay. Athymic mice bearing Caki-2 tumors were treated with clusterin ASO alone, clusterin ASO plus paclitaxel, and mismatch control oligonucleotides plus paclitaxel, over a period of 28 days with measurement of tumor volumes once weekly over 8 weeks. RESULTS: Immunohistochemistry of normal and malignant kidney tissue sections of 67 patients demonstrated positive clusterin staining for almost all RCC (98% and an overexpression, compared to normal tissue, in a majority of RCC (69%. Clusterin ASO, but not mismatch control oligonucleotides, decreased clusterin mRNA expression in Caki-2 cells in a dosedependent and sequence-specific manner. Pretreatment of Caki-2 cells with clusterin ASO significantly enhanced chemosensitivity to paclitaxel in vitro. Characteristic apoptotic DNA laddering was observed after combined treatment with ASO plus paclitaxel, but not with either agent alone. In vivo

  11. Polycation Induced Potential Dependent Structural Transitions of Oligonucleotide Monolayers on Au(111)-Surfaces

    DEFF Research Database (Denmark)

    Salvatore, Princia; Karlsen, Kasper Kannegård; Hansen, Allan Glargaard

    2012-01-01

    We have studied self-assembled molecular monolayers (SAMs) of several 3′-C3-SH conjugated single-strand (ss) and double-strand (ds) 20-base oligonucleotides (ONs) immobilized on single-crystal, atomically planar Au(111)-electrode surfaces in the presence of the triply positively charged base...... electrochemical potential control was used. Spd binding was found to increase notably the ds-ON melting temperature. CV displays capacitive features associated with ss- and ds-ON. A robust capacitive peak around −0.35 V versus saturated calomel electrode (SCE), specific to ds-ON and highly sensitive to base pair...

  12. A New Achiral Linker Reagent for the Incorporation of Multiple Amino Groups Into Oligonucleotides

    DEFF Research Database (Denmark)

    1997-01-01

    The present invention relates to a new functionalized achiral linker reagent for incorporating multiple primary amino groups or reporter groups into oligonucleotides following the phosphoramidite methodology. It is possible to substitute any ribodeoxynucleotide, deoxynucleotide, or nucleotide......-oxyl-2,2,5,5-tetramethylpyrrolidine), TEMPO (N-oxyl-2,2,6,6-tetramethylpiperidine), dinitrophenyl, texas red, tetramethyl rhodamine, 7-nitrobenzo-2-oxa-1-diazole (NBD), or pyrene. The present invention also relates to a solid phase support, e.g. a Controlled Pore Glass (CPG), immobilized linker reagent...

  13. Synthetic oligonucleotide antigens modified with locked nucleic acids detect disease specific antibodies

    DEFF Research Database (Denmark)

    Samuelsen, Simone V; Solov'yov, Ilia A.; Balboni, Imelda M.

    2016-01-01

    molecular dynamics we predict optimal nucleotide composition for targeting DNA-binding antibodies. As a proof of concept, we address a problem of detecting anti-DNA antibodies that are characteristic of systemic lupus erythematosus, a chronic autoimmune disease with multiple manifestations. We test the best...... oligonucleotide binders in surface plasmon resonance studies to analyze binding and kinetic aspects of interactions between antigens and target DNA. These DNA and LNA/DNA sequences showed improved binding in enzyme-linked immunosorbent assay using human samples of pediatric lupus patients. Our results suggest...

  14. Hierarchical Fuzzy Sets To Query Possibilistic Databases

    OpenAIRE

    Thomopoulos, Rallou; Buche, Patrice; Haemmerlé, Ollivier

    2008-01-01

    Within the framework of flexible querying of possibilistic databases, based on the fuzzy set theory, this chapter focuses on the case where the vocabulary used both in the querying language and in the data is hierarchically organized, which occurs in systems that use ontologies. We give an overview of previous works concerning two issues: firstly, flexible querying of imprecise data in the relational model; secondly, the introduction of fuzziness in hierarchies. Concerning the latter point, w...

  15. Hierarchically Structured Nanomaterials for Electrochemical Energy Conversion

    OpenAIRE

    Trogadas, P.; Ramani, V; Strasser, P.; T.F. Fuller; Coppens, M. O.

    2016-01-01

    © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Hierarchical nanomaterials are highly suitable as electrocatalysts and electrocatalyst supports in electrochemical energy conversion devices. The intrinsic kinetics of an electrocatalyst are associated with the nanostructure of the active phase and the support, while the overall properties are also affected by the mesostructure. Therefore, both structures need to be controlled. A comparative state-of-the-art review of catalysts and supports i...

  16. Pattern Recognition by Hierarchical Temporal Memory

    OpenAIRE

    Maltoni, Prof. Davide

    2011-01-01

    Hierarchical Temporal Memory (HTM) is still largely unknown by the pattern recognition community and only a few studies have been published in the scientific literature. This paper reviews HTM architecture and related learning algorithms by using formal notation and pseudocode description. Novel approaches are then proposed to encode coincidence-group membership (fuzzy grouping) and to derive temporal groups (maxstab temporal clustering). Systematic experiments on three line-drawing datasets ...

  17. Internet advertising effectiveness by using hierarchical model

    OpenAIRE

    Rahmani, Samaneh

    2015-01-01

    Abstract. Present paper has been developed with the title of internet advertising effectiveness by using hierarchical model. Presenting the question: Today Internet is an important channel in marketing and advertising. The reason for this could be the ability of the Internet to reduce costs and people’s access to online services[1]. Also advertisers can easily access a multitude of users and communicate with them at low cost [9]. On the other hand, compared to traditional advertising, interne...

  18. Predicting protein functions using incomplete hierarchical labels.

    Science.gov (United States)

    Yu, Guoxian; Zhu, Hailong; Domeniconi, Carlotta

    2015-01-16

    Protein function prediction is to assign biological or biochemical functions to proteins, and it is a challenging computational problem characterized by several factors: (1) the number of function labels (annotations) is large; (2) a protein may be associated with multiple labels; (3) the function labels are structured in a hierarchy; and (4) the labels are incomplete. Current predictive models often assume that the labels of the labeled proteins are complete, i.e. no label is missing. But in real scenarios, we may be aware of only some hierarchical labels of a protein, and we may not know whether additional ones are actually present. The scenario of incomplete hierarchical labels, a challenging and practical problem, is seldom studied in protein function prediction. In this paper, we propose an algorithm to Predict protein functions using Incomplete hierarchical LabeLs (PILL in short). PILL takes into account the hierarchical and the flat taxonomy similarity between function labels, and defines a Combined Similarity (ComSim) to measure the correlation between labels. PILL estimates the missing labels for a protein based on ComSim and the known labels of the protein, and uses a regularization to exploit the interactions between proteins for function prediction. PILL is shown to outperform other related techniques in replenishing the missing labels and in predicting the functions of completely unlabeled proteins on publicly available PPI datasets annotated with MIPS Functional Catalogue and Gene Ontology labels. The empirical study shows that it is important to consider the incomplete annotation for protein function prediction. The proposed method (PILL) can serve as a valuable tool for protein function prediction using incomplete labels. The Matlab code of PILL is available upon request.

  19. Hierarchical mixtures of naive Bayes classifiers

    OpenAIRE

    Wiering, M.A.

    2002-01-01

    Naive Bayes classifiers tend to perform very well on a large number of problem domains, although their representation power is quite limited compared to more sophisticated machine learning algorithms. In this pa- per we study combining multiple naive Bayes classifiers by using the hierar- chical mixtures of experts system. This system, which we call hierarchical mixtures of naive Bayes classifiers, is compared to a simple naive Bayes classifier and to using bagging and boosting for combining ...

  20. MANAJEMEN BANDWIDTH MENGGUNAKAN METODE HIERARCHICAL TOKEN BUCKET

    Directory of Open Access Journals (Sweden)

    Lisnawita Lisnawita

    2016-02-01

    Full Text Available AbstrakInternet pada era sekarang ini adalah salah satu kebutuhan manusia, karena itu dibutuhkan bandwidth yang besar supaya bisa download atau upload dengan cepat, tapi karena keterbatasan bandwidth yang ada, maka perlu Manajemen bandwidth dengan metode Hierarchical Token Bucket(HTB, metode ini banyak digunakan untuk mengatasi permasalahan pada koneksi Internet, memaksimalkan penggunaan bandwidth sehingga semua user dapat menggunakan bandwidth secara adil dan semua user mendapatkan kenyamanan dan kepuasan ketika browsing, Penelitian ini menggunakan metode Hierarchical Token Bucket(HTB yaitu salah satu metode yang dirancang untuk bisa melakukan manajemen bandwidth dengan baik, dimana algoritma ini menerapkan disiplin antrian yang mempunyai kelebihan dalam pembatasan trafik pada tiap level maupun klasifikasi, sehingga bandwidth yang tidak dipakai oleh level yang tinggi dapat digunakan atau dibagi oleh level yang lebih rendah.Kata Kunci: Bandwidth,Manajemen Bandwidth,HTBAbstractInternet in this era is one of human needs, because it takes a large bandwid thin order to download or upload quickly,but due to the limited bandwidth available, the necessary management bandwidth by using Hierarchical Token Bucket(HTB, this method is widely used for solve the problems on Internet connections, maximizing the use of bandwidth so that all users can use the bandwidth fairly and users get the comfort and satisfaction when browsing, this researchusing Hierarchical Token Bucket(HTB is one method that is designed to be able to perform bandwidth management,where the algorithm queuing discipline which have advantage sin traffic restriction soneach level and classification, so that the bandwidth is not used by a high level can be used or share by a low levelKeywords: Bandwidth, Bandwidth Management, HTB

  1. Hierarchical Parallel Evaluation of a Hamming Code

    Directory of Open Access Journals (Sweden)

    Shmuel T. Klein

    2017-04-01

    Full Text Available The Hamming code is a well-known error correction code and can correct a single error in an input vector of size n bits by adding logn parity checks. A new parallel implementation of the code is presented, using a hierarchical structure of n processors in logn layers. All the processors perform similar simple tasks, and need only a few bytes of internal memory.

  2. Factors influencing the electrokinetic injection of oligonucleotides in capillary gel electrophoresis when using laser-induced fluorescence detection.

    Science.gov (United States)

    Chen, Buyun; Chen, Guanhua; Bartlett, Michael G

    2014-03-01

    Capillary gel electrophoresis (CGE) is a powerful tool for the analysis of oligonucleotides owing to its extraordinary resolving power. However, the only feasible injection mode for CGE, electrokinetic injection, can cause bias of the injected amount and thus reproducibility issues for CGE methods. Although the source of the bias in electrokinetic injection for analysis of small molecules by capillary zone electrophoresis has long been identified, there are very few studies on electrokinetic injection issues for biological molecules analyzed by CGE. In this study, we report three issues related to electrokinetic injection for oligonucleotides. First, the relationship between the injection amount and the sample solution resistance is not always linear for oligonucleotides, as has been observed for small molecules. Second, the injecting water prior to an oligonucleotide sample dramatically improves the reproducibility of both the injected amount and resolution through a 'stacking-like' mechanism. Third, optimizing the gel concentration dramatically increases the amount of oligonucleotide that is injected into the column. Copyright © 2013 John Wiley & Sons, Ltd.

  3. Resolving prokaryotic taxonomy without rRNA: longer oligonucleotide word lengths improve genome and metagenome taxonomic classification.

    Directory of Open Access Journals (Sweden)

    Eric B Alsop

    Full Text Available Oligonucleotide signatures, especially tetranucleotide signatures, have been used as method for homology binning by exploiting an organism's inherent biases towards the use of specific oligonucleotide words. Tetranucleotide signatures have been especially useful in environmental metagenomics samples as many of these samples contain organisms from poorly classified phyla which cannot be easily identified using traditional homology methods, including NCBI BLAST. This study examines oligonucleotide signatures across 1,424 completed genomes from across the tree of life, substantially expanding upon previous work. A comprehensive analysis of mononucleotide through nonanucleotide word lengths suggests that longer word lengths substantially improve the classification of DNA fragments across a range of sizes of relevance to high throughput sequencing. We find that, at present, heptanucleotide signatures represent an optimal balance between prediction accuracy and computational time for resolving taxonomy using both genomic and metagenomic fragments. We directly compare the ability of tetranucleotide and heptanucleotide world lengths (tetranucleotide signatures are the current standard for oligonucleotide word usage analyses for taxonomic binning of metagenome reads. We present evidence that heptanucleotide word lengths consistently provide more taxonomic resolving power, particularly in distinguishing between closely related organisms that are often present in metagenomic samples. This implies that longer oligonucleotide word lengths should replace tetranucleotide signatures for most analyses. Finally, we show that the application of longer word lengths to metagenomic datasets leads to more accurate taxonomic binning of DNA scaffolds and have the potential to substantially improve taxonomic assignment and assembly of metagenomic data.

  4. Hierarchical unilamellar vesicles of controlled compositional heterogeneity.

    Directory of Open Access Journals (Sweden)

    Maik Hadorn

    Full Text Available Eukaryotic life contains hierarchical vesicular architectures (i.e. organelles that are crucial for material production and trafficking, information storage and access, as well as energy production. In order to perform specific tasks, these compartments differ among each other in their membrane composition and their internal cargo and also differ from the cell membrane and the cytosol. Man-made structures that reproduce this nested architecture not only offer a deeper understanding of the functionalities and evolution of organelle-bearing eukaryotic life but also allow the engineering of novel biomimetic technologies. Here, we show the newly developed vesicle-in-water-in-oil emulsion transfer preparation technique to result in giant unilamellar vesicles internally compartmentalized by unilamellar vesicles of different membrane composition and internal cargo, i.e. hierarchical unilamellar vesicles of controlled compositional heterogeneity. The compartmentalized giant unilamellar vesicles were subsequently isolated by a separation step exploiting the heterogeneity of the membrane composition and the encapsulated cargo. Due to the controlled, efficient, and technically straightforward character of the new preparation technique, this study allows the hierarchical fabrication of compartmentalized giant unilamellar vesicles of controlled compositional heterogeneity and will ease the development of eukaryotic cell mimics that resemble their natural templates as well as the fabrication of novel multi-agent drug delivery systems for combination therapies and complex artificial microreactors.

  5. A neural model of hierarchical reinforcement learning.

    Science.gov (United States)

    Rasmussen, Daniel; Voelker, Aaron; Eliasmith, Chris

    2017-01-01

    We develop a novel, biologically detailed neural model of reinforcement learning (RL) processes in the brain. This model incorporates a broad range of biological features that pose challenges to neural RL, such as temporally extended action sequences, continuous environments involving unknown time delays, and noisy/imprecise computations. Most significantly, we expand the model into the realm of hierarchical reinforcement learning (HRL), which divides the RL process into a hierarchy of actions at different levels of abstraction. Here we implement all the major components of HRL in a neural model that captures a variety of known anatomical and physiological properties of the brain. We demonstrate the performance of the model in a range of different environments, in order to emphasize the aim of understanding the brain's general reinforcement learning ability. These results show that the model compares well to previous modelling work and demonstrates improved performance as a result of its hierarchical ability. We also show that the model's behaviour is consistent with available data on human hierarchical RL, and generate several novel predictions.

  6. Hierarchical image classification in the bioscience literature.

    Science.gov (United States)

    Kim, Daehyun; Yu, Hong

    2009-11-14

    Our previous work has shown that images appearing in bioscience articles can be classified into five types: Gel-Image, Image-of-Thing, Graph, Model, and Mix. For this paper, we explored and analyzed features strongly associated with each image type and developed a hierarchical image classification approach for classifying an image into one of the five types. First, we applied texture features to separate images into two groups: 1) a texture group comprising Gel Image, Image-of-Thing, and Mix, and 2) a non-texture group comprising Graph and Model. We then applied entropy, skewness, and uniformity for the first group, and edge difference, uniformity, and smoothness for the second group to classify images into specific types. Our results show that hierarchical image classification accurately divided images into the two groups during the initial classification and that the overall accuracy of the image classification was higher than that of our previous approach. In particular, the recall of hierarchical image classification was greatly improved due to the high accuracy of the initial classification.

  7. The traveling salesman problem: a hierarchical model.

    Science.gov (United States)

    Graham, S M; Joshi, A; Pizlo, Z

    2000-10-01

    Our review of prior literature on spatial information processing in perception, attention, and memory indicates that these cognitive functions involve similar mechanisms based on a hierarchical architecture. The present study extends the application of hierarchical models to the area of problem solving. First, we report results of an experiment in which human subjects were tested on a Euclidean traveling salesman problem (TSP) with 6 to 30 cities. The subject's solutions were either optimal or near-optimal in length and were produced in a time that was, on average, a linear function of the number of cities. Next, the performance of the subjects is compared with that of five representative artificial intelligence and operations research algorithms, that produce approximate solutions for Euclidean problems. None of these algorithms was found to be an adequate psychological model. Finally, we present a new algorithm for solving the TSP, which is based on a hierarchical pyramid architecture. The performance of this new algorithm is quite similar to the performance of the subjects.

  8. Image Search Reranking With Hierarchical Topic Awareness.

    Science.gov (United States)

    Tian, Xinmei; Yang, Linjun; Lu, Yijuan; Tian, Qi; Tao, Dacheng

    2015-10-01

    With much attention from both academia and industrial communities, visual search reranking has recently been proposed to refine image search results obtained from text-based image search engines. Most of the traditional reranking methods cannot capture both relevance and diversity of the search results at the same time. Or they ignore the hierarchical topic structure of search result. Each topic is treated equally and independently. However, in real applications, images returned for certain queries are naturally in hierarchical organization, rather than simple parallel relation. In this paper, a new reranking method "topic-aware reranking (TARerank)" is proposed. TARerank describes the hierarchical topic structure of search results in one model, and seamlessly captures both relevance and diversity of the image search results simultaneously. Through a structured learning framework, relevance and diversity are modeled in TARerank by a set of carefully designed features, and then the model is learned from human-labeled training samples. The learned model is expected to predict reranking results with high relevance and diversity for testing queries. To verify the effectiveness of the proposed method, we collect an image search dataset and conduct comparison experiments on it. The experimental results demonstrate that the proposed TARerank outperforms the existing relevance-based and diversified reranking methods.

  9. Metal hierarchical patterning by direct nanoimprint lithography.

    Science.gov (United States)

    Radha, Boya; Lim, Su Hui; Saifullah, Mohammad S M; Kulkarni, Giridhar U

    2013-01-01

    Three-dimensional hierarchical patterning of metals is of paramount importance in diverse fields involving photonics, controlling surface wettability and wearable electronics. Conventionally, this type of structuring is tedious and usually involves layer-by-layer lithographic patterning. Here, we describe a simple process of direct nanoimprint lithography using palladium benzylthiolate, a versatile metal-organic ink, which not only leads to the formation of hierarchical patterns but also is amenable to layer-by-layer stacking of the metal over large areas. The key to achieving such multi-faceted patterning is hysteretic melting of ink, enabling its shaping. It undergoes transformation to metallic palladium under gentle thermal conditions without affecting the integrity of the hierarchical patterns on micro- as well as nanoscale. A metallic rice leaf structure showing anisotropic wetting behavior and woodpile-like structures were thus fabricated. Furthermore, this method is extendable for transferring imprinted structures to a flexible substrate to make them robust enough to sustain numerous bending cycles.

  10. Towards a hierarchical optimization modeling framework for ...

    Science.gov (United States)

    Background:Bilevel optimization has been recognized as a 2-player Stackelberg game where players are represented as leaders and followers and each pursue their own set of objectives. Hierarchical optimization problems, which are a generalization of bilevel, are especially difficult because the optimization is nested, meaning that the objectives of one level depend on solutions to the other levels. We introduce a hierarchical optimization framework for spatially targeting multiobjective green infrastructure (GI) incentive policies under uncertainties related to policy budget, compliance, and GI effectiveness. We demonstrate the utility of the framework using a hypothetical urban watershed, where the levels are characterized by multiple levels of policy makers (e.g., local, regional, national) and policy followers (e.g., landowners, communities), and objectives include minimization of policy cost, implementation cost, and risk; reduction of combined sewer overflow (CSO) events; and improvement in environmental benefits such as reduced nutrient run-off and water availability. Conclusions: While computationally expensive, this hierarchical optimization framework explicitly simulates the interaction between multiple levels of policy makers (e.g., local, regional, national) and policy followers (e.g., landowners, communities) and is especially useful for constructing and evaluating environmental and ecological policy. Using the framework with a hypothetical urba

  11. Hierarchical structure of Turkey's foreign trade

    Science.gov (United States)

    Kantar, Ersin; Deviren, Bayram; Keskin, Mustafa

    2011-10-01

    We examine the hierarchical structures of Turkey's foreign trade by using real prices of their commodity export and import move together over time. We obtain the topological properties among the countries based on Turkey's foreign trade during the 1996-2010 period by using the concept of hierarchical structure methods (minimal spanning tree, (MST) and hierarchical tree, (HT)). These periods are divided into two subperiods, such as 1996-2002 and 2003-2010, in order to test various time-window and observe the temporal evolution. We perform the bootstrap techniques to investigate a value of the statistical reliability to the links of the MSTs and HTs. We also use a clustering linkage procedure in order to observe the cluster structure much better. From the structural topologies of these trees, we identify different clusters of countries according to their geographical location and economic ties. Our results show that the DE (Germany), UK (United Kingdom), FR (France), IT (Italy) and RU (Russia) are more important within the network, due to a tighter connection with other countries. We have also found that these countries play a significant role for Turkey's foreign trade and have important implications for the design of portfolio and investment strategies.

  12. Hierarchical architecture for flexible energy storage.

    Science.gov (United States)

    Pan, H; Ma, J; Tao, J; Zhu, S

    2017-05-25

    The introduction of hierarchy and chirality into structure is of great interest, and can result in new optical and electronic properties due to the synergistic effect of helical and anisotropic structures. Herein, we demonstrate a simple and straightforward route toward the fabrication of hierarchical chiral materials based on the assembly of two-dimensional graphene oxide nanosheets (GO) and one-dimensional cellulose nanocrystals (CNCs). The unique layered structure of CNC/GO could be preserved in the solid state, allowing electrode active SnO 2 to be loaded for potential applications in energy storage. The resultant SnO 2 /CNC/reduced GO (SnO 2 /CNC/rGO) composite could be processed into film, fiber, and textile with an extremely high tensile strength of 100 MPa. The free-standing SnO 2 /CNC/rGO electrodes exhibit highly improved energy storage performance, with a reversible capacity of ∼500 mA h g -1 maintained for 1500 cycles in the film and ∼800 mA h g -1 maintained for 150 cycles in the textile at a current density of 500 mA g -1 . This is attributed to the prepared hierarchical chiral structures. The presented technique provides an effective approach to producing hierarchical functional materials from nanoparticles as building blocks, which might open an avenue for the creation of new flexible energy storage devices.

  13. Depth profiles and free volume in aircraft primer films

    Science.gov (United States)

    Van Horn, J. D.; Chen, H.; Jean, Y. C.; Zhang, W.; Jaworowski, M. R.

    2015-06-01

    Positron annihilation lifetime spectroscopy (PALS) and associated techniques provide non-destructive methods to study the free volume inside polymeric materials, and to study material characteristics over a depth profile. Cast free films of organic- or aqueous-based, non-chromated aerospace primers, when cured for about one week, had very different water vapour transport (through-plane) behaviour. In addition, both types of primer films showed strong anisotropic behaviour in in-plane versus through-plane water vapour transport rates. We report the differences between the organic- and aqueous-based aircraft primer films samples and their surface depth profiles. In bulk PALS measurements, an aged, organic-based film exhibited typical lifetimes and intensities for a particulate-containing polymer film on both faces. In contrast, aqueous-based films exhibited face oriented-dependent differences. In all aqueous- based samples, the I3 value of the back of the sample was smaller. The primer film samples were also evaluated with mono-energetic positron beam techniques to generate depth profile information. The heterogeneity in the samples was verified by Doppler broadening of energy spectroscopy (DBES). A model for the differences in the faces of the films, and their layered structure is discussed.

  14. Improved primer for bonding polyurethane adhesives to metals

    Science.gov (United States)

    Constanza, L. J.

    1969-01-01

    Primer ensures effective bonding integrity of polyurethane adhesives on metal surfaces at temperatures ranging from minus 423 degrees to plus 120 degrees F. It provides greater metal surface protection and bond strengths over this temperature range than could be attained with other adhesive systems.

  15. Barcoded primers used in multiplex amplicon pyrosequencing bias amplification.

    Science.gov (United States)

    Berry, David; Ben Mahfoudh, Karim; Wagner, Michael; Loy, Alexander

    2011-11-01

    "Barcode-tagged" PCR primers used for multiplex amplicon sequencing generate a thus-far-overlooked amplification bias that produces variable terminal restriction fragment length polymorphism (T-RFLP) and pyrosequencing data from the same environmental DNA template. We propose a simple two-step PCR approach that increases reproducibility and consistently recovers higher genetic diversity in pyrosequencing libraries.

  16. RUCS: Rapid identification of PCR primers for unique core sequences

    DEFF Research Database (Denmark)

    Thomsen, Martin Christen Frølund; Hasman, Henrik; Westh, Henrik

    2017-01-01

    in silico PCR simulation. We compared our method, which identifies the unique core sequences, against an existing tool called ssGeneFinder, and found that our method was 6.5-20 times more sensitive. We used RUCS to design primer pairs that would target a set of genomes known to contain the mcr-1 colistin...

  17. Sights, Sounds, Senses in Step with Reading. Primer.

    Science.gov (United States)

    Norris, Mildred W.; Messerli, John H.

    This Title III bibliography presents books, films, filmstrips, and records that correlate with stories from primers. The list of books is prepared from the suggested lists of the Ginn, Houghton Mifflin, Lippincott, and Scott, Foresman reading series. The bibliography includes sections about animals, birthdays, cowboys, fantasy, helpers, holidays,…

  18. Children on Medication: A Primer for School Personnel.

    Science.gov (United States)

    Gadow, Kenneth D.

    Intended as a primer for school personnel, the book discusses children whose various disorders require them to be on medication, and describes the behavioral effects of these drugs along with their major side effects. Fundamental concepts in pharmacotherapy are reviewed, including dosage adjustment and side effects, and a brief introduction to the…

  19. A Primer on Ways To Explore Item Bias.

    Science.gov (United States)

    Cromwell, Susan

    This paper provides an introductory primer on methods for exploring item bias. The purposes of item bias analysis are to investigate whether test scores are affected by different sources of variance in the various subpopulations, and if different sources of variance are found, to determine if an unfair advantage exists. The review discusses three…

  20. Onchocercal DNA amplification using beta actin gene primers ...

    African Journals Online (AJOL)

    Detecting any parts of the parasite stages such as antigens, enzymes and nucleic acids (DNA and RNA) is a definitive diagnosis and highly sensitive. This study was to evaluate the diagnostic reliability of the beta actin gene primer pair to confirm its suitability for validating presence or absence of skin microfilaria at ...