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Sample records for nonradioactive ultrasensitive site-specific

  1. Site Specific Verification Guidelines.

    SciTech Connect

    Harding, Steve; Gordon, Frederick M.; Kennedy, Mike

    1992-05-01

    The Bonneville Power Administration (BPA) and the Northwest region have moved from energy surplus to a time when demand for energy is likely to exceed available supplies. The Northwest Power Planning Council is calling for a major push to acquire new resources.'' To meet anticipated loads in the next decade, BPA and the region must more than double that rate at which we acquire conservation resources. BPA hopes to achieve some of this doubling by programs independently designed and implemented by utilities and other parties without intensive BPA involvement. BPA will accept proposals for programs using performance-based payments, in which BPA bases its reimbursement to the sponsor on measured energy savings rather than program costs. To receive payment for conservation projects developed under performance-based programs, utilities and other project developers must propose verification plans to measure the amount of energy savings. BPA has traditionally used analysis of billing histories, before and after measure installation, adjusted by a comparison group on non-participating customers to measure conservation savings. This approach does not work well for all conversation projects. For large or unusual facilities the comparison group approach is not reliable due to the absence of enough comparable non-participants to allow appropriate statistical analysis. For these facilities, which include large commercial and institutional buildings, industrial projects, and complex combinations of building types served by a single utility meter, savings must be verified on a site-specific basis. These guidelines were written to help proposers understand what Bonneville considers the important issues in site specific verification of conservation performance. It also provides a toolbox of methods with guidance on their application and use. 15 refs.

  2. Groundwater Monitoring Plan for the Nonradioactive Dangerous Waste Landfill

    SciTech Connect

    J.S. Lindberg; M.J. Hartman

    1999-08-17

    The Nonradioactive Dangerous Waste Landfill (NRDWL), which received nonradioactive hazardous waste between 1975 and 1985, is located in the central Hanford Site (Figure 1.1) in southeastern Washington State. The Solid Waste Landfill, which is regulated and monitored separately, is adjacent to the NRDWL. The NRDWL is regulated under the Resource Conservation and Recovery Act of 1976 (RCRA) and monitored by Pacific Northwest National Laboratory. Monitoring is done under interim-status, indicator-evaluation requirements (WAC 173-303 and by reference, 40 CFR 265.92). The well network includes three upgradient wells (one shared with the Solid Waste Landfill) and six downgradient wells. The wells are sampled semiannually for contaminant indicator parameters and site-specific parameters and annually for groundwater quality parameters.

  3. Ultrasensitive characterization of site-specific glycosylation of affinity-purified haptoglobin from lung cancer patient plasma using 10 μm i.d. porous layer open tubular liquid chromatography-linear ion trap collision-induced dissociation/electron transfer dissociation mass spectrometry.

    PubMed

    Wang, Dongdong; Hincapie, Marina; Rejtar, Tomas; Karger, Barry L

    2011-03-15

    Site-specific analysis of protein glycosylation is important for biochemical and clinical research efforts. Glycopeptide analysis using liquid chromatography-collision-induced dissociation/electron transfer dissociation mass spectrometry (LC-CID/ETD-MS) allows simultaneous characterization of the glycan structure and attached peptide site. However, due to the low ionization efficiency of glycopeptides during electrospray ionization, 200-500 fmol of sample per injection is needed for a single LC-MS run, which makes it challenging for the analysis of limited amounts of glycoprotein purified from biological matrixes. To improve the sensitivity of LC-MS analysis for glycopeptides, an ultranarrow porous layer open tubular (PLOT) LC column (2.5 m × 10 μm i.d.) was coupled to a linear ion trap (LTQ) collision-induced dissociation/electron transfer dissociation mass spectrometer to provide sensitive analysis of N-linked protein glycosylation heterogeneity. The potential of the developed method is demonstrated by the characterization of site-specific glycosylation using haptoglobin (Hpt) as a model protein. To limit the amount of haptoglobin to low picomole amounts of protein, we affinity purified it from 1 μL of pooled lung cancer patient plasma. A total of 26 glycoforms/glycan compositions on three Hpt tryptic glycopeptides were identified and quantified from 10 LC-MS runs with a consumption of 100 fmol of Hpt digest (13 ng of protein, 10 fmol per injection). Included in this analysis was the determination of the glycan occupancy level. At this sample consumption level, the high sensitivity of the PLOT LC-LTQ-CID/ETD-MS system allowed glycopeptide identification and structure determination, along with relative quantitation of glycans presented on the same peptide backbone, even for low abundant glycopeptides at the ∼100 amol level. The PLOT LC-MS system is shown to have sufficient sensitivity to allow characterization of site-specific protein glycosylation from trace

  4. Common uses of nonradioactive drugs in nuclear medicine

    SciTech Connect

    Ponto, J.A.; Hladik, W.B.

    1984-06-01

    A variety of nonradioactive pharmaceuticals commonly used in patients who receive nuclear medicine diagnostic tests are described. Nonradioactive drugs used in thyroid, brain, hepatobiliary, cardiac, renal, Meckel's diverticulum, gallium, adrenal, and hematological studies are described. Pharmaceutical necessities used as disinfectants, diluents, and anticoagulants are also described. Hospital pharmacists should be familiar with the uses of commonly prescribed nonradioactive drugs in nuclear medicine studies.

  5. DOE site-specific threat assessment

    SciTech Connect

    West, D.J.; Al-Ayat, R.A.; Judd, B.R.

    1985-07-12

    A facility manager faced with the challenges of protecting a nuclear facility against potential threats must consider the likelihood and consequences of such threats, know the capabilities of the facility safeguards and security systems, and make informed decisions about the cost-effectivness of safeguards and security upgrades. To help meet these challenges, the San Francisco Operations Office of the Department of Energy, in conjunction with the Lawrence Livermore Laboratory, has developed a site-specific threat assessment approach and a quantitative model to improve the quality and consistency of site-specific threat assessment and resultant security upgrade decisions at sensitive Department of Energy facilities. 5 figs.

  6. Site-Specific Infrared Probes of Proteins

    PubMed Central

    Ma, Jianqiang; Pazos, Ileana M.; Zhang, Wenkai; Culik, Robert M.; Gai, Feng

    2015-01-01

    Infrared spectroscopy has played an instrumental role in studying a wide variety of biological questions. However, in many cases it is impossible or difficult to rely on the intrinsic vibrational modes of biological molecules of interest, such as proteins, to reveal structural and/or environmental information in a site-specific manner. To overcome this limitation, many recent efforts have been dedicated to the development and application of various extrinsic vibrational probes that can be incorporated into biological molecules and used to site-specifically interrogate their structural and/or environmental properties. In this Review, we highlight some recent advancements of this rapidly growing research area. PMID:25580624

  7. Site-Specific Rules for Risk Assessment.

    ERIC Educational Resources Information Center

    Tucker, William; And Others

    1994-01-01

    This article examines the development of regulatory guidance upon which the technology of risk assessment has matured into a decision-making method of choice. It examines in particular the role of site-specific risk assessment at Superfund sites. (LZ)

  8. Site-Specific Rules for Risk Assessment.

    ERIC Educational Resources Information Center

    Tucker, William; And Others

    1994-01-01

    This article examines the development of regulatory guidance upon which the technology of risk assessment has matured into a decision-making method of choice. It examines in particular the role of site-specific risk assessment at Superfund sites. (LZ)

  9. Nanoparticles for Site Specific Genome Editing

    NASA Astrophysics Data System (ADS)

    McNeer, Nicole Ali

    Triplex-forming peptide nucleic acids (PNAs) can be used to coordinate the recombination of short 50-60 by "donor DNA" fragments into genomic DNA, resulting in site-specific correction of genetic mutations or the introduction of advantageous genetic modifications. Site-specific gene editing in hematopoietic stem and progenitor cells (HSPCs) could result in treatment or cure of inherited disorders of the blood such as beta-thalassemia. Gene editing in HSPCs and differentiated T cells could help combat HIV/AIDs by modifying receptors, such as CCR5, necessary for R5-tropic HIV entry. However, translation of genome modification technologies to clinical practice is limited by challenges in intracellular delivery, especially in difficult-to-transfect hematolymphoid cells. In vivo gene editing could also provide novel treatment for systemic monogenic disorders such as cystic fibrosis, an autosomal recessive disorder caused by mutations in the cystic fibrosis transmembrane receptor. Here, we have engineered biodegradable nanoparticles to deliver oligonucleotides for site-specific genome editing of disease-relevant genes in human cells, with high efficiency, low toxicity, and editing of clinically relevant cell types. We designed nanoparticles to edit the human beta-globin and CCR5 genes in hematopoietic cells. We show that poly(lactic-co-glycolic acid) (PLGA) nanoparticles can delivery PNA and donor DNA for site-specific gene modification in human hematopoietic cells in vitro and in vivo in NOD-scid IL2rgammanull mice. Nanoparticles delivered by tail vein localized to hematopoietic compartments in the spleen and bone marrow of humanized mice, resulting in modification of the beta-globin and CCR5 genes. Modification frequencies ranged from 0.005 to 20% of cells depending on the organ and cell type, without detectable toxicity. This project developed highly versatile methods for delivery of therapeutics to hematolymphoid cells and hematopoietic stem cells, and will help to

  10. Site-specific non-LTR retrotransposons.

    PubMed

    Fujiwara, Haruhiko

    2015-04-01

    Although most of non-long terminal repeat (non-LTR) retrotransposons are incorporated in the host genome almost randomly, some non-LTR retrotransposons are incorporated into specific sequences within a target site. On the basis of structural and phylogenetic features, non-LTR retrotransposons are classified into two large groups, restriction enzyme-like endonuclease (RLE)-encoding elements and apurinic/apyrimidinic endonuclease (APE)-encoding elements. All clades of RLE-encoding non-LTR retrotransposons include site-specific elements. However, only two of more than 20 APE-encoding clades, Tx1 and R1, contain site-specific non-LTR elements. Site-specific non-LTR retrotransposons usually target within multi-copy RNA genes, such as rRNA gene (rDNA) clusters, or repetitive genomic sequences, such as telomeric repeats; this behavior may be a symbiotic strategy to reduce the damage to the host genome. Site- and sequence-specificity are variable even among closely related non-LTR elements and appeared to have changed during evolution. In the APE-encoding elements, the primary determinant of the sequence- specific integration is APE itself, which nicks one strand of the target DNA during the initiation of target primed reverse transcription (TPRT). However, other factors, such as interaction between mRNA and the target DNA, and access to the target region in the nuclei also affect the sequence-specificity. In contrast, in the RLE-encoding elements, DNA-binding motifs appear to affect their sequence-specificity, rather than the RLE domain itself. Highly specific integration properties of these site-specific non-LTR elements make them ideal alternative tools for sequence-specific gene delivery, particularly for therapeutic purposes in human diseases.

  11. Site-Specific PEGylation of Therapeutic Proteins

    PubMed Central

    Dozier, Jonathan K.; Distefano, Mark D.

    2015-01-01

    The use of proteins as therapeutics has a long history and is becoming ever more common in modern medicine. While the number of protein-based drugs is growing every year, significant problems still remain with their use. Among these problems are rapid degradation and excretion from patients, thus requiring frequent dosing, which in turn increases the chances for an immunological response as well as increasing the cost of therapy. One of the main strategies to alleviate these problems is to link a polyethylene glycol (PEG) group to the protein of interest. This process, called PEGylation, has grown dramatically in recent years resulting in several approved drugs. Installing a single PEG chain at a defined site in a protein is challenging. Recently, there is has been considerable research into various methods for the site-specific PEGylation of proteins. This review seeks to summarize that work and provide background and context for how site-specific PEGylation is performed. After introducing the topic of site-specific PEGylation, recent developments using chemical methods are described. That is followed by a more extensive discussion of bioorthogonal reactions and enzymatic labeling. PMID:26516849

  12. Site-Specific PEGylation of Therapeutic Proteins.

    PubMed

    Dozier, Jonathan K; Distefano, Mark D

    2015-10-28

    The use of proteins as therapeutics has a long history and is becoming ever more common in modern medicine. While the number of protein-based drugs is growing every year, significant problems still remain with their use. Among these problems are rapid degradation and excretion from patients, thus requiring frequent dosing, which in turn increases the chances for an immunological response as well as increasing the cost of therapy. One of the main strategies to alleviate these problems is to link a polyethylene glycol (PEG) group to the protein of interest. This process, called PEGylation, has grown dramatically in recent years resulting in several approved drugs. Installing a single PEG chain at a defined site in a protein is challenging. Recently, there is has been considerable research into various methods for the site-specific PEGylation of proteins. This review seeks to summarize that work and provide background and context for how site-specific PEGylation is performed. After introducing the topic of site-specific PEGylation, recent developments using chemical methods are described. That is followed by a more extensive discussion of bioorthogonal reactions and enzymatic labeling.

  13. Site-Specific Carbon Isotopes in Organics

    NASA Astrophysics Data System (ADS)

    Piasecki, A.; Eiler, J. M.

    2012-12-01

    Natural organic molecules exhibit a wide range of internal site-specific isotope variation (i.e., molecules with same isotopic substitution type but different site). Such variations are generally unconstrained by bulk isotopic measurements. If known, site-specific variations might constrain temperatures of equilibrium, mechanisms of formation or consumption reactions, and possibly other details. For example, lipids can exhibit carbon isotope differences of up to 30‰ between adjacent carbon sites as a result of fractionations arising during decarboxylation of pyruvate and other steps in lipid biosynthesis(1). We present a method for site-specific carbon isotope analysis of propane, based on high-resolution, multi-collector gas source mass spectrometry, using a novel prototype instrument - the Thermo MAT 253 Ultra. This machine has an inlet system and electron bombardment ion source resembling those in conventional stable isotope gas source mass spectrometers, and the energy filter, magnet, and detector array resembling those in multi-collector ICPMS and TIMS. The detector array has 7 detector positions, 6 of which are movable, and each of which can collect ions with either a faraday cup (read through amplifiers ranging from 107-1012 ohms) or an SEM. High mass resolving power (up to 27,000, MRP = M/dM definition) is achieved through a narrow entrance slit, adjustable from 250 to 5 μm. Such resolution can cleanly separate isobaric interferences between isotopologues of organic molecules having the same cardinal mass (e.g., 13CH3 and 12CH2D). We use this technology to analyze the isotopologues and fragments of propane, and use such data to solve for the site-specific carbon isotope fractionation. By measuring isotopologues of both the one-carbon (13CH3) and the two-carbon (13C12CH4) fragment ion, we can solve for both bulk δ13C and the difference in δ13C between the terminal and central carbon position. We tested this method by analyzing mixtures between natural

  14. Savannah River Site's Site Specific Plan

    SciTech Connect

    Not Available

    1991-08-01

    This Site Specific Plan (SSP) has been prepared by the Savannah River Site (SRS) in order to show the Environmental Restoration and Waste Management activities that were identified during the preparation of the Department of Energy-Headquarters (DOE-HQ) Environmental Restoration and Waste Management Five-Year Plan (FYP) for FY 1992--1996. The SSP has been prepared in accordance with guidance received from DOE-HQ. DOE-SR is accountable to DOE-HQ for the implementation of this plan. The purpose of the SSP is to develop a baseline for policy, budget, and schedules for the DOE Environmental Restoration and Waste Management activities. The plan explains accomplishments since the Fiscal Year (FY) 1990 plan, demonstrates how present and future activities are prioritized, identifies currently funded activities and activities that are planned to be funded in the upcoming fiscal year, and describes future activities that SRS is considering.

  15. Examining site-specific GPCR phosphorylation.

    PubMed

    Butcher, Adrian J; Tobin, Andrew B; Kong, Kok Choi

    2011-01-01

    Phosphorylation of G protein-coupled receptors (GPCRs) is one of the most prominent post-translation modifications mediated by agonist stimulation. This process has been shown to result not only in receptor desensitisation but also, via the recruitment of arrestin adaptor proteins, to promote receptor coupling to numerous signalling pathways. Furthermore, there is now a growing body of evidence suggesting that GPCRs may employ phosphorylation as a mechanism to regulate their cell-type-specific signalling, hence generating tissue-specific functions. These advances have resulted partly from improved methods used in the determination of phospho-acceptor sites on GPCRs and improved analysis of the consequences of phosphorylation. This chapter aims to describe the methods used in our laboratory for the investigation of site-specific phosphorylation of the M₃-muscarinic receptor. These methods could easily be applied in the study of other receptors.

  16. The Connectivity Between Site-Specific Life Cycle Impact Assessment and Site-Specific Weighting

    EPA Science Inventory

    The goal of many LCIAs is to come to a single score with all of the impacts from a wide variety of impact assessments weighted to form this single score. My past experiences with developing site-specific impact assessment methodologies and how this can change the valuation porti...

  17. The Connectivity Between Site-Specific Life Cycle Impact Assessment and Site-Specific Weighting

    EPA Science Inventory

    The goal of many LCIAs is to come to a single score with all of the impacts from a wide variety of impact assessments weighted to form this single score. My past experiences with developing site-specific impact assessment methodologies and how this can change the valuation porti...

  18. RESRAD. Site-Specific Residual Radioactivity

    SciTech Connect

    Yu, C.

    1989-06-01

    RESRAD is designed to derive site-specific guidelines for allowable residual concentrations of radionuclides in soil. A guideline is defined as a radionuclide concentration or a level of radiation or radioactivity that is acceptable if a site is to be used without radiological restrictions. Guidelines are expressed as (1) concentrations of residual radionuclides in soil, (2) concentrations of airborne radon decay products, (3) levels of external gamma radiation, (4) levels of radioactivity from surface contamination, and (5) concentrations of residual radionuclides in air and water. Soil is defined as unconsolidated earth material, including rubble and debris that may be present. The controlling principles of all guidelines are (1) the annual radiation dose received by a member of the critical population group from the residual radioactive material - predicted by a realistic but reasonably conservative analysis and averaged over a 50 year period - should not exceed 100 mrem/yr, and (2) doses should be kept as low as reasonably achievable. All significant exposure pathways for the critical population group are considered in deriving soil guidelines. These pathways include direct exposure to external radiation from the contaminated soil material; internal radiation from inhalation of airborne radionuclides; and internal radiation from ingestion of plant foods grown in the contaminated soil, meat and milk from livestock fed with contaminated fodder and water, drinking water from a contaminated well, and fish from a contaminated pond.

  19. Site-specific codon bias in bacteria

    SciTech Connect

    Smith, J.M.; Smith, N.H.

    1996-03-01

    Sequences of the gapA and ompA genes from 10 genera of enterobacteria have been analyzed. There is strong bias in codon usage, but different synonymous codons are preferred at different sites in the same gene. Site-specific preference for unfavored codons is not confined to the first 100 codons and is usually manifest between two codons utilizing the same tRNA. Statistical analyses, based on conclusions reached in an accompanying paper, show that the use of an unfavored codon at a given site in different genera is not due to common descent and must therefore be caused either by sequence-specific mutation or sequence-specific selection. Reasons are given for thinking that sequence-specific mutation cannot be responsible. We are unable to explain the preference between synonymous codons ending in C or T, but synonymous choice between A and G at third sites is largely explained by avoidance of AG-G (where the hyphen indicates the boundary between codons). We also observed that the preferred codon for proline in Enterobacter cloacea has changed from CCG to CCA. 27 refs., 7 tabs.

  20. Statistical and Economic Techniques for Site-specific Nematode Management.

    PubMed

    Liu, Zheng; Griffin, Terry; Kirkpatrick, Terrence L

    2014-03-01

    Recent advances in precision agriculture technologies and spatial statistics allow realistic, site-specific estimation of nematode damage to field crops and provide a platform for the site-specific delivery of nematicides within individual fields. This paper reviews the spatial statistical techniques that model correlations among neighboring observations and develop a spatial economic analysis to determine the potential of site-specific nematicide application. The spatial econometric methodology applied in the context of site-specific crop yield response contributes to closing the gap between data analysis and realistic site-specific nematicide recommendations and helps to provide a practical method of site-specifically controlling nematodes.

  1. Statistical and Economic Techniques for Site-specific Nematode Management

    PubMed Central

    Liu, Zheng; Griffin, Terry; Kirkpatrick, Terrence L.

    2014-01-01

    Recent advances in precision agriculture technologies and spatial statistics allow realistic, site-specific estimation of nematode damage to field crops and provide a platform for the site-specific delivery of nematicides within individual fields. This paper reviews the spatial statistical techniques that model correlations among neighboring observations and develop a spatial economic analysis to determine the potential of site-specific nematicide application. The spatial econometric methodology applied in the context of site-specific crop yield response contributes to closing the gap between data analysis and realistic site-specific nematicide recommendations and helps to provide a practical method of site-specifically controlling nematodes. PMID:24643451

  2. Site-Specific, Climate-Friendly Farming

    NASA Astrophysics Data System (ADS)

    Brown, D. J.; Brooks, E. S.; Eitel, J.; Huggins, D. R.; Painter, K.; Rupp, R.; Smith, J. L.; Stockle, C.; Vierling, L. A.

    2011-12-01

    Of the four most important atmospheric greenhouse gasses (GHG) enriched through human activities, only nitrous oxide (N2O) emissions are due primarily to agriculture. However, reductions in the application of synthetic N fertilizers could have significant negative consequences for a growing world population given the crucial role that these fertilizers have played in cereal yield increases since WWII. Increasing N use efficiency (NUE) through precision management of agricultural N in space and time will therefore play a central role in the reduction of agricultural N2O emissions. Precision N management requires a greater understanding of the spatio-temporal variability of factors supporting N management decisions such as crop yield, water and N availability, utilization and losses. We present an overview of a large, collaborative, multi-disciplinary project designed to improve our basic understanding of nitrogen (N), carbon (C) and water (H2O) spatio-temporal dynamics for wheat-based cropping systems on complex landscapes, and develop management tools to optimize water- and nitrogen-use efficiency for these systems and landscapes. Major components of this project include: (a) cropping systems experiments addressing nitrogen application rate and seeding density for different landscape positions; (b) GHG flux experiments and monitoring; (c) soil microbial genetics and stable isotope analyses to elucidate biochemical pathways for N2O production; (d) proximal soil sensing for construction of detailed soil maps; (e) LiDAR and optical remote sensing for crop growth monitoring; (f) hydrologic experiments, monitoring, and modeling; (g) refining the CropSyst simulation model to estimate biophysical processes and GHG emissions under a variety of management and climatic scenarios; and (h) linking farm-scale enterprise budgets to simulation modeling in order to provide growers with economically viable site-specific climate-friendly farming guidance.

  3. Site-specific DNA alkylation and repair

    SciTech Connect

    Ezaz-Nikpay, K.

    1993-01-01

    This thesis describes a general method for the site-specific insertion of modified nucleotides into DNA and the application of this method to the study of N7-methyl-2[prime]-deoxyguanosine (m[sup 7]dG) in DNA. This thesis describes the chemical basis for the gap insertion/ligation method (GIL) and the use of this method to generate circularly permuted oligonucleotides. In this method, the synthesis of a single oligonucleotide leads to the formation of a double-stranded multimer with periodically-occurring gaps upon base-pairing in solution. The sequential action of a DNA polymerase and a DNA ligase leads to the insertion of a 2[prime]-deoxynucleoside-5[prime]-triphosphate into the gap, and formation of covalently-closed DNA. Finally, restriction endonucleases are used to generate oligonucleotides which contain the introduced nucleotide at symmetrically-related positions. The author describes the use of the GIL method for the insertion of m[sup 7]dG into various oligonucleotides and the Dickerson/Drew dodecamer respectively. The Dickerson/Drew dodecamer was chosen because it has been extensively studies both in its native and adduct bearing forms. The author describes the biophysical characterization of m[sup 7]dG in DNA, and concludes that the probe moiety in dimethyl-sulfate and template-directed interference footprinting of protein-DNA complexes in m[sup 7]dG and not a product of its decomposition. Further studies of m[sub 7]dG in DNA reveal that over long periods of time, the primary product of decomposition is an apurinic site. This dissertation describes the large-scale synthesis of the Dickerson/Drew dodecamer, and the characterization of its effect on DNA structure using nuclear magnetic resonance spectroscopy. The final chapter describes the overproduction, purification and crystallization of N3-methyladenine DNA glycosylase II (AlkA). AlkA is known to repair m[sup 7]dG residues in DNA.

  4. 32P-postlabeling analysis of non-radioactive aromatic carcinogen--DNA adducts.

    PubMed

    Gupta, R C; Reddy, M V; Randerath, K

    1982-01-01

    A newly developed enzymatic 32P-postlabeling method was applied to the analysis of DNA's containing non-radioactive arylamine, arylamide, and polycyclic aromatic hydrocarbon adducts. DNA reacted in vitro with N-hydroxy-2-amino-fluorene, N-acetoxy-2-acetylaminofluorene, and 7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene, respectively, as well as DNA preparations from the liver of rats treated with N-hydroxy-2-acetylaminofluorene and benzo[a]pyrene, respectively, were enzymatically digested to deoxyribonucleoside 3'-monophosphates, which were then converted to [5'-32P]deoxyribonucleoside 3',5'-bisphosphates by T4 polynucleotide kinase-catalyzed [32P]phosphate transfer from [gamma-32P]ATP. The 32P-labeled nucleotides were resolved by anion-exchange t.l.c. on polyethyleneimine-cellulose and detected by autoradiography. Aromatic adduct nucleotides were found to be retained at the origin in aqueous electrolyte solutions, but to migrate as distinct spots in solvents containing 7-8.5 M urea. Advantage was taken of this observation to remove 32P-labeled normal DNA nucleotides from adduct nucleotides. This purification enabled the detection of a single adduct in 10(7)-10(8) normal nucleotides. The method appears applicable to the ultrasensitive detection of a large number of carcinogen--DNA adducts of diverse structure without requiring radioactive carcinogens or specific antibodies.

  5. Computational identification of site-specific transcription factors in Drosophila.

    PubMed

    Adryan, Boris; Teichmann, Sarah A

    2007-01-01

    Site-specific transcription factors (TFs) recognise their target genes in a sequence- or conformation-dependent manor. In contrast to the basal TFs that are the general facilitators of gene expression, their site-specific interaction partners are tissue- or condition-specific. Thus, site-specific TFs constitute the major prerequisite for the modular expression programmes that drive metazoan development. This article deals with the computational identification of TFs (how to find them in genomes) and with online resources such as the FlyTF database of Drosophila site-specific TFs (how to find them online).

  6. Double clicking for site-specific coupling of multiple enzymes.

    PubMed

    Lim, Sung In; Cho, Jinhwan; Kwon, Inchan

    2015-09-14

    A method to site-specifically couple multiple enzymes is reported. The approach is based on the site-specific incorporation of a clickable non-natural amino acid into enzymes and two compatible click reactions. The multi-enzyme reaction system exhibited enhanced catalytic efficiency over the respective free enzymes.

  7. SITE-SPECIFIC CHARACTERIZATION OF SOIL RADON POTENTIALS

    EPA Science Inventory

    The report presents a theoretical basis for measuring site-specific radon potentials. However, the empirical measurements suggest that the precision of such measurements is marginal, leaving an uncertainty of about a factor of 2 in site-specific estimates. Although this may be us...

  8. Enhancing adoption of site-specific variable rate sprinkler systems

    USDA-ARS?s Scientific Manuscript database

    More than twenty years of private and public research on site-specific variable-rate sprinkler irrigation (SS-VRI) has resulted in very limited commercial adoption of the technology. Documented and proven water conservation strategies using site-specific irrigation are quite limited, and its cost-ef...

  9. Site-specific crop management using geophysical proximal sensors

    USDA-ARS?s Scientific Manuscript database

    Key components of site-specific crop management are (i) identifying the site-specific factors that influence within-field crop yield variation and (ii) spatially characterizing those factors. Geo-referenced measurements of apparent soil electrical conductivity (ECa) provide a potential means of cha...

  10. SITE-SPECIFIC CHARACTERIZATION OF SOIL RADON POTENTIALS

    EPA Science Inventory

    The report presents a theoretical basis for measuring site-specific radon potentials. However, the empirical measurements suggest that the precision of such measurements is marginal, leaving an uncertainty of about a factor of 2 in site-specific estimates. Although this may be us...

  11. Site Specific Management of Cotton Production in the United States

    USDA-ARS?s Scientific Manuscript database

    Site-specific management or precision agriculture, as it is evolving in large-scale crop production, offers promising new methods for managing cotton production for optimized yields, maximized profitability, and minimized environmental pollution. However, adaptation of site-specific theory and meth...

  12. DNA Hybridization: Nonradioactive Labeling Now Available for the Laboratory.

    ERIC Educational Resources Information Center

    Freeman, Lenore Gardner

    1984-01-01

    The advantages of DNA hybridization procedures for classroom and clinical use can now be realized by the recent development of nonradioactive DNA labeling/detection procedures. These methods (which are described) can replace the use of isotopes in standard DNA hybridization procedures. (JN)

  13. DNA Hybridization: Nonradioactive Labeling Now Available for the Laboratory.

    ERIC Educational Resources Information Center

    Freeman, Lenore Gardner

    1984-01-01

    The advantages of DNA hybridization procedures for classroom and clinical use can now be realized by the recent development of nonradioactive DNA labeling/detection procedures. These methods (which are described) can replace the use of isotopes in standard DNA hybridization procedures. (JN)

  14. An ELISA DYRK1A non-radioactive assay suitable for the characterization of inhibitors

    PubMed Central

    Liu, Yong; Adayev, Tatyana; Hwang, Yu-Wen

    2017-01-01

    The DYRK1A (dual specificity tyrosine phosphorylation-regulated kinase 1A) gene encodes a proline-directed Ser/Thr kinase. Elevated expression and/or altered distribution of the kinase have been implicated in the neurological impairments associated with Down syndrome (DS) and Alzheimer’s disease (AD). Consequently, DYRK1A inhibition has been of significant interest as a potential strategy for therapeutic intervention of DS and AD. Many classes of novel inhibitors have been described in the past decade. Although non-radioactive methods for analyzing DYRK1A inhibition have been developed, methods employing radioactive tracers are still commonly used for quantitative characterization of DYRK1A inhibitors. Here, we present a non-radioactive ELISA assay based on the detection of DYRK1A-phosphorylated dynamin 1a fragment using a phosphorylation site-specific antibody. The assay was verified by the use of two well-characterized DYRK1A inhibitors, epigallocatechin gallate (EGCG) and harmine. The IC 50s for EGCG and harmine determined by the ELISA method were found to be comparable to those previously measured by radioactive tracing methods.  Furthermore, we determined the mode of inhibition for EGCG and harmine by a modification of the ELISA assay. This assay confirms the mode of inhibition of EGCG (non-ATP-competitive) and harmine (ATP-competitive), as previously determined. We conclude that the ELISA platform demonstrated here is a viable alternative to the traditional radioactive tracer assays for analyzing DYRK1A inhibitors. PMID:28163906

  15. Complete Non-Radioactive Operability Tests for Cladding Hull Chlorination

    SciTech Connect

    Collins, Emory D; Johnson, Jared A.; Hylton, Tom D.; Brunson, Ronald Ray; Hunt, Rodney Dale; DelCul, Guillermo Daniel; Bradley, Eric Craig; Spencer, Barry B.

    2016-04-01

    Non-radioactive operability tests were made to test the metal chlorination reactor and condenser and their accessories using batch chlorinations of non-radioactive cladding samples and to identify optimum operating practices and components that need further modifications prior to installation of the equipment into the hot cell for tests on actual used nuclear fuel (UNF) cladding. The operability tests included (1) modifications to provide the desired heating and reactor temperature profile; and (2) three batch chlorination tests using, respectively, 100, 250, and 500 g of cladding. During the batch chlorinations, metal corrosion of the equipment was assessed, pressurization of the gas inlet was examined and the best method for maintaining solid salt product transfer through the condenser was determined. Also, additional accessing equipment for collection of residual ash and positioning of the unit within the hot cell were identified, designed, and are being fabricated.

  16. Criteria for the Certification of Non-Radioactive Hazardous Waste

    SciTech Connect

    Gagner, S D; Gaylord, R; Govers, R; Kennedy, W E; Hunnacek, M M; Kennedy, A M

    2003-04-10

    In 1991, in response to the Department of Energy (DOE) Moratorium on the shipment of hazardous waste from Radioactive Materials Management Areas (RMMAs), Lawrence Livermore National Laboratory (LLNL) developed a process to use a combination of generator knowledge and/or sampling and analyses to certify waste as non-radioactive. The analytical process used the minimum detectable activity (MDA) as the de minimus value. In the past twelve years, a great deal of operating experience has shown the LLNL certification process has serious limitations including: (1) Procedure-specified analytical methodologies have resulted in the inability to adopt new techniques and methods that are more rapid, safer, and produce less waste. (2) The characterization of materials as radioactive or non-radioactive is dependent on method-specific detection limits, not on an objective risk-based standard. (3) There are substantial differences in the limits for surface contamination, sewer discharges, and hazardous waste moratorium determinations, even though all of these methods are used to free-release materials from radiological controls. LLNL, in conjunction with the Chamberlain Group and Dade Moeller & Associates, Inc., is pursuing a risk-based approach to determine whether waste is non-radioactive, consistent with DOE guidance. This paper discusses the approach, which includes defining the radionuclides considered, establishing the exposure scenarios for the critical groups identified for each of three waste streams, defining the exposure pathways and key input data or assumptions, presenting radiation doses for unit concentrations of radionuclides in each waste stream, presenting radiation doses for unit concentrations of radionuclides in each waste stream, presenting the authorized limits for each waste stream, and discussing the results. Analytical values which fall below these authorization limits will be considered non-radioactive, with any individual dose maintained below 1 mrem/yr.

  17. Nonradioactive demonstration of the Alpha D and D Pilot Facility

    SciTech Connect

    Wobser, J.K.

    1983-01-01

    The Alpha-Contained Decontamination and Disassembly (AD and D) pilot facility was designed to demonstrate the process flowsheet under conditions typical to those expected in a production facility. To achieve this, nonradioactive waste items similar to those in retrievable storage at the Savannah River Plant burial ground (e.g. gloveboxes), were chemically sprayed and size reduced. During process runs, parameters such as feed rate, oxide removal, etching rate, and secondary waste generation were determined. The exhaust system was monitored during operation to ensure that exhaust from the facility was sufficiently filtered before release to the atmosphere. The strategy for decontamination techniques required development during the nonradioactive testing period. Under investigation during process runs were both once-through and recirculating washes, and their correlation to oxide removal and etching rates on the stainless steel feed items. Wash products of the decontamination process were analyzed for concentration of Ni, Cr, Fe, Mn, and Si, major components of stainless steel. Size reduction techniques were also developed during the nonradioactive testing period. An array of conventional power and pneumatic tools were tested and evaluated. Plasma arc torch operating parameters; standoff distance, ampere setting, and cutting angle were determined.

  18. Site-specific recombinases as tools for heterologous gene integration.

    PubMed

    Hirano, Nobutaka; Muroi, Tetsurou; Takahashi, Hideo; Haruki, Mitsuru

    2011-10-01

    Site-specific recombinases are the enzymes that catalyze site-specific recombination between two specific DNA sequences to mediate DNA integration, excision, resolution, or inversion and that play a pivotal role in the life cycles of many microorganisms including bacteria and bacteriophages. These enzymes are classified as tyrosine-type or serine-type recombinases based on whether a tyrosine or serine residue mediates catalysis. All known tyrosine-type recombinases catalyze the formation of a Holliday junction intermediate, whereas the catalytic mechanism of all known serine-type recombinases includes the 180° rotation and rejoining of cleaved substrate DNAs. Both recombinase families are further subdivided into two families; the tyrosine-type recombinases are subdivided by the recombination directionality, and the serine-type recombinases are subdivided by the protein size. Over more than two decades, many different site-specific recombinases have been applied to in vivo genome engineering, and some of them have been used successfully to mediate integration, deletion, or inversion in a wide variety of heterologous genomes, including those from bacteria to higher eukaryotes. Here, we review the recombination mechanisms of the best characterized recombinases in each site-specific recombinase family and recent advances in the application of these recombinases to genomic manipulation, especially manipulations involving site-specific gene integration into heterologous genomes.

  19. Utilization of Site-Specific Recombination in Biopharmaceutical Production

    PubMed Central

    Ahmadi, Maryam; Damavandi, Narges; Akbari, Mohammad Reza; Davami, Fatemeh

    2016-01-01

    Mammalian expression systems, due to their capacity in post-translational modification, are preferred systems for biopharmaceutical protein production. Several recombinant protein systems have been introduced to the market, most of which are under clinical development. In spite of significant improvements such as cell line engineering, introducing novel expression methods, gene silencing and process development, expression level is unpredictable and unstable because of the random location of integration in the genome. Site-specific recombination techniques are capable of producing stable and high producer clonal cells; therefore, they are gaining more importance in the biopharmaceutical production. Site-specific recombination methods increase the recombinant protein production by specifically inserting a vector at a locus with specific expression trait. The present review focused on the latest developments in site-specific recombination techniques, their specific features and comparisons. PMID:26602035

  20. Site specific protein labeling by enzymatic posttranslational modification.

    PubMed

    Sunbul, Murat; Yin, Jun

    2009-09-07

    Site specific protein labeling plays a key role in elucidating the function of the proteins at the molecular level by revealing their locations in the cell, their interaction networks with other cellular components and the dynamic mechanisms of their bio-generation, trafficking and degradation in response to regulatory signals in a biological system. Site specific protein labeling is, in essence, artificial modification of proteins with new chemical entities at the posttranslational stage. Based on the analogy between protein labeling and protein posttranslational modification, enzymatic tools have been developed for site specific and efficient labeling of target proteins with chemical probes of diverse structures and functionalities. This perspective surveys a number of protein labeling methods based on the application of protein posttranslational modification enzymes.

  1. Utilization of Site-Specific Recombination in Biopharmaceutical Production.

    PubMed

    Ahmadi, Maryam; Damavandi, Narges; Akbari Eidgahi, Mohammad Reza; Davami, Fatemeh

    2016-01-01

    Mammalian expression systems, due to their capacity in post-translational modification, are preferred systems for biopharmaceutical protein production. Several recombinant protein systems have been introduced to the market, most of which are under clinical development. In spite of significant improvements such as cell line engineering, introducing novel expression methods, gene silencing and process development, expression level is unpredictable and unstable because of the random location of integration in the genome. Site-specific recombination techniques are capable of producing stable and high producer clonal cells; therefore, they are gaining more importance in the biopharmaceutical production. Site-specific recombination methods increase the recombinant protein production by specifically inserting a vector at a locus with specific expression trait. The present review focused on the latest developments in site-specific recombination techniques, their specific features and comparisons.

  2. Ultrasensitive ultrasonic transducer studies

    SciTech Connect

    Dixon, R.; Darling, T.; Migliori, A.

    1996-09-01

    This is the final report of a one-year, Laboratory-Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). This project sought to continue development of the ultrasensitive ultrasonic transducers that won a 1994 R&D 100 Award. These transducers have a very smooth response across a broad frequency range and thus are extremely well- suited for resonant ultrasound spectroscopy as well as pulsed-echo and acoustic-emission applications. Current work on these transducers has indicated that bonding the piezoelectric and wear surface to a metal foil and attaching the foil to a body is less expensive and produces a transducer that is as good or better than commercially produced transducers. We have diffusion- bonded piezoelectric crystals and backings to stainless-steel-foil and wear surfaces. These are then attached onto stainless-steel tubes with electrical connectors to form the transducers. The transducers have been characterized using a reciprocity technique, electrical response, and optical interferometry. After characterization, the transducers have been compared to existing transducers by measuring and testing identical properties.

  3. Delineating site-specific management units with proximal sensors

    USDA-ARS?s Scientific Manuscript database

    Conventional farming uniformly manages fields with no consideration for spatial variability. This causes reduced productivity, misuse of finite resources (e.g., water and fertilizer), and detriment impacts on the environment. Site-specific management units (SSMUs) have been proposed as a means of ...

  4. Site-Specific Dance: Promoting Social Awareness in Choreography

    ERIC Educational Resources Information Center

    MacBean, Arianne

    2004-01-01

    Site-specific dance, which is often defined as dance that occurs outside of the conventional theater space, challenges choreographers to look at, listen to, feel, and think about the space in which the dance is performed. It also asks audiences to be active participants in the performance experience. The dances have to be informed by the space and…

  5. Engineering the mouse genome by site-specific recombination.

    PubMed

    Metzger, D; Feil, R

    1999-10-01

    Site-specific recombination systems are powerful tools for introducing predetermined modifications into eukaryotic genomes. Recent advances allow the manipulation of chromosomal DNA in a spatially and temporally controlled manner in mice, offering unprecedented possibilities for studying mammalian genome function and for generating animal models for human diseases.

  6. Uncertainty Analysis with Site Specific Groundwater Models: Experiences and Observations

    SciTech Connect

    Brewer, K.

    2003-07-15

    Groundwater flow and transport predictions are a major component of remedial action evaluations for contaminated groundwater at the Savannah River Site. Because all groundwater modeling results are subject to uncertainty from various causes; quantification of the level of uncertainty in the modeling predictions is beneficial to project decision makers. Complex site-specific models present formidable challenges for implementing an uncertainty analysis.

  7. Wireless Site-specific Irrigation - The Future of Intelligent Agriculture

    USDA-ARS?s Scientific Manuscript database

    A wireless site-specific irrigation system was developed with a distributed wireless sensor network. The system allows growers to remotely access field conditions and an irrigation operation at the home or office via wireless radio communication, directing individual sprinklers on how much water to ...

  8. Adoption of site-specific variable rate sprinkler irrigation systems

    USDA-ARS?s Scientific Manuscript database

    More than twenty years of private and public research on site-specific variable-rate sprinkler irrigation (SS-VRI) technology has resulted in limited commercial adoption of the technology. Competing patents, liability and proprietary software have affected industry’s willingness to move into a new t...

  9. Evaluation of potential water conservation using site-specific irrigation

    USDA-ARS?s Scientific Manuscript database

    With the advent of site-specific variable-rate irrigation (VRI) systems, irrigation can be spatially managed within sub-field-sized zones. Spatial irrigation management can optimize spatial water use efficiency and may conserve water. Spatial VRI systems are currently being managed by consultants ...

  10. Invasive Weed Management Is Site-Specific Weed Management.

    USDA-ARS?s Scientific Manuscript database

    Site-specific weed management in crops and invasive weed management in natural lands and rangelands appear to be unrelated research areas but there are many connections in the research problems, approaches and solutions. An obvious link is technology. The technology of precision agriculture - GPS, ...

  11. Site-Specific Dance: Promoting Social Awareness in Choreography

    ERIC Educational Resources Information Center

    MacBean, Arianne

    2004-01-01

    Site-specific dance, which is often defined as dance that occurs outside of the conventional theater space, challenges choreographers to look at, listen to, feel, and think about the space in which the dance is performed. It also asks audiences to be active participants in the performance experience. The dances have to be informed by the space and…

  12. Site-specific nicking within the adenovirus inverted terminal repetition.

    PubMed Central

    Chow, K C; Pearson, G D

    1984-01-01

    Site-specific nicking occurs on the l-strand, but not on the r-strand, of the adenovirus left inverted terminal repeat. Nicks are presumably introduced into double- or single-stranded DNA by a cellular endonuclease in an ATP-independent reaction. The consensus nick site has the sequence: (sequence in text). Images PMID:6322107

  13. Site-Specific Recombination Strategies for Engineering Actinomycete Genomes

    PubMed Central

    Herrmann, Simone; Siegl, Theresa; Luzhetska, Marta; Petzke, Lutz; Jilg, Caroline; Welle, Elisabeth; Erb, Annette; Leadlay, Peter F.; Bechthold, Andreas

    2012-01-01

    The feasibility of using technologies based on site-specific recombination in actinomycetes was shown several years ago. Despite their huge potential, these technologies mostly have been used for simple marker removal from a chromosome. In this paper, we present different site-specific recombination strategies for genome engineering in several actinomycetes belonging to the genera Streptomyces, Micromonospora, and Saccharothrix. Two different systems based on Cre/loxP and Dre/rox have been utilized for numerous applications. The activity of the Cre recombinase on the heterospecific loxLE and loxRE sites was similar to its activity on wild-type loxP sites. Moreover, an apramycin resistance marker flanked by the loxLERE sites was eliminated from the Streptomyces coelicolor M145 genome at a surprisingly high frequency (80%) compared to other bacteria. A synthetic gene encoding the Dre recombinase was constructed and successfully expressed in actinomycetes. We developed a marker-free expression method based on the combination of phage integration systems and site-specific recombinases. The Cre recombinase has been used in the deletion of huge genomic regions, including the phenalinolactone, monensin, and lipomycin biosynthetic gene clusters from Streptomyces sp. strain Tü6071, Streptomyces cinnamonensis A519, and Streptomyces aureofaciens Tü117, respectively. Finally, we also demonstrated the site-specific integration of plasmid and cosmid DNA into the chromosome of actinomycetes catalyzed by the Cre recombinase. We anticipate that the strategies presented here will be used extensively to study the genetics of actinomycetes. PMID:22247163

  14. Development of site-specific locking plates for acetabular fractures.

    PubMed

    Xu, Meng; Zhang, Li-Hai; Zhang, Ying-Ze; He, Chun-Qing; Zhang, Li-Cheng; Wang, Yan; Tang, Pei-Fu

    2013-05-01

    Site-specific locking plates have gained popularity for the treatment of fractures. However, the clinical use of a site-specific locking plate for acetabular fractures remains untested due to production limits. To design a universal site-specific locking plate for acetabular fractures, the 3-dimensional (3D) photographic records of 171 pelvises were retrospectively studied to generate a universal posterior innominate bone surface. Using 3D photographical processing software, the 3D coordinate system was reset according to bony landmarks and was scaled based on the acetabular diameter to allow a direct comparison between surfaces. The measured surface was separated into measurement units. At each measurement unit, the authors calculated the average z-axis values in all samples and obtained the 3D coordinate values of the point cloud that could be reconstructed into the universal surface. A plate was subsequently designed in 3D photographical processing software, and the orientation and distribution of locking screws was included. To manufacture a plate, the data were entered into Unigraphics NX version 6.0 software (Siemens PLM Software, Co, Ltd, Plano, Texas) and a CNC digital milling machine (FANUC Co, Ltd, Yamanashi, Japan). The resulting locking plate fit excellently with the reduced bone surface intraoperatively. Plate contouring was avoided intraoperatively. Universal 3.5-mm locking screws locked successfully into the plate, and their orientations were consistent with the design. No screw yielded to acetabular penetration. This method of designing a site-specific acetabular locking plate is practical, and the plates are suitable for clinical use. These site-specific locking plates may be an option for the treatment of acetabular fractures, particularly in elderly patients.

  15. Zeolite Vitrification Demonstration Program: characterization of nonradioactive demonstration product

    SciTech Connect

    Daniel, J L

    1982-09-01

    An extensive characterization of specimens of the glass produced during Run 4 (ZVDP-4) of the nonradioactive phase of the Three Mile Island (TMI) Zeolite Vitrification Demonstration Program was conducted by the Materials characterization Center (MCC) at Pacific Northwest Laboratory (PNL). The characteristics of the ZVDP-4 glass were compared with those of MCC's borosilicate reference glass, MCC 76-68. Tests included analyses of composition, density and phases present, the MCC-1P Static Leach Test Method, measurements of tensile strength and impact behavior, and evaluation of high-temperature vaporization.

  16. Adapter Reagents for Protein Site Specific Dye Labeling

    PubMed Central

    Thompson, Darren A.; Evans, Eric G. B.; Kasza, Tomas; Millhauser, Glenn L.; Dawson, Philip E.

    2016-01-01

    Chemoselective protein labeling remains a significant challenge in chemical biology. Although many selective labeling chemistries have been reported, the practicalities of matching the reaction with appropriately functionalized proteins and labeling reagents is often a challenge. For example, we encountered the challenge of site specifically labeling the cellular form of the murine Prion protein with a fluorescent dye. To facilitate this labeling, a protein was expressed with site specific p-acetylphenylalanine. However, the utility of this aceto-phenone reactive group is hampered by the severe lack of commercially available aminooxy fluorophores. Here we outline a general strategy for the efficient solid phase synthesis of adapter reagents capable of converting maleimido-labels into aminooxy or azide functional groups that can be further tuned for desired length or solubility properties. The utility of the adapter strategy is demonstrated in the context of fluorescent labeling of the murine Prion protein through an adapted aminooxy-Alexa dye. PMID:24599728

  17. Marker-free site-specific gene integration in plants.

    PubMed

    Srivastava, Vibha; Ow, David W

    2004-12-01

    For nearly 15 years, the use of site-specific recombination systems in plants has focused on the deletion or integration of DNA. Each of these applications offers practical solutions to two problems in biotechnology: the presence of unneeded DNA in the transgene locus and variation in the locus structure among independent transgenic lines. Given that each of these separate applications is becoming more practical for commercial use, it is time to consider combining them into an integrated technology. Here we propose a strategy for a "combined step" method that makes use of two site-specific recombination systems: one for integrating the DNA and a second for removing sequences that are not needed after DNA transfer. This strategy is based on published data and exemplifies the use of the Cre-lox, FLP-FRT and R-RS inducible systems.

  18. Site-specific magnetization reversal studies of magnetite

    SciTech Connect

    Cady, A.; Haskel, D.; Lang, J. C.; Islam, Z.; Srajer, G.; Ankudinov, A.; Subias, G.; Garcia, J.

    2006-04-01

    The mechanism of magnetization reversal in magnetite (Fe{sub 3}O{sub 4}) single crystals was studied using site-specific magnetic sensitive diffraction anomalous near-edge structure. By exploiting the angular dependence of the cross section, we are able to show that the mechanism of reversal involves a mixture of coherent rotation and domain formation. The results reveal additional details to that provided by XMCD measurements, which average over nonequivalent sites.

  19. Pretargeted PET Imaging Using a Site-Specifically Labeled Immunoconjugate.

    PubMed

    Cook, Brendon E; Adumeau, Pierre; Membreno, Rosemery; Carnazza, Kathryn E; Brand, Christian; Reiner, Thomas; Agnew, Brian J; Lewis, Jason S; Zeglis, Brian M

    2016-08-17

    In recent years, both site-specific bioconjugation techniques and bioorthogonal pretargeting strategies have emerged as exciting technologies with the potential to improve the safety and efficacy of antibody-based nuclear imaging. In the work at hand, we have combined these two approaches to create a pretargeted PET imaging strategy based on the rapid and bioorthogonal inverse electron demand Diels-Alder reaction between a (64)Cu-labeled tetrazine radioligand ((64)Cu-Tz-SarAr) and a site-specifically modified huA33-trans-cyclooctene immunoconjugate ((ss)huA33-PEG12-TCO). A bioconjugation strategy that harnesses enzymatic transformations and strain-promoted azide-alkyne click chemistry was used to site-specifically append PEGylated TCO moieties to the heavy chain glycans of the colorectal cancer-targeting huA33 antibody. Preclinical in vivo validation studies were performed in athymic nude mice bearing A33 antigen-expressing SW1222 human colorectal carcinoma xenografts. To this end, mice were administered (ss)huA33-PEG12-TCO via tail vein injection and-following accumulation intervals of 24 or 48 h-(64)Cu-Tz-SarAr. PET imaging and biodistribution studies reveal that this strategy clearly delineates tumor tissue as early as 1 h post-injection (6.7 ± 1.7%ID/g at 1 h p.i.), producing images with excellent contrast and high tumor-to-background activity concentration ratios (tumor:muscle = 21.5 ± 5.6 at 24 h p.i.). Furthermore, dosimetric calculations illustrate that this pretargeting approach produces only a fraction of the overall effective dose (0.0214 mSv/MBq; 0.079 rem/mCi) of directly labeled radioimmunoconjugates. Ultimately, this method effectively facilitates the high contrast pretargeted PET imaging of colorectal carcinoma using a site-specifically modified immunoconjugate.

  20. Site-Specific Biomolecule Labeling with Gold Clusters

    PubMed Central

    Ackerson, Christopher J.; Powell, Richard D.; Hainfeld, James F.

    2013-01-01

    Site-specific labeling of biomolecules in vitro with gold clusters can enhance the information content of electron cryomicroscopy experiments. This chapter provides a practical overview of well-established techniques for forming biomolecule/gold cluster conjugates. Three bioconjugation chemistries are covered: Linker-mediated bioconjugation, direct gold–biomolecule bonding, and coordination-mediated bonding of nickel(II) nitrilotriacetic acid (NTA)-derivatized gold clusters to polyhistidine (His)-tagged proteins. PMID:20887859

  1. Site-specific gene modification by PNAs conjugated to psoralen.

    PubMed

    Kim, Ki-Hyun; Nielsen, Peter E; Glazer, Peter M

    2006-01-10

    DNA-binding molecules, including triplex-forming oligonucleotides (TFOs) and peptide nucleic acids (PNAs), can be utilized to introduce site-specific mutations or to promote recombination at selected genomic sites. To further evaluate the utility of PNAs for site-specific gene modification, we tested dimeric bis-PNAs conjugated to psoralen. These PNAs are designed to form a triplex-invasion complex within the supF reporter gene in an episomal shuttle vector and to direct site-specific photoadduct formation by the conjugated psoralen. The psoralen-bis-PNA conjugate was found to direct photoadduct formation to the intended 5'-TpA base step next to the PNA-binding site, and the photoadduct formation efficiency displayed both concentration and UVA irradiation dependence. The effect of PNA-targeted photoadducts in a mammalian system was tested by SV40-based shuttle vector assay. After in vitro binding, we found that photoadducts directed by PNAs conjugated to psoralen-induced mutations at frequencies in the range of 0.46%, 6.5-fold above the background. In a protocol for intracellular gene targeting in the episomal shuttle vector, the psoralen-PNA-induced mutation frequency was 0.13%, 3.5-fold higher than the background. Most of the induced mutations were deletions and single-base-pair substitutions at or adjacent to the targeted PNA-binding and photoadduct-formation sites. When the results are taken together, they demonstrate the ability of bis-PNAs conjugated with psoralen to mediate site-specific gene modification, and they further support the development of PNAs as tools for gene-targeting applications.

  2. A general approach to site-specific antibody drug conjugates

    PubMed Central

    Tian, Feng; Lu, Yingchun; Manibusan, Anthony; Sellers, Aaron; Tran, Hon; Sun, Ying; Phuong, Trung; Barnett, Richard; Hehli, Brad; Song, Frank; DeGuzman, Michael J.; Ensari, Semsi; Pinkstaff, Jason K.; Sullivan, Lorraine M.; Biroc, Sandra L.; Cho, Ho; Schultz, Peter G.; DiJoseph, John; Dougher, Maureen; Ma, Dangshe; Dushin, Russell; Leal, Mauricio; Tchistiakova, Lioudmila; Feyfant, Eric; Gerber, Hans-Peter; Sapra, Puja

    2014-01-01

    Using an expanded genetic code, antibodies with site-specifically incorporated nonnative amino acids were produced in stable cell lines derived from a CHO cell line with titers over 1 g/L. Using anti-5T4 and anti-Her2 antibodies as model systems, site-specific antibody drug conjugates (NDCs) were produced, via oxime bond formation between ketones on the side chain of the incorporated nonnative amino acid and hydroxylamine functionalized monomethyl auristatin D with either protease-cleavable or noncleavable linkers. When noncleavable linkers were used, these conjugates were highly stable and displayed improved in vitro efficacy as well as in vivo efficacy and pharmacokinetic stability in rodent models relative to conventional antibody drug conjugates conjugated through either engineered surface-exposed or reduced interchain disulfide bond cysteine residues. The advantages of the oxime-bonded, site-specific NDCs were even more apparent when low–antigen-expressing (2+) target cell lines were used in the comparative studies. NDCs generated with protease-cleavable linkers demonstrated that the site of conjugation had a significant impact on the stability of these rationally designed prodrug linkers. In a single-dose rat toxicology study, a site-specific anti-Her2 NDC was well tolerated at dose levels up to 90 mg/kg. These experiments support the notion that chemically defined antibody conjugates can be synthesized in commercially relevant yields and can lead to antibody drug conjugates with improved properties relative to the heterogeneous conjugates formed by nonspecific chemical modification. PMID:24443552

  3. Site specific atomic polarizabilities in endohedral fullerenes and carbon onions.

    PubMed

    Zope, Rajendra R; Bhusal, Shusil; Basurto, Luis; Baruah, Tunna; Jackson, Koblar

    2015-08-28

    We investigate the polarizability of trimetallic nitride endohedral fullerenes by partitioning the total polarizability into site specific components. This analysis indicates that the polarizability of the endohedral fullerene is essentially due to the outer fullerene cage and has insignificant contribution from the encapsulated unit. Thus, the outer fullerene cages effectively shield the encapsulated clusters and behave like Faraday cages. The polarizability of endohedral fullerenes is slightly smaller than the polarizability of the corresponding bare carbon fullerenes. The application of the site specific polarizabilities to C60@C240 and C60@C180 onions shows that, compared to the polarizability of isolated C60 fullerene, the encapsulation of the C60 in C240 and C180 fullerenes reduces its polarizability by 75% and 83%, respectively. The differences in the polarizability of C60 in the two onions is a result of differences in the bonding (intershell electron transfer), fullerene shell relaxations, and intershell separations. The site specific analysis further shows that the outer atoms in a fullerene shell contribute most to the fullerene polarizability.

  4. Site specific atomic polarizabilities in endohedral fullerenes and carbon onions

    NASA Astrophysics Data System (ADS)

    Zope, Rajendra R.; Bhusal, Shusil; Basurto, Luis; Baruah, Tunna; Jackson, Koblar

    2015-08-01

    We investigate the polarizability of trimetallic nitride endohedral fullerenes by partitioning the total polarizability into site specific components. This analysis indicates that the polarizability of the endohedral fullerene is essentially due to the outer fullerene cage and has insignificant contribution from the encapsulated unit. Thus, the outer fullerene cages effectively shield the encapsulated clusters and behave like Faraday cages. The polarizability of endohedral fullerenes is slightly smaller than the polarizability of the corresponding bare carbon fullerenes. The application of the site specific polarizabilities to C60@C240 and C60@C180 onions shows that, compared to the polarizability of isolated C60 fullerene, the encapsulation of the C60 in C240 and C180 fullerenes reduces its polarizability by 75% and 83%, respectively. The differences in the polarizability of C60 in the two onions is a result of differences in the bonding (intershell electron transfer), fullerene shell relaxations, and intershell separations. The site specific analysis further shows that the outer atoms in a fullerene shell contribute most to the fullerene polarizability.

  5. Site-specifically radioiodinated antibody for targeting tumors

    SciTech Connect

    Rea, D.W.; Ultee, M.E.; Belinka, B.A. Jr.; Coughlin, D.J.; Alvarez, V.L. )

    1990-02-01

    Labeling of an antibody site specifically through its carbohydrate regions preserves its antigen-binding activity. Previously site-specific labeling studies have conjugated antibodies with metallic radioisotopes or drugs. We now report site-specific labeling with a new radioiodinated compound, 2-hydroxy-5-iodo-3-methylbenzoyl hydrazide, whose synthesis we described earlier. The compound is reacted with aldehyde groups produced by specific oxidation of the carbohydrate portion of the antibody with sodium m-periodate. Optimized conjugation conditions give good recovery of active antibody containing 10 groups per molecule. The conjugate is stable in solution for at least several weeks at both 4 and -70 degrees C. When injected into nude mice bearing LS174T human cancer xenografts, the conjugate of B72.3 antibody localizes well to tumor tissue, with low uptake by other organs. This biodistribution is similar to that of conjugate prepared by using solid-phase chloramine-T (Iodohead). There are only two significant differences. First, the carbohydrate conjugate is much less susceptible to dehalogenation, and thus shows much less thyroid uptake. Secondly, the biological half-life of the carbohydrate conjugate was about half that of the chloramine-T one. This could be due primarily to lysis of the hydrazine bond through which the antibody is attached to the compound, which would then be excreted rapidly by itself. The new reagent will be especially useful for antibodies which either cannot be labeled by chloramine-T methods, or whose activity is impaired by them.

  6. Site specific atomic polarizabilities in endohedral fullerenes and carbon onions

    SciTech Connect

    Zope, Rajendra R. Baruah, Tunna; Bhusal, Shusil; Basurto, Luis; Jackson, Koblar

    2015-08-28

    We investigate the polarizability of trimetallic nitride endohedral fullerenes by partitioning the total polarizability into site specific components. This analysis indicates that the polarizability of the endohedral fullerene is essentially due to the outer fullerene cage and has insignificant contribution from the encapsulated unit. Thus, the outer fullerene cages effectively shield the encapsulated clusters and behave like Faraday cages. The polarizability of endohedral fullerenes is slightly smaller than the polarizability of the corresponding bare carbon fullerenes. The application of the site specific polarizabilities to C{sub 60}@C{sub 240} and C{sub 60}@C{sub 180} onions shows that, compared to the polarizability of isolated C{sub 60} fullerene, the encapsulation of the C{sub 60} in C{sub 240} and C{sub 180} fullerenes reduces its polarizability by 75% and 83%, respectively. The differences in the polarizability of C{sub 60} in the two onions is a result of differences in the bonding (intershell electron transfer), fullerene shell relaxations, and intershell separations. The site specific analysis further shows that the outer atoms in a fullerene shell contribute most to the fullerene polarizability.

  7. Optimization under Uncertainty of Site-Specific Turbine Configurations: Preprint

    SciTech Connect

    Quick, Julian; Dykes, Katherine; Graf, Peter; Zahle, Frederik

    2016-11-01

    Uncertainty affects many aspects of wind energy plant performance and cost. In this study, we explore opportunities for site-specific turbine configuration optimization that accounts for uncertainty in the wind resource. As a demonstration, a simple empirical model for wind plant cost of energy is used in an optimization under uncertainty to examine how different risk appetites affect the optimal selection of a turbine configuration for sites of different wind resource profiles. If there is unusually high uncertainty in the site wind resource, the optimal turbine configuration diverges from the deterministic case and a generally more conservative design is obtained with increasing risk aversion on the part of the designer.

  8. Optimization Under Uncertainty of Site-Specific Turbine Configurations

    SciTech Connect

    Quick, J.; Dykes, K.; Graf, P.; Zahle, F.

    2016-10-03

    Uncertainty affects many aspects of wind energy plant performance and cost. In this study, we explore opportunities for site-specific turbine configuration optimization that accounts for uncertainty in the wind resource. As a demonstration, a simple empirical model for wind plant cost of energy is used in an optimization under uncertainty to examine how different risk appetites affect the optimal selection of a turbine configuration for sites of different wind resource profiles. Lastly, if there is unusually high uncertainty in the site wind resource, the optimal turbine configuration diverges from the deterministic case and a generally more conservative design is obtained with increasing risk aversion on the part of the designer.

  9. Site-specific labeling of proteins for electron microscopy

    PubMed Central

    Dambacher, Corey M.; Lander, Gabriel C.

    2015-01-01

    Electron microscopy is commonly employed to determine the subunit organization of large macromolecular assemblies. However, the field lacks a robust molecular labeling methodology for unambiguous identification of constituent subunits. We present a strategy that exploits the unique properties of an unnatural amino acid in order to enable site-specific attachment of a single, readily identifiable protein label at any solvent-exposed position on the macromolecular surface. Using this method, we show clear labeling of a subunit within the 19S proteasome lid subcomplex that has not been amenable to labeling by traditional approaches. PMID:26409249

  10. Site Specific Probable Maximum Precipitation Estimates and Professional Judgement

    NASA Astrophysics Data System (ADS)

    Hayes, B. D.; Kao, S. C.; Kanney, J. F.; Quinlan, K. R.; DeNeale, S. T.

    2015-12-01

    State and federal regulatory authorities currently rely upon the US National Weather Service Hydrometeorological Reports (HMRs) to determine probable maximum precipitation (PMP) estimates (i.e., rainfall depths and durations) for estimating flooding hazards for relatively broad regions in the US. PMP estimates for the contributing watersheds upstream of vulnerable facilities are used to estimate riverine flooding hazards while site-specific estimates for small water sheds are appropriate for individual facilities such as nuclear power plants. The HMRs are often criticized due to their limitations on basin size, questionable applicability in regions affected by orographic effects, their lack of consist methods, and generally by their age. HMR-51 for generalized PMP estimates for the United States east of the 105th meridian, was published in 1978 and is sometimes perceived as overly conservative. The US Nuclear Regulatory Commission (NRC), is currently reviewing several flood hazard evaluation reports that rely on site specific PMP estimates that have been commercially developed. As such, NRC has recently investigated key areas of expert judgement via a generic audit and one in-depth site specific review as they relate to identifying and quantifying actual and potential storm moisture sources, determining storm transposition limits, and adjusting available moisture during storm transposition. Though much of the approach reviewed was considered a logical extension of HMRs, two key points of expert judgement stood out for further in-depth review. The first relates primarily to small storms and the use of a heuristic for storm representative dew point adjustment developed for the Electric Power Research Institute by North American Weather Consultants in 1993 in order to harmonize historic storms for which only 12 hour dew point data was available with more recent storms in a single database. The second issue relates to the use of climatological averages for spatially

  11. Optimization Under Uncertainty of Site-Specific Turbine Configurations

    NASA Astrophysics Data System (ADS)

    Quick, J.; Dykes, K.; Graf, P.; Zahle, F.

    2016-09-01

    Uncertainty affects many aspects of wind energy plant performance and cost. In this study, we explore opportunities for site-specific turbine configuration optimization that accounts for uncertainty in the wind resource. As a demonstration, a simple empirical model for wind plant cost of energy is used in an optimization under uncertainty to examine how different risk appetites affect the optimal selection of a turbine configuration for sites of different wind resource profiles. If there is unusually high uncertainty in the site wind resource, the optimal turbine configuration diverges from the deterministic case and a generally more conservative design is obtained with increasing risk aversion on the part of the designer.

  12. Site-Specific, Sustained Release of Drugs to the Brain

    NASA Astrophysics Data System (ADS)

    Bodor, Nicholas; Farag, Hassan H.; Brewster, Marcus E.

    1981-12-01

    A dihydropyridine-pyridinium salt type of redox system is used in a general and flexible method for site-specific or sustained delivery (or both) of drugs to the brain. A biologically active compound linked to a lipoidal dihydropyridine carrier easily penetrates the blood-brain barrier. Oxidation of the carrier part in vivo to the ionic pyridinium salt prevents its elimination from the brain, while elimination from the general circulation is accelerated. Subsequent cleavage of the quaternary carrier-drug species results in sustained delivery of the drug in the brain and facile elimination of the carrier part.

  13. Site-specific tumor-targeted fluorescent contrast agents

    NASA Astrophysics Data System (ADS)

    Achilefu, Samuel I.; Bugaj, Joseph E.; Dorshow, Richard B.; Jimenez, Hermo N.; Rajagopalan, Raghavan; Wilhelm, R. Randy; Webb, Elizabeth G.; Erion, Jack L.

    2001-01-01

    Site-specific delivery of drugs and contrast agents to tumors protects normal tissues from the cytotoxic effect of drugs, and enhances the contrast between normal and diseased tissues. In optical medicine, biocompatible dyes can be used as photo therapeutics or as contrast agents. Previous studies have shown that the use of covalent or non-covalent dye conjugates of carries such as antibodies, liposomes, and polysaccharides improves the delivery of such molecules to tumors. However, large biomolecules can elicit adverse immunogenic reactions and also result in prolonged blood circulation times, delaying visualization of target tissues. A viable alternative to this strategy is to use small bioactive molecule-dye conjugates. These molecules have several advantages over large biomolecules, including ease of synthesis of a variety of high purity compounds for combinatorial screening of new targets, enhanced diffusivity to solid tumors, and the ability to affect the pharmocokinetics of the conjugates by minor structural changes. Thus, we conjugated a near IR light absorbing dye to bioactive peptides that specifically target over expressed tumor receptors in established rat tumor lines. High tumor uptake of the conjugates was obtained without loss of either the peptide receptor affinity or the dye fluorescence. These findings demonstrate the efficacy of a small peptide-dye conjugate strategy for in vivo tumor imaging. Site-specific delivery of photodynamic therapy agents may also benefit form this approach.

  14. Recent advances in covalent, site-specific protein immobilization

    PubMed Central

    Meldal, Morten; Schoffelen, Sanne

    2016-01-01

    The properties of biosensors, biomedical implants, and other materials based on immobilized proteins greatly depend on the method employed to couple the protein molecules to their solid support. Covalent, site-specific immobilization strategies are robust and can provide the level of control that is desired in this kind of application. Recent advances include the use of enzymes, such as sortase A, to couple proteins in a site-specific manner to materials such as microbeads, glass, and hydrogels. Also, self-labeling tags such as the SNAP-tag can be employed. Last but not least, chemical approaches based on bioorthogonal reactions, like the azide–alkyne cycloaddition, have proven to be powerful tools. The lack of comparative studies and quantitative analysis of these immobilization methods hampers the selection process of the optimal strategy for a given application. However, besides immobilization efficiency, the freedom in selecting the site of conjugation and the size of the conjugation tag and the researcher’s expertise regarding molecular biology and/or chemical techniques will be determining factors in this regard. PMID:27785356

  15. Temporally-controlled site-specific recombination in zebrafish.

    PubMed

    Hans, Stefan; Kaslin, Jan; Freudenreich, Dorian; Brand, Michael

    2009-01-01

    Conventional use of the site-specific recombinase Cre is a powerful technology in mouse, but almost absent in other vertebrate model organisms. In zebrafish, Cre-mediated recombination efficiency was previously very low. Here we show that using transposon-mediated transgenesis, Cre is in fact highly efficient in this organism. Furthermore, temporal control of recombination can be achieved by using the ligand-inducible CreER(T2). Site-specific recombination only occurs upon administration of the drug tamoxifen (TAM) or its active metabolite, 4-hydroxy-tamoxifen (4-OHT). Cre-mediated recombination is detectable already 4 or 2 hours after administration of TAM or 4-OHT, demonstrating fast recombination kinetics. In addition, low doses of TAM allow mosaic labeling of single cells. Combined, our results show that conditional Cre/lox will be a valuable tool for both, embryonic and adult zebrafish studies. Furthermore, single copy insertion transgenesis of Cre/lox constructs suggest a strategy suitable also for other organisms.

  16. Towards soft robotic devices for site-specific drug delivery.

    PubMed

    Alici, Gursel

    2015-01-01

    Considerable research efforts have recently been dedicated to the establishment of various drug delivery systems (DDS) that are mechanical/physical, chemical and biological/molecular DDS. In this paper, we report on the recent advances in site-specific drug delivery (site-specific, controlled, targeted or smart drug delivery are terms used interchangeably in the literature, to mean to transport a drug or a therapeutic agent to a desired location within the body and release it as desired with negligibly small toxicity and side effect compared to classical drug administration means such as peroral, parenteral, transmucosal, topical and inhalation) based on mechanical/physical systems consisting of implantable and robotic drug delivery systems. While we specifically focus on the robotic or autonomous DDS, which can be reprogrammable and provide multiple doses of a drug at a required time and rate, we briefly cover the implanted DDS, which are well-developed relative to the robotic DDS, to highlight the design and performance requirements, and investigate issues associated with the robotic DDS. Critical research issues associated with both DDSs are presented to describe the research challenges ahead of us in order to establish soft robotic devices for clinical and biomedical applications.

  17. Editing livestock genomes with site-specific nucleases.

    PubMed

    Carlson, Daniel F; Tan, Wenfang; Hackett, Perry B; Fahrenkrug, Scott C

    2013-01-01

    Over the past 5 years there has been a major transformation in our ability to precisely manipulate the genomes of animals. Efficiencies of introducing precise genetic alterations in large animal genomes have improved 100000-fold due to a succession of site-specific nucleases that introduce double-strand DNA breaks with a specificity of 10(-9). Herein we describe our applications of site-specific nucleases, especially transcription activator-like effector nucleases, to engineer specific alterations in the genomes of pigs and cows. We can introduce variable changes mediated by non-homologous end joining of DNA breaks to inactive genes. Alternatively, using homology-directed repair, we have introduced specific changes that support either precise alterations in a gene's encoded polypeptide, elimination of the gene or replacement by another unrelated DNA sequence. Depending on the gene and the mutation, we can achieve 10%-50% effective rates of precise mutations. Applications of the new precision genetics are extensive. Livestock now can be engineered with selected phenotypes that will augment their value and adaption to variable ecosystems. In addition, animals can be engineered to specifically mimic human diseases and disorders, which will accelerate the production of reliable drugs and devices. Moreover, animals can be engineered to become better providers of biomaterials used in the medical treatment of diseases and disorders.

  18. Site specific modification of the human plasma proteome by methylglyoxal.

    PubMed

    Kimzey, Michael J; Kinsky, Owen R; Yassine, Hussein N; Tsaprailis, George; Stump, Craig S; Monks, Terrence J; Lau, Serrine S

    2015-12-01

    Increasing evidence identifies dicarbonyl stress from reactive glucose metabolites, such as methylglyoxal (MG), as a major pathogenic link between hyperglycemia and complications of diabetes. MG covalently modifies arginine residues, yet the site specificity of this modification has not been thoroughly investigated. Sites of MG adduction in the plasma proteome were identified using LC-MS/MS analysis in vitro following incubation of plasma proteins with MG. Treatment of plasma proteins with MG yielded 14 putative MG hotspots from five plasma proteins (albumin [nine hotspots], serotransferrin, haptoglobin [2 hotspots], hemopexin, and Ig lambda-2 chain C regions). The search results revealed two versions of MG-arginine modification, dihydroxyimidazolidine (R+72) and hydroimidazolone (R+54) adducts. One of the sites identified was R257 in human serum albumin, which is a critical residue located in drug binding site I. This site was validated as a target for MG modification by a fluorescent probe displacement assay, which revealed significant drug dissociation at 300 μM MG from a prodan-HSA complex (75 μM). Moreover, twelve human plasma samples (six male, six female, with two type 2 diabetic subjects from both genders) were analyzed using multiple reaction monitoring (MRM) tandem mass spectrometry and revealed the presence of the MG-modified albumin R257 peptide. These data provide insights into the nature of the site-specificity of MG modification of arginine, which may be useful for therapeutic treatments that aim to prevent MG-mediated adverse responses in patients.

  19. Site-specific features influence sediment stability of intertidal flats

    NASA Astrophysics Data System (ADS)

    Defew, Emma C.; Tolhurst, Trevor J.; Paterson, David M.

    The factors that influence the sediment stability and the transport of estuarine mudflats are not yet fully understood but knowledge of them is essential in coastal engineering applications and pollution ecology studies. The suggestion that variation in predictive models of sediment stability might be due to site-specific characteristics is investigated using data from four estuarine mudflats (Eden Estuary, Scotland, the Biezelingsche Ham, Zandkreek, and Molenplaat mudflats in The Netherlands). These estuaries differ in their environmental conditions, macrofaunal species composition and local features (e.g. Enteromorpha mats, migratory biofilms). Stable and unstable sediments were compared, and mean chlorophyll-a concentrations and granulometry of the sediments were significantly different between the two groups. Step-wise multiple linear regressions were applied to the sediment stability data of all sites to establish the influences on erosion threshold of microphytobenthic biomass, water content, granulometry, organic carbon content and the abundance of dominant macrofaunal species. The stability of each site was influenced by different factors. Sediment stability of the Eden Estuary was affected by the Enteromorpha bloom; Biezelingsche Ham was influenced by the highly migratory nature of the diatom biofilms and the abundance of Corophium volutator; the polychaete worm Arenicola marina had a net negative effect on sediment stability of the Zandkreek; and the Molenplaat was influenced by microphytobenthic biomass. This research highlights the need for site-specific calibration of models and suggests that a universal proxy parameter for sediment stability is unlikely to be obtained.

  20. Site-Specific Genome Engineering in Human Pluripotent Stem Cells.

    PubMed

    Merkert, Sylvia; Martin, Ulrich

    2016-06-24

    The possibility to generate patient-specific induced pluripotent stem cells (iPSCs) offers an unprecedented potential of applications in clinical therapy and medical research. Human iPSCs and their differentiated derivatives are tools for diseases modelling, drug discovery, safety pharmacology, and toxicology. Moreover, they allow for the engineering of bioartificial tissue and are promising candidates for cellular therapies. For many of these applications, the ability to genetically modify pluripotent stem cells (PSCs) is indispensable, but efficient site-specific and safe technologies for genetic engineering of PSCs were developed only recently. By now, customized engineered nucleases provide excellent tools for targeted genome editing, opening new perspectives for biomedical research and cellular therapies.

  1. Optimization Under Uncertainty of Site-Specific Turbine Configurations

    DOE PAGES

    Quick, J.; Dykes, K.; Graf, P.; ...

    2016-10-03

    Uncertainty affects many aspects of wind energy plant performance and cost. In this study, we explore opportunities for site-specific turbine configuration optimization that accounts for uncertainty in the wind resource. As a demonstration, a simple empirical model for wind plant cost of energy is used in an optimization under uncertainty to examine how different risk appetites affect the optimal selection of a turbine configuration for sites of different wind resource profiles. Lastly, if there is unusually high uncertainty in the site wind resource, the optimal turbine configuration diverges from the deterministic case and a generally more conservative design is obtainedmore » with increasing risk aversion on the part of the designer.« less

  2. Synthesis of Site-Specifically (13)C Labeled Linoleic Acids.

    PubMed

    Offenbacher, Adam R; Zhu, Hui; Klinman, Judith P

    2016-10-12

    Soybean lipoxygenase-1 (SLO-1) catalyzes the C-H abstraction from the reactive carbon (C-11) in linoleic acid as the first and rate-determining step in the formation of alkylhydroperoxides. While previous labeling strategies have focused on deuterium labeling to ascertain the primary and secondary kinetic isotope effects for this reaction, there is an emerging interest and need for selectively enriched (13)C isotopologues. In this report, we present synthetic strategies for site-specific (13)C labeled linoleic acid substrates. We take advantage of a Corey-Fuchs formyl to terminal (13)C-labeled alkyne conversion, using (13)CBr4 as the labeling source, to reduce the number of steps from a previous fatty acid (13)C synthetic labeling approach. The labeled linoleic acid substrates are useful as nuclear tunneling markers and for extracting active site geometries of the enzyme-substrate complex in lipoxygenase.

  3. Methods for generating phosphorylation site-specific immunological reagents

    DOEpatents

    Anderson, Carl W.; Appella, Ettore; Sakaguchi, Kazuyasu

    2001-01-01

    The present invention provides methods for generating phosphorylation site-specific immunological reagents. More specifically, a phosphopeptide mimetic is incorporated into a polypeptide in place of a phosphorylated amino acid. The polypeptide is used as antigen by standard methods to generate either monoclonal or polyclonal antibodies which cross-react with the naturally phosphorylated polypeptide. The phosphopeptide mimetic preferably contains a non-hydrolyzable linkage from the appropriate carbon atom of the amino acid residue to a phosphate group. A preferred linkage is a CF.sub.2 group. Such a linkage is used to generate the phosphoserine mimetic F.sub.2 Pab, which is incorporated into a polypeptide sequence derived from p53 to produce antibodies which recognize a specific phosphorylation state of p53. A CF.sub.2 group linkage is also used to produce the phosphothreonine mimetic F.sub.2 Pmb, and to produce the phosphotyrosine mimetic, F.sub.2 Pmp.

  4. Site-Specific Genome Engineering in Human Pluripotent Stem Cells

    PubMed Central

    Merkert, Sylvia; Martin, Ulrich

    2016-01-01

    The possibility to generate patient-specific induced pluripotent stem cells (iPSCs) offers an unprecedented potential of applications in clinical therapy and medical research. Human iPSCs and their differentiated derivatives are tools for diseases modelling, drug discovery, safety pharmacology, and toxicology. Moreover, they allow for the engineering of bioartificial tissue and are promising candidates for cellular therapies. For many of these applications, the ability to genetically modify pluripotent stem cells (PSCs) is indispensable, but efficient site-specific and safe technologies for genetic engineering of PSCs were developed only recently. By now, customized engineered nucleases provide excellent tools for targeted genome editing, opening new perspectives for biomedical research and cellular therapies. PMID:27347935

  5. Site Specific Analyses of a Spent Nuclear Fuel Transportation Accident

    SciTech Connect

    Biwer, B. M.; Chen, S. Y.

    2003-02-24

    The number of spent nuclear fuel (SNF) shipments is expected to increase significantly during the time period that the United States' inventory of SNF is sent to a final disposal site. Prior work estimated that the highest accident risks of a SNF shipping campaign to the proposed geologic repository at Yucca Mountain were in the corridor states, such as Illinois. The largest potential human health impacts would be expected to occur in areas with high population densities such as urban settings. Thus, our current study examined the human health impacts from the most plausible severe SNF transportation accidents in the Chicago metropolitan area. The RISKIND 2.0 program was used to model site-specific data for an area where the largest impacts might occur. The results have shown that the radiological human health consequences of a severe SNF rail transportation accident on average might be similar to one year of exposure to natural background radiation for those persons living a nd working in the most affected areas downwind of the actual accident location. For maximally exposed individuals, an exposure similar to about two years of exposure to natural background radiation was estimated. In addition to the accident probabilities being very low (approximately 1 chance in 10,000 or less during the entire shipping campaign), the actual human health impacts are expected to be lower if any of the accidents considered did occur, because the results are dependent on the specific location and weather conditions, such as wind speed and direction, that were selected to maximize the results. Also, comparison of the results of longer duration accident scenarios against U.S. Environmental Protection Agency guidelines was made to demonstrate the usefulness of this site-specific analysis for emergency planning purposes.

  6. Performance of site-specific parameterizations of longwave radiation

    NASA Astrophysics Data System (ADS)

    Formetta, Giuseppe; Bancheri, Marialaura; David, Olaf; Rigon, Riccardo

    2016-11-01

    In this work 10 algorithms for estimating downwelling longwave atmospheric radiation (L↓) and 1 for upwelling longwave radiation (L) are integrated into the JGrass-NewAge modelling system. The algorithms are tested against energy flux measurements available for 24 sites in North America to assess their reliability. These new JGrass-NewAge model components are used (i) to evaluate the performances of simplified models (SMs) of L↓, as presented in literature formulations, and (ii) to determine by automatic calibration the site-specific parameter sets for L↓ in SMs. For locations where calibration is not possible because of a lack of measured data, we perform a multiple regression using on-site variables, i.e. mean annual air temperature, relative humidity, precipitation, and altitude. The regressions are verified through a leave-one-out cross validation, which also gathers information about the possible errors of estimation. Most of the SMs, when executed with parameters derived from the multiple regressions, give enhanced performances compared to the corresponding literature formulation. A sensitivity analysis is carried out for each SM to understand how small variations of a given parameter influence SM performance. Regarding the L↓ simulations, the Brunt (1932) and Idso (1981) SMs, in their literature formulations, provide the best performances in many of the sites. The site-specific parameter calibration improves SM performances compared to their literature formulations. Specifically, the root mean square error (RMSE) is almost halved and the Kling-Gupta efficiency is improved at all sites. Also in this case, Brunt (1932) and Idso (1981) SMs provided the best performances. The L SM is tested by using three different temperatures (surface soil temperature, air temperature at 2 m elevation, and soil temperature at 4 cm depth) and model performances are then assessed. Results show that the best performances are achieved using the

  7. Site-specific DNA Inversion by Serine Recombinases

    PubMed Central

    2015-01-01

    Reversible site-specific DNA inversion reactions are widely distributed in bacteria and their viruses. They control a range of biological reactions that most often involve alterations of molecules on the surface of cells or phage. These programmed DNA rearrangements usually occur at a low frequency, thereby preadapting a small subset of the population to a change in environmental conditions, or in the case of phages, an expanded host range. A dedicated recombinase, sometimes with the aid of additional regulatory or DNA architectural proteins, catalyzes the inversion of DNA. RecA or other components of the general recombination-repair machinery are not involved. This chapter discusses site-specific DNA inversion reactions mediated by the serine recombinase family of enzymes and focuses on the extensively studied serine DNA invertases that are stringently controlled by the Fis-bound enhancer regulatory system. The first section summarizes biological features and general properties of inversion reactions by the Fis/enhancer-dependent serine invertases and the recently described serine DNA invertases in Bacteroides. Mechanistic studies of reactions catalyzed by the Hin and Gin invertases are then discussed in more depth, particularly with regards to recent advances in our understanding of the function of the Fis/enhancer regulatory system, the assembly of the active recombination complex (invertasome) containing the Fis/enhancer, and the process of DNA strand exchange by rotation of synapsed subunit pairs within the invertasome. The role of DNA topological forces that function in concert with the Fis/enhancer controlling element in specifying the overwhelming bias for DNA inversion over deletion and intermolecular recombination is emphasized. PMID:25844275

  8. Ultrasensitivity in signaling cascades revisited: Linking local and global ultrasensitivity estimations.

    PubMed

    Altszyler, Edgar; Ventura, Alejandra C; Colman-Lerner, Alejandro; Chernomoretz, Ariel

    2017-01-01

    Ultrasensitive response motifs, capable of converting graded stimuli into binary responses, are well-conserved in signal transduction networks. Although it has been shown that a cascade arrangement of multiple ultrasensitive modules can enhance the system's ultrasensitivity, how a given combination of layers affects a cascade's ultrasensitivity remains an open question for the general case. Here, we introduce a methodology that allows us to determine the presence of sequestration effects and to quantify the relative contribution of each module to the overall cascade's ultrasensitivity. The proposed analysis framework provides a natural link between global and local ultrasensitivity descriptors and it is particularly well-suited to characterize and understand mathematical models used to study real biological systems. As a case study, we have considered three mathematical models introduced by O'Shaughnessy et al. to study a tunable synthetic MAPK cascade, and we show how our methodology can help modelers better understand alternative models.

  9. Ultrasensitivity in signaling cascades revisited: Linking local and global ultrasensitivity estimations

    PubMed Central

    Altszyler, Edgar; Ventura, Alejandra C.; Colman-Lerner, Alejandro; Chernomoretz, Ariel

    2017-01-01

    Ultrasensitive response motifs, capable of converting graded stimuli into binary responses, are well-conserved in signal transduction networks. Although it has been shown that a cascade arrangement of multiple ultrasensitive modules can enhance the system’s ultrasensitivity, how a given combination of layers affects a cascade’s ultrasensitivity remains an open question for the general case. Here, we introduce a methodology that allows us to determine the presence of sequestration effects and to quantify the relative contribution of each module to the overall cascade’s ultrasensitivity. The proposed analysis framework provides a natural link between global and local ultrasensitivity descriptors and it is particularly well-suited to characterize and understand mathematical models used to study real biological systems. As a case study, we have considered three mathematical models introduced by O’Shaughnessy et al. to study a tunable synthetic MAPK cascade, and we show how our methodology can help modelers better understand alternative models. PMID:28662096

  10. New Waste Calcining Facility Non-radioactive Process Decontamination

    SciTech Connect

    Swenson, Michael Clair

    2001-09-01

    This report documents the results of a test of the New Calcining Facility (NWCF) process decontamination system. The decontamination system test occurred in December 1981, during non-radioactive testing of the NWCF. The purpose of the decontamination system test was to identify equipment whose design prevented effective calcine removal and decontamination. Effective equipment decontamination was essential to reduce radiation fields for in-cell work after radioactive processing began. The decontamination system test began with a pre-decontamination inspection of the equipment. The pre-decontamination inspection documented the initial condition and cleanliness of the equipment. It provided a basis for judging the effectiveness of the decontamination. The decontamination consisted of a series of equipment flushes using nitric acid and water. A post-decontamination equipment inspection determined the effectiveness of the decontamination. The pre-decontamination and post-decontamination equipment inspections were documented with hotographs. The decontamination system was effective in removing calcine from most of the NWCF equipment as evidenced by little visible calcine residue in the equipment after decontamination. The decontamination test identified four areas where the decontamination system required improvement. These included the Calciner off-gas line, Cyclone off-gas line, fluidizing air line, and the Calciner baffle plates. Physical modifications to enhance decontamination were made to those areas, resulting in an effective NWCF decontamination system.

  11. New Waste Calcining Facility Non-Radioactive Process Decontamination

    SciTech Connect

    Swenson, Michael C.

    2001-09-30

    This report documents the results of a test of the New Calcining Facility (NWCF) process decontamination system. The decontamination system test occurred in December 1981, during non-radioactive testing of the NWCF. The purpose of the decontamination system test was to identify equipment whose design prevented effective calcine removal and decontamination. Effective equipment decontamination was essential to reduce radiation fields for in-cell work after radioactive processing began. The decontamination system test began with a pre-decontamination inspection of the equipment. The pre- decontamination inspection documented the initial condition and cleanliness of the equipment. It provided a basis for judging the effectiveness of the decontamination. The decontamination consisted of a series of equipment flushes using nitric acid and water. A post-decontamination equipment inspection determined the effectiveness of the decontamination. The pre-decontamination and post-decontamination equipment inspections were documented with photographs. The decontamination system was effective in removing calcine from most of the NWCF equipment as evidenced by little visible calcine residue in the equipment after decontamination. The decontamination test identified four areas where the decontamination system required improvement. These included the Calciner off-gas line, Cyclone off-gas line, fluidizing air line, and the Calciner baffle plates. Physical modifications to enhance decontamination were made to those areas, resulting in an effective NWCF decontamination system.

  12. Ascorbic acid: Nonradioactive extracellular space marker in canine heart

    SciTech Connect

    Reil, G.H.; Frombach, R.; Kownatzki, R.; Quante, W.; Lichtlen, P.R. )

    1987-11-01

    The distribution pattern of ascorbic acid and L-({sup 14}C)ascorbic acid in myocardial tissue was compared with those of the classical radioactive extracellular space markers ({sup 3}H)-inulin, ({sup 3}H)sucrose, and Na{sup 82}Br. A new polarographic techniques was developed for analogue registration of ascorbic acid concentration in coronary venous blood. The kinetic data of the markers were studied in an open-chest canine heart preparation during a constant tracer infusion of up to 9 min. Distribution volumes were calculated based on the mean transit time method of Zierler. The distribution volume of ascorbic acid as well as of L-({sup 14}C)ascorbic acid in myocardial tissue agreed closely with those of ({sup 3}H)inulin and ({sup 3}H)sucrose as well as {sup 82}Br. The obtained kinetic data confirmed that ascorbic acid exhibits the physicochemical properties of an extracellular space marker, though this compound was shown to leak slowly into myocardial cells. Favorable attributes of this indicator are its low molecular weight, high diffusibility in interstitial fluid, low binding affinity to macromolecules, and high transcapillary as well as low transplasmalemmal penetration rate. Therefore, this nonradioactive marker can be applied in a safe and simple fashion, and without untoward side effects in experimental animals as well as in patients.

  13. A fluorescence based non-radioactive electrophoretic mobility shift assay.

    PubMed

    Ruscher, K; Reuter, M; Kupper, D; Trendelenburg, G; Dirnagl, U; Meisel, A

    2000-03-10

    Electrophoretic mobility shift assay (EMSA) or gel shift assay is one of the most powerful methods for studying protein-DNA interactions. Typically, 32P-labeled DNA probes containing the sequence bound by the protein of interest are used in EMSA (rEMSA). Although rEMSA is sensitive and practicable, it relies on the handling of hazardous radioisotopes, and does not easily allow quantification. We developed a non-radioactive procedure using fluorescence (Cyano dye Cy5) labeled oligodeoxynucleotide duplexes as specific probes (fEMSA) and an automatic DNA sequencer for analysis. Testing different DNA-binding proteins (restriction endonuclease EcoRII, transcription factor NFkappaB and it's subunit p50) the results in fEMSA and rEMSA are similar in regard to quality, reproducibility, and sensitivity. fEMSA allows a semiquantitative screening of large amounts of samples for specific DNA binding activities and is, therefore, a high throughput technology for semiquantitative analysis of DNA-protein interaction.

  14. Simultaneous Site-Specific Dual Protein Labeling Using Protein Prenyltransferases.

    PubMed

    Zhang, Yi; Blanden, Melanie J; Sudheer, Ch; Gangopadhyay, Soumyashree A; Rashidian, Mohammad; Hougland, James L; Distefano, Mark D

    2015-12-16

    Site-specific protein labeling is an important technique in protein chemistry and is used for diverse applications ranging from creating protein conjugates to protein immobilization. Enzymatic reactions, including protein prenylation, have been widely exploited as methods to accomplish site-specific labeling. Enzymatic prenylation is catalyzed by prenyltransferases, including protein farnesyltransferase (PFTase) and geranylgeranyltransferase type I (GGTase-I), both of which recognize C-terminal CaaX motifs with different specificities and transfer prenyl groups from isoprenoid diphosphates to their respective target proteins. A number of isoprenoid analogues containing bioorthogonal functional groups have been used to label proteins of interest via PFTase-catalyzed reaction. In this study, we sought to expand the scope of prenyltransferase-mediated protein labeling by exploring the utility of rat GGTase-I (rGGTase-I). First, the isoprenoid specificity of rGGTase-I was evaluated by screening eight different analogues and it was found that those with bulky moieties and longer backbone length were recognized by rGGTase-I more efficiently. Taking advantage of the different substrate specificities of rat PFTase (rPFTase) and rGGTase-I, we then developed a simultaneous dual labeling method to selectively label two different proteins by using isoprenoid analogue and CaaX substrate pairs that were specific to only one of the prenyltransferases. Using two model proteins, green fluorescent protein with a C-terminal CVLL sequence (GFP-CVLL) and red fluorescent protein with a C-terminal CVIA sequence (RFP-CVIA), we demonstrated that when incubated together with both prenyltransferases and the selected isoprenoid analogues, GFP-CVLL was specifically modified with a ketone-functionalized analogue by rGGTase-I and RFP-CVIA was selectively labeled with an alkyne-containing analogue by rPFTase. By switching the ketone-containing analogue to an azide-containing analogue, it was

  15. Site-specific DOTA/europium-labeling of recombinant human relaxin-3 for receptor-ligand interaction studies.

    PubMed

    Zhang, Wei-Jie; Luo, Xiao; Liu, Ya-Li; Shao, Xiao-Xia; Wade, John D; Bathgate, Ross A D; Guo, Zhan-Yun

    2012-08-01

    Relaxin-3 (also known as INSL7) is a recently identified neuropeptide belonging to the insulin/relaxin superfamily. It has putative roles in the regulation of stress responses, food intake, and reproduction by activation of its cognate G-protein-coupled receptor RXFP3. It also binds and activates the relaxin family peptide receptors RXFP1 and RXFP4 in vitro. To obtain a europium-labeled relaxin-3 as tracer for studying the interaction of these receptors with various ligands, in the present work we propose a novel site-specific labeling strategy for the recombinant human relaxin-3 that has been previously prepared in our laboratory. First, the N-terminal 6 × His-tag of the single-chain relaxin-3 precursor was removed by Aeromonas aminopeptidase and all of the primary amines of the resultant peptide were reversibly blocked by citroconic anhydride. Second, the A-chain N-terminus of the blocked peptide was released by endoproteinase Asp-N cleavage that removed the linker peptide between the B- and A-chains. Third, an alkyne moiety was introduced to the newly released A-chain N-terminus by reaction with the highly active primary amine-specific N-hydroxysuccinimide ester. Fourth, after removal of the reversible blockage under mild acidic condition, europium-loaded DOTA with an azide moiety was introduced to the two-chain relaxin-3 carrying the alkyne moiety through click chemistry. Using this site-specific labeling strategy, homogeneous monoeuropium-labeled human relaxin-3 could be obtained with good overall yield. In contrast, conventional random labeling resulted in a complex mixture that was poorly resolved because human relaxin-3 has four primary amine moieties that all react with the modification reagent. Both saturation and competition binding assays demonstrated that the DOTA/Eu(3+)-labeled relaxin-3 retained high binding affinity for human RXFP3, RXFP4, and RXFP1 and was therefore a suitable non-radioactive and stable tracer to study the interaction of various

  16. Site-Specific Immunomodulator: A Novel Treatment for Crohn's Disease

    PubMed Central

    Bressler, Brian; Bethel, Kevin P.; Kleef, Ralf; Reynolds, Sophie L.; Sutcliffe, Simon; Mullins, David W.; Gunn, Hal

    2015-01-01

    We investigated the mechanism of action, safety, and efficacy of the Site-Specific Immunomodulator (SSI) QBECO, a novel immunotherapy for Crohn's disease (CD). Using human monocytic THP-1 cells, we demonstrate that SSI QBECO (derived from the common colon bacteria E. coli) activates macrophages to an M1 phenotype (associated with enhanced capacity to eliminate bacteria and activate innate immune responses). We assessed SSI QBECO in a compassionate use protocol of ten adult patients with active CD. Patients with moderate to severe clinical symptoms receiving conventional CD treatments and/or complementary therapies were included, except patients receiving anti-TNF medications. SSI QBECO was self-administered subcutaneously every second day, for a minimum of 2.5 months and a maximum of 11 months. All 10 patients reported improvement of symptoms while on the SSI QBECO treatment. Seven patients reported full resolution of clinical symptoms during a course of SSI QBECO of at least three months. Three patients have experienced ongoing sustained clinical remission after discontinuing all medications, including SSI treatment. The longest case of clinical remission is still ongoing (>4 years). No serious severe adverse clinical events were reported. Collectively, we conclude that treatment with the immunoactive SSI QBECO was well tolerated and effective for treatment of Crohn's disease in this case series. PMID:26064087

  17. Micro-tattoo guided OCT imaging of site specific inflammation

    NASA Astrophysics Data System (ADS)

    Phillips, Kevin G.; Choudhury, Niloy; Samatham, Ravikant V.; Singh, Harvinder; Jacques, Steven L.

    2010-02-01

    Epithelial biologists studying human skin diseases such as cancer formation and psoriasis commonly utilize mouse models to characterize the interplay among cells and intracellular signal transduction pathways that result in programmed changes in gene expression and cellular behaviors. The information obtained from animal models is useful only when phenotypic presentations of disease recapitulate those observed in humans. Excision of tissues followed by histochemical analysis is currently the primary means of establishing the morphological presentation. Non invasive imaging of animal models provides an alternate means to characterize tissue morphology associated with the disease of interest in vivo. While useful, the ability to perform in vivo imaging at different time points in the same tissue location has been a challenge. This information is key to understanding site specific changes as the imaged tissue can now be extracted and analyzed for mRNA expression. We present a method employing a micro-tattoo to guide optical coherence tomography (OCT) imaging of ultraviolet induced inflammation over time in the same tissue locations.

  18. Genetic fate mapping using site-specific recombinases.

    PubMed

    Legué, Emilie; Joyner, Alexandra L

    2010-01-01

    Understanding how cells are assembled in three dimensions to generate an organ, or a whole organism, is a pivotal question in developmental biology. Similarly, it is critical to understand how adult stem cells integrate into an existing organ during regeneration or in response to injury. Key to discovering the answers to these questions is being able to study the various behaviors of distinct cell types during development or regeneration. Fate mapping techniques are fundamental to studying cell behaviors such as proliferation, movement, and lineage segregation, as the techniques allow precursor cells to be marked and their descendants followed and characterized over time. The generation of transgenic mice, combined with the use of site-specific recombinases (SSR) in the mouse genome, has provided a means to develop powerful genetic fate mapping approaches. A key advantage of genetic fate mapping is that it allows cells to be genetically marked, and therefore the mark is transmitted to all the descendants of the initially marked cells. By making modifications to the SSRs that render their enzymatic activity inducible, and the development of an assortment of reporter alleles for marking cells, increasingly sophisticated genetic fate mapping studies can be performed. In this chapter, we review the four main genetic fate mapping methods that utilize intrachromosomal recombination to mark cells (cumulative, inducible, clonal, and intersectional) and one interchromosomal method, the tools required to carry out each approach, and the practical considerations that have to be taken into account before embarking on each type of genetic fate mapping study.

  19. An exactly solvable model of random site-specific recombinations

    PubMed Central

    Wei, Yi; Koulakov, Alexei A.

    2017-01-01

    Cre-lox and other systems are used as genetic tools to control site-specific recombination (SSR) events in genomic DNA. If multiple recombination sites are organized in a compact cluster within the same genome, a series of random recombination events may generate substantial cell specific genomic diversity. This diversity is used, for example, to distinguish neurons in the brain of the same multicellular mosaic organism, within the brainbow approach to neuronal connectome. In this paper we study an exactly solvable statistical model for SSR operating on a cluster of recombination sites. We consider two types of recombination events: inversions and excisions. Both of these events are available in the Cre-lox system. We derive three properties of the sequences generated by multiple recombination events. First, we describe the set of sequences that can in principle be generated by multiple inversions operating on the given initial sequence. We call this description the ergodicity theorem. On the basis of this description we calculate the number of sequences that can be generated from an initial sequence. This number of sequences is experimentally testable. Second, we demonstrate that after a large number of random inversions every sequence that can be generated is generated with equal probability. Lastly, we derive the equations for the probability to find a sequence as a function of time in the limit when excisions are much less frequent than inversions, such as in shufflon sequences. PMID:23151958

  20. Site-specific anticancer effects of dietary flavonoid quercetin.

    PubMed

    Sak, Katrin

    2014-01-01

    Food-derived flavonoid quercetin, widely distributed in onions, apples, and tea, is able to inhibit growth of various cancer cells indicating that this compound can be considered as a good candidate for anticancer therapy. Although the exact mechanism of this action is not thoroughly understood, behaving as antioxidant and/or prooxidant as well as modulating different intracellular signalling cascades may all play a certain role. Such inhibitory activity of quercetin has been shown to depend first of all on cell lines and cancer types; however, no comprehensive site-specific analysis of this effect has been published. In this review article, cytotoxicity constants of quercetin measured in various human malignant cell lines of different origin were compiled from literature and a clear cancer selective action was demonstrated. The most sensitive malignant sites for quercetin revealed to be cancers of blood, brain, lung, uterine, and salivary gland as well as melanoma whereas cytotoxic activity was higher in more aggressive cells compared to the slowly growing cells showing that the most harmful cells for the organism are probably targeted. More research is needed to overcome the issues of poor water solubility and relatively low bioavailability of quercetin as the major obstacles limiting its clinical use.

  1. Site-specific protein glycosylation analysis with glycan isomer differentiation.

    PubMed

    Hua, Serenus; Nwosu, Charles C; Strum, John S; Seipert, Richard R; An, Hyun Joo; Zivkovic, Angela M; German, J Bruce; Lebrilla, Carlito B

    2012-05-01

    Glycosylation is one of the most common yet diverse post-translational modifications. Information on glycan heterogeneity and glycosite occupancy is increasingly recognized as crucial to understanding glycoprotein structure and function. Yet, no approach currently exists with which to holistically consider both the proteomic and glycomic aspects of a system. Here, we developed a novel method of comprehensive glycosite profiling using nanoflow liquid chromatography/mass spectrometry (nano-LC/MS) that shows glycan isomer-specific differentiation on specific sites. Glycoproteins were digested by controlled non-specific proteolysis in order to produce informative glycopeptides. High-resolution, isomer-sensitive chromatographic separation of the glycopeptides was achieved using microfluidic chip-based capillaries packed with graphitized carbon. Integrated LC/MS/MS not only confirmed glycopeptide composition but also differentiated glycan and peptide isomers and yielded structural information on both the glycan and peptide moieties. Our analysis identified at least 13 distinct glycans (including isomers) corresponding to five compositions at the single N-glycosylation site on bovine ribonuclease B, 59 distinct glycans at five N-glycosylation sites on bovine lactoferrin, 13 distinct glycans at one N-glycosylation site on four subclasses of human immunoglobulin G, and 20 distinct glycans at five O-glycosylation sites on bovine κ-casein. Porous graphitized carbon provided effective separation of glycopeptide isomers. The integration of nano-LC with MS and MS/MS of non-specifically cleaved glycopeptides allows quantitative, isomer-sensitive, and site-specific glycoprotein analysis.

  2. Analysis of Chemokine Receptor Trafficking by Site-Specific Biotinylation

    PubMed Central

    Liebick, Marcel; Schläger, Christian; Oppermann, Martin

    2016-01-01

    Chemokine receptors undergo internalization and desensitization in response to ligand activation. Internalized receptors are either preferentially directed towards recycling pathways (e.g. CCR5) or sorted for proteasomal degradation (e.g. CXCR4). Here we describe a method for the analysis of receptor internalization and recycling based on specific Bir A-mediated biotinylation of an acceptor peptide coupled to the receptor, which allows a more detailed analysis of receptor trafficking compared to classical antibody-based detection methods. Studies on constitutive internalization of the chemokine receptors CXCR4 (12.1% ± 0.99% receptor internalization/h) and CCR5 (13.7% ± 0.68%/h) reveals modulation of these processes by inverse (TAK779; 10.9% ± 0.95%/h) or partial agonists (Met-CCL5; 15.6% ± 0.5%/h). These results suggest an actively driven internalization process. We also demonstrate the advantages of specific biotinylation compared to classical antibody detection during agonist-induced receptor internalization, which may be used for immunofluorescence analysis as well. Site-specific biotinylation may be applicable to studies on trafficking of transmembrane proteins, in general. PMID:27310579

  3. DNA origami metallized site specifically to form electrically conductive nanowires.

    PubMed

    Pearson, Anthony C; Liu, Jianfei; Pound, Elisabeth; Uprety, Bibek; Woolley, Adam T; Davis, Robert C; Harb, John N

    2012-09-06

    DNA origami is a promising tool for use as a template in the design and fabrication of nanoscale structures. The ability to engineer selected staple strands on a DNA origami structure provides a high density of addressable locations across the structure. Here we report a method using site-specific attachment of gold nanoparticles to modified staple strands and subsequent metallization to fabricate conductive wires from DNA origami templates. We have modified DNA origami structures by lengthening each staple strand in select regions with a 10-base nucleotide sequence and have attached DNA-modified gold nanoparticles to the lengthened staple strands via complementary base-pairing. The high density of extended staple strands allowed the gold nanoparticles to pack tightly in the modified regions of the DNA origami, where the measured median gap size between neighboring particles was 4.1 nm. Gold metallization processes were optimized so that the attached gold nanoparticles grew until gaps between particles were filled and uniform continuous nanowires were formed. Finally, electron beam lithography was used to pattern electrodes in order to measure the electrical conductivity of metallized DNA origami, which showed an average resistance of 2.4 kΩ per metallized structure.

  4. Site-specific group selection drives locally adapted group compositions.

    PubMed

    Pruitt, Jonathan N; Goodnight, Charles J

    2014-10-16

    Group selection may be defined as selection caused by the differential extinction or proliferation of groups. The socially polymorphic spider Anelosimus studiosus exhibits a behavioural polymorphism in which females exhibit either a 'docile' or 'aggressive' behavioural phenotype. Natural colonies are composed of a mixture of related docile and aggressive individuals, and populations differ in colonies' characteristic docile:aggressive ratios. Using experimentally constructed colonies of known composition, here we demonstrate that population-level divergence in docile:aggressive ratios is driven by site-specific selection at the group level--certain ratios yield high survivorship at some sites but not others. Our data also indicate that colonies responded to the risk of extinction: perturbed colonies tended to adjust their composition over two generations to match the ratio characteristic of their native site, thus promoting their long-term survival in their natal habitat. However, colonies of displaced individuals continued to shift their compositions towards mixtures that would have promoted their survival had they remained at their home sites, regardless of their contemporary environment. Thus, the regulatory mechanisms that colonies use to adjust their composition appear to be locally adapted. Our data provide experimental evidence of group selection driving collective traits in wild populations.

  5. Bone site-specific delivery of siRNA

    PubMed Central

    Liu, Xinli

    2016-01-01

    Abstract Small interfering RNAs (siRNA) have enormous potential as therapeutics to target and treat various bone disorders such as osteoporosis and cancer bone metastases. However, effective and specific delivery of siRNA therapeutics to bone and bone-specific cells in vivo is very challenging. To realize the full therapeutic potential of siRNA in treating bone disorders, a safe and efficient, tissue- and cell-specific delivery system must be developed. This review focuses on recent advances in bone site-specific delivery of siRNA at the tissue or cellular level. Bone-targeted nanoparticulate siRNA carriers and various bone-targeted moieties such as bisphosphonates, oligopeptides (Asp)8 and (AspSerSer)6, and aptamers are highlighted. Incorporation of these bone-seeking targeting moieties into siRNA carriers allows for recognition of different sub-tissue functional domains of bone and also specific cell types residing in bone tissue. It also provides a means for bone-formation surface-, bone-resorption surface-, or osteoblast-specific targeting and transportation of siRNA therapeutics. The discussion mainly focuses on systemic and local bone-specific delivery of siRNA in osteoporosis and bone metastasis preclinical models. PMID:26642236

  6. Cre-inducible site-specific recombination in zebrafish oligodendrocytes.

    PubMed

    Pinzon-Olejua, Alejandro; Welte, Cornelia; Chekuru, Avinash; Bosak, Viktoria; Brand, Michael; Hans, Stefan; Stuermer, Claudia A O

    2017-01-01

    The conditional Cre/lox system has recently emerged as a valuable tool for studies on both embryonic and adult Zebrafish. Temporal control and site-specific recombination are achieved by using the ligand-inducible CreER(T2) and administration of the drug tamoxifen (TAM) or its active metabolite, 4-Hydroxytamoxifen (4-OHT). Here we report the generation of a transgenic Zebrafish line, which expresses an mCherry-tagged variant of CreER(T2) under the control of the myelin basic protein a (mbpa) promoter. Our analysis shows that larval and adult expression of the transgene recapitulates the endogenous mbpa expression pattern in oligodendrocytes. Furthermore, combination with a Cre-dependent EGFP reporter results in EGFP-expressing oligodendrocytes in the spinal cord, brain, and optic nerve in TAM- or 4-OHT-treated larvae and 4-month-old fish, but not in untreated controls. The transgenic Zebrafish line Tg(mbpa:mCherry-T2A-CreER(T2) ) elicits CreER(T2) expression specifically in myelinating glia cells. Cre-inducible targeted recombination of genes in oligodendrocytes will be useful to elucidate cellular and molecular mechanisms of myelination in vivo during development (myelination) and regeneration (remyelination) after injury to the central nervous system (CNS). It will also allow targeted expression and overexpression of genes of interest (transgenes) in oligodendrocytes at defined developmental and adult stages. Developmental Dynamics 246:41-49, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  7. Site-specific PEGylation of lidamycin and its antitumor activity

    PubMed Central

    Li, Liang; Shang, Boyang; Hu, Lei; Shao, Rongguang; Zhen, Yongsu

    2015-01-01

    In this study, N-terminal site-specific mono-PEGylation of the recombinant lidamycin apoprotein (rLDP) of lidamycin (LDM) was prepared using a polyethyleneglycol (PEG) derivative (Mw 20 kDa) through a reactive terminal aldehyde group under weak acidic conditions (pH 5.5). The biochemical properties of mPEG-rLDP-AE, an enediyne-integrated conjugate, were analyzed by SDS-PAGE, RP-HPLC, SEC-HPLC and MALDI-TOF. Meanwhile, in vitro and in vivo antitumor activity of mPEG-rLDP-AE was evaluated by MTT assays and in xenograft model. The results indicated that mPEG-rLDP-AE showed significant antitumor activity both in vitro and in vivo. After PEGylation, mPEG-rLDP still retained the binding capability to the enediyne AE and presented the physicochemical characteristics similar to that of native LDP. It is of interest that the PEGylation did not diminish the antitumor efficacy of LDM, implying the possibility that this derivative may function as a payload to deliver novel tumor-targeted drugs. PMID:26579455

  8. Hanford facility dangerous waste permit application, 616 Nonradioactive Dangerous Waste Storage Facility. Revision 2A

    SciTech Connect

    Bowman, R.C.

    1994-04-01

    This permit application for the 616 Nonradioactive Dangerous Waste Storage Facility consists for 15 chapters. Topics of discussion include the following: facility description and general provisions; waste characteristics; process information; personnel training; reporting and record keeping; and certification.

  9. ASSESSMENT OF RADIOACTIVE AND NON-RADIOACTIVE CONTAMINANTS FOUND IN LOW LEVEL RADIOACTIVE WASTE STREAMS

    SciTech Connect

    R.H. Little, P.R. Maul, J.S.S. Penfoldag

    2003-02-27

    This paper describes and presents the findings from two studies undertaken for the European Commission to assess the long-term impact upon the environment and human health of non-radioactive contaminants found in various low level radioactive waste streams. The initial study investigated the application of safety assessment approaches developed for radioactive contaminants to the assessment of nonradioactive contaminants in low level radioactive waste. It demonstrated how disposal limits could be derived for a range of non-radioactive contaminants and generic disposal facilities. The follow-up study used the same approach but undertook more detailed, disposal system specific calculations, assessing the impacts of both the non-radioactive and radioactive contaminants. The calculations undertaken indicated that it is prudent to consider non-radioactive, as well as radioactive contaminants, when assessing the impacts of low level radioactive waste disposal. For some waste streams with relatively low concentrations of radionuclides, the potential post-closure disposal impacts from non-radioactive contaminants can be comparable with the potential radiological impacts. For such waste streams there is therefore an added incentive to explore options for recycling the materials involved wherever possible.

  10. Ascorbic acid: a nonradioactive extracellular space marker in canine heart.

    PubMed

    Reil, G H; Frombach, R; Kownatzki, R; Quante, W; Lichtlen, P R

    1987-11-01

    The distribution pattern of ascorbic acid and L-[14C]ascorbic acid in myocardial tissue was compared with those of the classical radioactive extracellular space markers [3H]-inulin, [3H]sucrose, and Na82Br. A new polarographic technique was developed for analogue registration of ascorbic acid concentration in coronary venous blood. The kinetic data of the markers were studied in an open-chest canine heart preparation (n = 17 dogs) during a constant tracer infusion of up to 9 min. Distribution volumes were calculated based on the mean transit time method of Zierler (K. L. Zierler, Circ. Res. 10:393-407, 1962). The distribution volume of ascorbic acid (23.6 ml/100 g wet wt) as well as of L-[14C]ascorbic acid (24.5 ml/100 g wet wt) in myocardial tissue agreed closely with those of [3H]inulin (18.6 ml/100 g wet wt) and [3H]sucrose (22.0 ml/100 g wet wt), as well as 82Br- (27.3 ml/100 g wet wt). The obtained kinetic data confirmed that ascorbic acid exhibits the physicochemical properties of an extracellular space marker, though this compound was shown to leak slowly into myocardial cells. Favorable attributes of this indicator are its low molecular weight, high diffusibility in interstitial fluid, low binding affinity to macromolecules, and high transcapillary (1.8 X 10(-5) cm/s) as well as low transplasmalemmal penetration rate (3.7 X 10(-8) cm/s). Therefore, this nonradioactive marker can be applied in a safe and simple fashion, and without untoward side effects in experimental animals as well as in patients.

  11. Site-specific hydrogen diffusion rates during clinopyroxene dehydration

    NASA Astrophysics Data System (ADS)

    Ferriss, Elizabeth; Plank, Terry; Walker, David

    2016-06-01

    The rate of hydrogen diffusion in clinopyroxene is relevant to interpreting hydrogen ("water") concentrations in xenoliths, phenocrysts, and clinopyroxene-hosted melt inclusions to provide insight into the deep-earth water cycle and volcanic explosivity. Here, we determine bulk and site-specific hydrogen diffusivities in two diopsides and an augite by heating initially homogeneous water-bearing samples in a 1-atm CO/CO2 gas-mixing furnace at 800-1000 °C and oxygen fugacity at the quartz-fayalite-magnetite buffer and observing H-loss profiles. The O-H stretching range between wavenumbers 3000 and 4000 cm-1 in FTIR spectra is resolved into 4-6 peaks, each of which is assumed to represent a distinct defect site for the hydrogen, to determine peak-specific diffusivities using our previously published whole-block method. For the diopside from the Kunlun Mts. in China, Arrhenius relations are reported for peaks at 3645, 3617, 3540, 3443, and 3355 cm-1 based on measurements at 816, 904, and 1000 °C. Bulk and site-specific diffusivities are determined for the same set of peaks at 904 °C for the second diopside (Jaipur). The augite (PMR-53) was a triangular thin slab, and hydrogen diffusivities were determined for bulk hydrogen and peaks at 3620, 3550, 3460, and 3355 cm-1 in the thickness direction at 800 °C. Bulk hydrogen diffusivity in the Jaipur diopside is consistent with previous work, and hydrogen diffusivity in augite PMR-53 is slightly lower than the fast direction diffusivities measured || [100] and [001]* in Jaipur diopside. Both diopsides show 1-2 orders of magnitude differences in the peaks-specific diffusivities, with the fastest diffusivities at 3450 cm-1 and the slowest at 3645 cm-1. However, the hydrogen diffusivities in Jaipur diopside are 2-4 orders of magnitude higher than those in Kunlun diopside for bulk hydrogen and all peaks. Thus, peak-specific differences cannot by themselves adequately explain the 5 orders of magnitude range in hydrogen

  12. Xer Site Specific Recombination: Double and Single Recombinase Systems

    PubMed Central

    Castillo, Fabio; Benmohamed, Amal; Szatmari, George

    2017-01-01

    The separation and segregation of newly replicated bacterial chromosomes can be constrained by the formation of circular chromosome dimers caused by crossing over during homologous recombination events. In Escherichia coli and most bacteria, dimers are resolved to monomers by site-specific recombination, a process performed by two Chromosomally Encoded tyrosine Recombinases (XerC and XerD). XerCD recombinases act at a 28 bp recombination site dif, which is located at the replication terminus region of the chromosome. The septal protein FtsK controls the initiation of the dimer resolution reaction, so that recombination occurs at the right time (immediately prior to cell division) and at the right place (cell division septum). XerCD and FtsK have been detected in nearly all sequenced eubacterial genomes including Proteobacteria, Archaea, and Firmicutes. However, in Streptococci and Lactococci, an alternative system has been found, composed of a single recombinase (XerS) genetically linked to an atypical 31 bp recombination site (difSL). A similar recombination system has also been found in 𝜀-proteobacteria such as Campylobacter and Helicobacter, where a single recombinase (XerH) acts at a resolution site called difH. Most Archaea contain a recombinase called XerA that acts on a highly conserved 28 bp sequence dif, which appears to act independently of FtsK. Additionally, several mobile elements have been found to exploit the dif/Xer system to integrate their genomes into the host chromosome in Vibrio cholerae, Neisseria gonorrhoeae, and Enterobacter cloacae. This review highlights the versatility of dif/Xer recombinase systems in prokaryotes and summarizes our current understanding of homologs of dif/Xer machineries. PMID:28373867

  13. Oxygen as a site specific structural probe in neutron diffraction

    SciTech Connect

    Neuefeind, Joerg C; Simonson, J Michael {Mike}; Salmon, Phil; Zeidler, Anita; Fischer, Henry E; Rauch, Helmut; Markland, Thomas; Lemmel, Hartmut

    2011-01-01

    Oxygen is a ubiquitous element, playing an essential role in most scientific and technological disciplines, and is often incorporated within a structurally disordered material where examples include molten silicates in planetary science, glasses used for lasers and optical communication, and water in biological processes. Establishing the structure of a liquid or glassy oxide and thereby its relation to the functional properties of a material is not, however, a trivial task owing to the complexity associated with atomic disorder. Here we approach this challenge by measuring the bound coherent neutron scattering lengths of the oxygen isotopes with the sensitive technique of neutron interferometry. We find that there is a small but finite contrast of 0.204(6) fm between the scattering lengths of the isotope 18O and oxygen of natural isotopic abundance natO, contrary to tables of recommended values. This has enabled us to investigate the structure of both light and heavy water by exploiting, for the first time, the method of oxygen isotope substitution in neutron diffraction, thus circumventing many of the significant problems associated with more traditional methods in which hydrogen is substituted by deuterium. We find a difference of ~0.5% between the O-H and O-D intra-molecular bond distances which is much smaller than recent estimates based on diffraction data and is found to be in excellent agreement with path integral molecular dynamics simulations made with a flexible polarisable water model. Our results demonstrate the potential for using oxygen isotope substitution as a powerful and effective site specific probe in a plethora of materials, of pertinence as instrumentation at next generation neutron sources comes online

  14. Improving Site-Specific Radiological Performance Assessments - 13431

    SciTech Connect

    Tauxe, John; Black, Paul; Catlett, Kate; Lee, Robert; Perona, Ralph; Stockton, Tom; Sully, Mike

    2013-07-01

    An improved approach is presented for conducting complete and defensible radiological site-specific performance assessments (PAs) to support radioactive waste disposal decisions. The basic tenets of PA were initiated some thirty years ago, focusing on geologic disposals and evaluating compliance with regulations. Some of these regulations were inherently probabilistic (i.e., addressing uncertainty in a quantitative fashion), such as the containment requirements of the U.S. Environmental Protection Agency's (EPA's) 40 CFR 191, Environmental Radiation Protection Standards for Management and Disposal of Spent Nuclear Fuel, High-Level and Transuranic Radioactive Wastes, Chap. 191.13 [1]. Methods of analysis were developed to meet those requirements, but at their core early PAs used 'conservative' parameter values and modeling approaches. This limited the utility of such PAs to compliance evaluation, and did little to inform decisions about optimizing disposal, closure and long-term monitoring and maintenance, or, in general, maintaining doses 'as low as reasonably achievable' (ALARA). This basic approach to PA development in the United States was employed essentially unchanged through the end of the 20. century, principally by the U.S. Department of Energy (DOE). Performance assessments developed in support of private radioactive waste disposal operations, regulated by the U.S. Nuclear Regulatory Commission (NRC) and its agreement states, were typically not as sophisticated. Discussion of new approaches to PA is timely, since at the time of this writing, the DOE is in the midst of revising its Order 435.1, Radioactive Waste Management [2], and the NRC is revising 10 CFR 61, Licensing Requirements for Land Disposal of Radioactive Waste [3]. Over the previous decade, theoretical developments and improved computational technology have provided the foundation for integrating decision analysis (DA) concepts and objective-focused thinking, plus a Bayesian approach to

  15. Site-Specific Seismic Site Response Model for the Waste Treatment Plant, Hanford, Washington

    SciTech Connect

    Rohay, Alan C.; Reidel, Steve P.

    2005-02-24

    This interim report documents the collection of site-specific geologic and geophysical data characterizing the Waste Treatment Plant site and the modeling of the site-specific structure response to earthquake ground motions.

  16. Effect of site-specific modification on restriction endonucleases and DNA modification methyltransferases.

    PubMed Central

    McClelland, M; Nelson, M; Raschke, E

    1994-01-01

    Restriction endonucleases have site-specific interactions with DNA that can often be inhibited by site-specific DNA methylation and other site-specific DNA modifications. However, such inhibition cannot generally be predicted. The empirically acquired data on these effects are tabulated for over 320 restriction endonucleases. In addition, a table of known site-specific DNA modification methyltransferases and their specificities is presented along with EMBL database accession numbers for cloned genes. PMID:7937074

  17. Site-specific retention of colloids at rough rock surfaces.

    PubMed

    Darbha, Gopala Krishna; Fischer, Cornelius; Luetzenkirchen, Johannes; Schäfer, Thorsten

    2012-09-04

    The spatial deposition of polystyrene latex colloids (d = 1 μm) at rough mineral and rock surfaces was investigated quantitatively as a function of Eu(III) concentration. Granodiorite samples from Grimsel test site (GTS), Switzerland, were used as collector surfaces for sorption experiments. At a scan area of 300 × 300 μm(2), the surface roughness (rms roughness, Rq) range was 100-2000 nm, including roughness contribution from asperities of several tens of nanometers in height to the sample topography. Although, an increase in both roughness and [Eu(III)] resulted in enhanced colloid deposition on granodiorite surfaces, surface roughness governs colloid deposition mainly at low Eu(III) concentrations (≤5 × 10(-7) M). Highest deposition efficiency on granodiorite has been found at walls of intergranular pores at surface sections with roughness Rq = 500-2000 nm. An about 2 orders of magnitude lower colloid deposition has been observed at granodiorite sections with low surface roughness (Rq < 500 nm), such as large and smooth feldspar or quartz crystal surface sections as well as intragranular pores. The site-specific deposition of colloids at intergranular pores is induced by small scale protrusions (mean height = 0.5 ± 0.3 μm). These protrusions diminish locally the overall DLVO interaction energy at the interface. The protrusions prevent further rolling over the surface by increasing the hydrodynamic drag required for detachment. Moreover, colloid sorption is favored at surface sections with high density of small protrusions (density (D) = 2.6 ± 0.55 μm(-1), asperity diameter (φ) = 0.6 ± 0.2 μm, height (h) = 0.4 ± 0.1 μm) in contrast to surface sections with larger asperities and lower asperity density (D = 1.2 ± 0.6 μm(-1), φ = 1.4 ± 0.4 μm, h = 0.6 ± 0.2 μm). The study elucidates the importance to include surface roughness parameters into predictive colloid-borne contaminant migration calculations.

  18. Feasibility Study of Non-Radioactive Tracers for Monitoring Injected Water in Oil Reservoirs

    NASA Astrophysics Data System (ADS)

    Sawamura, H.; Nishimura, K.; Mituishi, H.; Muta, T.; Schweitzer, J. S.

    This paper discusses the results of analyses conducted on non-radioactive tracers that can be used in combination with the sea water injected into a well for monitoring the water permeating through the oil reservoir by a nuclear logging tool utilizing a pulsed neutron generator. The model of the pulsed neutron tool is constructed to permit Monte Carlo Simulations to be performed of the tool response to the presence of non-radioactive tracers to achieve a desirable level of the neutron absorbing cross sections in the sea water injected into and permeating through the oil reservoirs. Sensitivity analyses of the tool response of the nuclear logging tool were performed for two types of non-radioactive tracers, ammonium tetraborate and gadolinium chloride.

  19. STATIC AND KINETIC SITE-SPECIFIC PROTEIN-DNA PHOTOCROSSLINKING: ANALYSIS OF BACTERIAL TRANSCRIPTION INITIATION COMPLEXES

    PubMed Central

    Naryshkin, Nikolai; Druzhinin, Sergei; Revyakin, Andrei; Kim, Younggyu; Mekler, Vladimir; Ebright, Richard H.

    2009-01-01

    Static site-specific protein-DNA photocrosslinking permits identification of protein-DNA interactions within multiprotein-DNA complexes. Kinetic site-specific protein-DNA photocrosslinking--involving rapid-quench-flow mixing and pulsed-laser irradiation--permits elucidation of pathways and kinetics of formation of protein-DNA interactions within multiprotein-DNA complexes. We present detailed protocols for application of static and kinetic site-specific protein-DNA photocrosslinking to bacterial transcription initiation complexes. PMID:19378179

  20. Bathophenanthroline-Ru(II) complexes as nonradioactive labels for dideoxy DNA sequencing.

    PubMed

    Bannwarth, W

    1989-09-01

    Bathophenanthroline-Ru(II) complexes represent interesting nonradioactive label molecules which can be measured in a time-resolved mode with high sensitivity. Here we demonstrate that Ru complex-labeled primers can be applied in dideoxy sequencing protocols giving the same sequencing patterns as unlabeled primers.

  1. 78 FR 12747 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-25

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... Participation: The EM SSAB, Idaho National Laboratory, welcomes the attendance of the public at its...

  2. 77 FR 53192 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

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    2012-08-31

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... Closeout Process Public Participation: The EM SSAB, Idaho National Laboratory, welcomes the attendance...

  3. 76 FR 25682 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

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    2011-05-05

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National ] Laboratory. The Federal Advisory Committee Act... National Laboratory, welcomes the attendance of the public at its advisory committee meetings and will...

  4. 76 FR 10018 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

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    2011-02-23

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... Public Participation: The EM SSAB, Idaho National Laboratory, welcomes the attendance of the public...

  5. 75 FR 39008 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

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    2010-07-07

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... Laboratory, welcomes the attendance of the public at its advisory committee meetings and will make...

  6. 76 FR 66917 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

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    2011-10-28

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... Environmental Assessment Public Participation: The EM SSAB, Idaho National Laboratory, welcomes the...

  7. 75 FR 7576 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

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    2010-02-22

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, March 10, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  8. 77 FR 38275 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-06-27

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy, DoE... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER INFORMATION CONTACT: Melyssa P. Noe, Federal...

  9. 76 FR 1415 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-01-10

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... January 12, 2011, ] meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Oak... Coordinator, Department of Energy Oak Ridge Operations Office, P.O. Box 2001, EM-90, Oak Ridge, TN 37831...

  10. 78 FR 63171 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation; Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-10-23

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation; Meeting AGENCY: Department of... Management Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  11. 76 FR 65190 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-10-20

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, November 9, 2011, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  12. 76 FR 4644 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-01-26

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, February 9, 2011, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  13. 77 FR 64494 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-22

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  14. 77 FR 29996 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-05-21

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER INFORMATION CONTACT: Melyssa P. Noe, Federal...

  15. 75 FR 24685 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-05

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, May 12, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge Turnpike...

  16. 76 FR 36101 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-06-21

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, July 13, 2011 at 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  17. 75 FR 35447 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-22

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, July 14, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge Turnpike...

  18. 75 FR 65466 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-25

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, November 10, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  19. 77 FR 18243 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-03-27

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER INFORMATION CONTACT: Melyssa P. Noe, Federal...

  20. 75 FR 51027 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-18

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, September 8, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  1. 78 FR 49738 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-15

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Center, Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR...

  2. 75 FR 13268 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-03-19

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, April 14, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  3. 78 FR 23241 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-04-18

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  4. 75 FR 71424 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-11-23

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, December 8, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  5. 76 FR 52944 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-24

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, September 14, 2011, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  6. 77 FR 49442 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-08-16

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Center, Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR...

  7. 78 FR 44942 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-25

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Coordinator, Department of Energy Oak Ridge Operations Office, P.O. Box 2001, EM-90, Oak Ridge, TN 37831...

  8. 76 FR 9572 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-18

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, March 9, 2011, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge Turnpike...

  9. 75 FR 3455 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-01-21

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, February 10, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  10. 76 FR 28759 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-18

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Thursday, May 26, 2011, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge Turnpike...

  11. 77 FR 58364 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-09-20

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  12. 76 FR 22388 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-21

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, May 11, 2011; 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge Turnpike...

  13. 75 FR 43518 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-07-26

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Department of Energy Oak Ridge Operations Office, P.O. Box 2001, EM-90, Oak Ridge, TN 37831. Phone (865) 576...

  14. 75 FR 27998 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-19

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, June 9, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge Turnpike...

  15. 76 FR 78908 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-20

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, January 11, 2012; 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  16. 78 FR 30911 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-05-23

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  17. 76 FR 29732 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-23

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, June 8, 2011, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge Turnpike...

  18. 77 FR 2714 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-01-19

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, February 8, 2012; 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  19. 78 FR 3890 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-01-17

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  20. 78 FR 17648 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-22

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  1. 77 FR 45345 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-31

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Oak Ridge Operations Office, P.O. Box 2001, EM-90, Oak Ridge, TN 37831. Phone (865) 241-3315; Fax (865...

  2. 76 FR 59393 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-26

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, October 12, 2011; 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  3. 78 FR 58292 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-23

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  4. 75 FR 57462 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-21

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, October 13, 2010, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  5. 77 FR 9219 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-16

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy, DoE... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER INFORMATION CONTACT: Melyssa P. Noe...

  6. 78 FR 12746 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-25

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  7. 78 FR 75552 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-12

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 ] Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  8. 76 FR 17637 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-30

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Register. DATES: Wednesday, April 13, 2011, 6 p.m. ADDRESSES: DOE Information Center, 475 Oak Ridge...

  9. 77 FR 74836 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-12-18

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub..., Office of Science and Technical Information, 1 Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER...

  10. 77 FR 23470 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-04-19

    ... Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Oak Ridge Reservation. The Federal Advisory Committee Act (Pub... Science.gov Way, Oak Ridge, Tennessee 37830. FOR FURTHER INFORMATION CONTACT: Melyssa P. Noe, Federal...

  11. 76 FR 19003 - Land Disposal Restrictions: Nevada and California; Site Specific Treatment Variances for...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-06

    ...: Proposed rule. SUMMARY: EPA is proposing to issue both a site-specific treatment variance to U.S. Ecology... proposes to issue both a site-specific treatment variance to U.S. Ecology Nevada (USEN) located in Beatty... section of this document. II. Does this action apply to me? This proposal applies only to U. S. Ecology...

  12. FLP recombinase-mediated site-specific recombination in silkworm, Bombyx mori

    USDA-ARS?s Scientific Manuscript database

    A comprehensive understanding of gene function and the production of site-specific genetically modified mutants are two major goals of genetic engineering in the post-genomic era. Although site-specific recombination systems have been powerful tools for genome manipulation of many organisms, they h...

  13. 77 FR 26273 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-05-03

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy, DoE... Site-Specific Advisory Board (EM SSAB), Northern New Mexico. The Federal Advisory Committee Act (Pub. L... New Mexico Citizens' Advisory Board (NNMCAB), 94 Cities of Gold Road, Santa Fe, NM 87506. Phone (505...

  14. 75 FR 2859 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-01-19

    ... Environmental Management Site-Specific Advisory Board, Hanford AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... management in the areas of environmental restoration, waste management, and related activities....

  15. 76 FR 39080 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-07-05

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... National Laboratory (INL) 101. INL EM Budget. Calcine Path Forward. Advanced Mixed Waste Treatment Project...

  16. 75 FR 56527 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-16

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act.... Overview Legacy Management--Long-Term Land Use at Idaho National Laboratory. Integrated Waste Treatment...

  17. 77 FR 65374 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-26

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... National Laboratory/ICP Public Involvement/ Communications Public Participation: The EM SSAB, Idaho...

  18. 76 FR 53888 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-30

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Site-Specific Advisory Board (EM SSAB), Idaho National Laboratory. The Federal Advisory Committee Act... Participation: The EM SSAB, Idaho National Laboratory, welcomes the attendance of the public at its advisory...

  19. 77 FR 76475 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-12-28

    ...] [FR Doc No: 2012-31173] DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board... notice announces a meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Idaho... the Board is to make recommendations to DOE-EM and site management in the areas of environmental...

  20. 76 FR 30695 - Environmental Management Site-Specific Advisory Board Chairs

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-26

    ... Environmental Management Site-Specific Advisory Board Chairs AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB) Chairs. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770) requires... Program Update, EM SSAB Chairs' Round Robin: Top Three Site-Specific Topics and Achievements, EM...

  1. 76 FR 38143 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-06-29

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy, DOE... Site-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L.... ADDRESSES: Savannah Rapids Pavilion, 3300 Evans to Locks Road, Martinez, GA 30907. FOR FURTHER...

  2. 77 FR 4027 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-01-26

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... and site management in the areas of environmental restoration, waste management, and...

  3. 78 FR 20311 - Environmental Management Site-Specific Advisory Board Chairs

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-04-04

    ... Environmental Management Site-Specific Advisory Board Chairs AGENCY: Department of Energy. ACTION: Notice of Open Webinar. SUMMARY: This notice announces a webinar of the Environmental Management Site-Specific... recommendations to DOE-EM and site management in the areas of environmental restoration, waste management,...

  4. 78 FR 53135 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-28

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... recommendations to DOE-EM and site management in the areas of environmental restoration, waste management,...

  5. 76 FR 80354 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-23

    ...] [FR Doc No: 2011-32910] DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board... meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Nevada. The Federal... environmental restoration, waste management, and related activities. Tentative Agenda: Fiscal Year...

  6. 76 FR 80355 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-23

    ...] [FR Doc No: 2011-32913] DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board... a meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Paducah. The... environmental restoration, waste management, and related activities. Tentative Agenda Call to Order...

  7. 76 FR 20651 - Environmental Management Site-Specific Advisory Board Chairs

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-13

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board Chairs AGENCY: Department of Energy. ACTION: Notice of... Environmental Management Site-Specific Advisory Board Chairs (76 FR 17118). This notice announces the...

  8. 76 FR 63613 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-10-13

    ...] [FR Doc No: 2011-26475] DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board... meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Hanford. The Federal... is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  9. 78 FR 26635 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-05-07

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy. ACTION: Notice of... of open meeting announcing a meeting on May 16, 2013 of the Environmental Management Site-Specific..., Deputy Committee Management Officer. BILLING CODE 6450-01-P ...

  10. 75 FR 56526 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-16

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION... Environmental Management Site-Specific Advisory Board (EM SSAB), Portsmouth. The Federal Advisory Committee Act... areas of environmental restoration, waste management and related activities. Tentative Agenda Portsmouth...

  11. 78 FR 59012 - Environmental Management Site-Specific Advisory Board Chairs

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-25

    ... Environmental Management Site-Specific Advisory Board Chairs AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... environmental restoration, waste management, and related activities. Tentative Agenda Topics: Wednesday, October...

  12. 75 FR 19630 - Environmental Management Site-Specific Advisory Board Charter Renewal

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-15

    ... No: 2010-8658] DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board Charter... site-specific issues include clean-up standards and environmental restoration; waste management and... determined to be essential to conduct Department of Energy business and to be in the public interest in...

  13. 29 CFR 1926.752 - Site layout, site-specific erection plan and construction sequence.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 29 Labor 8 2010-07-01 2010-07-01 false Site layout, site-specific erection plan and construction... Steel Erection § 1926.752 Site layout, site-specific erection plan and construction sequence. (a... intended minimum compressive design strength or sufficient strength to support the loads imposed...

  14. 29 CFR 1926.752 - Site layout, site-specific erection plan and construction sequence.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 29 Labor 8 2011-07-01 2011-07-01 false Site layout, site-specific erection plan and construction... Steel Erection § 1926.752 Site layout, site-specific erection plan and construction sequence. (a... intended minimum compressive design strength or sufficient strength to support the loads imposed...

  15. 76 FR 52944 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-24

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... (EIS) Committee of the Environmental Management Site- Specific Advisory Board (EM SSAB), Nevada. The... be announced in the Federal Register. DATES: Wednesday, September 7, 2011; 2 p.m. ADDRESSES:...

  16. 76 FR 50204 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-12

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... (EIS) Committee of the Environmental Management Site- Specific Advisory Board (EM SSAB), Nevada. The... be announced in the Federal Register. DATES: Wednesday, August 17, 2011, 2 p.m. ADDRESSES:...

  17. 76 FR 55370 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-07

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... (EIS) Committee of the Environmental Management Site- Specific Advisory Board (EM SSAB), Nevada. The... be announced in the Federal Register. DATES: Tuesday, October 4, 2011; 2 p.m. ADDRESSES: Nevada...

  18. Expanding the Scope of Site-Specific Recombinases for Genetic and Metabolic Engineering

    PubMed Central

    Gaj, Thomas; Sirk, Shannon J.; Barbas, Carlos F.

    2014-01-01

    Site-specific recombinases are tremendously valuable tools for basic research and genetic engineering. By promoting high-fidelity DNA modifications, site-specific recombination systems have empowered researchers with unprecedented control over diverse biological functions, enabling countless insights into cellular structure and function. The rigid target specificities of many sites-specific recombinases, however, have limited their adoption in fields that require highly flexible recognition abilities. As a result, intense effort has been directed toward altering the properties of site-specific recombination systems by protein engineering. Here, we review key developments in the rational design and directed molecular evolution of site-specific recombinases, highlighting the numerous applications of these enzymes across diverse fields of study. PMID:23982993

  19. How input noise limits biochemical sensing in ultrasensitive systems

    NASA Astrophysics Data System (ADS)

    Hu, Bo; Rappel, Wouter-Jan; Levine, Herbert

    2014-09-01

    Many biological processes are regulated by molecular devices that respond in an ultrasensitive fashion to upstream signals. An important question is whether such ultrasensitivity improves or limits its ability to read out the (noisy) input stimuli. Here, we develop a simple model to study the statistical properties of ultrasensitive signaling systems. We demonstrate that the output sensory noise is always bounded, in contrast to earlier theories using the small noise approximation, which tends to overestimate the impact of noise in ultrasensitive pathways. Our analysis also shows that the apparent sensitivity of the system is ultimately constrained by the input signal-to-noise ratio. Thus, ultrasensitivity can improve the precision of biochemical sensing only to a finite extent. This corresponds to a new limit for ultrasensitive signaling systems, which is strictly tighter than the Berg-Purcell limit.

  20. FLP recombinase-mediated site-specific recombination in silkworm, Bombyx mori.

    PubMed

    Long, Ding-Pei; Zhao, Ai-Chun; Chen, Xue-Jiao; Zhang, Yang; Lu, Wei-Jian; Guo, Qing; Handler, Alfred M; Xiang, Zhong-Huai

    2012-01-01

    A comprehensive understanding of gene function and the production of site-specific genetically modified mutants are two major goals of genetic engineering in the post-genomic era. Although site-specific recombination systems have been powerful tools for genome manipulation of many organisms, they have not yet been established for use in the manipulation of the silkworm Bombyx mori genome. In this study, we achieved site-specific excision of a target gene at predefined chromosomal sites in the silkworm using a FLP/FRT site-specific recombination system. We first constructed two stable transgenic target silkworm strains that both contain a single copy of the transgene construct comprising a target gene expression cassette flanked by FRT sites. Using pre-blastoderm microinjection of a FLP recombinase helper expression vector, 32 G3 site-specific recombinant transgenic individuals were isolated from five of 143 broods. The average frequency of FLP recombinase-mediated site-specific excision in the two target strains genome was approximately 3.5%. This study shows that it is feasible to achieve site-specific recombination in silkworms using the FLP/FRT system. We conclude that the FLP/FRT system is a useful tool for genome manipulation in the silkworm. Furthermore, this is the first reported use of the FLP/FRT system for the genetic manipulation of a lepidopteran genome and thus provides a useful reference for the establishment of genome manipulation technologies in other lepidopteran species.

  1. FLP Recombinase-Mediated Site-Specific Recombination in Silkworm, Bombyx mori

    PubMed Central

    Long, Ding-Pei; Zhao, Ai-Chun; Chen, Xue-Jiao; Zhang, Yang; Lu, Wei-Jian; Guo, Qing; Handler, Alfred M.; Xiang, Zhong-Huai

    2012-01-01

    A comprehensive understanding of gene function and the production of site-specific genetically modified mutants are two major goals of genetic engineering in the post-genomic era. Although site-specific recombination systems have been powerful tools for genome manipulation of many organisms, they have not yet been established for use in the manipulation of the silkworm Bombyx mori genome. In this study, we achieved site-specific excision of a target gene at predefined chromosomal sites in the silkworm using a FLP/FRT site-specific recombination system. We first constructed two stable transgenic target silkworm strains that both contain a single copy of the transgene construct comprising a target gene expression cassette flanked by FRT sites. Using pre-blastoderm microinjection of a FLP recombinase helper expression vector, 32 G3 site-specific recombinant transgenic individuals were isolated from five of 143 broods. The average frequency of FLP recombinase-mediated site-specific excision in the two target strains genome was approximately 3.5%. This study shows that it is feasible to achieve site-specific recombination in silkworms using the FLP/FRT system. We conclude that the FLP/FRT system is a useful tool for genome manipulation in the silkworm. Furthermore, this is the first reported use of the FLP/FRT system for the genetic manipulation of a lepidopteran genome and thus provides a useful reference for the establishment of genome manipulation technologies in other lepidopteran species. PMID:22768245

  2. Development of site-specific earthquake response spectra for eastern US sites

    SciTech Connect

    Beavers, J.E.; Brock, W.R.; Hunt, R.J.; Shaffer, K.E.

    1993-08-01

    Site-specific earthquake, uniform-hazard response spectra have been defined for the Department of Energy Oak Ridge, Tennessee, and Portsmouth, Ohio, sites for use in evaluating existing facilities and designing new facilities. The site-specific response spectra were defined from probabilistic and deterministic seismic hazard studies following the requirements in DOE-STD-1024-92, ``Guidelines for Probabilistic Seismic Hazard Curves at DOE Sites.` For these two sites, the results show that site-specific uniform-hazard response spectra are slightly higher in the high-frequency range and considerably lower in the low-frequency range compared with response spectra defined for these sites in the past.

  3. SUPPORTING THE REDEVELOPMENT OF BROWNFIELD SITES USING SITE-SPECIFIC MANAGEMENT APPROACHES AND REDEVELOPMENT TOOLS (SMART)

    EPA Science Inventory

    The Site-Specific Management Approaches and Redevelopment Tools (SMART) provides potential solutions for facilitating the redevelopment of brownfield sites. The term "brownfield site" refers to previously developed property whose reuse may be complicated by the presence of hazar...

  4. A METHOD FOR THE MEASUREMENT OF SITE-SPECIFIC TAUTOMERIC AND ZWITTERIONIC MICROSPECIES EQUILIBRIUM CONSTANTS

    EPA Science Inventory

    We describe a method for the individual measurement of simultaneously occurring, unimolecular, site-specific "microequilibrium" constants as in, for example, prototropic tautomerism and zwitterionic equilibria. Our method represents an elaboration of that of Nygren et al. (Anal. ...

  5. SUPPORTING THE REDEVELOPMENT OF BROWNFIELD SITES USING SITE-SPECIFIC MANAGEMENT APPROACHES AND REDEVELOPMENT TOOLS (SMART)

    EPA Science Inventory

    The Site-Specific Management Approaches and Redevelopment Tools (SMART) provides potential solutions for facilitating the redevelopment of brownfield sites. The term "brownfield site" refers to previously developed property whose reuse may be complicated by the presence of hazar...

  6. 77 FR 11516 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-27

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION... FURTHER INFORMATION CONTACT: Reinhard Knerr, Deputy Designated Federal Officer, Department of Energy...

  7. A METHOD FOR THE MEASUREMENT OF SITE-SPECIFIC TAUTOMERIC AND ZWITTERIONIC MICROSPECIES EQUILIBRIUM CONSTANTS

    EPA Science Inventory

    We describe a method for the individual measurement of simultaneously occurring, unimolecular, site-specific "microequilibrium" constants as in, for example, prototropic tautomerism and zwitterionic equilibria. Our method represents an elaboration of that of Nygren et al. (Anal. ...

  8. 78 FR 38969 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-06-28

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION... FURTHER INFORMATION CONTACT: Rachel Blumenfeld, Deputy Designated Federal Officer, Department of...

  9. 77 FR 4799 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-01-31

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION... FURTHER INFORMATION CONTACT: Reinhard Knerr, Deputy Designated Federal Officer, Department of...

  10. 78 FR 73519 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-06

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE) ACTION: Notice... FURTHER INFORMATION CONTACT: Rachel Blumenfeld, Deputy Designated Federal Officer, Department of...

  11. METHOD FOR THE MEASUREMENT OF SITE-SPECIFIC TAUTOMERIC AND ZWITTERIONIC MICROSPECIES EQUILIBRIUM CONSTANTS

    EPA Science Inventory

    We describe a method for the individual measurement of simultaneously occurring, unimolecular, site-specific “microequilibrium” constants as in, for example, prototropic tautomerism and zwitterionic equilibria. Our method represents an elaboration of that of Nygren et al. (Anal. ...

  12. METHOD FOR THE MEASUREMENT OF SITE-SPECIFIC TAUTOMERIC AND ZWITTERIONIC MICROSPECIES EQUILIBRIUM CONSTANTS

    EPA Science Inventory

    We describe a method for the individual measurement of simultaneously occurring, unimolecular, site-specific “microequilibrium” constants as in, for example, prototropic tautomerism and zwitterionic equilibria. Our method represents an elaboration of that of Nygren et al. (Anal. ...

  13. 75 FR 17701 - Environmental Management Site-Specific Advisory Board Chairs

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-07

    ... 28, 2010 EM Program Update, Priorities, and American Recovery and Reinvestment Act Discussion EM SSAB Chairs' Round Robin: Top Three Site-Specific Issues, EM SSAB Accomplishments, and Major Board...

  14. Separation of non-hazardous, non-radioactive components from ICPP calcine via chlorination

    SciTech Connect

    Nelson, L.O.

    1995-05-01

    A pyrochemical treatment method for separating non-radioactive from radioactive components in solid granular waste accumulated at the Idaho Chemical Processing Plant was investigated. The goal of this study was to obtain kinetic and chemical separation data on the reaction products of the chlorination of the solid waste, known as calcine. Thermodynamic equilibrium calculations were completed to verify that a separation of radioactive and non-radioactive calcine components was possible. Bench-scale chlorination experiments were completed subsequently in a variety of reactor configurations including: a fixed-bed reactor (reactive gases flowed around and not through the particle bed), a packed/fluidized-bed reactor, and a packed-bed reactor (reactive gases flowed through the particle bed). Chemical analysis of the reaction products generated during the chlorination experiments verified the predictions made by the equilibrium calculations. An empirical first-order kinetic rate expression was developed for each of the reactor configurations. 20 refs., 16 figs., 21 tabs.

  15. Nonradioactive sequence-tagged microsatellite site analyses: a method transferable to the tropics.

    PubMed

    Lagoda, P J; Dambier, D; Grapin, A; Baurens, F C; Lanaud, C; Noyer, J L

    1998-02-01

    Utilization of existing isozyme analysis facilities to detect sequence-tagged microsatellite site (STMS) polymorphism or any simple sequence repeat (SSR) variation is described. Different parameters concerning the difficulties in transferring molecular techniques to less sophisticated laboratory infrastructures (i.e. tropical outstations) are discussed (e.g. reproducibility, efficacy, precision). Nonradioactive STMS analysis is bound to foster collaborative research between "biodiversity" and "biotechnology" centers.

  16. Development of Ultrasensitive Plasmonic Nanosensors

    NASA Astrophysics Data System (ADS)

    Joshi, Gayatribahen K.

    -based LSPR nanosensors ("plasmonic nanosensors") have been developed for different sensing applications. Specifically, these plasmonic nanosensors displayed capacity to detect streptavidine, glucose, microRNA (cancer biomarker), as well as molecular and stimuli-responsive polymers conformational changes. In this study we found that the plasmonic nanosensors are exceptionally sensitive compared to other NSs and the sensitivity is highly edge length dependent. An ultrasensitive plasmonic nanosensor has been developed for the detection of microRNAs in crude plasma collected from pancreatic cancer patients. It shows that the LSPR-based nanosensor has the ability to detect and quantify the microRNA concentrations in clinical samples without any purification. The results presented here show potential for patients to commence treatment in early stage cancer diagnosis. The effect of various physiological medias and edge length of nanoprisms on the sensitivity of this nanosensor has been discussed. Second, molecular sensors have been developed by functionalization of azobenzene molecule contain alkanethiols onto the nanoprisms surface. Molecular conformational changes basis on a very less dielectric thickness changes have been detected through lambdaLSPR shift of nanoprisms and confirmed through surface enhanced Raman spectroscopy (SERS). In this study, the influence of resonance energy transfer between the molecule and nanoprisms onto the lambda LSPR shift and Raman intensity has been investigated by changing the distance between them. Finally, utilization of stimuli-responsive polymers structural change in the development of stimuli-responsive such as pH and temperature-responsive plasmonic nanosensors has been demonstrated. It was found that the stimuli-responsive nanosensors were able to detect very small R.I. change due to the polymers structural change. The enzymatic reaction between glucose and glucose oxidase has been used to detect glucose in bovine plasma using p

  17. Site-Specific Acetyl Lysine Antibodies Reveal Differential Regulation of Histone Acetylation upon Kinase Inhibition.

    PubMed

    Chen, Shi; Chen, Suping; Duan, Qianqian; Xu, Guoqiang

    2017-03-01

    Lysine acetylation regulates diverse biological functions for the modified proteins. Mass spectrometry-based proteomic approaches have identified thousands of lysine acetylation sites in cells and tissues. However, functional studies of these acetylation sites were limited by the lack of antibodies recognizing the specific modification sites. Here, we generated 55 site-specific acetyl lysine antibodies for the detection of this modification in cell lysates and evaluated the quality of these antibodies. Based on the immunoblotting analyses, we found that the nature of amino acid sequences adjacent to the modification sites affected the specificity of the site-specific acetyl lysine antibodies. Amino acids with charged, hydrophilic, small, or flexible side chains adjacent to the modification sites increase the likelihood of obtaining high quality site-specific acetyl lysine antibodies. This result may provide valuable insights in fine-tuning the amino acid sequences of the epitopes for the generation of site-specific acetyl lysine antibodies. Using the site-specific acetyl lysine antibodies, we further discovered that acetylation of histone 3 at four lysine residues was differentially regulated by kinase inhibitors. This result demonstrates the potential application of these antibodies in the study of new signaling pathways that lysine acetylation may participate in.

  18. In vivo and in vitro characterization of site-specific recombination of actinophage R4 integrase.

    PubMed

    Miura, Takamasa; Hosaka, Yayoi; Yan-Zhuo, Yang; Nishizawa, Tomoyasu; Asayama, Munehiko; Takahashi, Hideo; Shirai, Makoto

    2011-01-01

    The site-specific integrase of actinophage R4 belongs to the serine recombinase family. During the lysogenization process, it catalyzes site-specific recombination between the phage genome and the chromosome of Streptomyces parvulus 2297. An in vivo assay using Escherichia coli cells revealed that the minimum lengths of the recombination sites attB and attP are 50-bp and 49-bp, respectively, for efficient intramolecular recombination. The in vitro assay using overproduced R4 integrases as a hexahistidine (His(6))-glutathione-S-transferase (GST)-R4 integrase fusion protein, showed that the purified protein preparation retains the site-specific recombination activity which catalyzes the site-specific recombination between attP and attB in the intermolecular reaction. It also revealed that the inverted repeat within attP is essential for efficient in vitro intermolecular recombination. In addition, a gel shift assay showed that His(6)-GST-R4 integrase bound to the 50-bp attB and 49-bp attP specifically. Moreover, based on a detailed comparison analysis of amino acid sequences of serine integrases, we found the DNA binding region that is conserved in the serine recombinase containing the large C-terminal domain. Based on the results presented on this report, attachment sites needed in vitro and in vivo for site-specific recombination by the R4 integrase have been defined more precisely. This knowledge is useful for developing new genetic manipulation tools in the future.

  19. Site-specific water quality guidelines: 1. Derivation approaches based on physicochemical, ecotoxicological and ecological data.

    PubMed

    van Dam, R A; Humphrey, C L; Harford, A J; Sinclair, A; Jones, D R; Davies, S; Storey, A W

    2014-01-01

    Generic water quality guidelines (WQGs) are developed by countries/regions as broad scale tools to assist with the protection of aquatic ecosystems from the impacts of toxicants. However, since generic WQGs cannot adequately account for the many environmental factors that may affect toxicity at a particular site, site-specific WQGs are often needed, especially for high environmental value ecosystems. The Australian and New Zealand Guidelines for Fresh and Marine Water Quality provide comprehensive guidance on methods for refining or deriving WQGs for site-specific purposes. This paper describes three such methods for deriving site-specific WQGs, namely: (1) using local reference water quality data, (2) using biological effects data from laboratory-based toxicity testing, and (3) using biological effects data from field surveys. Two case studies related to the assessment of impacts arising from mining operations in northern Australia are used to illustrate the application of these methods. Finally, the potential of several emerging methods designed to assess thresholds of ecological change from field data for deriving site-specific WQGs is discussed. Ideally, multiple lines of evidence approaches, integrating both laboratory and field data, are recommended for deriving site-specific WQGs.

  20. A study to control chemical reactions using Si:2p core ionization: site-specific fragmentation.

    PubMed

    Nagaoka, Shin-ichi; Fukuzawa, Hironobu; Prümper, Georg; Takemoto, Mai; Takahashi, Osamu; Yamaguchi, Katsuhiro; Kakiuchi, Takuhiro; Tabayashi, Kiyohiko; Suzuki, Isao H; Harries, James R; Tamenori, Yusuke; Ueda, Kiyoshi

    2011-08-18

    In an aim to create a "sharp" molecular knife, we have studied site-specific fragmentation caused by Si:2p core photoionization of bridged trihalosilyltrimethylsilyl molecules in the vapor phase. Highly site-specific bond dissociation has been found to occur around the core-ionized Si site in some of the molecules studied. The site specificity in fragmentation and the 2p binding energy difference between the two Si sites depend in similar ways on the intersite bridge and the electronegativities of the included halogen atoms. The present experimental and computational results show that for efficient "cutting" the following conditions for the two atomic sites to be separated by the knife should be satisfied. First, the sites should be located far from each other and connected by a chain of saturated bonds so that intersite electron migration can be reduced. Second, the chemical environments of the atomic sites should be as different as possible.

  1. Industrial energy conservation-center-pivot site specific irrigation. Final report, January 1995--December 1996

    SciTech Connect

    McCann, I.R.; King, B.A.; Brady, R.

    1996-12-27

    This project was to aid in the development and commercial transfer of site specific technology to industry and farmers. This report contains the results of data collected during the fall of 1996 on a site near Aberdeen, Idaho. This site was equipped to apply water at variable rates predetermined by a digital control map that resided in a computer controller. The flow rates were then adjusted to maintain desired pressure with an Adjustable Speed Drive. Energy consumption was monitored during implementation of site specific irrigation practices and normal irrigation practices. This final report covers the impact that site specific irrigation has on energy use on a irrigated small grain crop near Aberdeen, Idaho. 3 refs., 20 figs., 1 tab.

  2. Bifunctional chelating agent for the design and development of site specific radiopharmaceuticals and biomolecule conjugation strategy

    DOEpatents

    Katti, Kattesh V.; Prabhu, Kandikere R.; Gali, Hariprasad; Pillarsetty, Nagavara Kishore; Volkert, Wynn A.

    2003-10-21

    There is provided a method of labeling a biomolecule with a transition metal or radiometal in a site specific manner to produce a diagnostic or therapeutic pharmaceutical compound by synthesizing a P.sub.2 N.sub.2 -bifunctional chelating agent intermediate, complexing the intermediate with a radio metal or a transition metal, and covalently linking the resulting metal-complexed bifunctional chelating agent with a biomolecule in a site specific manner. Also provided is a method of synthesizing the --PR.sub.2 containing biomolecules by synthesizing a P.sub.2 N.sub.2 -bifunctional chelating agent intermediate, complexing the intermediate with a radiometal or a transition metal, and covalently linking the resulting radio metal-complexed bifunctional chelating agent with a biomolecule in a site specific manner. There is provided a therapeutic or diagnostic agent comprising a --PR.sub.2 containing biomolecule.

  3. 76 FR 30027 - Land Disposal Restrictions: Site-Specific Treatment Variance for Hazardous Selenium-Bearing Waste...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-24

    ... Selenium-Bearing Waste Treated by U.S. Ecology Nevada in Beatty, NV and Withdrawal of Site-Specific... site-specific treatment variance to U.S. Ecology Nevada in Beatty, Nevada and withdrew an existing site... variance to U.S. Ecology Nevada in Beatty, Nevada and withdrawing an existing site-specific treatment...

  4. Nonlinear nanomechanical oscillators for ultrasensitive inertial detection

    DOEpatents

    Datskos, Panagiotis George; Lavrik, Nickolay V

    2013-08-13

    A system for ultrasensitive mass and/or force detection of this invention includes a mechanical oscillator driven to oscillate in a nonlinear regime. The mechanical oscillator includes a piezoelectric base with at least one cantilever resonator etched into the piezoelectric base. The cantilever resonator is preferably a nonlinear resonator which is driven to oscillate with a frequency and an amplitude. The system of this invention detects an amplitude collapse of the cantilever resonator at a bifurcation frequency as the cantilever resonator stimulated over a frequency range. As mass and/or force is introduced to the cantilever resonator, the bifurcation frequency shifts along a frequency axis in proportion to the added mass.

  5. High Speed, Ultrasensitive Trace Gas Sensing

    NASA Astrophysics Data System (ADS)

    Long, David A.; Fleisher, Adam J.; Plusquellic, David F.; Hodges, Joseph

    2014-06-01

    I will describe a variety of cavity-enhanced spectroscopic techniques, including frequency-agile rapid scanning spectroscopy (FARS) and heterodyne-detected cavity ring-down spectroscopy (HD-CRDS), which we have recently developed for rapid, ultrasensitive absorption measurements. Scanning rates that are limited only by the cavity response time itself as well as noise-equivalent detection limits as low as 6×10-14 cm-1 Hz-1/2 have been achieved. I will discuss the application of these techniques to current problems in atmospheric science including recent infrared measurements of analytes which are present at ultra-trace concentrations.

  6. Quantitation of pyrimidine dimer contents of nonradioactive deoxyribonucleic acid by electrophoresis in alkaline agarose gels

    SciTech Connect

    Sutherland, B.M.; Shih, A.G.

    1983-02-15

    We have developed a method of quantitating the pyrimidine dimer content of nonradioactive DNAs. DNA samples are treated with the UV-endonuclease from Micrococcus luteus and then separated according to molecular weight by electrophoresis on alkaline agarose gels. From their migration relative to known molecular weight standards, their median molecular weight and thus the number of dimers per DNA molecule in each sample can be calculated. Results of action spectra for dimer formation in T7 bacteriophage measured by this method agree well with action spectra for T7 killing. In addition, the method gives dimer yields in good agreement with those obtained by others using alkaline sucrose gradient sedimentation.

  7. Site-specific management of pH-induced iron chlorosis of maize

    USDA-ARS?s Scientific Manuscript database

    A study was conducted over nine site/years in Nebraska, USA between 2004 and 2005 to evaluate the potential to predict chlorosis-prone areas within fields which are relatively stable in space and time. The study also investigated the potential benefits of site-specific cultivar management according ...

  8. Environmental Restoration Site-Specific Plan for the Portsmouth Gaseous Diffusion Plant, FY 93

    SciTech Connect

    Not Available

    1993-01-15

    The purpose of this Site-Specific Plan (SSP) is to describe past, present, and future activities undertaken to implement Environmental Restoration and Waste Management goals at the Portsmouth Gaseous Diffusion Plant (PORTS). The SSP is presented in sections emphasizing Environmental Restoration description of activities, resources, and milestones.

  9. 75 FR 11872 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-03-12

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy..., Idaho National Laboratory to be held on March 16, 2010 75 FR 9590. In that notice, the meeting...

  10. 77 FR 47047 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-08-07

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... purpose of the Board is to make recommendations to DOE-EM and site management in the areas...

  11. 75 FR 82001 - Environmental Management Site-Specific Advisory Board, Oak Ridge Reservation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-12-29

    ... Board, Oak Ridge Reservation AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Oak... Center, 475 Oak Ridge Turnpike, Oak Ridge, Tennessee 37830. FOR FURTHER INFORMATION CONTACT: Patricia J...

  12. 76 FR 18921 - Land Disposal Restrictions: Nevada and California; Site Specific Treatment Variances for...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-06

    ... variance to U.S. Ecology Nevada (USEN) in Beatty, Nevada and to withdraw an existing site-specific.... Ecology Nevada located in Beatty, Nevada and to Chemical Waste Management, Inc. located in Kettleman Hills...-Bearing Waste II. Basis for This Determination III. Development of This Variance A. U.S. Ecology Nevada...

  13. Hellsgate Big Game Winter Range Wildlife Mitigation Site Specific Management Plan for the Hellsgate Project.

    SciTech Connect

    Berger, Matthew T.; Judd, Steven L.

    1999-01-01

    This report contains a detailed site-specific management plan for the Hellsgate Winter Range Wildlife Mitigation Project. The report provides background information about the mitigation process, the review process, mitigation acquisitions, Habitat Evaluation Procedures (HEP) and mitigation crediting, current habitat conditions, desired future habitat conditions, restoration/enhancements efforts and maps.

  14. 30 CFR 46.11 - Site-specific hazard awareness training.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Site-specific hazard awareness training. 46.11 Section 46.11 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING TRAINING AND RETRAINING OF MINERS ENGAGED IN SHELL DREDGING OR EMPLOYED AT SAND, GRAVEL, SURFACE...

  15. Site-specific ionisation edge fine-structure of Rutile in the electron microscope.

    PubMed

    Hetaba, Walid; Löffler, Stefan; Willinger, Marc-Georg; Schuster, Manfred Erwin; Schlögl, Robert; Schattschneider, Peter

    2014-08-01

    Combined Bloch-wave and density functional theory simulations are performed to investigate the effects of different channelling conditions on the fine-structure of electron energy-loss spectra. The simulated spectra compare well with experiments. Furthermore, we demonstrate that using this technique, the site-specific investigation of atomic orbitals is possible. This opens new possibilities for chemical analyses.

  16. 78 FR 10612 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-14

    ..., Santa Fe, NM 87506. Phone (505) 995-0393; Fax (505) 989-1752 or Email: msantistevan@doeal.gov... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Northern New Mexico (known locally as the Northern New Mexico Citizens...

  17. 75 FR 35446 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-22

    ..., 2010, 2 p.m.-4 p.m. ADDRESSES: NNMCAB Conference Room, 1660 Old Pecos Trail, Suite B, Santa Fe, NM... (NNMCAB), 1660 Old Pecos Trail, Suite B, Santa Fe, NM 87505. Phone (505) 995-0393; fax (505) 989-1752 or e... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...

  18. 40 CFR 60.3019 - What site-specific documentation is required?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 6 2010-07-01 2010-07-01 false What site-specific documentation is required? 60.3019 Section 60.3019 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR... operators that addresses the nine topics described in paragraphs (a)(1) through (9) of this section....

  19. 40 CFR 60.2910 - What site-specific documentation is required?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 6 2010-07-01 2010-07-01 false What site-specific documentation is required? 60.2910 Section 60.2910 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR... facility and readily accessible for all OSWI unit operators that addresses the nine topics described...

  20. 40 CFR 60.2095 - What site-specific documentation is required?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 6 2010-07-01 2010-07-01 false What site-specific documentation is required? 60.2095 Section 60.2095 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR... ten topics described in paragraphs (a)(1) through (10) of this section. You must maintain...

  1. 40 CFR 60.2660 - What site-specific documentation is required?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 6 2010-07-01 2010-07-01 false What site-specific documentation is required? 60.2660 Section 60.2660 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR... accessible for all CISWI unit operators that addresses the ten topics described in paragraphs (a)(1)...

  2. 40 CFR 60.2095 - What site-specific documentation is required?

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 7 2013-07-01 2013-07-01 false What site-specific documentation is required? 60.2095 Section 60.2095 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR... readily accessible for all CISWI unit operators that addresses the ten topics described in paragraphs...

  3. 78 FR 64932 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-10-30

    ..., November 20, 2013, 5:00 p.m. ADDRESSES: National Atomic Testing Museum, 755 E. Flamingo Road, Las Vegas... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  4. 78 FR 17192 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-20

    ... 17, 2013 5:00 p.m. ADDRESSES: National Atomic Testing Museum, 755 E. Flamingo Road, Las Vegas, Nevada... Underground Testing Area Activity Public Participation: The EM SSAB, Nevada, welcomes the attendance of the... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice...

  5. 77 FR 75627 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-12-21

    ... 16, 2013, 5:00 p.m. ADDRESSES: National Atomic Testing Museum, 755 E. Flamingo Road, Las Vegas... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  6. 75 FR 48662 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-11

    ...: Wednesday, September 1, 2010, 4 p.m. ADDRESSES: Atomic Testing Museum, 755 East Flamingo Road, Las Vegas... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada Test Site. The Federal Advisory Committee Act (Pub. L. 92-463, 86...

  7. 76 FR 21878 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-19

    ..., 2011, 5 p.m. ADDRESSES: Atomic Testing Museum, 755 East Flamingo Road, Las Vegas, Nevada 89119. FOR... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  8. 77 FR 49442 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-08-16

    ..., September 12, 2012, 4:00 p.m. ADDRESSES: Atomic Testing Museum, 755 E. Flamingo Road, Las Vegas, Nevada... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. No. 92-463, 86 Stat....

  9. 78 FR 23760 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-04-22

    ..., 2013, 5:00 p.m. ADDRESSES: National Atomic Testing Museum, 755 E. Flamingo Road, Las Vegas, Nevada... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  10. 76 FR 10343 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-24

    ..., 2011, 5 p.m. ADDRESSES: Atomic Testing Museum, 755 East Flamingo Road, Las Vegas, Nevada 89119. FOR... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  11. 76 FR 51362 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-18

    ..., September 14, 2011, 4 p.m. ADDRESSES: Atomic Testing Museum, 755 East Flamingo Road, Las Vegas, Nevada 89119... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  12. 77 FR 24694 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-04-25

    ..., 2012, 5 p.m. ADDRESSES: Atomic Testing Museum, 755 East Flamingo Road, Las Vegas, Nevada 89119. FOR... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  13. 77 FR 66962 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-11-08

    ..., November 28, 2012, 5:00 p.m. ADDRESSES: National Atomic Testing Museum, 755 E. Flamingo Road, Las Vegas... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat....

  14. Stoichiometry of site-specific lysine acetylation in an entire proteome.

    PubMed

    Baeza, Josue; Dowell, James A; Smallegan, Michael J; Fan, Jing; Amador-Noguez, Daniel; Khan, Zia; Denu, John M

    2014-08-01

    Acetylation of lysine ϵ-amino groups influences many cellular processes and has been mapped to thousands of sites across many organisms. Stoichiometric information of acetylation is essential to accurately interpret biological significance. Here, we developed and employed a novel method for directly quantifying stoichiometry of site-specific acetylation in the entire proteome of Escherichia coli. By coupling isotopic labeling and a novel pairing algorithm, our approach performs an in silico enrichment of acetyl peptides, circumventing the need for immunoenrichment. We investigated the function of the sole NAD(+)-dependent protein deacetylase, CobB, on both site-specific and global acetylation. We quantified 2206 peptides from 899 proteins and observed a wide distribution of acetyl stoichiometry, ranging from less than 1% up to 98%. Bioinformatic analysis revealed that metabolic enzymes, which either utilize or generate acetyl-CoA, and proteins involved in transcriptional and translational processes displayed the highest degree of acetylation. Loss of CobB led to increased global acetylation at low stoichiometry sites and induced site-specific changes at high stoichiometry sites, and biochemical analysis revealed altered acetyl-CoA metabolism. Thus, this study demonstrates that sirtuin deacetylase deficiency leads to both site-specific and global changes in protein acetylation stoichiometry, affecting central metabolism.

  15. 77 FR 50488 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-08-21

    ..., waste management and related activities. Tentative Agenda Call to Order, Introductions, Review of Agenda... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site...

  16. 76 FR 70121 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-11-10

    ..., waste management and related activities. Tentative Agenda Call to Order, Introductions, Review of Agenda... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site...

  17. 78 FR 16260 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-14

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  18. 77 FR 39234 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-02

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  19. 77 FR 16021 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-03-19

    ..., waste management and related activities. Tentative Agenda Call to Order, Introductions, Review of Agenda... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site...

  20. 76 FR 8359 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-14

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... purpose of the Board is to make recommendations to DOE-EM and site management in the areas of...

  1. 78 FR 49739 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-15

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... of the Board: The purpose of the Board is to make recommendations to DOE-EM and site management in...

  2. 78 FR 10611 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-14

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... purpose of the Board is to make recommendations to DOE-EM and site management in the areas of...

  3. 78 FR 25064 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-04-29

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  4. 76 FR 64329 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-10-18

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... purpose of the Board is to make recommendations to DOE-EM and site management in the areas of...

  5. 78 FR 45518 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-29

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  6. 78 FR 10612 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-14

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ] ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management... of the Board: The purpose of the Board is to make recommendations to DOE-EM and site management in...

  7. 77 FR 2283 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-01-17

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site..., Department of Energy Portsmouth/Paducah Project Office, Post Office Box 700, Piketon, Ohio 45661, (740)...

  8. 77 FR 6790 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-09

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site..., Department of Energy Portsmouth/Paducah Project Office, Post Office Box 700, Piketon, Ohio 45661, (740)...

  9. Use of GIS-based site-specific nitrogen management for improving energy efficiency

    USDA-ARS?s Scientific Manuscript database

    Nitrogen (N) is a significant energy component of in support of crop production but it can be highly variable within fields. To our knowledge, no efforts have been made to employ GIS-based site-specific N management (SSNM) to assess and improve energy costs and efficiency. We examine recent SSNM ca...

  10. Site-specific management of soil pH and nutrients in blueberry

    USDA-ARS?s Scientific Manuscript database

    Site-specific management of soil pH and fertilizers is one of the most promising strategies in precision agriculture and is potentially applicable to many horticultural crops, including blueberry. Unlike most fruit crops, blueberry is adapted to low soil pH conditions in the range of 4-5.5 and has ...

  11. 77 FR 31837 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-05-30

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... and site management in the areas of environmental restoration, waste management and related activities...

  12. SITE-SPECIFIC PROTOCOL FOR MEASURING SOIL RADON POTENTIALS FOR FLORIDA HOUSES

    EPA Science Inventory

    The report describes a protocol for site-specific measurement of radon potentials for Florida houses that is consistent with existing residential radon protection maps. The protocol gives further guidance on the possible need for radon-protective house construction features. In a...

  13. Site-specific mouth rinsing can improve oral odor by altering bacterial counts

    PubMed Central

    Alqumber, Mohammed A.; Arafa, Khaled A.

    2014-01-01

    Objectives: To determine whether site-specific mouth rinsing with oral disinfectants can improve oral odor beyond the traditional panoral mouth disinfection with mouth rinses by targeting specifically oral malodor implicated anaerobic bacteria Methods: Twenty healthy fasting subjects volunteered for a blinded prospective, descriptive correlational crossover cross-section clinical trial conducted during the month of Ramadan between July and August 2013 in Albaha province in Saudi Arabia involving the application of Listerine® Cool Mint® mouth rinse by either the traditional panoral rinsing method, or a site-specific disinfection method targeting the subgingival and supragingival plaque and the posterior third of the tongue dorsum, while avoiding the remaining locations within the oral cavity. The viable anaerobic and aerobic bacterial counts, volatile sulfur compounds (VSCs) levels, organoleptic assessment of oral odor, and the tongue-coating index were compared at baseline, one, 5, and 9 hours after the treatment. Results: The site-specific disinfection method reduced the VSCs and anaerobic bacterial loads while keeping the aerobic bacterial numbers higher than the traditional panoral rinsing method. Conclusion: Site-specific disinfection can more effectively maintain a healthy oral cavity by predominantly disinfecting the niches of anaerobic bacteria within the oral cavity. PMID:25399224

  14. LLNL Site Specific ASCI Software Quality Engineering Recommended Practices Overview Version 1.0

    SciTech Connect

    Peck, T; Sparkman, D; Storch, N

    2002-02-01

    ''The LLNL Site-Specific Advanced Simulation and Computing (ASCI) Software Quality Engineering Recommended Practices VI.I'' document describes a set of recommended software quality engineering (SQE) practices for ASCI code projects at Lawrence Livermore National Laboratory (LLNL). In this context, SQE is defined as the process of building quality into software products by applying the appropriate guiding principles and management practices. Continual code improvement and ongoing process improvement are expected benefits. Certain practices are recommended, although projects may select the specific activities they wish to improve, and the appropriate time lines for such actions. Additionally, projects can rely on the guidance of this document when generating ASCI Verification and Validation (VSrV) deliverables. ASCI program managers will gather information about their software engineering practices and improvement. This information can be shared to leverage the best SQE practices among development organizations. It will further be used to ensure the currency and vitality of the recommended practices. This Overview is intended to provide basic information to the LLNL ASCI software management and development staff from the ''LLNL Site-Specific ASCI Software Quality Engineering Recommended Practices VI.I'' document. Additionally the Overview provides steps to using the ''LLNL Site-Specific ASCI Software Quality Engineering Recommended Practices VI.I'' document. For definitions of terminology and acronyms, refer to the Glossary and Acronyms sections in the ''LLNL Site-Specific ASCI Software Quality Engineering Recommended Practices VI.I''.

  15. Summary of some feasibility studies for site-specific solar industrial process heat

    SciTech Connect

    1982-01-01

    Some feasibility studies for several different site specific solar industrial process heat applications are summarized. The followng applications are examined. Leather Tanning; Concrete Production: Lumber and Paper Processing; Milk Processing; Molding, Curing or Drying; Automobile Manufacture; and Food Processing and Preparation. For each application, site and process data, system design, and performance and cost estimates are summarized.

  16. 76 FR 24831 - Site-Specific Analyses for Demonstrating Compliance With Subpart C Performance Objectives

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-03

    ... proposing to amend its regulations to require low-level radioactive waste disposal facilities to conduct... safe disposal of low-level radioactive waste. The NRC is proposing additional changes to the... regulations to require low-level radioactive waste disposal facilities to conduct site-specific analyses...

  17. 76 FR 11772 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-03

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Northern New Mexico (known locally as the Northern New Mexico Citizens... Norte, Espanola, New Mexico 87532. FOR FURTHER INFORMATION CONTACT: Menice Santistevan, Northern New...

  18. 75 FR 53280 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-31

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Northern New Mexico. The Federal Advisory Committee Act (Pub. L. 92-463... Pueblo Sur, Taos, New Mexico 87571. FOR FURTHER INFORMATION CONTACT: Menice Santistevan, Northern New...

  19. 78 FR 23759 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-04-22

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Northern New Mexico. The Federal Advisory Committee Act (Pub. L. 92-463... Drive, Santa Fe, NM 87501. FOR FURTHER INFORMATION CONTACT: Menice Santistevan, Northern New Mexico...

  20. 78 FR 38305 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-06-26

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Northern New Mexico. The Federal Advisory Committee Act (Pub. L. 92-463... 87544. ] FOR FURTHER INFORMATION CONTACT: Menice Santistevan, Northern New Mexico Citizens' Advisory...

  1. 76 FR 11773 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-03

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Northern New Mexico. The Federal Advisory Committee Act (Pub. L. 92-463..., Santa Fe, New Mexico 87507. FOR FURTHER INFORMATION CONTACT: Menice Santistevan, Northern New Mexico...

  2. 76 FR 18540 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-04

    ... Environmental Management Site-Specific Advisory Board, Northern New Mexico AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Northern New Mexico (known locally as the Northern New Mexico Citizens....m.-4 p.m. ADDRESSES: Holiday Inn Express and Suites, 60 Entrada Drive, Los Alamos, New Mexico 87544...

  3. Design, synthesis, and characterization of nucleosomes containing site-specific DNA damage.

    PubMed

    Taylor, John-Stephen

    2015-12-01

    How DNA damaged is formed, recognized, and repaired in chromatin is an area of intense study. To better understand the structure activity relationships of damaged chromatin, mono and dinucleosomes containing site-specific damage have been prepared and studied. This review will focus on the design, synthesis, and characterization of model systems of damaged chromatin for structural, physical, and enzymatic studies.

  4. Automated measurement of site-specific N-glycosylation occupancy with SWATH-MS.

    PubMed

    Xu, Ying; Bailey, Ulla-Maja; Schulz, Benjamin L

    2015-07-01

    Asparagine-linked glycosylation is a common post-translational modification of proteins catalyzed by oligosaccharyltransferase that is important in regulating many aspects of protein function. Analysis of protein glycosylation, including glycoproteomic measurement of the site-specific extent of glycosylation, remains challenging. Here, we developed methods combining enzymatic deglycosylation and protease digestion with SWATH-MS to enable automated measurement of site-specific occupancy at many glycosylation sites. Deglycosylation with peptide-endoglycosidase H, leaving a remnant N-acetylglucosamine on asparagines previously carrying high-mannose glycans, followed by trypsin digestion allowed robust automated measurement of occupancy at many sites. Combining deglycosylation with the more general peptide-N-glycosidase F enzyme with AspN protease digest allowed robust automated differentiation of nonglycosylated and deglycosylated forms of a given glycosylation site. Ratiometric analysis of deglycosylated peptides and the total intensities of all peptides from the corresponding proteins allowed relative quantification of site-specific glycosylation occupancy between yeast strains with various isoforms of oligosaccharyltransferase. This approach also allowed robust measurement of glycosylation sites in human salivary glycoproteins. This method for automated relative quantification of site-specific glycosylation occupancy will be a useful tool for research with model systems and clinical samples.

  5. SITE-SPECIFIC PROTOCOL FOR MEASURING SOIL RADON POTENTIALS FOR FLORIDA HOUSES

    EPA Science Inventory

    The report describes a protocol for site-specific measurement of radon potentials for Florida houses that is consistent with existing residential radon protection maps. The protocol gives further guidance on the possible need for radon-protective house construction features. In a...

  6. CAN SITE-SPECIFIC TRENDS BE EXTRAPOLATED TO A REGION? AN ACIDIFICATION EXAMPLE FOR THE NORTHEAST

    EPA Science Inventory

    In the absence of true regional data on changes in the acid/base status of lakes in the northeastern United States, we explore the possibility of using site-specific trends information from a judgment sample of lakes to assess the efficacy of the Clean Air Act Amendments. A meta-...

  7. Derivation of site-specific skeletal masses within the current ICRP age series.

    PubMed

    Watchman, Christopher J; Hasenauer, Deanna; Bolch, Wesley E

    2007-06-07

    The calculation of absorbed dose to the radiosensitive tissues of the skeleton is routinely performed using reference masses provided in publications from the International Commission on Radiological Protection (ICRP). These values typically include total skeleton tissue masses by reference subject age, but not by individual bone site at a given age. Site-specific variations in absorbed fractions are known to occur for internal alpha-particle and beta-particle emitters, and in certain medical dose reconstructions, site-specific estimates of marrow dose may be desirable. Furthermore, bone-site-specific tissue masses are required to properly estimate skeletal-averaged absorbed fractions and, more importantly, specific absorbed fractions for internalized radionuclides and radiopharmaceuticals. Reference masses by skeletal site are also needed in the development of ICRP compliant tomographic phantoms, as this organ system is initially segmented from medical images only as a homogeneous tissue region. ICRP reference skeletal masses are assigned based upon several independent data sources, many of which may not be entirely consistent with one another. In this study, a methodology is presented, using data from the various ICRP publications, to derive site-specific skeletal tissue masses for each member of the ICRP age series. Active marrow masses are calculated and differences are shown with respect to ICRP Publications 70 and 89 values. New data for a revised surrogate tissue region for the osteoprogenitor cells within bone marrow is presented with estimates of its total mass throughout the skeleton and for different subject ages.

  8. Software Design for Wireless Sensor-based Site-specific Irrigation

    USDA-ARS?s Scientific Manuscript database

    In-field sensor-based site-specific irrigation management is of benefit to producers for efficient water management. Integration of the decision making process with the controls is a viable option for determining when and where to irrigate, and how much water to apply. This research presents the des...

  9. 78 FR 61348 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-10-03

    ...On September 16, 2013, in FR Doc. 2013-22453, on page 56871, the Department of Energy (DOE) published a notice of open meeting announcing a meeting on October 2, 2013 of the Environmental Management Site-Specific Advisory Board, Portsmouth (78 FR 56871). This notice announces the cancellation of this...

  10. CAN SITE-SPECIFIC TRENDS BE EXTRAPOLATED TO A REGION? AN ACIDIFICATION EXAMPLE FOR THE NORTHEAST

    EPA Science Inventory

    In the absence of true regional data on changes in the acid/base status of lakes in the northeastern United States, we explore the possibility of using site-specific trends information from a judgment sample of lakes to assess the efficacy of the Clean Air Act Amendments. A meta-...

  11. Use of GIS-based Site-specific Nitrogen Management for Improving Energy Efficiency

    USDA-ARS?s Scientific Manuscript database

    To our knowledge, geographical information system (GIS)-based site-specific nitrogen management (SSNM) techniques have not been used to assess agricultural energy costs and efficiency. This chapter uses SSNM case studies for corn (Zea mays L.) grown in Missouri and cotton (Gossypium hirsutum L.) gro...

  12. 75 FR 64718 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-20

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific.... Open Government Plan. Public Participation: The meeting is open to the public. The EM SSAB, Hanford....hanford.gov/page.cfm/hab . Issued at Washington, DC, on October 14, 2010. Rachel Samuel, Deputy...

  13. 78 FR 4139 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-01-18

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... Advisory Board (HAB) advice on the TC&WMFEIS. Status from Executive Issues Committee issues managers regarding HAB draft recommendations for Board diversity and other Board effectiveness issues....

  14. 77 FR 64112 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-18

    ... Committee Reports Board Business--Selection of new HAB Chair Public Participation: The meeting is open to... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... Protection Agency Draft White Paper on Hanford Advisory Board (HAB) Values Draft Letter/Advice--Other...

  15. 75 FR 6018 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-02-05

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... Discussion of HAB member comments on the TC and WM EIS Development of HAB advice principles Adjourn Public Participation: The meeting is open to the public. The EM SSAB, Hanford, welcomes the attendance of the public...

  16. 75 FR 8050 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-02-23

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... Environmental Impact Statement. Board Business. Public Participation: The meeting is open to the public. The EM.../page.cfm/hab . Issued at Washington, DC, on February 18, 2010. Rachel Samuel, Deputy...

  17. 77 FR 48131 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-08-13

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental ] Management Site-Specific... Board (HAB) Values White Paper Fiscal Year 2012 Board Accomplishments 2013 Tri-Party Agreement Priorities and HAB Work Plan Priorities 2013 HAB Meeting Calendar Board Business HAB Budget...

  18. 76 FR 14386 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-16

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific...: The meeting is open to the public. The EM SSAB, Hanford, welcomes the attendance of the public at its... site: http://www.hanford.gov/page.cfm/hab . Issued at Washington, DC, on March 11, 2011. LaTanya...

  19. 76 FR 28218 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-16

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... Discussions: Issue Managers. Advice Development. Public Participation: The meeting is open to the public. The... the following Web site: http://www.hanford.gov/page.cfm/hab . ] Issued at Washington, DC on May...

  20. 75 FR 13269 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-03-19

    ... Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific... meeting is open to the public. The EM SSAB, Hanford, welcomes the attendance of the public at its advisory.... Minutes will also be available at the following website: http://www.hanford.gov/page.cfm/hab . Issued...

  1. 75 FR 27999 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-19

    ... open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific..., M-91) 2012 Budget Request Board Business Public Participation: The meeting is open to the public... also be available at the following Web site: http://www.hanford.gov/page.cfm/hab . Issued at...

  2. Germinal transmission of site-specific excised genomic DNA by the bacterial ParA resolvase

    USDA-ARS?s Scientific Manuscript database

    Genome engineering is an essential tool in research and product development. Behind some of the recent advances in plant gene transfer is the development of site-specific recombination systems that enable the precise manipulation of DNA, e.g. the deletion, integration or translocation of DNA. DNA ...

  3. 40 CFR 170.232 - Knowledge of labeling and site-specific information.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Knowledge of labeling and site... (CONTINUED) PESTICIDE PROGRAMS WORKER PROTECTION STANDARD Standard for Pesticide Handlers § 170.232 Knowledge of labeling and site-specific information. (a) Knowledge of labeling information. (1) The handler...

  4. 40 CFR 170.232 - Knowledge of labeling and site-specific information.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Knowledge of labeling and site... (CONTINUED) PESTICIDE PROGRAMS WORKER PROTECTION STANDARD Standard for Pesticide Handlers § 170.232 Knowledge of labeling and site-specific information. (a) Knowledge of labeling information. (1) The handler...

  5. 40 CFR 170.232 - Knowledge of labeling and site-specific information.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Knowledge of labeling and site... (CONTINUED) PESTICIDE PROGRAMS WORKER PROTECTION STANDARD Standard for Pesticide Handlers § 170.232 Knowledge of labeling and site-specific information. (a) Knowledge of labeling information. (1) The handler...

  6. 40 CFR 170.232 - Knowledge of labeling and site-specific information.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Knowledge of labeling and site... (CONTINUED) PESTICIDE PROGRAMS WORKER PROTECTION STANDARD Standard for Pesticide Handlers § 170.232 Knowledge of labeling and site-specific information. (a) Knowledge of labeling information. (1) The handler...

  7. Site-specific risk factors for ray blight in Tasmanian pyrethrum fields

    USDA-ARS?s Scientific Manuscript database

    Ray blight of pyrethrum, caused by Phoma ligulicola var. inoxydablis can cause significant reductions in crop growth and pyrethrin yield. Weather and site-specific disease risk factors for ray blight have not been identified or quantified in terms of relative risk, which has limited the efficiency ...

  8. Site-Specific Amino Acid Preferences Are Mostly Conserved in Two Closely Related Protein Homologs

    PubMed Central

    Doud, Michael B.; Ashenberg, Orr; Bloom, Jesse D.

    2015-01-01

    Evolution drives changes in a protein’s sequence over time. The extent to which these changes in sequence lead to shifts in the underlying preference for each amino acid at each site is an important question with implications for comparative sequence-analysis methods, such as molecular phylogenetics. To quantify the extent that site-specific amino acid preferences shift during evolution, we performed deep mutational scanning on two homologs of human influenza nucleoprotein with 94% amino acid identity. We found that only a modest fraction of sites exhibited shifts in amino acid preferences that exceeded the noise in our experiments. Furthermore, even among sites that did exhibit detectable shifts, the magnitude tended to be small relative to differences between nonhomologous proteins. Given the limited change in amino acid preferences between these close homologs, we tested whether our measurements could inform site-specific substitution models that describe the evolution of nucleoproteins from more diverse influenza viruses. We found that site-specific evolutionary models informed by our experiments greatly outperformed nonsite-specific alternatives in fitting phylogenies of nucleoproteins from human, swine, equine, and avian influenza. Combining the experimental data from both homologs improved phylogenetic fit, partly because measurements in multiple genetic contexts better captured the evolutionary average of the amino acid preferences for sites with shifting preferences. Our results show that site-specific amino acid preferences are sufficiently conserved that measuring mutational effects in one protein provides information that can improve quantitative evolutionary modeling of nearby homologs. PMID:26226986

  9. 30 CFR 46.11 - Site-specific hazard awareness training.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 30 Mineral Resources 1 2014-07-01 2014-07-01 false Site-specific hazard awareness training. 46.11 Section 46.11 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING TRAINING AND RETRAINING OF MINERS ENGAGED IN SHELL DREDGING OR EMPLOYED AT SAND, GRAVEL, SURFACE...

  10. 30 CFR 46.11 - Site-specific hazard awareness training.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 30 Mineral Resources 1 2013-07-01 2013-07-01 false Site-specific hazard awareness training. 46.11 Section 46.11 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING TRAINING AND RETRAINING OF MINERS ENGAGED IN SHELL DREDGING OR EMPLOYED AT SAND, GRAVEL, SURFACE...

  11. 78 FR 30910 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-05-23

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management... the Board is to make recommendations to DOE-EM and site management in the areas of environmental...

  12. 75 FR 24685 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-05

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management... Board: The purpose of the Board is to make recommendations to DOE-EM and site management in the areas of...

  13. 78 FR 58294 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-23

    ... Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management... purpose of the Board is to make recommendations to DOE-EM and site management in the areas of...

  14. Integrated decision support, sensor networks and adaptive control for wireless site-specific sprinkler irrigation

    USDA-ARS?s Scientific Manuscript database

    The development of site-specific sprinkler irrigation water management systems will be a major factor in future efforts to improve the various efficiencies of water-use and to support a sustainable irrigated environment. The challenge is to develop fully integrated management systems with supporting...

  15. Integrated Decision Support, Sensor Networks and Adaptive Control for Wireless Site-specific Sprinkler Irrigation

    USDA-ARS?s Scientific Manuscript database

    The development of site-specific sprinkler irrigation water management systems will be a major factor in future efforts to improve the various efficiencies of water-use and to support a sustainable irrigated environment. The challenge is to develop fully integrated management systems with supporting...

  16. 75 FR 20832 - Environmental Management Site-Specific Advisory Board, Nevada Test Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-21

    ...This notice announces a meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Nevada Test Site. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770) requires that public notice of this meeting be announced in the Federal...

  17. 36 CFR 219.33 - Appeals of site-specific decisions.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 2 2011-07-01 2011-07-01 false Appeals of site-specific decisions. 219.33 Section 219.33 Parks, Forests, and Public Property FOREST SERVICE, DEPARTMENT OF AGRICULTURE PLANNING National Forest System Land and Resource Management Planning Objections and Appeals § 219...

  18. Site-specific management of cotton root rot using airborne and satellite imagery

    USDA-ARS?s Scientific Manuscript database

    Cotton root rot is a serious cotton disease that can now be effectively controlled with Topguard Terra Fungicide. The objectives of this research were to demonstrate how site-specific fungicide application could be implemented based on historical remote sensing imagery and variable rate technology. ...

  19. Stoichiometry of Site-specific Lysine Acetylation in an Entire Proteome*♦

    PubMed Central

    Baeza, Josue; Dowell, James A.; Smallegan, Michael J.; Fan, Jing; Amador-Noguez, Daniel; Khan, Zia; Denu, John M.

    2014-01-01

    Acetylation of lysine ϵ-amino groups influences many cellular processes and has been mapped to thousands of sites across many organisms. Stoichiometric information of acetylation is essential to accurately interpret biological significance. Here, we developed and employed a novel method for directly quantifying stoichiometry of site-specific acetylation in the entire proteome of Escherichia coli. By coupling isotopic labeling and a novel pairing algorithm, our approach performs an in silico enrichment of acetyl peptides, circumventing the need for immunoenrichment. We investigated the function of the sole NAD+-dependent protein deacetylase, CobB, on both site-specific and global acetylation. We quantified 2206 peptides from 899 proteins and observed a wide distribution of acetyl stoichiometry, ranging from less than 1% up to 98%. Bioinformatic analysis revealed that metabolic enzymes, which either utilize or generate acetyl-CoA, and proteins involved in transcriptional and translational processes displayed the highest degree of acetylation. Loss of CobB led to increased global acetylation at low stoichiometry sites and induced site-specific changes at high stoichiometry sites, and biochemical analysis revealed altered acetyl-CoA metabolism. Thus, this study demonstrates that sirtuin deacetylase deficiency leads to both site-specific and global changes in protein acetylation stoichiometry, affecting central metabolism. PMID:24917678

  20. Design, development and evaluation of a tree planting-site-specific fumigant applicator

    USDA-ARS?s Scientific Manuscript database

    The goal of this research was to use recent advances in the global positioning system (GPS) and computer technology to apply just the right amount of fumigant where it is most needed (i.e., tree-planting-site-specific application) to decrease the incidence of replant disease, and achieve the environ...

  1. 77 FR 76475 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-12-28

    ...This notice announces a meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Northern New Mexico. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770) requires that public notice of this meeting be announced in the Federal Register.

  2. 77 FR 10485 - Environmental Management Site-Specific Advisory Board, Idaho National Laboratory

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-22

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board, Idaho National Laboratory AGENCY: Department of Energy... Register. DATES: Wednesday, March 14, 2012, 8 a.m.-5 p.m. Opportunities for public participation will be...

  3. 77 FR 24695 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-04-25

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. . 92... Hilton Savannah DeSoto, 15 East Liberty Street Savannah, GA 31401. FOR FURTHER INFORMATION CONTACT:...

  4. 75 FR 24684 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-05

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463..., 601 East Bay Street, Savannah, Georgia 31401. FOR FURTHER INFORMATION CONTACT: Gerri Flemming,...

  5. 76 FR 25682 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-05

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463... Savannah, Two West Bay Street, Savannah, GA 31402. FOR FURTHER INFORMATION CONTACT: Gerri Flemming,...

  6. 77 FR 53193 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-08-31

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463... CONTACT: Gerri Flemming, Office of External Affairs, Department of Energy, Savannah River...

  7. 77 FR 13104 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-03-05

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463... of External Affairs, Department of Energy, Savannah River Operations Office, P.O. Box A, Aiken,...

  8. 78 FR 26005 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-05-03

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463...: Gerri Flemming, Office of External Affairs, Department of Energy, Savannah River Operations Office,...

  9. 78 FR 14088 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-04

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act requires that....m.-5:30 p.m.; Tuesday, March 26, 2013, 8:00 a.m.-4:30 p.m. ADDRESSES: Westin Savannah Harbor,...

  10. 76 FR 55369 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-07

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463... Flemming, Office of External Affairs, Department of Energy, Savannah River Operations Office, P.O. Box...

  11. 76 FR 65706 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-10-24

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463... Flemming, Office of External Affairs, Department of Energy, Savannah River Operations Office, P.O. Box...

  12. 78 FR 716 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-01-04

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463..., Office of External Affairs, Department of Energy, Savannah River Operations Office, P.O. Box A, Aiken,...

  13. 75 FR 39007 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-07-07

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463...: Gerri Flemming, Office of External Affairs, Department of Energy, Savannah River Operations Office,...

  14. 75 FR 82001 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-12-29

    ..., Savannah River Site AGENCY: Department of Energy. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Savannah River... FURTHER INFORMATION CONTACT: Gerri Flemming, Office of External Affairs, Department of Energy,...

  15. 78 FR 65979 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-04

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463..., Office of External Affairs, Department of Energy, Savannah River Operations Office, P.O. Box A, Aiken,...

  16. 75 FR 57462 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-21

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. No. 92...: Gerri Flemming, Office of External Affairs, Department of Energy, Savannah River Operations Office,...

  17. 76 FR 11772 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-03

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463... of External Affairs, Department of Energy, Savannah River Operations Office, P.O. Box A, Aiken,...

  18. 78 FR 16260 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-14

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board, Savannah River Site (78 FR 14088). This document makes a correction to that notice..., Savannah River Operations Office, P.O. ] Box A, Aiken, SC 29802; Phone: (803) 952-7886. Corrections In...

  19. 77 FR 60688 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-04

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463... Flemming, Office of External Affairs, Department of Energy, Savannah River Operations Office, P.O. Box...

  20. 78 FR 54461 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-04

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463.... ADDRESSES: Embassy Suites-Savannah, 145 Mulberry Boulevard, Savannah, GA 31322. FOR FURTHER...

  1. 77 FR 39235 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-02

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. 92-463...: Gerri Flemming, Office of External Affairs, Department of Energy, Savannah River Operations Office,...

  2. 78 FR 40130 - Environmental Management Site-Specific Advisory Board, Savannah River Site

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-03

    ... Environmental Management Site-Specific Advisory Board, Savannah River Site AGENCY: Department of Energy. ACTION...-Specific Advisory Board (EM SSAB), Savannah River Site. The Federal Advisory Committee Act (Pub. L. No. 92... CONTACT: Gerri Flemming, Office of External Affairs, Department of Energy, Savannah River...

  3. 30 CFR 46.11 - Site-specific hazard awareness training.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Site-specific hazard awareness training. 46.11 Section 46.11 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR EDUCATION AND TRAINING TRAINING AND RETRAINING OF MINERS ENGAGED IN SHELL DREDGING OR EMPLOYED AT SAND, GRAVEL,...

  4. 76 FR 78909 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-20

    ... Use Mapping and Data/ Wildlife Management Plan--Summary of Year 1 Field Work and Data Collection, Bob... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site...

  5. 78 FR 63172 - Environmental Management Site-Specific Advisory Board, Paducah; Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-10-23

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Environmental Management Site-Specific Advisory Board, Paducah; Meeting AGENCY: Department of Energy (DOE... of Energy Paducah Site Office, Post Office Box 1410, MS-103, Paducah, Kentucky 42001, (270)...

  6. Site-specific Topguard application based on aerial imagery for effective management of cotton root rot

    USDA-ARS?s Scientific Manuscript database

    Cotton root rot is a century-old cotton disease that can be controlled with Topguard Fungicide recently. As this disease tends to occur in the same general areas within fields in recurring years, site-specific application of the fungicide only to the infected areas can be more effective and economic...

  7. The parA resolvase performs site-specific genomic excision in Arabidopsis

    USDA-ARS?s Scientific Manuscript database

    We have designed a site-specific excision detection system in Arabidopsis to study the in planta activity of the small serine recombinase ParA. Using a transient expression assay as well as stable transgenic plant lines, we show that the ParA recombinase is catalytically active and capable of perfo...

  8. Development of Unmanned Aerial Vehicles for Site-Specific Crop Production Management

    USDA-ARS?s Scientific Manuscript database

    Unmanned Aerial Vehicles (UAV) have been developed and applied to support the practice of precision agriculture. Compared to piloted aircrafts, an Unmanned Aerial Vehicle can focus on much smaller crop fields with much lower flight altitude than regular airplanes to perform site-specific management ...

  9. Appreciating "Thirdspace": An Alternative Way of Viewing and Valuing Site-Specific Dance Performance

    ERIC Educational Resources Information Center

    Munjee, Tara

    2014-01-01

    Site-specific dance performance involves the presentation of choreography in connection with a site. The context of the site combined with a viewer's personal history, beliefs, and identity impact the reading and appreciation of the performance. Although both stage and site dance performance valuing elicit multiple interpretations of artistic…

  10. 36 CFR 219.33 - Appeals of site-specific decisions.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 2 2010-07-01 2010-07-01 false Appeals of site-specific decisions. 219.33 Section 219.33 Parks, Forests, and Public Property FOREST SERVICE, DEPARTMENT OF AGRICULTURE PLANNING National Forest System Land and Resource Management Planning Objections and Appeals §...

  11. 77 FR 43583 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-25

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities....

  12. 78 FR 78952 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-27

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities....

  13. Site-specific irrigation of peanuts on a Coastal Plain field

    USDA-ARS?s Scientific Manuscript database

    Irrigator-Pro is an expert system that prescribes irrigation for corn (Zea mays L.), cotton (Gossypium hirsutum L.) and peanut (Arachis hypogaea). We conducted an experiment in 2007 to evaluate Irrigator-Pro as a tool for variable rate irrigation of peanut using a site-specific center pivot irrigati...

  14. 76 FR 36100 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-06-21

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy. DOE. ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  15. 78 FR 32640 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-05-31

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  16. 75 FR 7577 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-02-22

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... environmental restoration, waste management and related activities. Tentative Agenda: Call to Order...

  17. 76 FR 48148 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-08

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  18. 75 FR 24686 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-05

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  19. 75 FR 66074 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-27

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of Open Meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  20. 76 FR 5147 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-01-28

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  1. 77 FR 2282 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-01-17

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy. ACTION: Notice of... of open meeting announcing a meeting on January 19, 2012, of the Environmental Management Site... Washington, DC, on January 12, 2012. LaTanya R. Butler, Acting Deputy Committee Management Officer. BILLING...

  2. 76 FR 57981 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-19

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  3. 75 FR 19379 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-14

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  4. 76 FR 48148 - Environmental Management Site-Specific Advisory Board, Hanford

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-08

    ... Environmental Management Site-Specific Advisory Board, Hanford AGENCY: Department of Energy (DoE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... environmental restoration, waste management, and related activities. Tentative Agenda: Annual Tri-Party Agency...

  5. 77 FR 59598 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-09-28

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  6. 77 FR 37390 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-06-21

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  7. 77 FR 29997 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-05-21

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  8. 75 FR 51026 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-18

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  9. 75 FR 54600 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-08

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE) ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  10. 77 FR 63300 - Environmental Management Site-Specific Advisory Board, Portsmouth

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-16

    ... Environmental Management Site-Specific Advisory Board, Portsmouth AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... Board is to make recommendations to DOE-EM and site management in the areas of environmental restoration...

  11. 75 FR 61711 - Environmental Management Site-Specific Advisory Board, Paducah

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-06

    ... Environmental Management Site-Specific Advisory Board, Paducah AGENCY: Department of Energy (DOE). ACTION: Notice of open meeting. SUMMARY: This notice announces a meeting of the Environmental Management Site... management in the areas of environmental restoration, waste management and related activities. Tentative...

  12. Appreciating "Thirdspace": An Alternative Way of Viewing and Valuing Site-Specific Dance Performance

    ERIC Educational Resources Information Center

    Munjee, Tara

    2014-01-01

    Site-specific dance performance involves the presentation of choreography in connection with a site. The context of the site combined with a viewer's personal history, beliefs, and identity impact the reading and appreciation of the performance. Although both stage and site dance performance valuing elicit multiple interpretations of artistic…

  13. 76 FR 80915 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-27

    ...This notice announces a meeting of the Environmental Management Site-Specific Advisory Board (EM SSAB), Northern New Mexico. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770) requires that public notice of this meeting be announced in the Federal Register.

  14. Evaluation of closed-loop site-specific irrigation with wireless sensor network

    USDA-ARS?s Scientific Manuscript database

    Automated site-specific sprinkler irrigation system can save water and maximize productivity, but implementing automated irrigation is challenging in system integration and decision making. A controllable irrigation system was integrated into a closed-loop control with a distributed wireless in-fiel...

  15. Evaluation of water-effect ratio methodology for establishing site-specific water quality criteria

    SciTech Connect

    Welsh, P.G.; Lipton, J.; Chapman, G.A.

    2000-06-01

    One approach outlined by the US Environmental Protection Agency (US EPA) for derivation of site-specific water quality criteria for metals in natural surface waters involves the development of water-effect ratios (WERs). This approach entails multiplying national water quality criteria by an experimentally derived WER, where the WER is defined as the ratio of the toxicity of the metal in the site water to the toxicity of the same metal in standard laboratory water. The authors discuss technical issues associated with test methods described in the US EPA WER guidance documents that may lead to inappropriate WERs. Critical issues include accounting for differences in calcium and magnesium concentrations (Ca:Mg ratios), alkalinity, and pH between site and laboratory waters; ensuring appropriate fish acclimation; and accounting for interspecies variability, multiple metals interactions, end-point variability, and temporal and spatial variability in the derivation of the WER. Failure to address these issues may have the unintended effect of deriving site-specific water quality criteria that are underprotective of aquatic life. The authors recommend that WER testing and future regulatory guidance for derivation of site-specific water quality criteria incorporate consideration of these potential confounding variables so that site-specific criteria can be established with greater confidence.

  16. 78 FR 75552 - Environmental Management Site-Specific Advisory Board, Northern New Mexico

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-12

    ...This notice announces a combined meeting of the Environmental Monitoring and Remediation Committee and Waste Management Committee of the Environmental Management Site-Specific Advisory Board (EM SSAB), Northern New Mexico (known locally as the Northern New Mexico Citizens' Advisory Board [NNMCAB]). The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770) requires that public notice of......

  17. The Bxb1 recombination system demonstrates heritable transmission of site-specific excision in Arabidopsis

    USDA-ARS?s Scientific Manuscript database

    Background: The mycobacteriophage large serine recombinase Bxb1 catalyzes site-specific recombination between its corresponding attP and attB recognition sites. Previously, we and others have shown that Bxb1 has catalytic activity in various eukaryotic species including Nicotiana tabacum, Schizosacc...

  18. 40 CFR 170.232 - Knowledge of labeling and site-specific information.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... requirements related to safe use of the pesticide, such as signal words, human hazard precautions, personal... (CONTINUED) PESTICIDE PROGRAMS WORKER PROTECTION STANDARD Standard for Pesticide Handlers § 170.232 Knowledge... of site-specific information. Whenever a handler who is employed by a commercial pesticide handling...

  19. Integration of aerial imaging and variable-rate technology for site-specific aerial herbicide application

    USDA-ARS?s Scientific Manuscript database

    As remote sensing and variable rate technology are becoming more available for aerial applicators, practical methodologies on effective integration of these technologies are needed for site-specific aerial applications of crop production and protection materials. The objectives of this study were to...

  20. 77 FR 16021 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-03-19

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770... Street, Pahrump, Nevada 89048. FOR FURTHER INFORMATION CONTACT: Denise Rupp, Board Administrator,...

  1. 75 FR 6370 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-02-09

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada Test Site. The Federal Advisory Committee Act (Pub. L. No. 92-463, 86 Stat..., Nevada 89131. FOR FURTHER INFORMATION CONTACT: Denise Rupp, Board Administrator, 232 Energy Way, M/S...

  2. 77 FR 39234 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-02

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770..., 2012, 5 p.m. ADDRESSES: Bob Ruud Community Center, 150 North Highway 160, Pahrump, Nevada 89060....

  3. 76 FR 12349 - Environmental Management Site-Specific Advisory Board, Nevada; Correction

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-07

    ... Environmental Management Site-Specific Advisory Board, Nevada; Correction AGENCY: Department of Energy. ACTION..., Nevada to be held on March 9, 2011 (76 FR 10343). This document makes several corrections to that notice... Vegas, Nevada 89030. Phone: (702) 657- 9088; Fax (702) 295-5300 or E-mail:...

  4. 76 FR 59393 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-26

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770... 12, 2011, 5 p.m. ADDRESSES: Las Vegas Country Club, 3000 Joe W. Brown Boulevard, Las Vegas,...

  5. 78 FR 46330 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-31

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ] ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770... 21, 2013 5:00 p.m. ADDRESSES: Bob Ruud Community Center, 150 N. Highway 160, Pahrump, Nevada...

  6. 77 FR 12044 - Environmental Management Site-Specific Advisory Board, Nevada

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-28

    ... Environmental Management Site-Specific Advisory Board, Nevada AGENCY: Department of Energy. ACTION: Notice of... Advisory Board (EM SSAB), Nevada. The Federal Advisory Committee Act (Pub. L. 92-463, 86 Stat. 770... Vegas, Nevada 89135. FOR FURTHER INFORMATION CONTACT: Denise Rupp, Board Administrator, 232 Energy...

  7. Insect pest densities across site-specific management zones of irrigated corn in northeastern Colorado.

    PubMed

    Davidson, Silas A; Peairs, Frank B; Khosla, Rajiv

    2007-06-01

    The ability to manage insect pests in a site-specific manner is hindered by the costs and time required to describe pest densities and distributions. The purpose of this study was to determine whether insect pest distributions are related to site-specific management zones (SSMZs). Site-specific management zones, as described in this study, delineate fields into three zones of similar yield potential: high, medium, and low productivity. If insect densities vary across SSMZs, it is possible that management decisions could be made at the SSMZ level instead of treating the whole field. This research was conducted during summers 2001 and 2002 on cooperators' farms in northeastern Colorado. Surveys were conducted within corn, Zea mays L., fields, so that densities of three common insect pests of Colorado corn could be compared across SSMZ. The three insect pests were western corn rootworm, Diabrotica virgifera virgifera LeConte; European corn borer, Ostrinia nubilalis (HiAbner); and western bean cutworm, Richia albicosta (Smith). D. v. virgifera larvae and adults were most common in the high-productivity SSMZ. O. nubilalis larval abundance was similar at three fields, whereas in a fourth field the larvae were most common in the high-productivity SSMZ. In one field that contained substantial numbers of R. albicosta, egg abundance was similar across SSMZs, whereas larvae were most common in the high-productivity SSMZ. Site-specific management zones seemed to correlate well with the abundance of some insect pests and might prove useful for managing insects in a site-specific manner.

  8. Site-Specifically Labeled Immunoconjugates for Molecular Imaging--Part 1: Cysteine Residues and Glycans.

    PubMed

    Adumeau, Pierre; Sharma, Sai Kiran; Brent, Colleen; Zeglis, Brian M

    2016-02-01

    Due to their remarkable selectivity and specificity for cancer biomarkers, immunoconjugates have emerged as extremely promising vectors for the delivery of diagnostic radioisotopes and fluorophores to malignant tissues. Paradoxically, however, these tools for precision medicine are synthesized in a remarkably imprecise way. Indeed, the vast majority of immunoconjugates are created via the random conjugation of bifunctional probes (e.g., DOTA-NCS) to amino acids within the antibody (e.g., lysines). Yet antibodies have multiple copies of these residues throughout their macromolecular structure, making control over the location of the conjugation reaction impossible. This lack of site specificity can lead to the formation of poorly defined, heterogeneous immunoconjugates with suboptimal in vivo behavior. Over the past decade, interest in the synthesis and development of site-specifically labeled immunoconjugates--both antibody-drug conjugates as well as constructs for in vivo imaging--has increased dramatically, and a number of reports have suggested that these better defined, more homogeneous constructs exhibit improved performance in vivo compared to their randomly modified cousins. In this two-part review, we seek to provide an overview of the various methods that have been developed to create site-specifically modified immunoconjugates for positron emission tomography, single photon emission computed tomography, and fluorescence imaging. We will begin with an introduction to the structure of antibodies and antibody fragments. This is followed by the core of the work: sections detailing the four different approaches to site-specific modification strategies based on cysteine residues, glycans, peptide tags, and unnatural amino acids. These discussions will be divided into two installments: cysteine residues and glycans will be detailed in Part 1 of the review, while peptide tags and unnatural amino acids will be addressed in Part 2. Ultimately, we sincerely hope

  9. An ultrasensitive photoelectrochemical nucleic acid biosensor

    PubMed Central

    Gao, Zhiqiang; Tansil, Natalia C.

    2005-01-01

    A simple and ultrasensitive procedure for non-labeling detection of nucleic acids is described in this study. It is based on the photoelectrochemical detection of target nucleic acids by forming a nucleic acid/photoreporter adduct layer on an ITO electrode. The target nucleic acids were hybridized with immobilized oligonucleotide capture probes on the ITO electrode. A subsequent binding of a photoreporter—a photoactive threading bis-intercalator consisting of two N,N′-bis(3-propyl-imidazole)-1,4,5,8-naphthalene diimides (PIND) linked by a Ru(bpy)22+ (bpy = 2,2′-bipyridine) complex (PIND–Ru–PIND)—allowed for photoelectrochemical detection of the target nucleic acids. The extremely low dissociation rate of the adduct and the highly reversible photoelectrochemical response under visible light illumination (490 nm) make it possible to conduct nucleic acid detection, with a sensitivity enhancement of four orders of magnitude over voltammetry. These results demonstrate for the first time the potential of photoelectrochemical biosensors for PCR-free ultrasensitive detection of nucleic acids. PMID:16061935

  10. Luminescent Quantum Dots as Ultrasensitive Biological Labels

    NASA Astrophysics Data System (ADS)

    Nie, Shuming

    2000-03-01

    Highly luminescent semiconductor quantum dots have been covalently coupled to biological molecules for use in ultrasensitive biological detection. This new class of luminescent labels is considerably brighter and more resistant againt photobleaching in comparison with organic dyes. Quantum dots labeled with the protein transferrin undergo receptor-mediated endocytosis (RME) in cultured HeLa cells, and those dots that were conjugated to immunomolecules recognize specific antibodies or antigens. In addition, we show that DNA functionalized quantum dots can be used to target specific genes by hybridization. We expect that quantum dot bioconjugates will have a broad range of biological applications, such as ligand-receptor interactions, real-time monitoring of molecular trafficking inside living cells, multicolor fluorescence in-situ hybridization (FISH), high-sensitivity detection in miniaturized devices (e.g., DNA chips), and fluorescent tagging of combinatorial chemical libraries. A potential clinical application is the use of quantum dots for ultrasensitive viral RNA detection, in which as low as 100 copies of hepatitis C and HIV viruses per ml blood should be detected.

  11. Applications of stable, nonradioactive isotope tracers in in vivo human metabolic research.

    PubMed

    Kim, Il-Young; Suh, Sang-Hoon; Lee, In-Kyu; Wolfe, Robert R

    2016-01-15

    The human body is in a constant state of turnover, that is, being synthesized, broken down and/or converted to different compounds. The dynamic nature of in vivo kinetics of human metabolism at rest and in stressed conditions such as exercise and pathophysiological conditions such as diabetes and cancer can be quantitatively assessed with stable, nonradioactive isotope tracers in conjunction with gas or liquid chromatography mass spectrometry and modeling. Although measurements of metabolite concentrations have been useful as general indicators of one's health status, critical information on in vivo kinetics of metabolites such as rates of production, appearance or disappearance of metabolites are not provided. Over the past decades, stable, nonradioactive isotope tracers have been used to provide information on dynamics of specific metabolites. Stable isotope tracers can be used in conjunction with molecular and cellular biology tools, thereby providing an in-depth dynamic assessment of metabolic changes, as well as simultaneous investigation of the molecular basis for the observed kinetic responses. In this review, we will introduce basic principles of stable isotope methodology for tracing in vivo kinetics of human or animal metabolism with examples of quantifying certain aspects of in vivo kinetics of carbohydrate, lipid and protein metabolism.

  12. Oak Ridge National Laboratory program plan for certification of nonradioactive hazardous waste

    SciTech Connect

    1996-05-01

    This document describes Oak Ridge National Laboratory`s (ORNL) Program for Certification of Nonradioactive Hazardous Waste (Program). The Program establishes the criteria that will be used by all ORNL divisions, offices, and programs for unrestricted release of hazardous waste to off-site commercial facilities. The certification process meets the requirements given in the Performance Objective for Certification of Non-Radioactive Hazardous Waste. The Program Plan has two main elements: (A) Establishing Radioactive Materials Management Areas (RMMAs). At ORNL, RMMAs are (1) Contamination Areas, High Contamination Areas, and Airborne Radioactivity Areas, (2) Radiological Buffer Areas established for contamination control, and (3) areas posted to prevent loss of control of activated items. (B) Certifying that hazardous waste originating in an RMMA is suitable for commercial treatment, storage, or disposal by process knowledge, surface contamination surveys, sampling and analysis, or a combination of these techniques. If process knowledge is insufficient, the hazardous waste must undergo sampling and analysis in addition to surface contamination surveys. This Program will reduce the impact to current ORNL operations by using current radiological area boundaries and existing plans and procedures to the greatest extent possible. New or revised procedures will be developed as necessary to implement this Program.

  13. Applications of stable, nonradioactive isotope tracers in in vivo human metabolic research

    PubMed Central

    Kim, Il-Young; Suh, Sang-Hoon; Lee, In-Kyu; Wolfe, Robert R

    2016-01-01

    The human body is in a constant state of turnover, that is, being synthesized, broken down and/or converted to different compounds. The dynamic nature of in vivo kinetics of human metabolism at rest and in stressed conditions such as exercise and pathophysiological conditions such as diabetes and cancer can be quantitatively assessed with stable, nonradioactive isotope tracers in conjunction with gas or liquid chromatography mass spectrometry and modeling. Although measurements of metabolite concentrations have been useful as general indicators of one's health status, critical information on in vivo kinetics of metabolites such as rates of production, appearance or disappearance of metabolites are not provided. Over the past decades, stable, nonradioactive isotope tracers have been used to provide information on dynamics of specific metabolites. Stable isotope tracers can be used in conjunction with molecular and cellular biology tools, thereby providing an in-depth dynamic assessment of metabolic changes, as well as simultaneous investigation of the molecular basis for the observed kinetic responses. In this review, we will introduce basic principles of stable isotope methodology for tracing in vivo kinetics of human or animal metabolism with examples of quantifying certain aspects of in vivo kinetics of carbohydrate, lipid and protein metabolism. PMID:26795236

  14. Microbial challenge tests on nonradioactive TiO2-based 68Ge/68Ga generator columns.

    PubMed

    Petrik, Milos; Schuessele, Andrea; Perkhofer, Susanne; Lass-Flörl, Cornelia; Becker, Dirk; Decristoforo, Clemens

    2012-08-01

    The aim of this study was to test the survival of microorganisms in eluents used for (68)Ge/(68)Ga generators and for regeneration of nonradioactive (68)Ge/(68)Ga generator columns after high microbial load. Nonradioactive generator columns were loaded with various microorganisms and tested to determine whether the microorganisms were proliferating or surviving in eluates of the columns. TiO(2) columns used in a commercially available (68)Ge/(68)Ga generators for absorption of (68)Ge were loaded with greater than 10(7) colony-forming units (CFU) of Staphylococcus aureus, Clostridium sporogenes, Candida albicans, Helicobacter pylori, Aspergillus niger and Deinococcus radiodurans. Columns were eluted with 0.1 N HCl under standard eluting conditions. Elutions, over a prolonged time period, were tested for endotoxins, sterility and number of CFU. Initial tests on the eluent (0.1 N HCl) already showed limited survival of microorganisms. Using TiO(2) generator columns, even with a higher load of microorganisms no survival of any microorganism could be detected in subsequent elutions, neither in colony-forming unit assays nor in sterility testing. In addition, endotoxin tests showed no elevated levels in any of the eluent samples, even days after incubation. The conditions used in this type of (68)Ge/(68)Ga generator are highly unfavourable for survival or growth of microorganisms. The risks associated with incidental microbial contaminations during the lifetime of such a generator are therefore very low.

  15. Ultra-sensitive transducer advances micro-measurement range

    NASA Technical Reports Server (NTRS)

    Rogallo, V. L.

    1964-01-01

    An ultrasensitive piezoelectric transducer, that converts minute mechanical forces into electrical impulses, measures the impact of micrometeoroids against space vehicles. It has uniform sensitivity over the entire target area and a high degree of stability.

  16. Ultrasensitivity and sharp threshold theorems for multisite systems

    NASA Astrophysics Data System (ADS)

    Dougoud, M.; Mazza, C.; Vinckenbosch, L.

    2017-02-01

    This work studies the ultrasensitivity of multisite binding processes where ligand molecules can bind to several binding sites. It considers more particularly recent models involving complex chemical reactions in allosteric phosphorylation processes and for transcription factors and nucleosomes competing for binding on DNA. New statistics-based formulas for the Hill coefficient and the effective Hill coefficient are provided and necessary conditions for a system to be ultrasensitive are exhibited. It is first shown that the ultrasensitivity of binding processes can be approached using sharp-threshold theorems which have been developed in applied probability theory and statistical mechanics for studying sharp threshold phenomena in reliability theory, random graph theory and percolation theory. Special classes of binding process are then introduced and are described as density dependent birth and death process. New precise large deviation results for the steady state distribution of the process are obtained, which permits to show that switch-like ultrasensitive responses are strongly related to the multi-modality of the steady state distribution. Ultrasensitivity occurs if and only if the entropy of the dynamical system has more than one global minimum for some critical ligand concentration. In this case, the Hill coefficient is proportional to the number of binding sites, and the system is highly ultrasensitive. The classical effective Hill coefficient I is extended to a new cooperativity index I q , for which we recommend the computation of a broad range of values of q instead of just the standard one I  =  I 0.9 corresponding to the 10%-90% variation in the dose-response. It is shown that this single choice can sometimes mislead the conclusion by not detecting ultrasensitivity. This new approach allows a better understanding of multisite ultrasensitive systems and provides new tools for the design of such systems.

  17. Site-specific analysis of heteronuclear Overhauser effects in microcrystalline proteins.

    PubMed

    del Amo, Juan Miguel Lopez; Agarwal, Vipin; Sarkar, Riddhiman; Porter, Justin; Asami, Sam; Rübbelke, Martin; Fink, Uwe; Xue, Yi; Lange, Oliver F; Reif, Bernd

    2014-08-01

    Relaxation parameters such as longitudinal relaxation are susceptible to artifacts such as spin diffusion, and can be affected by paramagnetic impurities as e.g. oxygen, which make a quantitative interpretation difficult. We present here the site-specific measurement of [(1)H](13)C and [(1)H](15)N heteronuclear rates in an immobilized protein. For methyls, a strong effect is expected due to the three-fold rotation of the methyl group. Quantification of the [(1)H](13)C heteronuclear NOE in combination with (13)C-R 1 can yield a more accurate analysis of side chain motional parameters. The observation of significant [(1)H](15)N heteronuclear NOEs for certain backbone amides, as well as for specific asparagine/glutamine sidechain amides is consistent with MD simulations. The measurement of site-specific heteronuclear NOEs is enabled by the use of highly deuterated microcrystalline protein samples in which spin diffusion is reduced in comparison to protonated samples.

  18. Recent Developments in the Site-Specific Immobilization of Proteins onto Solid Supports

    SciTech Connect

    Camarero, J A

    2007-02-21

    Immobilization of proteins onto surfaces is of great importance in numerous applications, including protein analysis, drug screening, and medical diagnostics, among others. The success of all these technologies relies on the immobilization technique employed to attach a protein to the corresponding surface. Non-specific physical adsorption or chemical cross-linking with appropriate surfaces results in the immobilization of the protein in random orientations. Site-specific covalent attachment, on the other hand, leads to molecules being arranged in a definite, orderly fashion and allows the use of spacers and linkers to help minimize steric hindrances between the protein and the surface. The present work reviews the latest chemical and biochemical developments for the site-specific covalent attachment of proteins onto solid supports.

  19. Site-Specific Integration of Exogenous Genes Using Genome Editing Technologies in Zebrafish.

    PubMed

    Kawahara, Atsuo; Hisano, Yu; Ota, Satoshi; Taimatsu, Kiyohito

    2016-05-13

    The zebrafish (Danio rerio) is an ideal vertebrate model to investigate the developmental molecular mechanism of organogenesis and regeneration. Recent innovation in genome editing technologies, such as zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) system, have allowed researchers to generate diverse genomic modifications in whole animals and in cultured cells. The CRISPR/Cas9 and TALEN techniques frequently induce DNA double-strand breaks (DSBs) at the targeted gene, resulting in frameshift-mediated gene disruption. As a useful application of genome editing technology, several groups have recently reported efficient site-specific integration of exogenous genes into targeted genomic loci. In this review, we provide an overview of TALEN- and CRISPR/Cas9-mediated site-specific integration of exogenous genes in zebrafish.

  20. Engineering Bacillus thuringiensis bioinsecticides with an indigenous site-specific recombination system.

    PubMed Central

    Baum, J A; Kakefuda, M; Gawron-Burke, C

    1996-01-01

    The cry genes of Bacillus thuringiensis encode a diverse group of crystal-forming proteins that exhibit insecticidal activity, particularly against the larvae of lepidopteran, coleopteran, and dipteran insects. The efficacy of B. thuringiensis-based biopesticides may be improved through the genetic manipulation of these genes. A gene transfer system has been developed for the introduction and maintenance of cloned insecticidal cry genes on small plasmids in B. thuringiensis. This vector system combines a B. thuringiensis plasmid replicon and an indigenous site-specific recombination system that allows for the selective removal of ancillary or foreign DNA from the recombinant bacterium after introduction of the Cry-encoding plasmid. The site-specific recombination system is useful for engineering strains with unique combinations of cry genes, resulting in new active ingredients with improved insecticidal properties. PMID:8953709

  1. Engineering Bacillus thuringiensis bioinsecticides with an indigenous site-specific recombination system.

    PubMed

    Baum, J A; Kakefuda, M; Gawron-Burke, C

    1996-12-01

    The cry genes of Bacillus thuringiensis encode a diverse group of crystal-forming proteins that exhibit insecticidal activity, particularly against the larvae of lepidopteran, coleopteran, and dipteran insects. The efficacy of B. thuringiensis-based biopesticides may be improved through the genetic manipulation of these genes. A gene transfer system has been developed for the introduction and maintenance of cloned insecticidal cry genes on small plasmids in B. thuringiensis. This vector system combines a B. thuringiensis plasmid replicon and an indigenous site-specific recombination system that allows for the selective removal of ancillary or foreign DNA from the recombinant bacterium after introduction of the Cry-encoding plasmid. The site-specific recombination system is useful for engineering strains with unique combinations of cry genes, resulting in new active ingredients with improved insecticidal properties.

  2. Site-specifically modified oligodeoxyribonucleotides as templates for Escherichia coli DNA polymerase I

    SciTech Connect

    O'Connor, D.; Stoehrer, G.

    1985-04-01

    Oligodeoxyribonucleotides with site-specific modifications have been used as substrates for Escherichia coli DNA polymerase I holoenzyme and Klenow fragment. Modifications included the bulky guanine-8-aminofluorene adduct and a guanine oxidation product resembling the product of photosensitized DNA oxidation. By a combination of primers and nick-mers, conditions of single-strand-directed DNA synthesis and nick-translation could be created. The results show that the polymerase can bypass both types of lesions. Bypass occurs on a single-stranded template but is facilitated on a nicked, double-stranded template. Only purines, with guanine more favored than adenine, are incorporated across both lesions. The results indicate that site-specifically modified oligonucleotides can be sensitive probes for the action of polymerases on damaged templates. They also suggest a function for polymerase I, in its nick-translation capacity, during DNA repair and mutagenesis.

  3. 2-Cyanobenzothiazole (CBT) condensation for site-specific labeling of proteins at the terminal cysteine residues.

    PubMed

    Cui, Lina; Rao, Jianghong

    2015-01-01

    Site specificity is pivotal in obtaining homogeneously labeled proteins without batch-to-batch variations. More importantly, precisely controlled modification at specific sites avoids potential pitfalls that could otherwise interfere with protein folding, structure, and function. Inspired by the chemical synthesis of D-luciferin, we have developed an efficient strategy (second-order rate constant k 2 = 9.2 M(-1) s(-1)) for labeling of proteins containing 1,2-aminothiol via reaction with 2-cyanobenzothiazole (CBT). In addition, the CBT condensation enjoys the convenience of protein engineering, as production of N-terminal cysteine-containing proteins has been well developed for native chemical ligation. This protocol describes the preparation of Renilla luciferase (rLuc) with 1,2-aminothiol at either its N- or C-terminus, and site-specific labeling of rLuc with fluorescein or (18)F via CBT condensation.

  4. Site-Specific Integration of Exogenous Genes Using Genome Editing Technologies in Zebrafish

    PubMed Central

    Kawahara, Atsuo; Hisano, Yu; Ota, Satoshi; Taimatsu, Kiyohito

    2016-01-01

    The zebrafish (Danio rerio) is an ideal vertebrate model to investigate the developmental molecular mechanism of organogenesis and regeneration. Recent innovation in genome editing technologies, such as zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) system, have allowed researchers to generate diverse genomic modifications in whole animals and in cultured cells. The CRISPR/Cas9 and TALEN techniques frequently induce DNA double-strand breaks (DSBs) at the targeted gene, resulting in frameshift-mediated gene disruption. As a useful application of genome editing technology, several groups have recently reported efficient site-specific integration of exogenous genes into targeted genomic loci. In this review, we provide an overview of TALEN- and CRISPR/Cas9-mediated site-specific integration of exogenous genes in zebrafish. PMID:27187373

  5. Site-specific seismic-hazard analysis that is completely probabilistic

    USGS Publications Warehouse

    Cramer, C.H.

    2003-01-01

    When a site-specific probabilistic ground-motion estimate is required, the full site-amplification distribution should be used instead of a single deterministic median value. A probabilistic methodology using site-amplification distributions to modify rock ground-motion attenuation relations into site-specific relations prior to calculating seismic hazard has been developed and applied at two selected sites in the central United States: Memphis, Tennessee, and Paducah, Kentucky. The use of a completely probabilistic approach can make about a 10% difference in ground-motion estimates over simply multiplying a bedrock probabilistic ground motion by a median site-amplification factor at a 1 in 2475 annual probability of exceedance and even larger differences at smaller probabilites of exceedance. The value of this approach is that a probabilistic answer incorporating the uncertainty in our knowledge of site amplification of ground motions can be calculated.

  6. Recent Advances in Site Specific Conjugations of Antibody Drug Conjugates (ADCs).

    PubMed

    Gao, Wenlong; Zhang, Jingxin; Xiang, Jun; Zhang, Lei; Wu, Chengbin; Dhal, Pradeep K; Chen, Bo

    2016-01-01

    Antibody-drug conjugates (ADCs) take the advantage of antigen specificity of monoclonal antibodies to deliver highly potent cytotoxic drugs selectively to antigen-expressing tumor cells. The recent approval of Adcetris™ and Kadcyla™ as well as emerging data from numerous ongoing clinical trials underscore the role of ADCs as a new therapeutic option for cancer patients. However, conventional conjugation methods generally result in a heterogeneous mixture of ADCs, which can result in significant therapeutic liabilities and can lead to complicated manufacturing processes. The increased understanding from the clinical investigation of current ADCs and site-specific bioconjugation technologies has enabled scientists to accelerate the discovery and development of the next generation ADCs with defined and homogeneous composition. The present manuscript reviews the recent advances and trends in the research and development of novel ADCs obtained by site-specific conjugation method.

  7. Generating site-specifically modified proteins via a versatile and stable nucleophilic carbon ligation.

    PubMed

    Kudirka, Romas; Barfield, Robyn M; McFarland, Jesse; Albers, Aaron E; de Hart, Gregory W; Drake, Penelope M; Holder, Patrick G; Banas, Stefanie; Jones, Lesley C; Garofalo, Albert W; Rabuka, David

    2015-02-19

    There is a need for facile chemistries that allow for chemo- and regioselectivity in bioconjugation reactions. To address this need, we are pioneering site-specific bioconjugation methods that use formylglycine as a bioorthogonal handle on a protein surface. Here we introduce aldehyde-specific bioconjugation chemistry, the trapped-Knoevenagel ligation. The speed and stability of the trapped-Knoevenagel ligation further advances the repertoire of aldehyde-based bioconjugations and expands the toolbox for site-specific protein modifications. The trapped-Knoevenagel ligation reaction can be run at near neutral pH in the absence of catalysts to produce conjugates that are stable under physiological conditions. Using this new ligation, we generated an antibody-drug conjugate that demonstrates excellent efficacy in vitro and in vivo.

  8. Comparative fitness assessment of Anopheles stephensi transgenic lines receptive to site-specific integration.

    PubMed

    Amenya, D A; Bonizzoni, M; Isaacs, A T; Jasinskiene, N; Chen, H; Marinotti, O; Yan, G; James, A A

    2010-04-01

    Genetically modified mosquitoes that are unable to transmit pathogens offer opportunities for controlling vector-borne diseases such as malaria and dengue. Site-specific gene recombination technologies are advantageous in the development of these insects because antipathogen effector genes can be inserted at integration sites in the genome that cause the least alteration in mosquito fitness. Here we describe Anopheles stephensi transgenic lines containing phi C31 attP'docking' sites linked to a fluorescent marker gene. Chromosomal insertion sites were determined and life-table parameters were assessed for transgenic mosquitoes of each line. No significant differences in fitness between the transgenic and nontransgenic mosquitoes were detected in this study. These transgenic lines are suitable for future site-specific integrations of antiparasite transgenes into the attP sites.

  9. Site-Specific Earthquake Response Analysis for Portsmouth Gaseous Diffusion Plant, Portsmouth, Ohio

    DTIC Science & Technology

    1993-08-01

    PAPER Miscellaneous Paper GL-93-13 August 1993 Site-Specific Earthquke Response Analysis for Portsmouth Gaseous Diffusion Plant, Portsmouth, Ohio by David...been predicted . The use of other computer models may be necessary to define peak response values. 68. Six different figures and various tables are used...NJ, pg 85. Zen, K. and Higuchi, Y. 1984. " Prediction of Vibratory Shear Modulus and Damping Ratio for Cohesive Soils, Proc.. Eighth Int’l Conference

  10. Site-Specific Differentiation of Fibroblasts in Normal and Scleroderma Skin

    DTIC Science & Technology

    2010-06-01

    expression of one HOX lncRNA, termed nc-HOXC10, is correlated with fibrotic gene expression in fibroblasts. In our initial study, we found nc-HOXC10...anatomic expression pattern of Hox genes from embryonic development through adulthood. The ongoing Hox expression endowed fibroblasts with site...specific inductive activities that control the homeostasis and regeneration of epithelia throughout the body. The epigenetic memory of Hox genes depend on

  11. Use of a water effect ratio to develop an estuarine copper site-specific standard

    SciTech Connect

    Brosnan, T.

    1995-12-31

    Development of a copper site-specific standard in New York Harbor involved several steps: (1) EPA`s Indicator Species Procedure was used to develop a Water Effect Ratio (WER), which produces a biologically-based adjustment to the existing criteria. Samples from seven stations were collected during high and low river-flow conditions in 1992--93. Toxicity testing included embryo-larval development tests on two bivalves, an urchin, and a shrimp. A red algae was used for a sexual reproduction test. This testing yielded WER`s of 1.33 2.1 7, with an average of 1.49. When multiplied by the existing EPA criteria of 2.9 ug/L (total recoverable), this yielded a preliminary acute site specific standard of 4.3 ug/L (total recoverable); (2) Further analysis of these data, combined with a literature search of similar data, resulted in a recalculation of the original national acute criteria value, from 2.9 ug/L (total recoverable) to 5.3 ug/L (dissolved). Multiplying the WER by the recalculated national criteria yielded a final acute site, specific criteria of 7.9 ug/L (dissolved); (3) Finally, EPA`s original criteria indicated that achieving the acute criterion would be protective of chronic effects (i.e. the acute:chronic ratio (ACR) was 2). However, EPA, recently decided that an ACR of 2 was no longer valid, and decided to use the freshwater ACR of 2.8. This resulted in a recalculation of the national chronic criteria to 3.75 ug/L (dissolved). Combined with the WER of 1.49, this yielded a final chronic site specific standard of 5.6 ug/L. The new standard eliminated ambient violations in most of the harbor, and removed the need for wasteload allocations at 12 of NYC`s 14 sewage treatment plants.

  12. Site-specific cancer risk in the Baltic cohort of Chernobyl cleanup workers, 1986-2007.

    PubMed

    Rahu, Kaja; Hakulinen, Timo; Smailyte, Giedre; Stengrevics, Aivars; Auvinen, Anssi; Inskip, Peter D; Boice, John D; Rahu, Mati

    2013-09-01

    To assess site-specific cancer risk in the Baltic cohort of Chernobyl cleanup workers, 1986-2007. The Baltic cohort includes 17,040 men from Estonia, Latvia and Lithuania who participated in the environmental cleanup after the accident at the Chernobyl Nuclear Power Station in 1986-1991 and who were followed up for cancer incidence until the end of 2007. Cancer cases diagnosed in the cohort and in the male population of each country were identified from the respective national cancer registers. The proportional incidence ratio (PIR) with 95% confidence interval (CI) was used to estimate the site-specific cancer risk in the cohort. For comparison and as it was possible, the site-specific standardised incidence ratio (SIR) was calculated for the Estonian sub-cohort, which was not feasible for the other countries. Overall, 756 cancer cases were reported during 1986-2007. A higher proportion of thyroid cancers in relation to the male population was found (PIR=2.76; 95%CI 1.63-4.36), especially among those who started their mission shortly after the accident, in April-May 1986 (PIR=6.38; 95%CI 2.34-13.89). Also, an excess of oesophageal cancers was noted (PIR=1.52; 95% CI 1.06-2.11). No increased PIRs for leukaemia or radiation-related cancer sites combined were observed. PIRs and SIRs for the Estonian sub-cohort demonstrated the same site-specific cancer risk pattern. Consistent evidence of an increase in radiation-related cancers in the Baltic cohort was not observed with the possible exception of thyroid cancer, where conclusions are hampered by known medical examination including thyroid screening among cleanup workers. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Site-specific cancer risk in the Baltic cohort of Chernobyl cleanup workers, 1986–2007

    PubMed Central

    Rahu, Kaja; Hakulinen, Timo; Smailyte, Giedre; Stengrevics, Aivars; Auvinen, Anssi; Inskip, Peter D.; Boice, John D.; Rahu, Mati

    2013-01-01

    Objective To assess site-specific cancer risk in the Baltic cohort of Chernobyl cleanup workers 1986–2007. Methods The Baltic cohort includes 17,040 men from Estonia, Latvia and Lithuania who participated in the environmental cleanup after the accident at the Chernobyl Nuclear Power Station in 1986–1991, and who were followed for cancer incidence until the end of 2007. Cancer cases diagnosed in the cohort and in the male population of each country were identified from the respective national cancer registers. The proportional incidence ratio (PIR) with 95% confidence interval (CI) was used to estimate the site-specific cancer risk in the cohort. For comparison and as it was possible, the site-specific standardized incidence ratio (SIR) was calculated for the Estonian sub-cohort, which was not feasible for the other countries. Results Overall, 756 cancer cases were reported during 1986–2007. A higher proportion of thyroid cancers in relation to the male population was found (PIR=2.76; 95%CI 1.63–4.36), especially among those who started their mission shortly after the accident, in April–May 1986 (PIR=6.38; 95% CI 2.34–13.89). Also, an excess of oesophageal cancers was noted (PIR=1.52; 95% CI 1.06–2.11). No increased PIRs for leukaemia or radiation-related cancer sites combined were observed. PIRs and SIRs for the Estonian sub-cohort demonstrated the same site-specific cancer risk pattern. Conclusion Consistent evidence of an increase in radiation-related cancers in the Baltic cohort was not observed with the possible exception of thyroid cancer, where conclusions are hampered by known medical examination including thyroid screening among cleanup workers. PMID:23683549

  14. Locked by Design: A Conformationally Constrained Transglutaminase Tag Enables Efficient Site-Specific Conjugation.

    PubMed

    Siegmund, Vanessa; Schmelz, Stefan; Dickgiesser, Stephan; Beck, Jan; Ebenig, Aileen; Fittler, Heiko; Frauendorf, Holm; Piater, Birgit; Betz, Ulrich A K; Avrutina, Olga; Scrima, Andrea; Fuchsbauer, Hans-Lothar; Kolmar, Harald

    2015-11-02

    Based on the crystal structure of a natural protein substrate for microbial transglutaminase, an enzyme that catalyzes protein crosslinking, a recognition motif for site-specific conjugation was rationally designed. Conformationally locked by an intramolecular disulfide bond, this structural mimic of a native conjugation site ensured efficient conjugation of a reporter cargo to the therapeutic monoclonal antibody cetuximab without erosion of its binding properties. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Site-specific functionalization of proteins and their applications to therapeutic antibodies

    PubMed Central

    van Vught, Remko; Pieters, Roland J; Breukink, Eefjan

    2014-01-01

    Protein modifications are often required to study structure and function relationships. Instead of the random labeling of lysine residues, methods have been developed to (sequence) specific label proteins. Next to chemical modifications, tools to integrate new chemical groups for bioorthogonal reactions have been applied. Alternatively, proteins can also be selectively modified by enzymes. Herein we review the methods available for site-specific modification of proteins and their applications for therapeutic antibodies. PMID:24757499

  16. Site-specific fatty acid-conjugation to prolong protein half-life in vivo

    PubMed Central

    Lim, Sung In; Mizuta, Yukina; Takasu, Akinori; Hahn, Young S.; Kim, Yong Hwan; Kwon, Inchan

    2015-01-01

    Therapeutic proteins are indispensable in treating numerous human diseases. However, therapeutic proteins often suffer short serum half-life. In order to extend the serum half-life, a natural albumin ligand (a fatty acid) has been conjugated to small therapeutic peptides resulting in a prolonged serum half-life via binding to patients' serum albumin in vivo. However, fatty acid-conjugation has limited applicability due to lack of site-specificity resulting in the heterogeneity of conjugated proteins and a significant loss in pharmaceutical activity. In order to address these issues, we exploited the site-specific fatty acid-conjugation to a permissive site of a protein, using copper-catalyzed alkyne-azide cycloaddition, by linking a fatty acid derivative to p-ethynylphenylalanine incorporated into a protein using an engineered pair of yeast tRNA/aminoacyl tRNA synthetase. As a proof-of-concept, we show that single palmitic acid conjugated to superfolder green fluorescent protein (sfGFP) in a site-specific manner enhanced a protein's albumin-binding in vitro about 20 times and the serum half-life in vivo 5 times when compared to those of the unmodified sfGFP. Furthermore, the fatty acid conjugation did not cause a significant reduction in the fluorescence of sfGFP. Therefore, these results clearly indicate that the site-specific fatty acid-conjugation is a very promising strategy to prolong protein serum half-life in vivo without compromising its folded structure and activity. PMID:23735573

  17. Efficient Preparation of Site-Specific Antibody-Drug Conjugates Using Cysteine Insertion.

    PubMed

    Dimasi, Nazzareno; Fleming, Ryan; Zhong, Haihong; Bezabeh, Binyam; Kinneer, Krista; Christie, Ronald J; Fazenbaker, Christine; Wu, Herren; Gao, Changshou

    2017-05-01

    Antibody-drug conjugates (ADCs) are a class of biopharmaceuticals that combine the specificity of antibodies with the high-potency of cytotoxic drugs. Engineering cysteine residues in the antibodies using mutagenesis is a common method to prepare site-specific ADCs. With this approach, solvent accessible amino acids in the antibody have been selected for substitution with cysteine for conjugating maleimide-bearing cytotoxic drugs, resulting in homogeneous and stable site-specific ADCs. Here we describe a cysteine engineering approach based on the insertion of cysteines before and after selected sites in the antibody, which can be used for site-specific preparation of ADCs. Cysteine-inserted antibodies have expression level and monomeric content similar to the native antibodies. Conjugation to a pyrrolobenzodiazepine dimer (SG3249) resulted in comparable efficiency of site-specific conjugation between cysteine-inserted and cysteine-substituted antibodies. Cysteine-inserted ADCs were shown to have biophysical properties, FcRn, and antigen binding affinity similar to the cysteine-substituted ADCs. These ADCs were comparable for serum stability to the ADCs prepared using cysteine-mutagenesis and had selective and potent cytotoxicity against human prostate cancer cells. Two of the cysteine-inserted variants abolish binding of the resulting ADCs to FcγRs in vitro, thereby potentially preventing non-target mediated uptake of the ADCs by cells of the innate immune system that express FcγRs, which may result in mitigating off-target toxicities. A selected cysteine-inserted ADC demonstrated potent dose-dependent anti-tumor activity in a xenograph tumor mouse model of human breast adenocarcinoma expressing the oncofetal antigen 5T4.

  18. Complete Genome Sequence of Streptomyces parvulus 2297, Integrating Site-Specifically with Actinophage R4

    PubMed Central

    Miura, Takamasa; Harada, Chizuko; Guo, Yong; Narisawa, Kazuhiko; Ohta, Hiroyuki; Takahashi, Hideo; Shirai, Makoto

    2016-01-01

    Streptomyces parvulus 2297, which is a host for site-specific recombination according to actinophage R4, is derived from the type strain ATCC 12434. Species of S. parvulus are known as producers of polypeptide antibiotic actinomycins and have been considered for industrial applications. We herein report for the first time the complete genome sequence of S. parvulus 2297. PMID:27563047

  19. Pattern development in cellular automata triggered by site-specific reactive processes

    NASA Astrophysics Data System (ADS)

    Abowd, Gregory D.; Garza-López, Roberto A.; Kozak, John J.

    1988-02-01

    The (discretized) evolution of regular and fractal patterns, initiated by a site-specific event, is studied via simulation on a simple model. Consistent with recent theories of diffusion on percolation networks, our results show that the development of fractal patterns is distinctly slower than for regular (euclidean) patterns. The possible relevance of our results to the evolution of symmetry-breaking instabilities is brought out.

  20. Intruder scenarios for site-specific low-level radioactive waste classification

    SciTech Connect

    Kennedy, W.E. Jr.; Peloquin, R.A.

    1988-09-01

    The US Department of Energy (DOE) has revised its low-level radioactive waste (LLW) management requirements and guidelines for waste generated at its facilities supporting defense missions. Specifically, draft DOE Order 5820.2A, Chapter 3 describes the purpose, policy, and requirements necessary for the management of defense LLW. The draft DOE policy calls for LLW operations to be managed to protect the health and safety of the public, preserve the environment, and ensure that no remedial action will be necessary after termination of operations. The basic approach used by DOE is to establish overall performance objectives, in terms of groundwater protection and public radiation dose limits, and to require site-specific performance assessments to determine compliance. As a result of these performance assessments, each site will develop waste acceptance criteria that define the allowable quantities and concentrations of specific radioisotopes. Additional limitations on waste disposal design, waste form, and waste treatment will also be developed on a site-specific basis. As a key step in the site-specific performance assessments, an evaluation must be conducted of potential radiation doses to intruders who may inadvertently move onto a closed DOE LLW disposal site after loss of institutional controls. This report (1) describes the types of intruder scenarios that should be considered when performing this step of the site-specific performance assessment, (2) provides the results of generic calculations performed using unit concentrations of various radionuclides as a comparison of the magnitude of importance of the various intruder scenarios, and (3) shows the relationship between the generic doses and waste classification limits for defense wastes.

  1. Site-specific investigations and preliminary design for the Gorleben repository

    SciTech Connect

    Brennecke, P.; Kranz, H.; Schneider, H. )

    1992-01-01

    In the Federal Republic of Germany, the disposal of radioactive waste is being planned for deep geologic formations only. A repository will be constructed in the Gorleben salt dome for all kinds of radioactive waste, in particular, heat-generating waste from reprocessing and spent fuel. Pusuant to German safety criteria, a detailed site characterization program including above-ground and underground investigations must be performed to provide all necessary data for the site-specific safety assessment.

  2. Site-specific gene integration in rice genome mediated by the FLP-FRT recombination system.

    PubMed

    Nandy, Soumen; Srivastava, Vibha

    2011-08-01

    Plant transformation based on random integration of foreign DNA often generates complex integration structures. Precision in the integration process is necessary to ensure the formation of full-length, single-copy integration. Site-specific recombination systems are versatile tools for precise genomic manipulations such as DNA excision, inversion or integration. The yeast FLP-FRT recombination system has been widely used for DNA excision in higher plants. Here, we report the use of FLP-FRT system for efficient targeting of foreign gene into the engineered genomic site in rice. The transgene vector containing a pair of directly oriented FRT sites was introduced by particle bombardment into the cells containing the target locus. FLP activity generated by the co-bombarded FLP gene efficiently separated the transgene construct from the vector-backbone and integrated the backbone-free construct into the target site. Strong FLP activity, derived from the enhanced FLP protein, FLPe, was important for the successful site-specific integration (SSI). The majority of the transgenic events contained a precise integration and expressed the transgene. Interestingly, each transgenic event lacked the co-bombarded FLPe gene, suggesting reversion of the integration structure in the presence of the constitutive FLPe expression. Progeny of the precise transgenic lines inherited the stable SSI locus and expressed the transgene. This work demonstrates the application of FLP-FRT system for site-specific gene integration in plants using rice as a model.

  3. Generic guidelines versus site-specific assessments: Does marriage make sense?

    SciTech Connect

    Gaudet, C.L.; Keenleyside, K.A.; Smith, S.L.; Kent, R.A.; Wong, M.P.

    1995-12-31

    The maintenance, protection and restoration of a high level of environmental quality requires the availability of practical scientific tools. Environmental quality guidelines (also called criteria) are one such scientific tool that help measure progress towards these goals. These guidelines provide scientific benchmarks that can offer consistency and clarity in defining scientific measures for environmental quality that are easily understood, communicated, and implemented as the basis for management decisions. At the same time, debate exists over the use of generic guidelines versus site-specific risk assessments. It is the contention that generic and site-specific approaches are not mutually exclusive, but complementary decision-support tools and that any apparent controversy stems from an incomplete understanding of the nature and intent of generic environmental quality guidelines or from the use of guidelines in the absence of a coherent framework. The authors advocate an approach that marries the strengths of the generic and site-specific approaches and promotes consistent, scientifically-defensible decisions that support broad societal goals for environmental protection. Using Canadian environmental quality guidelines as an example, they provide an overview of the role of environmental quality guidelines in decision-making, with concrete examples of their implementation in addressing specific environmental quality issues.

  4. Efficient Genome Manipulation by Variants of Site-Specific Recombinases R and TD.

    PubMed

    Voziyanova, Eugenia; Anderson, Rachelle P; Shah, Riddhi; Li, Feng; Voziyanov, Yuri

    2016-02-27

    Genome engineering benefits from the availability of DNA modifying enzymes that have different target specificities and have optimized performance in different cell types. This variety of site-specific enzymes can be used to develop complex genome engineering applications at multiple loci. Although eight yeast site-specific tyrosine recombinases are known, only Flp is actively used in genome engineering. To expand the pool of the yeast site-specific tyrosine recombinases capable of mediating genome manipulations in mammalian cells, we engineered and analyzed variants of two tyrosine recombinases: R and TD. The activity of the evolved variants, unlike the activity of the native R and TD recombinases, is suitable for genome engineering in Escherichia coli and mammalian cells. Unexpectedly, we found that R recombinase benefits from the shortening of its C-terminus. We also found that the activity of wild-type R can be modulated by its non-consensus "head" sequence but this modulation became not apparent in the evolved R variants. The engineered recombinase variants were found to be active in all recombination reactions tested: excision, integration, and dual recombinase-mediated cassette exchange. The analysis of the latter reaction catalyzed by the R/TD recombinase pair shows that the condition supporting the most efficient replacement reaction favors efficient TD-mediated integration reaction while favoring efficient R-mediated integration and deletion reactions. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Characterizing site specific considerations for protecting aircraft during LGS operations at W. M. Keck Observatory

    NASA Astrophysics Data System (ADS)

    Stomski, Paul J., Jr.; Campbell, Randy; McCann, Kevin; Shimko, Steve

    2010-07-01

    W. M. Keck Observatory (WMKO) routinely operates laser guide star (LGS) Adaptive Optics (AO) systems at the telescope facility on the Big Island of Hawaii. One of the operational requirements for the LGS system is that a safety system to prevent nearby aircraft from being adversely affected by the laser must be provided. We will support operations in the near term with human aircraft spotters until we can successfully develop and get the appropriate approvals needed for an Automated, Integrated and Reliable System for an Aircraft Friendly Environment (AIRSAFE). This report describes some of the preliminary requirements development work at WMKO in support of the future development of AIRSAFE. We discuss the results of recent work to characterize site specific considerations that impact requirements development. The site specific considerations include the proximity of WMKO laser operations to nearby commercial airports, the implications of military operations in the area and the character of the air traffic volume and flight patterns over the telescope facility. Finally, we discuss how the design and implementation of AIRSAFE will be impacted by these site specific considerations.

  6. Hypoxia drives transient site-specific copy gain and drug-resistant gene expression

    PubMed Central

    Black, Joshua C.; Atabakhsh, Elnaz; Kim, Jaegil; Biette, Kelly M.; Van Rechem, Capucine; Ladd, Brendon; Burrowes, Paul d.; Donado, Carlos; Mattoo, Hamid; Kleinstiver, Benjamin P.; Song, Bing; Andriani, Grasiella; Joung, J. Keith; Iliopoulos, Othon; Montagna, Cristina; Pillai, Shiv; Getz, Gad

    2015-01-01

    Copy number heterogeneity is a prominent feature within tumors. The molecular basis for this heterogeneity remains poorly characterized. Here, we demonstrate that hypoxia induces transient site-specific copy gains (TSSGs) in primary, nontransformed, and transformed human cells. Hypoxia-driven copy gains are not dependent on HIF1α or HIF2α; however, they are dependent on the KDM4A histone demethylase and are blocked by inhibition of KDM4A with a small molecule or the natural metabolite succinate. Furthermore, this response is conserved at a syntenic region in zebrafish cells. Regions with site-specific copy gain are also enriched for amplifications in hypoxic primary tumors. These tumors exhibited amplification and overexpression of the drug resistance gene CKS1B, which we recapitulated in hypoxic breast cancer cells. Our results demonstrate that hypoxia provides a biological stimulus to create transient site-specific copy alterations that could result in heterogeneity within tumors and cell populations. These findings have major implications in our understanding of copy number heterogeneity and the emergence of drug resistance genes in cancer. PMID:25995187

  7. Biologically Derived Nanoparticle Arrays via a Site-Specific Reconstitution of Ferritin and their Electrochemistry

    NASA Technical Reports Server (NTRS)

    Kim, Jae-Woo; Choi, Sang H.; Lillehei, Peter T.; King, Glen C.; Elliott, James R.; Chu, Sang-Hyon; Park, Yeonjoon; Watt, Gerald D.

    2004-01-01

    Nanoparticle arrays biologically derived from an electrochemically-controlled site-specific biomineralization were fabricated on a gold substrate through the immobilization process of biomolecules. The work reported herein includes the immobilization of ferritin with various surface modifications, the electrochemical biomineralization of ferritins with different inorganic cores, the fabrication of self-assembled arrays with the immobilized ferritin, and the electrochemical characterization of various core materials. Protein immobilization on the substrate is achieved by anchoring ferritins with dithiobis-N-succinimidyl propionate (DTSP). A reconstitution process of electrochemical site-specific biomineralization with a protein cage loads ferritins with different core materials such as Pt, Co, Mn, and Ni. The ferritin acts as a nano-scale template, a biocompatible cage, and a separator between the nanoparticles. The nano-sized metalcored ferritins on a gold substrate displayed a good electrochemical activity for the electron transport and storage, which is suitable for bioelectronics applications such as biofuel cell, bionanobattery, biosensors, etc. Keywords: Ferritin, immobilization, site-specific reconstitution, biomineralization, and bioelectronics

  8. Synthesis of DNA Oligodeoxynucleotides Containing Site-Specific 1,3-Butadiene- Deoxyadenosine Lesions

    PubMed Central

    Wickramaratne, Susith; Seiler, Christopher L.

    2016-01-01

    Post-oligomerization synthesis is a useful technique for preparing site-specifically modified DNA oligomers. This approach involves site-specific incorporation of inherently reactive halogenated nucleobases into DNA strands using standard solid phase synthesis, followed by post-oligomerization nucleophilic aromatic substitution (SNAr) reactions with carcinogen-derived synthons. In these reactions, the inherent reactivities of DNA and carcinogen-derived species are reversed: the modified DNA nucleobase acts as an electrophile, while the carcinogen-derived species acts as a nucleophile. In the present protocol, we describe the use of the post-oligomerization approach to prepare DNA strands containing site- and stereospecific N6-adenine and N1, N6-adenine adducts induced by epoxide metabolites of the known human and animal carcinogen, 1,3-butadiene (BD). The resulting oligomers containing site specific, structurally defined DNA adducts can be used in structural and biological studies to reveal the roles of specific BD adducts in carcinogenesis and mutagenesis. PMID:26344227

  9. Generation of site-specific mutant mice using the CRISPR/Cas9 system.

    PubMed

    Min, Bai; Qi, Li; Yanjiao, Shao; Yuanhua, Huang; Dali, Li; Yanlin, Ma

    2015-10-01

    The CRISPR/Cas9 system is a recently developed important technology for genome editing in cellular and animal models. Here we established a CRISPR/Cas9-based system of generating site-specific mutant mice using DNA double-strand breaks (DSBs) induced homologous recombination (HR)-dependent or independent repair mechanism. Through co-microinjection of Cas9 mRNA and single-guide RNA (sgRNA) targeting genomic DNA sequence corresponding to enzyme activity of lysine (K)-specific demethylase 2b (Kdm2b), both a frame-shifted Kdm2b null mutant and a Kdm2b enzyme activity disrupted mouse strain were obtained simultaneously. Moreover, sgRNA targeting flavin containing monooxygenases3 (Fmo3) gene and the corresponding single strand oligonucleotides (ssODN) donor template with point mutation were co-injected into the male pronucleus of one-cell mouse embryos stimulated HR-mediated repair mechanism. Genomic sequence analysis of F0 mice showed that frame-shifted Fmo3 knockout mouse and site-specific Fmo3 knock-in mouse with single base substitution were successfully generated, and these mutations could be stably transmitted to the next generation. Therefore, we successfully generated mouse strains containing site-specific mutations through HR-dependent and -independent DSB repair using the CRISPR/Cas9 system.

  10. Comparisons of CAP88PC version 2.0 default parameters to site specific inputs

    SciTech Connect

    Lehto, M. A.; Courtney, J. C.; Charter, N.; Egan, T.

    2000-03-02

    The effects of varying the input for the CAP88PC Version 2.0 program on the total effective dose equivalents (TEDEs) were determined for hypothetical releases from the Hot Fuel Examination Facility (HFEF) located at the Argonne National Laboratory site on the Idaho National Engineering and Environmental Laboratory (INEEL). Values for site specific meteorological conditions and agricultural production parameters were determined for the 80 km radius surrounding the HFEF. Four nuclides, {sup 3}H, {sup 85}Kr, {sup 129}I, and {sup 137}Cs (with its short lived progeny, {sup 137m}Ba) were selected for this study; these are the radioactive materials most likely to be released from HFEF under normal or abnormal operating conditions. Use of site specific meteorological parameters of annual precipitation, average temperature, and the height of the inversion layer decreased the TEDE from {sup 137}Cs-{sup 137m}Ba up to 36%; reductions for other nuclides were less than 3%. Use of the site specific agricultural parameters reduced TEDE values between 7% and 49%, depending on the nuclide. Reductions are associated with decreased committed effective dose equivalents (CEDEs) from the ingestion pathway. This is not surprising since the HFEF is located well within the INEEL exclusion area, and the surrounding area closest to the release point is a high desert with limited agricultural diversity. Livestock and milk production are important in some counties at distances greater than 30 km from the HFEF.

  11. Lack of site-specific recombination between mitochondrial genomes of petite mutants of yeast.

    PubMed

    Rayko, E; Goursot, R; Bernardi, G

    1993-10-15

    Previous work from our laboratory showed that mitochondrial (mt) genomes, with tandem repeat units, from spontaneous, cytoplasmic petite mutants of Saccharomyces cerevisiae do not exhibit site-specific recombination in petite x petite crosses [Rayko et al., Gene 63 (1988) 213-226]. Here, we have extended and confirmed these observations by studying other crosses of petites with tandem repeat units, as well as crosses in which one of the parents was, instead, an unstable petite, a-15/4/1, having a palindromic mt genome. In no case was site-specific recombination of the parental mt genomes observed. Progeny cells harbored mt genomes derived from either one or both of the two parents, as shown by analysis of restriction fragments. In the case of biparental inheritance, extensive subcloning of the diploids showed that this was due to a persistent heteroplasmic state and not to intermolecular recombination. The 'new' restriction fragments present in the mt DNA from some diploids were shown to be derived from the unstable parental genome, a-15/4/1, by a secondary excision process. Lack of site-specific recombination is, therefore, not only a feature of crosses involving petite genomes made up of tandem repeat units, but also of crosses in which one parental genome consists of inverted repeats and frequently originates secondary petite genomes formed by tandem repeats. Previous reports of mt recombination in petite mutants are discussed in light of these results.

  12. Hypoxia drives transient site-specific copy gain and drug-resistant gene expression.

    PubMed

    Black, Joshua C; Atabakhsh, Elnaz; Kim, Jaegil; Biette, Kelly M; Van Rechem, Capucine; Ladd, Brendon; Burrowes, Paul D; Donado, Carlos; Mattoo, Hamid; Kleinstiver, Benjamin P; Song, Bing; Andriani, Grasiella; Joung, J Keith; Iliopoulos, Othon; Montagna, Cristina; Pillai, Shiv; Getz, Gad; Whetstine, Johnathan R

    2015-05-15

    Copy number heterogeneity is a prominent feature within tumors. The molecular basis for this heterogeneity remains poorly characterized. Here, we demonstrate that hypoxia induces transient site-specific copy gains (TSSGs) in primary, nontransformed, and transformed human cells. Hypoxia-driven copy gains are not dependent on HIF1α or HIF2α; however, they are dependent on the KDM4A histone demethylase and are blocked by inhibition of KDM4A with a small molecule or the natural metabolite succinate. Furthermore, this response is conserved at a syntenic region in zebrafish cells. Regions with site-specific copy gain are also enriched for amplifications in hypoxic primary tumors. These tumors exhibited amplification and overexpression of the drug resistance gene CKS1B, which we recapitulated in hypoxic breast cancer cells. Our results demonstrate that hypoxia provides a biological stimulus to create transient site-specific copy alterations that could result in heterogeneity within tumors and cell populations. These findings have major implications in our understanding of copy number heterogeneity and the emergence of drug resistance genes in cancer. © 2015 Black et al.; Published by Cold Spring Harbor Laboratory Press.

  13. Multiple DNA binding activities of the novel site-specific recombinase, Piv, from Moraxella lacunata.

    PubMed

    Tobiason, D M; Lenich, A G; Glasgow, A C

    1999-04-02

    The recombinase, Piv, is essential for site-specific DNA inversion of the type IV pilin DNA segment in Moraxella lacunata and Moraxella bovis. Piv shows significant homology with the transposases of the IS110/IS492 family of insertion elements, but, surprisingly, Piv contains none of the conserved amino acid motifs of the lambda Int or Hin/Res families of site-specific recombinases. Therefore, Piv may mediate site-specific recombination by a novel mechanism. To begin to determine how Piv may assemble a synaptic nucleoprotein structure for DNA cleavage and strand exchange, we have characterized the interaction of Piv with the DNA inversion region of M. lacunata. Gel shift and nuclease/chemical protection assays, competition and dissociation rate analyses, and cooperativity studies indicate that Piv binds two distinct recognition sequences. One recognition sequence, found at multiple sites within and outside of the invertible segment, is bound by Piv protomers with high affinity. The second recognition sequence is located at the recombination cross-over sites at the ends of the invertible element; Piv interacts with this sequence as an oligomer with apparent low affinity. A model is proposed for the role of the different Piv binding sites of the M. lacunata inversion region in the formation of an active synaptosome.

  14. Site-specific parameter values for the Nuclear Regulatory Commission's food pathway dose model

    SciTech Connect

    Hamby, D.M. )

    1992-02-01

    Routine operations at the Savannah River Site (SRS) in Western South Carolina result in radionuclide releases to the atmosphere and to the Savannah River. The resulting radiation doses to the off-site maximum individual and the off-site population within 80 km of the SRS are estimated on a yearly basis. These estimates are currently generated using dose models prescribed for the commercial nuclear power industry by the Nuclear Regulatory Commission (NRC). The NRC provides default values for dose-model parameters for facilities without resources to develop site-specific values. A survey of land- and water-use characteristics for the Savannah River area has been conducted to determine site-specific values for water recreation, consumption, and agricultural parameters used in the NRC Regulatory Guide 1.109 (1977) dosimetric models. These site parameters include local characteristics of meat, milk, and vegetable production; recreational and commercial activities on the Savannah River; and meat, milk, vegetable, and seafood consumption rates. This paper describes how parameter data were obtained at the Savannah River Site and the impacts of such data on off-site dose. Dose estimates using site-specific parameter values are compared to estimates using the NRC default values.

  15. The Bxb1 recombination system demonstrates heritable transmission of site-specific excision in Arabidopsis

    PubMed Central

    2012-01-01

    Background The mycobacteriophage large serine recombinase Bxb1 catalyzes site-specific recombination between its corresponding attP and attB recognition sites. Previously, we and others have shown that Bxb1 has catalytic activity in various eukaryotic species including Nicotiana tabacum, Schizosaccharomyces pombe, insects and mammalian cells. Results In this work, the Bxb1 recombinase gene was transformed and constitutively expressed in Arabidopsis thaliana plants harboring a chromosomally integrated attP and attB-flanked target sequence. The Bxb1 recombinase successfully excised the target sequence in a conservative manner and the resulting recombination event was heritably transmitted to subsequent generations in the absence of the recombinase transgene. In addition, we also show that Bxb1 recombinase expressing plants can be manually crossed with att-flanked target transgenic plants to generate excised progeny. Conclusion The Bxb1 large serine recombinase performs site-specific recombination in Arabidopsis thaliana germinal tissue, producing stable lines free of unwanted DNA. The precise site-specific deletion produced by Bxb1 in planta demonstrates that this enzyme can be a useful tool for the genetic engineering of plants without selectable marker transgenes or other undesirable exogenous sequences. PMID:22436504

  16. Site-Specific N-Glycosylation of Endothelial Cell Receptor Tyrosine Kinase VEGFR-2.

    PubMed

    Chandler, Kevin Brown; Leon, Deborah R; Meyer, Rosana D; Rahimi, Nader; Costello, Catherine E

    2017-02-03

    Vascular endothelial growth factor receptor-2 (VEGFR-2) is an important receptor tyrosine kinase (RTK) that plays critical roles in both physiologic and pathologic angiogenesis. The extracellular domain of VEGFR-2 is composed of seven immunoglobulin-like domains, each with multiple potential N-glycosylation sites (sequons). N-glycosylation plays a central role in RTK ligand binding, trafficking, and stability. However, despite its importance, the functional role of N-glycosylation of VEGFR-2 remains poorly understood. The objectives of the present study were to characterize N-glycosylation sites in VEGFR-2 via enzymatic release of the glycans and concomitant incorporation of (18)O into formerly N-glycosylated sites followed by tandem mass spectrometry (MS/MS) analysis to determine N-glycosylation site occupancy and the site-specific N-glycan heterogeneity of VEGFR-2 glycopeptides. The data demonstrated that all seven VEGFR-2 immunoglobulin-like domains have at least one occupied N-glycosylation site. MS/MS analyses of glycopeptides and deamidated, deglycosylated (PNGase F-treated) peptides from ectopically expressed VEGFR-2 in porcine aortic endothelial (PAE) cells identified N-glycans at the majority of the 17 potential N-glycosylation sites on VEGFR-2 in a site-specific manner. The data presented here provide direct evidence for site-specific, heterogeneous N-glycosylation and N-glycosylation site occupancy on VEGFR-2. The study has important implications for the therapeutic targeting of VEGFR-2, ligand binding, trafficking, and signaling.

  17. Cancer registries in Japan: National Clinical Database and site-specific cancer registries.

    PubMed

    Anazawa, Takayuki; Miyata, Hiroaki; Gotoh, Mitsukazu

    2015-02-01

    The cancer registry is an essential part of any rational program of evidence-based cancer control. The cancer control program is required to strategize in a systematic and impartial manner and efficiently utilize limited resources. In Japan, the National Clinical Database (NCD) was launched in 2010. It is a nationwide prospective registry linked to various types of board certification systems regarding surgery. The NCD is a nationally validated database using web-based data collection software; it is risk adjusted and outcome based to improve the quality of surgical care. The NCD generalizes site-specific cancer registries by taking advantage of their excellent organizing ability. Some site-specific cancer registries, including pancreatic, breast, and liver cancer registries have already been combined with the NCD. Cooperation between the NCD and site-specific cancer registries can establish a valuable platform to develop a cancer care plan in Japan. Furthermore, the prognosis information of cancer patients arranged using population-based and hospital-based cancer registries can help in efficient data accumulation on the NCD. International collaboration between Japan and the USA has recently started and is expected to provide global benchmarking and to allow a valuable comparison of cancer treatment practices between countries using nationwide cancer registries in the future. Clinical research and evidence-based policy recommendation based on accurate data from the nationwide database may positively impact the public.

  18. Confident assignment of site-specific glycosylation in complex glycoproteins in a single step.

    PubMed

    Khatri, Kshitij; Staples, Gregory O; Leymarie, Nancy; Leon, Deborah R; Turiák, Lilla; Huang, Yu; Yip, Shun; Hu, Han; Heckendorf, Christian F; Zaia, Joseph

    2014-10-03

    A glycoprotein may contain several sites of glycosylation, each of which is heterogeneous. As a consequence of glycoform diversity and signal suppression from nonglycosylated peptides that ionize more efficiently, typical reversed-phase LC-MS and bottom-up proteomics database searching workflows do not perform well for identification of site-specific glycosylation for complex glycoproteins. We present an LC-MS system for enrichment, separation, and analysis of glycopeptides from complex glycoproteins (>4 N-glycosylation sequons) in a single step. This system uses an online HILIC enrichment trap prior to reversed-phase C18-MS analysis. We demonstrated the effectiveness of the system using a set of glycoproteins including human transferrin (2 sequons), human alpha-1-acid glycoprotein (5 sequons), and influenza A virus hemagglutinin (9 sequons). The online enrichment renders glycopeptides the most abundant ions detected, thereby facilitating the generation of high-quality data-dependent tandem mass spectra. The tandem mass spectra exhibited product ions from both glycan and peptide backbone dissociation for a majority of the glycopeptides tested using collisionally activated dissociation that served to confidently assign site-specific glycosylation. We demonstrated the value of our system to define site-specific glycosylation using a hemagglutinin containing 9 N-glycosylation sequons from a single HILIC-C18-MS acquisition.

  19. GIS-based procedure for site-specific risk assessment of pesticides for aquatic ecosystems.

    PubMed

    Sala, Serenella; Vighi, Marco

    2008-01-01

    The EU Water Framework Directive states that the management of surface water must be based on a site-specific assessment of water quality, that is dependent on land use. As a result, to develop a robust chemical management policy for aquatic ecosystems, the ecotoxicological risk must be strictly related to the local conditions and characteristics of the system. This paper presents a methodology developed to assess the ecotoxicological risk of pesticides to site-specific aquatic ecosystems. Spatial and relational databases, provisional models and risk indices were integrated into Geographical Information Systems (GIS) to produce maps of exposure, effect and risk at watershed scale. Each active ingredient is characterised by a data set that includes input data as well as results represented by a risk assessment cartography. The aim of this procedure is to perform a site-specific risk assessment by integrating geographical distribution of predicted environmental concentrations (PECs), ecotoxicological effects and the potential/actual quality of the exposed ecosystem. Examples of pesticide risk maps for surface waters in Lombardia Region (Northern Italy) are shown.

  20. Site-specific protein labeling with PRIME and chelation-assisted Click chemistry

    PubMed Central

    Uttamapinant, Chayasith; Sanchez, Mateo I.; Liu, Daniel S.; Yao, Jennifer Z.; White, Katharine A.; Grecian, Scott; Clarke, Scott; Gee, Kyle R.; Ting, Alice Y.

    2016-01-01

    This protocol describes an efficient method to site-specifically label cell-surface or purified proteins with chemical probes in two steps: PRobe Incorporation Mediated by Enzymes (PRIME) followed by chelation-assisted copper-catalyzed azide-alkyne cycloaddition (CuAAC). In the PRIME step, Escherichia coli lipoic acid ligase site-specifically attaches a picolyl azide derivative to a 13-amino acid recognition sequence that has been genetically fused onto the protein of interest. Proteins bearing picolyl azide are chemoselectively derivatized with an alkyne-probe conjugate by chelation-assisted CuAAC in the second step. We describe herein the optimized protocols to synthesize picolyl azide, perform PRIME labeling, and achieve CuAAC derivatization of picolyl azide on live cells, fixed cells, and purified proteins. Reagent preparations, including synthesis of picolyl azide probes and expression of lipoic acid ligase, take 12 d, while the procedure to perform site-specific picolyl azide ligation and CuAAC on cells or on purified proteins takes 40 min-3 h. PMID:23887180

  1. Ultrasensitive and selective DNA detection by hydroxylamine assisted gold nanoparticle amplification.

    PubMed

    Cai, Sheng; Xin, Liang; Lau, Choiwan; Lu, Jianzhong; Zhang, Xueji

    2011-06-07

    A new nanoparticle-based chemiluminescent (CL) method has been developed for the ultrasensitive detection of DNA hybridization which can achieve ultra-sensitivity up to approximately 30 zmol, i.e. 300 aM.

  2. Ultra-Sensitive Photoreceiver Boosts Data Transmission

    NASA Technical Reports Server (NTRS)

    2007-01-01

    NASA depends on advanced, ultra-sensitive photoreceivers and photodetectors to provide high-data communications and pinpoint image-detection and -recognition capabilities from great distances. In 2003, Epitaxial Technologies LLC was awarded a Small Business Innovation Research (SBIR) contract from Goddard Space Flight Center to address needs for advanced sensor components. Epitaxial developed a photoreciever capable of single proton sensitivity that is also smaller, lighter, and requires less power than its predecessor. This receiver operates in several wavelength ranges; will allow data rate transmissions in the terabit range; and will enhance Earth-based missions for remote sensing of crops and other natural resources, including applications for fluorescence and phosphorescence detection. Widespread military and civilian applications are anticipated, especially through enhancing fiber optic communications, laser imaging, and laser communications.

  3. Site-Specific Analyses for Demonstrating Compliance with 10 CFR 61 Performance Objectives - 12179

    SciTech Connect

    Grossman, C.J.; Esh, D.W.; Yadav, P.; Carrera, A.G.

    2012-07-01

    The U.S. Nuclear Regulatory Commission (NRC) is proposing to amend its regulations at 10 CFR Part 61 to require low-level radioactive waste disposal facilities to conduct site-specific analyses to demonstrate compliance with the performance objectives in Subpart C. The amendments would require licensees to conduct site-specific analyses for protection of the public and inadvertent intruders as well as analyses for long-lived waste. The amendments would ensure protection of public health and safety, while providing flexibility to demonstrate compliance with the performance objectives, for current and potential future waste streams. NRC staff intends to submit proposed rule language and associated regulatory basis to the Commission for its approval in early 2012. The NRC staff also intends to develop associated guidance to accompany any proposed amendments. The guidance is intended to supplement existing low-level radioactive waste guidance on issues pertinent to conducting site-specific analyses to demonstrate compliance with the performance objectives. The guidance will facilitate implementation of the proposed amendments by licensees and assist competent regulatory authorities in reviewing the site-specific analyses. Specifically, the guidance provides staff recommendations on general considerations for the site-specific analyses, modeling issues for assessments to demonstrate compliance with the performance objectives including the performance assessment, intruder assessment, stability assessment, and analyses for long-lived waste. This paper describes the technical basis for changes to the rule language and the proposed guidance associated with implementation of the rule language. The NRC staff, per Commission direction, intends to propose amendments to 10 CFR Part 61 to require licensees to conduct site-specific analyses to demonstrate compliance with performance objectives for the protection of public health and the environment. The amendments would require a

  4. In vivo kinetics and nonradioactive imaging of rapidly proliferating cells in graft-versus-host disease.

    PubMed

    Buxbaum, Nataliya P; Farthing, Donald E; Maglakelidze, Natella; Lizak, Martin; Merkle, Hellmut; Carpenter, Andrea C; Oliver, Brittany U; Kapoor, Veena; Castro, Ehydel; Swan, Gregory A; Dos Santos, Liliane M; Bouladoux, Nicolas J; Bare, Catherine V; Flomerfelt, Francis A; Eckhaus, Michael A; Telford, William G; Belkaid, Yasmine; Bosselut, Remy J; Gress, Ronald E

    2017-06-15

    Hematopoietic stem cell transplantation (HSCT) offers a cure for cancers that are refractory to chemotherapy and radiation. Most HSCT recipients develop chronic graft-versus-host disease (cGVHD), a systemic alloimmune attack on host organs. Diagnosis is based on clinical signs and symptoms, as biopsies are risky. T cells are central to the biology of cGVHD. We found that a low Treg/CD4+ T effector memory (Tem) ratio in circulation, lymphoid, and target organs identified early and established mouse cGVHD. Using deuterated water labeling to measure multicompartment in vivo kinetics of these subsets, we show robust Tem and Treg proliferation in lymphoid and target organs, while Tregs undergo apoptosis in target organs. Since deuterium enrichment into DNA serves as a proxy for cell proliferation, we developed a whole-body clinically relevant deuterium MRI approach to nonradioactively detect cGVHD and potentially allow imaging of other diseases characterized by rapidly proliferating cells.

  5. Fast pulsed operation of a small non-radioactive electron source with continuous emission current control

    SciTech Connect

    Cochems, P.; Kirk, A. T.; Bunert, E.; Runge, M.; Goncalves, P.; Zimmermann, S.

    2015-06-15

    Non-radioactive electron sources are of great interest in any application requiring the emission of electrons at atmospheric pressure, as they offer better control over emission parameters than radioactive electron sources and are not subject to legal restrictions. Recently, we published a simple electron source consisting only of a vacuum housing, a filament, and a single control grid. In this paper, we present improved control electronics that utilize this control grid in order to focus and defocus the electron beam, thus pulsing the electron emission at atmospheric pressure. This allows short emission pulses and excellent stability of the emitted electron current due to continuous control, both during pulsed and continuous operations. As an application example, this electron source is coupled to an ion mobility spectrometer. Here, the pulsed electron source allows experiments on gas phase ion chemistry (e.g., ion generation and recombination kinetics) and can even remove the need for a traditional ion shutter.

  6. Detection of adenovirus in nasopharyngeal specimens by radioactive and nonradioactive DNA probes

    SciTech Connect

    Hyypiae, T.

    1985-05-01

    The presence of adenovirus DNA in clinical specimens was analyzed by nucleic acid hybridization assays by both radioactive and enzymatic detection systems. The sensitivity of the hybridization tests was in the range of 10 to 100 pg of homologous adenovirus DNA. Minimal background was noticed with unrelated viral and nonviral DNA. Twenty-four nasopharyngeal mucus aspirate specimens, collected from children with acute respiratory infection, were assayed in the hybridization tests and also by an enzyme immunoassay for adenovirus hexon antigen which was used as a reference test. Sixteen specimens positive by the enzyme immunoassay also were positive in the two nucleic acid hybridization tests, and the remaining eight specimens were negative in all of the tests. The results indicate that nucleid acid hybridization tests with both radioactive and nonradioactive probes can be used for diagnosis of microbial infections.

  7. Nonradioactive Isotopic Labeling and Tracing of Selenoproteins in Cultured Cell Lines.

    PubMed

    Sonet, Jordan; Mounicou, Sandra; Chavatte, Laurent

    2018-01-01

    Selenium (Se) is an essential component of genetically encoded selenoproteins, in the form of a rare amino acid, namely the selenocysteine (Sec). Radioactive (75)Se has been widely used to trace selenoproteins in vitro and in vivo (cell models and animals). Alternatively, its unique isotopic pattern can be used to detect and characterize nonradioactive Se-compounds in cellular extracts using molecular or elemental mass spectrometry at ppm levels. However, when studying trace levels of Se-compounds, such as selenoproteins (ppt levels), the distribution of the signal between its six naturally abundant isotopes reduces its sensitivity. Here, we describe the use of isotopically enriched forms of Se as an alternative strategy to radioactive (75)Se, for the labeling and tracing of selenoproteins in cultured cell lines.

  8. In vivo kinetics and nonradioactive imaging of rapidly proliferating cells in graft-versus-host disease

    PubMed Central

    Buxbaum, Nataliya P.; Farthing, Donald E.; Maglakelidze, Natella; Lizak, Martin; Merkle, Hellmut; Carpenter, Andrea C.; Oliver, Brittany U.; Kapoor, Veena; Castro, Ehydel; Swan, Gregory A.; dos Santos, Liliane M.; Bouladoux, Nicolas J.; Bare, Catherine V.; Flomerfelt, Francis A.; Eckhaus, Michael A.; Telford, William G.; Belkaid, Yasmine; Bosselut, Remy J.; Gress, Ronald E.

    2017-01-01

    Hematopoietic stem cell transplantation (HSCT) offers a cure for cancers that are refractory to chemotherapy and radiation. Most HSCT recipients develop chronic graft-versus-host disease (cGVHD), a systemic alloimmune attack on host organs. Diagnosis is based on clinical signs and symptoms, as biopsies are risky. T cells are central to the biology of cGVHD. We found that a low Treg/CD4+ T effector memory (Tem) ratio in circulation, lymphoid, and target organs identified early and established mouse cGVHD. Using deuterated water labeling to measure multicompartment in vivo kinetics of these subsets, we show robust Tem and Treg proliferation in lymphoid and target organs, while Tregs undergo apoptosis in target organs. Since deuterium enrichment into DNA serves as a proxy for cell proliferation, we developed a whole-body clinically relevant deuterium MRI approach to nonradioactively detect cGVHD and potentially allow imaging of other diseases characterized by rapidly proliferating cells. PMID:28614804

  9. Distinguishing persistent insulin autoantibodies with differential risk: nonradioactive bivalent proinsulin/insulin autoantibody assay.

    PubMed

    Yu, Liping; Miao, Dongmei; Scrimgeour, Laura; Johnson, Kelly; Rewers, Marian; Eisenbarth, George S

    2012-01-01

    A subset of children develops persistent insulin autoantibodies (IAA; almost always as the only islet autoantibody) without evidence of progression to diabetes. The aim of the current study was the development and characterization of the performance of a nonradioactive fluid phase IAA assay in relation to standard IAA radioassay. We developed a nonradioactive IAA assay where bivalent IAA cross-link two insulin moieties in a fluid phase. The serum samples positive for anti-islet autoantibodies from 150 newly diagnosed patients with diabetes (Barbara Davis Center plus Diabetes Autoantibody Standardization Program [DASP] workshop) and 70 prediabetic subjects who were followed to diabetes were studied. In addition, sequential samples from 64 nondiabetic subjects who were persistently IAA(+) were analyzed. With 99th percentile of specificity, the new assay with the technology from Meso Scale Discovery Company (MSD-IAA) detects as positive 61% (61 of 100) of new-onset patients and 80% (56 of 70) of prediabetic patients compared with our current fluid phase micro-IAA radioassay (mIAA; 44 and 74%, respectively). In addition, MSD-IAA demonstrated better sensitivity than our mIAA from blinded DASP workshop (68 vs. 56% with the same 99% specificity). Of 64 IAA(+) nondiabetic subjects, 25% (8 of 32) who had only IAA and thus the low risk for progression to diabetes were positive with MSD-IAA assay. In contrast, 100% (32 of 32) high-risk children (IAA plus other islet autoantibodies) were positive with MSD-IAA. The IAA detectable by radioassay, but not MSD-IAA, were usually of lower affinity compared with the IAA of the high-risk children. These data suggest that a subset of IAA with current radioassay (not MSD-IAA) represents biologic false positives in terms of autoimmunity leading to diabetes. We hypothesize that factors related to the mechanism of loss of tolerance leading to diabetes determine high affinity and MSD-IAA reactivity.

  10. Synthesis and nonradioactive micro-analysis of diphosphoinositol phosphates by HPLC with postcolumn complexometry.

    PubMed

    Lin, Hongying; Lindner, Karsten; Mayr, Georg W

    2010-01-01

    A nonradioactive high-performance anion-exchange chromatographic method based on MDD-HPLC (Mayr Biochem. J. 254:585-591, 1988) was developed for the separation of inositol hexakisphosphate (InsP(6), phytic acid) and most isomers of pyrophosphorylated inositol phosphates, such as diphosphoinositol pentakisphosphate (PPInsP(5) or InsP(7)) and bis-diphosphoinositol tetrakisphosphate (bisPPInsP(4) or InsP(8)). With an acidic elution, the anion-exchange separation led to the resolution of four separable PPInsP(5) isomers (including pairs of enantiomers) into three peaks and of nine separable bisPPInsP(4) isomers into nine peaks. The whole separation procedure was completed within 20-36 min after optimization. Reference standards of all bisPPInsP(4) isomers were generated by a nonenzymatic shotgun synthesis from InsP(6). Hereby, the phosphorylation was brought about nonenzymatically when concentrated InsP(6) bound to the solid surface of anion-exchange beads was incubated with creatine phosphate under optimal pH conditions. From the mixture of pyrophosphorylated InsP(6) derivatives containing all theoretically possible isomers of PPInsP(5), bisPPInsP(4), and also some isomers of trisPPInsP(3), isomers were separated by anion-exchange chromatography and fractions served as reference standards of bisPPInsP(4) isomers for further investigation. Their isomeric nature could be partly assigned by comparison with position specifically synthesized or NMR-characterized purified protozoan reference compounds and partly by limited hydrolysis to PPInsP(5) isomers. By applying this nonradioactive analysis technique to cellular studies, the isomeric nature of the major bisPPInsP(4) in mammalian cells could be identified without the need to obtain sufficient material for NMR analysis.

  11. Safe genetic modification of cardiac stem cells using a site-specific integration technique.

    PubMed

    Lan, Feng; Liu, Junwei; Narsinh, Kazim H; Hu, Shijun; Han, Leng; Lee, Andrew S; Karow, Marisa; Nguyen, Patricia K; Nag, Divya; Calos, Michele P; Robbins, Robert C; Wu, Joseph C

    2012-09-11

    Human cardiac progenitor cells (hCPCs) are a promising cell source for regenerative repair after myocardial infarction. Exploitation of their full therapeutic potential may require stable genetic modification of the cells ex vivo. Safe genetic engineering of stem cells, using facile methods for site-specific integration of transgenes into known genomic contexts, would significantly enhance the overall safety and efficacy of cellular therapy in a variety of clinical contexts. We used the phiC31 site-specific recombinase to achieve targeted integration of a triple fusion reporter gene into a known chromosomal context in hCPCs and human endothelial cells. Stable expression of the reporter gene from its unique chromosomal integration site resulted in no discernible genomic instability or adverse changes in cell phenotype. Namely, phiC31-modified hCPCs were unchanged in their differentiation propensity, cellular proliferative rate, and global gene expression profile when compared with unaltered control hCPCs. Expression of the triple fusion reporter gene enabled multimodal assessment of cell fate in vitro and in vivo using fluorescence microscopy, bioluminescence imaging, and positron emission tomography. Intramyocardial transplantation of genetically modified hCPCs resulted in significant improvement in myocardial function 2 weeks after cell delivery, as assessed by echocardiography (P=0.002) and MRI (P=0.001). We also demonstrated the feasibility and therapeutic efficacy of genetically modifying differentiated human endothelial cells, which enhanced hind limb perfusion (P<0.05 at day 7 and 14 after transplantation) on laser Doppler imaging. The phiC31 integrase genomic modification system is a safe, efficient tool to enable site-specific integration of reporter transgenes in progenitor and differentiated cell types.

  12. Site-specific modification of ED-B-targeting antibody using intein-fusion technology

    PubMed Central

    2011-01-01

    Background A promising new approach in cancer therapy is the use of tumor specific antibodies coupled to cytotoxic agents. Currently these immunoconjugates are prepared by rather unspecific coupling chemistries, resulting in heterogeneous products. As the drug load is a key parameter for the antitumor activity, site-specific strategies are desired. Expressed protein ligation (EPL) and protein trans-splicing (PTS) are methods for the specific C-terminal modification of a target protein. Both include the expression as an intein fusion protein, followed by the exchange of the intein for a functionalized moiety. Results A full-length IgG specific for fibronectin ED-B was expressed as fusion protein with an intein (Mxe GyrA or Npu DnaE) attached to each heavy chain. In vitro protocols were established to site-specifically modify the antibodies in high yields by EPL or PTS, respectively. Although reducing conditions had to be employed during the process, the integrity or affinity of the antibody was not affected. The protocols were used to prepare immunoconjugates containing two biotin molecules per antibody, attached to the C-termini of the heavy chains. Conclusion Full-length antibodies can be efficiently and site-specifically modified at the C-termini of their heavy chains by intein-fusion technologies. The described protocols can be used to prepare immunoconjugates of high homogeneity and with a defined drug load of two. The attachment to the C-termini is expected to retain the affinity and effector functions of the antibodies. PMID:21777442

  13. Synthesis of Site-Specific Radiolabeled Antibodies for Radioimmunotherapy via Genetic Code Expansion.

    PubMed

    Wu, Yiming; Zhu, Hua; Zhang, Bo; Liu, Fei; Chen, Jingxian; Wang, Yufei; Wang, Yan; Zhang, Ziwei; Wu, Ling; Si, Longlong; Xu, Huan; Yao, Tianzhuo; Xiao, Sulong; Xia, Qing; Zhang, Lihe; Yang, Zhi; Zhou, Demin

    2016-10-19

    Radioimmunotherapy (RIT) delivers radioisotopes to antigen-expressing cells via monoantibodies for the imaging of lesions or medical therapy. The chelates are typically conjugated to the antibody through cysteine or lysine residues, resulting in heterogeneous chelate-to-antibody ratios and various conjugation sites. To overcome this heterogeneity, we have developed an approach for site-specific radiolabeling of antibodies by combination of genetic code expansion and click chemistry. As a proof-of-concept study, model systems including anti-CD20 antibody rituximab, positron-emitting isotope (64)Cu, and a newly synthesized bifunctional linker (4-dibenzocyclooctynol-1,4,7,10-tetraazacyclotetradecane-1,4,7,10-tetraacetic acid, DIBO-DOTA) were used. The approach consists of three steps: (1) site-specific incorporation of an azido group-bearing amino acid (NEAK) via the genetic code expansion technique at the defined sites of the antibody as a "chemical handle"; (2) site-specific and quantitative conjugation of bifunctional linkers with the antibodies under a mild condition; and (3) radiolabeling of the chelate-modified antibodies with the appropriate isotope. We used heavy-chain A122NEAK rituximab as proof-of-concept and obtained a homogeneous radioconjugate with precisely two chelates per antibody, incorporated only at the chosen sites. The conjugation did not alter the binding and pharmacokinetics of the rituximab, as indicated by in vitro assays and in vivo PET imaging. We believe our research is a good supplement to the genetic code expansion technique for the development of novel radioimmunoconjugates.

  14. Site-specific bacterial chromosome engineering mediated by IntA integrase from Rhizobium etli.

    PubMed

    Hernández-Tamayo, Rogelio; Torres-Tejerizo, Gonzalo; Brom, Susana; Romero, David

    2016-06-29

    The bacterial chromosome may be used to stably maintain foreign DNA in the mega-base range. Integration into the chromosome circumvents issues such as plasmid replication, stability, incompatibility, and copy number variance. The site-specific integrase IntA from Rhizobium etli CFN42 catalyzes a direct recombination between two specific DNA sites: attA and attD (23 bp). This recombination is stable. The aim of this work was to develop a R. etli derivative that may be used as recipient for the integration of foreign DNA in the chromosome, adapting the IntA catalyzed site-specific recombination system. To fulfill our aim, we designed a Rhizobium etli CFN42 derivative, containing a "landing pad" (LP) integrated into the chromosome. The LP sector consists of a green fluorescent protein gene under the control of the lacZ promoter and a spectinomycin resistance gene. Between the lacZ promoter and the GFP gene we inserted an IntA attachment site, which does not affect transcription from the lac promoter. Also, a mobilizable donor vector was generated, containing an attA site and a kanamycin resistance gene; to facilitate insertion of foreign DNA, this vector also contains a multicloning site. There are no promoters flanking the multicloning site. A biparental mating protocol was used to transfer the donor vector into the landing pad strain; insertion of the donor vector into the landing pad sector via IntA-mediated attA X attA recombination thereby interrupted the expression of the green fluorescent protein, generating site-specific cointegrants. Cointegrants were easily recognized by screening for antibiotic sensitivity and lack of GFP expression, and were obtained with an efficiency of 6.18 %. Integration of foreign DNA in Rhizobium, lacking any similarity with the genome, can be easily achieved by IntA-mediated recombination. This protocol contains the mating and selection procedures for creating and isolating integrants.

  15. Risks of all-cause and site-specific fractures among hospitalized patients with COPD

    PubMed Central

    Liao, Kuang-Ming; Liang, Fu-Wen; Li, Chung-Yi

    2016-01-01

    Abstract Patients with chronic obstructive pulmonary disease (COPD) have a high prevalence of osteoporosis. The clinical sequel of osteoporosis is fracture. Patients with COPD who experience a fracture also have increased morbidity and mortality. Currently, the types of all-cause and site-specific fracture among patients with COPD are unknown. Thus, we elucidated the all-cause and site-specific fractures among patients with COPD. A retrospective, population-based, cohort study was conducted utilizing the Taiwan Longitudinal Health Insurance Database. Patients with COPD were defined as those who were hospitalized with an International Classification of Diseases, Ninth Revision, Clinical Modification code of 490 to 492 or 496 between 2001 and 2011. The index date was set as the date of discharge. The study patients were followed from the index date to the date when they sought care for any type of fracture, date of death, date of health insurance policy termination, or the last day of 2013. The types of fracture analyzed in this study included vertebral, rib, humeral, radial and ulnar/wrist, pelvic, femoral, and tibial and fibular fractures. The cohort consisted of 11,312 patients with COPD. Among these patients, 1944 experienced fractures. The most common site-specific fractures were vertebral, femoral, rib, and forearm fractures (radius, ulna, and wrist) at 32.4%, 31%, 12%, and 11.8%, respectively. The adjusted hazard ratios of fracture were 1.71 [95% confidence interval (95% CI) = 1.56–1.87] for female patient with COPD and 1.50 (95% CI = 1.39–1.52) for patients with osteoporosis after covariate adjustment. Vertebral and hip fractures are common among patients with COPD, especially among males with COPD. Many comorbidities contribute to the high risk of fracture among patients with COPD. PMID:27749576

  16. Spatial Distribution and Site-Specific Spraying of Main Sucking Pests of Elm Trees.

    PubMed

    Karimzadeh, R; Iranipour, S

    2016-11-09

    Elm trees are important landscape trees and sucking insects weaken the elm trees and produce large amounts of honeydew. The main objectives of this study were to identify main honeydew-producing pests of elm trees and do site-specific spraying against these pests. To map the spatial distribution of the sucking pests in the large scale, the study area was divided into 40 × 40 m grids and one tree was chosen randomly from each grid (a total of 55 trees). These trees were sampled twice a year in 2011 and 2012. Each sample was a 30-cm branch terminal. Eight samples were taken from each tree in four cardinal directions and two canopy levels. The number of sucking insects and leaves of each sample were counted and recorded. Spatial analysis of the data was carried out using geostatistics. Kriging was used for producing prediction maps. Insecticide application was restricted to the regions with populations higher than threshold. To identify within-tree distribution of the honeydew-producing pests, six and four elm trees were chosen in 2011 and 2012 respectively, and sampled weekly. These trees were sampled as described previously. European elm scale (EES), Gossyparia spuria (Modeer) and two species of aphids were the dominant honeydew-producing pests. The results revealed that the effects of direction, canopy level and their interactions on insect populations were not statistically significant (P < 0.05). Site-specific spraying decreased the amount of insecticides used by ca. 20%, while satisfactory control of the sucking pests and honeydew excretion was obtained. Considering the environmental and economic benefits of site-specific spraying, it is worth doing more complementary works in this area.

  17. Site-specific methylated reporter constructs for functional analysis of DNA methylation.

    PubMed

    Han, Weiguo; Shi, Miao; Spivack, Simon D

    2013-11-01

    Methods to experimentally alter and functionally evaluate cytosine methylation in a site-specific manner have proven elusive. We describe a site-specific DNA methylation method, using synthetically methylated primers and high fidelity PCR coupled with ligation of reporter constructs. We applied this method to introduce methylated cytosines into fragments of the respective DAPK and RASSF1A promoters that had been cloned into luciferase reporters. We found that methylation of 3-7 residue CpG clusters that were 5' adjacent to the transcription start site (TSS) of the DAPK gene produced up to a 54% decrease in promoter activity (p<0.01). Similarly, for RASSF1A promoter reporter constructs, the methylation of either of two clusters of four CpGs each, but not an intervening cluster, produced a 63% decrease in promoter activity (p<0.01), suggesting that precise mCpG position is crucial, and factors other than simple proximity to the TSS are at play. Chromatin immunoprecipitation analysis of these reporter constructs demonstrated that transcription factor Oct-1 and Sp1 preferentially bound the unmethylated vs. methylated DAPK or RASSF1A promoter reporter constructs at the functional CpG sites. Histone H1, hnRNP1, and MeCP2 showed preferential binding to methylated sequence at functional sites in these reporter constructs, as well as highly preferential (> 8-80-fold) binding to native methylated vs. unmethylated chromatin. These results suggest that: (1) site-specific, precision DNA methylation of a reporter construct can be used for functional analysis of commonly observed gene promoter methylation patterns; (2) the reporter system contains key elements of the endogenous chromatin machinery.

  18. Site-specific data confirm arsenic exposure predicted by the U.S. Environmental Protection Agency.

    PubMed Central

    Walker, S; Griffin, S

    1998-01-01

    The EPA uses an exposure assessment model to estimate daily intake to chemicals of potential concern. At the Anaconda Superfund site in Montana, the EPA exposure assessment model was used to predict total and speciated urinary arsenic concentrations. Predicted concentrations were then compared to concentrations measured in children living near the site. When site-specific information on concentrations of arsenic in soil, interior dust, and diet, site-specific ingestion rates, and arsenic absorption rates were used, measured and predicted urinary arsenic concentrations were in reasonable agreement. The central tendency exposure assessment model successfully described the measured urinary arsenic concentration for the majority of children at the site. The reasonable maximum exposure assessment model successfully identified the uppermost exposed population. While the agreement between measured and predicted urinary arsenic is good, it is not exact. The variables that were identified which influenced agreement included soil and dust sample collection methodology, daily urinary volume, soil ingestion rate, and the ability to define the exposure unit. The concentration of arsenic in food affected agreement between measured and predicted total urinary arsenic, but was not considered when comparing measured and predicted speciated urinary arsenic. Speciated urinary arsenic is the recommended biomarker for recent inorganic arsenic exposure. By using site-specific data in the exposure assessment model, predicted risks from exposure to arsenic were less than predicted risks would have been if the EPA's default values had been used in the exposure assessment model. This difference resulted in reduced magnitude and cost of remediation while still protecting human health. Images Figure 1 PMID:9452415

  19. Safe Genetic Modification of Cardiac Stem Cells Using a Site-Specific Integration Technique

    PubMed Central

    Lan, Feng; Liu, Junwei; Narsinh, Kazim H.; Hu, Shijun; Han, Leng; Lee, Andrew S.; Karow, Marisa; Nguyen, Patricia K.; Nag, Divya; Calos, Michele P.; Robbins, Robert C.; Wu, Joseph C.

    2012-01-01

    Background Human cardiac progenitor cells (hCPCs) are a promising cell source for regenerative repair after myocardial infarction. Exploitation of their full therapeutic potential may require stable genetic modification of the cells ex vivo. Safe genetic engineering of stem cells, using facile methods for site-specific integration of transgenes into known genomic contexts, would significantly enhance the overall safety and efficacy of cellular therapy in a variety of clinical contexts. Methods and Results We employed the phiC31 site-specific recombinase to achieve targeted integration of a triple fusion reporter gene into a known chromosomal context in hCPCs and human endothelial cells (hECs). Stable expression of the reporter gene from its unique chromosomal integration site resulted in no discernible genomic instability or adverse changes in cell phenotype. Namely, phiC31-modified hCPCs were unchanged in their differentiation propensity, cellular proliferative rate, and global gene expression profile when compared to unaltered control hCPCs. Expression of the triple fusion reporter gene enabled multimodal assessment of cell fate in vitro and in vivo using fluorescence microscopy, bioluminescence imaging (BLI), and positron emission tomography (PET). Intramyocardial transplantation of genetically modified hCPCs resulted in significant improvement in myocardial function two weeks after cell delivery, as assessed by echocardiography (P = 0.002) and magnetic resonance imaging (P = 0.001). We also demonstrated the feasibility and therapeutic efficacy of genetically modifying differentiated hECs, which enhanced hindlimb perfusion (P<0.05 at day 7 and 14 after transplantation) on laser Doppler imaging. Conclusions The phiC31 integrase genomic modification system is a safe, efficient tool to enable site-specific integration of reporter transgenes in progenitor and differentiated cell types. PMID:22965984

  20. Environmental restoration and waste management Site-Specific Plan for the Oak Ridge Reservation. FY 1993

    SciTech Connect

    Not Available

    1993-01-15

    The United States Department of Energy (DOE) is committed to achieving and maintaining environmental regulatory compliance while responding to public concerns and emphasizing waste minimization. DOE publishes the Environmental Restoration and Waste Management Five-Year Plan (FYP) annually to document its progress towards these goals. The purpose of this Site-Specific Plan (SSP) is to describe the activities undertaken to implement the FYP goals at the DOE Oak Ridge Field Office (DOE/OR) installations and programs specifically for the Oak Ridge Reservation (ORR) and surrounding areas. This SSP addresses activities and goals to be accomplished during FY93 even through the FYP focuses on FY94.

  1. Site-specific Chemical Modification of a Glycoprotein Fragment Expressed in Yeast

    PubMed Central

    Xiao, Junpeng; Tolbert, Thomas J.

    2011-01-01

    Site-specific modification of glycoproteins has wide application in both biochemical and biophysical studies. This method describes the conjugation of synthetic molecules to the N-terminus of a glycoprotein fragment: immunoglobulin G subclass 1 fragment crystallizable (IgG1 Fc) by native chemical ligation. The glycosylated IgG1 Fc is expressed in a glycosylation deficient yeast strain. The N-terminal cysteine is generated by the endogenous yeast protease Kex2 in the yeast secretory pathway. The N-terminal cysteine is then conjugated with a biotin thioester to produce a biotinylated, glycosylated IgG1 Fc using native chemical ligation. PMID:21674341

  2. Site-specific DNA-antibody conjugates for specific and sensitive immuno-PCR

    PubMed Central

    Kazane, Stephanie A.; Sok, Devin; Cho, Edward H.; Uson, Maria Loressa; Kuhn, Peter; Schultz, Peter G.; Smider, Vaughn V.

    2012-01-01

    Antibody conjugates are widely used as diagnostics and imaging reagents. However, many such conjugates suffer losses in sensitivity and specificity due to nonspecific labeling techniques. We have developed methodology to site-specifically conjugate oligonucleotides to antibodies containing a genetically encoded unnatural amino acid with orthogonal chemical reactivity. These oligobody molecules were used in immuno-PCR assays to detect Her2+ cells with greater sensitivity and specificity than nonspecifically coupled fragments, and can detect extremely rare Her2+ cells in a complex cellular environment. Such designed antibody-oligonucleotide conjugates should provide sensitive and specific reagents for diagnostics, as well as enable other unique applications based on oligobody building blocks. PMID:22345566

  3. Physical activity and cancer risk: dose-response and cancer, all sites and site-specific.

    PubMed

    Thune, I; Furberg, A S

    2001-06-01

    The association between physical activity and overall and site-specific cancer risk is elaborated in relation to whether any observed dose-response association between physical activity and cancer can be interpreted in terms of how much physical activity (type, intensity, duration, frequency) is needed to influence site- and gender-specific cancer risk. Observational studies were reviewed that have examined the independent effect of the volume of occupational physical activity (OPA) and/or leisure time physical activity (LPA) on overall and site-specific cancer risk. The evidence of cohort and case-control studies suggests that both leisure time and occupational physical activity protect against overall cancer risk, with a graded dose-response association suggested in both sexes. Confounding effects such as diet, body weight, and parity are often included as a covariate in the analyses, with little influence on the observed associations. A crude graded inverse dose-response association was observed between physical activity and colon cancer in 48 studies including 40,674 colon/colorectal cancer cases for both sexes. A dose-response effect of physical activity on colon cancer risk was especially observed, when participation in activities of at least moderate activity (>4.5 MET) and demonstrated by activities expressed as MET-hours per week. An observed inverse association with a dose-response relationship between physical activity and breast cancer was also identified in the majority of the 41 studies including 108,031 breast cancer cases. The dose-response relationship was in particular observed in case-control studies and supported by observations in cohort studies when participation in activities of at least moderate activity (>4.5 MET) and demonstrated by activities expressed by MET-hours per week. This association between physical activity and breast cancer risk is possibly dependent on age at exposure, age at diagnosis, menopausal status and other effect

  4. Analysis of site-specific N-glycan remodeling in the endoplasmic reticulum and the Golgi

    PubMed Central

    Hang, Ivan; Lin, Chia-wei; Grant, Oliver C; Fleurkens, Susanna; Villiger, Thomas K; Soos, Miroslav; Morbidelli, Massimo; Woods, Robert J; Gauss, Robert; Aebi, Markus

    2015-01-01

    The hallmark of N-linked protein glycosylation is the generation of diverse glycan structures in the secretory pathway. Dynamic, non-template-driven processes of N-glycan remodeling in the endoplasmic reticulum and the Golgi provide the cellular setting for structural diversity. We applied newly developed mass spectrometry-based analytics to quantify site-specific N-glycan remodeling of the model protein Pdi1p expressed in insect cells. Molecular dynamics simulation, mutational analysis, kinetic studies of in vitro processing events and glycan flux analysis supported the defining role of the protein in N-glycan processing. PMID:26240167

  5. Site-Specific Labeling of scVEGF with Fluorine-18 for Positron Emission Tomography Imaging

    PubMed Central

    Wang, Hui; Gao, Haokao; Guo, Ning; Niu, Gang; Ma, Ying; Kiesewetter, Dale O.; Chen, Xiaoyuan

    2012-01-01

    Vascular endothelial growth factor (VEGF) is one of the most important mediators of angiogenesis. Single-chain (sc)-VEGF protein containing an N-terminal Cys-tag has been designed for site-specific modification with a variety of imaging and therapeutic moieties. Site-specific labeling of scVEGF with thiol-reactive prosthetic group, N-[2-(4-18F-fluorobenzamido) ethyl] maleimide ([18F]FBEM) for positron emission tomography (PET) imaging of VEFGR may provide a new tracer which has great potential for clinical translation. Methods: [18F]FBEM-scVEGF was synthesized by site-specific conjugation of 18F-FBEM to a thiol group in Cys-tag of scVEGF at room temperature. The functional activity after labeling was tested by immunofluorescence staining, cellular uptake and efflux. The tumor targeting and in vivo properties were evaluated by biodistribution and microPET studies in tumor-bearing mice. Results: The radiolabeling yield and specific activity of [18F]FBEM-scVEGF were 20.6 ± 15.1% (based on starting [18F]FBEM, uncorrected, n = 5) and 58.8 ± 12.4 GBq/µmol, respectively. Noninvasive microPET and direct tissue sampling experiments demonstrated that [18F]FBEM-scVEGF had VEGFR specific tumor uptake in MDA-MB-435, U87MG and 4T1 xenograft models. The optimal tumor uptake was achieved at 2 h p.i., which can be partially, but significantly blocked by co-injection of non-labeled scVEGF protein. Overall, [18F]FBEM-scVEGF showed VEGFR specific tumor uptake. Conclusion: The scVEGF was site-specifically labeled with 18F via [18F]FBEM prosthetic group and the tracer [18F]FBEM-scVEGF exhibited high receptor binding affinity and tumor targeting efficacy. Further study of [18F] FBEM-scVEGF to evaluate angiogenesis in cancer and other disease types is warranted. PMID:22768028

  6. Site specific incorporation of heavy atom-containing unnatural amino acids into proteins for structure determination

    DOEpatents

    Xie, Jianming [San Diego, CA; Wang, Lei [San Diego, CA; Wu, Ning [Boston, MA; Schultz, Peter G [La Jolla, CA

    2008-07-15

    Translation systems and other compositions including orthogonal aminoacyl tRNA-synthetases that preferentially charge an orthogonal tRNA with an iodinated or brominated amino acid are provided. Nucleic acids encoding such synthetases are also described, as are methods and kits for producing proteins including heavy atom-containing amino acids, e.g., brominated or iodinated amino acids. Methods of determining the structure of a protein, e.g., a protein into which a heavy atom has been site-specifically incorporated through use of an orthogonal tRNA/aminoacyl tRNA-synthetase pair, are also described.

  7. Site specific chemoselective labelling of proteins with robust and highly sensitive Ru(II) bathophenanthroline complexes.

    PubMed

    Uzagare, Matthew C; Claussnitzer, Iris; Gerrits, Michael; Bannwarth, Willi

    2012-03-21

    The bioorthogonal and chemoselective fluorescence labelling of several cell-free synthesized proteins containing a site-specifically incorporated azido amino acid was possible using different alkyne-functionalized Ru(II) bathophenanthroline complexes. We were able to achieve a selective labelling even in complex mixtures of proteins despite the fact that ruthenium dyes normally show a high tendency for unspecific interactions with proteins and are commonly used for total staining of proteins. Since the employed Ru complexes are extremely robust, photo-stable and highly sensitive, the approach should be applicable to the production of labelled proteins for single molecule spectroscopy and fluorescence-based interaction studies.

  8. Coverage intervals for trace elements in human scalp hair are site specific.

    PubMed

    Tamburo, E; Varrica, D; Dongarrà, G

    2015-01-01

    Coverage intervals for trace elements in human scalp hair commonly provide the basis for interpreting laboratory results and also in comparative decision-making processes regarding exposure risk assessment. This short communication documents, by some examples, that those computed for human hair are to be considered site specific, as they reflect local environmental conditions; also each geographic area has a typical profile of hair elemental composition of its inhabitants. Therefore, the levels of trace elements in hair are not strictly comparable between different areas of the world. This issue is particularly relevant when identification of anomalous environmental exposures are requested or even in detecting physiological disorders.

  9. The species- and site-specific acid-base properties of penicillamine and its homodisulfide

    NASA Astrophysics Data System (ADS)

    Mirzahosseini, Arash; Szilvay, András; Noszál, Béla

    2014-08-01

    Penicillamine, penicillamine disulfide and 4 related compounds were studied by 1H NMR-pH titrations and case-tailored evaluation methods. The resulting acid-base properties are quantified in terms of 14 macroscopic and 28 microscopic protonation constants and the concomitant 7 interactivity parameters. The species- and site-specific basicities are interpreted by means of inductive and shielding effects through various intra- and intermolecular comparisons. The thiolate basicities determined this way are key parameters and exclusive means for the prediction of thiolate oxidizabilities and chelate forming properties in order to understand and influence chelation therapy and oxidative stress at the molecular level.

  10. Enhanced Aqueous Suzuki–Miyaura Coupling Allows Site-Specific Polypeptide 18F-Labeling

    PubMed Central

    2013-01-01

    The excesses of reagents used in protein chemistry are often incompatible with the reduced or even inverse stoichiometries used for efficient radiolabeling. Analysis and screening of aqueous Pd(0) ligand systems has revealed the importance of a guanidine core and the discovery of 1,1-dimethylguanidine as an enhanced ligand for aqueous Suzuki–Miyaura cross-coupling. This novel Pd catalyst system has now allowed the labeling of small molecules, peptides, and proteins with the fluorine-18 prosthetic [18F]4-fluorophenylboronic acid. These findings now enable site-specific protein 18F-labeling under biologically compatible conditions using a metal-triggered reaction. PMID:23991754

  11. Functional polymer-based siRNA delivery carrier that recognizes site-specific biosignals.

    PubMed

    Takemoto, Hiroyasu; Nishiyama, Nobuhiro

    2017-09-18

    Responsive molecular designs to specific biosignals in microenvironments endow site-specific functionalities with associated polymers. Thus, the construction of small interfering RNA (siRNA) carriers with functional polymers enables smart programs that are triggered by sequential biosignals in a pathway to the targeted cytosol for effective gene silencing. In this review, we explain rational strategies for the design of functional polymers with responsiveness to biosignals and describe the examples of smart carriers for siRNA delivery. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Site Specific Synthesis and in-situ Immobilization of Fluorescent Silver Nanoclusters on DNA Nanoscaffolds Using Tollens Reaction

    SciTech Connect

    Pal, Suchetan; Varghese, R.; Deng, Z.; Zhao, Z.; Kumar, A.; Yan, Hao; Liu, Yan

    2011-04-06

    DNA strands with specific sequences and covalently attached sugar moieties were used for the site-specific incorporation of the sugar units on a DNA origami scaffold. This approach enabled the subsequent site-specific synthesis and in situ immobilization of fluorescent Ag clusters at predefined positions on the DNA nanoscaffold by treatment with the Tollens reagent.

  13. Development of Combined Site-Specific MESA and LEPA Methods on a Linear Move Sprinkler Irrigation System

    USDA-ARS?s Scientific Manuscript database

    A site-specific controller, hardware and software systems were developed with the capability to switch between either mid-elevation spray application (MESA) or low energy precision application (LEPA) methods. These systems were field tested and used to manage site-specific irrigations under a linear...

  14. Evaluation of site-specific tactics using bifenazate and Neoseiulus californicus for management of Tetranychus urticae (Acari: Tetranychidae) in strawberries.

    PubMed

    Liu, Ruohan; Nyoike, Teresia W; Liburd, Oscar E

    2016-10-01

    Greenhouse and field experiments were conducted to evaluate the effectiveness of site-specific tactics for management of the twospotted spider mite, Tetranychus urticae Koch, a major pest of greenhouse and field-grown strawberries (Fragaria x ananassa Duchesne). Two site-specific (spot) treatments, the miticide bifenazate (Acramite(®)) and the predatory mite Neoseiulus californicus McGregor, were compared with whole-plot treatments of bifenazate or N. californicus to determine whether T. urticae could be effectively managed in field-grown strawberry using only site-specific tactics. Additionally, the cost of site-specific tactics was compared with whole-plot treatments to determine the economic value of using site-specific management tactics for T. urticae in strawberries. In the greenhouse, all treatments equivalently reduced the number of T. urticae below control. In the field during the 2011-2012 season, more T. urticae eggs and motiles were in the whole-plot treatments of both N. californicus and bifenazate in the mid-season and late season, respectively, compared with the spot treatments. With the exception of site-specific N. californicus during the 2011-2012 field season, there were no differences in marketable yields between plots with site-specific treatments and whole-plot management. An economic analysis demonstrated a significant cost savings (75.3 %) with site-specific treatments of N. californicus compared with whole-plot application of N. californicus. Similarly, a 24.7 % reduction in cost was achieved in using site-specific bifenazate compared with whole-plot application of bifenazate. The findings indicate that site-specific treatments with N. californicus and bifenazate are competitive alternatives to whole-field application for T. urticae management in strawberries.

  15. New developments for the site-specific attachment of protein to surfaces

    SciTech Connect

    Camarero, J A

    2005-05-12

    Protein immobilization on surfaces is of great importance in numerous applications in biology and biophysics. The key for the success of all these applications relies on the immobilization technique employed to attach the protein to the corresponding surface. Protein immobilization can be based on covalent or noncovalent interaction of the molecule with the surface. Noncovalent interactions include hydrophobic interactions, hydrogen bonding, van der Waals forces, electrostatic forces, or physical adsorption. However, since these interactions are weak, the molecules can get denatured or dislodged, thus causing loss of signal. They also result in random attachment of the protein to the surface. Site-specific covalent attachment of proteins onto surfaces, on the other hand, leads to molecules being arranged in a definite, orderly fashion and uses spacers and linkers to help minimize steric hindrances between the protein surface. This work reviews in detail some of the methods most commonly used as well as the latest developments for the site-specific covalent attachment of protein to solid surfaces.

  16. Synthesis of Site-Specific DNA–Protein Conjugates and Their Effects on DNA Replication

    PubMed Central

    2015-01-01

    DNA–protein cross-links (DPCs) are bulky, helix-distorting DNA lesions that form in the genome upon exposure to common antitumor drugs, environmental/occupational toxins, ionizing radiation, and endogenous free-radical-generating systems. As a result of their considerable size and their pronounced effects on DNA–protein interactions, DPCs can interfere with DNA replication, transcription, and repair, potentially leading to mutagenesis, genotoxicity, and cytotoxicity. However, the biological consequences of these ubiquitous lesions are not fully understood due to the difficulty of generating DNA substrates containing structurally defined, site-specific DPCs. In the present study, site-specific cross-links between the two biomolecules were generated by copper-catalyzed [3 + 2] Huisgen cycloaddition (click reaction) between an alkyne group from 5-(octa-1,7-diynyl)-uracil in DNA and an azide group within engineered proteins/polypeptides. The resulting DPC substrates were subjected to in vitro primer extension in the presence of human lesion bypass DNA polymerases η, κ, ν, and ι. We found that DPC lesions to the green fluorescent protein and a 23-mer peptide completely blocked DNA replication, while the cross-link to a 10-mer peptide was bypassed. These results indicate that the polymerases cannot read through the larger DPC lesions and further suggest that proteolytic degradation may be required to remove the replication block imposed by bulky DPC adducts. PMID:24918113

  17. Development of a site-specific marine water quality standard for cyanide

    SciTech Connect

    Arredondo, L.A.; Brix, K.V.; Cardwell, R.D.; Marsden, A.

    1995-12-31

    A study was conducted to develop a site-specific marine standard for cyanide. The generic cyanide standard of 1 {micro}g/L is ``driven`` by toxicity data for eastern rock crab (Cancer irroratus) zoeae. The reported LC50 for C. irroratus is 4.9 {micro}g/L cyanide and is six times more sensitive that any other marine species tested. In order to develop a site-specific standard for Washington state, cyanide toxicity tests were conducted using the first stage zoeae of Cancer magister and Cancer oregonensis, two Cancer resident to Puget Sound, in accordance with standard ASTM test methods. Testing with C. magister and C. oregonensis resulted in Species Mean Acute Values (SMAVS) of 68 and 131 {micro}g/L cyanide based on measured test concentrations. This is considerably higher than that reported for C. irroratus, is more consistent with cyanide toxicity values for other species tested, and results in a water quality criterion of 9.85 {micro}g/L cyanide with inclusion of these values in the data set. This paper presents the test methods used and the potential effects the test results may have on the marine water quality criterion for cyanide.

  18. Site specific rates of mitochondrial genomes and the phylogeny of eutheria

    PubMed Central

    Kjer, Karl M; Honeycutt, Rodney L

    2007-01-01

    Background Traditionally, most studies employing data from whole mitochondrial genomes to diagnose relationships among the major lineages of mammals have attempted to exclude regions that potentially complicate phylogenetic analysis. Components generally excluded are 3rd codon positions of protein-encoding genes, the control region, rRNAs, tRNAs, and the ND6 gene (encoded on the opposite strand). We present an approach that includes all the data, with the exception of the control region. This approach is based on a site-specific rate model that accommodates excessive homoplasy and that utilizes secondary structure as a reference for proper alignment of rRNAs and tRNAs. Results Mitochondrial genomic data for 78 eutherian mammals, 8 metatherians, and 3 monotremes were analyzed with a Bayesian analysis and our site specific rate model. The resultant phylogeny revealed strong support for most nodes and was highly congruent with more recent phylogenies based on nuclear DNA sequences. In addition, many of the conflicting relationships observed by earlier mitochondrial-based analyses were resolved without need for the exclusion of large subsets of the data. Conclusion Rather than exclusion of data to minimize presumed noise associated with non-protein encoding genes in the mitochondrial genome, our results indicate that selection of an appropriate model that accommodates rate heterogeneity across data partitions and proper treatment of RNA genes can result in a mitochondrial genome-based phylogeny of eutherian mammals that is reasonably congruent with recent phylogenies derived from nuclear genes. PMID:17254354

  19. Site-specific emergency response concept plans for the Chemical Stockpile Disposal Program

    SciTech Connect

    Carnes, S.A.

    1989-12-01

    Site-specific emergency response concept plans were developed to help initiate enhanced emergency preparedness for continued storage of the stockpile and the Chemical Stockpile Disposal Program (CSDP) at the eight army installations storing the unitary chemical stockpile -- Aberdeen Proving Ground, Anniston Army Depot, Lexington-Blue Grass Army Depot, Newport Army Ammunition Plant, Pine Bluff Arsenal, Pueblo Depot Activity, Tooele Army Depot, and Umatilla Depot Activity. This document summarizes the emergency response plans for all the sites and highlights similarities and differences among them. Section 2 summarizes site-specific differences in stockpile hazard and risk by showing differences in planning-basis accident categories and distributions of topographical features, meteorological conditions, and populations at risk. Section 3 presents a summary of the methodology used to identify the emergency planning zones for each site and the actual recommended boundaries of those zones for the eight sites. Section 4 identifies feasible and recommended protective actions for the sites and explains reasons for differences in them. Finally, Section 5 notes the dependence of protective action effectiveness on the development and implementation of command and control and warning systems that can be implemented in a timely manner, it also identifies the differences in recommended lead times (i.e., from the onset of an accidental release) needed at the sites for effective implementation of protective actions. 17 refs., 11 figs. , 12 tabs.

  20. Site-specific accumulation and dynamic change of flavonoids in Apocyni Veneti Folium.

    PubMed

    Chen, Cui-Hua; Xu, Hu; Liu, Xun-Hong; Zou, Li-Si; Wang, Mei; Liu, Zi-Xiu; Fu, Xing-Sheng; Zhao, Hui; Yan, Ying

    2017-09-01

    Site-specific accumulation of flavonoids in Apocyni Veneti Folium was determined by laser scanning confocal microscope (LSCM) and the localization of catechins also was observed via vanillin-HCl staining under the conventional optical microscope. The contents of five flavonoids in Apocyni Veneti Folium from different harvest times and growth parts were measured using HPLC method. LSCM observation showed that flavonoids are accumulated in cuticle of epidermal cells and vessel walls, especially in protoplasts and nucleolus of the collenchyma cells and the epidermal cells. Catechins are localized in the palisade parenchyma cells and vessel walls, particularly in the laticifers found in the phloem. On the basis of the difference of the maximal emission wavelength between quercetin and kaempferol derivatives which have fluorescence behavior by appropriate treatment, kaempferol and its derivatives are localized exclusively in the cuticle. Results showed that the content of astragalin in Apocyni Veneti Folium from different parts revealed the decreasing trend, while hyperin and isoquercitrin were higher in June and July analyzed by HPLC. In summary, the site-specific accumulation of flavonoids in Apocyni Veneti Folium can be determined by LSCM and vanillin-HCl staining. The contents of flavonoids in Apocyni Veneti Folium are correlated with harvest times and growth parts. © 2017 Wiley Periodicals, Inc.

  1. Effective and site-specific phosphoramidation reaction for universally labeling nucleic acids.

    PubMed

    Su, Yu-Chih; Chen, Hsing-Yin; Ko, Ni Chien; Hwang, Chi-Ching; Wu, Min Hui; Wang, Li-Fang; Wang, Yun-Ming; Chang, Sheng-Nan; Wang, Eng-Chi; Wang, Tzu-Pin

    2014-03-15

    Here we report efficient and selective postsynthesis labeling strategies, based on an advanced phosphoramidation reaction, for nucleic acids of either synthetic or enzyme-catalyzed origin. The reactions provided phosphorimidazolide intermediates of DNA or RNA which, whether reacted in one pot (one-step) or purified (two-step), were directly or indirectly phosphoramidated with label molecules. The acquired fluorophore-labeled nucleic acids, prepared from the phosphoramidation reactions, demonstrated labeling efficacy by their F/N ratio values (number of fluorophores per molecule of nucleic acid) of 0.02-1.2 which are comparable or better than conventional postsynthesis fluorescent labeling methods for DNA and RNA. Yet, PCR and UV melting studies of the one-step phosphoramidation-prepared FITC-labeled DNA indicated that the reaction might facilitate nonspecific hybridization in nucleic acids. Intrinsic hybridization specificity of nucleic acids was, however, conserved in the two-step phosphoramidation reaction. The reaction of site-specific labeling nucleic acids at the 5'-end was supported by fluorescence quenching and UV melting studies of fluorophore-labeled DNA. The two-step phosphoramidation-based, effective, and site-specific labeling method has the potential to expedite critical research including visualization, quantification, structural determination, localization, and distribution of nucleic acids in vivo and in vitro.

  2. Cre recombinase-mediated site-specific recombination between plant chromosomes.

    PubMed Central

    Qin, M; Bayley, C; Stockton, T; Ow, D W

    1994-01-01

    We report the use of the bacteriophage P1 Cre-lox system for generating conservative site-specific recombination between tobacco chromosomes. Two constructs, one containing a promoterless hygromycin-resistance gene preceded by a lox site (lox-hpt) and the other containing a cauliflower mosaic virus 35S promoter linked to a lox sequence and the cre coding region (35S-lox-cre), were introduced separately into tobacco plants. Crosses between plants harboring either construct produced plants with the two constructs situated on different chromosomes. Plants with recombination events were identified by selecting for hygromycin resistance, a phenotype expressed upon recombination. Molecular analysis showed that these recombination events occurred specifically at the lox sites and resulted in the reciprocal exchange of flanking host DNA. Progenies of these plants showed 67-100% cotransmission of the new transgenes, 35S-lox-hpt and lox-cre, consistent with the preferential cosegregation of translocated chromosomes. These results illustrate that site-specific recombination systems can be useful tools for the large-scale manipulation of eukaryotic chromosomes in vivo. Images PMID:8127869

  3. The Skeletal Site-Specific Role of Connective Tissue Growth Factor in Prenatal Osteogenesis

    PubMed Central

    Lambi, Alex G.; Pankratz, Talia L.; Mundy, Christina; Gannon, Maureen; Barbe, Mary F.; Richtsmeier, Joan T.; Popoff, Steven N.

    2013-01-01

    Background Connective tissue growth factor (CTGF/CCN2) is a matricellular protein that is highly expressed during bone development. Mice with global CTGF ablation (knockout, KO) have multiple skeletal dysmorphisms and perinatal lethality. A quantitative analysis of the bone phenotype has not been conducted. Results We demonstrated skeletal site-specific changes in growth plate organization, bone microarchitecture, and shape and gene expression levels in CTGF KO compared with wild-type mice. Growth plate malformations included reduced proliferation zone and increased hypertrophic zone lengths. Appendicular skeletal sites demonstrated decreased metaphyseal trabecular bone, while having increased mid-diaphyseal bone and osteogenic expression markers. Axial skeletal analysis showed decreased bone in caudal vertebral bodies, mandibles, and parietal bones in CTGF KO mice, with decreased expression of osteogenic markers. Analysis of skull phenotypes demonstrated global and regional differences in CTGF KO skull shape resulting from allometric (size-based) and nonallometric shape changes. Localized differences in skull morphology included increased skull width and decreased skull length. Dysregulation of the transforming growth factor-β-CTGF axis coupled with unique morphologic traits provides a potential mechanistic explanation for the skull phenotype. Conclusions We present novel data on a skeletal phenotype in CTGF KO mice, in which ablation of CTGF causes site-specific aberrations in bone formation. PMID:23073844

  4. High throughput peptide mapping method for analysis of site specific monoclonal antibody oxidation.

    PubMed

    Li, Xiaojuan; Xu, Wei; Wang, Yi; Zhao, Jia; Liu, Yan-Hui; Richardson, Daisy; Li, Huijuan; Shameem, Mohammed; Yang, Xiaoyu

    2016-08-19

    Oxidation of therapeutic monoclonal antibodies (mAbs) often occurs on surface exposed methionine and tryptophan residues during their production in cell culture, purification, and storage, and can potentially impact the binding to their targets. Characterization of site specific oxidation is critical for antibody quality control. Antibody oxidation is commonly determined by peptide mapping/LC-MS methods, which normally require a long (up to 24h) digestion step. The prolonged sample preparation procedure could result in oxidation artifacts of susceptible methionine and tryptophan residues. In this paper, we developed a rapid and simple UV based peptide mapping method that incorporates an 8-min trypsin in-solution digestion protocol for analysis of oxidation. This method is able to determine oxidation levels at specific residues of a mAb based on the peptide UV traces within <1h, from either TBHP treated or UV light stressed samples. This is the simplest and fastest method reported thus far for site specific oxidation analysis, and can be applied for routine or high throughput analysis of mAb oxidation during various stability and degradation studies. By using the UV trace, the method allows more accurate measurement than mass spectrometry and can be potentially implemented as a release assay. It has been successfully used to monitor antibody oxidation in real time stability studies.

  5. Simultaneous non-contiguous deletions using large synthetic DNA and site-specific recombinases

    PubMed Central

    Krishnakumar, Radha; Grose, Carissa; Haft, Daniel H.; Zaveri, Jayshree; Alperovich, Nina; Gibson, Daniel G.; Merryman, Chuck; Glass, John I.

    2014-01-01

    Toward achieving rapid and large scale genome modification directly in a target organism, we have developed a new genome engineering strategy that uses a combination of bioinformatics aided design, large synthetic DNA and site-specific recombinases. Using Cre recombinase we swapped a target 126-kb segment of the Escherichia coli genome with a 72-kb synthetic DNA cassette, thereby effectively eliminating over 54 kb of genomic DNA from three non-contiguous regions in a single recombination event. We observed complete replacement of the native sequence with the modified synthetic sequence through the action of the Cre recombinase and no competition from homologous recombination. Because of the versatility and high-efficiency of the Cre-lox system, this method can be used in any organism where this system is functional as well as adapted to use with other highly precise genome engineering systems. Compared to present-day iterative approaches in genome engineering, we anticipate this method will greatly speed up the creation of reduced, modularized and optimized genomes through the integration of deletion analyses data, transcriptomics, synthetic biology and site-specific recombination. PMID:24914053

  6. Site-specific fluorescent labeling to visualize membrane translocation of a myristoyl switch protein

    PubMed Central

    Yang, Sung-Tae; Lim, Sung In; Kiessling, Volker; Kwon, Inchan; Tamm, Lukas K.

    2016-01-01

    Fluorescence approaches have been widely used for elucidating the dynamics of protein-membrane interactions in cells and model systems. However, non-specific multi-site fluorescent labeling often results in a loss of native structure and function, and single cysteine labeling is not feasible when native cysteines are required to support a protein’s folding or catalytic activity. Here, we develop a method using genetic incorporation of non-natural amino acids and bio-orthogonal chemistry to site-specifically label with a single fluorescent small molecule or protein the myristoyl-switch protein recoverin, which is involved in rhodopsin-mediated signaling in mammalian visual sensory neurons. We demonstrate reversible Ca2+-responsive translocation of labeled recoverin to membranes and show that recoverin favors membranes with negative curvature and high lipid fluidity in complex heterogeneous membranes, which confers spatio-temporal control over down-stream signaling events. The site-specific orthogonal labeling technique is promising for structural, dynamical, and functional studies of many lipid-anchored membrane protein switches. PMID:27605302

  7. Monitoring protein misfolding by site-specific labeling of proteins in vivo.

    PubMed

    Hsieh, Tzung-yang; Nillegoda, Nadinath B; Tyedmers, Jens; Bukau, Bernd; Mogk, Axel; Kramer, Günter

    2014-01-01

    Incorporating fluorescent amino acids by suppression of the TAG amber codon is a useful tool for site-specific labeling of proteins and visualizing their localization in living cells. Here we use a plasmid encoded orthogonal tRNA/aminoacyl-tRNA synthetase pair to site-specifically label firefly luciferase with the environmentally sensitive fluorescent amino acid, 3-(6-acetylnaphthalen-2-ylamino)-2- aminopropanoic acid (ANAP) and explore the detectability of conformational changes in labeled luciferase in the yeast cytoplasm. We find that ANAP labeling efficiency is greatly increased in [PSI+] cells and show that analysis of the ANAP fluorescence emission by confocal imaging allows for tracking the thermal unfolding and aggregation of luciferase in vivo. Furthermore we demonstrate that flow cytometry can be used to study conformational changes in luciferase and chaperone-mediated refolding in quantitative terms and at the level of single cells. This experimental setup for the first time allows for the direct analysis of the folding state of a protein in living cells and may serve as valuable new tool for examining mechanisms of protein folding, misfolding and aggregation.

  8. Monitoring Protein Misfolding by Site-Specific Labeling of Proteins In Vivo

    PubMed Central

    Hsieh, Tzung-yang; Nillegoda, Nadinath B.; Tyedmers, Jens; Bukau, Bernd; Mogk, Axel; Kramer, Günter

    2014-01-01

    Incorporating fluorescent amino acids by suppression of the TAG amber codon is a useful tool for site-specific labeling of proteins and visualizing their localization in living cells. Here we use a plasmid encoded orthogonal tRNA/aminoacyl-tRNA synthetase pair to site-specifically label firefly luciferase with the environmentally sensitive fluorescent amino acid, 3-(6-acetylnaphthalen-2-ylamino)-2- aminopropanoic acid (ANAP) and explore the detectability of conformational changes in labeled luciferase in the yeast cytoplasm. We find that ANAP labeling efficiency is greatly increased in [PSI+] cells and show that analysis of the ANAP fluorescence emission by confocal imaging allows for tracking the thermal unfolding and aggregation of luciferase in vivo. Furthermore we demonstrate that flow cytometry can be used to study conformational changes in luciferase and chaperone-mediated refolding in quantitative terms and at the level of single cells. This experimental setup for the first time allows for the direct analysis of the folding state of a protein in living cells and may serve as valuable new tool for examining mechanisms of protein folding, misfolding and aggregation. PMID:24915041

  9. Coulomb nanoradiator-mediated, site-specific thrombolytic proton treatment with a traversing pristine Bragg peak

    NASA Astrophysics Data System (ADS)

    Jeon, Jae-Kun; Han, Sung-Mi; Min, Soon-Ki; Seo, Seung-Jun; Ihm, Kyuwook; Chang, Won-Seok; Kim, Jong-Ki

    2016-11-01

    Traversing proton beam-irradiated, mid/high-Z nanoparticles produce site-specific enhancement of X-ray photon-electron emission via the Coulomb nanoradiator (CNR) effect, resulting in a nano- to micro-scale therapeutic effect at the nanoparticle-uptake target site. Here, we demonstrate the uptake of iron oxide nanoparticles (IONs) and nanoradiator-mediated, site-specific thrombolysis without damaging the vascular endothelium in an arterial thrombosis mouse model. The enhancement of low-energy electron (LEE) emission and reactive oxygen species (ROS) production from traversing proton beam-irradiated IONs was examined. Flow recovery was only observed in CNR-treated mice, and greater than 50% removal of the thrombus was achieved. A 2.5-fold greater reduction in the thrombus-enabled flow recovery was observed in the CNR group compared with that observed in the untreated ION-only and proton-only control groups (p < 0.01). Enhancement of the X-ray photon-electron emission was evident from both the pronounced Shirley background in the electron yield and the 1.2- to 2.5-fold enhanced production of ROS by the proton-irradiated IONs, which suggests chemical degradation of the thrombus without potent emboli.

  10. Generation of Food-Grade Recombinant Lactic Acid Bacterium Strains by Site-Specific Recombination

    PubMed Central

    Martín, M. Cruz; Alonso, Juan C.; Suárez, Juan E.; Alvarez, Miguel A.

    2000-01-01

    The construction of a delivery and clearing system for the generation of food-grade recombinant lactic acid bacterium strains, based on the use of an integrase (Int) and a resolvo-invertase (β-recombinase) and their respective target sites (attP-attB and six, respectively) is reported. The delivery system contains a heterologous replication origin and antibiotic resistance markers surrounded by two directly oriented six sites, a multiple cloning site where passenger DNA could be inserted (e.g., the cI gene of bacteriophage A2), the int gene, and the attP site of phage A2. The clearing system provides a plasmid-borne gene encoding β-recombinase. The nonreplicative vector-borne delivery system was transformed into Lactobacillus casei ATCC 393 and, by site-specific recombination, integrated as a single copy in an orientation- and Int-dependent manner into the attB site present in the genome of the host strain. The transfer of the clearing system into this strain, with the subsequent expression of the β-recombinase, led to site-specific DNA resolution of the non-food-grade DNA. These methods were validated by the construction of a stable food-grade L. casei ATCC 393-derived strain completely immune to phage A2 infection during milk fermentation. PMID:10831443

  11. Environmental Restoration and Waste Management Site-Specific Plan (SSP) for fiscal year 1992 (FY92)

    SciTech Connect

    Not Available

    1991-09-01

    The FY-92 Site-Specific Plan (FY-92 SSP) for environmental restoration and waste management at the Idaho National Engineering Laboratory (INEL) is designed to provide the reader with easy access to the status of environmental restoration and waste management activities at INEL. The first chapter provides background on INIEL`s physical environment, site history and mission, and general information about the site and its facilities. In addition, this chapter discusses the inter-relationships between the Site Specific Plan, the Environmental Restoration and Waste Management Five-Year Plan, the environmental restoration and waste management prioritization systems, and the Activity Data Sheets (ADSs) for environmental restoration and waste management. This discussion should help readers understand what the SSP is and how it fits into the environmental restoration and waste management process at INEL. This understanding should provide the reader with a better context for understanding the discussions in the SSP as well as a better feel for how and what to comment on during the public comment period that will be held from the first of September through the end of October 1991.

  12. Environmental Restoration and Waste Management Site-Specific Plan (SSP) for fiscal year 1992 (FY92)

    SciTech Connect

    Not Available

    1991-09-01

    The FY-92 Site-Specific Plan (FY-92 SSP) for environmental restoration and waste management at the Idaho National Engineering Laboratory (INEL) is designed to provide the reader with easy access to the status of environmental restoration and waste management activities at INEL. The first chapter provides background on INIEL's physical environment, site history and mission, and general information about the site and its facilities. In addition, this chapter discusses the inter-relationships between the Site Specific Plan, the Environmental Restoration and Waste Management Five-Year Plan, the environmental restoration and waste management prioritization systems, and the Activity Data Sheets (ADSs) for environmental restoration and waste management. This discussion should help readers understand what the SSP is and how it fits into the environmental restoration and waste management process at INEL. This understanding should provide the reader with a better context for understanding the discussions in the SSP as well as a better feel for how and what to comment on during the public comment period that will be held from the first of September through the end of October 1991.

  13. A phage integrase directs efficient site-specific integration in human cells

    PubMed Central

    Groth, Amy C.; Olivares, Eric C.; Thyagarajan, Bhaskar; Calos, Michele P.

    2000-01-01

    The integrase from the Streptomyces phage φC31 carries out efficient recombination between the attP site in the phage genome and the attB site in the host bacterial chromosome. In this paper, we show that the enzyme also functions in human cells. A plasmid assay system was constructed that measured intramolecular integration of attP into attB. This assay was used to demonstrate that in the presence of the φC31 integrase, precise unidirectional integration occurs with an efficiency of 100% in Escherichia coli and >50% in human cells. This assay system was also used to define the minimal sizes of attB and attP at 34 bp and 39 bp, respectively. Furthermore, precise and efficient intermolecular integration of an incoming plasmid bearing attP into an established Epstein–Barr virus plasmid bearing attB was documented in human cells. This work is a demonstration of efficient, site-specific, unidirectional integration in mammalian cells. These observations form the basis for site-specific integration strategies potentially useful in a broad range of genetic engineering applications. PMID:10801973

  14. Topoisomerase IV, not gyrase, decatenates products of site-specific recombination in Escherichia coli

    PubMed Central

    Zechiedrich, E. Lynn; Khodursky, Arkady B.; Cozzarelli, Nicholas R.

    1997-01-01

    DNA replication and recombination generate intertwined DNA intermediates that must be decatenated for chromosome segregation to occur. We showed recently that topoisomerase IV (topo IV) is the only important decatenase of DNA replication intermediates in bacteria. Earlier results, however, indicated that DNA gyrase has the primary role in unlinking the catenated products of site-specific recombination. To address this discordance, we constructed a set of isogenic strains that enabled us to inhibit selectively with the quinolone norfloxacin topo IV, gyrase, both enzymes, or neither enzyme in vivo. We obtained identical results for the decatenation of the products of two different site-specific recombination enzymes, phage λ integrase and transposon Tn3 resolvase. Norfloxacin blocked decatenation in wild-type strains, but had no effect in strains with drug-resistance mutations in both gyrase and topo IV. When topo IV alone was inhibited, decatenation was almost completely blocked. If gyrase alone were inhibited, most of the catenanes were unlinked. We showed that topo IV is the primary decatenase in vivo and that this function is dependent on the level of DNA supercoiling. We conclude that the role of gyrase in decatenation is to introduce negative supercoils into DNA, which makes better substrates for topo IV. We also discovered that topo IV has an unexpectedly strong DNA relaxation activity that, together with gyrase and topo I, is able to set the supercoiling levels in Escherichia coli. PMID:9334322

  15. Hybrid TiO II nanoparticles: an approach for developing site specific DNA cleavage

    NASA Astrophysics Data System (ADS)

    Liu, J.; Saponjic, Z.; Dimitrijevic, N. M.; Luo, S.; Preuss, D.; Rajh, T.

    2006-02-01

    We have developed hybrid light responsive TiO II nanoparticles electronically linked to PNA oligonucleotides that site specifically bind to double stranded target DNA. This opens a new opportunity for the development of a highly efficient "artificial restriction enzyme" whose activity can be controlled by using light. The work focuses on the use of TiO II nanocomposites as analogs of restriction enzymes with unique specificity that does not exist in current biological approaches. TiO II nanoparticles electronically linked to DNA or PNA adapters have been site-specifically attached along double stranded λ DNA vectors. Illumination of this assembly results in selective oxidation of DNA at the deepest "thermodynamic traps" located closest to the nanoparticle surface, causing DNA cleavage. We investigate the effect of the sequence and length of DNA and PNA adapters on the specificity of DNA cleavage. Related to this issue, the potential use of TiO II/DNA nanocomposites as "rare cutters" that cleave DNA in the places not achieved with existing protein-based enzymes is investigated.

  16. Coulomb nanoradiator-mediated, site-specific thrombolytic proton treatment with a traversing pristine Bragg peak

    PubMed Central

    Jeon, Jae-Kun; Han, Sung-Mi; Min, Soon-Ki; Seo, Seung-Jun; Ihm, Kyuwook; Chang, Won-Seok; Kim, Jong-Ki

    2016-01-01

    Traversing proton beam-irradiated, mid/high-Z nanoparticles produce site-specific enhancement of X-ray photon-electron emission via the Coulomb nanoradiator (CNR) effect, resulting in a nano- to micro-scale therapeutic effect at the nanoparticle-uptake target site. Here, we demonstrate the uptake of iron oxide nanoparticles (IONs) and nanoradiator-mediated, site-specific thrombolysis without damaging the vascular endothelium in an arterial thrombosis mouse model. The enhancement of low-energy electron (LEE) emission and reactive oxygen species (ROS) production from traversing proton beam-irradiated IONs was examined. Flow recovery was only observed in CNR-treated mice, and greater than 50% removal of the thrombus was achieved. A 2.5-fold greater reduction in the thrombus-enabled flow recovery was observed in the CNR group compared with that observed in the untreated ION-only and proton-only control groups (p < 0.01). Enhancement of the X-ray photon-electron emission was evident from both the pronounced Shirley background in the electron yield and the 1.2- to 2.5-fold enhanced production of ROS by the proton-irradiated IONs, which suggests chemical degradation of the thrombus without potent emboli. PMID:27897205

  17. Applying nitrogen site-specifically using soil electrical conductivity maps and precision agriculture technology.

    PubMed

    Lund, E D; Wolcott, M C; Hanson, G P

    2001-10-16

    Soil texture varies significantly within many agricultural fields. The physical properties of soil, such as soil texture, have a direct effect on water holding capacity, cation exchange capacity, crop yield, production capability, and nitrogen (N) loss variations within a field. In short, mobile nutrients are used, lost, and stored differently as soil textures vary. A uniform application of N to varying soils results in a wide range of N availability to the crop. N applied in excess of crop usage results in a waste of the grower"s input expense, a potential negative effect on the environment, and in some crops a reduction of crop quality, yield, and harvestability. Inadequate N levels represent a lost opportunity for crop yield and profit. The global positioning system (GPS)-referenced mapping of bulk soil electrical conductivity (EC) has been shown to serve as an effective proxy for soil texture and other soil properties. Soils with a high clay content conduct more electricity than coarser textured soils, which results in higher EC values. This paper will describe the EC mapping process and provide case studies of site-specific N applications based on EC maps. Results of these case studies suggest that N can be managed site-specifically using a variety of management practices, including soil sampling, variable yield goals, and cropping history.

  18. Photoaffinity site-specific covalent labeling of human corticosteroid-binding globulin.

    PubMed Central

    Marver, D; Chiu, W; Wolff, M E; Edelman, I S

    1976-01-01

    A method was developed for the synthesis of high-specific-activity 21-diazo-21-[6,7-(3)H]deoxycorticosterone, an analog of corticosterone. This analog was used as a photoaffinity label of a high affinity steroid-binding protein, human corticosteroid-binding globulin. Based on direct binding studies and crosscompetition experiments, this diazo derivative exhibited the requisite affinity (within a factor of 1.5 times that of corticosterone) and site specificity to qualify as an affinity labeling legand. Irradiation of corticosteroid-binding globulin with the 21-diazo derivative resulted in irreversible binding to corticosteroid-binding globulin, identified by polyacrylamide gel electrophoresis. Specificity of covalent binding to corticosteroid-binding globulin was established by competition analysis with various steroids. Irreversibility of photodependent binding was shown by persistence of the complex on electrophoresis (in contrast to the noncovalently linked complex), and resistance to exchange with corticosterone or pregnanediol and to solvent extraction. Site specificity of covalent binding was inferred from the effects of a scavenger, Tris-HC1, and fluorescence quenching of a neighboring tryptophan. PMID:1069998

  19. Criteria and Processes for the Certification of Non-Radioactive Hazardous and Non-Hazardous Wastes

    SciTech Connect

    Dominick, J

    2008-12-18

    This document details Lawrence Livermore National Laboratory's (LLNL) criteria and processes for determining if potentially volumetrically contaminated or potentially surface contaminated wastes are to be managed as material containing residual radioactivity or as non-radioactive. This document updates and replaces UCRL-AR-109662, Criteria and Procedures for the Certification of Nonradioactive Hazardous Waste (Reference 1), also known as 'The Moratorium', and follows the guidance found in the U.S. Department of Energy (DOE) document, Performance Objective for Certification of Non-Radioactive Hazardous Waste (Reference 2). The 1992 Moratorium document (UCRL-AR-109662) is three volumes and 703 pages. The first volume provides an overview of the certification process and lists the key radioanalytical methods and their associated Limits of Sensitivities. Volumes Two and Three contain supporting documents and include over 30 operating procedures, QA plans, training documents and organizational charts that describe the hazardous and radioactive waste management system in place in 1992. This current document is intended to update the previous Moratorium documents and to serve as the top-tier LLNL institutional Moratorium document. The 1992 Moratorium document was restricted to certification of Resource Conservation and Recovery Act (RCRA), State and Toxic Substances Control Act (TSCA) hazardous waste from Radioactive Material Management Areas (RMMA). This still remains the primary focus of the Moratorium; however, this document increases the scope to allow use of this methodology to certify other LLNL wastes and materials destined for off-site disposal, transfer, and re-use including non-hazardous wastes and wastes generated outside of RMMAs with the potential for DOE added radioactivity. The LLNL organization that authorizes off-site transfer/disposal of a material or waste stream is responsible for implementing the requirements of this document. The LLNL Radioactive and

  20. Ultrasensitivity in Phosphorylation-Dephosphorylation Cycles with Little Substrate

    PubMed Central

    Martins, Bruno M. C.; Swain, Peter S.

    2013-01-01

    Cellular decision-making is driven by dynamic behaviours, such as the preparations for sunrise enabled by circadian rhythms and the choice of cell fates enabled by positive feedback. Such behaviours are often built upon ultrasensitive responses where a linear change in input generates a sigmoidal change in output. Phosphorylation-dephosphorylation cycles are one means to generate ultrasensitivity. Using bioinformatics, we show that in vivo levels of kinases and phosphatases frequently exceed the levels of their corresponding substrates in budding yeast. This result is in contrast to the conditions often required by zero-order ultrasensitivity, perhaps the most well known means for how such cycles become ultrasensitive. We therefore introduce a mechanism to generate ultrasensitivity when numbers of enzymes are higher than numbers of substrates. Our model combines distributive and non-distributive actions of the enzymes with two-stage binding and concerted allosteric transitions of the substrate. We use analytical and numerical methods to calculate the Hill number of the response. For a substrate with phosphosites, we find an upper bound of the Hill number of , and so even systems with a single phosphosite can be ultrasensitive. Two-stage binding, where an enzyme must first bind to a binding site on the substrate before it can access the substrate's phosphosites, allows the enzymes to sequester the substrate. Such sequestration combined with competition for each phosphosite provides an intuitive explanation for the sigmoidal shifts in levels of phosphorylated substrate. Additionally, we find cases for which the response is not monotonic, but shows instead a peak at intermediate levels of input. Given its generality, we expect the mechanism described by our model to often underlay decision-making circuits in eukaryotic cells. PMID:23950701