Sample records for lagging strand synthesis

  1. Strand-Specific Analysis of DNA Synthesis and Proteins Association with DNA Replication Forks in Budding Yeast.

    PubMed

    Yu, Chuanhe; Gan, Haiyun; Zhang, Zhiguo

    2018-01-01

    DNA replication initiates at DNA replication origins after unwinding of double-strand DNA(dsDNA) by replicative helicase to generate single-stranded DNA (ssDNA) templates for the continuous synthesis of leading-strand and the discontinuous synthesis of lagging-strand. Therefore, methods capable of detecting strand-specific information will likely yield insight into the association of proteins at leading and lagging strand of DNA replication forks and the regulation of leading and lagging strand synthesis during DNA replication. The enrichment and Sequencing of Protein-Associated Nascent DNA (eSPAN), which measure the relative amounts of proteins at nascent leading and lagging strands of DNA replication forks, is a step-wise procedure involving the chromatin immunoprecipitation (ChIP) of a protein of interest followed by the enrichment of protein-associated nascent DNA through BrdU immunoprecipitation. The isolated ssDNA is then subjected to strand-specific sequencing. This method can detect whether a protein is enriched at leading or lagging strand of DNA replication forks. In addition to eSPAN, two other strand-specific methods, (ChIP-ssSeq), which detects potential protein-ssDNA binding and BrdU-IP-ssSeq, which can measure synthesis of both leading and lagging strand, were developed along the way. These methods can provide strand-specific and complementary information about the association of the target protein with DNA replication forks as well as synthesis of leading and lagging strands genome wide. Below, we describe the detailed eSPAN, ChIP-ssSeq, and BrdU-IP-ssSeq protocols.

  2. Bacillus subtilis DNA polymerases, PolC and DnaE, are required for both leading and lagging strand synthesis in SPP1 origin-dependent DNA replication

    PubMed Central

    Seco, Elena M.

    2017-01-01

    Abstract Firmicutes have two distinct replicative DNA polymerases, the PolC leading strand polymerase, and PolC and DnaE synthesizing the lagging strand. We have reconstituted in vitro Bacillus subtilis bacteriophage SPP1 θ-type DNA replication, which initiates unidirectionally at oriL. With this system we show that DnaE is not only restricted to lagging strand synthesis as previously suggested. DnaG primase and DnaE polymerase are required for initiation of DNA replication on both strands. DnaE and DnaG synthesize in concert a hybrid RNA/DNA ‘initiation primer’ on both leading and lagging strands at the SPP1 oriL region, as it does the eukaryotic Pol α complex. DnaE, as a RNA-primed DNA polymerase, extends this initial primer in a reaction modulated by DnaG and one single-strand binding protein (SSB, SsbA or G36P), and hands off the initiation primer to PolC, a DNA-primed DNA polymerase. Then, PolC, stimulated by DnaG and the SSBs, performs the bulk of DNA chain elongation at both leading and lagging strands. Overall, these modulations by the SSBs and DnaG may contribute to the mechanism of polymerase switch at Firmicutes replisomes. PMID:28575448

  3. Alternative lengthening of telomeres can be maintained by preferential elongation of lagging strands

    PubMed Central

    Min, Jaewon; Wright, Woodring E.

    2017-01-01

    Abstract Alternative lengthening of telomeres (ALT) is a telomerase independent telomere maintenance mechanism that occurs in ∼15% of cancers. The potential mechanism of ALT is homology-directed telomere synthesis, but molecular mechanisms of how ALT maintains telomere length in human cancer is poorly understood. Here, we generated TERC (telomerase RNA) gene knockouts in telomerase positive cell lines that resulted in long-term surviving clones acquiring the ALT pathway but at a very low frequency. By comparing these ALT cells with parental telomerase positive cells, we observed that ALT cells possess excessively long telomeric overhangs derived from telomere elongation processes that mostly occur during S phase. ALT cells exhibited preferential elongation of the telomeric lagging strands, whereas telomerase positive cells exhibited similar elongation between leading and lagging strands. We propose that the ALT pathway preferentially occurs at telomeric lagging strands leading to heterogeneous telomere lengths observed in most ALT cancers. PMID:28082393

  4. Chromatin Controls DNA Replication Origin Selection, Lagging-Strand Synthesis, and Replication Fork Rates.

    PubMed

    Kurat, Christoph F; Yeeles, Joseph T P; Patel, Harshil; Early, Anne; Diffley, John F X

    2017-01-05

    The integrity of eukaryotic genomes requires rapid and regulated chromatin replication. How this is accomplished is still poorly understood. Using purified yeast replication proteins and fully chromatinized templates, we have reconstituted this process in vitro. We show that chromatin enforces DNA replication origin specificity by preventing non-specific MCM helicase loading. Helicase activation occurs efficiently in the context of chromatin, but subsequent replisome progression requires the histone chaperone FACT (facilitates chromatin transcription). The FACT-associated Nhp6 protein, the nucleosome remodelers INO80 or ISW1A, and the lysine acetyltransferases Gcn5 and Esa1 each contribute separately to maximum DNA synthesis rates. Chromatin promotes the regular priming of lagging-strand DNA synthesis by facilitating DNA polymerase α function at replication forks. Finally, nucleosomes disrupted during replication are efficiently re-assembled into regular arrays on nascent DNA. Our work defines the minimum requirements for chromatin replication in vitro and shows how multiple chromatin factors might modulate replication fork rates in vivo. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  5. DNA-directed mutations. Leading and lagging strand specificity

    NASA Technical Reports Server (NTRS)

    Sinden, R. R.; Hashem, V. I.; Rosche, W. A.

    1999-01-01

    The fidelity of replication has evolved to reproduce B-form DNA accurately, while allowing a low frequency of mutation. The fidelity of replication can be compromised, however, by defined order sequence DNA (dosDNA) that can adopt unusual or non B-DNA conformations. These alternative DNA conformations, including hairpins, cruciforms, triplex DNAs, and slipped-strand structures, may affect enzyme-template interactions that potentially lead to mutations. To analyze the effect of dosDNA elements on spontaneous mutagenesis, various mutational inserts containing inverted repeats or direct repeats were cloned in a plasmid containing a unidirectional origin of replication and a selectable marker for the mutation. This system allows for analysis of mutational events that are specific for the leading or lagging strands during DNA replication in Escherichia coli. Deletions between direct repeats, involving misalignment stabilized by DNA secondary structure, occurred preferentially on the lagging strand. Intermolecular strand switch events, correcting quasipalindromes to perfect inverted repeats, occurred preferentially during replication of the leading strand.

  6. A major role of DNA polymerase δ in replication of both the leading and lagging DNA strands

    PubMed Central

    Prakash, Louise; Prakash, Satya

    2015-01-01

    SUMMARY Genetic studies with S. cerevisiae Polδ (pol3-L612M) and Polε (pol2-M644G) mutant alleles, each of which display a higher rate for the generation of a specific mismatch, have led to the conclusion that Polε is the primary leading strand replicase and that Polδ is restricted to replicating the lagging strand template. Contrary to this widely accepted view, here we show that Polδ plays a major role in the replication of both DNA strands, and that the paucity of pol3-L612M generated errors on the leading strand results from their more proficient removal. Thus, the apparent lack of Polδ contribution to leading strand replication is due to differential mismatch removal rather than differential mismatch generation. Altogether, our genetic studies with Pol3 and Pol2 mutator alleles support the conclusion that Polδ, and not Polε, is the major DNA polymerase for carrying out both leading and lagging DNA synthesis. PMID:26145172

  7. Dynamics of Leading-strand Lesion Skipping by the Replisome

    PubMed Central

    Yeeles, Joseph T.P.; Marians, Kenneth J.

    2013-01-01

    SUMMARY The E. coli replisome stalls transiently when it encounters a lesion in the leading-strand template, skipping over the damage by reinitiating replication at a new primer synthesized downstream by the primase. We report here that template unwinding and lagging-strand synthesis continue downstream of the lesion at a reduced rate after replisome stalling, that one replisome is capable of skipping multiple lesions, and that the rate limiting steps of replication restart involve the synthesis and activation of the new primer downstream. We also find little support for the concept that polymerase uncoupling, where extensive lagging-strand synthesis proceeds downstream in the absence of leading-strand synthesis, involves physical separation of the leading-strand polymerase from the replisome. Instead, our data indicate that extensive uncoupled replication likely results from a failure of the leading-strand polymerase still associated with the DNA helicase and the lagging-strand polymerase that are proceeding downstream to reinitiate synthesis. PMID:24268579

  8. Reconstitution of a eukaryotic replisome reveals suppression mechanisms that define leading/lagging strand operation

    PubMed Central

    Georgescu, Roxana E; Schauer, Grant D; Yao, Nina Y; Langston, Lance D; Yurieva, Olga; Zhang, Dan; Finkelstein, Jeff; O'Donnell, Mike E

    2015-01-01

    We have reconstituted a eukaryotic leading/lagging strand replisome comprising 31 distinct polypeptides. This study identifies a process unprecedented in bacterial replisomes. While bacteria and phage simply recruit polymerases to the fork, we find that suppression mechanisms are used to position the distinct eukaryotic polymerases on their respective strands. Hence, Pol ε is active with CMG on the leading strand, but it is unable to function on the lagging strand, even when Pol δ is not present. Conversely, Pol δ-PCNA is the only enzyme capable of extending Okazaki fragments in the presence of Pols ε and α. We have shown earlier that Pol δ-PCNA is suppressed on the leading strand with CMG (Georgescu et al., 2014). We propose that CMG, the 11-subunit helicase, is responsible for one or both of these suppression mechanisms that spatially control polymerase occupancy at the fork. DOI: http://dx.doi.org/10.7554/eLife.04988.001 PMID:25871847

  9. The rate of polymerase release upon filling the gap between Okazaki fragments is inadequate to support cycling during lagging strand synthesis.

    PubMed

    Dohrmann, Paul R; Manhart, Carol M; Downey, Christopher D; McHenry, Charles S

    2011-11-18

    Upon completion of synthesis of an Okazaki fragment, the lagging strand replicase must recycle to the next primer at the replication fork in under 0.1 s to sustain the physiological rate of DNA synthesis. We tested the collision model that posits that cycling is triggered by the polymerase encountering the 5'-end of the preceding Okazaki fragment. Probing with surface plasmon resonance, DNA polymerase III holoenzyme initiation complexes were formed on an immobilized gapped template. Initiation complexes exhibit a half-life of dissociation of approximately 15 min. Reduction in gap size to 1 nt increased the rate of dissociation 2.5-fold, and complete filling of the gap increased the off-rate an additional 3-fold (t(1/2)~2 min). An exogenous primed template and ATP accelerated dissociation an additional 4-fold in a reaction that required complete filling of the gap. Neither a 5'-triphosphate nor a 5'-RNA terminated oligonucleotide downstream of the polymerase accelerated dissociation further. Thus, the rate of polymerase release upon gap completion and collision with a downstream Okazaki fragment is 1000-fold too slow to support an adequate rate of cycling and likely provides a backup mechanism to enable polymerase release when the other cycling signals are absent. Kinetic measurements indicate that addition of the last nucleotide to fill the gap is not the rate-limiting step for polymerase release and cycling. Modest (approximately 7 nt) strand displacement is observed after the gap between model Okazaki fragments is filled. To determine the identity of the protein that senses gap filling to modulate affinity of the replicase for the template, we performed photo-cross-linking experiments with highly reactive and non-chemoselective diazirines. Only the α subunit cross-linked, indicating that it serves as the sensor. Copyright © 2011 Elsevier Ltd. All rights reserved.

  10. Arranging eukaryotic nuclear DNA polymerases for replication: Specific interactions with accessory proteins arrange Pols α, δ, and ϵ in the replisome for leading-strand and lagging-strand DNA replication.

    PubMed

    Kunkel, Thomas A; Burgers, Peter M J

    2017-08-01

    Biochemical and cryo-electron microscopy studies have just been published revealing interactions among proteins of the yeast replisome that are important for highly coordinated synthesis of the two DNA strands of the nuclear genome. These studies reveal key interactions important for arranging DNA polymerases α, δ, and ϵ for leading and lagging strand replication. The CMG (Mcm2-7, Cdc45, GINS) helicase is central to this interaction network. These are but the latest examples of elegant studies performed in the recent past that lead to a much better understanding of how the eukaryotic replication fork achieves efficient DNA replication that is accurate enough to prevent diseases yet allows evolution. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

  11. RPA prevents G-rich structure formation at lagging-strand telomeres to allow maintenance of chromosome ends.

    PubMed

    Audry, Julien; Maestroni, Laetitia; Delagoutte, Emmanuelle; Gauthier, Tiphaine; Nakamura, Toru M; Gachet, Yannick; Saintomé, Carole; Géli, Vincent; Coulon, Stéphane

    2015-07-14

    Replication protein A (RPA) is a highly conserved heterotrimeric single-stranded DNA-binding protein involved in DNA replication, recombination, and repair. In fission yeast, the Rpa1-D223Y mutation provokes telomere shortening. Here, we show that this mutation impairs lagging-strand telomere replication and leads to the accumulation of secondary structures and recruitment of the homologous recombination factor Rad52. The presence of these secondary DNA structures correlates with reduced association of shelterin subunits Pot1 and Ccq1 at telomeres. Strikingly, heterologous expression of the budding yeast Pif1 known to efficiently unwind G-quadruplex rescues all the telomeric defects of the D223Y cells. Furthermore, in vitro data show that the identical D to Y mutation in human RPA specifically affects its ability to bind G-quadruplex. We propose that RPA prevents the formation of G-quadruplex structures at lagging-strand telomeres to promote shelterin association and facilitate telomerase action at telomeres. © 2015 The Authors.

  12. Cryo-EM structure of Mcm2-7 double hexamer on DNA suggests a lagging-strand DNA extrusion model.

    PubMed

    Noguchi, Yasunori; Yuan, Zuanning; Bai, Lin; Schneider, Sarah; Zhao, Gongpu; Stillman, Bruce; Speck, Christian; Li, Huilin

    2017-11-07

    During replication initiation, the core component of the helicase-the Mcm2-7 hexamer-is loaded on origin DNA as a double hexamer (DH). The two ring-shaped hexamers are staggered, leading to a kinked axial channel. How the origin DNA interacts with the axial channel is not understood, but the interaction could provide key insights into Mcm2-7 function and regulation. Here, we report the cryo-EM structure of the Mcm2-7 DH on dsDNA and show that the DNA is zigzagged inside the central channel. Several of the Mcm subunit DNA-binding loops, such as the oligosaccharide-oligonucleotide loops, helix 2 insertion loops, and presensor 1 (PS1) loops, are well defined, and many of them interact extensively with the DNA. The PS1 loops of Mcm 3, 4, 6, and 7, but not 2 and 5, engage the lagging strand with an approximate step size of one base per subunit. Staggered coupling of the two opposing hexamers positions the DNA right in front of the two Mcm2-Mcm5 gates, with each strand being pressed against one gate. The architecture suggests that lagging-strand extrusion initiates in the middle of the DH that is composed of the zinc finger domains of both hexamers. To convert the Mcm2-7 DH structure into the Mcm2-7 hexamer structure found in the active helicase, the N-tier ring of the Mcm2-7 hexamer in the DH-dsDNA needs to tilt and shift laterally. We suggest that these N-tier ring movements cause the DNA strand separation and lagging-strand extrusion. Copyright © 2017 the Author(s). Published by PNAS.

  13. Identification of cis-acting elements on positive-strand subgenomic mRNA required for the synthesis of negative-strand counterpart in bovine coronavirus.

    PubMed

    Yeh, Po-Yuan; Wu, Hung-Yi

    2014-07-30

    It has been demonstrated that, in addition to genomic RNA, sgmRNA is able to serve as a template for the synthesis of the negative-strand [(-)-strand] complement. However, the cis-acting elements on the positive-strand [(+)-strand] sgmRNA required for (-)-strand sgmRNA synthesis have not yet been systematically identified. In this study, we employed real-time quantitative reverse transcription polymerase chain reaction to analyze the cis-acting elements on bovine coronavirus (BCoV) sgmRNA 7 required for the synthesis of its (-)-strand counterpart by deletion mutagenesis. The major findings are as follows. (1) Deletion of the 5'-terminal leader sequence on sgmRNA 7 decreased the synthesis of the (-)-strand sgmRNA complement. (2) Deletions of the 3' untranslated region (UTR) bulged stem-loop showed no effect on (-)-strand sgmRNA synthesis; however, deletion of the 3' UTR pseudoknot decreased the yield of (-)-strand sgmRNA. (3) Nucleotides positioned from -15 to -34 of the sgmRNA 7 3'-terminal region are required for efficient (-)-strand sgmRNA synthesis. (4) Nucleotide species at the 3'-most position (-1) of sgmRNA 7 is correlated to the efficiency of (-)-strand sgmRNA synthesis. These results together suggest, in principle, that the 5'- and 3'-terminal sequences on sgmRNA 7 harbor cis-acting elements are critical for efficient (-)-strand sgmRNA synthesis in BCoV.

  14. Strand displacement synthesis by yeast DNA polymerase ε.

    PubMed

    Ganai, Rais A; Zhang, Xiao-Ping; Heyer, Wolf-Dietrich; Johansson, Erik

    2016-09-30

    DNA polymerase ε (Pol ε) is a replicative DNA polymerase with an associated 3'-5' exonuclease activity. Here, we explored the capacity of Pol ε to perform strand displacement synthesis, a process that influences many DNA transactions in vivo We found that Pol ε is unable to carry out extended strand displacement synthesis unless its 3'-5' exonuclease activity is removed. However, the wild-type Pol ε holoenzyme efficiently displaced one nucleotide when encountering double-stranded DNA after filling a gap or nicked DNA. A flap, mimicking a D-loop or a hairpin structure, on the 5' end of the blocking primer inhibited Pol ε from synthesizing DNA up to the fork junction. This inhibition was observed for Pol ε but not with Pol δ, RB69 gp43 or Pol η. Neither was Pol ε able to extend a D-loop in reconstitution experiments. Finally, we show that the observed strand displacement synthesis by exonuclease-deficient Pol ε is distributive. Our results suggest that Pol ε is unable to extend the invading strand in D-loops during homologous recombination or to add more than two nucleotides during long-patch base excision repair. Our results support the hypothesis that Pol ε participates in short-patch base excision repair and ribonucleotide excision repair. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  15. Strand displacement synthesis by yeast DNA polymerase ε

    PubMed Central

    Ganai, Rais A.; Zhang, Xiao-Ping; Heyer, Wolf-Dietrich; Johansson, Erik

    2016-01-01

    DNA polymerase ε (Pol ε) is a replicative DNA polymerase with an associated 3′–5′ exonuclease activity. Here, we explored the capacity of Pol ε to perform strand displacement synthesis, a process that influences many DNA transactions in vivo. We found that Pol ε is unable to carry out extended strand displacement synthesis unless its 3′–5′ exonuclease activity is removed. However, the wild-type Pol ε holoenzyme efficiently displaced one nucleotide when encountering double-stranded DNA after filling a gap or nicked DNA. A flap, mimicking a D-loop or a hairpin structure, on the 5′ end of the blocking primer inhibited Pol ε from synthesizing DNA up to the fork junction. This inhibition was observed for Pol ε but not with Pol δ, RB69 gp43 or Pol η. Neither was Pol ε able to extend a D-loop in reconstitution experiments. Finally, we show that the observed strand displacement synthesis by exonuclease-deficient Pol ε is distributive. Our results suggest that Pol ε is unable to extend the invading strand in D-loops during homologous recombination or to add more than two nucleotides during long-patch base excision repair. Our results support the hypothesis that Pol ε participates in short-patch base excision repair and ribonucleotide excision repair. PMID:27325747

  16. CMG helicase and DNA polymerase ε form a functional 15-subunit holoenzyme for eukaryotic leading-strand DNA replication.

    PubMed

    Langston, Lance D; Zhang, Dan; Yurieva, Olga; Georgescu, Roxana E; Finkelstein, Jeff; Yao, Nina Y; Indiani, Chiara; O'Donnell, Mike E

    2014-10-28

    DNA replication in eukaryotes is asymmetric, with separate DNA polymerases (Pol) dedicated to bulk synthesis of the leading and lagging strands. Pol α/primase initiates primers on both strands that are extended by Pol ε on the leading strand and by Pol δ on the lagging strand. The CMG (Cdc45-MCM-GINS) helicase surrounds the leading strand and is proposed to recruit Pol ε for leading-strand synthesis, but to date a direct interaction between CMG and Pol ε has not been demonstrated. While purifying CMG helicase overexpressed in yeast, we detected a functional complex between CMG and native Pol ε. Using pure CMG and Pol ε, we reconstituted a stable 15-subunit CMG-Pol ε complex and showed that it is a functional polymerase-helicase on a model replication fork in vitro. On its own, the Pol2 catalytic subunit of Pol ε is inefficient in CMG-dependent replication, but addition of the Dpb2 protein subunit of Pol ε, known to bind the Psf1 protein subunit of CMG, allows stable synthesis with CMG. Dpb2 does not affect Pol δ function with CMG, and thus we propose that the connection between Dpb2 and CMG helps to stabilize Pol ε on the leading strand as part of a 15-subunit leading-strand holoenzyme we refer to as CMGE. Direct binding between Pol ε and CMG provides an explanation for specific targeting of Pol ε to the leading strand and provides clear mechanistic evidence for how strand asymmetry is maintained in eukaryotes.

  17. CMG helicase and DNA polymerase ε form a functional 15-subunit holoenzyme for eukaryotic leading-strand DNA replication

    PubMed Central

    Langston, Lance D.; Zhang, Dan; Yurieva, Olga; Georgescu, Roxana E.; Finkelstein, Jeff; Yao, Nina Y.; Indiani, Chiara; O’Donnell, Mike E.

    2014-01-01

    DNA replication in eukaryotes is asymmetric, with separate DNA polymerases (Pol) dedicated to bulk synthesis of the leading and lagging strands. Pol α/primase initiates primers on both strands that are extended by Pol ε on the leading strand and by Pol δ on the lagging strand. The CMG (Cdc45-MCM-GINS) helicase surrounds the leading strand and is proposed to recruit Pol ε for leading-strand synthesis, but to date a direct interaction between CMG and Pol ε has not been demonstrated. While purifying CMG helicase overexpressed in yeast, we detected a functional complex between CMG and native Pol ε. Using pure CMG and Pol ε, we reconstituted a stable 15-subunit CMG–Pol ε complex and showed that it is a functional polymerase–helicase on a model replication fork in vitro. On its own, the Pol2 catalytic subunit of Pol ε is inefficient in CMG-dependent replication, but addition of the Dpb2 protein subunit of Pol ε, known to bind the Psf1 protein subunit of CMG, allows stable synthesis with CMG. Dpb2 does not affect Pol δ function with CMG, and thus we propose that the connection between Dpb2 and CMG helps to stabilize Pol ε on the leading strand as part of a 15-subunit leading-strand holoenzyme we refer to as CMGE. Direct binding between Pol ε and CMG provides an explanation for specific targeting of Pol ε to the leading strand and provides clear mechanistic evidence for how strand asymmetry is maintained in eukaryotes. PMID:25313033

  18. The importance of becoming double-stranded: Innate immunity and the kinetic model of HIV-1 central plus strand synthesis

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

    Poeschla, Eric, E-mail: poeschla.eric@mayo.edu

    Central initiation of plus strand synthesis is a conserved feature of lentiviruses and certain other retroelements. This complication of the standard reverse transcription mechanism produces a transient “central DNA flap” in the viral cDNA, which has been proposed to mediate its subsequent nuclear import. This model has assumed that the important feature is the flapped DNA structure itself rather than the process that produces it. Recently, an alternative kinetic model was proposed. It posits that central plus strand synthesis functions to accelerate conversion to the double-stranded state, thereby helping HIV-1 to evade single-strand DNA-targeting antiviral restrictions such as APOBEC3 proteins,more » and perhaps to avoid innate immune sensor mechanisms. The model is consistent with evidence that lentiviruses must often synthesize their cDNAs when dNTP concentrations are limiting and with data linking reverse transcription and uncoating. There may be additional kinetic advantages for the artificial genomes of lentiviral gene therapy vectors. - Highlights: • Two main functional models for HIV central plus strand synthesis have been proposed. • In one, a transient central DNA flap in the viral cDNA mediates HIV-1 nuclear import. • In the other, multiple kinetic consequences are emphasized. • One is defense against APOBEC3G, which deaminates single-stranded DNA. • Future questions pertain to antiviral restriction, uncoating and nuclear import.« less

  19. Regulation of yeast DNA polymerase δ-mediated strand displacement synthesis by 5′-flaps

    PubMed Central

    Koc, Katrina N.; Stodola, Joseph L.; Burgers, Peter M.; Galletto, Roberto

    2015-01-01

    The strand displacement activity of DNA polymerase δ is strongly stimulated by its interaction with proliferating cell nuclear antigen (PCNA). However, inactivation of the 3′–5′ exonuclease activity is sufficient to allow the polymerase to carry out strand displacement even in the absence of PCNA. We have examined in vitro the basic biochemical properties that allow Pol δ-exo− to carry out strand displacement synthesis and discovered that it is regulated by the 5′-flaps in the DNA strand to be displaced. Under conditions where Pol δ carries out strand displacement synthesis, the presence of long 5′-flaps or addition in trans of ssDNA suppress this activity. This suggests the presence of a secondary DNA binding site on the enzyme that is responsible for modulation of strand displacement activity. The inhibitory effect of a long 5′-flap can be suppressed by its interaction with single-stranded DNA binding proteins. However, this relief of flap-inhibition does not simply originate from binding of Replication Protein A to the flap and sequestering it. Interaction of Pol δ with PCNA eliminates flap-mediated inhibition of strand displacement synthesis by masking the secondary DNA site on the polymerase. These data suggest that in addition to enhancing the processivity of the polymerase PCNA is an allosteric modulator of other Pol δ activities. PMID:25813050

  20. Programmable autonomous synthesis of single-stranded DNA

    NASA Astrophysics Data System (ADS)

    Kishi, Jocelyn Y.; Schaus, Thomas E.; Gopalkrishnan, Nikhil; Xuan, Feng; Yin, Peng

    2018-02-01

    DNA performs diverse functional roles in biology, nanotechnology and biotechnology, but current methods for autonomously synthesizing arbitrary single-stranded DNA are limited. Here, we introduce the concept of primer exchange reaction (PER) cascades, which grow nascent single-stranded DNA with user-specified sequences following prescribed reaction pathways. PER synthesis happens in a programmable, autonomous, in situ and environmentally responsive fashion, providing a platform for engineering molecular circuits and devices with a wide range of sensing, monitoring, recording, signal-processing and actuation capabilities. We experimentally demonstrate a nanodevice that transduces the detection of a trigger RNA into the production of a DNAzyme that degrades an independent RNA substrate, a signal amplifier that conditionally synthesizes long fluorescent strands only in the presence of a particular RNA signal, molecular computing circuits that evaluate logic (AND, OR, NOT) combinations of RNA inputs, and a temporal molecular event recorder that records in the PER transcript the order in which distinct RNA inputs are sequentially detected.

  1. Programmable autonomous synthesis of single-stranded DNA.

    PubMed

    Kishi, Jocelyn Y; Schaus, Thomas E; Gopalkrishnan, Nikhil; Xuan, Feng; Yin, Peng

    2018-02-01

    DNA performs diverse functional roles in biology, nanotechnology and biotechnology, but current methods for autonomously synthesizing arbitrary single-stranded DNA are limited. Here, we introduce the concept of primer exchange reaction (PER) cascades, which grow nascent single-stranded DNA with user-specified sequences following prescribed reaction pathways. PER synthesis happens in a programmable, autonomous, in situ and environmentally responsive fashion, providing a platform for engineering molecular circuits and devices with a wide range of sensing, monitoring, recording, signal-processing and actuation capabilities. We experimentally demonstrate a nanodevice that transduces the detection of a trigger RNA into the production of a DNAzyme that degrades an independent RNA substrate, a signal amplifier that conditionally synthesizes long fluorescent strands only in the presence of a particular RNA signal, molecular computing circuits that evaluate logic (AND, OR, NOT) combinations of RNA inputs, and a temporal molecular event recorder that records in the PER transcript the order in which distinct RNA inputs are sequentially detected.

  2. BLM helicase facilitates telomere replication during leading strand synthesis of telomeres

    PubMed Central

    Kosiyatrakul, Settapong T.

    2015-01-01

    Based on its in vitro unwinding activity on G-quadruplex (G4) DNA, the Bloom syndrome–associated helicase BLM is proposed to participate in telomere replication by aiding fork progression through G-rich telomeric DNA. Single molecule analysis of replicated DNA (SMARD) was used to determine the contribution of BLM helicase to telomere replication. In BLM-deficient cells, replication forks initiating from origins within the telomere, which copy the G-rich strand by leading strand synthesis, moved slower through the telomere compared with the adjacent subtelomere. Fork progression through the telomere was further slowed in the presence of a G4 stabilizer. Using a G4-specific antibody, we found that deficiency of BLM, or another G4-unwinding helicase, the Werner syndrome-associated helicase WRN, resulted in increased G4 structures in cells. Importantly, deficiency of either helicase led to greater increases in G4 DNA detected in the telomere compared with G4 seen genome-wide. Collectively, our findings are consistent with BLM helicase facilitating telomere replication by resolving G4 structures formed during copying of the G-rich strand by leading strand synthesis. PMID:26195664

  3. Process of infection with bacteriophage phi chi 174. XL. Viral DNA replication of phi chi 174 mutants blocked in progeny single-stranded DNA synthesis.

    PubMed Central

    Fukuda, A; Sinsheimer, R L

    1976-01-01

    Mutation in several different cistrons of bacteriophage phi chi 174 blocks net progeny single-stranded DNA synthesis at the late period of infection (15). For the study of the functions of these cistrons in single-stranded DNA synthesis, asymmetric replication of replicative form DNA was examined at the late period of infection with amber mutants of these cistrons. While the normal, rapid process of asymmetric single-stranded viral DNA synthesis is blocked at the late period of these mutant infections, an asymmetric synthesis of the viral strand of replicative-form DNA is observed in this period, though at a reduced level, together with degradation of prelabeled viral strand. Some intermediate replicative-form molecules were also detected. Asymmetric synthesis of the viral strand of replicative-form DNA at the late period of phi chi infection is completely inhibited in the presence of a low concentration (35mug/ml) of chloramphenicol (which also blocks net single-stranded viral DNA synthesis). These results are discussed in terms of the possible role of the specific viral proteins for normal single-stranded DNA synthesis. PMID:1255871

  4. Human Cell Assays for Synthesis-Dependent Strand Annealing and Crossing over During Double-Strand Break Repair.

    PubMed

    Zapotoczny, Grzegorz; Sekelsky, Jeff

    2017-04-03

    DNA double-strand breaks (DSBs) are one of the most deleterious types of lesions to the genome. Synthesis-dependent strand annealing (SDSA) is thought to be a major pathway of DSB repair, but direct tests of this model have only been conducted in budding yeast and Drosophila To better understand this pathway, we developed an SDSA assay for use in human cells. Our results support the hypothesis that SDSA is an important DSB repair mechanism in human cells. We used siRNA knockdown to assess the roles of a number of helicases suggested to promote SDSA. None of the helicase knockdowns reduced SDSA, but knocking down BLM or RTEL1 increased SDSA. Molecular analysis of repair products suggests that these helicases may prevent long-tract repair synthesis. Since the major alternative to SDSA (repair involving a double-Holliday junction intermediate) can lead to crossovers, we also developed a fluorescent assay that detects crossovers generated during DSB repair. Together, these assays will be useful in investigating features and mechanisms of SDSA and crossover pathways in human cells. Copyright © 2017 Zapotoczny and Sekelsky.

  5. Human Cell Assays for Synthesis-Dependent Strand Annealing and Crossing over During Double-Strand Break Repair

    PubMed Central

    Zapotoczny, Grzegorz; Sekelsky, Jeff

    2017-01-01

    DNA double-strand breaks (DSBs) are one of the most deleterious types of lesions to the genome. Synthesis-dependent strand annealing (SDSA) is thought to be a major pathway of DSB repair, but direct tests of this model have only been conducted in budding yeast and Drosophila. To better understand this pathway, we developed an SDSA assay for use in human cells. Our results support the hypothesis that SDSA is an important DSB repair mechanism in human cells. We used siRNA knockdown to assess the roles of a number of helicases suggested to promote SDSA. None of the helicase knockdowns reduced SDSA, but knocking down BLM or RTEL1 increased SDSA. Molecular analysis of repair products suggests that these helicases may prevent long-tract repair synthesis. Since the major alternative to SDSA (repair involving a double-Holliday junction intermediate) can lead to crossovers, we also developed a fluorescent assay that detects crossovers generated during DSB repair. Together, these assays will be useful in investigating features and mechanisms of SDSA and crossover pathways in human cells. PMID:28179392

  6. The RNA synthesis machinery of negative-stranded RNA viruses

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

    Ortín, Juan, E-mail: jortin@cnb.csic.es; Martín-Benito, Jaime, E-mail: jmartinb@cnb.csic.es

    The group of Negative-Stranded RNA Viruses (NSVs) includes many human pathogens, like the influenza, measles, mumps, respiratory syncytial or Ebola viruses, which produce frequent epidemics of disease and occasional, high mortality outbreaks by transmission from animal reservoirs. The genome of NSVs consists of one to several single-stranded, negative-polarity RNA molecules that are always assembled into mega Dalton-sized complexes by association to many nucleoprotein monomers. These RNA-protein complexes or ribonucleoproteins function as templates for transcription and replication by action of the viral RNA polymerase and accessory proteins. Here we review our knowledge on these large RNA-synthesis machines, including the structure ofmore » their components, the interactions among them and their enzymatic activities, and we discuss models showing how they perform the virus transcription and replication programmes. - Highlights: • Overall organisation of NSV RNA synthesis machines. • Structure and function of the ribonucleoprotein components: Atomic structure of the RNA polymerase complex. • Commonalities and differences between segmented- and non-segmented NSVs. • Transcription versus replication programmes.« less

  7. Genomic mapping of single-stranded DNA in hydroxyurea-challenged yeasts identifies origins of replication.

    PubMed

    Feng, Wenyi; Collingwood, David; Boeck, Max E; Fox, Lindsay A; Alvino, Gina M; Fangman, Walton L; Raghuraman, Mosur K; Brewer, Bonita J

    2006-02-01

    During DNA replication one or both strands transiently become single stranded: first at the sites where initiation of DNA synthesis occurs (known as origins of replication) and subsequently on the lagging strands of replication forks as discontinuous Okazaki fragments are generated. We report a genome-wide analysis of single-stranded DNA (ssDNA) formation in the presence of hydroxyurea during DNA replication in wild-type and checkpoint-deficient rad53 Saccharomyces cerevisiae cells. In wild-type cells, ssDNA was first observed at a subset of replication origins and later 'migrated' bi-directionally, suggesting that ssDNA formation is associated with continuously moving replication forks. In rad53 cells, ssDNA was observed at virtually every known origin, but remained there over time, suggesting that replication forks stall. Telomeric regions seemed to be particularly sensitive to the loss of Rad53 checkpoint function. Replication origins in Schizosaccharomyces pombe were also mapped using our method.

  8. Strand displacement by DNA polymerase III occurs through a tau-psi-chi link to single-stranded DNA-binding protein coating the lagging strand template.

    PubMed

    Yuan, Quan; McHenry, Charles S

    2009-11-13

    In addition to the well characterized processive replication reaction catalyzed by the DNA polymerase III holoenzyme on single-stranded DNA templates, the enzyme possesses an intrinsic strand displacement activity on flapped templates. The strand displacement activity is distinguished from the single-stranded DNA-templated reaction by a high dependence upon single-stranded DNA binding protein and an inability of gamma-complex to support the reaction in the absence of tau. However, if gamma-complex is present to load beta(2), a truncated tau protein containing only domains III-V will suffice. This truncated protein is sufficient to bind both the alpha subunit of DNA polymerase (Pol) III and chipsi. This is reminiscent of the minimal requirements for Pol III to replicate short single-stranded DNA-binding protein (SSB)-coated templates where tau is only required to serve as a scaffold to hold Pol III and chi in the same complex (Glover, B., and McHenry, C. (1998) J. Biol. Chem. 273, 23476-23484). We propose a model in which strand displacement by DNA polymerase III holoenzyme depends upon a Pol III-tau-psi-chi-SSB binding network, where SSB is bound to the displaced strand, stabilizing the Pol III-template interaction. The same interaction network is probably important for stabilizing the leading strand polymerase interactions with authentic replication forks. The specificity constant (k(cat)/K(m)) for the strand displacement reaction is approximately 300-fold less favorable than reactions on single-stranded templates and proceeds with a slower rate (150 nucleotides/s) and only moderate processivity (approximately 300 nucleotides). PriA, the initiator of replication restart on collapsed or misassembled replication forks, blocks the strand displacement reaction, even if added to an ongoing reaction.

  9. Unveiling the mystery of mitochondrial DNA replication in yeasts.

    PubMed

    Chen, Xin Jie; Clark-Walker, George Desmond

    2018-01-01

    Conventional DNA replication is initiated from specific origins and requires the synthesis of RNA primers for both the leading and lagging strands. In contrast, the replication of yeast mitochondrial DNA is origin-independent. The replication of the leading strand is likely primed by recombinational structures and proceeded by a rolling circle mechanism. The coexistent linear and circular DNA conformers facilitate the recombination-based initiation. The replication of the lagging strand is poorly understood. Re-evaluation of published data suggests that the rolling circle may also provide structures for the synthesis of the lagging-strand by mechanisms such as template switching. Thus, the coupling of recombination with rolling circle replication and possibly, template switching, may have been selected as an economic replication mode to accommodate the reductive evolution of mitochondria. Such a replication mode spares the need for conventional replicative components, including those required for origin recognition/remodelling, RNA primer synthesis and lagging-strand processing. Copyright © 2017 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

  10. RNA-primed complementary-sense DNA synthesis of the geminivirus African cassava mosaic virus.

    PubMed Central

    Saunders, K; Lucy, A; Stanley, J

    1992-01-01

    The plant DNA virus African cassava mosaic virus (ACMV) is believed to replicate by a rolling circle mechanism. To investigate complementary-sense DNA (lagging strand) synthesis, we have analysed the heterogenous form of complementary-sense DNA (H3 DNA) from infected Nicotiana benthamiana by two-dimensional agarose gel electrophoresis and blot hybridisation. The presence of an RNA moeity is demonstrated by comparison of results for nucleic acids resolved on neutral/alkaline and neutral/formamide gels, suggesting that complementary-sense DNA synthesis on the virus-sense single-stranded DNA template is preceded by the synthesis of an RNA primer. Hybridisation with probes to specific parts of ACMV DNA A genome indicates that synthesis of the putative RNA primer initiates between nucleotides 2581-221, a region that includes intergenic sequences that have been implicated in geminivirus DNA replication and the control of gene expression. Images PMID:1475192

  11. Correlated Template-Switching Events during Minus-Strand DNA Synthesis: a Mechanism for High Negative Interference during Retroviral Recombination

    PubMed Central

    Anderson, Jeffrey A.; Teufel, Ronald J.; Yin, Philip D.; Hu, Wei-Shau

    1998-01-01

    Two models for the mechanism of retroviral recombination have been proposed: forced copy choice (minus-strand recombination) and strand displacement-assimilation (plus-strand recombination). Each minus-strand recombination event results in one template switch, whereas each plus-strand recombination event results in two template switches. Recombinant proviruses with one and more than one template switches were previously observed. Recombinants with one template switch were generated by minus-strand recombination, while recombinants containing more than one template switch may have been generated by plus-strand recombination or by correlated minus-strand recombination. We recently observed that retroviral recombination exhibits high negative interference whereby the frequency of recombinants containing multiple template-switching events is higher than expected. To delineate the mechanism that generates recombinants with more than one template switch, we devised a system that permits only minus-strand recombination. Two highly homologous vectors, WH204 and WH221, containing eight different restriction site markers were used. The primer binding site (PBS) of WH221 was deleted; although reverse transcription cannot initiate from WH221 RNA, it can serve as a template for DNA synthesis in heterozygotic virions. After one round of retroviral replication, the structures of the recombinant proviruses were examined. Recombinants containing two, three, four, and five template switches were observed at 1.4-, 10-, 65-, and 50-fold-higher frequencies, respectively, than expected. This indicates that minus-strand recombination events are correlated and can generate proviruses with multiple template switches efficiently. The frequencies of recombinants containing multiple template switches were similar to those observed in the previous system, which allowed both minus- and plus-strand recombination. Thus, the previously reported high negative interference during retroviral

  12. Analysis of Duck Hepatitis B Virus Reverse Transcription Indicates a Common Mechanism for the Two Template Switches during Plus-Strand DNA Synthesis

    PubMed Central

    Havert, Michael B.; Ji, Lin; Loeb, Daniel D.

    2002-01-01

    The synthesis of the hepadnavirus relaxed circular DNA genome requires two template switches, primer translocation and circularization, during plus-strand DNA synthesis. Repeated sequences serve as donor and acceptor templates for these template switches, with direct repeat 1 (DR1) and DR2 for primer translocation and 5′r and 3′r for circularization. These donor and acceptor sequences are at, or near, the ends of the minus-strand DNA. Analysis of plus-strand DNA synthesis of duck hepatitis B virus (DHBV) has indicated that there are at least three other cis-acting sequences that make contributions during the synthesis of relaxed circular DNA. These sequences, 5E, M, and 3E, are located near the 5′ end, the middle, and the 3′ end of minus-strand DNA, respectively. The mechanism by which these sequences contribute to the synthesis of plus-strand DNA was unclear. Our aim was to better understand the mechanism by which 5E and M act. We localized the DHBV 5E element to a short sequence of approximately 30 nucleotides that is 100 nucleotides 3′ of DR2 on minus-strand DNA. We found that the new 5E mutants were partially defective for primer translocation/utilization at DR2. They were also invariably defective for circularization. In addition, examination of several new DHBV M variants indicated that they too were defective for primer translocation/utilization and circularization. Thus, this analysis indicated that 5E and M play roles in both primer translocation/utilization and circularization. In conjunction with earlier findings that 3E functions in both template switches, our findings indicate that the processes of primer translocation and circularization share a common underlying mechanism. PMID:11861843

  13. Role of the Pepino mosaic virus 3'-untranslated region elements in negative-strand RNA synthesis in vitro.

    PubMed

    Osman, Toba A M; Olsthoorn, René C L; Livieratos, Ioannis C

    2014-09-22

    Pepino mosaic virus (PepMV) is a mechanically-transmitted positive-strand RNA potexvirus, with a 6410 nt long single-stranded (ss) RNA genome flanked by a 5'-methylguanosine cap and a 3' poly-A tail. Computer-assisted folding of the 64 nt long PepMV 3'-untranslated region (UTR) resulted in the prediction of three stem-loop structures (hp1, hp2, and hp3 in the 3'-5' direction). The importance of these structures and/or sequences for promotion of negative-strand RNA synthesis and binding to the RNA dependent RNA polymerase (RdRp) was tested in vitro using a specific RdRp assay. Hp1, which is highly variable among different PepMV isolates, appeared dispensable for negative-strand synthesis. Hp2, which is characterized by a large U-rich loop, tolerated base-pair changes in its stem as long as they maintained the stem integrity but was very sensitive to changes in the U-rich loop. Hp3, which harbours the conserved potexvirus ACUUAA hexamer motif, was essential for template activity. Template-RNA polymerase binding competition experiments showed that the ACUUAA sequence represents a high-affinity RdRp binding element. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. RNA primer–primase complexes serve as the signal for polymerase recycling and Okazaki fragment initiation in T4 phage DNA replication

    PubMed Central

    Spiering, Michelle M.; Hanoian, Philip; Gannavaram, Swathi; Benkovic, Stephen J.

    2017-01-01

    The opposite strand polarity of duplex DNA necessitates that the leading strand is replicated continuously whereas the lagging strand is replicated in discrete segments known as Okazaki fragments. The lagging-strand polymerase sometimes recycles to begin the synthesis of a new Okazaki fragment before finishing the previous fragment, creating a gap between the Okazaki fragments. The mechanism and signal that initiate this behavior—that is, the signaling mechanism—have not been definitively identified. We examined the role of RNA primer–primase complexes left on the lagging ssDNA from primer synthesis in initiating early lagging-strand polymerase recycling. We show for the T4 bacteriophage DNA replication system that primer–primase complexes have a residence time similar to the timescale of Okazaki fragment synthesis and the ability to block a holoenzyme synthesizing DNA and stimulate the dissociation of the holoenzyme to trigger polymerase recycling. The collision with primer–primase complexes triggering the early termination of Okazaki fragment synthesis has distinct advantages over those previously proposed because this signal requires no transmission to the lagging-strand polymerase through protein or DNA interactions, the mechanism for rapid dissociation of the holoenzyme is always collision, and no unique characteristics need to be assigned to either identical polymerase in the replisome. We have modeled repeated cycles of Okazaki fragment initiation using a collision with a completed Okazaki fragment or primer–primase complexes as the recycling mechanism. The results reproduce experimental data, providing insights into events related to Okazaki fragment initiation and the overall functioning of DNA replisomes. PMID:28507156

  15. RNA primer-primase complexes serve as the signal for polymerase recycling and Okazaki fragment initiation in T4 phage DNA replication.

    PubMed

    Spiering, Michelle M; Hanoian, Philip; Gannavaram, Swathi; Benkovic, Stephen J

    2017-05-30

    The opposite strand polarity of duplex DNA necessitates that the leading strand is replicated continuously whereas the lagging strand is replicated in discrete segments known as Okazaki fragments. The lagging-strand polymerase sometimes recycles to begin the synthesis of a new Okazaki fragment before finishing the previous fragment, creating a gap between the Okazaki fragments. The mechanism and signal that initiate this behavior-that is, the signaling mechanism-have not been definitively identified. We examined the role of RNA primer-primase complexes left on the lagging ssDNA from primer synthesis in initiating early lagging-strand polymerase recycling. We show for the T4 bacteriophage DNA replication system that primer-primase complexes have a residence time similar to the timescale of Okazaki fragment synthesis and the ability to block a holoenzyme synthesizing DNA and stimulate the dissociation of the holoenzyme to trigger polymerase recycling. The collision with primer-primase complexes triggering the early termination of Okazaki fragment synthesis has distinct advantages over those previously proposed because this signal requires no transmission to the lagging-strand polymerase through protein or DNA interactions, the mechanism for rapid dissociation of the holoenzyme is always collision, and no unique characteristics need to be assigned to either identical polymerase in the replisome. We have modeled repeated cycles of Okazaki fragment initiation using a collision with a completed Okazaki fragment or primer-primase complexes as the recycling mechanism. The results reproduce experimental data, providing insights into events related to Okazaki fragment initiation and the overall functioning of DNA replisomes.

  16. The N-terminal strand modulates immunoglobulin light chain fibrillogenesis

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

    Pozo-Yauner, Luis del, E-mail: ldelpozo@inmegen.gob.mx; Wall, Jonathan S.; González Andrade, Martín

    2014-01-10

    Highlights: •We evaluated the impact of mutations in the N-terminal strand of 6aJL2 protein. •Mutations destabilized the protein in a position-dependent manner. •Destabilizing mutations accelerated the fibrillogenesis by shortening the lag time. •The effect on the kinetic of fibril elongation by seeding was of different nature. •The N-terminal strand is buried in the fibrillar state of 6aJL2 protein. -- Abstract: It has been suggested that the N-terminal strand of the light chain variable domain (V{sub L}) protects the molecule from aggregation by hindering spurious intermolecular contacts. We evaluated the impact of mutations in the N-terminal strand on the thermodynamic stabilitymore » and kinetic of fibrillogenesis of the V{sub L} protein 6aJL2. Mutations in this strand destabilized the protein in a position-dependent manner, accelerating the fibrillogenesis by shortening the lag time; an effect that correlated with the extent of destabilization. In contrast, the effect on the kinetics of fibril elongation, as assessed in seeding experiments was of different nature, as it was not directly dependant on the degree of destabilization. This finding suggests different factors drive the nucleation-dependent and elongation phases of light chain fibrillogenesis. Finally, taking advantage of the dependence of the Trp fluorescence upon environment, four single Trp substitutions were made in the N-terminal strand, and changes in solvent exposure during aggregation were evaluated by acrylamide-quenching. The results suggest that the N-terminal strand is buried in the fibrillar state of 6aJL2 protein. This finding suggest a possible explanation for the modulating effect exerted by the mutations in this strand on the aggregation behavior of 6aJL2 protein.« less

  17. The impact of environmental factors on marine turtle stranding rates

    PubMed Central

    Flint, Mark; Limpus, Colin J.; Mills, Paul C.

    2017-01-01

    Globally, tropical and subtropical regions have experienced an increased frequency and intensity in extreme weather events, ranging from severe drought to protracted rain depressions and cyclones, these coincided with an increased number of marine turtles subsequently reported stranded. This study investigated the relationship between environmental variables and marine turtle stranding. The environmental variables examined in this study, in descending order of importance, were freshwater discharge, monthly mean maximum and minimum air temperatures, monthly average daily diurnal air temperature difference and rainfall for the latitudinal hotspots (-27°, -25°, -23°, -19°) along the Queensland coast as well as for major embayments within these blocks. This study found that marine turtle strandings can be linked to these environmental variables at different lag times (3–12 months), and that cumulative (months added together for maximum lag) and non-cumulative (single month only) effects cause different responses. Different latitudes also showed different responses of marine turtle strandings, both in response direction and timing.Cumulative effects of freshwater discharge in all latitudes resulted in increased strandings 10–12 months later. For latitudes -27°, -25° and -23° non-cumulative effects for discharge resulted in increased strandings 7–12 months later. Latitude -19° had different results for the non-cumulative bay with strandings reported earlier (3–6 months). Monthly mean maximum and minimum air temperatures, monthly average daily diurnal air temperature difference and rainfall had varying results for each examined latitude. This study will allow first responders and resource managers to be better equipped to deal with increased marine turtle stranding rates following extreme weather events. PMID:28771635

  18. Single molecular biology: coming of age in DNA replication.

    PubMed

    Liu, Xiao-Jing; Lou, Hui-Qiang

    2017-09-20

    DNA replication is an essential process of the living organisms. To achieve precise and reliable replication, DNA polymerases play a central role in DNA synthesis. Previous investigations have shown that the average rates of DNA synthesis on the leading and lagging strands in a replisome must be similar to avoid the formation of significant gaps in the nascent strands. The underlying mechanism has been assumed to be coordination between leading- and lagging-strand polymerases. However, Kowalczykowski's lab members recently performed single molecule techniques in E. coli and showed the real-time behavior of a replisome. The leading- and lagging-strand polymerases function stochastically and independently. Furthermore, when a DNA polymerase is paused, the helicase slows down in a self-regulating fail-safe mechanism, akin to a ''dead-man's switch''. Based on the real-time single-molecular observation, the authors propose that leading- and lagging-strand polymerases synthesize DNA stochastically within a Gaussian distribution. Along with the development and application of single-molecule techniques, we will witness a new age of DNA replication and other biological researches.

  19. CdS nanowires formed by chemical synthesis using conjugated single-stranded DNA molecules

    NASA Astrophysics Data System (ADS)

    Sarangi, S. N.; Sahu, S. N.; Nozaki, S.

    2018-03-01

    CdS nanowires were successfully grown by chemical synthesis using two conjugated single-stranded (ss) DNA molecules, poly G (30) and poly C (30), as templates. During the early stage of the synthesis with the DNA molecules, the Cd 2+ interacts with Poly G and Poly C and produces the (Cd 2+)-Poly GC complex. As the growth proceeds, it results in nanowires. The structural analysis by grazing angle x-ray diffraction and transmission electron microscopy confirmed the zinc-blende CdS nanowires with the growth direction of <220>. Although the nanowires are well surface-passivated with the DNA molecules, the photoluminescence quenching was caused by the electron transfer from the nanowires to the DNA molecules. The quenching can be used to detect and label the DNAs.

  20. Linear nicking endonuclease-mediated strand-displacement DNA amplification.

    PubMed

    Joneja, Aric; Huang, Xiaohua

    2011-07-01

    We describe a method for linear isothermal DNA amplification using nicking endonuclease-mediated strand displacement by a DNA polymerase. The nicking of one strand of a DNA target by the endonuclease produces a primer for the polymerase to initiate synthesis. As the polymerization proceeds, the downstream strand is displaced into a single-stranded form while the nicking site is also regenerated. The combined continuous repetitive action of nicking by the endonuclease and strand-displacement synthesis by the polymerase results in linear amplification of one strand of the DNA molecule. We demonstrate that DNA templates up to 5000 nucleotides can be linearly amplified using a nicking endonuclease with 7-bp recognition sequence and Sequenase version 2.0 in the presence of single-stranded DNA binding proteins. We also show that a mixture of three templates of 500, 1000, and 5000 nucleotides in length is linearly amplified with the original molar ratios of the templates preserved. Moreover, we demonstrate that a complex library of hydrodynamically sheared genomic DNA from bacteriophage lambda can be amplified linearly. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Linear nicking endonuclease-mediated strand displacement DNA amplification

    PubMed Central

    Joneja, Aric; Huang, Xiaohua

    2011-01-01

    We describe a method for linear isothermal DNA amplification using nicking endonuclease-mediated strand displacement by a DNA polymerase. The nicking of one strand of a DNA target by the endonuclease produces a primer for the polymerase to initiate synthesis. As the polymerization proceeds, the downstream strand is displaced into a single-stranded form while the nicking site is also regenerated. The combined continuous repetitive action of nicking by the endonuclease and strand displacement synthesis by the polymerase results in linear amplification of one strand of the DNA molecule. We demonstrate that DNA templates up to five thousand nucleotides can be linearly amplified using a nicking endonuclease with seven base-pair recognition sequence and Sequenase version 2.0 in the presence of single-stranded DNA binding proteins. We also show that a mixture of three templates of 500, 1000, and 5000 nucleotides in length are linearly amplified with the original molar ratios of the templates preserved. Moreover, we demonstrate that a complex library of hydrodynamically sheared genomic DNA from bacteriophage lambda can be amplified linearly. PMID:21342654

  2. Mitochondrial transcription terminator family members mTTF and mTerf5 have opposing roles in coordination of mtDNA synthesis.

    PubMed

    Jõers, Priit; Lewis, Samantha C; Fukuoh, Atsushi; Parhiala, Mikael; Ellilä, Simo; Holt, Ian J; Jacobs, Howard T

    2013-01-01

    All genomes require a system for avoidance or handling of collisions between the machineries of DNA replication and transcription. We have investigated the roles in this process of the mTERF (mitochondrial transcription termination factor) family members mTTF and mTerf5 in Drosophila melanogaster. The two mTTF binding sites in Drosophila mtDNA, which also bind mTerf5, were found to coincide with major sites of replication pausing. RNAi-mediated knockdown of either factor resulted in mtDNA depletion and developmental arrest. mTTF knockdown decreased site-specific replication pausing, but led to an increase in replication stalling and fork regression in broad zones around each mTTF binding site. Lagging-strand DNA synthesis was impaired, with extended RNA/DNA hybrid segments seen in replication intermediates. This was accompanied by the accumulation of recombination intermediates and nicked/broken mtDNA species. Conversely, mTerf5 knockdown led to enhanced replication pausing at mTTF binding sites, a decrease in fragile replication intermediates containing single-stranded segments, and the disappearance of species containing segments of RNA/DNA hybrid. These findings indicate an essential and previously undescribed role for proteins of the mTERF family in the integration of transcription and DNA replication, preventing unregulated collisions and facilitating productive interactions between the two machineries that are inferred to be essential for completion of lagging-strand DNA synthesis.

  3. Mitochondrial Transcription Terminator Family Members mTTF and mTerf5 Have Opposing Roles in Coordination of mtDNA Synthesis

    PubMed Central

    Jõers, Priit; Lewis, Samantha C.; Fukuoh, Atsushi; Parhiala, Mikael; Ellilä, Simo; Holt, Ian J.; Jacobs, Howard T.

    2013-01-01

    All genomes require a system for avoidance or handling of collisions between the machineries of DNA replication and transcription. We have investigated the roles in this process of the mTERF (mitochondrial transcription termination factor) family members mTTF and mTerf5 in Drosophila melanogaster. The two mTTF binding sites in Drosophila mtDNA, which also bind mTerf5, were found to coincide with major sites of replication pausing. RNAi-mediated knockdown of either factor resulted in mtDNA depletion and developmental arrest. mTTF knockdown decreased site-specific replication pausing, but led to an increase in replication stalling and fork regression in broad zones around each mTTF binding site. Lagging-strand DNA synthesis was impaired, with extended RNA/DNA hybrid segments seen in replication intermediates. This was accompanied by the accumulation of recombination intermediates and nicked/broken mtDNA species. Conversely, mTerf5 knockdown led to enhanced replication pausing at mTTF binding sites, a decrease in fragile replication intermediates containing single-stranded segments, and the disappearance of species containing segments of RNA/DNA hybrid. These findings indicate an essential and previously undescribed role for proteins of the mTERF family in the integration of transcription and DNA replication, preventing unregulated collisions and facilitating productive interactions between the two machineries that are inferred to be essential for completion of lagging-strand DNA synthesis. PMID:24068965

  4. Extended Minus-Strand DNA as Template for R-U5-Mediated Second-Strand Transfer in Recombinational Rescue of Primer Binding Site-Modified Retroviral Vectors

    PubMed Central

    Mikkelsen, Jacob Giehm; Lund, Anders H.; Dybkær, Karen; Duch, Mogens; Pedersen, Finn Skou

    1998-01-01

    We have previously demonstrated recombinational rescue of primer binding site (PBS)-impaired Akv murine leukemia virus-based vectors involving initial priming on endogenous viral sequences and template switching during cDNA synthesis to obtain PBS complementarity in second-strand transfer of reverse transcription (Mikkelsen et al., J. Virol. 70:1439–1447, 1996). By use of the same forced recombination system, we have now found recombinant proviruses of different structures, suggesting that PBS knockout vectors may be rescued through initial priming on endogenous virus RNA, read-through of the mutated PBS during minus-strand synthesis, and subsequent second-strand transfer mediated by the R-U5 complementarity of the plus strand and the extended minus-strand DNA acceptor template. Mechanisms for R-U5-mediated second-strand transfer and its possible role in retrovirus replication and evolution are discussed. PMID:9499117

  5. RNA synthesis is modulated by G-quadruplex formation in Hepatitis C virus negative RNA strand.

    PubMed

    Chloé, Jaubert; Amina, Bedrat; Laura, Bartolucci; Carmelo, Di Primo; Michel, Ventura; Jean-Louis, Mergny; Samir, Amrane; Marie-Line, Andreola

    2018-05-25

    DNA and RNA guanine-rich oligonucleotides can form non-canonical structures called G-quadruplexes or "G4" that are based on the stacking of G-quartets. The role of DNA and RNA G4 is documented in eukaryotic cells and in pathogens such as viruses. Yet, G4 have been identified only in a few RNA viruses, including the Flaviviridae family. In this study, we analysed the last 157 nucleotides at the 3'end of the HCV (-) strand. This sequence is known to be the minimal sequence required for an efficient RNA replication. Using bioinformatics and biophysics, we identified a highly conserved G4-prone sequence located in the stem-loop IIy' of the negative strand. We also showed that the formation of this G-quadruplex inhibits the in vitro RNA synthesis by the RdRp. Furthermore, Phen-DC3, a specific G-quadruplex binder, is able to inhibit HCV viral replication in cells in conditions where no cytotoxicity was measured. Considering that this domain of the negative RNA strand is well conserved among HCV genotypes, G4 ligands could be of interest for new antiviral therapies.

  6. Eukaryotic DNA Replication Fork.

    PubMed

    Burgers, Peter M J; Kunkel, Thomas A

    2017-06-20

    This review focuses on the biogenesis and composition of the eukaryotic DNA replication fork, with an emphasis on the enzymes that synthesize DNA and repair discontinuities on the lagging strand of the replication fork. Physical and genetic methodologies aimed at understanding these processes are discussed. The preponderance of evidence supports a model in which DNA polymerase ε (Pol ε) carries out the bulk of leading strand DNA synthesis at an undisturbed replication fork. DNA polymerases α and δ carry out the initiation of Okazaki fragment synthesis and its elongation and maturation, respectively. This review also discusses alternative proposals, including cellular processes during which alternative forks may be utilized, and new biochemical studies with purified proteins that are aimed at reconstituting leading and lagging strand DNA synthesis separately and as an integrated replication fork.

  7. CTC1-STN1 coordinates G- and C-strand synthesis to regulate telomere length.

    PubMed

    Gu, Peili; Jia, Shuting; Takasugi, Taylor; Smith, Eric; Nandakumar, Jayakrishnan; Hendrickson, Eric; Chang, Sandy

    2018-05-17

    Coats plus (CP) is a rare autosomal recessive disorder caused by mutations in CTC1, a component of the CST (CTC1, STN1, and TEN1) complex important for telomere length maintenance. The molecular basis of how CP mutations impact upon telomere length remains unclear. The CP CTC1 L1142H mutation has been previously shown to disrupt telomere maintenance. In this study, we used CRISPR/Cas9 to engineer this mutation into both alleles of HCT116 and RPE cells to demonstrate that CTC1:STN1 interaction is required to repress telomerase activity. CTC1 L1142H interacts poorly with STN1, leading to telomerase-mediated telomere elongation. Impaired interaction between CTC1 L1142H :STN1 and DNA Pol-α results in increased telomerase recruitment to telomeres and further telomere elongation, revealing that C:S binding to DNA Pol-α is required to fully repress telomerase activity. CP CTC1 mutants that fail to interact with DNA Pol-α resulted in loss of C-strand maintenance and catastrophic telomere shortening. Our findings place the CST complex as an important regulator of both G-strand extensions by telomerase and C-strand synthesis by DNA Pol-α. © 2018 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

  8. Simple and rapid enzymatic method for the synthesis of single-strand oligonucleotides containing trifluorothymidine.

    PubMed

    Suzuki, Norihiko; Fukushima, Masakazu

    2010-11-01

    To investigate the mechanism of trifluorothymidine (TFT)-induced DNA damage, we developed an enzymatic method for the synthesis of single-strand oligonucleotides containing TFT-monophosphate residues. Sixteen-mer oligonucleotides and 14-mer 5'-phosphorylated oligonucleotides were annealed to the template of 25-mer, so as to empty one nucleotide site. TFT-triphosphate was incorporated into the site by DNA polymerase and then ligated to 5'-phosphorylated oligonucleotides by DNA ligase. The synthesized 31-mer oligonucleotides containing TFT residues were isolated from the 25-mer complementary template by denaturing polyacrylamide electrophoresis. Using these single-strand oligonucleotides containing TFT residues, the cleavage of TFT residues from DNA, using mismatch uracil-DNA glycosylase (MUG) of E.coli origin, was compared with that of 5-fluorouracil (5FU) and 5-bromodeoxyuridine (BrdU). The TFT/A pair was not cleaved by MUG, while the other pairs, namely, 5FU/A, 5FU/G, BrdU/A, BrdU/G, and TFT/G, were easily cleaved from each synthesized DNA. Thus, this method is useful for obtaining some site-specifically modified oligonucleotides.

  9. Simultaneous binding to the tracking strand, displaced strand and the duplex of a DNA fork enhances unwinding by Dda helicase

    PubMed Central

    Aarattuthodiyil, Suja; Byrd, Alicia K.; Raney, Kevin D.

    2014-01-01

    Interactions between helicases and the tracking strand of a DNA substrate are well-characterized; however, the role of the displaced strand is a less understood characteristic of DNA unwinding. Dda helicase exhibited greater processivity when unwinding a DNA fork compared to a ss/ds DNA junction substrate. The lag phase in the unwinding progress curve was reduced for the forked DNA compared to the ss/ds junction. Fewer kinetic steps were required to unwind the fork compared to the ss/ds junction, suggesting that binding to the fork leads to disruption of the duplex. DNA footprinting confirmed that interaction of Dda with a fork leads to two base pairs being disrupted whereas no disruption of base pairing was observed with the ss/ds junction. Neutralization of the phosphodiester backbone resulted in a DNA-footprinting pattern similar to that observed with the ss/ds junction, consistent with disruption of the interaction between Dda and the displaced strand. Several basic residues in the 1A domain which were previously proposed to bind to the incoming duplex DNA were replaced with alanines, resulting in apparent loss of interaction with the duplex. Taken together, these results suggest that Dda interaction with the tracking strand, displaced strand and duplex coordinates DNA unwinding. PMID:25249618

  10. Comparison of Polymerase Subunits from Double-Stranded RNA Bacteriophages

    PubMed Central

    Yang, Hongyan; Makeyev, Eugene V.; Bamford, Dennis H.

    2001-01-01

    The family Cystoviridae comprises several bacteriophages with double-stranded RNA (dsRNA) genomes. We have previously purified the catalytic polymerase subunit (Pol) of one of the Cystoviridae members, bacteriophage φ6, and shown that the protein can catalyze RNA synthesis in vitro. In this reaction, both bacteriophage-specific and heterologous RNAs can serve as templates, but those containing 3′ termini from the φ6 minus strands are favored. This provides a molecular basis for the observation that only plus strands, not minus strands, are transcribed from φ6 dsRNA segments in vivo. To test whether such a regulatory mechanism is also found in other dsRNA viruses, we purified recombinant Pol subunits from the φ6-related bacteriophages φ8 and φ13 and assayed their polymerase activities in vitro. The enzymes catalyze template-dependent RNA synthesis using both single-stranded-RNA (ssRNA) and dsRNA templates. However, they differ from each other as well as from φ6 Pol in certain biochemical properties. Notably, each polymerase demonstrates a distinct preference for ssRNAs bearing short 3′-terminal sequences from the virus-specific minus strands. This suggests that, in addition to other factors, RNA transcription in Cystoviridae is controlled by the template specificity of the polymerase subunit. PMID:11602748

  11. Initiation of poliovirus plus-strand RNA synthesis in a membrane complex of infected HeLa cells

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

    Takeda, N.; Kuhn, R.J.; Yang, C.F.

    1986-10-01

    An in vitro poliovirus RNA-synthesizing system derived from a crude membrance fraction of infected HeLa cells was used to analyze the mechanism of initiation of poliovirus plus-strand RNA synthesis. This system contains an activity that synthesizes the nucleotidyl proteins VPg-pU and VPg-pUpU. These molecules represent the 5'-terminal structure of nascent RNA molecules and of virion RNA. The membranous replication complex is also capable of synthesizing mucleotidyl proteins containing nine or more of the poliovirus 5'-proximal nucleotides as assayed by the formation of the RNase T/sub 1/-resistant oligonucleotide VPg-pUUAAAACAGp or by fingerprint analysis of the in vitro-synthesized /sup 32/P-RNA. Incubation ofmore » preformed VPg-pUpU with unlabeled nucleoside triphosphates resulted in the formation of VPg-pUUAAAACAGp. This reaction, which appeared to be an elongation of VPg-pUpU, was stimulated by the addition of a soluble fraction (S-10) obtained from uninfected HeLa cells. Preformed VPg-pU could be chased into VPg-pUpU in the presence of UTP. The data are consistent with a model that VPg-pU can function as a primer for poliovirus plus-strand RNA synthesis in the membranous replication complex and that the elongation reaction may be stimulated by a host cellular factor.« less

  12. CTC1-mediated C-strand fill-in is an essential step in telomere length maintenance

    PubMed Central

    Feng, Xuyang; Hsu, Shih-Jui; Kasbek, Christopher; Chaiken, Mary

    2017-01-01

    Abstract To prevent progressive telomere shortening as a result of conventional DNA replication, new telomeric DNA must be added onto the chromosome end. The de novo DNA synthesis involves elongation of the G-rich strand of the telomere by telomerase. In human cells, the CST complex (CTC1-STN1-TEN1) also functions in telomere replication. CST first aids in duplication of the telomeric dsDNA. Then after telomerase has extended the G-rich strand, CST facilitates fill-in synthesis of the complementary C-strand. Here, we analyze telomere structure after disruption of human CTC1 and demonstrate that functional CST is essential for telomere length maintenance due to its role in mediating C-strand fill-in. Removal of CTC1 results in elongation of the 3΄ overhang on the G-rich strand. This leads to accumulation of RPA and telomeric DNA damage signaling. G-overhang length increases with time after CTC1 disruption and at early times net G-strand growth is apparent, indicating telomerase-mediated G-strand extension. In contrast, C-strand length decreases continuously, indicating a deficiency in C-strand fill-in synthesis. The lack of C-strand maintenance leads to gradual shortening of the telomeric dsDNA, similar to that observed in cells lacking telomerase. Thus, telomerase-mediated G-strand extension and CST-mediated C-strand fill-in are equally important for telomere length maintenance. PMID:28334750

  13. Mechanism for priming DNA synthesis by yeast DNA Polymerase α

    PubMed Central

    Perera, Rajika L; Torella, Rubben; Klinge, Sebastian; Kilkenny, Mairi L; Maman, Joseph D; Pellegrini, Luca

    2013-01-01

    The DNA Polymerase α (Pol α)/primase complex initiates DNA synthesis in eukaryotic replication. In the complex, Pol α and primase cooperate in the production of RNA-DNA oligonucleotides that prime synthesis of new DNA. Here we report crystal structures of the catalytic core of yeast Pol α in unliganded form, bound to an RNA primer/DNA template and extending an RNA primer with deoxynucleotides. We combine the structural analysis with biochemical and computational data to demonstrate that Pol α specifically recognizes the A-form RNA/DNA helix and that the ensuing synthesis of B-form DNA terminates primer synthesis. The spontaneous release of the completed RNA-DNA primer by the Pol α/primase complex simplifies current models of primer transfer to leading- and lagging strand polymerases. The proposed mechanism of nucleotide polymerization by Pol α might contribute to genomic stability by limiting the amount of inaccurate DNA to be corrected at the start of each Okazaki fragment. DOI: http://dx.doi.org/10.7554/eLife.00482.001 PMID:23599895

  14. Single-molecule visualization of Saccharomyces cerevisiae leading-strand synthesis reveals dynamic interaction between MTC and the replisome

    PubMed Central

    Lewis, Jacob S.; Spenkelink, Lisanne M.; Schauer, Grant D.; Hill, Flynn R.; Georgescu, Roxanna E.; O’Donnell, Michael E.; van Oijen, Antoine M.

    2017-01-01

    The replisome, the multiprotein system responsible for genome duplication, is a highly dynamic complex displaying a large number of different enzyme activities. Recently, the Saccharomyces cerevisiae minimal replication reaction has been successfully reconstituted in vitro. This provided an opportunity to uncover the enzymatic activities of many of the components in a eukaryotic system. Their dynamic behavior and interactions in the context of the replisome, however, remain unclear. We use a tethered-bead assay to provide real-time visualization of leading-strand synthesis by the S. cerevisiae replisome at the single-molecule level. The minimal reconstituted leading-strand replisome requires 24 proteins, forming the CMG helicase, the Pol ε DNA polymerase, the RFC clamp loader, the PCNA sliding clamp, and the RPA single-stranded DNA binding protein. We observe rates and product lengths similar to those obtained from ensemble biochemical experiments. At the single-molecule level, we probe the behavior of two components of the replication progression complex and characterize their interaction with active leading-strand replisomes. The Minichromosome maintenance protein 10 (Mcm10), an important player in CMG activation, increases the number of productive replication events in our assay. Furthermore, we show that the fork protection complex Mrc1–Tof1–Csm3 (MTC) enhances the rate of the leading-strand replisome threefold. The introduction of periods of fast replication by MTC leads to an average rate enhancement of a factor of 2, similar to observations in cellular studies. We observe that the MTC complex acts in a dynamic fashion with the moving replisome, leading to alternating phases of slow and fast replication. PMID:28923950

  15. The N-terminal strand modulates immunoglobulin light chain fibrillogenesis.

    PubMed

    del Pozo-Yauner, Luis; Wall, Jonathan S; González Andrade, Martín; Sánchez-López, Rosana; Rodríguez-Ambriz, Sandra L; Pérez Carreón, Julio I; Ochoa-Leyva, Adrián; Fernández-Velasco, D Alejandro

    2014-01-10

    It has been suggested that the N-terminal strand of the light chain variable domain (V(L)) protects the molecule from aggregation by hindering spurious intermolecular contacts. We evaluated the impact of mutations in the N-terminal strand on the thermodynamic stability and kinetic of fibrillogenesis of the V(L) protein 6aJL2. Mutations in this strand destabilized the protein in a position-dependent manner, accelerating the fibrillogenesis by shortening the lag time; an effect that correlated with the extent of destabilization. In contrast, the effect on the kinetics of fibril elongation, as assessed in seeding experiments was of different nature, as it was not directly dependant on the degree of destabilization. This finding suggests different factors drive the nucleation-dependent and elongation phases of light chain fibrillogenesis. Finally, taking advantage of the dependence of the Trp fluorescence upon environment, four single Trp substitutions were made in the N-terminal strand, and changes in solvent exposure during aggregation were evaluated by acrylamide-quenching. The results suggest that the N-terminal strand is buried in the fibrillar state of 6aJL2 protein. This finding suggest a possible explanation for the modulating effect exerted by the mutations in this strand on the aggregation behavior of 6aJL2 protein. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. 5′CAG and 5′CTG Repeats Create Differential Impediment to the Progression of a Minimal Reconstituted T4 Replisome Depending on the Concentration of dNTPs

    PubMed Central

    Delagoutte, Emmanuelle; Baldacci, Giuseppe

    2011-01-01

    Instability of repetitive sequences originates from strand misalignment during repair or replicative DNA synthesis. To investigate the activity of reconstituted T4 replisomes across trinucleotide repeats (TNRs) during leading strand DNA synthesis, we developed a method to build replication miniforks containing a TNR unit of defined sequence and length. Each minifork consists of three strands, primer, leading strand template, and lagging strand template with a 5′ single-stranded (ss) tail. Each strand is prepared independently, and the minifork is assembled by hybridization of the three strands. Using these miniforks and a minimal reconstituted T4 replisome, we show that during leading strand DNA synthesis, the dNTP concentration dictates which strand of the structure-forming 5′CAG/5′CTG repeat creates the strongest impediment to the minimal replication complex. We discuss this result in the light of the known fluctuation of dNTP concentration during the cell cycle and cell growth and the known concentration balance among individual dNTPs. PMID:22096622

  17. Importance of the efficiency of double-stranded DNA formation in cDNA synthesis for the imprecision of microarray expression analysis.

    PubMed

    Thormar, Hans G; Gudmundsson, Bjarki; Eiriksdottir, Freyja; Kil, Siyoen; Gunnarsson, Gudmundur H; Magnusson, Magnus Karl; Hsu, Jason C; Jonsson, Jon J

    2013-04-01

    The causes of imprecision in microarray expression analysis are poorly understood, limiting the use of this technology in molecular diagnostics. Two-dimensional strandness-dependent electrophoresis (2D-SDE) separates nucleic acid molecules on the basis of length and strandness, i.e., double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), and RNA·DNA hybrids. We used 2D-SDE to measure the efficiency of cDNA synthesis and its importance for the imprecision of an in vitro transcription-based microarray expression analysis. The relative amount of double-stranded cDNA formed in replicate experiments that used the same RNA sample template was highly variable, ranging between 0% and 72% of the total DNA. Microarray experiments showed an inverse relationship between the difference between sample pairs in probe variance and the relative amount of dsDNA. Approximately 15% of probes showed between-sample variation (P < 0.05) when the dsDNA percentage was between 12% and 35%. In contrast, only 3% of probes showed between-sample variation when the dsDNA percentage was 69% and 72%. Replication experiments of the 35% dsDNA and 72% dsDNA samples were used to separate sample variation from probe replication variation. The estimated SD of the sample-to-sample variation and of the probe replicates was lower in 72% dsDNA samples than in 35% dsDNA samples. Variation in the relative amount of double-stranded cDNA synthesized can be an important component of the imprecision in T7 RNA polymerase-based microarray expression analysis. © 2013 American Association for Clinical Chemistry

  18. Replicase activity of purified recombinant protein P2 of double-stranded RNA bacteriophage phi6.

    PubMed

    Makeyev, E V; Bamford, D H

    2000-01-04

    In nature, synthesis of both minus- and plus-sense RNA strands of all the known double-stranded RNA viruses occurs in the interior of a large protein assembly referred to as the polymerase complex. In addition to other proteins, the complex contains a putative polymerase possessing characteristic sequence motifs. However, none of the previous studies has shown template-dependent RNA synthesis directly with an isolated putative polymerase protein. In this report, recombinant protein P2 of double-stranded RNA bacteriophage phi6 was purified and demonstrated in an in vitro enzymatic assay to act as the replicase. The enzyme efficiently utilizes phage-specific, positive-sense RNA substrates to produce double-stranded RNA molecules, which are formed by newly synthesized, full-length minus-strands base paired with the plus-strand templates. P2-catalyzed replication is also shown to be very effective with a broad range of heterologous single-stranded RNA templates. The importance and implications of these results are discussed.

  19. Characterization of human translesion DNA synthesis across a UV-induced DNA lesion

    PubMed Central

    Hedglin, Mark; Pandey, Binod; Benkovic, Stephen J

    2016-01-01

    Translesion DNA synthesis (TLS) during S-phase uses specialized TLS DNA polymerases to replicate a DNA lesion, allowing stringent DNA synthesis to resume beyond the offending damage. Human TLS involves the conjugation of ubiquitin to PCNA clamps encircling damaged DNA and the role of this post-translational modification is under scrutiny. A widely-accepted model purports that ubiquitinated PCNA recruits TLS polymerases such as pol η to sites of DNA damage where they may also displace a blocked replicative polymerase. We provide extensive quantitative evidence that the binding of pol η to PCNA and the ensuing TLS are both independent of PCNA ubiquitination. Rather, the unique properties of pols η and δ are attuned to promote an efficient and passive exchange of polymerases during TLS on the lagging strand. DOI: http://dx.doi.org/10.7554/eLife.19788.001 PMID:27770570

  20. Synthesis of triple-stranded complexes using bis(dipyrromethene) ligands.

    PubMed

    Zhang, Zhan; Dolphin, David

    2010-12-20

    The reaction of an α-free, β,β'-linked bis(dipyrromethene) ligand with Fe(3+) or Co(3+) led to noninterconvertible triple-stranded helicates and mesocates. In the present context, a stable α-free ligand 2 has been developed and complexation of ligands 1 and 2 with diamagnetic Co(3+), Ga(3+), and In(3+) has been studied. The triple-stranded M(2)1(3) (M = Ga, In) and M(2)2(3) (M = Co, Ga, In) complexes were characterized using matrix-assisted laser desorption ionization time-of-flight spectrometry, (1)H NMR and UV-vis spectroscopy, and X-ray crystallography. Again, the (1)H NMR analysis showed that both the triple-stranded helicates and mesocates were generated in this metal-directed assembly. Consistent with our previous finding on coordinatively inert Co(3+) complexes, variable-temperature NMR spectroscopy indicated that the triple-stranded helicate and mesocate of labile In(3+) did not interconvert in solution, either. However, the diastereoselectivity of the M(2)2(3) complexes was found to improve with an increase in the reaction temperature. Taken together, this study complements the coordination chemistry of poly(dipyrromethene) ligands and provides further insight into the formation of helicates versus mesocates.

  1. Terminations of DNA synthesis on 'proflavine and light'-treated phi X174 single-stranded DNA.

    PubMed

    Piette, J; Calberg-Bacq, C M; Lopez, M; van de Vorst, A

    1984-04-05

    Bacteriophage phi X174 single-stranded DNA molecules were primed with five different restriction fragments and irradiated with visible light in the presence of proflavine. This photodamaged DNA was used as template for the in vitro complementary chain synthesis by E. coli DNA polymerase I (Klenow fragment). Chain terminations were observed by polyacrylamide gel electrophoresis of the synthesized products and localized by comparison with standard sequencing performed simultaneously on the untreated template. 90% of the chain terminations occurred one nucleotide before a guanine residue in the template strand. More than 80% of the sequenced guanine residues were blocking lesions demonstrating the absence of 'hot-spots' for the photodamaging effect of proflavine. At a defined position, the chain termination frequency increased linearly with the irradiation time and was directly influenced by the proflavine concentration present. An important part of lesions resulted from the action of singlet oxygen produced by excited proflavine as shown by the effect that both NaN3 and 2H2O exerted on the reaction. The induced blocking lesions must be important in vivo since no complete replicative forms could be extracted from cell infected with bacteriophages inactivated by 'proflavine and light' treatment.

  2. In vitro synthesis of minus-strand RNA by an isolated cereal yellow dwarf virus RNA-dependent RNA polymerase requires VPg and a stem-loop structure at the 3' end of the virus RNA.

    PubMed

    Osman, Toba A M; Coutts, Robert H A; Buck, Kenneth W

    2006-11-01

    Cereal yellow dwarf virus (CYDV) RNA has a 5'-terminal genome-linked protein (VPg). We have expressed the VPg region of the CYDV genome in bacteria and used the purified protein (bVPg) to raise an antiserum which was able to detect free VPg in extracts of CYDV-infected oat plants. A template-dependent RNA-dependent RNA polymerase (RdRp) has been produced from a CYDV membrane-bound RNA polymerase by treatment with BAL 31 nuclease. The RdRp was template specific, being able to utilize templates from CYDV plus- and minus-strand RNAs but not those of three unrelated viruses, Red clover necrotic mosaic virus, Cucumber mosaic virus, and Tobacco mosaic virus. RNA synthesis catalyzed by the RdRp required a 3'-terminal GU sequence and the presence of bVPg. Additionally, synthesis of minus-strand RNA on a plus-strand RNA template required the presence of a putative stem-loop structure near the 3' terminus of CYDV RNA. The base-paired stem, a single-nucleotide (A) bulge in the stem, and the sequence of a tetraloop were all required for the template activity. Evidence was produced showing that minus-strand synthesis in vitro was initiated by priming by bVPg at the 3' end of the template. The data are consistent with a model in which the RdRp binds to the stem-loop structure which positions the active site to recognize the 3'-terminal GU sequence for initiation of RNA synthesis by the addition of an A residue to VPg.

  3. In Vitro Synthesis of Minus-Strand RNA by an Isolated Cereal Yellow Dwarf Virus RNA-Dependent RNA Polymerase Requires VPg and a Stem-Loop Structure at the 3′ End of the Virus RNA▿

    PubMed Central

    Osman, Toba A. M.; Coutts, Robert H. A.; Buck, Kenneth W.

    2006-01-01

    Cereal yellow dwarf virus (CYDV) RNA has a 5′-terminal genome-linked protein (VPg). We have expressed the VPg region of the CYDV genome in bacteria and used the purified protein (bVPg) to raise an antiserum which was able to detect free VPg in extracts of CYDV-infected oat plants. A template-dependent RNA-dependent RNA polymerase (RdRp) has been produced from a CYDV membrane-bound RNA polymerase by treatment with BAL 31 nuclease. The RdRp was template specific, being able to utilize templates from CYDV plus- and minus-strand RNAs but not those of three unrelated viruses, Red clover necrotic mosaic virus, Cucumber mosaic virus, and Tobacco mosaic virus. RNA synthesis catalyzed by the RdRp required a 3′-terminal GU sequence and the presence of bVPg. Additionally, synthesis of minus-strand RNA on a plus-strand RNA template required the presence of a putative stem-loop structure near the 3′ terminus of CYDV RNA. The base-paired stem, a single-nucleotide (A) bulge in the stem, and the sequence of a tetraloop were all required for the template activity. Evidence was produced showing that minus-strand synthesis in vitro was initiated by priming by bVPg at the 3′ end of the template. The data are consistent with a model in which the RdRp binds to the stem-loop structure which positions the active site to recognize the 3′-terminal GU sequence for initiation of RNA synthesis by the addition of an A residue to VPg. PMID:16928757

  4. C-terminal Phenylalanine of Bacteriophage T7 Single-stranded DNA-binding Protein Is Essential for Strand Displacement Synthesis by T7 DNA Polymerase at a Nick in DNA*

    PubMed Central

    Ghosh, Sharmistha; Marintcheva, Boriana; Takahashi, Masateru; Richardson, Charles C.

    2009-01-01

    Single-stranded DNA-binding protein (gp2.5), encoded by gene 2.5 of bacteriophage T7, plays an essential role in DNA replication. Not only does it remove impediments of secondary structure in the DNA, it also modulates the activities of the other replication proteins. The acidic C-terminal tail of gp2.5, bearing a C-terminal phenylalanine, physically and functionally interacts with the helicase and DNA polymerase. Deletion of the phenylalanine or substitution with a nonaromatic amino acid gives rise to a dominant lethal phenotype, and the altered gp2.5 has reduced affinity for T7 DNA polymerase. Suppressors of the dominant lethal phenotype have led to the identification of mutations in gene 5 that encodes the T7 DNA polymerase. The altered residues in the polymerase are solvent-exposed and lie in regions that are adjacent to the bound DNA. gp2.5 lacking the C-terminal phenylalanine has a lower affinity for gp5-thioredoxin relative to the wild-type gp2.5, and this affinity is partially restored by the suppressor mutations in DNA polymerase. gp2.5 enables T7 DNA polymerase to catalyze strand displacement DNA synthesis at a nick in DNA. The resulting 5′-single-stranded DNA tail provides a loading site for T7 DNA helicase. gp2.5 lacking the C-terminal phenylalanine does not support this event with wild-type DNA polymerase but does to a limited extent with T7 DNA polymerase harboring the suppressor mutations. PMID:19726688

  5. C-terminal phenylalanine of bacteriophage T7 single-stranded DNA-binding protein is essential for strand displacement synthesis by T7 DNA polymerase at a nick in DNA.

    PubMed

    Ghosh, Sharmistha; Marintcheva, Boriana; Takahashi, Masateru; Richardson, Charles C

    2009-10-30

    Single-stranded DNA-binding protein (gp2.5), encoded by gene 2.5 of bacteriophage T7, plays an essential role in DNA replication. Not only does it remove impediments of secondary structure in the DNA, it also modulates the activities of the other replication proteins. The acidic C-terminal tail of gp2.5, bearing a C-terminal phenylalanine, physically and functionally interacts with the helicase and DNA polymerase. Deletion of the phenylalanine or substitution with a nonaromatic amino acid gives rise to a dominant lethal phenotype, and the altered gp2.5 has reduced affinity for T7 DNA polymerase. Suppressors of the dominant lethal phenotype have led to the identification of mutations in gene 5 that encodes the T7 DNA polymerase. The altered residues in the polymerase are solvent-exposed and lie in regions that are adjacent to the bound DNA. gp2.5 lacking the C-terminal phenylalanine has a lower affinity for gp5-thioredoxin relative to the wild-type gp2.5, and this affinity is partially restored by the suppressor mutations in DNA polymerase. gp2.5 enables T7 DNA polymerase to catalyze strand displacement DNA synthesis at a nick in DNA. The resulting 5'-single-stranded DNA tail provides a loading site for T7 DNA helicase. gp2.5 lacking the C-terminal phenylalanine does not support this event with wild-type DNA polymerase but does to a limited extent with T7 DNA polymerase harboring the suppressor mutations.

  6. Interaction between the cellular protein eEF1A and the 3'-terminal stem-loop of West Nile virus genomic RNA facilitates viral minus-strand RNA synthesis.

    PubMed

    Davis, William G; Blackwell, Jerry L; Shi, Pei-Yong; Brinton, Margo A

    2007-09-01

    RNase footprinting and nitrocellulose filter binding assays were previously used to map one major and two minor binding sites for the cell protein eEF1A on the 3'(+) stem-loop (SL) RNA of West Nile virus (WNV) (3). Base substitutions in the major eEF1A binding site or adjacent areas of the 3'(+) SL were engineered into a WNV infectious clone. Mutations that decreased, as well as ones that increased, eEF1A binding in in vitro assays had a negative effect on viral growth. None of these mutations affected the efficiency of translation of the viral polyprotein from the genomic RNA, but all of the mutations that decreased in vitro eEF1A binding to the 3' SL RNA also decreased viral minus-strand RNA synthesis in transfected cells. Also, a mutation that increased the efficiency of eEF1A binding to the 3' SL RNA increased minus-strand RNA synthesis in transfected cells, which resulted in decreased synthesis of genomic RNA. These results strongly suggest that the interaction between eEF1A and the WNV 3' SL facilitates viral minus-strand synthesis. eEF1A colocalized with viral replication complexes (RC) in infected cells and antibody to eEF1A coimmunoprecipitated viral RC proteins, suggesting that eEF1A facilitates an interaction between the 3' end of the genome and the RC. eEF1A bound with similar efficiencies to the 3'-terminal SL RNAs of four divergent flaviviruses, including a tick-borne flavivirus, and colocalized with dengue virus RC in infected cells. These results suggest that eEF1A plays a similar role in RNA replication for all flaviviruses.

  7. CMG–Pol epsilon dynamics suggests a mechanism for the establishment of leading-strand synthesis in the eukaryotic replisome

    PubMed Central

    Janska, Agnieszka; Goswami, Panchali; Renault, Ludovic; Abid Ali, Ferdos; Kotecha, Abhay; Costa, Alessandro

    2017-01-01

    The replisome unwinds and synthesizes DNA for genome duplication. In eukaryotes, the Cdc45–MCM–GINS (CMG) helicase and the leading-strand polymerase, Pol epsilon, form a stable assembly. The mechanism for coupling DNA unwinding with synthesis is starting to be elucidated, however the architecture and dynamics of the replication fork remain only partially understood, preventing a molecular understanding of chromosome replication. To address this issue, we conducted a systematic single-particle EM study on multiple permutations of the reconstituted CMG–Pol epsilon assembly. Pol epsilon contains two flexibly tethered lobes. The noncatalytic lobe is anchored to the motor of the helicase, whereas the polymerization domain extends toward the side of the helicase. We observe two alternate configurations of the DNA synthesis domain in the CMG-bound Pol epsilon. We propose that this conformational switch might control DNA template engagement and release, modulating replisome progression. PMID:28373564

  8. Polypyrimidine tract-binding protein influences negative strand RNA synthesis of dengue virus.

    PubMed

    Jiang, Linbin; Yao, Huiling; Duan, Xiaoqun; Lu, Xi; Liu, Yongming

    2009-07-24

    Flavivirus non-structural protein 4A (NS4A) induces membrane rearrangements to form viral replication complex and functions as interferon antagonist. However, other non-structural roles of NS4A protein in relation to virus life-cycle are poorly defined. This study elucidated if dengue virus (DENV) NS4A protein interacts with host proteins and contributes to viral pathogenesis by screening human liver cDNA yeast-two-hybrid library. Our study identified polypyrimidine tract-binding protein (PTB) as a novel interacting partner of DENV NS4A protein. We reported for the first time that PTB influenced DENV production. Gene-silencing studies showed that PTB did not have an effect on DENV entry and DENV RNA translation. Further functional studies revealed that PTB influenced DENV production by modulating negative strand RNA synthesis. This is the first study that enlightens the interaction of DENV NS4A protein with PTB, in addition to demonstrating the novel role of PTB in relation to mosquito-borne flavivirus life-cycle.

  9. Analysis of strand transfer and template switching mechanisms of DNA gap repair by homologous recombination in Escherichia coli: predominance of strand transfer.

    PubMed

    Izhar, Lior; Goldsmith, Moshe; Dahan, Ronny; Geacintov, Nicholas; Lloyd, Robert G; Livneh, Zvi

    2008-09-12

    Daughter strand gaps formed upon interruption of replication at DNA lesions in Escherichia coli can be repaired by either translesion DNA synthesis or homologous recombination (HR) repair. Using a plasmid-based assay system that enables discrimination between strand transfer and template switching (information copying) modes of HR gap repair, we found that approximately 80% of strand gaps were repaired by physical strand transfer from the donor, whereas approximately 20% appear to be repaired by template switching. HR gap repair operated on both small and bulky lesions and largely depended on RecA and RecF but not on the RecBCD nuclease. In addition, we found that HR was mildly reduced in cells lacking the RuvABC and RecG proteins involved in resolution of Holliday junctions. These results, obtained for the first time under conditions that detect the two HR gap repair mechanisms, provide in vivo high-resolution molecular evidence for the predominance of the strand transfer mechanism in HR gap repair. A small but significant portion of HR gap repair appears to occur via a template switching mechanism.

  10. Stabilized NADH as a Countermeasure for Jet Lag

    NASA Technical Reports Server (NTRS)

    Kay, Gary G.; Viirre, Erik; Clark, Jonathan

    2001-01-01

    Current remedies for jet lag (phototherapy, melatonin, stimulant, and sedative medications) are limited in efficacy and practicality. The efficacy of a stabilized, sublingual form of reduced nicotin amide adenine dinucleotide (NADH, ENADAlert, Menuco Corp.) as a countermeasure for jet lag was examined. Because NADH increases cellular production of ATP and facilitates dopamine synthesis, it may counteract the effects of jet lag on cognitive functioning and sleepiness. Thirty-five healthy, employed subjects participated in this double-blind, placebo-controlled study. Training and baseline testing were conducted on the West Coast before subjects flew overnight to the East Coast, where they would experience a 3-hour time difference. Upon arrival, individuals were randomly assigned to receive either 20 mg of sublingual stabilized ADH (n=18) or identical placebo tablets (n=17). All participants completed computer-administered tests (including CogScreen7) to assess changes in cognitive functioning, mood, and sleepiness in the morning and afternoon. Jet lag resulted in increased sleepiness for over half the participants and deterioration of cognitive functioning for approximately one third. The morning following the flight, subjects experienced lapses of attention in addition to disruptions in working memory, divided attention, and visual perceptual speed. Individuals who received NADH performed significantly better on 5 of 8 cognitive and psychomotor test measures (P less than or equal to 0.5) and showed a trend for better performance on the other three measures (P less than or equal to .l0). Subjects also reported less sleepiness compared with those who received placebo. No adverse effects were observed with NADH treatment. Stabilized NADH significantly reduced jet lag-induced disruptions of cognitive functioning, was easily administered, and was found to have no adverse side effects.

  11. Rolling Circle Amplification For Spatially Directed Synthesis Of A Solid Phase Anchored Single-Stranded DNA Molecule

    NASA Astrophysics Data System (ADS)

    Reiß, Edda; Hölzel, Ralph; von Nickisch-Rosenegk, Markus; Bier, Frank F.

    2006-09-01

    In this article the usefulness of the enzyme phi29 DNA polymerase and the principle of rolling circle amplification (RCA) for creating single-stranded DNA (ssDNA) nanostructures is described. Currently we are working on the spatial orientation of a growing ssDNA molecule during its RCA-based synthesis by the application of a hydrodynamic force. Starting at an immobilized primer at single molecule level, the aim is to construct a nanostructure of known location and orientation, providing multiple repeating binding sites that can be addressed via complementary base-pairing. Proof-of-principle experiments demonstrate the potential of the enzymatic reaction. ssDNA molecules of more than 20 μm length were created at an immobilized primer and detected by means of fluorescence microscopy.

  12. Potent Inhibitors of the Hepatitis C Virus NS3 Protease: Design and Synthesis of Macrocyclic Substrate-Based [beta]-Strand Mimics

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

    Goudreau, Nathalie; Brochu, Christian; Cameron, Dale R.

    2008-06-30

    The virally encoded NS3 protease is essential to the life cycle of the hepatitis C virus (HCV), an important human pathogen causing chronic hepatitis, cirrhosis of the liver, and hepatocellular carcinoma. The design and synthesis of 15-membered ring {beta}-strand mimics which are capable of inhibiting the interactions between the HCV NS3 protease enzyme and its polyprotein substrate will be described. The binding interactions between a macrocyclic ligand and the enzyme were explored by NMR and molecular dynamics, and a model of the ligand/enzyme complex was developed.

  13. Abnormal, Error-Prone Bypass of Photoproducts by Xeroderma Pigmentosum Variant Cell Extracts Results in Extreme Strand Bias for the Kinds of Mutations Induced by UV Light

    PubMed Central

    McGregor, W. Glenn; Wei, Dong; Maher, Veronica M.; McCormick, J. Justin

    1999-01-01

    Xeroderma pigmentosum (XP) is a rare genetic disease characterized by a greatly increased susceptibility to sunlight-induced skin cancer. Cells from the majority of patients are defective in nucleotide excision repair. However, cells from one set of patients, XP variants, exhibit normal repair but are abnormally slow in replicating DNA containing UV photoproducts. The frequency of UV radiation-induced mutations in the XP variant cells is significantly higher than that in normal human cells. Furthermore, the kinds of UV-induced mutations differ very significantly from normal. Instead of transitions, mainly C→T, 30% of the base substitutions consist of C→A transversions, all arising from photoproducts located in one strand. Mutations involving cytosine in the other strand are almost all C→T transitions. Forty-five percent of the substitutions involve thymine, and the majority are transversions. To test the hypothesis that the UV hypermutability and the abnormal spectrum of mutations result from abnormal bypass of photoproducts in DNA, we compared extracts from XP variant cells with those from HeLa cells and a fibroblast cell strain, MSU-1.2, for the ability to replicate a UV-irradiated form I M13 phage. The M13 template contains a simian virus 40 origin of replication located directly to the left or to the right of the target gene, lacZα, so that the template for the leading and lagging strands of DNA replication is defined. Reduction of replication to ∼37% of the control value required only 1 photoproduct per template for XP variant cell extracts, but ∼2.2 photoproducts for HeLa or MSU-1.2 cell extracts. The frequency of mutants induced was four times higher with XP variant cell extracts than with HeLa or MSU-1.2 cell extracts. With XP variant cell extracts, the proportion of C→A transversions reached as high as 43% with either M13 template and arose from photoproducts located in the template for leading-strand synthesis; with HeLa or MSU-1.2 cell

  14. Nucleocapsid Protein Annealing of a Primer-Template Enhances (+)-Strand DNA Synthesis and Fidelity by HIV-1 Reverse Transcriptase†

    PubMed Central

    Kim, Jiae; Roberts, Anne; Yuan, Hua; Xiong, Yong; Anderson, Karen S.

    2012-01-01

    Human immunodeficiency virus type-1 (HIV-1) requires reverse transcriptase (RT) and HIV-1 nucleocapsid protein (NCp7) for proper viral replication. HIV-1 NCp7 has been shown to enhance various steps in reverse transcription including tRNA initiation and strand transfer which may be mediated through interactions with RT as well as RNA and DNA oligonucleotides. With the use of DNA oligonucleotides, we have examined the interaction of NCp7 with RT and the kinetics of reverse transcription during (+)-strand synthesis with an NCp7-facilitated annealed primer-template. Using a pre-steady state kinetics approach, the NCp7-annealed primer-template has a substantial increase (3-7 fold) in the rate of incorporation (kpol) by RT as compared to heat annealed primer-template with single nucleotide incorporation. There was also a 2-fold increase in the binding affinity constant (Kd) of the nucleotide. These differences in kpol and Kd were not through direct interactions between HIV-1 RT and NCp7. When examining extension by RT, the data suggests that the NCp7-annealed primer-template facilitates the formation of a longer product more quickly compared to the heat annealed primer-template. This enhancement in rate is mediated through interactions with NCp7’s zinc fingers and N-terminal domain and nucleic acids. The NCp7-annealed primer-template also enhances the fidelity of RT (3-fold) by slowing the rate of incorporation of an incorrect nucleotide. Taken together, this study elucidates a new role of NCp7 by facilitating DNA-directed DNA synthesis during reverse transcription by HIV-1 RT that may translate into enhanced viral fitness and offers an avenue to exploit for targeted therapeutic intervention against HIV. PMID:22210155

  15. Time-lapse crystallography snapshots of a double-strand break repair polymerase in action.

    PubMed

    Jamsen, Joonas A; Beard, William A; Pedersen, Lars C; Shock, David D; Moon, Andrea F; Krahn, Juno M; Bebenek, Katarzyna; Kunkel, Thomas A; Wilson, Samuel H

    2017-08-15

    DNA polymerase (pol) μ is a DNA-dependent polymerase that incorporates nucleotides during gap-filling synthesis in the non-homologous end-joining pathway of double-strand break repair. Here we report time-lapse X-ray crystallography snapshots of catalytic events during gap-filling DNA synthesis by pol μ. Unique catalytic intermediates and active site conformational changes that underlie catalysis are uncovered, and a transient third (product) metal ion is observed in the product state. The product manganese coordinates phosphate oxygens of the inserted nucleotide and PP i . The product metal is not observed during DNA synthesis in the presence of magnesium. Kinetic analyses indicate that manganese increases the rate constant for deoxynucleoside 5'-triphosphate insertion compared to magnesium. The likely product stabilization role of the manganese product metal in pol μ is discussed. These observations provide insight on structural attributes of this X-family double-strand break repair polymerase that impact its biological function in genome maintenance.DNA polymerase (pol) μ functions in DNA double-strand break repair. Here the authors use time-lapse X-ray crystallography to capture the states of pol µ during the conversion from pre-catalytic to product complex and observe a third transiently bound metal ion in the product state.

  16. The buzz-lag effect.

    PubMed

    Cellini, Cristiano; Scocchia, Lisa; Drewing, Knut

    2016-10-01

    In the flash-lag illusion, a brief visual flash and a moving object presented at the same location appear to be offset with the flash trailing the moving object. A considerable amount of studies investigated the visual flash-lag effect, and flash-lag-like effects have also been observed in audition, and cross-modally between vision and audition. In the present study, we investigate whether a similar effect can also be observed when using only haptic stimuli. A fast vibration (or buzz, lasting less than 20 ms) was applied to the moving finger of the observers and employed as a "haptic flash." Participants performed a two-alternative forced-choice (2AFC) task where they had to judge whether the moving finger was located to the right or to the left of the stationary finger at the time of the buzz. We used two different movement velocities (Slow and Fast conditions). We found that the moving finger was systematically misperceived to be ahead of the stationary finger when the two were physically aligned. This result can be interpreted as a purely haptic analogue of the flash-lag effect, which we refer to as "buzz-lag effect." The buzz-lag effect can be well accounted for by the temporal-sampling explanation of flash-lag-like effects.

  17. Intramedullary nails with two lag screws.

    PubMed

    Brown, C J; Wang, C J; Yettram, A L; Procter, P

    2004-06-01

    To investigate the structural integrity of intramedullary nails with two lag screws, and to give guidance to orthopaedic surgeons in the choice of appropriate devices. Alternative designs of the construct are considered, and the use of a slotted upper lag screw insertion hole is analysed. Intramedullary fixation devices with a single lag screw have been known to fail at the lag screw insertion hole. Using two lag screws is considered. It has also been proposed to use a slot in the nail for the upper lag screw to prevent the upper lag screw from sticking. Bending and torsion load cases are analysed using finite element method. Consideration of both load conditions is essential. The results present the overall stiffness of the assembly, the load sharing between lag screws, and the possibility for cut-out to occur. While the slot for the upper lag screw might be advantageous with regard to the stresses in the lag screws, it could be detrimental for cut-out occurring adjacent to the lag screws. Comparative analyses demonstrate that two lag screws may be advantageous in patients whose cancellous bone quality is good and who impose large loads on the lag screw/nail interface. However, the use of two screws might pre-dispose to failure by cut-out of the lag screws. The addition of a slotted hole for the upper lag screw appears to do nothing significant to reduce the risk of such a failure. Copyright 2004 Elsevier Ltd.

  18. Switch from translation to RNA replication in a positive-stranded RNA virus

    PubMed Central

    Gamarnik, Andrea V.; Andino, Raul

    1998-01-01

    In positive-stranded viruses, the genomic RNA serves as a template for both translation and RNA replication. Using poliovirus as a model, we examined the interaction between these two processes. We show that the RNA polymerase is unable to replicate RNA templates undergoing translation. We discovered that an RNA structure at the 5′ end of the viral genome, next to the internal ribosomal entry site, carries signals that control both viral translation and RNA synthesis. The interaction of this RNA structure with the cellular factor PCBP up-regulates viral translation, while the binding of the viral protein 3CD represses translation and promotes negative-strand RNA synthesis. We propose that the interaction of 3CD with this RNA structure controls whether the genomic RNA is used for translation or RNA replication. PMID:9694795

  19. The effect of bifocal add on accommodative lag in myopic children with high accommodative lag.

    PubMed

    Berntsen, David A; Mutti, Donald O; Zadnik, Karla

    2010-12-01

    To determine the effect of a bifocal add and manifest correction on accommodative lag in myopic children with high accommodative lag, who have been reported to have the greatest reduction in myopia progression with progressive addition lenses (PALs). Monocular accommodative lag to a 4-D Badal stimulus was measured on two occasions 6 months apart in 83 children (mean ± SD age, 9.9 ± 1.3 years) with high lag randomized to wearing single-vision lenses (SVLs) or PALs. Accommodative lag was measured with the following corrections: habitual, manifest, manifest with +2.00-D add, and habitual with +2.00-D add (6-month visit only). At baseline, accommodative lag was higher (1.72 ± 0.37 D; mean ± SD) when measured with manifest correction than with habitual correction (1.51 ± 0.50; P < 0.05). This higher lag with manifest correction correlated with a larger amount of habitual undercorrection at baseline (r = -0.29, P = 0.009). A +2.00-D add over the manifest correction reduced lag by 0.45 ± 0.34 D at baseline and 0.33 ± 0.38 D at the 6-month visit. Lag results at 6 months were not different between PAL and SVL wearers (P = 0.92). A +2.00-D bifocal add did not eliminate accommodative lag and reduced lag by less than 25% of the bifocal power, indicating that children mainly responded to a bifocal by decreasing accommodation. If myopic progression is substantial, measuring lag with full correction can overestimate the hyperopic retinal blur that a child most recently experienced. (ClinicalTrials.gov number, NCT00335049.).

  20. The Effect of Bifocal Add on Accommodative Lag in Myopic Children with High Accommodative Lag

    PubMed Central

    Mutti, Donald O.; Zadnik, Karla

    2010-01-01

    Purpose. To determine the effect of a bifocal add and manifest correction on accommodative lag in myopic children with high accommodative lag, who have been reported to have the greatest reduction in myopia progression with progressive addition lenses (PALs). Methods. Monocular accommodative lag to a 4-D Badal stimulus was measured on two occasions 6 months apart in 83 children (mean ± SD age, 9.9 ± 1.3 years) with high lag randomized to wearing single-vision lenses (SVLs) or PALs. Accommodative lag was measured with the following corrections: habitual, manifest, manifest with +2.00-D add, and habitual with +2.00-D add (6-month visit only). Results. At baseline, accommodative lag was higher (1.72 ± 0.37 D; mean ± SD) when measured with manifest correction than with habitual correction (1.51 ± 0.50; P < 0.05). This higher lag with manifest correction correlated with a larger amount of habitual undercorrection at baseline (r = −0.29, P = 0.009). A +2.00-D add over the manifest correction reduced lag by 0.45 ± 0.34 D at baseline and 0.33 ± 0.38 D at the 6-month visit. Lag results at 6 months were not different between PAL and SVL wearers (P = 0.92). Conclusions. A +2.00-D bifocal add did not eliminate accommodative lag and reduced lag by less than 25% of the bifocal power, indicating that children mainly responded to a bifocal by decreasing accommodation. If myopic progression is substantial, measuring lag with full correction can overestimate the hyperopic retinal blur that a child most recently experienced. (ClinicalTrials.gov number, NCT00335049.) PMID:20688729

  1. Influence of oxidized purine processing on strand directionality of mismatch repair.

    PubMed

    Repmann, Simone; Olivera-Harris, Maite; Jiricny, Josef

    2015-04-17

    Replicative DNA polymerases are high fidelity enzymes that misincorporate nucleotides into nascent DNA with a frequency lower than [1/10(5)], and this precision is improved to about [1/10(7)] by their proofreading activity. Because this fidelity is insufficient to replicate most genomes without error, nature evolved postreplicative mismatch repair (MMR), which improves the fidelity of DNA replication by up to 3 orders of magnitude through correcting biosynthetic errors that escaped proofreading. MMR must be able to recognize non-Watson-Crick base pairs and excise the misincorporated nucleotides from the nascent DNA strand, which carries by definition the erroneous genetic information. In eukaryotes, MMR is believed to be directed to the nascent strand by preexisting discontinuities such as gaps between Okazaki fragments in the lagging strand or breaks in the leading strand generated by the mismatch-activated endonuclease of the MutL homologs PMS1 in yeast and PMS2 in vertebrates. We recently demonstrated that the eukaryotic MMR machinery can make use also of strand breaks arising during excision of uracils or ribonucleotides from DNA. We now show that intermediates of MutY homolog-dependent excision of adenines mispaired with 8-oxoguanine (G(O)) also act as MMR initiation sites in extracts of human cells or Xenopus laevis eggs. Unexpectedly, G(O)/C pairs were not processed in these extracts and failed to affect MMR directionality, but extracts supplemented with exogenous 8-oxoguanine DNA glycosylase (OGG1) did so. Because OGG1-mediated excision of G(O) might misdirect MMR to the template strand, our findings suggest that OGG1 activity might be inhibited during MMR. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Ethylene Control of Anthocyanin Synthesis in Sorghum

    PubMed Central

    Craker, L. E.; Standley, L. A.; Starbuck, M. J.

    1971-01-01

    Light-induced anthocyanin synthesis in Sorghum vulgare L. seedlings was both promoted and inhibited by ethylene treatment. The rate of anthocyanin formation in sorghum tissue was dependent upon the time of ethylene treatment in relation to light exposure and the stage of the anthocyanin synthesis process. Those plants receiving ethylene treatment during the early lag phase of anthocyanin synthesis had higher anthocyanin content at 24 hours than control plants receiving no ethylene treatment. Plants receiving ethylene treatment after the lag phase had lower anthocyanin content at 24 hours than control plants receiving no ethylene treatment. PMID:16657796

  3. Two distinct mechanisms ensure transcriptional polarity in double-stranded RNA bacteriophages.

    PubMed

    Yang, Hongyan; Makeyev, Eugene V; Butcher, Sarah J; Gaidelyte, Ausra; Bamford, Dennis H

    2003-01-01

    In most double-stranded RNA (dsRNA) viruses, RNA transcription occurs inside a polymerase (Pol) complex particle, which contains an RNA-dependent RNA Pol subunit as a minor component. Only plus- but not minus-sense copies of genomic segments are produced during this reaction. In the case of phi6, a dsRNA bacteriophage from the Cystoviridae family, isolated Pol synthesizes predominantly plus strands using virus-specific dsRNAs in vitro, thus suggesting that Pol template preferences determine the transcriptional polarity. Here, we dissect transcription reactions catalyzed by Pol complexes and Pol subunits of two other cystoviruses, phi8 and phi13. While both Pol complexes synthesize exclusively plus strands over a wide range of conditions, isolated Pol subunits can be stimulated by Mn(2+) to produce minus-sense copies on phi13 dsRNA templates. Importantly, all three Pol subunits become more prone to the native-like plus-strand synthesis when the dsRNA templates (including phi13 dsRNA) are activated by denaturation before the reaction. Based on these and earlier observations, we propose a model of transcriptional polarity in Cystoviridae controlled on two independent levels: Pol affinity to plus-strand initiation sites and accessibility of these sites to the Pol in a single-stranded form.

  4. Two Distinct Mechanisms Ensure Transcriptional Polarity in Double-Stranded RNA Bacteriophages

    PubMed Central

    Yang, Hongyan; Makeyev, Eugene V.; Butcher, Sarah J.; Gaidelyte·, Aušra; Bamford, Dennis H.

    2003-01-01

    In most double-stranded RNA (dsRNA) viruses, RNA transcription occurs inside a polymerase (Pol) complex particle, which contains an RNA-dependent RNA Pol subunit as a minor component. Only plus- but not minus-sense copies of genomic segments are produced during this reaction. In the case of φ6, a dsRNA bacteriophage from the Cystoviridae family, isolated Pol synthesizes predominantly plus strands using virus-specific dsRNAs in vitro, thus suggesting that Pol template preferences determine the transcriptional polarity. Here, we dissect transcription reactions catalyzed by Pol complexes and Pol subunits of two other cystoviruses, φ8 and φ13. While both Pol complexes synthesize exclusively plus strands over a wide range of conditions, isolated Pol subunits can be stimulated by Mn2+ to produce minus-sense copies on φ13 dsRNA templates. Importantly, all three Pol subunits become more prone to the native-like plus-strand synthesis when the dsRNA templates (including φ13 dsRNA) are activated by denaturation before the reaction. Based on these and earlier observations, we propose a model of transcriptional polarity in Cystoviridae controlled on two independent levels: Pol affinity to plus-strand initiation sites and accessibility of these sites to the Pol in a single-stranded form. PMID:12502836

  5. Pausing kinetics dominates strand-displacement polymerization by reverse transcriptase

    PubMed Central

    Malik, Omri; Khamis, Hadeel; Rudnizky, Sergei; Marx, Ailie

    2017-01-01

    Abstract Reverse transcriptase (RT) catalyzes the conversion of the viral RNA into an integration-competent double-stranded DNA, with a variety of enzymatic activities that include the ability to displace a non-template strand concomitantly with polymerization. Here, using high-resolution optical tweezers to follow the activity of the murine leukemia Virus RT, we show that strand-displacement polymerization is frequently interrupted. Abundant pauses are modulated by the strength of the DNA duplex ∼8 bp ahead, indicating the existence of uncharacterized RT/DNA interactions, and correspond to backtracking of the enzyme, whose recovery is also modulated by the duplex strength. Dissociation and reinitiation events, which induce long periods of inactivity and are likely the rate-limiting step in the synthesis of the genome in vivo, are modulated by the template structure and the viral nucleocapsid protein. Our results emphasize the potential regulatory role of conserved structural motifs, and may provide useful information for the development of potent and specific inhibitors. PMID:28973474

  6. Purification of Single-Stranded cDNA Based on RNA Degradation Treatment and Adsorption Chromatography.

    PubMed

    Trujillo-Esquivel, Elías; Franco, Bernardo; Flores-Martínez, Alberto; Ponce-Noyola, Patricia; Mora-Montes, Héctor M

    2016-08-02

    Analysis of gene expression is a common research tool to study networks controlling gene expression, the role of genes with unknown function, and environmentally induced responses of organisms. Most of the analytical tools used to analyze gene expression rely on accurate cDNA synthesis and quantification to obtain reproducible and quantifiable results. Thus far, most commercial kits for isolation and purification of cDNA target double-stranded molecules, which do not accurately represent the abundance of transcripts. In the present report, we provide a simple and fast method to purify single-stranded cDNA, exhibiting high purity and yield. This method is based on the treatment with RNase H and RNase A after cDNA synthesis, followed by separation in silica spin-columns and ethanol precipitation. In addition, our method avoids the use of DNase I to eliminate genomic DNA from RNA preparations, which improves cDNA yield. As a case report, our method proved to be useful in the purification of single-stranded cDNA from the pathogenic fungus Sporothrix schenckii.

  7. SNMIB/Apollo protects leading-strand telomeres against NHEJ-mediated repair.

    PubMed

    Lam, Yung C; Akhter, Shamima; Gu, Peili; Ye, Jing; Poulet, Anaïs; Giraud-Panis, Marie-Josèphe; Bailey, Susan M; Gilson, Eric; Legerski, Randy J; Chang, Sandy

    2010-07-07

    Progressive telomere attrition or deficiency of the protective shelterin complex elicits a DNA damage response as a result of a cell's inability to distinguish dysfunctional telomeric ends from DNA double-strand breaks. SNMIB/Apollo is a shelterin-associated protein and a member of the SMN1/PSO2 nuclease family that localizes to telomeres through its interaction with TRF2. Here, we generated SNMIB/Apollo knockout mouse embryo fibroblasts (MEFs) to probe the function of SNMIB/Apollo at mammalian telomeres. SNMIB/Apollo null MEFs exhibit an increased incidence of G2 chromatid-type fusions involving telomeres created by leading-strand DNA synthesis, reflective of a failure to protect these telomeres after DNA replication. Mutations within SNMIB/Apollo's conserved nuclease domain failed to suppress this phenotype, suggesting that its nuclease activity is required to protect leading-strand telomeres. SNMIB/Apollo(-/-)ATM(-/-) MEFs display robust telomere fusions when Trf2 is depleted, indicating that ATM is dispensable for repair of uncapped telomeres in this setting. Our data implicate the 5'-3' exonuclease function of SNM1B/Apollo in the generation of 3' single-stranded overhangs at newly replicated leading-strand telomeres to protect them from engaging the non-homologous end-joining pathway.

  8. Synthesis of double-stranded RNA in a virus-enriched fraction from Agaricus bisporus

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

    Sriskantha, A.; Wach, P.; Schlagnhaufer, B.

    Partially purified virus preparations from sporophores of Agaricus bisporus affected with LaFrance disease had up to a 15-fold-higher RNA-dependent RNA polymerase activity than did comparable preparations from health sporophores. Enzyme activity was dependent upon the presence of Mg/sup 2 +/ and the four nucleoside triphosphates and was insensitive to actinomycin D, ..cap alpha..-amanitin, and rifampin. The /sup 3/H-labeled enzyme reaction products were double-stranded RNA (dsRNA) as indicated by CF-11 cellulose column chromatography and by their ionic-strength-dependent sensitivity to hydrolysis by RNase A. The principal dsRNA products had estimated molecular weights of 4.3 /times/ 10/sup 6/ and 1.4 /times/ 10/sup 6/.more » Cs/sub 2/SO/sub 4/ equilibrium centrifugation of the virus preparation resolved a single peak of RNA polymerase activity that banded with a 35-nm spherical virus particle containing dsRNAs with molecular weights of 4.3 /times/ 10/sup 6/ and 1.4 /times/ 10/sup 6/. The data suggest that the RNA-dependent RNA polymerase associated with the 35-nm spherical virus is a replicase which catalyzes the synthesis of the genomic dsRNAs.« less

  9. Srs2 promotes synthesis-dependent strand annealing by disrupting DNA polymerase δ-extending D-loops

    PubMed Central

    Liu, Jie; Ede, Christopher; Wright, William D; Gore, Steven K; Jenkins, Shirin S; Freudenthal, Bret D; Todd Washington, M; Veaute, Xavier; Heyer, Wolf-Dietrich

    2017-01-01

    Synthesis-dependent strand annealing (SDSA) is the preferred mode of homologous recombination in somatic cells leading to an obligatory non-crossover outcome, thus avoiding the potential for chromosomal rearrangements and loss of heterozygosity. Genetic analysis identified the Srs2 helicase as a prime candidate to promote SDSA. Here, we demonstrate that Srs2 disrupts D-loops in an ATP-dependent fashion and with a distinct polarity. Specifically, we partly reconstitute the SDSA pathway using Rad51, Rad54, RPA, RFC, DNA Polymerase δ with different forms of PCNA. Consistent with genetic data showing the requirement for SUMO and PCNA binding for the SDSA role of Srs2, Srs2 displays a slight but significant preference to disrupt extending D-loops over unextended D-loops when SUMOylated PCNA is present, compared to unmodified PCNA or monoubiquitinated PCNA. Our data establish a biochemical mechanism for the role of Srs2 in crossover suppression by promoting SDSA through disruption of extended D-loops. DOI: http://dx.doi.org/10.7554/eLife.22195.001 PMID:28535142

  10. Interstrand cross-links arising from strand breaks at true abasic sites in duplex DNA

    PubMed Central

    Yang, Zhiyu; Price, Nathan E.; Johnson, Kevin M.

    2017-01-01

    Abstract Interstrand cross-links are exceptionally bioactive DNA lesions. Endogenous generation of interstrand cross-links in genomic DNA may contribute to aging, neurodegeneration, and cancer. Abasic (Ap) sites are common lesions in genomic DNA that readily undergo spontaneous and amine-catalyzed strand cleavage reactions that generate a 2,3-didehydro-2,3-dideoxyribose sugar remnant (3’ddR5p) at the 3’-terminus of the strand break. Interestingly, this strand scission process leaves an electrophilic α,β-unsaturated aldehyde residue embedded within the resulting nicked duplex. Here we present evidence that 3’ddR5p derivatives generated by spermine-catalyzed strand cleavage at Ap sites in duplex DNA can react with adenine residues on the opposing strand to generate a complex lesion consisting of an interstrand cross-link adjacent to a strand break. The cross-link blocks DNA replication by ϕ29 DNA polymerase, a highly processive polymerase enzyme that couples synthesis with strand displacement. This suggests that 3’ddR5p-derived cross-links have the potential to block critical cellular DNA transactions that require strand separation. LC-MS/MS methods developed herein provide powerful tools for studying the occurrence and properties of these cross-links in biochemical and biological systems. PMID:28531327

  11. Genome instabilities arising from ribonucleotides in DNA.

    PubMed

    Klein, Hannah L

    2017-08-01

    Genomic DNA is transiently contaminated with ribonucleotide residues during the process of DNA replication through misincorporation by the replicative DNA polymerases α, δ and ε, and by the normal replication process on the lagging strand, which uses RNA primers. These ribonucleotides are efficiently removed during replication by RNase H enzymes and the lagging strand synthesis machinery. However, when ribonucleotides remain in DNA they can distort the DNA helix, affect machineries for DNA replication, transcription and repair, and can stimulate genomic instabilities which are manifest as increased mutation, recombination and chromosome alterations. The genomic instabilities associated with embedded ribonucleotides are considered here, along with a discussion of the origin of the lesions that stimulate particular classes of instabilities. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Effect of lag time distribution on the lag phase of bacterial growth - a Monte Carlo analysis

    USDA-ARS?s Scientific Manuscript database

    The objective of this study is to use Monte Carlo simulation to evaluate the effect of lag time distribution of individual bacterial cells incubated under isothermal conditions on the development of lag phase. The growth of bacterial cells of the same initial concentration and mean lag phase durati...

  13. A way around the Nyquist lag

    NASA Astrophysics Data System (ADS)

    Penland, C.

    2017-12-01

    One way to test for the linearity of a multivariate system is to perform Linear Inverse Modeling (LIM) to a multivariate time series. LIM yields an estimated operator by combining a lagged covariance matrix with the contemporaneous covariance matrix. If the underlying dynamics is linear, the resulting dynamical description should not depend on the particular lag at which the lagged covariance matrix is estimated. This test is known as the "tau test." The tau test will be severely compromised if the lag at which the analysis is performed is approximately half the period of an internal oscillation frequency. In this case, the tau test will fail even though the dynamics are actually linear. Thus, until now, the tau test has only been possible for lags smaller than this "Nyquist lag." In this poster, we investigate the use of Hilbert transforms as a way to avoid the problems associated with Nyquist lags. By augmenting the data with dimensions orthogonal to those spanning the original system, information that would be inaccessible to LIM in its original form may be sampled.

  14. Mechanism for CCC DNA synthesis in hepadnaviruses.

    PubMed

    Sohn, Ji A; Litwin, Samuel; Seeger, Christoph

    2009-11-30

    Hepadnavirus replication requires the synthesis of a covalently closed circular (CCC) DNA from the relaxed circular (RC) viral genome by an unknown mechanism. CCC DNA formation could require enzymatic activities of the viral reverse transcriptase (RT), or cellular DNA repair enzymes, or both. Physical mapping of the 5' and 3' ends of RC DNA and sequence analysis of CCC DNA revealed that CCC DNA synthesis requires the removal of the RT and an RNA oligomer from the 5' ends of minus and plus strand DNA, respectively, removal of sequences from the terminally redundant minus strand, completion of the less than full-length plus strand, and ligation of the ends. Two models have been proposed that could explain CCC DNA formation. The first (model 1) invokes a role for the RT to catalyze a cleavage-ligation reaction leading to the formation of a unit length minus strand in CCC DNA and a DNA repair reaction for the completion and ligation of plus strand DNA; the second (model 2) predicts that CCC DNA formation depends entirely on cellular DNA repair enzymes. To determine which mechanism is utilized, we developed cell lines expressing duck hepatitis B virus genomes carrying mutations permitting us to follow the fate of viral DNA sequences during their conversion from RC to CCC DNA. Our results demonstrated that the oligomer at the 5' end of minus strand DNA is completely or at least partially removed prior to CCC DNA synthesis. The results indicated that both RC DNA strands undergo DNA repair reactions carried out by the cellular DNA repair machinery as predicted by model 2. Thus, our study provided the basis for the identification of the cellular components required for CCC DNA formation.

  15. The binding efficiency of RPA to telomeric G-strands folded into contiguous G-quadruplexes is independent of the number of G4 units.

    PubMed

    Lancrey, Astrid; Safa, Layal; Chatain, Jean; Delagoutte, Emmanuelle; Riou, Jean-François; Alberti, Patrizia; Saintomé, Carole

    2018-03-01

    Replication protein A (RPA) is a single-stranded DNA binding protein involved in replication and in telomere maintenance. During telomere replication, G-quadruplexes (G4) can accumulate on the lagging strand template and need to be resolved. It has been shown that human RPA is able to unfold a single G4. Nevertheless, the G-strand of human telomeres is prone to fold into higher-order structures formed by contiguous G-quadruplexes. To understand how RPA deals with these structures, we studied its interaction with telomeric G-strands folding into an increasing number of contiguous G4s. The aim of this study was to determine whether the efficiency of binding/unfolding of hRPA to telomeric G-strands depends on the number of G4 units. Our data show that the number n of contiguous G4 units (n ≥ 2) does not affect the efficiency of hRPA to coat transiently exposed single-stranded telomeric G-strands. This feature may be essential in preventing instability due to G4 structures during telomere replication. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  16. Strand specific RNA-sequencing and membrane lipid profiling reveals growth phase-dependent cold stress response mechanisms in Listeria monocytogenes.

    PubMed

    Hingston, Patricia; Chen, Jessica; Allen, Kevin; Truelstrup Hansen, Lisbeth; Wang, Siyun

    2017-01-01

    The human pathogen Listeria monocytogenes continues to pose a challenge in the food industry, where it is known to contaminate ready-to-eat foods and grow during refrigerated storage. Increased knowledge of the cold-stress response of this pathogen will enhance the ability to control it in the food-supply-chain. This study utilized strand-specific RNA sequencing and whole cell fatty acid (FA) profiling to characterize the bacterium's cold stress response. RNA and FAs were extracted from a cold-tolerant strain at five time points between early lag phase and late stationary-phase, both at 4°C and 20°C. Overall, more genes (1.3×) were suppressed than induced at 4°C. Late stationary-phase cells exhibited the greatest number (n = 1,431) and magnitude (>1,000-fold) of differentially expressed genes (>2-fold, p<0.05) in response to cold. A core set of 22 genes was upregulated at all growth phases, including nine genes required for branched-chain fatty acid (BCFA) synthesis, the osmolyte transporter genes opuCBCD, and the internalin A and D genes. Genes suppressed at 4°C were largely associated with cobalamin (B12) biosynthesis or the production/export of cell wall components. Antisense transcription accounted for up to 1.6% of total mapped reads with higher levels (2.5×) observed at 4°C than 20°C. The greatest number of upregulated antisense transcripts at 4°C occurred in early lag phase, however, at both temperatures, antisense expression levels were highest in late stationary-phase cells. Cold-induced FA membrane changes included a 15% increase in the proportion of BCFAs and a 15% transient increase in unsaturated FAs between lag and exponential phase. These increases probably reduced the membrane phase transition temperature until optimal levels of BCFAs could be produced. Collectively, this research provides new information regarding cold-induced membrane composition changes in L. monocytogenes, the growth-phase dependency of its cold-stress regulon, and the

  17. Interstrand cross-links arising from strand breaks at true abasic sites in duplex DNA.

    PubMed

    Yang, Zhiyu; Price, Nathan E; Johnson, Kevin M; Wang, Yinsheng; Gates, Kent S

    2017-06-20

    Interstrand cross-links are exceptionally bioactive DNA lesions. Endogenous generation of interstrand cross-links in genomic DNA may contribute to aging, neurodegeneration, and cancer. Abasic (Ap) sites are common lesions in genomic DNA that readily undergo spontaneous and amine-catalyzed strand cleavage reactions that generate a 2,3-didehydro-2,3-dideoxyribose sugar remnant (3'ddR5p) at the 3'-terminus of the strand break. Interestingly, this strand scission process leaves an electrophilic α,β-unsaturated aldehyde residue embedded within the resulting nicked duplex. Here we present evidence that 3'ddR5p derivatives generated by spermine-catalyzed strand cleavage at Ap sites in duplex DNA can react with adenine residues on the opposing strand to generate a complex lesion consisting of an interstrand cross-link adjacent to a strand break. The cross-link blocks DNA replication by ϕ29 DNA polymerase, a highly processive polymerase enzyme that couples synthesis with strand displacement. This suggests that 3'ddR5p-derived cross-links have the potential to block critical cellular DNA transactions that require strand separation. LC-MS/MS methods developed herein provide powerful tools for studying the occurrence and properties of these cross-links in biochemical and biological systems. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  18. Compiler-aided systematic construction of large-scale DNA strand displacement circuits using unpurified components

    NASA Astrophysics Data System (ADS)

    Thubagere, Anupama J.; Thachuk, Chris; Berleant, Joseph; Johnson, Robert F.; Ardelean, Diana A.; Cherry, Kevin M.; Qian, Lulu

    2017-02-01

    Biochemical circuits made of rationally designed DNA molecules are proofs of concept for embedding control within complex molecular environments. They hold promise for transforming the current technologies in chemistry, biology, medicine and material science by introducing programmable and responsive behaviour to diverse molecular systems. As the transformative power of a technology depends on its accessibility, two main challenges are an automated design process and simple experimental procedures. Here we demonstrate the use of circuit design software, combined with the use of unpurified strands and simplified experimental procedures, for creating a complex DNA strand displacement circuit that consists of 78 distinct species. We develop a systematic procedure for overcoming the challenges involved in using unpurified DNA strands. We also develop a model that takes synthesis errors into consideration and semi-quantitatively reproduces the experimental data. Our methods now enable even novice researchers to successfully design and construct complex DNA strand displacement circuits.

  19. The Weighted-Average Lagged Ensemble.

    PubMed

    DelSole, T; Trenary, L; Tippett, M K

    2017-11-01

    A lagged ensemble is an ensemble of forecasts from the same model initialized at different times but verifying at the same time. The skill of a lagged ensemble mean can be improved by assigning weights to different forecasts in such a way as to maximize skill. If the forecasts are bias corrected, then an unbiased weighted lagged ensemble requires the weights to sum to one. Such a scheme is called a weighted-average lagged ensemble. In the limit of uncorrelated errors, the optimal weights are positive and decay monotonically with lead time, so that the least skillful forecasts have the least weight. In more realistic applications, the optimal weights do not always behave this way. This paper presents a series of analytic examples designed to illuminate conditions under which the weights of an optimal weighted-average lagged ensemble become negative or depend nonmonotonically on lead time. It is shown that negative weights are most likely to occur when the errors grow rapidly and are highly correlated across lead time. The weights are most likely to behave nonmonotonically when the mean square error is approximately constant over the range forecasts included in the lagged ensemble. An extreme example of the latter behavior is presented in which the optimal weights vanish everywhere except at the shortest and longest lead times.

  20. 47 CFR 52.103 - Lag times.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 47 Telecommunication 3 2010-10-01 2010-10-01 false Lag times. 52.103 Section 52.103... Free Numbers § 52.103 Lag times. (a) Definitions. As used in this section, the following definitions... and an exchange carrier intercept recording is being provided. (3) Lag Time. The interval between a...

  1. 47 CFR 52.103 - Lag times.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 47 Telecommunication 3 2012-10-01 2012-10-01 false Lag times. 52.103 Section 52.103... Free Numbers § 52.103 Lag times. (a) Definitions. As used in this section, the following definitions... and an exchange carrier intercept recording is being provided. (3) Lag Time. The interval between a...

  2. 47 CFR 52.103 - Lag times.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 47 Telecommunication 3 2011-10-01 2011-10-01 false Lag times. 52.103 Section 52.103... Free Numbers § 52.103 Lag times. (a) Definitions. As used in this section, the following definitions... and an exchange carrier intercept recording is being provided. (3) Lag Time. The interval between a...

  3. 47 CFR 52.103 - Lag times.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 47 Telecommunication 3 2013-10-01 2013-10-01 false Lag times. 52.103 Section 52.103... Free Numbers § 52.103 Lag times. (a) Definitions. As used in this section, the following definitions... and an exchange carrier intercept recording is being provided. (3) Lag Time. The interval between a...

  4. Whole Genome Sequence Analysis of Mutations Accumulated in rad27Δ Yeast Strains with Defects in the Processing of Okazaki Fragments Indicates Template-Switching Events

    PubMed Central

    Omer, Sumita; Lavi, Bar; Mieczkowski, Piotr A.; Covo, Shay; Hazkani-Covo, Einat

    2017-01-01

    Okazaki fragments that are formed during lagging strand DNA synthesis include an initiating primer consisting of both RNA and DNA. The RNA fragment must be removed before the fragments are joined. In Saccharomyces cerevisiae, a key player in this process is the structure-specific flap endonuclease, Rad27p (human homolog FEN1). To obtain a genomic view of the mutational consequence of loss of RAD27, a S. cerevisiae rad27Δ strain was subcultured for 25 generations and sequenced using Illumina paired-end sequencing. Out of the 455 changes observed in 10 colonies isolated the two most common types of events were insertions or deletions (INDELs) in simple sequence repeats (SSRs) and INDELs mediated by short direct repeats. Surprisingly, we also detected a previously neglected class of 21 template-switching events. These events were presumably generated by quasi-palindrome to palindrome correction, as well as palindrome elongation. The formation of these events is best explained by folding back of the stalled nascent strand and resumption of DNA synthesis using the same nascent strand as a template. Evidence of quasi-palindrome to palindrome correction that could be generated by template switching appears also in yeast genome evolution. Out of the 455 events, 55 events appeared in multiple isolates; further analysis indicates that these loci are mutational hotspots. Since Rad27 acts on the lagging strand when the leading strand should not contain any gaps, we propose a mechanism favoring intramolecular strand switching over an intermolecular mechanism. We note that our results open new ways of understanding template switching that occurs during genome instability and evolution. PMID:28974572

  5. Determinants of Base-Pair Substitution Patterns Revealed by Whole-Genome Sequencing of DNA Mismatch Repair Defective Escherichia coli.

    PubMed

    Foster, Patricia L; Niccum, Brittany A; Popodi, Ellen; Townes, Jesse P; Lee, Heewook; MohammedIsmail, Wazim; Tang, Haixu

    2018-06-15

    Mismatch repair (MMR) is a major contributor to replication fidelity, but its impact varies with sequence context and the nature of the mismatch. Mutation accumulation experiments followed by whole-genome sequencing of MMR-defective E. coli strains yielded ≈30,000 base-pair substitutions, revealing mutational patterns across the entire chromosome. The base-pair substitution spectrum was dominated by A:T > G:C transitions, which occurred predominantly at the center base of 5'N A C3'+5'G T N3' triplets. Surprisingly, growth on minimal medium or at low temperature attenuated these mutations. Mononucleotide runs were also hotspots for base-pair substitutions, and the rate at which these occurred increased with run length. Comparison with ≈2000 base-pair substitutions accumulated in MMR-proficient strains revealed that both kinds of hotspots appeared in the wild-type spectrum and so are likely to be sites of frequent replication errors. In MMR-defective strains transitions were strand biased, occurring twice as often when A and C rather than T and G were on the lagging-strand template. Loss of nucleotide diphosphate kinase increases the cellular concentration of dCTP, which resulted in increased rates of mutations due to misinsertion of C opposite A and T. In an mmr ndk double mutant strain, these mutations were more frequent when the template A and T were on the leading strand, suggesting that lagging-strand synthesis was more error-prone or less well corrected by proofreading than was leading strand synthesis. Copyright © 2018, Genetics.

  6. Robust Short-Lag Spatial Coherence Imaging.

    PubMed

    Nair, Arun Asokan; Tran, Trac Duy; Bell, Muyinatu A Lediju

    2018-03-01

    Short-lag spatial coherence (SLSC) imaging displays the spatial coherence between backscattered ultrasound echoes instead of their signal amplitudes and is more robust to noise and clutter artifacts when compared with traditional delay-and-sum (DAS) B-mode imaging. However, SLSC imaging does not consider the content of images formed with different lags, and thus does not exploit the differences in tissue texture at each short-lag value. Our proposed method improves SLSC imaging by weighting the addition of lag values (i.e., M-weighting) and by applying robust principal component analysis (RPCA) to search for a low-dimensional subspace for projecting coherence images created with different lag values. The RPCA-based projections are considered to be denoised versions of the originals that are then weighted and added across lags to yield a final robust SLSC (R-SLSC) image. Our approach was tested on simulation, phantom, and in vivo liver data. Relative to DAS B-mode images, the mean contrast, signal-to-noise ratio (SNR), and contrast-to-noise ratio (CNR) improvements with R-SLSC images are 21.22 dB, 2.54, and 2.36, respectively, when averaged over simulated, phantom, and in vivo data and over all lags considered, which corresponds to mean improvements of 96.4%, 121.2%, and 120.5%, respectively. When compared with SLSC images, the corresponding mean improvements with R-SLSC images were 7.38 dB, 1.52, and 1.30, respectively (i.e., mean improvements of 14.5%, 50.5%, and 43.2%, respectively). Results show great promise for smoothing out the tissue texture of SLSC images and enhancing anechoic or hypoechoic target visibility at higher lag values, which could be useful in clinical tasks such as breast cyst visualization, liver vessel tracking, and obese patient imaging.

  7. The influence of DNA inhibitor synthesis on the induction and repair of double-strand DNA breaks in human lymphocytes under action of radiation with a different linear energy transfer

    NASA Astrophysics Data System (ADS)

    Boreyko, A. V.; Chausov, V. N.; Krasavin, E. A.; Ravnachka, I.; Stukova, S. I.

    2011-07-01

    The influence that inhibitors of repair and replicative DNA synthesis, 1-β-D-arabinofuranosyl-cytosine and hydroxyurea, have on the formation and repair kinetics of double-strand breaks (DSBs) in peripheral human blood lymphocytes under the influence of radiation with a different linear energy transfer (LET) (gamma quanta and accelerated heavy ions) is studied. It is demonstrated that lithium and boron ions with LETs of 20 and 40 keV/μm, respectively, possess higher biological effectiveness with respect to the DNA DSB induction criterion. The value of the relative biological effectiveness of accelerated lithium and boron ions is 1.5 ± 0.1 and 1.6 ± 0.1, respectively. It is found that, upon cell irradiation by gamma quanta in the absence of inhibitors, efficient DNA DSB repair is observed during incubation. Under the conditions of cell incubation and in the presence of inhibitors, some growth in the number of DNA DSBs, rather than a reduction, is observed after 5-h incubation. In the case of the action of accelerated boron ions (as well as gamma quanta), under normal conditions, the efficient repair of induced DNA lesions takes place. Unlike the action of gamma quanta, in the case of cell incubation in the presence of radiomodifiers, the number of induced DNA DSBs falls. These results may testify to the fact that the repair of double-strand DNS breaks takes place under the action of ionizing radiation with a different LET on mammalian cells in the presence of DNA synthesis inhibitors Ara-C and HU. It is concluded that, for cells subject to gamma irradiation, no DNA DSB repair is observed due to the large contribution of single-strand incision DNA breaks formed in the postradiation period in the course of excision nucleotide repair.

  8. Single-stranded DNA-binding Protein in Vitro Eliminates the Orientation-dependent Impediment to Polymerase Passage on CAG/CTG Repeats*

    PubMed Central

    Delagoutte, Emmanuelle; Goellner, Geoffrey M.; Guo, Jie; Baldacci, Giuseppe; McMurray, Cynthia T.

    2008-01-01

    Small insertions and deletions of trinucleotide repeats (TNRs) can occur by polymerase slippage and hairpin formation on either template or newly synthesized strands during replication. Although not predicted by a slippage model, deletions occur preferentially when 5′-CTG is in the lagging strand template and are highly favored over insertion events in rapidly replicating cells. The mechanism for the deletion bias and the orientation dependence of TNR instability is poorly understood. We report here that there is an orientation-dependent impediment to polymerase progression on 5′-CAG and 5′-CTG repeats that can be relieved by the binding of single-stranded DNA-binding protein. The block depends on the primary sequence of the TNR but does not correlate with the thermodynamic stability of hairpins. The orientation-dependent block of polymerase passage is the strongest when 5′-CAG is the template. We propose a “template-push” model in which the slow speed of DNA polymerase across the 5′-CAG leading strand template creates a threat to helicase-polymerase coupling. To prevent uncoupling, the TNR template is pushed out and by-passed. Hairpins do not cause the block, but appear to occur as a consequence of polymerase pass-over. PMID:18263578

  9. Ultrastructure of the replication sites of positive-strand RNA viruses

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

    Harak, Christian; Lohmann, Volker, E-mail: volker_lohmann@med.uni-heidelberg.de

    2015-05-15

    Positive strand RNA viruses replicate in the cytoplasm of infected cells and induce intracellular membranous compartments harboring the sites of viral RNA synthesis. These replication factories are supposed to concentrate the components of the replicase and to shield replication intermediates from the host cell innate immune defense. Virus induced membrane alterations are often generated in coordination with host factors and can be grouped into different morphotypes. Recent advances in conventional and electron microscopy have contributed greatly to our understanding of their biogenesis, but still many questions remain how viral proteins capture membranes and subvert host factors for their need. Inmore » this review, we will discuss different representatives of positive strand RNA viruses and their ways of hijacking cellular membranes to establish replication complexes. We will further focus on host cell factors that are critically involved in formation of these membranes and how they contribute to viral replication. - Highlights: • Positive strand RNA viruses induce massive membrane alterations. • Despite the great diversity, replication complexes share many similarities. • Host factors play a pivotal role in replication complex biogenesis. • Use of the same host factors by several viruses hints to similar functions.« less

  10. Mapping vaccinia virus DNA replication origins at nucleotide level by deep sequencing.

    PubMed

    Senkevich, Tatiana G; Bruno, Daniel; Martens, Craig; Porcella, Stephen F; Wolf, Yuri I; Moss, Bernard

    2015-09-01

    Poxviruses reproduce in the host cytoplasm and encode most or all of the enzymes and factors needed for expression and synthesis of their double-stranded DNA genomes. Nevertheless, the mode of poxvirus DNA replication and the nature and location of the replication origins remain unknown. A current but unsubstantiated model posits only leading strand synthesis starting at a nick near one covalently closed end of the genome and continuing around the other end to generate a concatemer that is subsequently resolved into unit genomes. The existence of specific origins has been questioned because any plasmid can replicate in cells infected by vaccinia virus (VACV), the prototype poxvirus. We applied directional deep sequencing of short single-stranded DNA fragments enriched for RNA-primed nascent strands isolated from the cytoplasm of VACV-infected cells to pinpoint replication origins. The origins were identified as the switching points of the fragment directions, which correspond to the transition from continuous to discontinuous DNA synthesis. Origins containing a prominent initiation point mapped to a sequence within the hairpin loop at one end of the VACV genome and to the same sequence within the concatemeric junction of replication intermediates. These findings support a model for poxvirus genome replication that involves leading and lagging strand synthesis and is consistent with the requirements for primase and ligase activities as well as earlier electron microscopic and biochemical studies implicating a replication origin at the end of the VACV genome.

  11. Annealing of Complementary DNA Sequences During Double-Strand Break Repair in Drosophila Is Mediated by the Ortholog of SMARCAL1.

    PubMed

    Holsclaw, Julie Korda; Sekelsky, Jeff

    2017-05-01

    DNA double-strand breaks (DSBs) pose a serious threat to genomic integrity. If unrepaired, they can lead to chromosome fragmentation and cell death. If repaired incorrectly, they can cause mutations and chromosome rearrangements. DSBs are repaired using end-joining or homology-directed repair strategies, with the predominant form of homology-directed repair being synthesis-dependent strand annealing (SDSA). SDSA is the first defense against genomic rearrangements and information loss during DSB repair, making it a vital component of cell health and an attractive target for chemotherapeutic development. SDSA has also been proposed to be the primary mechanism for integration of large insertions during genome editing with CRISPR/Cas9. Despite the central role for SDSA in genome stability, little is known about the defining step: annealing. We hypothesized that annealing during SDSA is performed by the annealing helicase SMARCAL1, which can anneal RPA-coated single DNA strands during replication-associated DNA damage repair. We used unique genetic tools in Drosophila melanogaster to test whether the fly ortholog of SMARCAL1, Marcal1, mediates annealing during SDSA. Repair that requires annealing is significantly reduced in Marcal1 null mutants in both synthesis-dependent and synthesis-independent (single-strand annealing) assays. Elimination of the ATP-binding activity of Marcal1 also reduced annealing-dependent repair, suggesting that the annealing activity requires translocation along DNA. Unlike the null mutant, however, the ATP-binding defect mutant showed reduced end joining, shedding light on the interaction between SDSA and end-joining pathways. Copyright © 2017 by the Genetics Society of America.

  12. Reliable method for generating double-stranded DNA vectors containing site-specific base modifications.

    PubMed

    Brégeon, Damien; Doetsch, Paul W

    2004-11-01

    Cells of all living organisms are continuously exposed to physical and chemical agents that damage DNA and alter the integrity of their genomes. Despite the relatively high efficiency of the different repair pathways, some lesions remain in DNA when it is replicated or transcribed. Lesion bypass by DNA and RNA polymerases has been the subject of numerous investigations. However, knowledge of the in vivo mechanism of transcription lesion bypass is very limited because no robust methodology is available. Here we describe a protocol based on the synthesis of a complementary strand of a circular, single-stranded DNA molecule, which allows for the production of large amounts of double-stranded DNA containing a lesion at a specific position in a transcribed sequence. Such constructs can subsequently be used for lesion bypass studies in vivo by RNA polymerase and to ascertain how these events can be affected by the genetic background of the cells.

  13. Single-cell template strand sequencing by Strand-seq enables the characterization of individual homologs.

    PubMed

    Sanders, Ashley D; Falconer, Ester; Hills, Mark; Spierings, Diana C J; Lansdorp, Peter M

    2017-06-01

    The ability to distinguish between genome sequences of homologous chromosomes in single cells is important for studies of copy-neutral genomic rearrangements (such as inversions and translocations), building chromosome-length haplotypes, refining genome assemblies, mapping sister chromatid exchange events and exploring cellular heterogeneity. Strand-seq is a single-cell sequencing technology that resolves the individual homologs within a cell by restricting sequence analysis to the DNA template strands used during DNA replication. This protocol, which takes up to 4 d to complete, relies on the directionality of DNA, in which each single strand of a DNA molecule is distinguished based on its 5'-3' orientation. Culturing cells in a thymidine analog for one round of cell division labels nascent DNA strands, allowing for their selective removal during genomic library construction. To preserve directionality of template strands, genomic preamplification is bypassed and labeled nascent strands are nicked and not amplified during library preparation. Each single-cell library is multiplexed for pooling and sequencing, and the resulting sequence data are aligned, mapping to either the minus or plus strand of the reference genome, to assign template strand states for each chromosome in the cell. The major adaptations to conventional single-cell sequencing protocols include harvesting of daughter cells after a single round of BrdU incorporation, bypassing of whole-genome amplification, and removal of the BrdU + strand during Strand-seq library preparation. By sequencing just template strands, the structure and identity of each homolog are preserved.

  14. Repetition and lag effects in movement recognition.

    PubMed

    Hall, C R; Buckolz, E

    1982-03-01

    Whether repetition and lag improve the recognition of movement patterns was investigated. Recognition memory was tested for one repetition, two-repetitions massed, and two-repetitions distributed with movement patterns at lags of 3, 5, 7, and 13. Recognition performance was examined both immediately afterwards and following a 48 hour delay. Both repetition and lag effects failed to be demonstrated, providing some support for the claim that memory is unaffected by repetition at a constant level of processing (Craik & Lockhart, 1972). There was, as expected, a significant decrease in recognition memory following the retention interval, but this appeared unrelated to repetition or lag.

  15. Strand specific RNA-sequencing and membrane lipid profiling reveals growth phase-dependent cold stress response mechanisms in Listeria monocytogenes

    PubMed Central

    Hingston, Patricia; Chen, Jessica; Allen, Kevin; Truelstrup Hansen, Lisbeth

    2017-01-01

    The human pathogen Listeria monocytogenes continues to pose a challenge in the food industry, where it is known to contaminate ready-to-eat foods and grow during refrigerated storage. Increased knowledge of the cold-stress response of this pathogen will enhance the ability to control it in the food-supply-chain. This study utilized strand-specific RNA sequencing and whole cell fatty acid (FA) profiling to characterize the bacterium’s cold stress response. RNA and FAs were extracted from a cold-tolerant strain at five time points between early lag phase and late stationary-phase, both at 4°C and 20°C. Overall, more genes (1.3×) were suppressed than induced at 4°C. Late stationary-phase cells exhibited the greatest number (n = 1,431) and magnitude (>1,000-fold) of differentially expressed genes (>2-fold, p<0.05) in response to cold. A core set of 22 genes was upregulated at all growth phases, including nine genes required for branched-chain fatty acid (BCFA) synthesis, the osmolyte transporter genes opuCBCD, and the internalin A and D genes. Genes suppressed at 4°C were largely associated with cobalamin (B12) biosynthesis or the production/export of cell wall components. Antisense transcription accounted for up to 1.6% of total mapped reads with higher levels (2.5×) observed at 4°C than 20°C. The greatest number of upregulated antisense transcripts at 4°C occurred in early lag phase, however, at both temperatures, antisense expression levels were highest in late stationary-phase cells. Cold-induced FA membrane changes included a 15% increase in the proportion of BCFAs and a 15% transient increase in unsaturated FAs between lag and exponential phase. These increases probably reduced the membrane phase transition temperature until optimal levels of BCFAs could be produced. Collectively, this research provides new information regarding cold-induced membrane composition changes in L. monocytogenes, the growth-phase dependency of its cold-stress regulon, and

  16. Crystal structure of an Okazaki fragment at 2-A resolution

    NASA Technical Reports Server (NTRS)

    Egli, M.; Usman, N.; Zhang, S. G.; Rich, A.

    1992-01-01

    In DNA replication, Okazaki fragments are formed as double-stranded intermediates during synthesis of the lagging strand. They are composed of the growing DNA strand primed by RNA and the template strand. The DNA oligonucleotide d(GGGTATACGC) and the chimeric RNA-DNA oligonucleotide r(GCG)d(TATACCC) were combined to form a synthetic Okazaki fragment and its three-dimensional structure was determined by x-ray crystallography. The fragment adopts an overall A-type conformation with 11 residues per turn. Although the base-pair geometry, particularly in the central TATA part, is distorted, there is no evidence for a transition from the A- to the B-type conformation at the junction between RNA.DNA hybrid and DNA duplex. The RNA trimer may, therefore, lock the complete fragment in an A-type conformation.

  17. Both DNA Polymerases δ and ε Contact Active and Stalled Replication Forks Differently

    PubMed Central

    Yu, Chuanhe; Gan, Haiyun

    2017-01-01

    ABSTRACT Three DNA polymerases, polymerases α, δ, and ε (Pol α, Pol δ, and Pol ε), are responsible for eukaryotic genome duplication. When DNA replication stress is encountered, DNA synthesis stalls until the stress is ameliorated. However, it is not known whether there is a difference in the association of each polymerase with active and stalled replication forks. Here, we show that each DNA polymerase has a distinct pattern of association with active and stalled replication forks. Pol α is enriched at extending Okazaki fragments of active and stalled forks. In contrast, although Pol δ contacts the nascent lagging strands of active and stalled forks, it binds to only the matured (and not elongating) Okazaki fragments of stalled forks. Pol ε has greater contact with the nascent single-stranded DNA (ssDNA) of the leading strand on active forks than on stalled forks. We propose that the configuration of DNA polymerases at stalled forks facilitates the resumption of DNA synthesis after stress removal. PMID:28784720

  18. Spectral lags in different episodes of gamma-ray bursts

    NASA Astrophysics Data System (ADS)

    Jia, LanWei; Yi, TingFeng; Liang, EnWei

    2013-08-01

    A systematical analysis of the spectral lags in different episodes within a gamma-ray burst (GRB) for the BATSE GRB sample is given. The identified episodes are usually a single pulse with mixing of small fluctuations. The spectral lags were calculated for lightcurves in the 25-55 keV and 110-320 keV bands. No universal spectral lag evolution feature in different episodes within a GRB were found for most GRBs. Among 362 bright GRBs that have at least three well-identified episodes, 19 of them show long-to-short lag and 19 of them show short-to-long lag in successive episodes. The other 324 GRBs have no clear evolution trend. Defining the specified lag with the ratio of the spectral lag to the episode duration in 110-320 keV band, no prominent case of specified lag was found showing clear evolution features. The results suggest that the observed spectral lag may contribute to the dynamics and/or the radiation physics of a given emission episode.

  19. Long-Lag, Wide-pulse Gamma-Ray Bursts

    NASA Technical Reports Server (NTRS)

    Norris, J. P.; Bonnell, J. T.; Kazanas, D.; Scargle, . D.; Hakkila, J.; Giblin, T. W.

    2004-01-01

    Currently, the best available probe of the early phase of gamma-ray burst (GRB) jet attributes is the prompt gamma-ray emission, in which several intrinsic and extrinsic variables determine GRB pulse evolution. Bright, usually complex bursts have many narrow pulses that are difficult to model due to overlap. However, the relatively simple, long spectral lag, wide-pulse bursts may have simpler physics and are easier to model. In this work we analyze the temporal and spectral behavior of wide pulses in 24 long-lag bursts, using a pulse model with two shape parameters - width and asymmetry - and the Band spectral model with three shape parameters. We find that pulses in long-lag bursts are distinguished both temporally and spectrally from those in bright bursts: the pulses in long spectral lag bursts are few in number, and approximately 100 times wider (10s of seconds), have systematically lower peaks in vF(v), harder low-energy spectra and softer high-energy spectra. We find that these five pulse descriptors are essentially uncorrelated for our long-lag sample, suggesting that at least approximately 5 parameters are needed to model burst temporal and spectral behavior. However, pulse width is strongly correlated with spectral lag; hence these two parameters may be viewed as mutual surrogates. We infer that accurate formulations for estimating GRB luminosity and total energy will depend on several gamma-ray attributes, at least for long-lag bursts. The prevalence of long-lag bursts near the BATSE trigger threshold, their predominantly low vF(v) spectral peaks, and relatively steep upper power-law spectral indices indicate that Swift will detect many such bursts.

  20. Exonuclease of human DNA polymerase gamma disengages its strand displacement function.

    PubMed

    He, Quan; Shumate, Christie K; White, Mark A; Molineux, Ian J; Yin, Y Whitney

    2013-11-01

    Pol γ, the only DNA polymerase found in human mitochondria, functions in both mtDNA repair and replication. During mtDNA base-excision repair, gaps are created after damaged base excision. Here we show that Pol γ efficiently gap-fills except when the gap is only a single nucleotide. Although wild-type Pol γ has very limited ability for strand displacement DNA synthesis, exo(-) (3'-5' exonuclease-deficient) Pol γ has significantly high activity and rapidly unwinds downstream DNA, synthesizing DNA at a rate comparable to that of the wild-type enzyme on a primer-template. The catalytic subunit Pol γA alone, even when exo(-), is unable to synthesize by strand displacement, making this the only known reaction of Pol γ holoenzyme that has an absolute requirement for the accessory subunit Pol γB. © 2013. Published by Elsevier B.V.

  1. Enzymatic synthesis of long double-stranded DNA labeled with haloderivatives of nucleobases in a precisely pre-determined sequence

    PubMed Central

    2011-01-01

    Background Restriction endonucleases are widely applied in recombinant DNA technology. Among them, enzymes of class IIS, which cleave DNA beyond recognition sites, are especially useful. We use BsaI enzyme for the pinpoint introduction of halogen nucleobases into DNA. This has been done for the purpose of anticancer radio- and phototherapy that is our long-term objective. Results An enzymatic method for synthesizing long double-stranded DNA labeled with the halogen derivatives of nucleobases (Hal-NBs) with 1-bp accuracy has been put forward and successfully tested on three different DNA fragments containing the 5-bromouracil (5-BrU) residue. The protocol assumes enzymatic cleavage of two Polymerase-Chain-Reaction (PCR) fragments containing two recognition sequences for the same or different class IIS restriction endonucleases, where each PCR fragment has a partially complementary cleavage site. These sites are introduced using synthetic DNA primers or are naturally present in the sequence used. The cleavage sites are not compatible, and therefore not susceptible to ligation until they are partially filled with a Hal-NB or original nucleobase, resulting in complementary cohesive end formation. Ligation of these fragments ultimately leads to the required Hal-NB-labeled DNA duplex. With this approach, a synthetic, extremely long DNA fragment can be obtained by means of a multiple assembly reaction (n × maximum PCR product length: n × app. 50 kb). Conclusions The long, precisely labeled DNA duplexes obtained behave in very much the same manner as natural DNA and are beyond the range of chemical synthesis. Moreover, the conditions of synthesis closely resemble the natural ones, and all the artifacts accompanying the chemical synthesis of DNA are thus eliminated. The approach proposed seems to be completely general and could be used to label DNA at multiple pre-determined sites and with halogen derivatives of any nucleobase. Access to DNAs labeled with Hal-NBs at

  2. Enzymatic synthesis of long double-stranded DNA labeled with haloderivatives of nucleobases in a precisely pre-determined sequence.

    PubMed

    Sobolewski, Ireneusz; Polska, Katarzyna; Zylicz-Stachula, Agnieszka; Jeżewska-Frąckowiak, Joanna; Rak, Janusz; Skowron, Piotr

    2011-08-24

    Restriction endonucleases are widely applied in recombinant DNA technology. Among them, enzymes of class IIS, which cleave DNA beyond recognition sites, are especially useful. We use BsaI enzyme for the pinpoint introduction of halogen nucleobases into DNA. This has been done for the purpose of anticancer radio- and phototherapy that is our long-term objective. An enzymatic method for synthesizing long double-stranded DNA labeled with the halogen derivatives of nucleobases (Hal-NBs) with 1-bp accuracy has been put forward and successfully tested on three different DNA fragments containing the 5-bromouracil (5-BrU) residue. The protocol assumes enzymatic cleavage of two Polymerase-Chain-Reaction (PCR) fragments containing two recognition sequences for the same or different class IIS restriction endonucleases, where each PCR fragment has a partially complementary cleavage site. These sites are introduced using synthetic DNA primers or are naturally present in the sequence used. The cleavage sites are not compatible, and therefore not susceptible to ligation until they are partially filled with a Hal-NB or original nucleobase, resulting in complementary cohesive end formation. Ligation of these fragments ultimately leads to the required Hal-NB-labeled DNA duplex. With this approach, a synthetic, extremely long DNA fragment can be obtained by means of a multiple assembly reaction (n × maximum PCR product length: n × app. 50 kb). The long, precisely labeled DNA duplexes obtained behave in very much the same manner as natural DNA and are beyond the range of chemical synthesis. Moreover, the conditions of synthesis closely resemble the natural ones, and all the artifacts accompanying the chemical synthesis of DNA are thus eliminated. The approach proposed seems to be completely general and could be used to label DNA at multiple pre-determined sites and with halogen derivatives of any nucleobase. Access to DNAs labeled with Hal-NBs at specific position is an

  3. Human DNA polymerase η accommodates RNA for strand extension.

    PubMed

    Su, Yan; Egli, Martin; Guengerich, F Peter

    2017-11-03

    Ribonucleotides are the natural analogs of deoxyribonucleotides, which can be misinserted by DNA polymerases, leading to the most abundant DNA lesions in genomes. During replication, DNA polymerases tolerate patches of ribonucleotides on the parental strands to different extents. The majority of human DNA polymerases have been reported to misinsert ribonucleotides into genomes. However, only PrimPol, DNA polymerase α, telomerase, and the mitochondrial human DNA polymerase (hpol) γ have been shown to tolerate an entire RNA strand. Y-family hpol η is known for translesion synthesis opposite the UV-induced DNA lesion cyclobutane pyrimidine dimer and was recently found to incorporate ribonucleotides into DNA. Here, we report that hpol η is able to bind DNA/DNA, RNA/DNA, and DNA/RNA duplexes with similar affinities. In addition, hpol η, as well as another Y-family DNA polymerase, hpol κ, accommodates RNA as one of the two strands during primer extension, mainly by inserting dNMPs opposite unmodified templates or DNA lesions, such as 8-oxo-2'-deoxyguanosine or cyclobutane pyrimidine dimer, even in the presence of an equal amount of the DNA/DNA substrate. The discovery of this RNA-accommodating ability of hpol η redefines the traditional concept of human DNA polymerases and indicates potential new functions of hpol η in vivo . © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. Statistical properties of Fourier-based time-lag estimates

    NASA Astrophysics Data System (ADS)

    Epitropakis, A.; Papadakis, I. E.

    2016-06-01

    Context. The study of X-ray time-lag spectra in active galactic nuclei (AGN) is currently an active research area, since it has the potential to illuminate the physics and geometry of the innermost region (I.e. close to the putative super-massive black hole) in these objects. To obtain reliable information from these studies, the statistical properties of time-lags estimated from data must be known as accurately as possible. Aims: We investigated the statistical properties of Fourier-based time-lag estimates (I.e. based on the cross-periodogram), using evenly sampled time series with no missing points. Our aim is to provide practical "guidelines" on estimating time-lags that are minimally biased (I.e. whose mean is close to their intrinsic value) and have known errors. Methods: Our investigation is based on both analytical work and extensive numerical simulations. The latter consisted of generating artificial time series with various signal-to-noise ratios and sampling patterns/durations similar to those offered by AGN observations with present and past X-ray satellites. We also considered a range of different model time-lag spectra commonly assumed in X-ray analyses of compact accreting systems. Results: Discrete sampling, binning and finite light curve duration cause the mean of the time-lag estimates to have a smaller magnitude than their intrinsic values. Smoothing (I.e. binning over consecutive frequencies) of the cross-periodogram can add extra bias at low frequencies. The use of light curves with low signal-to-noise ratio reduces the intrinsic coherence, and can introduce a bias to the sample coherence, time-lag estimates, and their predicted error. Conclusions: Our results have direct implications for X-ray time-lag studies in AGN, but can also be applied to similar studies in other research fields. We find that: a) time-lags should be estimated at frequencies lower than ≈ 1/2 the Nyquist frequency to minimise the effects of discrete binning of the

  5. The prion protein has DNA strand transfer properties similar to retroviral nucleocapsid protein.

    PubMed

    Gabus, C; Auxilien, S; Péchoux, C; Dormont, D; Swietnicki, W; Morillas, M; Surewicz, W; Nandi, P; Darlix, J L

    2001-04-06

    The transmissible spongiform encephalopathies are fatal neurodegenerative diseases that are associated with the accumulation of a protease-resistant form of the cellular prion protein (PrP). Although PrP is highly conserved and widely expressed in vertebrates, its function remains a matter of speculation. Indeed PrP null mice develop normally and are healthy. Recent results show that PrP binds to nucleic acids in vitro and is found associated with retroviral particles. Furthermore, in mice the scrapie infectious process appears to be accelerated by MuLV replication. These observations prompted us to further investigate the interaction between PrP and nucleic acids, and compare it with that of the retroviral nucleocapsid protein (NC). As the major nucleic acid-binding protein of the retroviral particle, NC protein is tightly associated with the genomic RNA in the virion nucleocapsid, where it chaperones proviral DNA synthesis by reverse transcriptase. Our results show that the human prion protein (huPrP) functionally resembles NCp7 of HIV-1. Both proteins form large nucleoprotein complexes upon binding to DNA. They accelerate the hybridization of complementary DNA strands and chaperone viral DNA synthesis during the minus and plus DNA strand transfers necessary to generate the long terminal repeats. The DNA-binding and strand transfer properties of huPrP appear to map to the N-terminal fragment comprising residues 23 to 144, whereas the C-terminal domain is inactive. These findings suggest that PrP could be involved in nucleic acid metabolism in vivo. Copyright 2001 Academic Press.

  6. 46 CFR 36.30-1 - Lagged tanks-TB/ALL.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 1 2010-10-01 2010-10-01 false Lagged tanks-TB/ALL. 36.30-1 Section 36.30-1 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY TANK VESSELS ELEVATED TEMPERATURE CARGOES Periodic Inspections § 36.30-1 Lagged tanks—TB/ALL. (a) Lagged tanks shall have part of the lagging removed on the...

  7. 46 CFR 36.30-1 - Lagged tanks-TB/ALL.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 1 2012-10-01 2012-10-01 false Lagged tanks-TB/ALL. 36.30-1 Section 36.30-1 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY TANK VESSELS ELEVATED TEMPERATURE CARGOES Periodic Inspections § 36.30-1 Lagged tanks—TB/ALL. (a) Lagged tanks shall have part of the lagging removed on the...

  8. 46 CFR 36.30-1 - Lagged tanks-TB/ALL.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 1 2014-10-01 2014-10-01 false Lagged tanks-TB/ALL. 36.30-1 Section 36.30-1 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY TANK VESSELS ELEVATED TEMPERATURE CARGOES Periodic Inspections § 36.30-1 Lagged tanks—TB/ALL. (a) Lagged tanks shall have part of the lagging removed on the...

  9. 46 CFR 36.30-1 - Lagged tanks-TB/ALL.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 1 2011-10-01 2011-10-01 false Lagged tanks-TB/ALL. 36.30-1 Section 36.30-1 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY TANK VESSELS ELEVATED TEMPERATURE CARGOES Periodic Inspections § 36.30-1 Lagged tanks—TB/ALL. (a) Lagged tanks shall have part of the lagging removed on the...

  10. 46 CFR 36.30-1 - Lagged tanks-TB/ALL.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 1 2013-10-01 2013-10-01 false Lagged tanks-TB/ALL. 36.30-1 Section 36.30-1 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY TANK VESSELS ELEVATED TEMPERATURE CARGOES Periodic Inspections § 36.30-1 Lagged tanks—TB/ALL. (a) Lagged tanks shall have part of the lagging removed on the...

  11. Comparing lagged linear correlation, lagged regression, Granger causality, and vector autoregression for uncovering associations in EHR data.

    PubMed

    Levine, Matthew E; Albers, David J; Hripcsak, George

    2016-01-01

    Time series analysis methods have been shown to reveal clinical and biological associations in data collected in the electronic health record. We wish to develop reliable high-throughput methods for identifying adverse drug effects that are easy to implement and produce readily interpretable results. To move toward this goal, we used univariate and multivariate lagged regression models to investigate associations between twenty pairs of drug orders and laboratory measurements. Multivariate lagged regression models exhibited higher sensitivity and specificity than univariate lagged regression in the 20 examples, and incorporating autoregressive terms for labs and drugs produced more robust signals in cases of known associations among the 20 example pairings. Moreover, including inpatient admission terms in the model attenuated the signals for some cases of unlikely associations, demonstrating how multivariate lagged regression models' explicit handling of context-based variables can provide a simple way to probe for health-care processes that confound analyses of EHR data.

  12. Role of Escherichia coli dnaG function in coliphage M13 DNA synthesis

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

    Dasgupta, S.; Mitra, S.

    Examination of the role of Escherichia coli dnaG function in different stages of M13 phage DNA synthesis by ultracentrifugal analysis of intracellular phage DNA in a thermosensitive dnaG mutant shows that: (a) the formation of parental double-strand replicative-form DNA (rfDNA) from the infecting virus is independent of dnaG function; (b) the synthesis of progeny rfDNA requires dnaG product; (c) after a pool of rfDNA is made up, dnaG function is not required for the progeny single-strand DNA (ssDNA) synthesis. The ssDNAs produced under nonpermissive condition are mostly circular and biologically functional.

  13. Helical self-organization and hierarchical self-assembly of an oligoheterocyclic pyridine-pyridazine strand into extended supramolecular fibers.

    PubMed

    Cuccia, Louis A; Ruiz, Eliseo; Lehn, Jean-Marie; Homo, Jean-Claude; Schmutz, Marc

    2002-08-02

    The synthesis and characterization of an alternating pyridine-pyridazine strand comprising thirteen heterocycles are described. Spontaneous folding into a helical secondary structure is based on a general molecular self-organization process enforced by the conformational information encoded within the primary structure of the molecular strand itself. Conformational control based on heterocyclic "helicity codons" illustrates a strategy for designing folding properties into synthetic oligomers (foldamers). Strong intermolecular interactions of the highly ordered lock-washer subunits of compound 3 results in hierarchical supramolecular self-assembly into protofibrils and fibrils. Compound 3 also forms mechanically stable two-dimensional Langmuir-Blodgett and cast thin films.

  14. Codon usage in Chlamydia trachomatis is the result of strand-specific mutational biases and a complex pattern of selective forces

    PubMed Central

    Romero, Héctor; Zavala, Alejandro; Musto, Héctor

    2000-01-01

    The patterns of synonymous codon choices of the completely sequenced genome of the bacterium Chlamydia trachomatis were analysed. We found that the most important source of variation among the genes results from whether the sequence is located on the leading or lagging strand of replication, resulting in an over representation of G or C, respectively. This can be explained by different mutational biases associated to the different enzymes that replicate each strand. Next we found that most highly expressed sequences are located on the leading strand of replication. From this result, replicational-transcriptional selection can be invoked. Then, when the genes located on the leading strand are studied separately, the correspondence analysis detects a principal trend which discriminates between lowly and highly expressed sequences, the latter displaying a different codon usage pattern than the former, suggesting selection for translation, which is reinforced by the fact that Ks values between orthologous sequences from C.trachomatis and Chlamydia pneumoniae are much smaller in highly expressed genes. Finally, synonymous codon choices appear to be influenced by the hydropathy of each encoded protein and by the degree of amino acid conservation. Therefore, synonymous codon usage in C.trachomatis seems to be the result of a very complex balance among different factors, which rises the problem of whether the forces driving codon usage patterns among microorganisms are rather more complex than generally accepted. PMID:10773076

  15. Codon usage in Chlamydia trachomatis is the result of strand-specific mutational biases and a complex pattern of selective forces.

    PubMed

    Romero, H; Zavala, A; Musto, H

    2000-05-15

    The patterns of synonymous codon choices of the completely sequenced genome of the bacterium Chlamydia trachomatis were analysed. We found that the most important source of variation among the genes results from whether the sequence is located on the leading or lagging strand of replication, resulting in an over representation of G or C, respectively. This can be explained by different mutational biases associated to the different enzymes that replicate each strand. Next we found that most highly expressed sequences are located on the leading strand of replication. From this result, replicational-transcriptional selection can be invoked. Then, when the genes located on the leading strand are studied separately, the correspondence analysis detects a principal trend which discriminates between lowly and highly expressed sequences, the latter displaying a different codon usage pattern than the former, suggesting selection for translation, which is reinforced by the fact that Ks values between orthologous sequences from C. trachomatis and Chlamydia pneumoniae are much smaller in highly expressed genes. Finally, synonymous codon choices appear to be influenced by the hydropathy of each encoded protein and by the degree of amino acid conservation. Therefore, synonymous codon usage in C.trachomatis seems to be the result of a very complex balance among different factors, which rises the problem of whether the forces driving codon usage patterns among microorganisms are rather more complex than generally accepted.

  16. A Novel Leu92 Mutant of HIV-1 Reverse Transcriptase with a Selective Deficiency in Strand Transfer Causes a Loss of Viral Replication.

    PubMed

    Herzig, Eytan; Voronin, Nickolay; Kucherenko, Nataly; Hizi, Amnon

    2015-08-01

    The process of reverse transcription (RTN) in retroviruses is essential to the viral life cycle. This key process is catalyzed exclusively by the viral reverse transcriptase (RT) that copies the viral RNA into DNA by its DNA polymerase activity, while concomitantly removing the original RNA template by its RNase H activity. During RTN, the combination between DNA synthesis and RNA hydrolysis leads to strand transfers (or template switches) that are critical for the completion of RTN. The balance between these RT-driven activities was considered to be the sole reason for strand transfers. Nevertheless, we show here that a specific mutation in HIV-1 RT (L92P) that does not affect the DNA polymerase and RNase H activities abolishes strand transfer. There is also a good correlation between this complete loss of the RT's strand transfer to the loss of the DNA clamp activity of the RT, discovered recently by us. This finding indicates a mechanistic linkage between these two functions and that they are both direct and unique functions of the RT (apart from DNA synthesis and RNA degradation). Furthermore, when the RT's L92P mutant was introduced into an infectious HIV-1 clone, it lost viral replication, due to inefficient intracellular strand transfers during RTN, thus supporting the in vitro data. As far as we know, this is the first report on RT mutants that specifically and directly impair RT-associated strand transfers. Therefore, targeting residue Leu92 may be helpful in selectively blocking this RT activity and consequently HIV-1 infectivity and pathogenesis. Reverse transcription in retroviruses is essential for the viral life cycle. This multistep process is catalyzed by viral reverse transcriptase, which copies the viral RNA into DNA by its DNA polymerase activity (while concomitantly removing the RNA template by its RNase H activity). The combination and balance between synthesis and hydrolysis lead to strand transfers that are critical for reverse transcription

  17. Assembling of G-strands into novel tetra-molecular parallel G4-DNA nanostructures using avidin-biotin recognition.

    PubMed

    Borovok, Natalia; Iram, Natalie; Zikich, Dragoslav; Ghabboun, Jamal; Livshits, Gideon I; Porath, Danny; Kotlyar, Alexander B

    2008-09-01

    We describe a method for the preparation of novel long (hundreds of nanometers), uniform, inter-molecular G4-DNA molecules composed of four parallel G-strands. The only long continuous G4-DNA reported so far are intra-molecular structures made of a single G-strand. To enable a tetra-molecular assembly of the G-strands we developed a novel approach based on avidin-biotin biological recognition. The steps of the G4-DNA production include: (i) Enzymatic synthesis of long poly(dG)-poly(dC) molecules with biotinylated poly(dG)-strand; (ii) Formation of a complex between avidin-tetramer and four biotinylated poly(dG)-poly(dC) molecules; (iii) Separation of the poly(dC) strands from the poly(dG)-strands, which are connected to the avidin; (iv) Assembly of the four G-strands attached to the avidin into tetra-molecular G4-DNA. The average contour length of the formed structures, as measured by AFM, is equal to that of the initial poly(dG)-poly(dC) molecules, suggesting a tetra-molecular mechanism of the G-strands assembly. The height of tetra-molecular G4-nanostructures is larger than that of mono-molecular G4-DNA molecules having similar contour length. The CD spectra of the tetra- and mono-molecular G4-DNA are markedly different, suggesting different structural organization of these two types of molecules. The tetra-molecular G4-DNA nanostructures showed clear electrical polarizability. This suggests that they may be useful for molecular electronics.

  18. LAG3 Expression in Active Mycobacterium tuberculosis Infections

    PubMed Central

    Phillips, Bonnie L.; Mehra, Smriti; Ahsan, Muhammad H.; Selman, Moises; Khader, Shabaana A.; Kaushal, Deepak

    2016-01-01

    Mycobacterium tuberculosis (MTB) is a highly successful pathogen because of its ability to persist in human lungs for long periods of time. MTB modulates several aspects of the host immune response. Lymphocyte-activation gene 3 (LAG3) is a protein with a high affinity for the CD4 receptor and is expressed mainly by regulatory T cells with immunomodulatory functions. To understand the function of LAG3 during MTB infection, a nonhuman primate model of tuberculosis, which recapitulates key aspects of natural human infection in rhesus macaques (Macaca mulatta), was used. We show that the expression of LAG3 is highly induced in the lungs and particularly in the granulomatous lesions of macaques experimentally infected with MTB. Furthermore, we show that LAG3 expression is not induced in the lungs and lung granulomas of animals exhibiting latent tuberculosis infection. However, simian immunodeficiency virus–induced reactivation of latent tuberculosis infection results in an increased expression of LAG3 in the lungs. This response is not observed in nonhuman primates infected with non-MTB bacterial pathogens, nor with simian immunodeficiency virus alone. Our data show that LAG3 was expressed primarily on CD4+ T cells, presumably by regulatory T cells but also by natural killer cells. The expression of LAG3 coincides with high bacterial burdens and changes in the host type 1 helper T-cell response. PMID:25549835

  19. Current-voltage characteristics of double stranded versus single stranded DNA molecules

    NASA Astrophysics Data System (ADS)

    Hartzell, B.; Chen, Hong; Heremans, J. J.; McCord, B.; Soghomonian, V.

    2004-03-01

    Investigation of DNA conductivity has focused on the native, duplex structure, with controversial results. Here, we present the influence of the double-helical structure on charge transport through lambda DNA molecules. The current-voltage (I-V) characteristics of both disulfide-labeled double stranded DNA (dsDNA) and disulfide-labeled single stranded DNA (ssDNA) were measured. The ssDNA was formed from the dsDNA using two different methods for comparison purposes: a thermal/chemical denaturation and enzymatic digestion utilizing lambda exonuclease. Resulting I-V characteristics of both the double stranded and single stranded samples were close-to-linear when measured at room temperature. However, the ssDNA samples consistently gave conductivity values about two orders of magnitude smaller in amplitude. Our results suggest an integral relationship between the native structure of DNA with its stacked base pairs and the molecule's ability to support charge transport.(NSF NIRT 0103034)

  20. Long-Lag, Wide-pulse Gamma-Ray Bursts

    NASA Technical Reports Server (NTRS)

    Norris, J. P.; Bonnell, J. T.; Kazanas, D.; Scargie, J. D.; Hakkila, J.; Giblin, T. W.

    2005-01-01

    The best available probe of the early phase of gamma-ray burst (GRB) jet attributes is the prompt gamma-ray emission, in which several intrinsic and extrinsic variables determine observed GRB pulse evolution, including at least: jet opening angle, profiles of Lorentz factor and matter/field density, distance of emission region from central source, and viewing angle. Bright, usually complex bursts have many narrow pulses that are difficult to model due to overlap. However, the relatively simple, long spectral lag, wide-pulse bursts may have simpler physics and are easier to model. We have analyzed the temporal and spectral behavior of wide pulses in 24 long-lag bursts from the BATSE sample, using a pulse model with two shape parameters - width and asymmetry - and the Band spectral model with three shape parameters. We find that pulses in long-lag bursts are distinguished both temporally and spectrally from those in bright bursts: the pulses in long spectral lag bursts are few in number, and approximately 100 times wider (10s of seconds), have systemtically lower peaks in nu*F(nu), harder low-energy spectra and softer high-energy spectra. These five pulse descriptors are essentially uncorrelated for our long-lag sample, suggesting that at least approximately 5 parameters are needed to model burst temporal and spectral behavior, roughly commensurate with the theoretical phase space. However, we do find that pulse width is strongly correlated with spectral lag; hence these two parameters may be viewed as mutual surrogates. The prevalence of long-lag bursts near the BATSE trigger threshold, their predominantly low nu*F(nu) spectral peaks, and relatively steep upper power-law spectral indices indicate that Swiift will detect many such bursts.

  1. Behavioral Processes in Long-Lag Intervention Studies.

    PubMed

    Miller, Dale T; Dannals, Jennifer E; Zlatev, Julian J

    2017-05-01

    We argue that psychologists who conduct experiments with long lags between the manipulation and the outcome measure should pay more attention to behavioral processes that intervene between the manipulation and the outcome measure. Neglect of such processes, we contend, stems from psychology's long tradition of short-lag lab experiments where there is little scope for intervening behavioral processes. Studying process in the lab invariably involves studying psychological processes, but in long-lag field experiments it is important to study causally relevant behavioral processes as well as psychological ones. To illustrate the roles that behavioral processes can play in long-lag experiments we examine field experiments motivated by three policy-relevant goals: prejudice reduction, health promotion, and educational achievement. In each of the experiments discussed we identify various behavioral pathways through which the manipulated psychological state could have produced the observed outcome. We argue that if psychologists conducting long-lag interventions posited a theory of change that linked manipulated psychological states to outcomes via behavioral pathways, the result would be richer theory and more practically useful research. Movement in this direction would also permit more opportunities for productive collaborations between psychologists and other social scientists interested in similar social problems.

  2. X-ray time lags in PG 1211+143

    NASA Astrophysics Data System (ADS)

    Lobban, A. P.; Vaughan, S.; Pounds, K.; Reeves, J. N.

    2018-05-01

    We investigate the X-ray time lags of a recent ˜630 ks XMM-Newton observation of PG 1211+143. We find well-correlated variations across the XMM-Newton EPIC bandpass, with the first detection of a hard lag in this source with a mean time delay of up to ˜3 ks at the lowest frequencies. We find that the energy-dependence of the low-frequency hard lag scales approximately linearly with log(E) when averaged over all orbits, consistent with the propagating fluctuations model. However, we find that the low-frequency lag behaviour becomes more complex on time-scales longer than a single orbit, suggestive of additional modes of variability. We also detect a high-frequency soft lag at ˜10-4 Hz with the magnitude of the delay peaking at ≲ 0.8 ks, consistent with previous observations, which we discuss in terms of small-scale reverberation.

  3. Nanocrystal synthesis in microfluidic reactors: where next?

    PubMed

    Phillips, Thomas W; Lignos, Ioannis G; Maceiczyk, Richard M; deMello, Andrew J; deMello, John C

    2014-09-07

    The past decade has seen a steady rise in the use of microfluidic reactors for nanocrystal synthesis, with numerous studies reporting improved reaction control relative to conventional batch chemistry. However, flow synthesis procedures continue to lag behind batch methods in terms of chemical sophistication and the range of accessible materials, with most reports having involved simple one- or two-step chemical procedures directly adapted from proven batch protocols. Here we examine the current status of microscale methods for nanocrystal synthesis, and consider what role microreactors might ultimately play in laboratory-scale research and industrial production.

  4. Cdc45 (cell division cycle protein 45) guards the gate of the Eukaryote Replisome helicase stabilizing leading strand engagement

    PubMed Central

    Petojevic, Tatjana; Pesavento, James J.; Costa, Alessandro; Liang, Jingdan; Wang, Zhijun; Berger, James M.; Botchan, Michael R.

    2015-01-01

    DNA replication licensing is now understood to be the pathway that leads to the assembly of double hexamers of minichromosome maintenance (Mcm2–7) at origin sites. Cell division control protein 45 (Cdc45) and GINS proteins activate the latent Mcm2–7 helicase by inducing allosteric changes through binding, forming a Cdc45/Mcm2-7/GINS (CMG) complex that is competent to unwind duplex DNA. The CMG has an active gate between subunits Mcm2 and Mcm5 that opens and closes in response to nucleotide binding. The consequences of inappropriate Mcm2/5 gate actuation and the role of a side channel formed between GINS/Cdc45 and the outer edge of the Mcm2–7 ring for unwinding have remained unexplored. Here we uncover a novel function for Cdc45. Cross-linking studies trace the path of the DNA with the CMG complex at a fork junction between duplex and single strands with the bound CMG in an open or closed gate conformation. In the closed state, the lagging strand does not pass through the side channel, but in the open state, the leading strand surprisingly interacts with Cdc45. Mutations in the recombination protein J fold of Cdc45 that ablate this interaction diminish helicase activity. These data indicate that Cdc45 serves as a shield to guard against occasional slippage of the leading strand from the core channel. PMID:25561522

  5. LAG3 expression in active Mycobacterium tuberculosis infections.

    PubMed

    Phillips, Bonnie L; Mehra, Smriti; Ahsan, Muhammad H; Selman, Moises; Khader, Shabaana A; Kaushal, Deepak

    2015-03-01

    Mycobacterium tuberculosis (MTB) is a highly successful pathogen because of its ability to persist in human lungs for long periods of time. MTB modulates several aspects of the host immune response. Lymphocyte-activation gene 3 (LAG3) is a protein with a high affinity for the CD4 receptor and is expressed mainly by regulatory T cells with immunomodulatory functions. To understand the function of LAG3 during MTB infection, a nonhuman primate model of tuberculosis, which recapitulates key aspects of natural human infection in rhesus macaques (Macaca mulatta), was used. We show that the expression of LAG3 is highly induced in the lungs and particularly in the granulomatous lesions of macaques experimentally infected with MTB. Furthermore, we show that LAG3 expression is not induced in the lungs and lung granulomas of animals exhibiting latent tuberculosis infection. However, simian immunodeficiency virus-induced reactivation of latent tuberculosis infection results in an increased expression of LAG3 in the lungs. This response is not observed in nonhuman primates infected with non-MTB bacterial pathogens, nor with simian immunodeficiency virus alone. Our data show that LAG3 was expressed primarily on CD4(+) T cells, presumably by regulatory T cells but also by natural killer cells. The expression of LAG3 coincides with high bacterial burdens and changes in the host type 1 helper T-cell response. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  6. Spacing and lag effects in free recall of pure lists.

    PubMed

    Kahana, Michael J; Howard, Marc W

    2005-02-01

    Repeating list items leads to better recall when the repetitions are separated by several unique items than when they are presented successively; the spacing effect refers to improved recall for spaced versus successive repetition (lag > 0 vs. lag = 0); the lag effect refers to improved recall for long lags versus short lags. Previous demonstrations of the lag effect have utilized lists containing a mixture of items with varying degrees of spacing. Because differential rehearsal of items in mixed lists may exaggerate any effects of spacing, it is important to demonstrate these effects in pure lists. As in Toppino and Schneider (1999), we found an overall advantage for recall of spaced lists. We further report the first demonstration of a lag effect in pure lists, with significantly better recall for lists with widely spaced repetitions than for those with moderately spaced repetitions.

  7. Inhibition of adenovirus DNA synthesis in vitro by sera from patients with systemic lupus erythematosus

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

    Horwitz, M.S.; Friefeld, B.R.; Keiser, H.D.

    1982-12-01

    Sera containing antinuclear antibodies from patients with systemic lupus erythematosus (SLE) and related disorders were tested for their effect on the synthesis of adenovirus (Ad) DNA in an in vitro replication system. After being heated at 60/sup 0/C for 1 h, some sera from patients with SLE inhibited Ad DNA synthesis by 60 to 100%. Antibodies to double-stranded DNA were present in 15 of the 16 inhibitory sera, and inhibitory activity copurified with anti-double-stranded DNA in the immunoglobulin G fraction. These SLE sera did not inhibit the DNA polymerases ..cap alpha.., BETA, ..gamma.. and had no antibody to the 72,000-daltonmore » DNA-binding protein necessary for Ad DNA synthesis. The presence of antibodies to single-stranded DNA and a variety of saline-extractable antigens (Sm, Ha, nRNP, and rRNP) did not correlate with SLE serum inhibitory activity. Methods previously developed for studying the individual steps in Ad DNA replication were used to determine the site of inhibition by the SLE sera that contained antibody to double-stranded DNA. Concentrations of the SLE inhibitor that decreased the elongation of Ad DNA by greater than 85% had no effect on either the initiation of Ad DNA synthesis or the polymerization of the first 26 deoxyribonucleotides.« less

  8. Five-Strand versus Four-Strand Hamstring Tendon Graft Technique for Anterior Cruciate Ligament Reconstruction: A Biomechanical Comparison.

    PubMed

    Vaillant, Eric R; Parks, Brent G; Camire, Lyn M; Hinton, Richard Y

    2017-11-01

    The aim of this article is to compare diameter and stiffness, displacement, and strain in a five-strand versus four-strand hamstring graft for anterior cruciate ligament reconstruction. Eight matched pairs of lower extremities underwent four-strand or five-strand hamstring graft reconstruction. Diameter was significantly higher in the five-strand versus the four-strand construct ( p  = 0.002). No significant difference was found between the groups in construct displacement or stiffness. Significantly higher strain was observed in the inner limb versus the outer limb in the four-strand construct ( p  = 0.001) and in the inner limb versus the fifth limb in the 5-strand construct ( p  = 0.004). A fifth limb added to a four-strand hamstring graft significantly increased graft diameter but did not significantly change stiffness or displacement, suggesting that attachment of additional graft material via suture did not provide for full incorporation of the added limb into the graft at time zero. The inner limb in both constructs absorbed significantly greater load than did other limbs. The use of suture to attach additional material to a four-strand hamstring graft may not contribute to improved biomechanical qualities of the graft at time zero. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

  9. Excess single-stranded DNA inhibits meiotic double-strand break repair.

    PubMed

    Johnson, Rebecca; Borde, Valérie; Neale, Matthew J; Bishop-Bailey, Anna; North, Matthew; Harris, Sheila; Nicolas, Alain; Goldman, Alastair S H

    2007-11-01

    During meiosis, self-inflicted DNA double-strand breaks (DSBs) are created by the protein Spo11 and repaired by homologous recombination leading to gene conversions and crossovers. Crossover formation is vital for the segregation of homologous chromosomes during the first meiotic division and requires the RecA orthologue, Dmc1. We analyzed repair during meiosis of site-specific DSBs created by another nuclease, VMA1-derived endonuclease (VDE), in cells lacking Dmc1 strand-exchange protein. Turnover and resection of the VDE-DSBs was assessed in two different reporter cassettes that can repair using flanking direct repeat sequences, thereby obviating the need for a Dmc1-dependent DNA strand invasion step. Access of the single-strand binding complex replication protein A, which is normally used in all modes of DSB repair, was checked in chromatin immunoprecipitation experiments, using antibody against Rfa1. Repair of the VDE-DSBs was severely inhibited in dmc1Delta cells, a defect that was associated with a reduction in the long tract resection required to initiate single-strand annealing between the flanking repeat sequences. Mutants that either reduce Spo11-DSB formation or abolish resection at Spo11-DSBs rescued the repair block. We also found that a replication protein A component, Rfa1, does not accumulate to expected levels at unrepaired single-stranded DNA (ssDNA) in dmc1Delta cells. The requirement of Dmc1 for VDE-DSB repair using flanking repeats appears to be caused by the accumulation of large quantities of ssDNA that accumulate at Spo11-DSBs when Dmc1 is absent. We propose that these resected DSBs sequester both resection machinery and ssDNA binding proteins, which in wild-type cells would normally be recycled as Spo11-DSBs repair. The implication is that repair proteins are in limited supply, and this could reflect an underlying mechanism for regulating DSB repair in wild-type cells, providing protection from potentially harmful effects of overabundant

  10. Excess Single-Stranded DNA Inhibits Meiotic Double-Strand Break Repair

    PubMed Central

    Bishop-Bailey, Anna; North, Matthew; Harris, Sheila; Nicolas, Alain; Goldman, Alastair S. H

    2007-01-01

    During meiosis, self-inflicted DNA double-strand breaks (DSBs) are created by the protein Spo11 and repaired by homologous recombination leading to gene conversions and crossovers. Crossover formation is vital for the segregation of homologous chromosomes during the first meiotic division and requires the RecA orthologue, Dmc1.We analyzed repair during meiosis of site-specific DSBs created by another nuclease, VMA1-derived endonuclease (VDE), in cells lacking Dmc1 strand-exchange protein. Turnover and resection of the VDE-DSBs was assessed in two different reporter cassettes that can repair using flanking direct repeat sequences, thereby obviating the need for a Dmc1-dependent DNA strand invasion step. Access of the single-strand binding complex replication protein A, which is normally used in all modes of DSB repair, was checked in chromatin immunoprecipitation experiments, using antibody against Rfa1. Repair of the VDE-DSBs was severely inhibited in dmc1Δ cells, a defect that was associated with a reduction in the long tract resection required to initiate single-strand annealing between the flanking repeat sequences. Mutants that either reduce Spo11-DSB formation or abolish resection at Spo11-DSBs rescued the repair block. We also found that a replication protein A component, Rfa1, does not accumulate to expected levels at unrepaired single-stranded DNA (ssDNA) in dmc1Δ cells. The requirement of Dmc1 for VDE-DSB repair using flanking repeats appears to be caused by the accumulation of large quantities of ssDNA that accumulate at Spo11-DSBs when Dmc1 is absent. We propose that these resected DSBs sequester both resection machinery and ssDNA binding proteins, which in wild-type cells would normally be recycled as Spo11-DSBs repair. The implication is that repair proteins are in limited supply, and this could reflect an underlying mechanism for regulating DSB repair in wild-type cells, providing protection from potentially harmful effects of overabundant repair

  11. Exposure-lag-response in Longitudinal Studies: Application of Distributed Lag Non-linear Models in an Occupational Cohort.

    PubMed

    Neophytou, Andreas M; Picciotto, Sally; Brown, Daniel M; Gallagher, Lisa E; Checkoway, Harvey; Eisen, Ellen A; Costello, Sadie

    2018-02-13

    Prolonged exposures can have complex relationships with health outcomes, as timing, duration, and intensity of exposure are all potentially relevant. Summary measures such as cumulative exposure or average intensity of exposure may not fully capture these relationships. We applied penalized and unpenalized distributed lag non-linear models (DLNMs) with flexible exposure-response and lag-response functions in order to examine the association between crystalline silica exposure and mortality from lung cancer and non-malignant respiratory disease in a cohort study of 2,342 California diatomaceous earth workers, followed 1942-2011. We also assessed associations using simple measures of cumulative exposure assuming linear exposure-response and constant lag-response. Measures of association from DLNMs were generally higher than from simpler models. Rate ratios from penalized DLNMs corresponding to average daily exposures of 0.4 mg/m3 during lag years 31-50 prior to the age of observed cases were 1.47 (95% confidence interval (CI) 0.92, 2.35) for lung cancer and 1.80 (95% CI: 1.14, 2.85) for non-malignant respiratory disease. Rate ratios from the simpler models for the same exposure scenario were 1.15 (95% CI: 0.89-1.48) and 1.23 (95% CI: 1.03-1.46) respectively. Longitudinal cohort studies of prolonged exposures and chronic health outcomes should explore methods allowing for flexibility and non-linearities in the exposure-lag-response. © The Author(s) 2018. Published by Oxford University Press on behalf of the Johns Hopkins Bloomberg School of Public Health.

  12. Can a double stranded DNA be unzipped by pulling a single strand?: phases of adsorbed DNA.

    PubMed

    Kapri, Rajeev

    2009-04-14

    We study the unzipping of a double stranded DNA (dsDNA) by applying an external force on a single strand while leaving the other strand free. We find that the dsDNA can be unzipped to two single strands if the external force exceeds a critical value. We obtain the phase diagram, which is found to be different from the phase diagram of unzipping by pulling both the strands in opposite directions. In the presence of an attractive surface near DNA, the phase diagram gets modified drastically and shows richer surprises including a critical end point and a triple point.

  13. To lag or not to lag? How to compare indices of stock markets that operate on different times

    NASA Astrophysics Data System (ADS)

    Sandoval, Leonidas

    2014-06-01

    Financial markets worldwide do not have the same working hours. As a consequence, the study of correlation or causality between financial market indices becomes dependent on whether we should use all indices on the same day or lagged indices in computations of correlation matrices. The answer this article proposes is that we should consider both, by representing original and lagged indices in the same network. We then obtain a better understanding of how indices that operate on different hours relate to each other. We use a diverse range of 79 stock market indices from around the world and study their correlation structure, the eigenvalues and eigenvectors of their correlations under different time periods and volatility, as well as the differences between the working hours of the stock exchanges in order to analyze the possible time zone effects and suggest ways to remove them. We also analyze the enlarged correlation matrix obtained from original and lagged indices and examine a network structure derived from it, thus showing connections between lagged and original indices that could not be well represented before.

  14. SAW1 is required for SDSA double-strand break repair in S. cerevisiae.

    PubMed

    Diamante, Graciel; Phan, Claire; Celis, Angie S; Krueger, Jonas; Kelson, Eric P; Fischhaber, Paula L

    2014-03-14

    SAW1, coding for Saw1, is required for single-strand annealing (SSA) DNA double-strand break (DSB) repair in Saccharomycescerevisiae. Saw1 physically associates with Rad1 and Rad52 and recruits the Rad1-Rad10 endonuclease. Herein we show by fluorescence microscopy that SAW1 is similarly required for recruitment of Rad10 to sites of Synthesis-Dependent Strand Annealing (SDSA) and associates with sites of SDSA repair in a manner temporally overlapped with Rad10. The magnitude of induction of colocalized Saw1-CFP/Rad10-YFP/DSB-RFP foci in SDSA is more dramatic in S and G2 phase cells than in M phase, consistent with the known mechanism of SDSA. We observed a substantial fraction of foci in which Rad10 was localized to the repair site without Saw1, but few DSB sites that contained Saw1 without Rad10. Together these data are consistent with a model in which Saw1 recruits Rad1-Rad10 to SDSA sites, possibly even binding as a protein-protein complex, but departs the repair site in advance of Rad1-Rad10. Copyright © 2014 Elsevier Inc. All rights reserved.

  15. Why Do Rereading Lag Effects Depend on Test Delay?

    ERIC Educational Resources Information Center

    Rawson, Katherine A.

    2012-01-01

    In previous research, rereading after a long lag versus a short lag led to greater performance on delayed tests but not on immediate tests. The current study tested two accounts of why the effects of rereading lag depend on test delay. The "levels of representation" ("LOR") "hypothesis" states that the effects reflect…

  16. Implications of Lag-Luminosity Relationship for Unified GRB Paradigms

    NASA Technical Reports Server (NTRS)

    Norris, J. P.; White, Nicholas E. (Technical Monitor)

    2002-01-01

    Spectral lags (tau(sub lag)) are deduced for 1437 long (T(sub 90) greater than 2 s) BATSE gamma-ray bursts (GRBs) with peak flux F(sub p) greater than 0.25 photons cm(sup -2)/s, near to the BATSE trigger threshold. The lags are modeled to approximate the observed distribution in the F(sub p)-T(sub lag) plane, realizing a noise-free representation. Assuming a two-branch lag-luminosity relationship, the lags are self-consistently corrected for cosmological effects to yield distributions in luminosity, distance, and redshift. The results have several consequences for GRB populations and for unified gamma-ray/afterglow scenarios which would account for afterglow break times and gamma-ray spectral evolution in terms of jet opening angle, viewing angle, or a profiled jet with variable Lorentz factor: A component of the burst sample is identified - those with few, wide pulses, lags of a few tenths to several seconds, and soft spectra - whose Log[N]-Log[F(sub p)] distribution approximates a -3/2 power-law, suggesting homogeneity and thus relatively nearby sources. The proportion of these long-lag bursts increases from negligible among bright BATSE bursts to approx. 50% at trigger threshold. Bursts with very long lags, approx. 1-2 less than tau(sub lag) (S) less than 10, show a tendency to concentrate near the Supergalactic Plane with a quadrupole moment of approx. -0.10 +/- 0.04. GRB 980425 (SN 1998bw) is a member of this subsample of approx. 90 bursts with estimated distances less than 100 Mpc. The frequency of the observed ultra-low luminosity bursts is approx. 1/4 that of SNe Ib/c within the same volume. If truly nearby, the core-collapse events associated with these GRBs might produce gravitational radiation detectable by LIGO-II. Such nearby bursts might also help explain flattening of the cosmic ray spectrum at ultra-high energies, as observed by AGASA.

  17. Classifying Saturn's F Ring Strands

    NASA Astrophysics Data System (ADS)

    Albers, Nicole; Sremcevic, M.; Esposito, L. W.; Colwell, J. E.

    2009-09-01

    The Cassini Ultraviolet Imaging Spectrograph (UVIS) High Speed Photometer (HSP) has recorded more than 113 stellar occultations by Saturn's F ring providing measurements with ring plane resolutions of a few dozen meters and better. Inner and outer F ring strands have been seen throughout the Cassini mission where they revealed themselves as non-continuous, azimuthally and temporally highly variable structures. In the light of a more accurate orbit description of the F ring core we find evidence for a ring that becomes dynamically more active as the system approaches anti-apse alignment with Prometheus. This is consistent with the observed increased strand activity. A recent strand that morphologically resembles the core is the strongest seen to date and points to the intricate relation between core and strands indicating the strands' violent creation. Using more than 150 identifications of various strands, we trace their kinematics and infer dynamical timescales and photometric properties. Implications for the dynamical evolution of the F ring will be discussed. This research was supported by the Cassini Project.

  18. Enzymatic Synthesis of Self-assembled Dicer Substrate RNA Nanostructures for Programmable Gene Silencing.

    PubMed

    Jang, Bora; Kim, Boyoung; Kim, Hyunsook; Kwon, Hyokyoung; Kim, Minjeong; Seo, Yunmi; Colas, Marion; Jeong, Hansaem; Jeong, Eun Hye; Lee, Kyuri; Lee, Hyukjin

    2018-06-08

    Enzymatic synthesis of RNA nanostructures is achieved by isothermal rolling circle transcription (RCT). Each arm of RNA nanostructures provides a functional role of Dicer substrate RNA inducing sequence specific RNA interference (RNAi). Three different RNAi sequences (GFP, RFP, and BFP) are incorporated within the three-arm junction RNA nanostructures (Y-RNA). The template and helper DNA strands are designed for the large-scale in vitro synthesis of RNA strands to prepare self-assembled Y-RNA. Interestingly, Dicer processing of Y-RNA is highly influenced by its physical structure and different gene silencing activity is achieved depending on its arm length and overhang. In addition, enzymatic synthesis allows the preparation of various Y-RNA structures using a single DNA template offering on demand regulation of multiple target genes.

  19. A critique of the cross-lagged panel model.

    PubMed

    Hamaker, Ellen L; Kuiper, Rebecca M; Grasman, Raoul P P P

    2015-03-01

    The cross-lagged panel model (CLPM) is believed by many to overcome the problems associated with the use of cross-lagged correlations as a way to study causal influences in longitudinal panel data. The current article, however, shows that if stability of constructs is to some extent of a trait-like, time-invariant nature, the autoregressive relationships of the CLPM fail to adequately account for this. As a result, the lagged parameters that are obtained with the CLPM do not represent the actual within-person relationships over time, and this may lead to erroneous conclusions regarding the presence, predominance, and sign of causal influences. In this article we present an alternative model that separates the within-person process from stable between-person differences through the inclusion of random intercepts, and we discuss how this model is related to existing structural equation models that include cross-lagged relationships. We derive the analytical relationship between the cross-lagged parameters from the CLPM and the alternative model, and use simulations to demonstrate the spurious results that may arise when using the CLPM to analyze data that include stable, trait-like individual differences. We also present a modeling strategy to avoid this pitfall and illustrate this using an empirical data set. The implications for both existing and future cross-lagged panel research are discussed. (c) 2015 APA, all rights reserved).

  20. Quality control mechanisms exclude incorrect polymerases from the eukaryotic replication fork

    PubMed Central

    Schauer, Grant D.; O’Donnell, Michael E.

    2017-01-01

    The eukaryotic genome is primarily replicated by two DNA polymerases, Pol ε and Pol δ, that function on the leading and lagging strands, respectively. Previous studies have established recruitment mechanisms whereby Cdc45-Mcm2-7-GINS (CMG) helicase binds Pol ε and tethers it to the leading strand, and PCNA (proliferating cell nuclear antigen) binds tightly to Pol δ and recruits it to the lagging strand. The current report identifies quality control mechanisms that exclude the improper polymerase from a particular strand. We find that the replication factor C (RFC) clamp loader specifically inhibits Pol ε on the lagging strand, and CMG protects Pol ε against RFC inhibition on the leading strand. Previous studies show that Pol δ is slow and distributive with CMG on the leading strand. However, Saccharomyces cerevisiae Pol δ–PCNA is a rapid and processive enzyme, suggesting that CMG may bind and alter Pol δ activity or position it on the lagging strand. Measurements of polymerase binding to CMG demonstrate Pol ε binds CMG with a Kd value of 12 nM, but Pol δ binding CMG is undetectable. Pol δ, like bacterial replicases, undergoes collision release upon completing replication, and we propose Pol δ–PCNA collides with the slower CMG, and in the absence of a stabilizing Pol δ–CMG interaction, the collision release process is triggered, ejecting Pol δ on the leading strand. Hence, by eviction of incorrect polymerases at the fork, the clamp machinery directs quality control on the lagging strand and CMG enforces quality control on the leading strand. PMID:28069954

  1. Do subjective symptoms predict our perception of jet-lag?

    PubMed

    Waterhouse, J; Edwards, B; Nevill, A; Atkinson, G; Reilly, T; Davies, P; Godfrey, R

    2000-10-01

    A total of 39 subjects were studied after a flight from the UK to either Sydney or Brisbane (10 time-zones to the east). Subjects varied widely in their age, their athletic ability, whether or not they were taking melatonin, and in their objectives when in Australia. For the first 6 days after arrival, subjects scored their jet-lag five times per day and other subjective variables up to five times per day, using visual analogue scales. For jet-lag, the scale was labelled 0 = no jet-lag to 10 = very bad jet-lag; the extremes of the other scales were labelled - 5 and + 5, indicating marked changes compared with normal, and the centrepoint was labelled 0 indicating 'normal'. Mean daily values for jet-lag and fatigue were initially high (+ 3.65 +/- 0.35 and + 1.55 +/- 0.22 on day 1, respectively) and fell progressively on subsequent days, but were still raised significantly (p < 0.05) on day 5 (fatigue) or day 6 (jet-lag). In addition, times of waking were earlier on all days. By contrast, falls in concentration and motivation, and rises in irritability and nocturnal wakings, had recovered by day 4 or earlier, and bowel activity was less frequent, with harder stools, on days 1 and 2 only. Also, on day 1, there was a decrease in the ease of getting to sleep (- 1.33 +/- 0.55), but this changed to an increase from day 2 onwards (for example, + 0.75 +/- 0.25 on day 6). Stepwise regression analysis was used to investigate predictors of jet-lag. The severity of jet-lag at all the times that were measured was strongly predicted by fatigue ratings made at the same time. Its severity at 08:00 h was predicted by an earlier time of waking, by feeling less alert 30 min after waking and, marginally, by the number of waking episodes. Jet-lag at 12:00 and 16:00 h was strongly predicted by a fall of concentration at these times; jet-lag at mealtimes (12:00, 16:00 and 20:00 h) was predicted by the amount of feeling bloated. Such results complicate an exact interpretation that can be

  2. Subnuclear systems for synthesis of simian virus 40 DNA in vitro.

    PubMed Central

    Edenberg, H J; Waqar, M A; Huberman, J A

    1976-01-01

    We have developed two subnuclear systems for synthesis of DNA of simian virus 40 in vitro. We prepare chromatin from infected cells by the method of Hancock [(1974) J. Mol. Biol. 86, 649-663]; these "chromatin bodies" can be disrupted and large debris can be pelleted, leaving a supernatant ("soluble system"). Both chromatin bodies and the soluble system incorporate deoxyribonucleoside triphosphates into nucleoprotein complexes that contain simian virus 40 DNA. The DNA labeled in short pulses sediments in neutral sucrose gradients slightly faster than mature simian virus 40 DNA, as expected for replicating intermediate. When rebanded in alkaline sucrose gradients, about half of the radioactivity is found in short strands (200-300 nucleotides) and half in longer strands (up to full viral size). When these systems are supplemented with a cytoplasmic preparation from HeLa cells, synthesis is stimulated about 5-fold, and the short strands are converted into strands of up to full viral length as well as into covalently closed circles. These subnuclear DNA-replicating systems should be useful for biochemical fractionation and characterization of some of the proteins required for DNA replication. PMID:188037

  3. Bubbles in live-stranded dolphins.

    PubMed

    Dennison, S; Moore, M J; Fahlman, A; Moore, K; Sharp, S; Harry, C T; Hoppe, J; Niemeyer, M; Lentell, B; Wells, R S

    2012-04-07

    Bubbles in supersaturated tissues and blood occur in beaked whales stranded near sonar exercises, and post-mortem in dolphins bycaught at depth and then hauled to the surface. To evaluate live dolphins for bubbles, liver, kidneys, eyes and blubber-muscle interface of live-stranded and capture-release dolphins were scanned with B-mode ultrasound. Gas was identified in kidneys of 21 of 22 live-stranded dolphins and in the hepatic portal vasculature of 2 of 22. Nine then died or were euthanized and bubble presence corroborated by computer tomography and necropsy, 13 were released of which all but two did not re-strand. Bubbles were not detected in 20 live wild dolphins examined during health assessments in shallow water. Off-gassing of supersaturated blood and tissues was the most probable origin for the gas bubbles. In contrast to marine mammals repeatedly diving in the wild, stranded animals are unable to recompress by diving, and thus may retain bubbles. Since the majority of beached dolphins released did not re-strand it also suggests that minor bubble formation is tolerated and will not lead to clinically significant decompression sickness.

  4. Recent trends for drug lag in clinical development of oncology drugs in Japan: does the oncology drug lag still exist in Japan?

    PubMed

    Maeda, Hideki; Kurokawa, Tatsuo

    2015-12-01

    This study exhaustively and historically investigated the status of drug lag for oncology drugs approved in Japan. We comprehensively investigated oncology drugs approved in Japan between April 2001 and July 2014, using publicly available information. We also examined changes in the status of drug lag between Japan and the United States, as well as factors influencing drug lag. This study included 120 applications for approval of oncology drugs in Japan. The median difference over a 13-year period in the approval date between the United States and Japan was 875 days (29.2 months). This figure peaked in 2002, and showed a tendency to decline gradually each year thereafter. In 2014, the median approval lag was 281 days (9.4 months). Multiple regression analysis identified the following potential factors that reduce drug lag: "Japan's participation in global clinical trials"; "bridging strategies"; "designation of priority review in Japan"; and "molecularly targeted drugs". From 2001 to 2014, molecularly targeted drugs emerged as the predominant oncology drug, and the method of development has changed from full development in Japan or bridging strategy to global simultaneous development by Japan's taking part in global clinical trials. In line with these changes, the drug lag between the United States and Japan has significantly reduced to less than 1 year.

  5. On the phase lag of turbulent dissipation in rotating tidal flows

    NASA Astrophysics Data System (ADS)

    Zhang, Qianjiang; Wu, Jiaxue

    2018-03-01

    Field observations of rotating tidal flows in a shallow tidally swept sea reveal that a notable phase lag of both shear production and turbulent dissipation increases with height above the seafloor. These vertical delays of turbulent quantities are approximately equivalent in magnitude to that of squared mean shear. The shear production approximately equals turbulent dissipation over the phase-lag column, and thus a main mechanism of phase lag of dissipation is mean shear, rather than vertical diffusion of turbulent kinetic energy. By relating the phase lag of dissipation to that of the mean shear, a simple formulation with constant eddy viscosity is developed to describe the phase lag in rotating tidal flows. An analytical solution indicates that the phase lag increases linearly with height subjected to a combined effect of tidal frequency, Coriolis parameter and eddy viscosity. The vertical diffusion of momentum associated with eddy viscosity produces the phase lag of squared mean shear, and resultant delay of turbulent quantities. Its magnitude is inhibited by Earth's rotation. Furthermore, a theoretical formulation of the phase lag with a parabolic eddy viscosity profile can be constructed. A first-order approximation of this formulation is still a linear function of height, and its magnitude is approximately 0.8 times that with constant viscosity. Finally, the theoretical solutions of phase lag with realistic viscosity can be satisfactorily justified by realistic phase lags of dissipation.

  6. Lag Times and Peak Coefficients for Rural Watersheds in Kansas

    DOT National Transportation Integrated Search

    1999-10-01

    Lag time is an essential input to the most common synthetic unit-hydrograph models. The lag time for an ungaged stream must be estimated from the physical characteristics of the stream and its watershed. In this study, a lag-time formula for small ru...

  7. Mitigating shear lag in tall buildings

    NASA Astrophysics Data System (ADS)

    Gaur, Himanshu; Goliya, Ravindra K.

    2015-09-01

    As the height of building increases, effect of shear lag also becomes considerable in the design of high-rise buildings. In this paper, shear lag effect in tall buildings of heights, i.e., 120, 96, 72, 48 and 36 stories of which aspect ratio ranges from 3 to 10 is studied. Tube-in-tube structural system with façade bracing is used for designing the building of height 120 story. It is found that bracing system considerably reduces the shear lag effect and hence increases the building stiffness to withstand lateral loads. Different geometric patterns of bracing system are considered. The best effective geometric configuration of bracing system is concluded in this study. Lateral force, as wind load is applied on the buildings as it is the most dominating lateral force for such heights. Wind load is set as per Indian standard code of practice IS 875 Part-3. For analysis purpose SAP 2000 software program is used.

  8. The Broken Belt: Meteorite Concentrations on Stranded Ice

    NASA Technical Reports Server (NTRS)

    Harvey, R. P.

    2003-01-01

    Since the first Antarctic meteorite concentrations were discovered more than 25 years ago, many theories regarding the role of iceflow in the production of meteorite concentrations have been put forward, and most agree on the basic principles. These models suggest that as the East Antarctic icesheet flows toward the margins of the continent, meteorites randomly located within the volume of ice are transported toward the icesheet margin. Where mountains or subsurface obstructions block glacial flow, diversion of ice around or over an obstruction reduces horizontal ice movement rates adjacent to the barriers and creates a vertical (upward) component of movement. If local mechanisms for ice loss (ablation) exist at such sites, an equilibrium surface will develop according to the balance between ice supply and loss, and the cargo of meteorites is exhumed on a blue ice surface. The result is a conceptual conveyor belt bringing meteorite-bearing volumes of ice from the interior of the continent to stagnant or slowmoving surfaces where ice is then lost and a precious cargo is left as a lag deposit. Cassidy et al. provides an excellent overview of how this model has been adapted to several Antarctic stranding surfaces.

  9. Flavivirus RNA Synthesis in vitro

    PubMed Central

    Padmanabhan, Radhakrishnan; Takhampunya, Ratree; Teramoto, Tadahisa; Choi, Kyung H.

    2015-01-01

    Summary Establishment of in vitro systems to study mechanisms of RNA synthesis for positive strand RNA viruses have been very useful in the past and have shed light on the composition of protein and RNA components, optimum conditions, the nature of the products formed, cis-acting RNA elements and trans-acting protein factors required for efficient synthesis. In this review, we summarize our current understanding regarding the requirements for flavivirus RNA synthesis in vitro. We describe details of reaction conditions, the specificity of template used by either the multi-component membrane-bound viral replicase complex or by purified, recombinant RNA-dependent RNA polymerase. We also discuss future perspectives to extend the boundaries of our knowledge. PMID:26272247

  10. Kinetic analysis of elastomeric lag damper for helicopter rotors

    NASA Astrophysics Data System (ADS)

    Liu, Yafang; Wang, Jidong; Tong, Yan

    2018-02-01

    The elastomeric lag dampers suppress the ground resonance and air resonance that play a significant role in the stability of the helicopter. In this paper, elastomeric lag damper which is made from silicone rubber is built. And a series of experiments are conducted on this elastomeric lag damper. The stress-strain curves of elastomeric lag dampers employed shear forces at different frequency are obtained. And a finite element model is established based on Burgers model. The result of simulation and tests shows that the simple, linear model will yield good predictions of damper energy dissipation and it is adequate for predicting the stress-strain hysteresis loop within the operating frequency and a small-amplitude oscillation.

  11. GHRH excess and blockade in X-LAG syndrome.

    PubMed

    Daly, Adrian F; Lysy, Philippe A; Desfilles, Céline; Rostomyan, Liliya; Mohamed, Amira; Caberg, Jean-Hubert; Raverot, Veronique; Castermans, Emilie; Marbaix, Etienne; Maiter, Dominique; Brunelle, Chloe; Trivellin, Giampaolo; Stratakis, Constantine A; Bours, Vincent; Raftopoulos, Christian; Beauloye, Veronique; Barlier, Anne; Beckers, Albert

    2016-03-01

    X-linked acrogigantism (X-LAG) syndrome is a newly described form of inheritable pituitary gigantism that begins in early childhood and is usually associated with markedly elevated GH and prolactin secretion by mixed pituitary adenomas/hyperplasia. Microduplications on chromosome Xq26.3 including the GPR101 gene cause X-LAG syndrome. In individual cases random GHRH levels have been elevated. We performed a series of hormonal profiles in a young female sporadic X-LAG syndrome patient and subsequently undertook in vitro studies of primary pituitary tumor culture following neurosurgical resection. The patient demonstrated consistently elevated circulating GHRH levels throughout preoperative testing, which was accompanied by marked GH and prolactin hypersecretion; GH demonstrated a paradoxical increase following TRH administration. In vitro, the pituitary cells showed baseline GH and prolactin release that was further stimulated by GHRH administration. Co-incubation with GHRH and the GHRH receptor antagonist, acetyl-(d-Arg(2))-GHRH (1-29) amide, blocked the GHRH-induced GH stimulation; the GHRH receptor antagonist alone significantly reduced GH release. Pasireotide, but not octreotide, inhibited GH secretion. A ghrelin receptor agonist and an inverse agonist led to modest, statistically significant increases and decreases in GH secretion, respectively. GHRH hypersecretion can accompany the pituitary abnormalities seen in X-LAG syndrome. These data suggest that the pathology of X-LAG syndrome may include hypothalamic dysregulation of GHRH secretion, which is in keeping with localization of GPR101 in the hypothalamus. Therapeutic blockade of GHRH secretion could represent a way to target the marked hormonal hypersecretion and overgrowth that characterizes X-LAG syndrome. © 2016 Society for Endocrinology.

  12. Dietary choline deficiency causes DNA strand breaks and alters epigenetic marks on DNA and histones.

    PubMed

    Zeisel, Steven H

    2012-05-01

    Dietary choline is an important modulator of gene expression (via epigenetic marks) and of DNA integrity. Choline was discovered to be an essential nutrient for some humans approximately one decade ago. This requirement is diminished in young women because estrogen drives endogenous synthesis of phosphatidylcholine, from which choline can be derived. Almost half of women have a single nucleotide polymorphism that abrogates estrogen-induction of endogenous synthesis, and these women require dietary choline just as do men. In the US, dietary intake of choline is marginal. Choline deficiency in people is associated with liver and muscle dysfunction and damage, with apoptosis, and with increased DNA strand breaks. Several mechanisms explain these modifications to DNA. Choline deficiency increases leakage of reactive oxygen species from mitochondria consequent to altered mitochondrial membrane composition and enhanced fatty acid oxidation. Choline deficiency impairs folate metabolism, resulting in decreased thymidylate synthesis and increased uracil misincorporation into DNA, with strand breaks resulting during error-prone repair attempts. Choline deficiency alters DNA methylation, which alters gene expression for critical genes involved in DNA mismatch repair, resulting in increased mutation rates. Any dietary deficiency which increases mutation rates should be associated with increased risk of cancers, and this is the case for choline deficiency. In rodent models, diets low in choline and methyl-groups result in spontaneous hepatocarcinomas. In human epidemiological studies, there are interesting data that suggest that this also may be the case for humans, especially those with SNPs that increase the dietary requirement for choline. Copyright © 2011 Elsevier B.V. All rights reserved.

  13. Dietary Choline Deficiency causes DNA Strand Breaks and Alters Epigenetic Marks on DNA and Histones

    PubMed Central

    Zeisel, Steven H.

    2011-01-01

    Dietary choline is an important modulator of gene expression (via epigenetic marks) and of DNA integrity. Choline was discovered to be an essential nutrient for some humans approximately one decade ago. This requirement is diminished in young women because estrogen drives endogenous synthesis of phosphatidylcholine, from which choline can be derived. Almost half of women have a single nucleotide polymorphism that abrogates estrogen-induction of endogenous synthesis, and these women require dietary choline just as do men. In the US, dietary intake of choline is marginal. Choline deficiency in people is associated with liver and muscle dysfunction and damage, with apoptosis, and with increased DNA strand breaks. Several mechanisms explain these modifications to DNA. Choline deficiency increases leakage of reactive oxygen species from mitochondria consequent to altered mitochondrial membrane composition and enhanced fatty acid oxidation. Choline deficiency impairs folate metabolism, resulting in decreased thymidylate synthesis and increased uracil misincorporation into DNA, with strand breaks resulting during error-prone repair attempts. Choline deficiency alters DNA methylation, which alters gene expression for critical genes involved in DNA mismatch repair, resulting in increased mutation rates. Any dietary deficiency which increases mutation rates should be associated with increased risk of cancers, and this is the case for choline deficiency. In rodent models, diets low in choline and methyl-groups result in spontaneous hepatocarcinomas. In human epidemiological studies, there are interesting data that suggest that this also may be the case for humans, especially those with SNPs that increase the dietary requirement for choline. PMID:22041500

  14. Modelling toehold-mediated RNA strand displacement.

    PubMed

    Šulc, Petr; Ouldridge, Thomas E; Romano, Flavio; Doye, Jonathan P K; Louis, Ard A

    2015-03-10

    We study the thermodynamics and kinetics of an RNA toehold-mediated strand displacement reaction with a recently developed coarse-grained model of RNA. Strand displacement, during which a single strand displaces a different strand previously bound to a complementary substrate strand, is an essential mechanism in active nucleic acid nanotechnology and has also been hypothesized to occur in vivo. We study the rate of displacement reactions as a function of the length of the toehold and temperature and make two experimentally testable predictions: that the displacement is faster if the toehold is placed at the 5' end of the substrate; and that the displacement slows down with increasing temperature for longer toeholds. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  15. Mechanism of asymmetric polymerase assembly at the eukaryotic replication fork

    PubMed Central

    Georgescu, Roxana E; Langston, Lance; Yao, Nina Y; Yurieva, Olga; Zhang, Dan; Finkelstein, Jeff; Agarwal, Tani; O’Donnell, Mike E

    2015-01-01

    Eukaryotes use distinct polymerases for leading- and lagging-strand replication, but how they target their respective strands is uncertain. We reconstituted Saccharomyces cerevisiae replication forks and found that CMG helicase selects polymerase (Pol) ε to the exclusion of Pol δ on the leading strand. Even if Pol δ assembles on the leading strand, Pol ε rapidly replaces it. Pol δ–PCNA is distributive with CMG, in contrast to its high stability on primed ssDNA. Hence CMG will not stabilize Pol δ, instead leaving the leading strand accessible for Pol ε and stabilizing Pol ε. Comparison of Pol ε and Pol δ on a lagging-strand model DNA reveals the opposite. Pol δ dominates over excess Pol ε on PCNA-primed ssDNA. Thus, PCNA strongly favors Pol δ over Pol ε on the lagging strand, but CMG over-rides and flips this balance in favor of Pol ε on the leading strand. PMID:24997598

  16. Lags in the response of mountain plant communities to climate change.

    PubMed

    Alexander, Jake M; Chalmandrier, Loïc; Lenoir, Jonathan; Burgess, Treena I; Essl, Franz; Haider, Sylvia; Kueffer, Christoph; McDougall, Keith; Milbau, Ann; Nuñez, Martin A; Pauchard, Aníbal; Rabitsch, Wolfgang; Rew, Lisa J; Sanders, Nathan J; Pellissier, Loïc

    2018-02-01

    Rapid climatic changes and increasing human influence at high elevations around the world will have profound impacts on mountain biodiversity. However, forecasts from statistical models (e.g. species distribution models) rarely consider that plant community changes could substantially lag behind climatic changes, hindering our ability to make temporally realistic projections for the coming century. Indeed, the magnitudes of lags, and the relative importance of the different factors giving rise to them, remain poorly understood. We review evidence for three types of lag: "dispersal lags" affecting plant species' spread along elevational gradients, "establishment lags" following their arrival in recipient communities, and "extinction lags" of resident species. Variation in lags is explained by variation among species in physiological and demographic responses, by effects of altered biotic interactions, and by aspects of the physical environment. Of these, altered biotic interactions could contribute substantially to establishment and extinction lags, yet impacts of biotic interactions on range dynamics are poorly understood. We develop a mechanistic community model to illustrate how species turnover in future communities might lag behind simple expectations based on species' range shifts with unlimited dispersal. The model shows a combined contribution of altered biotic interactions and dispersal lags to plant community turnover along an elevational gradient following climate warming. Our review and simulation support the view that accounting for disequilibrium range dynamics will be essential for realistic forecasts of patterns of biodiversity under climate change, with implications for the conservation of mountain species and the ecosystem functions they provide. © 2017 John Wiley & Sons Ltd.

  17. The multiple personalities of Watson and Crick strands.

    PubMed

    Cartwright, Reed A; Graur, Dan

    2011-02-08

    In genetics it is customary to refer to double-stranded DNA as containing a "Watson strand" and a "Crick strand." However, there seems to be no consensus in the literature on the exact meaning of these two terms, and the many usages contradict one another as well as the original definition. Here, we review the history of the terminology and suggest retaining a single sense that is currently the most useful and consistent. The Saccharomyces Genome Database defines the Watson strand as the strand which has its 5'-end at the short-arm telomere and the Crick strand as its complement. The Watson strand is always used as the reference strand in their database. Using this as the basis of our standard, we recommend that Watson and Crick strand terminology only be used in the context of genomics. When possible, the centromere or other genomic feature should be used as a reference point, dividing the chromosome into two arms of unequal lengths. Under our proposal, the Watson strand is standardized as the strand whose 5'-end is on the short arm of the chromosome, and the Crick strand as the one whose 5'-end is on the long arm. Furthermore, the Watson strand should be retained as the reference (plus) strand in a genomic database. This usage not only makes the determination of Watson and Crick unambiguous, but also allows unambiguous selection of reference stands for genomics. This article was reviewed by John M. Logsdon, Igor B. Rogozin (nominated by Andrey Rzhetsky), and William Martin.

  18. The multiple personalities of Watson and Crick strands

    PubMed Central

    2011-01-01

    Background In genetics it is customary to refer to double-stranded DNA as containing a "Watson strand" and a "Crick strand." However, there seems to be no consensus in the literature on the exact meaning of these two terms, and the many usages contradict one another as well as the original definition. Here, we review the history of the terminology and suggest retaining a single sense that is currently the most useful and consistent. Proposal The Saccharomyces Genome Database defines the Watson strand as the strand which has its 5'-end at the short-arm telomere and the Crick strand as its complement. The Watson strand is always used as the reference strand in their database. Using this as the basis of our standard, we recommend that Watson and Crick strand terminology only be used in the context of genomics. When possible, the centromere or other genomic feature should be used as a reference point, dividing the chromosome into two arms of unequal lengths. Under our proposal, the Watson strand is standardized as the strand whose 5'-end is on the short arm of the chromosome, and the Crick strand as the one whose 5'-end is on the long arm. Furthermore, the Watson strand should be retained as the reference (plus) strand in a genomic database. This usage not only makes the determination of Watson and Crick unambiguous, but also allows unambiguous selection of reference stands for genomics. Reviewers This article was reviewed by John M. Logsdon, Igor B. Rogozin (nominated by Andrey Rzhetsky), and William Martin. PMID:21303550

  19. Spectral evolution of GRBs with negative spectral lag using Fermi GBM observations

    NASA Astrophysics Data System (ADS)

    Chakrabarti, Arundhati; Chaudhury, Kishor; Sarkar, Samir K.; Bhadra, Arunava

    2018-06-01

    The positive spectral lag of Gamma Ray Bursts (GRBs) is often explained in terms of hard-to-soft spectral evolution of GRB pulses. While positive lags of GRBs is very common, there are few GRB pulses that exhibits negative spectral lags. In the present work we examine whether negative lags of GRBs also can be interpreted in terms of spectral evolution of GRB pulses or not. Using Fermi-GBM data, we identify two GRBs, GRB 090426C and GRB 150213A, with clean pulses that exhibit negative spectral lag. An indication of soft to hard transition has been noticed for the negative spectral lag events from the spectral evolution study. The implication of the present findings on the models of GRB spectral lags are discussed.

  20. Transcriptional requirements of the distal heavy-strand promoter of mtDNA

    PubMed Central

    Zollo, Ornella; Tiranti, Valeria; Sondheimer, Neal

    2012-01-01

    The heavy strand of mtDNA contains two promoters with nonoverlapping functions. The role of the minor heavy-strand promoter (HSP2) is controversial, because the promoter has been difficult to activate in an in vitro system. We have isolated HSP2 by excluding its interaction with the more powerful HSP1 promoter, and we find that it is transcribed efficiently by recombinant mtRNA polymerase and mitochondrial transcription factor B2. The mitochondrial transcription factor A is not required for initiation, but it has the ability to alternatively activate and repress the HSP2 transcriptional unit depending on the ratio between mitochondrial transcription factor A and other transcription factors. The positioning of transcriptional initiation agrees with our current understanding of HSP2 activity in vivo. Serial deletion of HSP2 shows that only proximal sequences are required. Several mutations, including the disruption of a polycytosine track upstream of the HSP2 initiation site, influence transcriptional activity. Transcription from HSP2 is also observed when HeLa cell mitochondrial extract is used as the source of mitochondrial polymerase, and this transcription is maintained when HSP2 is provided in proper spacing and context to the HSP1 promoter. Studies of the linked heavy-strand promoters show that they are differentially regulated by ATP dosage. We conclude that HSP2 is transcribed and has features that allow it to regulate mitochondrial mRNA synthesis. PMID:22454497

  1. In vivo production of RNA nanostructures via programmed folding of single-stranded RNAs.

    PubMed

    Li, Mo; Zheng, Mengxi; Wu, Siyu; Tian, Cheng; Liu, Di; Weizmann, Yossi; Jiang, Wen; Wang, Guansong; Mao, Chengde

    2018-06-06

    Programmed self-assembly of nucleic acids is a powerful approach for nano-constructions. The assembled nanostructures have been explored for various applications. However, nucleic acid assembly often requires chemical or in vitro enzymatical synthesis of DNA or RNA, which is not a cost-effective production method on a large scale. In addition, the difficulty of cellular delivery limits the in vivo applications. Herein we report a strategy that mimics protein production. Gene-encoded DNA duplexes are transcribed into single-stranded RNAs, which self-fold into well-defined RNA nanostructures in the same way as polypeptide chains fold into proteins. The resulting nanostructure contains only one component RNA molecule. This approach allows both in vitro and in vivo production of RNA nanostructures. In vivo synthesized RNA strands can fold into designed nanostructures inside cells. This work not only suggests a way to synthesize RNA nanostructures on a large scale and at a low cost but also facilitates the in vivo applications.

  2. Emergence and temporal structure of Lead-Lag correlations in collective stock dynamics

    NASA Astrophysics Data System (ADS)

    Xia, Lisi; You, Daming; Jiang, Xin; Chen, Wei

    2018-07-01

    Understanding the correlations among stock returns is crucial for reducing the risk of investment in stock markets. As an important stylized correlation, lead-lag effect plays a major role in analyzing market volatility and deriving trading strategies. Here, we explore historical lead-lag relationships among stocks in the Chinese stock market. Strongly positive lagged correlations can be empirically observed. We demonstrate this lead-lag phenomenon is not constant but temporally emerges during certain periods. By introducing moving time window method, we transform the lead-lag dynamics into a series of asymmetric lagged correlation matrices. Dynamic lead-lag structures are uncovered in the form of temporal network structures. We find that the size of lead-lag group experienced a rapid drop during the year 2012, which signaled a re-balance of the stock market. On the daily timescale, we find the lead-lag structure exhibits several persistent patterns, which can be characterized by the Jaccard matrix. We show significant market events can be distinguished in the Jaccard matrix diagram. Taken together, we study an integration of all the temporal networks and identify several leading stock sectors, which are in accordance with the common Chinese economic fundamentals.

  3. Burst Statistics Using the Lag-Luminosity Relationship

    NASA Technical Reports Server (NTRS)

    Band, D. L.; Norris, J. P.; Bonnell, J. T.

    2003-01-01

    Using the lag-luminosity relation and various BATSE catalogs we create a large catalog of burst redshifts, peak luminosities and emitted energies. These catalogs permit us to evaluate the lag-luminosity relation, and to study the burst energy distribution. We find that this distribution can be described as a power law with an index of alpha = 1.76 +/- 0.05 (95% confidence), close to the alpha = 2 predicted by the original quasi-universal jet model.

  4. Synthesis of bacteriophage phiC DNA in dna mutants of Esherichia coli.

    PubMed

    Kodaira, K I; Taketo, A

    1978-06-01

    Host dna functions involved in the replication of microvirid phage phiC DNA were investigated in vivo. Although growth of this phage was markedly inhibited even at 35-37 degrees C even in dna+ host, conversion of the infecting single-stranded DNA into the double-stranded parental replicative form (stage I synthesis) occurred normally at 43 degrees C in dna+, dnaA, dnaB, dnaC(D), and dnaE cells. In dnaG mutant, the stage I synthesis was severely inhibited at 43 degrees C but not at 30 degrees C. The stage I replication of phiC DNA was clearly thermosensitive in dnaZ cells incubated in nutrient broth. In Tris-casamino acids-glucose medium, however, the dnaZ mutant sufficiently supported synthesis of the parental replicative form. At 43 degrees C, synthesis of the progeny replicative form DNA (stage II replication) was significantly inhibited even in dna+ cells and was nearly completely blocked in dnaB or dnaC(D) mutant. At 37 degrees C, the stage II replication proceeded normally in dna+ bacteria.

  5. Mechanisms of Zero-Lag Synchronization in Cortical Motifs

    PubMed Central

    Gollo, Leonardo L.; Mirasso, Claudio; Sporns, Olaf; Breakspear, Michael

    2014-01-01

    Zero-lag synchronization between distant cortical areas has been observed in a diversity of experimental data sets and between many different regions of the brain. Several computational mechanisms have been proposed to account for such isochronous synchronization in the presence of long conduction delays: Of these, the phenomenon of “dynamical relaying” – a mechanism that relies on a specific network motif – has proven to be the most robust with respect to parameter mismatch and system noise. Surprisingly, despite a contrary belief in the community, the common driving motif is an unreliable means of establishing zero-lag synchrony. Although dynamical relaying has been validated in empirical and computational studies, the deeper dynamical mechanisms and comparison to dynamics on other motifs is lacking. By systematically comparing synchronization on a variety of small motifs, we establish that the presence of a single reciprocally connected pair – a “resonance pair” – plays a crucial role in disambiguating those motifs that foster zero-lag synchrony in the presence of conduction delays (such as dynamical relaying) from those that do not (such as the common driving triad). Remarkably, minor structural changes to the common driving motif that incorporate a reciprocal pair recover robust zero-lag synchrony. The findings are observed in computational models of spiking neurons, populations of spiking neurons and neural mass models, and arise whether the oscillatory systems are periodic, chaotic, noise-free or driven by stochastic inputs. The influence of the resonance pair is also robust to parameter mismatch and asymmetrical time delays amongst the elements of the motif. We call this manner of facilitating zero-lag synchrony resonance-induced synchronization, outline the conditions for its occurrence, and propose that it may be a general mechanism to promote zero-lag synchrony in the brain. PMID:24763382

  6. Evidence for a Single-Stranded Adenovirus-Associated Virus Genome: Isolation and Separation of Complementary Single Strands

    PubMed Central

    Berns, K. I.; Rose, J. A.

    1970-01-01

    Single-stranded adenovirus-associated virus type 2 deoxyribonucleic acid (AAV-2 DNA) has been isolated from the virion after enzymatic pretreatment of the particles by heating at 53 C for 1 hr in 0.015 m NaCl plus 0.0015 m sodium citrate in the presence of 1% sodium dodecyl sulfate. Double-stranded AAV-2 DNA present as a marker is not denatured by this treatment. AAV-2 single-stranded DNA is composed of two complementary species which can be separated in neutral CsCl when 5-bromodeoxyuridine has been substituted for thymidine in the DNA. The present report is the first documented instance of the separation of complementary strands of an animal virus DNA. PMID:5429749

  7. Monthly ENSO Forecast Skill and Lagged Ensemble Size

    PubMed Central

    DelSole, T.; Tippett, M.K.; Pegion, K.

    2018-01-01

    Abstract The mean square error (MSE) of a lagged ensemble of monthly forecasts of the Niño 3.4 index from the Climate Forecast System (CFSv2) is examined with respect to ensemble size and configuration. Although the real‐time forecast is initialized 4 times per day, it is possible to infer the MSE for arbitrary initialization frequency and for burst ensembles by fitting error covariances to a parametric model and then extrapolating to arbitrary ensemble size and initialization frequency. Applying this method to real‐time forecasts, we find that the MSE consistently reaches a minimum for a lagged ensemble size between one and eight days, when four initializations per day are included. This ensemble size is consistent with the 8–10 day lagged ensemble configuration used operationally. Interestingly, the skill of both ensemble configurations is close to the estimated skill of the infinite ensemble. The skill of the weighted, lagged, and burst ensembles are found to be comparable. Certain unphysical features of the estimated error growth were tracked down to problems with the climatology and data discontinuities. PMID:29937973

  8. Monthly ENSO Forecast Skill and Lagged Ensemble Size

    NASA Astrophysics Data System (ADS)

    Trenary, L.; DelSole, T.; Tippett, M. K.; Pegion, K.

    2018-04-01

    The mean square error (MSE) of a lagged ensemble of monthly forecasts of the Niño 3.4 index from the Climate Forecast System (CFSv2) is examined with respect to ensemble size and configuration. Although the real-time forecast is initialized 4 times per day, it is possible to infer the MSE for arbitrary initialization frequency and for burst ensembles by fitting error covariances to a parametric model and then extrapolating to arbitrary ensemble size and initialization frequency. Applying this method to real-time forecasts, we find that the MSE consistently reaches a minimum for a lagged ensemble size between one and eight days, when four initializations per day are included. This ensemble size is consistent with the 8-10 day lagged ensemble configuration used operationally. Interestingly, the skill of both ensemble configurations is close to the estimated skill of the infinite ensemble. The skill of the weighted, lagged, and burst ensembles are found to be comparable. Certain unphysical features of the estimated error growth were tracked down to problems with the climatology and data discontinuities.

  9. Lag and anticipating synchronization without time-delay coupling.

    PubMed

    Corron, Ned J; Blakely, Jonathan N; Pethel, Shawn D

    2005-06-01

    We describe a new method for achieving approximate lag and anticipating synchronization in unidirectionally coupled chaotic oscillators. The method uses a specific parameter mismatch between the drive and response that is a first-order approximation to true time-delay coupling. As a result, an adjustable lag or anticipation effect can be achieved without the need for a variable delay line, making the method simpler and more economical to implement in many physical systems. We present a stability analysis, demonstrate the method numerically, and report experimental observation of the effect in radio-frequency electronic oscillators. In the circuit experiments, both lag and anticipation are controlled by tuning a single capacitor in the response oscillator.

  10. Signatures of Steady Heating in Time Lag Analysis of Coronal Emission

    NASA Technical Reports Server (NTRS)

    Viall, Nicholeen M.; Klimchuk, James A.

    2016-01-01

    Among the multitude of methods used to investigate coronal heating, the time lag method of Viall Klimchuk is becoming increasingly prevalent as an analysis technique that is complementary to those that are traditionally used.The time lag method cross correlates light curves at a given spatial location obtained in spectral bands that sample different temperature plasmas. It has been used most extensively with data from the Atmospheric Imaging Assembly on the Solar Dynamics Observatory. We have previously applied the time lag method to entire active regions and surrounding the quiet Sun and created maps of the results. We find that the majority of time lags are consistent with the cooling of coronal plasma that has been impulsively heated. Additionally, a significant fraction of the map area has a time lag of zero. This does not indicate a lack of variability. Rather, strong variability must be present, and it must occur in phase between the different channels. We have previously shown that these zero time lags are consistent with the transition region response to coronal nanoflares, although other explanations are possible. A common misconception is that the zero time lag indicates steady emission resulting from steady heating. Using simulated and observed light curves, we demonstrate here that highly correlated light curves at zero time lag are not compatible with equilibrium solutions. Such light curves can only be created by evolution

  11. RPA Stabilization of Single-Stranded DNA Is Critical for Break-Induced Replication.

    PubMed

    Ruff, Patrick; Donnianni, Roberto A; Glancy, Eleanor; Oh, Julyun; Symington, Lorraine S

    2016-12-20

    DNA double-strand breaks (DSBs) are cytotoxic lesions that must be accurately repaired to maintain genome stability. Replication protein A (RPA) plays an important role in homology-dependent repair of DSBs by protecting the single-stranded DNA (ssDNA) intermediates formed by end resection and by facilitating Rad51 loading. We found that hypomorphic mutants of RFA1 that support intra-chromosomal homologous recombination are profoundly defective for repair processes involving long tracts of DNA synthesis, in particular break-induced replication (BIR). The BIR defects of the rfa1 mutants could be partially suppressed by eliminating the Sgs1-Dna2 resection pathway, suggesting that Dna2 nuclease attacks the ssDNA formed during end resection when not fully protected by RPA. Overexpression of Rad51 was also found to suppress the rfa1 BIR defects. We suggest that Rad51 binding to the ssDNA formed by excessive end resection and during D-loop migration can partially compensate for dysfunctional RPA. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  12. A strand graph semantics for DNA-based computation

    PubMed Central

    Petersen, Rasmus L.; Lakin, Matthew R.; Phillips, Andrew

    2015-01-01

    DNA nanotechnology is a promising approach for engineering computation at the nanoscale, with potential applications in biofabrication and intelligent nanomedicine. DNA strand displacement is a general strategy for implementing a broad range of nanoscale computations, including any computation that can be expressed as a chemical reaction network. Modelling and analysis of DNA strand displacement systems is an important part of the design process, prior to experimental realisation. As experimental techniques improve, it is important for modelling languages to keep pace with the complexity of structures that can be realised experimentally. In this paper we present a process calculus for modelling DNA strand displacement computations involving rich secondary structures, including DNA branches and loops. We prove that our calculus is also sufficiently expressive to model previous work on non-branching structures, and propose a mapping from our calculus to a canonical strand graph representation, in which vertices represent DNA strands, ordered sites represent domains, and edges between sites represent bonds between domains. We define interactions between strands by means of strand graph rewriting, and prove the correspondence between the process calculus and strand graph behaviours. Finally, we propose a mapping from strand graphs to an efficient implementation, which we use to perform modelling and simulation of DNA strand displacement systems with rich secondary structure. PMID:27293306

  13. LAG-3 Confers a Competitive Disadvantage upon Antiviral CD8+ T Cell Responses.

    PubMed

    Cook, Kevin D; Whitmire, Jason K

    2016-07-01

    Ongoing clinical trials are evaluating the benefits of systemic blockade of lymphocyte activation gene-3 (LAG-3) signals to improve immunity to tumors. Those studies are founded on the well-established inhibitory role of LAG-3 in regulating CD8(+) T cells during chronic virus infection and antitumor responses. However, the T cell response in LAG-3-deficient mice is similar in size and function to that in wild type animals, suggesting LAG-3 has nuanced immune-regulatory functions. We performed a series of adoptive transfer experiments in mice to better understand the T cell-intrinsic functions of LAG-3 in the regulation of CD8(+) T cell responses. Our results indicate that LAG-3 expression by CD8(+) T cells inhibits their competitive fitness and results in a slightly reduced rate of cell division in comparison with LAG-3-deficient cells. This cell-intrinsic effect of LAG-3 was consistent across both acute and chronic virus infections. These data show that LAG-3 directly modulates the size of the T cell response and support the use of LAG-3 blockade regimens to enhance CD8(+) T cell responses. Copyright © 2016 by The American Association of Immunologists, Inc.

  14. [Results of flexor tendon sutures of the fingers with 2-strand (40 tendons) and 4-strand (64 tendons) core sutures].

    PubMed

    Winkel, R; Kalbhenn, O; Hoffmann, R

    2012-06-01

    This retrospective examination compares the results of finger flexor tendon sutures with 2 strands and 4 strands. It was checked, whether and how 2 more strands influenced the rupture rate, the movement of the finger and the contentment of the patients. From 1996 to 2000 for the core suture of the flexor tendon of fingers we used 2 strands. 35 patients with 40 tendon sutures of 73 patients were examined. From 2001 to 2005 we used for the core suture 2 loop threads. 53 patients with 64 tendon sutures from a total of 111 patients were examined. At least 12 months had passed between operation and the examination. The rupture rate and the range of movement of each finger joint and the total mobility of the affected fingers were evaluated. Each case was compared to the uninjured opposite hand. The functional result was judged according to the score of Buck-Gramcko. The patient's contentment was recorded by the DASH (disability of arm, shoulder and hand) score. Effects of gender, age, accompanying injuries, zone of the injury and their influence on the results were analysed. The Buck-Gramcko score showed in the 2-strand group a distribution from summarised 70% "excellent" and "good" and 30% "fair" and "poor". In the 4-strand-group the relation was 93.7% "excellent" and "good", 6.3% "fair", one "poor". In the 2-strand group 2/40 (5%) of the tendon sutures ruptured, in the 4-strand group 1/64 (1.6%) ruptured. The average DASH value in the 2-strands-group was 16.6/100, in the 4-strands-group 18.1/100 when 0 is the best possible result and 100 the worst. The patient judgement in the 2-strand group was summarised to 70% for "excellent" and "good" and 30% "fair" and "poor". In the 4-strand group the patient's judgment was summarised in 75% "excellent" and "good" and in 25% "fair". The results of flexor tendon sutures with 4-strand core sutures have been superior to the results with 2-strand core suture according to range of motion of the fingers (P <0.005). © Georg Thieme

  15. Non-encapsidation Activities of the Capsid Proteins of Positive-strand RNA Viruses

    PubMed Central

    Ni, Peng; Kao, C. Cheng

    2013-01-01

    Viral capsid proteins (CPs) are characterized by their role in forming protective shells around viral genomes. However, CPs have additional and important roles in the virus infection cycles and in the cellular response to infection. These activities involve CP binding to RNAs in both sequence-specific and nonspecific manners as well as association with other proteins. This review focuses on CPs of both plant and animal-infecting viruses with positive-strand RNA genomes. We summarize the structural features of CPs and describe their modulatory roles in viral translation, RNA-dependent RNA synthesis, and host defense responses. PMID:24074574

  16. [Expression and purification of a novel thermophilic bacterial single-stranded DNA-binding protein and enhancement the synthesis of DNA and cDNA].

    PubMed

    Jia, Xiao-Wei; Zhang, Guo-Hui; Shi, Hai-Yan

    2012-12-01

    Express a novel species of single-stranded DNA-binding protein (SSB) derived from Thermococcus kodakarensis KOD1, abbreviated kod-ssb. And evaluate the effect of kod-ssb on PCR-based DNA amplification and reverse transcription. We express kod-ssb with the Transrtta (DE3), and kod-ssb was purified by affinity chromatography on a Ni2+ Sepharose column, detected by SDS-PAGE. To evaluate the effect of kod-ssb on PCR-based DNA amplification, the human beta globin gene was used as template to amplify a 5-kb, 9-kb and 13-kb. And to detect the effect of kod-ssb on reverse transcription, we used RNA from flu cell culture supernatant extraction as templates to implement qRT-PCR reaction. The plasmid pET11a-kod was transformed into Transetta (DE3) and the recombinant strain Transetta (pET11 a-kod) was obtained. The kod-ssb was highly expressed when the recombinant strain Transetta(pET11a-kod) was induced by IPTG. The specific protein was detected by SDS-PAGE. To confirm that kod-ssb can enhance target DNA synthesis and reduce PCR by-products, 5-, 9-, and 13-kb human beta globin gene fragments were used as templates for PCR. When PCR reactions did not include SSB proteins, the specific PCR product was contaminated with non-specific products. When kod -ssb was added, kod-ssb significantly enhanced amplification of the 5-, 9-and 13-kb target product and minimised the non-specific PCR products. To confirm that kod-ssb can enhance target cDNA synthesis, RNA from flu cell culture supernatant extraction was used as templates for qRT-PCR reaction. The results was that when kod-ssb was added, kod-ssb significantly enhanced the synthesis of cDNA, average Ct value is 19.42, and the average Ct value without kod-ssb is 22.15. kod-ssb may in future be used to enhance DNA and cDNA amplification.

  17. Strand-specific transcriptome profiling with directly labeled RNA on genomic tiling microarrays

    PubMed Central

    2011-01-01

    Background With lower manufacturing cost, high spot density, and flexible probe design, genomic tiling microarrays are ideal for comprehensive transcriptome studies. Typically, transcriptome profiling using microarrays involves reverse transcription, which converts RNA to cDNA. The cDNA is then labeled and hybridized to the probes on the arrays, thus the RNA signals are detected indirectly. Reverse transcription is known to generate artifactual cDNA, in particular the synthesis of second-strand cDNA, leading to false discovery of antisense RNA. To address this issue, we have developed an effective method using RNA that is directly labeled, thus by-passing the cDNA generation. This paper describes this method and its application to the mapping of transcriptome profiles. Results RNA extracted from laboratory cultures of Porphyromonas gingivalis was fluorescently labeled with an alkylation reagent and hybridized directly to probes on genomic tiling microarrays specifically designed for this periodontal pathogen. The generated transcriptome profile was strand-specific and produced signals close to background level in most antisense regions of the genome. In contrast, high levels of signal were detected in the antisense regions when the hybridization was done with cDNA. Five antisense areas were tested with independent strand-specific RT-PCR and none to negligible amplification was detected, indicating that the strong antisense cDNA signals were experimental artifacts. Conclusions An efficient method was developed for mapping transcriptome profiles specific to both coding strands of a bacterial genome. This method chemically labels and uses extracted RNA directly in microarray hybridization. The generated transcriptome profile was free of cDNA artifactual signals. In addition, this method requires fewer processing steps and is potentially more sensitive in detecting small amount of RNA compared to conventional end-labeling methods due to the incorporation of more

  18. Intracellular generation of single-strand template increases the knock-in efficiency by combining CRISPR/Cas9 with AAV.

    PubMed

    Xiao, Qing; Min, Taishan; Ma, Shuangping; Hu, Lingna; Chen, Hongyan; Lu, Daru

    2018-04-18

    Targeted integration of transgenes facilitates functional genomic research and holds prospect for gene therapy. The established microhomology-mediated end-joining (MMEJ)-based strategy leads to the precise gene knock-in with easily constructed donor, yet the limited efficiency remains to be further improved. Here, we show that single-strand DNA (ssDNA) donor contributes to efficient increase of knock-in efficiency and establishes a method to achieve the intracellular linearization of long ssDNA donor. We identified that the CRISPR/Cas9 system is responsible for breaking double-strand DNA (dsDNA) of palindromic structure in inverted terminal repeats (ITRs) region of recombinant adeno-associated virus (AAV), leading to the inhibition of viral second-strand DNA synthesis. Combing Cas9 plasmids targeting genome and ITR with AAV donor delivery, the precise knock-in of gene cassette was achieved, with 13-14% of the donor insertion events being mediated by MMEJ in HEK 293T cells. This study describes a novel method to integrate large single-strand transgene cassettes into the genomes, increasing knock-in efficiency by 13.6-19.5-fold relative to conventional AAV-mediated method. It also provides a comprehensive solution to the challenges of complicated production and difficult delivery with large exogenous fragments.

  19. Marine mammal strandings in the New Caledonia region, Southwest Pacific.

    PubMed

    Borsa, Philippe

    2006-04-01

    Four hundred twenty three marine mammals, in 72 stranding events, were recorded between 1877 and 2005 in New Caledonia, the Loyalty Islands, and Vanuatu in the southwest Pacific. Sixteen species were represented in this count, including: minke whale, Balaenoptera acutorostrata (1 single stranding), sei whale, B. borealis (1 single stranding), blue whale, B. musculus (1 single stranding), humpback whale, Megaptera novaeangliae (2 single strandings), giant sperm whale, Physeter macrocephalus (18 single strandings, 2 pair strandings), pygmy sperm whale, Kogia breviceps (5 single strandings), dwarf sperm whale, K. sima (2 single strandings, 1 triple stranding), Blainville's beaked whale, Mesoplodon densirostris (2 single strandings), short-finned pilot whale, Globicephala macrorhynchus (4 strandings, 56 individuals), melon-headed whale, Peponocephala electra (1 single stranding and 2 mass strandings totalling 231 individuals), common dolphin, Delphinus delphis (1 single stranding), spinner dolphin, Stenella longirostris (1 pair stranding and 2 mass strandings of groups of approximately 30 individuals each), Indian Ocean bottlenose dolphin, Tursiops aduncus (2 single strandings), dugong, Dugong dugon (14 single strandings), and New Zealand fur seal, Arctocephalus forsteri (3 single strandings). A stranded rorqual identified as an Antarctic minke whale (B. bonaerensis), with coloration patterns that did not match known descriptions, was also reported. Sei whale was recorded for the first time in the tropical Southwest Pacific region and Antarctic minke whale, melon-headed whale, and Indian Ocean bottlenose dolphin were recorded for the first time in New Caledonia. Strandings of sperm whales were most frequent in the spring, but also occurred in autumn months, suggesting a seasonal pattern of occurrence possibly related to seasonal migration. One stranded humpback whale bore the scars of a killer whale's attack and one dugong was injured by a shark. Scars left by

  20. The drug lag: an interpretive review of the literature.

    PubMed

    Schifrin, L G; Tayan, J R

    1977-01-01

    This articel reviews the literature pertinent to the "drug lag" issue, in order to evaluate the evidence underlying the thesis that the U.S. has suffered from a slow-down in the rate and timing of new drug introductions, to the detriment of patients, because of the stricter requirements since 1962 regarding proof of safety and efficacy for new drugs. Comparing the post-1962 record in the U.S. with (a) the U.S. record before 1962 and (b) the post-1962 record in other, mainly Western European, countries, the weight of evidence and argument falls on the side of those who see a lag existing in the U.S. Other evidence, more subjective, supports the view that this lag imposes net positive costs on U.S. patients. However, it is not clear that the 1962 Drug Act is the sole, or even the main, cause of the lag. Instead, the drug information system and its inefficiencies emerge as the fundamental reason for the existence of a lag; hence, changes in the administration of the law by the Food and Drug Administration and greater efficiency by companies in supporting New Drug Applications can help close the gap, but probably only in minor degree. The solution to the lag problem lies in formulating and implementing a drug information system that quickly and accurately gathers, interprets, and disseminates information on the positive and negative effects of newly introduced drugs; with such a system, protection of drug users from undue risk is compatible with a greater rate and more rapid appearance of new drug discoveries.

  1. How jet lag impairs Major League Baseball performance.

    PubMed

    Song, Alex; Severini, Thomas; Allada, Ravi

    2017-02-07

    Laboratory studies have demonstrated that circadian clocks align physiology and behavior to 24-h environmental cycles. Examination of athletic performance has been used to discern the functions of these clocks in humans outside of controlled settings. Here, we examined the effects of jet lag, that is, travel that shifts the alignment of 24-h environmental cycles relative to the endogenous circadian clock, on specific performance metrics in Major League Baseball. Accounting for potential differences in home and away performance, travel direction, and team confounding variables, we observed that jet-lag effects were largely evident after eastward travel with very limited effects after westward travel, consistent with the >24-h period length of the human circadian clock. Surprisingly, we found that jet lag impaired major parameters of home-team offensive performance, for example, slugging percentage, but did not similarly affect away-team offensive performance. On the other hand, jet lag impacted both home and away defensive performance. Remarkably, the vast majority of these effects for both home and away teams could be explained by a single measure, home runs allowed. Rather than uniform effects, these results reveal surprisingly specific effects of circadian misalignment on athletic performance under natural conditions.

  2. Improving the effectiveness of an interruption lag by inducing a memory-based strategy.

    PubMed

    Morgan, Phillip L; Patrick, John; Tiley, Leyanne

    2013-01-01

    The memory for goals model (Altmann & Trafton, 2002) posits the importance of a short delay (the 'interruption lag') before an interrupting task to encode suspended goals for retrieval post-interruption. Two experiments used the theory of soft constraints (Gray, Simms, Fu & Schoelles, 2006) to investigate whether the efficacy of an interruption lag could be improved by increasing goal-state access cost to induce a more memory-based encoding strategy. Both experiments used a copying task with three access cost conditions (Low, Medium, and High) and a 5-s interruption lag with a no lag control condition. Experiment 1 found that the participants in the High access cost condition resumed more interrupted trials and executed more actions correctly from memory when coupled with an interruption lag. Experiment 2 used a prospective memory test post-interruption and an eyetracker recorded gaze activity during the interruption lag. The participants in the High access cost condition with an interruption lag were best at encoding target information during the interruption lag, evidenced by higher scores on the prospective memory measure and more gaze activity on the goal-state during the interruption lag. Theoretical and practical issues regarding the use of goal-state access cost and an interruption lag are discussed. Copyright © 2012. Published by Elsevier B.V.

  3. Protected DNA strand displacement for enhanced single nucleotide discrimination in double-stranded DNA.

    PubMed

    Khodakov, Dmitriy A; Khodakova, Anastasia S; Huang, David M; Linacre, Adrian; Ellis, Amanda V

    2015-03-04

    Single nucleotide polymorphisms (SNPs) are a prime source of genetic diversity. Discriminating between different SNPs provides an enormous leap towards the better understanding of the uniqueness of biological systems. Here we report on a new approach for SNP discrimination using toehold-mediated DNA strand displacement. The distinctiveness of the approach is based on the combination of both 3- and 4-way branch migration mechanisms, which allows for reliable discrimination of SNPs within double-stranded DNA generated from real-life human mitochondrial DNA samples. Aside from the potential diagnostic value, the current study represents an additional way to control the strand displacement reaction rate without altering other reaction parameters and provides new insights into the influence of single nucleotide substitutions on 3- and 4-way branch migration efficiency and kinetics.

  4. Eating Disorder Symptomatology and Identity Formation in Adolescence: A Cross-Lagged Longitudinal Approach.

    PubMed

    Verschueren, Margaux; Claes, Laurence; Bogaerts, Annabel; Palmeroni, Nina; Gandhi, Amarendra; Moons, Philip; Luyckx, Koen

    2018-01-01

    Introduction: Eating disorder symptomatology, comprising both psychological and behavioral aspects of subclinical eating concerns, constitutes a clear precursor of developing eating disorders. It is crucial to investigate its antecedents and correlates to subsequently inform eating disorder prevention programs. The present study focused on identity formation, a core developmental task in adolescence, that has increasingly been linked to eating disorder development. Our main aim was to examine the temporal sequence between eating disorder symptomatology and identity formation. Methods: Data on eating disorder symptomatology and identity formation were collected in 530 high school students (at Time 1: mean age = 15 years; SD = 1.84; range: 12-18 years; 50.6% females) using self-report questionnaires at three annual measurement points. Cross-lagged structural equation modeling was performed to examine the directionality of effects. Results: Results indicated bidirectional effects between eating disorder symptomatology and identity formation. Identity confusion seemed to increase vulnerability to body dissatisfaction and bulimia symptoms, whereas identity synthesis seemed to protect against their development. Additionally, identity synthesis seemed to protect against the development of drive for thinness as well. At the same time, body dissatisfaction and bulimia symptoms positively predicted identity confusion and negatively predicted identity synthesis over time. Conclusion: The present study adds to the growing body of literature on identity and eating disorders by focusing on their temporal interplay in a community sample of adolescents. As bidirectional effects emerged, a greater emphasis on identity formation in eating disorder prevention programs is advocated.

  5. Eating Disorder Symptomatology and Identity Formation in Adolescence: A Cross-Lagged Longitudinal Approach

    PubMed Central

    Verschueren, Margaux; Claes, Laurence; Bogaerts, Annabel; Palmeroni, Nina; Gandhi, Amarendra; Moons, Philip; Luyckx, Koen

    2018-01-01

    Introduction: Eating disorder symptomatology, comprising both psychological and behavioral aspects of subclinical eating concerns, constitutes a clear precursor of developing eating disorders. It is crucial to investigate its antecedents and correlates to subsequently inform eating disorder prevention programs. The present study focused on identity formation, a core developmental task in adolescence, that has increasingly been linked to eating disorder development. Our main aim was to examine the temporal sequence between eating disorder symptomatology and identity formation. Methods: Data on eating disorder symptomatology and identity formation were collected in 530 high school students (at Time 1: mean age = 15 years; SD = 1.84; range: 12–18 years; 50.6% females) using self-report questionnaires at three annual measurement points. Cross-lagged structural equation modeling was performed to examine the directionality of effects. Results: Results indicated bidirectional effects between eating disorder symptomatology and identity formation. Identity confusion seemed to increase vulnerability to body dissatisfaction and bulimia symptoms, whereas identity synthesis seemed to protect against their development. Additionally, identity synthesis seemed to protect against the development of drive for thinness as well. At the same time, body dissatisfaction and bulimia symptoms positively predicted identity confusion and negatively predicted identity synthesis over time. Conclusion: The present study adds to the growing body of literature on identity and eating disorders by focusing on their temporal interplay in a community sample of adolescents. As bidirectional effects emerged, a greater emphasis on identity formation in eating disorder prevention programs is advocated. PMID:29915548

  6. A homogeneous nucleic acid hybridization assay based on strand displacement.

    PubMed Central

    Vary, C P

    1987-01-01

    A homogeneous nucleic acid hybridization assay which is conducted in solution and requires no separation steps is described. The assay is based on the concept of strand displacement. In the strand displacement assay, an RNA "signal strand" is hybridized within a larger DNA strand termed the "probe strand", which is, in turn, complementary to the target nucleic acid of interest. Hybridization of the target nucleic acid with the probe strand ultimately results in displacement of the RNA signal strand. Strand displacement, therefore, causes conversion of the RNA from double to single-stranded form. The single-strand specificity of polynucleotide phosphorylase (EC 2.7.7.8) allows discrimination between double-helical and single-stranded forms of the RNA signal strand. As displacement proceeds, free RNA signal strands are preferentially phosphorolyzed to component nucleoside diphosphates, including adenosine diphosphate. The latter nucleotide is converted to ATP by pyruvate kinase(EC 2.7.1.40). Luciferase catalyzed bioluminescence is employed to measure the ATP generated as a result of strand displacement. Images PMID:3309890

  7. Modulation of mutagenesis in eukaryotes by DNA replication fork dynamics and quality of nucleotide pools

    PubMed Central

    Waisertreiger, Irina S.-R.; Liston, Victoria G.; Menezes, Miriam R.; Kim, Hyun-Min; Lobachev, Kirill S.; Stepchenkova, Elena I.; Tahirov, Tahir H.; Rogozin, Igor B.; Pavlov, Youri. I.

    2014-01-01

    The rate of mutations in eukaryotes depends on a plethora of factors and is not immediately derived from the fidelity of DNA polymerases (Pols). Replication of chromosomes containing the anti-parallel strands of duplex DNA occurs through the copying of leading and lagging strand templates by a trio of Pols α, δ and ε, with the assistance of Pol ζ and Y-family Pols at difficult DNA template structures or sites of DNA damage. The parameters of the synthesis at a given location are dictated by the quality and quantity of nucleotides in the pools, replication fork architecture, transcription status, regulation of Pol switches, and structure of chromatin. The result of these transactions is a subject of survey and editing by DNA repair. PMID:23055184

  8. Chromatin Constrains the Initiation and Elongation of DNA Replication.

    PubMed

    Devbhandari, Sujan; Jiang, Jieqing; Kumar, Charanya; Whitehouse, Iestyn; Remus, Dirk

    2017-01-05

    Eukaryotic chromosomal DNA is faithfully replicated in a complex series of cell-cycle-regulated events that are incompletely understood. Here we report the reconstitution of DNA replication free in solution with purified proteins from the budding yeast Saccharomyces cerevisiae. The system recapitulates regulated bidirectional origin activation; synthesis of leading and lagging strands by the three replicative DNA polymerases Pol α, Pol δ, and Pol ε; and canonical maturation of Okazaki fragments into continuous daughter strands. We uncover a dual regulatory role for chromatin during DNA replication: promoting origin dependence and determining Okazaki fragment length by restricting Pol δ progression. This system thus provides a functional platform for the detailed mechanistic analysis of eukaryotic chromosome replication. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Modelling of capital asset pricing by considering the lagged effects

    NASA Astrophysics Data System (ADS)

    Sukono; Hidayat, Y.; Bon, A. Talib bin; Supian, S.

    2017-01-01

    In this paper the problem of modelling the Capital Asset Pricing Model (CAPM) with the effect of the lagged is discussed. It is assumed that asset returns are analysed influenced by the market return and the return of risk-free assets. To analyse the relationship between asset returns, the market return, and the return of risk-free assets, it is conducted by using a regression equation of CAPM, and regression equation of lagged distributed CAPM. Associated with the regression equation lagged CAPM distributed, this paper also developed a regression equation of Koyck transformation CAPM. Results of development show that the regression equation of Koyck transformation CAPM has advantages, namely simple as it only requires three parameters, compared with regression equation of lagged distributed CAPM.

  10. DNA synthesis arrest sites at the right terminus of rat long interspersed repeated (LINE or L1Rn) DNA family members.

    PubMed Central

    d'Ambrosio, E; Furano, A V

    1987-01-01

    An approximately equal to 150-bp GC-rich (approximately equal to 60%) region is at the right end of rat long interspersed repeated DNA (LINE or L1Rn) family members. We report here that one of the DNA strands from this region contains several non-palindromic sites that strongly arrest DNA synthesis in vitro by the prokaryotic Klenow and T4 DNA polymerases, the eukaryotic alpha polymerase, and AMV reverse transcriptase. The strongest arrest sites are G-rich (approximately equal to 70%) homopurine stretches of 18 or more residues. Shorter homopurine stretches (12 residues or fewer) did not arrest DNA synthesis even if the stretch contains 11/12 G residues. Arrest of the prokaryotic polymerases was not affected by their respective single strand binding proteins or polymerase accessory proteins. The region of duplex DNA which contains DNA synthesis arrest sites reacts with bromoacetaldehyde when present in negatively supercoiled molecules. By contrast, homopurine stretches that do not arrest DNA synthesis do not react with bromoacetaldehyde. The presence of bromoacetaldehyde-reactive bases in a G-rich homopurine-containing duplex under torsional stress is thought to be caused by base stacking in the homopurine strand. Therefore, we suggest that base-stacked regions of the template arrest DNA synthesis. Images PMID:2436148

  11. Unraveling the strands of Saturn's F ring

    USGS Publications Warehouse

    Murray, C.D.; Gordon, M.K.; Giuliatti, Winter S.M.

    1997-01-01

    Several high-resolution Voyager 2 images of Saturn's F ring show that it is composed of at least four separate, non-intersecting strands extending ~45?? in longitude. Voyager 1 images show that the two brightest strands appear to intersect, giving rise to a "braided" morphology. From a study of all available Voyager images the detectable radial structure is cataloged and reviewed. Previous indications that there is fine material interior to the orbit of the F ring are confirmed. Evidence is presented that a model of four strands with comparable eccentricities and nearly aligned perichrones is consistent with all the Voyager observations. The observed perichrone offset of the two brightest strands suggests a minimum radial separation of ~20 km, which implies intersection of these strands when their finite radial widths are taken into account. The longitude range of such an intersection includes that observed in the Voyager 1 "braid" images. The proximity of these two strands at some longitudes may account for the apparent differences in the ring between the Voyager encounters, as well as provide a source for the short-lived features detected in the Hubble Space Telescope images of the F ring. There is no evidence that the locations of the individual strands are determined by resonant perturbations with known satellites. It is proposed that the radial structure is formed by the localized action of small satellites orbiting within the strand region. ?? 1997 Academic Press.

  12. Fish stranding in freshwater systems: sources, consequences, and mitigation.

    PubMed

    Nagrodski, Alexander; Raby, Graham D; Hasler, Caleb T; Taylor, Mark K; Cooke, Steven J

    2012-07-30

    Fish can become stranded when water levels decrease, often rapidly, as a result of anthropogenic (e.g., canal drawdown, hydropeaking, vessel wakes) and natural (e.g., floods, drought, winter ice dynamics) events. We summarize existing research on stranding of fish in freshwater, discuss the sources, consequences, and mitigation options for stranding, and report current knowledge gaps. Our literature review revealed that ∼65.5% of relevant peer-reviewed articles were found to focus on stranding associated with hydropower operations and irrigation projects. In fact, anthropogenic sources of fish stranding represented 81.8% of available literature compared to only 19.9% attributed to natural fish stranding events. While fish mortality as a result of stranding is well documented, our analysis revealed that little is known about the sublethal and long-term consequences of stranding on growth and population dynamics. Furthermore, the contribution of stranding to annual mortality rates is poorly understood as are the potential ecosystem-scale impacts. Mitigation strategies available to deal with stranding include fish salvage, ramping rate limitations, and physical habitat works (e.g., to contour substrate to minimize stranding). However, a greater knowledge of the factors that cause fish stranding would promote the development and refinement of mitigation strategies that are economically and ecologically sustainable. Copyright © 2012 Elsevier Ltd. All rights reserved.

  13. Strand-specific, real-time RT-PCR assays for quantification of genomic and positive-sense RNAs of the fish rhabdovirus, Infectious hematopoietic necrosis virus

    USGS Publications Warehouse

    Purcell, Maureen K.; Hart, S. Alexandra; Kurath, Gael; Winton, James R.

    2006-01-01

    The fish rhabdovirus, Infectious hematopoietic necrosis virus (IHNV), is an important pathogen of salmonids. Cell culture assays have traditionally been used to quantify levels of IHNV in samples; however, real-time or quantitative RT-PCR assays have been proposed as a rapid alternative. For viruses having a single-stranded, negative-sense RNA genome, standard qRT-PCR assays do not distinguish between the negative-sense genome and positive-sense RNA species including mRNA and anti-genome. Thus, these methods do not determine viral genome copy number. This study reports development of strand-specific, qRT-PCR assays that use tagged primers for enhancing strand specificity during cDNA synthesis and quantitative PCR. Protocols were developed for positive-strand specific (pss-qRT-PCR) and negative-strand specific (nss-qRT-PCR) assays for IHNV glycoprotein (G) gene sequences. Validation with synthetic RNA transcripts demonstrated the assays could discriminate the correct strand with greater than 1000-fold fidelity. The number of genome copies in livers of IHNV-infected fish determined by nss-qRT-PCR was, on average, 8000-fold greater than the number of infectious units as determined by plaque assay. We also compared the number of genome copies with the quantity of positive-sense RNA and determined that the ratio of positive-sense molecules to negative-sense genome copies was, on average, 2.7:1. Potential future applications of these IHNV strand-specific qRT-PCR assays are discussed.

  14. Time-lag effects of global vegetation responses to climate change.

    PubMed

    Wu, Donghai; Zhao, Xiang; Liang, Shunlin; Zhou, Tao; Huang, Kaicheng; Tang, Bijian; Zhao, Wenqian

    2015-09-01

    Climate conditions significantly affect vegetation growth in terrestrial ecosystems. Due to the spatial heterogeneity of ecosystems, the vegetation responses to climate vary considerably with the diverse spatial patterns and the time-lag effects, which are the most important mechanism of climate-vegetation interactive effects. Extensive studies focused on large-scale vegetation-climate interactions use the simultaneous meteorological and vegetation indicators to develop models; however, the time-lag effects are less considered, which tends to increase uncertainty. In this study, we aim to quantitatively determine the time-lag effects of global vegetation responses to different climatic factors using the GIMMS3g NDVI time series and the CRU temperature, precipitation, and solar radiation datasets. First, this study analyzed the time-lag effects of global vegetation responses to different climatic factors. Then, a multiple linear regression model and partial correlation model were established to statistically analyze the roles of different climatic factors on vegetation responses, from which the primary climate-driving factors for different vegetation types were determined. The results showed that (i) both the time-lag effects of the vegetation responses and the major climate-driving factors that significantly affect vegetation growth varied significantly at the global scale, which was related to the diverse vegetation and climate characteristics; (ii) regarding the time-lag effects, the climatic factors explained 64% variation of the global vegetation growth, which was 11% relatively higher than the model ignoring the time-lag effects; (iii) for the area with a significant change trend (for the period 1982-2008) in the global GIMMS3g NDVI (P < 0.05), the primary driving factor was temperature; and (iv) at the regional scale, the variation in vegetation growth was also related to human activities and natural disturbances. Considering the time-lag effects is quite

  15. Lambda Red Mediated Gap Repair Utilizes a Novel Replicative Intermediate in Escherichia coli

    PubMed Central

    Reddy, Thimma R.; Fevat, Léna M. S.; Munson, Sarah E.; Stewart, A. Francis; Cowley, Shaun M.

    2015-01-01

    The lambda phage Red recombination system can mediate efficient homologous recombination in Escherichia coli, which is the basis of the DNA engineering technique termed recombineering. Red mediated insertion of DNA requires DNA replication, involves a single-stranded DNA intermediate and is more efficient on the lagging strand of the replication fork. Lagging strand recombination has also been postulated to explain the Red mediated repair of gapped plasmids by an Okazaki fragment gap filling model. Here, we demonstrate that gap repair involves a different strand independent mechanism. Gap repair assays examining the strand asymmetry of recombination did not show a lagging strand bias. Directly testing an ssDNA plasmid showed lagging strand recombination is possible but dsDNA plasmids did not employ this mechanism. Insertional recombination combined with gap repair also did not demonstrate preferential lagging strand bias, supporting a different gap repair mechanism. The predominant recombination route involved concerted insertion and subcloning though other routes also operated at lower frequencies. Simultaneous insertion of DNA resulted in modification of both strands and was unaffected by mutations to DNA polymerase I, responsible for Okazaki fragment maturation. The lower efficiency of an alternate Red mediated ends-in recombination pathway and the apparent lack of a Holliday junction intermediate suggested that gap repair does not involve a different Red recombination pathway. Our results may be explained by a novel replicative intermediate in gap repair that does not involve a replication fork. We exploited these observations by developing a new recombineering application based on concerted insertion and gap repair, termed SPI (subcloning plus insertion). SPI selected against empty vector background and selected for correct gap repair recombinants. We used SPI to simultaneously insert up to four different gene cassettes in a single recombineering reaction

  16. Improving strand pairing prediction through exploring folding cooperativity

    PubMed Central

    Jeong, Jieun; Berman, Piotr; Przytycka, Teresa M.

    2008-01-01

    The topology of β-sheets is defined by the pattern of hydrogen-bonded strand pairing. Therefore, predicting hydrogen bonded strand partners is a fundamental step towards predicting β-sheet topology. At the same time, finding the correct partners is very difficult due to long range interactions involved in strand pairing. Additionally, patterns of aminoacids observed in β-sheet formations are very general and therefore difficult to use for computational recognition of specific contacts between strands. In this work, we report a new strand pairing algorithm. To address above mentioned difficulties, our algorithm attempts to mimic elements of the folding process. Namely, in addition to ensuring that the predicted hydrogen bonded strand pairs satisfy basic global consistency constraints, it takes into account hypothetical folding pathways. Consistently with this view, introducing hydrogen bonds between a pair of strands changes the probabilities of forming hydrogen bonds between other pairs of strand. We demonstrate that this approach provides an improvement over previously proposed algorithms. We also compare the performance of this method to that of a global optimization algorithm that poses the problem as integer linear programming optimization problem and solves it using ILOG CPLEX™ package. PMID:18989036

  17. Understanding DNA replication by the bacteriophage T4 replisome.

    PubMed

    Benkovic, Stephen J; Spiering, Michelle M

    2017-11-10

    The T4 replisome has provided a unique opportunity to investigate the intricacies of DNA replication. We present a comprehensive review of this system focusing on the following: its 8-protein composition, their individual and synergistic activities, and assembly in vitro and in vivo into a replisome capable of coordinated leading/lagging strand DNA synthesis. We conclude with a brief comparison with other replisomes with emphasis on how coordinated DNA replication is achieved. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. SYNTHETIC STRANDS OF CARDIAC MUSCLE

    PubMed Central

    Purdy, Joyce E.; Lieberman, Melvyn; Roggeveen, Anne E.; Kirk, R. Gary

    1972-01-01

    Spontaneously active bundles of cardiac muscle (synthetic strands) were prepared from isolated cells of 11–13-day old embryonic chick hearts which were disaggregated with trypsin. Linear orientation of the cells was obtained by plating them on agar-coated culture dishes in which either grooves were cut in the agar film or a thin line of palladium was deposited over the agar. The influence of cell-to-cell and cell-to-substrate interactions was observed with time lapse cinematography and the formation of the synthetic strand was shown to involve both random and guided cell movements, enlargement of aggregates by accretion and coalescence, and the compact linear arrangement of cells along paths of preferential adhesion. Electron microscope investigations of these strands showed that a dispersed population of heart cells organized into an inner core of muscle cells and an outer sheath of fibroblast-like cells. The muscle cells contained well-developed, but widely spaced myofibrils, a developing sarcoplasmic reticulum associated in part with the myofibrils and in part with the sarcolemma, an abundance of nonmembrane bound ribosomes and glycogen, and a prominent Golgi complex. Numerous specialized contacts were observed between the muscle cells in the strand, e.g., fasciae adherentes, desmosomes, and nexuses. A distinct type of muscle cell characterized by its pale appearance was regularly observed in the strand and was noted to be similar to Purkinje cells described in the adult avian conduction system and in developing chick myocardium. The present findings were compared with other observations of the developing myocardium, in situ, and it was concluded that, by a number or criteria, the muscle cells of the strand were differentiating normally and suitably organized for electrophysiological studies. PMID:4656702

  19. How jet lag impairs Major League Baseball performance

    PubMed Central

    Song, Alex; Severini, Thomas; Allada, Ravi

    2017-01-01

    Laboratory studies have demonstrated that circadian clocks align physiology and behavior to 24-h environmental cycles. Examination of athletic performance has been used to discern the functions of these clocks in humans outside of controlled settings. Here, we examined the effects of jet lag, that is, travel that shifts the alignment of 24-h environmental cycles relative to the endogenous circadian clock, on specific performance metrics in Major League Baseball. Accounting for potential differences in home and away performance, travel direction, and team confounding variables, we observed that jet-lag effects were largely evident after eastward travel with very limited effects after westward travel, consistent with the >24-h period length of the human circadian clock. Surprisingly, we found that jet lag impaired major parameters of home-team offensive performance, for example, slugging percentage, but did not similarly affect away-team offensive performance. On the other hand, jet lag impacted both home and away defensive performance. Remarkably, the vast majority of these effects for both home and away teams could be explained by a single measure, home runs allowed. Rather than uniform effects, these results reveal surprisingly specific effects of circadian misalignment on athletic performance under natural conditions. PMID:28115724

  20. LAG-3 confers a competitive disadvantage upon antiviral CD8+ T cell responses1

    PubMed Central

    Cook, Kevin D.; Whitmire, Jason K.

    2016-01-01

    Ongoing clinical trials are evaluating the benefits of systemic blockade of lymphocyte activation gene-3 (LAG-3) signals to improve immunity to tumors. Those studies are founded on the well-established inhibitory role of LAG-3 in regulating CD8+ T cells during chronic virus infection and anti-tumor responses. However, the T cell response in LAG-3 deficient mice is similar in size and function to that in wild type animals, suggesting LAG-3 has nuanced immune-regulatory functions. We performed a series of adoptive transfer experiments in mice to better understand the T cell-intrinsic functions of LAG-3 in the regulation of CD8+ T cell responses. Our results indicate that LAG-3 expression by CD8+ T cells inhibits their competitive fitness and results in a slightly reduced rate of cell division in comparison to LAG-3 deficient cells. This cell-intrinsic effect of LAG-3 was consistent across both acute and chronic virus infections. These data show that LAG-3 directly modulates the size of the T cell response and support the use of LAG-3 blockade regimens to enhance CD8+ T cell responses. PMID:27206765

  1. A Race Against Time: Time Lags in Terrestrial-Aquatic Linkages

    NASA Astrophysics Data System (ADS)

    Basu, N. B.

    2017-12-01

    Unprecedented decreases in atmospheric nitrogen (N) deposition together with increases in agricultural N-use efficiency have led to decreases in net anthropogenic N inputs in many eastern U.S. and Canadian watersheds as well as in Europe. Despite such decreases, N concentrations in streams and rivers continue to increase, and problems of coastal eutrophication remain acute. Such a mismatch between N inputs and outputs can arise due to legacy N accumulation and subsequent lag times between implementation of conservation measures and improvements in water quality. In the present study, we quantified such lag times by pairing long-term N input trajectories with stream N concentration data in multiple watersheds in North America. Results show significant nonlinearity between N inputs and outputs, with a strong hysteresis effect indicative of decadal-scale lag times. Lag times were found to be negatively correlated with both tile drainage and watershed slope, with tile drainage being a dominant control in fall and watershed slope being significant during the spring snowmelt period. Quantification of such lags will be crucial to policy-makers as they struggle to set appropriate goals for water quality improvement in human-impacted watersheds.

  2. When Long-Range Zero-Lag Synchronization is Feasible in Cortical Networks

    PubMed Central

    Viriyopase, Atthaphon; Bojak, Ingo; Zeitler, Magteld; Gielen, Stan

    2012-01-01

    Many studies have reported long-range synchronization of neuronal activity between brain areas, in particular in the beta and gamma bands with frequencies in the range of 14–30 and 40–80 Hz, respectively. Several studies have reported synchrony with zero phase lag, which is remarkable considering the synaptic and conduction delays inherent in the connections between distant brain areas. This result has led to many speculations about the possible functional role of zero-lag synchrony, such as for neuronal communication, attention, memory, and feature binding. However, recent studies using recordings of single-unit activity and local field potentials report that neuronal synchronization may occur with non-zero phase lags. This raises the questions whether zero-lag synchrony can occur in the brain and, if so, under which conditions. We used analytical methods and computer simulations to investigate which connectivity between neuronal populations allows or prohibits zero-lag synchrony. We did so for a model where two oscillators interact via a relay oscillator. Analytical results and computer simulations were obtained for both type I Mirollo–Strogatz neurons and type II Hodgkin–Huxley neurons. We have investigated the dynamics of the model for various types of synaptic coupling and importantly considered the potential impact of Spike-Timing Dependent Plasticity (STDP) and its learning window. We confirm previous results that zero-lag synchrony can be achieved in this configuration. This is much easier to achieve with Hodgkin–Huxley neurons, which have a biphasic phase response curve, than for type I neurons. STDP facilitates zero-lag synchrony as it adjusts the synaptic strengths such that zero-lag synchrony is feasible for a much larger range of parameters than without STDP. PMID:22866034

  3. Collective Dynamics of Oscillator Networks: Why do we suffer from heavy jet lag?

    NASA Astrophysics Data System (ADS)

    Kori, Hiroshi

    The circadian rhythm of the entire body in mammals is orchestrated by a small tissue in the brain called the suprachiamatic nucleus (SCN). The SCN consists of a population of neurons, each of which exhibit circadian (i.e., approximately 24 h) gene expression. Neurons form a complex network and interact with each other using various types of neurotransmitters. The rhythmic gene expressions of individual cells in the SCN synchronize through such interaction. Jet-lag symptoms arise from temporal mismatch between the internal circadian clock orchestrated by the SCN and external solar time. It may take about one week or even longer to recover from jet lag after a long-distance trip. We recently found that recovery from jet lag is considerably accelerated in the knocked-out (KO) mice lacking the receptors of a certain neurotransmitter in the SCN. Importantly, all other properties of mice including sleep-awake rhythms and breeding seem to be intact. Only the response to the jet lag changes. It was also found that after a few days of jet lag, cells in the SCN desynchronize in the wild type (WT) mice, whereas they do not in KO mice. This desynchrony might be a main reason for heavy jet lag symptoms. To understand the mechanism underlying jet lag, we propose a simple model of the SCN, which is a network of phase oscillators. Despite its simplicity, this model can reproduce important dynamical properties of the SCN. For example, this model reproduces the desynchrony of oscillators after jet lag. Moreover, when intercellular interaction is weaker, this desynchrony is suppressed and the recover from jet lag is considerably accelerated. Our mathematical study provides a deeper understanding of jet lag and an idea how to circumvent heavy jet lag symptoms

  4. Dynamic DNA nanotechnology using strand-displacement reactions

    NASA Astrophysics Data System (ADS)

    Zhang, David Yu; Seelig, Georg

    2011-02-01

    The specificity and predictability of Watson-Crick base pairing make DNA a powerful and versatile material for engineering at the nanoscale. This has enabled the construction of a diverse and rapidly growing set of DNA nanostructures and nanodevices through the programmed hybridization of complementary strands. Although it had initially focused on the self-assembly of static structures, DNA nanotechnology is now also becoming increasingly attractive for engineering systems with interesting dynamic properties. Various devices, including circuits, catalytic amplifiers, autonomous molecular motors and reconfigurable nanostructures, have recently been rationally designed to use DNA strand-displacement reactions, in which two strands with partial or full complementarity hybridize, displacing in the process one or more pre-hybridized strands. This mechanism allows for the kinetic control of reaction pathways. Here, we review DNA strand-displacement-based devices, and look at how this relatively simple mechanism can lead to a surprising diversity of dynamic behaviour.

  5. Heat transfer characteristics of an emergent strand

    NASA Technical Reports Server (NTRS)

    Simon, W. E.; Witte, L. C.; Hedgcoxe, P. G.

    1974-01-01

    A mathematical model was developed to describe the heat transfer characteristics of a hot strand emerging into a surrounding coolant. A stable strand of constant efflux velocity is analyzed, with a constant (average) heat transfer coefficient on the sides and leading surface of the strand. After developing a suitable governing equation to provide an adequate description of the physical system, the dimensionless governing equation is solved with Laplace transform methods. The solution yields the temperature within the strand as a function of axial distance and time. Generalized results for a wide range of parameters are presented, and the relationship of the results and experimental observations is discussed.

  6. Comparison of specific binding sites for Escherichia coli RNA polymerase with naturally occurring hairpin regions in single-stranded DNA of coliphage M13. [Aspergillus oryzae

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

    Niyogi, S.K.; Mitra, S.

    Escherichia coli RNA polymerase binds specifically to the single-stranded circular DNA of coliphage M13 in the presence of a saturating concentration of the bacterial DNA binding protein presumably as an essential step in the synthesis of the RNA primer required for synthesizing the complementary DNA strand in parental replicative-form DNA. The RNA polymerase-protected DNA regions were isolated after extensive digestion with pancreatic DNase, S1 endonuclease of Aspergillus oryzae, and exonuclease I of E. coli. The physicochemical properties of the RNA polymerase-protected segments (called PI and PII) were compared with those of the naturally occurring hairpin regions.

  7. G-quadruplexes Significantly Stimulate Pif1 Helicase-catalyzed Duplex DNA Unwinding*

    PubMed Central

    Duan, Xiao-Lei; Liu, Na-Nv; Yang, Yan-Tao; Li, Hai-Hong; Li, Ming; Dou, Shuo-Xing; Xi, Xu-Guang

    2015-01-01

    The evolutionarily conserved G-quadruplexes (G4s) are faithfully inherited and serve a variety of cellular functions such as telomere maintenance, gene regulation, DNA replication initiation, and epigenetic regulation. Different from the Watson-Crick base-pairing found in duplex DNA, G4s are formed via Hoogsteen base pairing and are very stable and compact DNA structures. Failure of untangling them in the cell impedes DNA-based transactions and leads to genome instability. Cells have evolved highly specific helicases to resolve G4 structures. We used a recombinant nuclear form of Saccharomyces cerevisiae Pif1 to characterize Pif1-mediated DNA unwinding with a substrate mimicking an ongoing lagging strand synthesis stalled by G4s, which resembles a replication origin and a G4-structured flap in Okazaki fragment maturation. We find that the presence of G4 may greatly stimulate the Pif1 helicase to unwind duplex DNA. Further studies reveal that this stimulation results from G4-enhanced Pif1 dimerization, which is required for duplex DNA unwinding. This finding provides new insights into the properties and functions of G4s. We discuss the observed activation phenomenon in relation to the possible regulatory role of G4s in the rapid rescue of the stalled lagging strand synthesis by helping the replicator recognize and activate the replication origin as well as by quickly removing the G4-structured flap during Okazaki fragment maturation. PMID:25627683

  8. Lags in the response of mountain plant communities to climate change

    PubMed Central

    Alexander, Jake M.; Chalmandrier, Loïc; Lenoir, Jonathan; Burgess, Treena I.; Essl, Franz; Haider, Sylvia; Kueffer, Christoph; McDougall, Keith; Milbau, Ann; Nuñez, Martin A.; Pauchard, Aníbal; Rabitsch, Wolfgang; Rew, Lisa J.; Sanders, Nathan J.; Pellissier, Loïc

    2018-01-01

    Rapid climatic changes and increasing human influence at high elevations around the world will have profound impacts on mountain biodiversity. However, forecasts from statistical models (e.g. species distribution models) rarely consider that plant community changes could substantially lag behind climatic changes, hindering our ability to make temporally realistic projections for the coming century. Indeed, the magnitudes of lags, and the relative importance of the different factors giving rise to them, remain poorly understood. We review evidence for three types of lag: “dispersal lags” affecting plant species’ spread along elevational gradients, “establishment lags” following their arrival in recipient communities, and “extinction lags” of resident species. Variation in lags is explained by variation among species in physiological and demographic responses, by effects of altered biotic interactions, and by aspects of the physical environment. Of these, altered biotic interactions could contribute substantially to establishment and extinction lags, yet impacts of biotic interactions on range dynamics are poorly understood. We develop a mechanistic community model to illustrate how species turnover in future communities might lag behind simple expectations based on species’ range shifts with unlimited dispersal. The model shows a combined contribution of altered biotic interactions and dispersal lags to plant community turnover along an elevational gradient following climate warming. Our review and simulation support the view that accounting for disequilibrium range dynamics will be essential for realistic forecasts of patterns of biodiversity under climate change, with implications for the conservation of mountain species and the ecosystem functions they provide. PMID:29112781

  9. Method for producing labeled single-stranded nucleic acid probes

    DOEpatents

    Dunn, John J.; Quesada, Mark A.; Randesi, Matthew

    1999-10-19

    Disclosed is a method for the introduction of unidirectional deletions in a cloned DNA segment. More specifically, the method comprises providing a recombinant DNA construct comprising a DNA segment of interest inserted in a cloning vector, the cloning vector having an f1 endonuclease recognition sequence adjacent to the insertion site of the DNA segment of interest. The recombinant DNA construct is then contacted with the protein pII encoded by gene II of phage f1 thereby generating a single-stranded nick. The nicked DNA is then contacted with E. coli Exonuclease III thereby expanding the single-stranded nick into a single-stranded gap. The single-stranded gapped DNA is then contacted with a single-strand-specific endonuclease thereby producing a linearized DNA molecule containing a double-stranded deletion corresponding in size to the single-stranded gap. The DNA treated in this manner is then incubated with DNA ligase under conditions appropriate for ligation. Also disclosed is a method for producing single-stranded DNA probes. In this embodiment, single-stranded gapped DNA, produced as described above, is contacted with a DNA polymerase in the presence of labeled nucleotides to fill in the gap. This DNA is then linearized by digestion with a restriction enzyme which cuts outside the DNA segment of interest. The product of this digestion is then denatured to produce a labeled single-stranded nucleic acid probe.

  10. Virus-Specific RNA Synthesis in Cells Infected by Infectious Pancreatic Necrosis Virus

    PubMed Central

    Somogyi, Paul; Dobos, Peter

    1980-01-01

    Pulse-labeling experiments with [3H]uridine revealed that the rate of infections pancreatic necrosis virus-specific RNA synthesis was maximal at 8 to 10 h after infection and was completely diminished by 12 to 14 h. Three forms of RNA intermediates were detected: (i) a putative transcription intermediate (TRI) which comigrated in acrylamide gels with virion double-stranded RNA (dsRNA) after RNase treatment; (ii) a 24S genome length mRNA which could be resolved into two bands by polyacrylamide gel electrophoresis; and (iii) a 14S dsRNA component indistinguishable from virion RNA by gradient centrifugation and gel electrophoresis. The TRI (i) was LiCl precipitable; (ii) sedimented slightly faster and broader (14 to 16S) than the 14S virion dsRNA; (iii) had a lower electrophoretic mobility in acrylamide gels than dsRNA, barely entering acrylamide gels as a heterogenous component; (iv) yielded genome-sized pieces of dsRNA after RNase digestion; and (v) was the most abundant RNA form early in the infectious cycle. The 24S single-stranded RNA was thought to be the viral mRNA since it: (i) became labeled during short pulses; (ii) was found in the polysomal fraction of infected cells; and (iii) hybridized to denatured viral RNA, forming two segments of RNase-resistant RNA that comigrated with virion dsRNA in gels. The 24S mRNA component was formed before the synthesis of dsRNA, and radioactivity could be chased from 24S single-stranded RNA to dsRNA, indicating that 24S RNA may serve as template for the synthesis of complementary strands to form dsRNA. Similar to reovirus, infectious pancreatic necrosis viral 24S mRNA contained no polyadenylic acid tracts. Images PMID:16789184

  11. Role of Escherichia coli dnaA gene and its integrative suppression in M13 Coliphage DNA synthesis

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

    Mitra, S.; Stallions, D.R.

    An F/sup +/ derivative of Escherichia coli E508 thermosensitive in dnaA function (involved in DNA synthesis initiation), its revertant and an Hfr derivative of E508(ts) in which the temperature-sensitive phenotype is suppressed by integrative suppression have been compared for their ability to support M13 phage DNA synthesis at the nonpermissive temperature. Upon infection at the nonpermissive temperature, both the revertant and the Hfr strain support normal phage replication while the temperature-sensitive mutant does not. However, when infection is carried out at a permissive temperature and the temperature is shifted up after infection, phage synthesis occurs in the temperature-sensitive mutant also,more » but in lesser quantity than in the revertant strain. Analysis of intracellular labeled phage DNA indicates: (a) parental replicative form DNA synthesis is not dependent on dnaA function; (b) progeny replicative form DNA synthesis is strongly inhibited in the temperature-sensitive dnaA mutant at the nonpermissive temperature; (c) progeny single-strand DNA synthesis does not absolutely require dnaA function; (d) progeny single-strand DNA is present in the circular form. The implication of the host DNA replication in M13 DNA synthesis is discussed.« less

  12. The Synthesis of Quinolone Natural Products from Pseudonocardia sp.

    PubMed Central

    Salvaggio, Flavia; Hodgkinson, James T.; Carro, Laura; Geddis, Stephen M.; Galloway, Warren R. J. D.; Welch, Martin

    2015-01-01

    Abstract The synthesis of four quinolone natural products from the actinomycete Pseudonocardia sp. is reported. The key step involved a sp2–sp3 Suzuki–Miyaura reaction between a common boronic ester lateral chain and various functionalised quinolone cores. The quinolones slowed growth of E. coli and S. aureus by inducing extended lag phases.

  13. Effective Iterated Greedy Algorithm for Flow-Shop Scheduling Problems with Time lags

    NASA Astrophysics Data System (ADS)

    ZHAO, Ning; YE, Song; LI, Kaidian; CHEN, Siyu

    2017-05-01

    Flow shop scheduling problem with time lags is a practical scheduling problem and attracts many studies. Permutation problem(PFSP with time lags) is concentrated but non-permutation problem(non-PFSP with time lags) seems to be neglected. With the aim to minimize the makespan and satisfy time lag constraints, efficient algorithms corresponding to PFSP and non-PFSP problems are proposed, which consist of iterated greedy algorithm for permutation(IGTLP) and iterated greedy algorithm for non-permutation (IGTLNP). The proposed algorithms are verified using well-known simple and complex instances of permutation and non-permutation problems with various time lag ranges. The permutation results indicate that the proposed IGTLP can reach near optimal solution within nearly 11% computational time of traditional GA approach. The non-permutation results indicate that the proposed IG can reach nearly same solution within less than 1% computational time compared with traditional GA approach. The proposed research combines PFSP and non-PFSP together with minimal and maximal time lag consideration, which provides an interesting viewpoint for industrial implementation.

  14. Drug Lag and Key Regulatory Barriers in the Emerging Markets

    PubMed Central

    Wileman, Harriet; Mishra, Arun

    2010-01-01

    There have been numerous investigations targeted at identifying whether a drug lag exists in the mature markets of the US, EU and Japan. This work focuses on the emerging markets because of the potential they hold for the future of the pharmaceutical industry as a consequence of rapid economic and political development. The aims of this work are to ascertain whether a drug lag exists in the emerging markets and how it has changed over time from the 1960s to the 2000s. It will also highlight key regulatory barriers which may contribute to drug lag. The date of the marketing authorisation (MA) approval by the US Food and Drug Administration (FDA) was used as a reference point. A comparison against the company database regarding emerging market specific approval enabled the difference in time and thus the drug lag for that particular market to be calculated. This work concludes that the overall relative drug lag in the emerging markets has decreased over time and that there are seven key regulatory barriers which need to be targeted in order to make further improvements; ‘Western Approval’, local clinical development (LCD), Certificate of Pharmaceutical Product (CPP), Good Manufacturing Practice (GMP), pricing approval, document authentication and harmonisation. PMID:21829782

  15. Hearing Loss in Stranded Odontocete Dolphins and Whales

    PubMed Central

    Mann, David; Hill-Cook, Mandy; Manire, Charles; Greenhow, Danielle; Montie, Eric; Powell, Jessica; Wells, Randall; Bauer, Gordon; Cunningham-Smith, Petra; Lingenfelser, Robert; DiGiovanni, Robert; Stone, Abigale; Brodsky, Micah; Stevens, Robert; Kieffer, George; Hoetjes, Paul

    2010-01-01

    The causes of dolphin and whale stranding can often be difficult to determine. Because toothed whales rely on echolocation for orientation and feeding, hearing deficits could lead to stranding. We report on the results of auditory evoked potential measurements from eight species of odontocete cetaceans that were found stranded or severely entangled in fishing gear during the period 2004 through 2009. Approximately 57% of the bottlenose dolphins and 36% of the rough-toothed dolphins had significant hearing deficits with a reduction in sensitivity equivalent to severe (70–90 dB) or profound (>90 dB) hearing loss in humans. The only stranded short-finned pilot whale examined had profound hearing loss. No impairments were detected in seven Risso's dolphins from three different stranding events, two pygmy killer whales, one Atlantic spotted dolphin, one spinner dolphin, or a juvenile Gervais' beaked whale. Hearing impairment could play a significant role in some cetacean stranding events, and the hearing of all cetaceans in rehabilitation should be tested. PMID:21072206

  16. Hearing loss in stranded odontocete dolphins and whales.

    PubMed

    Mann, David; Hill-Cook, Mandy; Manire, Charles; Greenhow, Danielle; Montie, Eric; Powell, Jessica; Wells, Randall; Bauer, Gordon; Cunningham-Smith, Petra; Lingenfelser, Robert; DiGiovanni, Robert; Stone, Abigale; Brodsky, Micah; Stevens, Robert; Kieffer, George; Hoetjes, Paul

    2010-11-03

    The causes of dolphin and whale stranding can often be difficult to determine. Because toothed whales rely on echolocation for orientation and feeding, hearing deficits could lead to stranding. We report on the results of auditory evoked potential measurements from eight species of odontocete cetaceans that were found stranded or severely entangled in fishing gear during the period 2004 through 2009. Approximately 57% of the bottlenose dolphins and 36% of the rough-toothed dolphins had significant hearing deficits with a reduction in sensitivity equivalent to severe (70-90 dB) or profound (>90 dB) hearing loss in humans. The only stranded short-finned pilot whale examined had profound hearing loss. No impairments were detected in seven Risso's dolphins from three different stranding events, two pygmy killer whales, one Atlantic spotted dolphin, one spinner dolphin, or a juvenile Gervais' beaked whale. Hearing impairment could play a significant role in some cetacean stranding events, and the hearing of all cetaceans in rehabilitation should be tested.

  17. Method and apparatus for testing a forward-moving strand

    DOEpatents

    Ducommun, Joel; Vulliens, Philippe

    1980-01-01

    In a method for testing a continuously forward-moving strand a light beam which passes along a plane that extends approximately perpendicularly to the longitudinal axis of the strand is introduced into the strand. The brightness value is measured on a place of the strand exterior which is distal from the light incidence place by means of at least one photoelectronic element disposed directly on the strand exterior and the measured result is evaluated in a gating circuit which is electrically connected to the photoelectronic element.

  18. Flap-Lag-Torsion Stability in Forward Flight

    NASA Technical Reports Server (NTRS)

    Panda, B.; Chopra, I.

    1985-01-01

    An aeroelastic stability of three-degree flap-lag-torsion blade in forward flight is examined. Quasisteady aerodynamics with a dynamic inflow model is used. The nonlinear time dependent periodic blade response is calculated using an iterative procedure based on Floquet theory. The periodic perturbation equations are solved for stability using Floquet transition matrix theory as well as constant coefficient approximation in the fixed reference frame. Results are presented for both stiff-inplane and soft-inplane blade configurations. The effects of several parameters on blade stability are examined, including structural coupling, pitch-flap and pitch-lag coupling, torsion stiffness, steady inflow distribution, dynamic inflow, blade response solution and constant coefficient approximation.

  19. Fungal mycelia show lag time before re-growth on endogenous carbon.

    PubMed

    Pollack, Judith K; Li, Zheng Jian; Marten, Mark R

    2008-06-15

    Nutrient starvation is a common occurrence for filamentous fungi. To better understand the effects of starvation, we used a parallel plate flow chamber to study individual fungal mycelia when subjected to a step change in glucose concentration. We report the presence of a finite "lag time" in starved mycelia during which they ceased to grow/extend while switching from growth on exogenous carbon to re-growth on endogenous carbon. This lag time precedes other morphological or physiological changes such as change in growth rate (50-70% reduction), vacuolation (up to 16%), and decreased hyphal diameter (almost 50% reduction). Data suggests that during lag time, vacuolar degradation produces sufficient endogenous carbon to support survival and restart hyphal extension. Lag time is inversely related to the size of the mycelium at the time of starvation, which suggests a critical flow of endogenous carbon to the apical tip. We present a mathematical model consistent with our experimental observations that relate lag time, area, and flow of endogenous carbon. (c) 2008 Wiley Periodicals, Inc.

  20. Information-theoretic approach to lead-lag effect on financial markets

    NASA Astrophysics Data System (ADS)

    Fiedor, Paweł

    2014-08-01

    Recently the interest of researchers has shifted from the analysis of synchronous relationships of financial instruments to the analysis of more meaningful asynchronous relationships. Both types of analysis are concentrated mostly on Pearson's correlation coefficient and consequently intraday lead-lag relationships (where one of the variables in a pair is time-lagged) are also associated with them. Under the Efficient-Market Hypothesis such relationships are not possible as all information is embedded in the prices, but in real markets we find such dependencies. In this paper we analyse lead-lag relationships of financial instruments and extend known methodology by using mutual information instead of Pearson's correlation coefficient. Mutual information is not only a more general measure, sensitive to non-linear dependencies, but also can lead to a simpler procedure of statistical validation of links between financial instruments. We analyse lagged relationships using New York Stock Exchange 100 data not only on an intraday level, but also for daily stock returns, which have usually been ignored.

  1. Clinical introduction of image lag correction for a cone beam CT system.

    PubMed

    Stankovic, Uros; Ploeger, Lennert S; Sonke, Jan-Jakob; van Herk, Marcel

    2016-03-01

    Image lag in the flat-panel detector used for Linac integrated cone beam computed tomography (CBCT) has a degrading effect on CBCT image quality. The most prominent visible artifact is the presence of bright semicircular structure in the transverse view of the scans, known also as radar artifact. Several correction strategies have been proposed, but until now the clinical introduction of such corrections remains unreported. In November 2013, the authors have clinically implemented a previously proposed image lag correction on all of their machines at their main site in Amsterdam. The purpose of this study was to retrospectively evaluate the effect of the correction on the quality of CBCT images and evaluate the required calibration frequency. Image lag was measured in five clinical CBCT systems (Elekta Synergy 4.6) using an in-house developed beam interrupting device that stops the x-ray beam midway through the data acquisition of an unattenuated beam for calibration. A triple exponential falling edge response was fitted to the measured data and used to correct image lag from projection images with an infinite response. This filter, including an extrapolation for saturated pixels, was incorporated in the authors' in-house developed clinical cbct reconstruction software. To investigate the short-term stability of the lag and associated parameters, a series of five image lag measurement over a period of three months was performed. For quantitative analysis, the authors have retrospectively selected ten patients treated in the pelvic region. The apparent contrast was quantified in polar coordinates for scans reconstructed using the parameters obtained from different dates with and without saturation handling. Visually, the radar artifact was minimal in scans reconstructed using image lag correction especially when saturation handling was used. In patient imaging, there was a significant reduction of the apparent contrast from 43 ± 16.7 to 15.5 ± 11.9 HU without the

  2. Clinical introduction of image lag correction for a cone beam CT system

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

    Stankovic, Uros; Ploeger, Lennert S.; Sonke, Jan-Jakob, E-mail: j.sonke@nki.nl

    Purpose: Image lag in the flat-panel detector used for Linac integrated cone beam computed tomography (CBCT) has a degrading effect on CBCT image quality. The most prominent visible artifact is the presence of bright semicircular structure in the transverse view of the scans, known also as radar artifact. Several correction strategies have been proposed, but until now the clinical introduction of such corrections remains unreported. In November 2013, the authors have clinically implemented a previously proposed image lag correction on all of their machines at their main site in Amsterdam. The purpose of this study was to retrospectively evaluate themore » effect of the correction on the quality of CBCT images and evaluate the required calibration frequency. Methods: Image lag was measured in five clinical CBCT systems (Elekta Synergy 4.6) using an in-house developed beam interrupting device that stops the x-ray beam midway through the data acquisition of an unattenuated beam for calibration. A triple exponential falling edge response was fitted to the measured data and used to correct image lag from projection images with an infinite response. This filter, including an extrapolation for saturated pixels, was incorporated in the authors’ in-house developed clinical CBCT reconstruction software. To investigate the short-term stability of the lag and associated parameters, a series of five image lag measurement over a period of three months was performed. For quantitative analysis, the authors have retrospectively selected ten patients treated in the pelvic region. The apparent contrast was quantified in polar coordinates for scans reconstructed using the parameters obtained from different dates with and without saturation handling. Results: Visually, the radar artifact was minimal in scans reconstructed using image lag correction especially when saturation handling was used. In patient imaging, there was a significant reduction of the apparent contrast from 43

  3. Ctf4 Is a Hub in the Eukaryotic Replisome that Links Multiple CIP-Box Proteins to the CMG Helicase.

    PubMed

    Villa, Fabrizio; Simon, Aline C; Ortiz Bazan, Maria Angeles; Kilkenny, Mairi L; Wirthensohn, David; Wightman, Mel; Matak-Vinkovíc, Dijana; Pellegrini, Luca; Labib, Karim

    2016-08-04

    Replisome assembly at eukaryotic replication forks connects the DNA helicase to DNA polymerases and many other factors. The helicase binds the leading-strand polymerase directly, but is connected to the Pol α lagging-strand polymerase by the trimeric adaptor Ctf4. Here, we identify new Ctf4 partners in addition to Pol α and helicase, all of which contain a "Ctf4-interacting-peptide" or CIP-box. Crystallographic analysis classifies CIP-boxes into two related groups that target different sites on Ctf4. Mutations in the CIP-box motifs of the Dna2 nuclease or the rDNA-associated protein Tof2 do not perturb DNA synthesis genome-wide, but instead lead to a dramatic shortening of chromosome 12 that contains the large array of rDNA repeats. Our data reveal unexpected complexity of Ctf4 function, as a hub that connects multiple accessory factors to the replisome. Most strikingly, Ctf4-dependent recruitment of CIP-box proteins couples other processes to DNA synthesis, including rDNA copy-number regulation. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  4. Granular resistive force theory explains the neuromechanical phase lag during sand-swimming

    NASA Astrophysics Data System (ADS)

    Ding, Yang; Sharpe, Sarah; Goldman, Daniel

    2012-11-01

    Undulatory locomotion is a common gait used by a diversity of animals in a range of environments. This mode of locomotion is characterized by the propagation of a traveling wave of body bending, which propels the animal in the opposite direction of the wave. Previous studies of undulatory locomotion in fluids, on land, and even within sand revealed that the wave of muscle activation progresses faster than the traveling wave of curvature. This leads to an increasing phase lag between activation and curvature at more posterior segments, known as the neuromechanical phase lag. In this study, we compare biological measurements of phase lag during the sand-swimming of the sandfish lizard to predictions of a simple model of undulatory swimming that consists of prescribed kinematics and granular resistive forces. The neuromechanical phase lag measured using electromyography (EMG) quantitatively matches the predicted phase lag between the local body curvature and torque exerted by granular resistive forces. Two effects are responsible for the phase lag in this system: the yaw motion of the body and different integration length over a traveling force pattern for different positions along the body.

  5. Connecting localized DNA strand displacement reactions

    NASA Astrophysics Data System (ADS)

    Mullor Ruiz, Ismael; Arbona, Jean-Michel; Lad, Amitkumar; Mendoza, Oscar; Aimé, Jean-Pierre; Elezgaray, Juan

    2015-07-01

    Logic circuits based on DNA strand displacement reactions have been shown to be versatile enough to compute the square root of four-bit numbers. The implementation of these circuits as a set of bulk reactions faces difficulties which include leaky reactions and intrinsically slow, diffusion-limited reaction rates. In this paper, we consider simple examples of these circuits when they are attached to platforms (DNA origamis). As expected, constraining distances between DNA strands leads to faster reaction rates. However, it also induces side-effects that are not detectable in the solution-phase version of this circuitry. Appropriate design of the system, including protection and asymmetry between input and fuel strands, leads to a reproducible behaviour, at least one order of magnitude faster than the one observed under bulk conditions.Logic circuits based on DNA strand displacement reactions have been shown to be versatile enough to compute the square root of four-bit numbers. The implementation of these circuits as a set of bulk reactions faces difficulties which include leaky reactions and intrinsically slow, diffusion-limited reaction rates. In this paper, we consider simple examples of these circuits when they are attached to platforms (DNA origamis). As expected, constraining distances between DNA strands leads to faster reaction rates. However, it also induces side-effects that are not detectable in the solution-phase version of this circuitry. Appropriate design of the system, including protection and asymmetry between input and fuel strands, leads to a reproducible behaviour, at least one order of magnitude faster than the one observed under bulk conditions. Electronic supplementary information (ESI) available. See DOI: 10.1039/C5NR02434J

  6. Poly(ADP-ribose) polymerases covalently modify strand break termini in DNA fragments in vitro

    PubMed Central

    Talhaoui, Ibtissam; Lebedeva, Natalia A.; Zarkovic, Gabriella; Saint-Pierre, Christine; Kutuzov, Mikhail M.; Sukhanova, Maria V.; Matkarimov, Bakhyt T.; Gasparutto, Didier; Saparbaev, Murat K.; Lavrik, Olga I.; Ishchenko, Alexander A.

    2016-01-01

    Poly(ADP-ribose) polymerases (PARPs/ARTDs) use nicotinamide adenine dinucleotide (NAD+) to catalyse the synthesis of a long branched poly(ADP-ribose) polymer (PAR) attached to the acceptor amino acid residues of nuclear proteins. PARPs act on single- and double-stranded DNA breaks by recruiting DNA repair factors. Here, in in vitro biochemical experiments, we found that the mammalian PARP1 and PARP2 proteins can directly ADP-ribosylate the termini of DNA oligonucleotides. PARP1 preferentially catalysed covalent attachment of ADP-ribose units to the ends of recessed DNA duplexes containing 3′-cordycepin, 5′- and 3′-phosphate and also to 5′-phosphate of a single-stranded oligonucleotide. PARP2 preferentially ADP-ribosylated the nicked/gapped DNA duplexes containing 5′-phosphate at the double-stranded termini. PAR glycohydrolase (PARG) restored native DNA structure by hydrolysing PAR-DNA adducts generated by PARP1 and PARP2. Biochemical and mass spectrometry analyses of the adducts suggested that PARPs utilise DNA termini as an alternative to 2′-hydroxyl of ADP-ribose and protein acceptor residues to catalyse PAR chain initiation either via the 2′,1″-O-glycosidic ribose-ribose bond or via phosphodiester bond formation between C1′ of ADP-ribose and the phosphate of a terminal deoxyribonucleotide. This new type of post-replicative modification of DNA provides novel insights into the molecular mechanisms underlying biological phenomena of ADP-ribosylation mediated by PARPs. PMID:27471034

  7. Stranded cost securitization: Analytical considerations

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

    Abbott, S.

    1997-10-01

    Securitization is a promising financing approach by which utilities may recover their stranded costs while lowering their cost of capital, permitting them to offer rate reductions to customers. However, there are important issues to analyze before determining that securitization will be an attractive option for bondholders. To facilitate the transition to a competitive electric market, numerous state legislatures have passed or are considering legislation that, while mandating competition, allows utilities to recover their stranded costs through the imposition of a competitive transition fee. To accommodate securitization of revenues from the fees, statutes typically designate as a property right the futuremore » revenues from these fees and the utility may sell, assign, or transfer the rights to a financing vehicle. Securities may be issued by a trust or other special purpose vehicle supported by future revenues from these fees. Because of the unique characteristics of the highly regulated utility industry and the {open_quotes}asset{close_quotes} that is securitized, the credit analysis of stranded cost securities differs from that of most other assets. For example, underwriting and servicing issues, which are key items of interest in other segments of the ABS market, are less of a concern in a stranded cost context.« less

  8. Longitudinal Links Between Identity Consolidation and Psychosocial Problems in Adolescence: Using Bi-Factor Latent Change and Cross-Lagged Effect Models.

    PubMed

    Hatano, Kai; Sugimura, Kazumi; Schwartz, Seth J

    2018-04-01

    Most previous identity research has focused on relationships between identity synthesis, confusion, and psychosocial problems. However, these studies did not take into account Erikson's notion of identity consolidation, that is, the dynamic interplay between identity synthesis and confusion. This study aimed to examine longitudinal relationships and the directionality of the effects between identity consolidation and psychosocial problems during adolescence, using two waves of longitudinal data from 793 Japanese adolescents (49.7% girls; ages 13-14 and 16-17 at Time 1). A bi-factor latent change model revealed that levels and changes in identity consolidation were negatively associated with levels and changes in psychosocial problems. Furthermore, a bi-factor cross-lagged effects model provided evidence that identity consolidation negatively predicted psychosocial problems, and vice versa. Our study facilitates a better understanding of the importance of identity consolidation in the relations between identity components and psychosocial problems.

  9. Homozygous diploid deletion strains of Saccharomyces cerevisiae that determine lag phase and dehydration tolerance.

    PubMed

    D'Elia, Riccardo; Allen, Patricia L; Johanson, Kelly; Nickerson, Cheryl A; Hammond, Timothy G

    2005-06-01

    This study identifies genes that determine length of lag phase, using the model eukaryotic organism, Saccharomyces cerevisiae. We report growth of a yeast deletion series following variations in the lag phase induced by variable storage times after drying-down yeast on filters. Using a homozygous diploid deletion pool, lag times ranging from 0 h to 90 h were associated with increased drop-out of mitochondrial genes and increased survival of nuclear genes. Simple linear regression (R2 analysis) shows that there are over 500 genes for which > 70% of the variation can be explained by lag alone. In the genes with a positive correlation, such that the gene abundance increases with lag and hence the deletion strain is suitable for survival during prolonged storage, there is a strong predominance of nucleonic genes. In the genes with a negative correlation, such that the gene abundance decreases with lag and hence the strain may be critical for getting yeast out of the lag phase, there is a strong predominance of glycoproteins and transmembrane proteins. This study identifies yeast deletion strains with survival advantage on prolonged storage and amplifies our understanding of the genes critical for getting out of the lag phase.

  10. Homozygous diploid deletion strains of Saccharomyces cerevisiae that determine lag phase and dehydration tolerance

    NASA Technical Reports Server (NTRS)

    D'Elia, Riccardo; Allen, Patricia L.; Johanson, Kelly; Nickerson, Cheryl A.; Hammond, Timothy G.

    2005-01-01

    This study identifies genes that determine length of lag phase, using the model eukaryotic organism, Saccharomyces cerevisiae. We report growth of a yeast deletion series following variations in the lag phase induced by variable storage times after drying-down yeast on filters. Using a homozygous diploid deletion pool, lag times ranging from 0 h to 90 h were associated with increased drop-out of mitochondrial genes and increased survival of nuclear genes. Simple linear regression (R2 analysis) shows that there are over 500 genes for which > 70% of the variation can be explained by lag alone. In the genes with a positive correlation, such that the gene abundance increases with lag and hence the deletion strain is suitable for survival during prolonged storage, there is a strong predominance of nucleonic genes. In the genes with a negative correlation, such that the gene abundance decreases with lag and hence the strain may be critical for getting yeast out of the lag phase, there is a strong predominance of glycoproteins and transmembrane proteins. This study identifies yeast deletion strains with survival advantage on prolonged storage and amplifies our understanding of the genes critical for getting out of the lag phase.

  11. SOFT LAGS IN NEUTRON STAR kHz QUASI-PERIODIC OSCILLATIONS: EVIDENCE FOR REVERBERATION?

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

    Barret, Didier, E-mail: didier.barret@irap.omp.eu; CNRS, Institut de Recherche en Astrophysique et Planetologie, 9 Av. colonel Roche, BP 44346, F-31028 Toulouse cedex 4

    2013-06-10

    High frequency soft reverberation lags have now been detected from stellar mass and supermassive black holes. Their interpretation involves reflection of a hard source of photons onto an accretion disk, producing a delayed reflected emission, with a time lag consistent with the light travel time between the irradiating source and the disk. Independently of the location of the clock, the kHz quasi-periodic oscillation (QPO) emission is thought to arise from the neutron star boundary layer. Here, we search for the signature of reverberation of the kHz QPO emission, by measuring the soft lags and the lag energy spectrum of themore » lower kHz QPOs from 4U1608-522. Soft lags, ranging from {approx}15 to {approx}40 {mu}s, between the 3-8 keV and 8-30 keV modulated emissions are detected between 565 and 890 Hz. The soft lags are not constant with frequency and show a smooth decrease between 680 Hz and 890 Hz. The broad band X-ray spectrum is modeled as the sum of a disk and a thermal Comptonized component, plus a broad iron line, expected from reflection. The spectral parameters follow a smooth relationship with the QPO frequency, in particular the fitted inner disk radius decreases steadily with frequency. Both the bump around the iron line in the lag energy spectrum and the consistency between the lag changes and the inferred changes of the inner disk radius, from either spectral fitting or the QPO frequency, suggest that the soft lags may indeed involve reverberation of the hard pulsating QPO source on the disk.« less

  12. Time-lagged response of the Antarctic atmosphere to tropical MJO convection

    NASA Astrophysics Data System (ADS)

    Henderson, G. R.; Barrett, B.

    2017-12-01

    Intraseasonal tropical climate variability has important implications on mid- and high- latitude climate, and in recent studies has been shown to modulate a number of weather processes in the Northern Hemisphere, such as snow depth, sea ice concentration, precipitation, atmospheric rivers, and air temperature. In such studies, the leading mode of tropical intraseasonal variability, the Madden-Julian Oscillation (MJO), has tended to lag tropical convection by approximately 7 days. However, no such consensus exists when considering the relationship and lag between the MJO and the Antarctic atmosphere. This study builds on previous work by further examining the time-lagged response of Southern Hemisphere tropospheric circulation to tropical MJO forcing, with specific focus on the latitude belt associated with the Antarctic Oscillation, during the months of June (Austral winter) and December (Austral summer) using NCEP-DOE Reanalysis 2 data for the years 1970-2016. Principal findings indicate that the time lag with strongest height anomalies depends on both the location of the MJO convection (e.g., the MJO phase) and the season, and that the lagged height anomalies in the Antarctic atmosphere are fairly consistent across different vertical levels and latitudinal bands. In addition, certain MJO phases in December displayed lagged height anomalies indicative of blocking-type atmospheric patterns, with an approximate wavenumber of 4, whereas in June most phases were associated with progressive height anomaly centers displaying a wavenumber-3 type pattern.

  13. Structure of the replication fork in ultraviolet light-irradiated human cells.

    PubMed Central

    Cordeiro-Stone, M; Schumacher, R I; Meneghini, R

    1979-01-01

    The DNA extracted from xeroderma pigmentosum human fibroblasts previously irradiated with 12.5 J/m2 of UV light and pulse-labeled for 45 min with radioactive and (or) heavy precursors, was used to determine the structural characteristics of the replication fork. Density equilibrium centrifugation experiments showed that a fork moved 6 micrometer in 45 min and bypassed 3 pyrimidine dimers in both strands. The same length was covered in 15-20 min in control cells. The delay in irradiated cells was apparently due to pyrimidine dimers acting as temporary blocks to the fork movement. Evidence for this interpretation comes from kinetics of incorporation of [3H]thymidine into DNA, which show that the time necessary to attain a new stable level of DNA synthesis in irradiated cells is equivalent to that required for the replication fork to cover the interdimer distance in one strand. On the other hand, the action of S1 nuclease on DNA synthesized soon after irradiation gives rise to a bimodal distribution in neutral sucrose gradients, one peak corresponding to 43 X 10(6) daltons and the other to 3 X 10(6) daltons. These two DNA species are generated by the attack of the S1 nuclease on single-stranded regions associated with the replication fork. A possible explanation for these results is given by a model according to which there is a delayed bypass of the dimer in the leading strand and the appearance of gaps opposite pyrimidine dimers in the lagging strand, as a direct consequence of the discontinuous mode of DNA replication. In terms of the model, the DNA of 43 X 10(6) daltons corresponds to the leading strand, linked to the unreplicated branch of the forks, whereas the piece of 3 X 10(6) daltons is the intergap DNA coming from the lagging strand. Pulse and chase experiments reveal that the low molecular weight DNA grows in a pattern that suggests that more than one gap may be formed per replication fork. PMID:233582

  14. Structure of the replication fork in ultraviolet light-irradiated human cells.

    PubMed

    Cordeiro-Stone, M; Schumacher, R I; Meneghini, R

    1979-08-01

    The DNA extracted from xeroderma pigmentosum human fibroblasts previously irradiated with 12.5 J/m2 of UV light and pulse-labeled for 45 min with radioactive and (or) heavy precursors, was used to determine the structural characteristics of the replication fork. Density equilibrium centrifugation experiments showed that a fork moved 6 micrometer in 45 min and bypassed 3 pyrimidine dimers in both strands. The same length was covered in 15-20 min in control cells. The delay in irradiated cells was apparently due to pyrimidine dimers acting as temporary blocks to the fork movement. Evidence for this interpretation comes from kinetics of incorporation of [3H]thymidine into DNA, which show that the time necessary to attain a new stable level of DNA synthesis in irradiated cells is equivalent to that required for the replication fork to cover the interdimer distance in one strand. On the other hand, the action of S1 nuclease on DNA synthesized soon after irradiation gives rise to a bimodal distribution in neutral sucrose gradients, one peak corresponding to 43 X 10(6) daltons and the other to 3 X 10(6) daltons. These two DNA species are generated by the attack of the S1 nuclease on single-stranded regions associated with the replication fork. A possible explanation for these results is given by a model according to which there is a delayed bypass of the dimer in the leading strand and the appearance of gaps opposite pyrimidine dimers in the lagging strand, as a direct consequence of the discontinuous mode of DNA replication. In terms of the model, the DNA of 43 X 10(6) daltons corresponds to the leading strand, linked to the unreplicated branch of the forks, whereas the piece of 3 X 10(6) daltons is the intergap DNA coming from the lagging strand. Pulse and chase experiments reveal that the low molecular weight DNA grows in a pattern that suggests that more than one gap may be formed per replication fork.

  15. LagLoc - a new surgical technique for locking plate systems.

    PubMed

    Triana, Miguel; Gueorguiev, Boyko; Sommer, Christoph; Stoffel, Karl; Agarwal, Yash; Zderic, Ivan; Helfen, Tobias; Krieg, James C; Krause, Fabian; Knobe, Matthias; Richards, R Geoff; Lenz, Mark

    2018-06-19

    Treatment of oblique and spiral fractures remains challenging. The aim of this study was to introduce and investigate the new LagLoc technique for locked plating with generation of interfragmentary compression, combining the advantages of lag-screw and locking-head-screw techniques. Oblique fracture was simulated in artificial diaphyseal bones, assigned to three groups for plating with a 7-hole locking compression plate. Group I was plated with three locking screws in holes 1, 4 and 7. The central screw crossed the fracture line. In group II the central hole was occupied with a lag screw perpendicular to fracture line. Group III was instrumented applying the LagLoc technique as follows. Hole 4 was predrilled perpendicularly to the plate, followed by overdrilling of the near cortex and insertion of a locking screw whose head was covered by a holding sleeve to prevent temporarily the locking in the plate hole and generate interfragmentary compression. Subsequently, the screw head was released and locked in the plate hole. Holes 1 and 7 were occupied with locking screws. Interfragmentary compression in the fracture gap was measured using pressure sensors. All screws in the three groups were tightened with 4Nm torque. Interfragmentary compression in group I (167 ± 25N) was significantly lower in comparison to groups II (431 ± 21N) and III (379 ± 59N), p≤0.005. The difference in compression between groups II and III remained not significant (p = 0.999). The new LagLoc technique offers an alternative tool to generate interfragmentary compression with the application of locking plates by combining the biomechanical advantages of lag screw and locking screw fixations. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  16. Markers of Decompression Stress of Mass Stranded/Live Caught and Released vs. Single Stranded Marine Mammals

    DTIC Science & Technology

    2014-09-30

    Caught and Released vs. Single Stranded Marine Mammals Michael Moore Biology Department Woods Hole Oceanographic Institution Woods Hole, MA 02543...analyze blood samples from captive, wild-caught, and stranded marine mammals in order to compare concentrations of Microparticles (MPs). If confirmed...military sonar or during seismic exploration, may harm marine animals. It has been suggested that alteration in physiology or diving behavior may

  17. Inhibition of GABA synthesis in the prefrontal cortex increases locomotor activity but does not affect attention in the 5-choice serial reaction time task.

    PubMed

    Asinof, Samuel K; Paine, Tracie A

    2013-02-01

    Attention deficits are a core cognitive symptom of schizophrenia; the neuropathology underlying these deficits is not known. Attention is regulated, at least in part, by the prefrontal cortex (PFC), a brain area in which pathology of γ-aminobutyric acid (GABA) neurons has been consistently observed in post-mortem analysis of the brains of people with schizophrenia. Specifically, expression of the 67-kD isoform of the GABA synthesis enzyme glutamic acid decarboxylase (GAD67) is reduced in parvalbumin-containing fast-spiking GABA interneurons. Thus it is hypothesized that reduced cortical GABA synthesis and release may contribute to the attention deficits in schizophrenia. Here the effect of reducing cortical GABA synthesis with l-allylglycine (LAG) on attention was tested using three different versions of the 5-choice serial reaction time task (5CSRTT). Because 5CSRTT performance can be affected by locomotor activity, we also measured this behavior in an open field. Finally, the expression of Fos protein was used as an indirect measure of reduced GABA synthesis. Intra-cortical LAG (10 μg/0.5 μl/side) infusions increased Fos expression and resulted in hyperactivity in the open field. Intra-cortical LAG infusions did not affect attention in any version of the 5CSRTT. These results suggest that a general decrease in GABA synthesis is not sufficient to cause attention deficits. It remains to be tested whether a selective decrease in GABA synthesis in parvalbumin-containing GABA neurons could cause attention deficits. Decreased cortical GABA synthesis did increase locomotor activity; this may reflect the positive symptoms of schizophrenia. Copyright © 2012 Elsevier Ltd. All rights reserved.

  18. The economics of stranded investment - a two-way street

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

    Cearley, R.; McKinzie, L.

    In the transition to deregulation, the risk, costs and benefits of utility assets are transferred from the customer to the investor, creating potential stranded benefits as well as stranded costs. Investors may be better or worse off depending on whether an asset`s cost is below or above market. Regulators can minimize unintended wealth transfers by compensating each potential loser in the transition. The amount of investment stranded - i.e., the portion of plant that is above market value - does seem to be a murky issue. This article sets a framework for evaluating stranded investment and traces the possible welfaremore » effects of different policies to address it. It defines {open_quote}stranded costs,{close_quote} {open_quote}stranded investment,{close_quote} and {open_quote}stranded benefits.{close_quote} It addresses their interrelationship, and shows that the redefinition of property rights during the transition to a competitive market is what leads to stranded investment. The elimination of the utility`s exclusive franchise - i.e., its obligation to serve and customers` obligation to pay - leads to the redefinition of those property rights as they pertain to the costs, benefits and risks associated with existing utility generation. Finally, the authors address the possible welfare implications from this transition.« less

  19. Translocation of double strand DNA into a biological nanopore

    NASA Astrophysics Data System (ADS)

    Chatkaew, Sunita; Mlayeh, Lamia; Leonetti, Marc; Homble, Fabrice

    2009-03-01

    Translocation of double strand DNA across a unique mitochondrial biological nanopore (VDAC) is observed by an electrophysiological method. Characteristics of opened and sub-conductance states of VDAC are studied. When the applied electric potential is beyond ± 20 mV, VDAC transits to a sub-conductance state. Plasmids (circular double strand DNA) with a diameter greater than that of the channel shows the current reduction into the channel during the interaction but the state with zero-current is not observed. On the contrary, the interaction of linear double strand DNA with the channel shows the current reduction along with the zero-current state. These show the passages of linear double strand DNA across the channel and the electrostatic effect due to the surface charges of double strand DNA and channel for circular and linear double strand DNA.

  20. Time lag and communication in changing unpopular norms.

    PubMed

    Gërxhani, Klarita; Bruggeman, Jeroen

    2015-01-01

    Humans often coordinate their social lives through norms. When a large majority of people are dissatisfied with an existing norm, it seems obvious that they will change it. Often, however, this does not occur. We investigate how a time lag between individual support of a norm change and the change itself hinders such change, related to the critical mass of supporters needed to effectuate the change, and the (im)possibility of communicating about it. To isolate these factors, we utilize a laboratory experiment. As predicted, we find unambiguous effects of time lag on precluding norm change; a higher threshold for a critical mass does so as well. Communication facilitates choosing superior norms but it does not necessarily lead to norm change when the uncertainty on whether there will be a norm change in the future is high. Communication seems to help coordination on actions at the present but not the future. Hence, the uncertainty driven by time lag makes individuals choose the status quo, here the unpopular norm.

  1. Time Lag and Communication in Changing Unpopular Norms

    PubMed Central

    Gërxhani, Klarita; Bruggeman, Jeroen

    2015-01-01

    Humans often coordinate their social lives through norms. When a large majority of people are dissatisfied with an existing norm, it seems obvious that they will change it. Often, however, this does not occur. We investigate how a time lag between individual support of a norm change and the change itself hinders such change, related to the critical mass of supporters needed to effectuate the change, and the (im)possibility of communicating about it. To isolate these factors, we utilize a laboratory experiment. As predicted, we find unambiguous effects of time lag on precluding norm change; a higher threshold for a critical mass does so as well. Communication facilitates choosing superior norms but it does not necessarily lead to norm change when the uncertainty on whether there will be a norm change in the future is high. Communication seems to help coordination on actions at the present but not the future. Hence, the uncertainty driven by time lag makes individuals choose the status quo, here the unpopular norm. PMID:25880200

  2. Shear-lag analysis about an internally-dropped ply

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

    Vizzini, A.J.

    1995-12-31

    The region around a terminated ply is modeled as several elastic layers separated by shear regions. A shear-lag analysis is then performed allowing for the thickness of the elastic and shear layers to vary. Boundary conditions, away for the ply drop, are based on the deflections determined by a finite element model. The interlaminar stresses are compared against those generated by the finite element model for tapered laminates under pure extension, pure bending, and extension-bending coupling. The shear-lag analysis predicts the interlaminar shear at and near the ply drop for pure extension and in cases involving bending if the deflectionsmore » due to bending are removed. The interlaminar shear stress and force equilibrium are used to determine the interlaminar normal stress. The trends in the interlaminar normal stress shown by the finite element model are partially captured by the shear-lag analysis. This simple analysis indicates that the mechanism for load transfer about a ply drop is primarily due to shear transfer through the resin rich areas.« less

  3. Low-residue euthanasia of stranded mysticetes.

    PubMed

    Harms, Craig A; McLellan, William A; Moore, Michael J; Barco, Susan G; Clarke, Elsburgh O; Thayer, Victoria G; Rowles, Teresa K

    2014-01-01

    Euthanasia of stranded large whales poses logistic, safety, pharmaceutical, delivery, public relations, and disposal challenges. Reasonable arguments may be made for allowing a stranded whale to expire naturally. However, slow cardiovascular collapse from gravitational effects outside of neutral buoyancy, often combined with severely debilitating conditions, motivate humane efforts to end the animal's suffering. The size of the animal and prevailing environmental conditions often pose safety concerns for stranding personnel, which take priority over other considerations. When considering chemical euthanasia, the size of the animal also necessitates large quantities of euthanasia agents. Drug residues are a concern for relay toxicity to scavengers, particularly for pentobarbital-containing euthanasia solutions. Pentobarbital is also an environmental concern because of its stability and long persistence in aquatic environments. We describe a euthanasia technique for stranded mysticetes using readily available, relatively inexpensive, preanesthetic and anesthetic drugs (midazolam, acepromazine, xylazine) followed by saturated KCl delivered via custom-made needles and a low-cost, basic, pressurized canister. This method provides effective euthanasia while moderating personnel exposure to hazardous situations and minimizing drug residues of concern for relay toxicity.

  4. An evaluation of the lag of accommodation using photorefraction.

    PubMed

    Seidemann, Anne; Schaeffel, Frank

    2003-02-01

    The lag of accommodation which occurs in most human subjects during reading has been proposed to explain the association between reading and myopia. However, the measured lags are variable among different published studies and current knowledge on its magnitude rests largely on measurements with the Canon R-1 autorefractor. Therefore, we have measured it with another technique, eccentric infrared photorefraction (the PowerRefractor), and studied how it can be modified. Particular care was taken to ensure correct calibration of the instrument. Ten young adult subjects were refracted both in the fixation axis of the right eye and from the midline between both eyes, while they read text both monocularly and binocularly at 1.5, 2, 3, 4 and 5 D distance ("group 1"). A second group of 10 subjects ("group 2"), measured from the midline between both eyes, was studied to analyze the effects of binocular vs monocular vision, addition of +1 or +2 D lenses, and of letter size. Spherical equivalents (SE) were analyzed in all cases. The lag of accommodation was variable among subjects (standard deviations among groups and viewing distances ranging from 0.18 to 1.07 D) but was significant when the measurements were done in the fixation axis (0.35 D at 3 D target distance to 0.60 D at 5 D with binocular vision; p<0.01 or better all cases). Refracting from the midline between both eyes tended to underestimate the lag of accommodation although this was significant only at 5 D (ANOVA: p<0.0001, post hoc t-test: p<0.05). There was a small improvement in accommodation precision with binocular compared to monocular viewing but significance was reached only for the 5 D reading target (group 1--lags for a 3/4/5 D target: 0.35 vs 0.41 D/0.48 vs 0.47 D/0.60 vs 0.66 D, ANOVA: p<0.0001, post hoc t-test: p<0.05; group 2--0.29 vs 0.12 D, 0.33 vs 0.16 D, 0.23 vs -0.31 D, ANOVA: p<0.0001, post hoc t-test: p<0.05). Adjusting the letter height for constant angular subtense (0.2 deg) induced

  5. Enzymatic production of 'monoclonal stoichiometric' single-stranded DNA oligonucleotides.

    PubMed

    Ducani, Cosimo; Kaul, Corinna; Moche, Martin; Shih, William M; Högberg, Björn

    2013-07-01

    Single-stranded oligonucleotides are important as research tools, as diagnostic probes, in gene therapy and in DNA nanotechnology. Oligonucleotides are typically produced via solid-phase synthesis, using polymer chemistries that are limited relative to what biological systems produce. The number of errors in synthetic DNA increases with oligonucleotide length, and the resulting diversity of sequences can be a problem. Here we present the 'monoclonal stoichiometric' (MOSIC) method for enzyme-mediated production of DNA oligonucleotides. We amplified oligonucleotides from clonal templates derived from single bacterial colonies and then digested cutter hairpins in the products, which released pools of oligonucleotides with precisely controlled relative stoichiometric ratios. We prepared 14-378-nucleotide MOSIC oligonucleotides either by in vitro rolling-circle amplification or by amplification of phagemid DNA in Escherichia coli. Analyses of the formation of a DNA crystal and folding of DNA nanostructures confirmed the scalability, purity and stoichiometry of the produced oligonucleotides.

  6. Protein stabilization by introduction of cross-strand disulfides.

    PubMed

    Chakraborty, Kausik; Thakurela, Sudhir; Prajapati, Ravindra Singh; Indu, S; Ali, P Shaik Syed; Ramakrishnan, C; Varadarajan, Raghavan

    2005-11-08

    Disulfides cross-link residues in a protein that are separated in primary sequence and stabilize the protein through entropic destabilization of the unfolded state. While the removal of naturally occurring disulfides leads to protein destabilization, introduction of engineered disulfides does not always lead to significant stabilization of a protein. We have analyzed naturally occurring disulfides that span adjacent antiparallel strands of beta sheets (cross-strand disulfides). Cross-strand disulfides have recently been implicated as redox-based conformational switches in proteins such as gp120 and CD4. The propensity of these disulfides to act as conformational switches was postulated on the basis of the hypothesis that this class of disulfide is conformationally strained. In the present analysis, there was no evidence to suggest that cross-strand disulfides are more strained compared to other disulfides as assessed by their torsional energy. It was also observed that these disulfides occur solely at non-hydrogen-bonded (NHB) registered pairs of adjacent antiparallel strands and not at hydrogen-bonded (HB) positions as suggested previously. One of the half-cystines involved in cross-strand disulfide formation often occurs at an edge strand. Experimental confirmation of the stabilizing effects of such disulfides was carried out in Escherichia coli thioredoxin. Four pairs of cross-strand cysteines were introduced, two at HB and two at NHB pairs. Disulfides were formed in all four cases. However, as predicted from our analysis, disulfides at NHB positions resulted in an increase in melting temperature of 7-10 degrees C, while at HB positions there was a corresponding decrease of -7 degrees C. The reduced state of all proteins had similar stability.

  7. The shear-lag effect of thin-walled box girder under vertical earthquake excitation

    NASA Astrophysics Data System (ADS)

    Zhai, Zhipeng; Li, Yaozhuang; Guo, Wei

    2017-03-01

    The variation method based on the energy variation principle is proved to be accurate and valid for analyzing the shear lag effect of box girder under static and dynamic load. Meanwhile, dynamic problems gradually become the key factors in engineering practice. Therefore, a method for calculating the shear lag effect in thin-walled box girder under vertical seismic excitation is proposed by applying Hamilton Principle in this paper. The Timoshenko shear deformation is taken into account. And a new definition of shear lag ratio for box girder is given. What's more, some conclusions are drawn by analysis of numerical example. The results show that small amplitude of earthquake ground motion can generate high stress and obvious shear lag, especially in the region of resonance. And the influence of rotary inertia cannot be ignored for analyzing the shear lag effect. With the increase of span to width ratio, shear lag effect becomes smaller and smaller. These research conclusions will be useful for the engineering practice and enrich the theoretical studies of box girders.

  8. A nonlinear lag correction algorithm for a-Si flat-panel x-ray detectors

    PubMed Central

    Starman, Jared; Star-Lack, Josh; Virshup, Gary; Shapiro, Edward; Fahrig, Rebecca

    2012-01-01

    Purpose: Detector lag, or residual signal, in a-Si flat-panel (FP) detectors can cause significant shading artifacts in cone-beam computed tomography reconstructions. To date, most correction models have assumed a linear, time-invariant (LTI) model and correct lag by deconvolution with an impulse response function (IRF). However, the lag correction is sensitive to both the exposure intensity and the technique used for determining the IRF. Even when the LTI correction that produces the minimum error is found, residual artifact remains. A new non-LTI method was developed to take into account the IRF measurement technique and exposure dependencies. Methods: First, a multiexponential (N = 4) LTI model was implemented for lag correction. Next, a non-LTI lag correction, known as the nonlinear consistent stored charge (NLCSC) method, was developed based on the LTI multiexponential method. It differs from other nonlinear lag correction algorithms in that it maintains a consistent estimate of the amount of charge stored in the FP and it does not require intimate knowledge of the semiconductor parameters specific to the FP. For the NLCSC method, all coefficients of the IRF are functions of exposure intensity. Another nonlinear lag correction method that only used an intensity weighting of the IRF was also compared. The correction algorithms were applied to step-response projection data and CT acquisitions of a large pelvic phantom and an acrylic head phantom. The authors collected rising and falling edge step-response data on a Varian 4030CB a-Si FP detector operating in dynamic gain mode at 15 fps at nine incident exposures (2.0%–92% of the detector saturation exposure). For projection data, 1st and 50th frame lag were measured before and after correction. For the CT reconstructions, five pairs of ROIs were defined and the maximum and mean signal differences within a pair were calculated for the different exposures and step-response edge techniques. Results: The LTI

  9. Why does lag affect the durability of memory-based automaticity: loss of memory strength or interference?

    PubMed

    Wilkins, Nicolas J; Rawson, Katherine A

    2013-10-01

    In Rickard, Lau, and Pashler's (2008) investigation of the lag effect on memory-based automaticity, response times were faster and proportion of trials retrieved was higher at the end of practice for short lag items than for long lag items. However, during testing after a delay, response times were slower and proportion of trials retrieved was lower for short lag items than for long lag items. The current study investigated the extent to which the lag effect on the durability of memory-based automaticity is due to interference or to the loss of memory strength with time. Participants repeatedly practiced alphabet subtraction items in short lag and long lag conditions. After practice, half of the participants were immediately tested and the other half were tested after a 7-day delay. Results indicate that the lag effect on the durability of memory-based automaticity is primarily due to interference. We discuss potential modification of current memory-based processing theories to account for these effects. © 2013.

  10. Accommodative Lag by Autorefraction and Two Dynamic Retinoscopy Methods

    PubMed Central

    2008-01-01

    Purpose To evaluate two clinical procedures, MEM and Nott retinoscopy, for detecting accommodative lags 1.00 diopter (D) or greater in children as identified by an open-field autorefractor. Methods 168 children 8 to <12 years old with low myopia, normal visual acuity, and no strabismus participated as part of an ancillary study within the screening process for a randomized trial. Accommodative response to a 3.00 D demand was first assessed by MEM and Nott retinoscopy, viewing binocularly with spherocylindrical refractive error corrected, with testing order randomized and each performed by a different masked examiner. The response was then determined viewing monocularly with spherical equivalent refractive error corrected, using an open-field autorefractor, which was the gold standard used for eligibility for the clinical trial. Sensitivity and specificity for accommodative lags of 1.00 D or more were calculated for each retinoscopy method compared to the autorefractor. Results 116 (69%) of the 168 children had accommodative lag of 1.00 D or more by autorefraction. MEM identified 66 children identified by autorefraction for a sensitivity of 57% (95% CI = 47% to 66%) and a specificity of 63% (95% CI = 49% to 76%). Nott retinoscopy identified 35 children for a sensitivity of 30% (95% CI = 22% to 39%) and a specificity of 81% (95% CI = 67% to 90%). Analysis of receiver operating characteristic (ROC) curves constructed for MEM and for Nott retinoscopy failed to reveal alternate cut points that would improve the combination of sensitivity and specificity for identifying accommodative lag ≥ 1.00 D as defined by autorefraction. Conclusions Neither MEM nor Nott retinoscopy provided adequate sensitivity and specificity to identify myopic children with accommodative lag ≥ 1.00 D as determined by autorefraction. A variety of methodological differences between the techniques may contribute to the modest to poor agreement. PMID:19214130

  11. Psoralen interstrand cross-link repair is specifically altered by an adjacent triple-stranded structure

    PubMed Central

    Guillonneau, F.; Guieysse, A. L.; Nocentini, S.; Giovannangeli, C.; Praseuth, D.

    2004-01-01

    Targeting DNA-damaging agents to specific DNA sites by using sequence-specific DNA ligands has been successful in directing genomic modifications. The understanding of repair processing of such targeted damage and the influence of the adjacent complex is largely unknown. In this way, directed interstrand cross-links (ICLs) have already been generated by psoralen targeting. The mechanisms responsible for ICL removal are far from being understood in mammalian cells, with the proposed involvement of both mutagenic and recombinogenic pathways. Here, a unique ICL was introduced at a selected site by photoactivation of a psoralen moiety with the use of psoralen conjugates of triplex-forming oligonucleotides. The processing of psoralen ICL was evaluated in vitro and in cells for two types of cross-linked substrates, either containing a psoralen ICL alone or with an adjacent triple-stranded structure. We show that the presence of a neighbouring triplex structure interferes with different stages of psoralen ICL processing: (i) the ICL-induced DNA repair synthesis in HeLa cell extracts is inhibited by the triplex structure, as measured by the efficiency of ‘true’ and futile repair synthesis, stopping at the ICL site; (ii) in HeLa cells, the ICL removal via a nucleotide excision repair (NER) pathway is delayed in the presence of a neighbouring triplex; and (iii) the binding to ICL of recombinant xeroderma pigmentosum A protein, which is involved in pre-incision recruitment of NER factors is impaired by the presence of the third DNA strand. These data characterize triplex-induced modulation of ICL repair pathways at specific steps, which might have implications for the controlled induction of targeted genomic modifications and for the associated cellular responses. PMID:14966263

  12. Circadian rhythms, athletic performance, and jet lag

    PubMed Central

    Manfredini, R.; Manfredini, F.; Fersini, C.; Conconi, F.

    1998-01-01

    Rapid air travel across several time zones exposes the traveller to a shift in his/her internal biological clock. The result is a transient desynchronisation of the circadian rhythm, called jet lag, lasting until the rhythm is rephased to the new environmental conditions. The most commonly experienced symptoms are sleep disorders, difficulties with concentrating, irritability, depression, fatigue, disorientation, loss of appetite, and gastrointestinal disturbance. Apart from the decrements in mental and physical performance directly consequent on such symptoms, competitive athletes are also exposed to the additional negative consequences of a shift from the optimal circadian window of performance. A brief summary of the possible negative effects of jet lag on athletic performance and potentially alleviating strategies is given. 




 PMID:9631214

  13. Yeast Pif1 Accelerates Annealing of Complementary DNA Strands

    PubMed Central

    2015-01-01

    Pif1 is a helicase involved in the maintenance of nuclear and mitochondrial genomes in eukaryotes. Here we report a new activity of Saccharomyces cerevisiae Pif1, annealing of complementary DNA strands. We identified preferred substrates for annealing as those that generate a duplex product with a single-stranded overhang relative to a blunt end duplex. Importantly, we show that Pif1 can anneal DNA in the presence of ATP and Mg2+. Pif1-mediated annealing also occurs in the presence of single-stranded DNA binding proteins. Additionally, we show that partial duplex substrates with 3′-single-stranded overhangs such as those generated during double-strand break repair can be annealed by Pif1. PMID:25393406

  14. Yeast Pif1 accelerates annealing of complementary DNA strands.

    PubMed

    Ramanagoudr-Bhojappa, Ramanagouda; Byrd, Alicia K; Dahl, Christopher; Raney, Kevin D

    2014-12-09

    Pif1 is a helicase involved in the maintenance of nuclear and mitochondrial genomes in eukaryotes. Here we report a new activity of Saccharomyces cerevisiae Pif1, annealing of complementary DNA strands. We identified preferred substrates for annealing as those that generate a duplex product with a single-stranded overhang relative to a blunt end duplex. Importantly, we show that Pif1 can anneal DNA in the presence of ATP and Mg(2+). Pif1-mediated annealing also occurs in the presence of single-stranded DNA binding proteins. Additionally, we show that partial duplex substrates with 3'-single-stranded overhangs such as those generated during double-strand break repair can be annealed by Pif1.

  15. Method of preparing and applying single stranded DNA probes to double stranded target DNAs in situ

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel

    1991-01-01

    A method is provided for producing single stranded non-self-complementary nucleic acid probes, and for treating target DNA for use therewith. Probe is constructed by treating DNA with a restriction enzyme and an exonuclease to form template/primers for a DNA polymerase. The digested strand is resynthesized in the presence of labeled nucleoside triphosphate precursor. Labeled single stranded fragments are separated from the resynthesized fragments to form the probe. Target DNA is treated with the same restriction enzyme used to construct the probe, and is treated with an exonuclease before application of the probe. The method significantly increases the efficiency and specificity of hybridization mixtures by increasing effective probe concentration by eliminating self-hybridization between both probe and target DNAs, and by reducing the amount of target DNA available for mismatched hybridizations.

  16. 49 CFR 230.64 - Leaks under lagging.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ..., DEPARTMENT OF TRANSPORTATION STEAM LOCOMOTIVE INSPECTION AND MAINTENANCE STANDARDS Boilers and Appurtenances Steam Leaks § 230.64 Leaks under lagging. The steam locomotive owner and/or operator shall take out of...

  17. Aeromechanical stability augmentation using semi-active friction-based lead-lag damper

    NASA Astrophysics Data System (ADS)

    Agarwal, Sandeep

    2005-11-01

    Lead-lag dampers are present in most rotors to provide the required level of damping in all flight conditions. These dampers are a critical component of the rotor system, but they also represent a major source of maintenance cost. In present rotor systems, both hydraulic and elastomeric lead-lag dampers have been used. Hydraulic dampers are complex mechanical components that require hydraulic fluids and have high associated maintenance costs. Elastomeric dampers are conceptually simpler and provide a "dry" rotor, but are rather costly. Furthermore, their damping characteristics can degrade with time without showing external signs of failure. Hence, the dampers must be replaced on a regular basis. A semi-active friction based lead-lag damper is proposed as a replacement for hydraulic and elastomeric dampers. Damping is provided by optimized energy dissipation due to frictional forces in semi-active joints. An actuator in the joint modulates the normal force that controls energy dissipation at the frictional interfaces, resulting in large hysteretic loops. Various selective damping strategies are developed and tested for a simple system containing two different frequency modes in its response, one of which needs to be damped out. The system reflects the situation encountered in rotor response where 1P excitation is present along with the potentially unstable regressive lag motion. Simulation of the system response is obtained to compare their effectiveness. Next, a control law governing the actuation in the lag damper is designed to generate the desired level of damping for performing adaptive selective damping of individual blade lag motion. Further, conceptual design of a piezoelectric friction based lag damper for a full-scale rotor is presented and various factors affecting size, design and maintenance cost, damping capacity, and power requirements of the damper are discussed. The selective semi-active damping strategy is then studied in the context of classical

  18. Lead-Lag Control for Helicopter Vibration and Noise Reduction

    NASA Technical Reports Server (NTRS)

    Gandhi, Farhan

    1995-01-01

    . Both schemes cause an increase in pitch link loads. Trailing Edge Flap (TEF) deployment can also used to generate unsteady aerodynamic forces and moments that counter the original vibratory loads, and thereby reduce rotor vibrations. While the vibrations absorbers, HHC, IBC, and TEF concepts discussed above attempt to reduce the vibratory loads, they do not specifically address the phenomena causing the vibrations at high advance ratios. One passive method that attempts to directly alleviate compressibility and stall, instead of reducing the ensuing vibrations, is the use of advanced tip designs. Taper, sweep, anhedral, and the manipulation of other geometric properties of the blade tips can reduce the severity of stall and compressibility effects , as well as reduce rotor power. A completely different approach to solve these problems is the tiltrotor configuration. As the forward velocity of the aircraft increases, the rotors, in this case, are tilted forward until they are perpendicular to the flow and act as propellers. This eliminates the edgewise flow encountered by conventional rotors and circumvents all the problems associated with flow asymmetry. However, the success involves a tremendous increase in cost and complexity of the aircraft. Another possible approach that has been proposed for the alleviation of vibratory loads at high forward flight speeds involves the use of controlled lead-lag motions to reduce the asymmetry in flow. A correctly phased 1/rev controlled lag motion could be introduced such that it produces a backward velocity on the advancing side and a forward velocity on the retreating side, to delay compressibility effects and stall to a higher advance ratio. Using a large enough lead-lag amplitude, the tip velocities could be reduced to levels encountered in hover. This concept was examined by two groups in the 1950's and early 1960's. In the United States, the Research Labs Division of United Aircraft developed a large lead-lag motion rotor

  19. Development of strand burner for solid propellant burning rate studies

    NASA Astrophysics Data System (ADS)

    Aziz, A.; Mamat, R.; Ali, W. K. Wan

    2013-12-01

    It is well-known that a strand burner is an apparatus that provides burning rate measurements of a solid propellant at an elevated pressure in order to obtain the burning characteristics of a propellant. This paper describes the facilities developed by author that was used in his studies. The burning rate characteristics of solid propellant have be evaluated over five different chamber pressures ranging from 1 atm to 31 atm using a strand burner. The strand burner has a mounting stand that allows the propellant strand to be mounted vertically. The strand was ignited electrically using hot wire, and the burning time was recorded by electronic timer. Wire technique was used to measure the burning rate. Preliminary results from these techniques are presented. This study shows that the strand burner can be used on propellant strands to obtain accurate low pressure burning rate data.

  20. Influence of several factors on ignition lag in a compression-ignition engine

    NASA Technical Reports Server (NTRS)

    Gerrish, Harold C; Voss, Fred

    1932-01-01

    This investigation was made to determine the influence of fuel quality, injection advance angle, injection valve-opening pressure, inlet-air pressure, compression ratio, and engine speed on the time lag of auto-ignition of a Diesel fuel oil in a single-cylinder compression-ignition engine as obtained from an analysis of indicator diagrams. Three cam-operated fuel-injection pumps, two pumps cams, and an automatic injection valve with two different nozzles were used. Ignition lag was considered to be the interval between the start of injection of the fuel as determined with a Stroborama and the start of effective combustion as determined from the indicator diagram, the latter being the point where 4.0 x 10(exp-6) pound of fuel had been effectively burned. For this particular engine and fuel it was found that: (1) for a constant start and the same rate of fuel injection up the point of cut-off, a variation in fuel quantity from 1.2 x 10(exp-4) to 4.1 x 10(exp-4) pound per cycle has no appreciable effect on the ignition lag; (2) injection advance angle increases or decreases the lag according to whether density, temperature, or turbulence has the controlling influence; (3) increase in valve-opening pressure slightly increases the lag; and (4) increase of inlet-air pressure, compression ratio, and engine speed reduces the lag.

  1. Chlamydomonas chloroplasts can use short dispersed repeats and multiple pathways to repair a double-strand break in the genome.

    PubMed

    Odom, Obed W; Baek, Kwang-Hyun; Dani, Radhika N; Herrin, David L

    2008-03-01

    Certain group I introns insert into intronless DNA via an endonuclease that creates a double-strand break (DSB). There are two models for intron homing in phage: synthesis-dependent strand annealing (SDSA) and double-strand break repair (DSBR). The Cr.psbA4 intron homes efficiently from a plasmid into the chloroplast psbA gene in Chlamydomonas, but little is known about the mechanism. Analysis of co-transformants selected using a spectinomycin-resistant 16S gene (16S(spec)) provided evidence for both pathways. We also examined the consequences of the donor DNA having only one-sided or no homology with the psbA gene. When there was no homology with the donor DNA, deletions of up to 5 kb involving direct repeats that flank the psbA gene were obtained. Remarkably, repeats as short as 15 bp were used for this repair, which is consistent with the single-strand annealing (SSA) pathway. When the donor had one-sided homology, the DSB in most co-transformants was repaired using two DNAs, the donor and the 16S(spec) plasmid, which, coincidentally, contained a region that is repeated upstream of psbA. DSB repair using two separate DNAs provides further evidence for the SDSA pathway. These data show that the chloroplast can repair a DSB using short dispersed repeats located proximally, distally, or even on separate molecules relative to the DSB. They also provide a rationale for the extensive repertoire of repeated sequences in this genome.

  2. Double stranded nucleic acid biochips

    DOEpatents

    Chernov, Boris; Golova, Julia

    2006-05-23

    This invention describes a new method of constructing double-stranded DNA (dsDNA) microarrays based on the use of pre-synthesized or natural DNA duplexes without a stem-loop structure. The complementary oligonucleotide chains are bonded together by a novel connector that includes a linker for immobilization on a matrix. A non-enzymatic method for synthesizing double-stranded nucleic acids with this novel connector enables the construction of inexpensive and robust dsDNA/dsRNA microarrays. DNA-DNA and DNA-protein interactions are investigated using the microarrays.

  3. The (not so) immortal strand hypothesis.

    PubMed

    Tomasetti, Cristian; Bozic, Ivana

    2015-03-01

    Non-random segregation of DNA strands during stem cell replication has been proposed as a mechanism to minimize accumulated genetic errors in stem cells of rapidly dividing tissues. According to this hypothesis, an "immortal" DNA strand is passed to the stem cell daughter and not the more differentiated cell, keeping the stem cell lineage replication error-free. After it was introduced, experimental evidence both in favor and against the hypothesis has been presented. Using a novel methodology that utilizes cancer sequencing data we are able to estimate the rate of accumulation of mutations in healthy stem cells of the colon, blood and head and neck tissues. We find that in these tissues mutations in stem cells accumulate at rates strikingly similar to those expected without the protection from the immortal strand mechanism. Utilizing an approach that is fundamentally different from previous efforts to confirm or refute the immortal strand hypothesis, we provide evidence against non-random segregation of DNA during stem cell replication. Our results strongly suggest that parental DNA is passed randomly to stem cell daughters and provides new insight into the mechanism of DNA replication in stem cells. Copyright © 2015. Published by Elsevier B.V.

  4. The (not so) Immortal Strand Hypothesis

    PubMed Central

    Tomasetti, Cristian; Bozic, Ivana

    2015-01-01

    Background Non-random segregation of DNA strands during stem cell replication has been proposed as a mechanism to minimize accumulated genetic errors in stem cells of rapidly dividing tissues. According to this hypothesis, an “immortal” DNA strand is passed to the stem cell daughter and not the more differentiated cell, keeping the stem cell lineage replication error-free. After it was introduced, experimental evidence both in favor and against the hypothesis has been presented. Principal Findings Using a novel methodology that utilizes cancer sequencing data we are able to estimate the rate of accumulation of mutations in healthy stem cells of the colon, blood and head and neck tissues. We to find that in these tissues mutations in stem cells accumulate at rates strikingly similar to those expected without the protection from the immortal strand mechanism. Significance Utilizing an approach that is fundamentally different from previous efforts to confirm or refute the immortal strand hypothesis, we provide strong evidence against non-random segregation of DNA during stem cell replication. Our results strongly suggest that parental DNA is passed randomly to stem cell daughters and provides new insight into the mechanism of DNA replication in stem cells. PMID:25700960

  5. Constraining Anisotropic Lorentz Violation via the Spectral-lag Transition of GRB 160625B

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

    Wei, Jun-Jie; Wu, Xue-Feng; Shao, Lang

    Violations of Lorentz invariance can lead to an energy-dependent vacuum dispersion of light, which results in arrival-time differences of photons with different energies arising from a given transient source. In this work, direction-dependent dispersion constraints are obtained on nonbirefringent Lorentz-violating effects using the observed spectral lags of the gamma-ray burst GRB 160625B. This burst has unusually large high-energy photon statistics, so we can obtain constraints from the true spectral time lags of bunches of high-energy photons rather than from the rough time lag of a single highest-energy photon. Also, GRB 160625B is the only burst to date having a well-definedmore » transition from positive lags to negative lags, providing a unique opportunity to distinguish Lorentz-violating effects from any source-intrinsic time lag in the emission of photons of different energy bands. Our results place comparatively robust two-sided constraints on a variety of isotropic and anisotropic coefficients for Lorentz violation, including the first bounds on Lorentz-violating effects from operators of mass dimension 10 in the photon sector.« less

  6. The effect of control and display lag on unmanned air system internal pilot manual landing performance

    NASA Astrophysics Data System (ADS)

    Lloyd, Marshall Everett

    An important characteristic of UASs is lag because it can become a considerable challenge to successful human-in-the-loop control. As such, UASs are designed and configured to minimize system lag, though this can increase acquisition and operation costs considerably. In an effort to cut costs, an organization may choose to accept greater risk and deploy a UAS with high system lag. Before this risk can be responsibly accepted, it must be quantified. While many studies have examined system lag, very few have been able to quantify the risk that various levels of lag pose to an internally piloted, manually landed UAS. This study attempted to do so by evaluating pilot landing performance in a simulator with 0 ms, 240 ms, and 1000 ms of additional lag. Various measures were used, including a novel coding technique. Results indicated that 1000 ms of lag was unsafe by all measures. They also indicate that 240 ms of lag degrades performance, but participants were able to successfully land the simulated aircraft. This study showed the utility of using several measures to evaluate the effect of lag on landing performance and it helped demonstrate that while 1000 ms poses a high risk, 240 ms of lag may be a much more manageable risk. Future research suggested by this research includes: investigating lag between 240 ms and 1000 ms, introducing different weather phenomena, developing system lag training techniques for operators, and investigating the effect of aides such as predictive displays and autopilot-assisted recovery.

  7. Method of preparing and applying single stranded DNA probes to double stranded target DNAs in situ

    DOEpatents

    Gray, J.W.; Pinkel, D.

    1991-07-02

    A method is provided for producing single stranded non-self-complementary nucleic acid probes, and for treating target DNA for use therewith. The probe is constructed by treating DNA with a restriction enzyme and an exonuclease to form template/primers for a DNA polymerase. The digested strand is resynthesized in the presence of labeled nucleoside triphosphate precursor. Labeled single stranded fragments are separated from the resynthesized fragments to form the probe. Target DNA is treated with the same restriction enzyme used to construct the probe, and is treated with an exonuclease before application of the probe. The method significantly increases the efficiency and specificity of hybridization mixtures by increasing effective probe concentration by eliminating self-hybridization between both probe and target DNAs, and by reducing the amount of target DNA available for mismatched hybridizations. No Drawings

  8. An intercalation-locked parallel-stranded DNA tetraplex

    DOE PAGES

    Tripathi, S.; Zhang, D.; Paukstelis, P. J.

    2015-01-27

    DNA has proved to be an excellent material for nanoscale construction because complementary DNA duplexes are programmable and structurally predictable. However, in the absence of Watson–Crick pairings, DNA can be structurally more diverse. Here, we describe the crystal structures of d(ACTCGGATGAT) and the brominated derivative, d(AC BrUCGGA BrUGAT). These oligonucleotides form parallel-stranded duplexes with a crystallographically equivalent strand, resulting in the first examples of DNA crystal structures that contains four different symmetric homo base pairs. Two of the parallel-stranded duplexes are coaxially stacked in opposite directions and locked together to form a tetraplex through intercalation of the 5'-most A–A basemore » pairs between adjacent G–G pairs in the partner duplex. The intercalation region is a new type of DNA tertiary structural motif with similarities to the i-motif. 1H– 1H nuclear magnetic resonance and native gel electrophoresis confirmed the formation of a parallel-stranded duplex in solution. Finally, we modified specific nucleotide positions and added d(GAY) motifs to oligonucleotides and were readily able to obtain similar crystals. This suggests that this parallel-stranded DNA structure may be useful in the rational design of DNA crystals and nanostructures.« less

  9. Template-switching during DNA synthesis by Thermus aquaticus DNA polymerase I.

    PubMed Central

    Odelberg, S J; Weiss, R B; Hata, A; White, R

    1995-01-01

    Recombinant DNA molecules are often generated during the polymerase chain reaction (PCR) when partially homologous templates are available [e.g., see Pääbo et al. (1990) J. Biol. Chem. 265, 4718-4721]. It has been suggested that these recombinant molecules are a consequence of truncated extension products annealing to partially homologous templates on subsequent PCR cycles. However, we demonstrate here that recombinants can be generated during a single round of primer extension in the absence of subsequent heat denaturation, indicating that template-switching produces some of these recombinant molecules. Two types of template-switches were observed: (i) switches to pre-existing templates and (ii) switches to the complementary nascent strand. Recombination is reduced several fold when the complementary template strands are physically separated by attachment to streptavidin magnetic beads. This result supports the hypothesis that either the polymerase or at least one of the two extending strands switches templates during DNA synthesis and that interaction between the complementary template strands is necessary for efficient template-switching. Images PMID:7596836

  10. AN INVESTIGATION OF TIME LAG MAPS USING THREE-DIMENSIONAL SIMULATIONS OF HIGHLY STRATIFIED HEATING

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

    Winebarger, Amy R.; Lionello, Roberto; Downs, Cooper

    2016-11-10

    The location and frequency of coronal energy release provide a significant constraint on the coronal heating mechanism. The evolution of the intensity observed in coronal structures found from time lag analysis of Atmospheric Imaging Assembly (AIA) data has been used to argue that heating must occur sporadically. Recently, we have demonstrated that quasi-steady, highly stratified (footpoint) heating can produce results qualitatively consistent with the evolution of observed coronal structures. The goals of this paper are to demonstrate that time lag analysis of 3D simulations of footpoint heating are qualitatively consistent with time lag analysis of observations and to use themore » 3D simulations to further understand whether time lag analysis is a useful tool in defining the evolution of coronal structures. We find the time lag maps generated from simulated data are consistent with the observed time lag maps. We next investigate several example points. In some cases, the calculated time lag reflects the evolution of a unique loop along the line of sight, though there may be additional evolving structures along the line of sight. We confirm that using the multi-peak AIA channels can produce time lags that are difficult to interpret. We suggest using a different high temperature channel, such as an X-ray channel. Finally, we find that multiple evolving structures along the line of sight can produce time lags that do not represent the physical properties of any structure along the line of sight, although the cross-correlation coefficient of the lightcurves is high. Considering the projected geometry of the loops may reduce some of the line-of-sight confusion.« less

  11. DNA strand displacement system running logic programs.

    PubMed

    Rodríguez-Patón, Alfonso; Sainz de Murieta, Iñaki; Sosík, Petr

    2014-01-01

    The paper presents a DNA-based computing model which is enzyme-free and autonomous, not requiring a human intervention during the computation. The model is able to perform iterated resolution steps with logical formulae in conjunctive normal form. The implementation is based on the technique of DNA strand displacement, with each clause encoded in a separate DNA molecule. Propositions are encoded assigning a strand to each proposition p, and its complementary strand to the proposition ¬p; clauses are encoded comprising different propositions in the same strand. The model allows to run logic programs composed of Horn clauses by cascading resolution steps. The potential of the model is demonstrated also by its theoretical capability of solving SAT. The resulting SAT algorithm has a linear time complexity in the number of resolution steps, whereas its spatial complexity is exponential in the number of variables of the formula. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  12. The immortal strand hypothesis: still non-randomly segregating opinions.

    PubMed

    Wakeman, Jane A; Hmadcha, Abdelkrim; Soria, Bernat; McFarlane, Ramsay J

    2012-06-01

    Abstract Cairns first suggested a mechanism for protecting the genomes of stem cells (SCs) from replicative errors some 40 years ago when he proposed the immortal strand hypothesis, which argued for the inheritance of a so-called immortal strand by an SC following asymmetric SC divisions. To date, the existence of immortal strands remains contentious with published evidence arguing in favour of and against the retention of an immortal strand by asymmetrically dividing SCs. The conflicting evidence is derived from a diverse array of studies on adult SC types and is predominantly based on following the fate of labelled DNA strands during asymmetric cell division events. Here, we review current data, highlighting limitations of such labelling techniques, and suggest how interpretation of such data may be improved in the future.

  13. Enzymatic Production of Monoclonal Stoichiometric Single-Stranded DNA Oligonucleotides

    PubMed Central

    Ducani, Cosimo; Kaul, Corinna; Moche, Martin; Shih, William M.; Högberg, Björn

    2013-01-01

    Single-stranded oligonucleotides are important as research tools as probes for diagnostics and gene therapy. Today, production of oligonucleotides is done via solid-phase synthesis. However, the capabilities of current polymer chemistry are limited in comparison to what can be produced in biological systems. The errors in synthetic DNA increases with oligonucleotide length, and sequence diversity can often be a problem. Here, we present the Monoclonal Stoichiometric (MOSIC) method for enzymatic DNA oligonucleotide production. Using this method, we amplify oligonucleotides from clonal templates followed by digestion of a cutter-hairpin, resulting in pools of monoclonal oligonucleotides with precisely controlled relative stoichiometric ratios. We present data where MOSIC oligonucleotides, 14–378 nt long, were prepared either by in vitro rolling-circle amplification, or by amplification in Escherichia coli in the form of phagemid DNA. The formation of a DNA crystal and folding of DNA nanostructures confirmed the scalability, purity and stoichiometry of the produced oligonucleotides. PMID:23727986

  14. 46 CFR 98.25-30 - Lagging.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 4 2012-10-01 2012-10-01 false Lagging. 98.25-30 Section 98.25-30 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) CARGO AND MISCELLANEOUS VESSELS SPECIAL CONSTRUCTION, ARRANGEMENT, AND OTHER PROVISIONS FOR CERTAIN DANGEROUS CARGOES IN BULK Anhydrous Ammonia in Bulk § 98.25-30...

  15. 46 CFR 98.25-30 - Lagging.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 4 2011-10-01 2011-10-01 false Lagging. 98.25-30 Section 98.25-30 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) CARGO AND MISCELLANEOUS VESSELS SPECIAL CONSTRUCTION, ARRANGEMENT, AND OTHER PROVISIONS FOR CERTAIN DANGEROUS CARGOES IN BULK Anhydrous Ammonia in Bulk § 98.25-30...

  16. 46 CFR 98.25-30 - Lagging.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 4 2010-10-01 2010-10-01 false Lagging. 98.25-30 Section 98.25-30 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) CARGO AND MISCELLANEOUS VESSELS SPECIAL CONSTRUCTION, ARRANGEMENT, AND OTHER PROVISIONS FOR CERTAIN DANGEROUS CARGOES IN BULK Anhydrous Ammonia in Bulk § 98.25-30...

  17. 46 CFR 98.25-30 - Lagging.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 4 2013-10-01 2013-10-01 false Lagging. 98.25-30 Section 98.25-30 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) CARGO AND MISCELLANEOUS VESSELS SPECIAL CONSTRUCTION, ARRANGEMENT, AND OTHER PROVISIONS FOR CERTAIN DANGEROUS CARGOES IN BULK Anhydrous Ammonia in Bulk § 98.25-30...

  18. 46 CFR 98.25-30 - Lagging.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 4 2014-10-01 2014-10-01 false Lagging. 98.25-30 Section 98.25-30 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) CARGO AND MISCELLANEOUS VESSELS SPECIAL CONSTRUCTION, ARRANGEMENT, AND OTHER PROVISIONS FOR CERTAIN DANGEROUS CARGOES IN BULK Anhydrous Ammonia in Bulk § 98.25-30...

  19. Tissue strands as "bioink" for scale-up organ printing.

    PubMed

    Yu, Yin; Ozbolat, Ibrahim T

    2014-01-01

    Organ printing, takes tissue spheroids as building blocks together with additive manufacturing technique to engineer tissue or organ replacement parts. Although a wide array of cell aggregation techniques has been investigated, and gained noticeable success, the application of tissue spheroids for scale-up tissue fabrication is still worth investigation. In this paper, we introduce a new micro-fabrication technique to create tissue strands at the scale of 500-700μm as a "bioink" for future robotic tissue printing. Printable alginate micro-conduits are used as semi-permeable capsules for tissue strand fabrication. Mouse insulinoma beta TC3 cell tissue strands were formed upon 4 days post fabrication with reasonable mechanical strength, high cell viability close to 90%, and tissue specific markers expression. Fusion was readily observed between strands when placing them together as early as 24h. Also, tissue strands were deposited with human umbilical vein smooth muscle cells (HUVSMCs) vascular conduits together to fabricated miniature pancreatic tissue analog. Our study provided a novel technique using tissue strands as "bioink" for scale-up bioprinting of tissues or organs.

  20. Modeling eye gaze patterns in clinician-patient interaction with lag sequential analysis.

    PubMed

    Montague, Enid; Xu, Jie; Chen, Ping-Yu; Asan, Onur; Barrett, Bruce P; Chewning, Betty

    2011-10-01

    The aim of this study was to examine whether lag sequential analysis could be used to describe eye gaze orientation between clinicians and patients in the medical encounter. This topic is particularly important as new technologies are implemented into multiuser health care settings in which trust is critical and nonverbal cues are integral to achieving trust. This analysis method could lead to design guidelines for technologies and more effective assessments of interventions. Nonverbal communication patterns are important aspects of clinician-patient interactions and may affect patient outcomes. The eye gaze behaviors of clinicians and patients in 110 videotaped medical encounters were analyzed using the lag sequential method to identify significant behavior sequences. Lag sequential analysis included both event-based lag and time-based lag. Results from event-based lag analysis showed that the patient's gaze followed that of the clinician, whereas the clinician's gaze did not follow the patient's. Time-based sequential analysis showed that responses from the patient usually occurred within 2 s after the initial behavior of the clinician. Our data suggest that the clinician's gaze significantly affects the medical encounter but that the converse is not true. Findings from this research have implications for the design of clinical work systems and modeling interactions. Similar research methods could be used to identify different behavior patterns in clinical settings (physical layout, technology, etc.) to facilitate and evaluate clinical work system designs.

  1. Modeling Eye Gaze Patterns in Clinician-Patient Interaction with Lag Sequential Analysis

    PubMed Central

    Montague, E; Xu, J; Asan, O; Chen, P; Chewning, B; Barrett, B

    2011-01-01

    Objective The aim of this study was to examine whether lag-sequential analysis could be used to describe eye gaze orientation between clinicians and patients in the medical encounter. This topic is particularly important as new technologies are implemented into multi-user health care settings where trust is critical and nonverbal cues are integral to achieving trust. This analysis method could lead to design guidelines for technologies and more effective assessments of interventions. Background Nonverbal communication patterns are important aspects of clinician-patient interactions and may impact patient outcomes. Method Eye gaze behaviors of clinicians and patients in 110-videotaped medical encounters were analyzed using the lag-sequential method to identify significant behavior sequences. Lag-sequential analysis included both event-based lag and time-based lag. Results Results from event-based lag analysis showed that the patients’ gaze followed that of clinicians, while clinicians did not follow patients. Time-based sequential analysis showed that responses from the patient usually occurred within two seconds after the initial behavior of the clinician. Conclusion Our data suggest that the clinician’s gaze significantly affects the medical encounter but not the converse. Application Findings from this research have implications for the design of clinical work systems and modeling interactions. Similar research methods could be used to identify different behavior patterns in clinical settings (physical layout, technology, etc.) to facilitate and evaluate clinical work system designs. PMID:22046723

  2. Effect of Escherichia coli DNA binding protein on the transcription of single-stranded phage M13 DNA by Escherichia coli RNA polymerase

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

    Niyogi, S.K.; Ratrie, H. III; Datta, A.K.

    E. coli DNA binding protein strongly inhibits the transcription of single-stranded rather than double-stranded phage M13 DNA by E. coli RNA polymerase. This inhibition cannot be significantly overcome by increasing the concentration of RNA polymerase. Nor does the order of addition of binding protein affect its inhibitory property: inhibition is evident whether binding protein is added before or after the formation of the RNA polymerase--DNA complex. Inhibition is also observed if binding protein is added at various times after initiation of RNA synthesis. Maximal inhibition occurs at a binding protein-to-DNA ratio (w/w) of about 8:1. This corresponds to one bindingmore » protein molecule covering about 30 nucleotides, in good agreement with values obtained by physical measurements.« less

  3. A new method for determining the optimal lagged ensemble

    PubMed Central

    DelSole, T.; Tippett, M. K.; Pegion, K.

    2017-01-01

    Abstract We propose a general methodology for determining the lagged ensemble that minimizes the mean square forecast error. The MSE of a lagged ensemble is shown to depend only on a quantity called the cross‐lead error covariance matrix, which can be estimated from a short hindcast data set and parameterized in terms of analytic functions of time. The resulting parameterization allows the skill of forecasts to be evaluated for an arbitrary ensemble size and initialization frequency. Remarkably, the parameterization also can estimate the MSE of a burst ensemble simply by taking the limit of an infinitely small interval between initialization times. This methodology is applied to forecasts of the Madden Julian Oscillation (MJO) from version 2 of the Climate Forecast System version 2 (CFSv2). For leads greater than a week, little improvement is found in the MJO forecast skill when ensembles larger than 5 days are used or initializations greater than 4 times per day. We find that if the initialization frequency is too infrequent, important structures of the lagged error covariance matrix are lost. Lastly, we demonstrate that the forecast error at leads ≥10 days can be reduced by optimally weighting the lagged ensemble members. The weights are shown to depend only on the cross‐lead error covariance matrix. While the methodology developed here is applied to CFSv2, the technique can be easily adapted to other forecast systems. PMID:28580050

  4. Telomeric 3′ overhangs derive from resection by Exo1 and Apollo and fill-in by POT1b-associated CST

    PubMed Central

    Wu, Peng; Takai, Hiroyuki; de Lange, Titia

    2012-01-01

    SUMMARY A 3′ overhang is critical for the protection and maintenance of mammalian telomeres. How these overhangs are generated and whether different processing steps modify telomeres synthesized by leading- and lagging-strand DNA replication was not known. Here we evaluate changes in the telomeric overhangs through the cell cycle and at leading- and lagging-end telomeres in mouse cells lacking relevant genes. Apollo, a nuclease bound to the shelterin subunit TRF2, initiated formation of the 3′ overhang at leading-, but not lagging-end telomeres. Hyper-resection by Apollo was blocked at both ends by the shelterin protein POT1b. Exo1 extensively resected both telomere ends, generating long 3′ overhangs that transiently occurred in S/G2. CST/AAF, a DNA polymeraseα. primase accessory factor related to yeast CST, bound POT1b and shortened the extended overhangs produced by Exo1, most likely through fill-in synthesis. The results establish 3′ overhang formation as a multi-step, shelterin-controlled process that ensures functional telomeric overhangs at all chromosome ends. PMID:22748632

  5. A methodology to identify stranded generation facilities and estimate stranded costs for Louisiana's electric utility industry

    NASA Astrophysics Data System (ADS)

    Cope, Robert Frank, III

    1998-12-01

    The electric utility industry in the United States is currently experiencing a new and different type of growing pain. It is the pain of having to restructure itself into a competitive business. Many industry experts are trying to explain how the nation as a whole, as well as individual states, will implement restructuring and handle its numerous "transition problems." One significant transition problem for federal and state regulators rests with determining a utility's stranded costs. Stranded generation facilities are assets which would be uneconomic in a competitive environment or costs for assets whose regulated book value is greater than market value. At issue is the methodology which will be used to estimate stranded costs. The two primary methods are known as "Top-Down" and "Bottom-Up." The "Top-Down" approach simply determines the present value of the losses in revenue as the market price for electricity changes over a period of time into the future. The problem with this approach is that it does not take into account technical issues associated with the generation and wheeling of electricity. The "Bottom-Up" approach computes the present value of specific strandable generation facilities and compares the resulting valuations with their historical costs. It is regarded as a detailed and difficult, but more precise, approach to identifying stranded assets and their associated costs. This dissertation develops a "Bottom-Up" quantitative, optimization-based approach to electric power wheeling within the state of Louisiana. It optimally evaluates all production capabilities and coordinates the movement of bulk power through transmission interconnections of competing companies in and around the state. Sensitivity analysis to this approach is performed by varying seasonal consumer demand, electric power imports, and transmission inter-connection cost parameters. Generation facility economic dispatch and transmission interconnection bulk power transfers, specific

  6. LAG-3 Represents a Marker of CD4+ T Cells with Regulatory Activity in Patients with Bone Fracture.

    PubMed

    Wang, Jun; Ti, Yunfan; Wang, Yicun; Guo, Guodong; Jiang, Hui; Chang, Menghan; Qian, Hongbo; Zhao, Jianning; Sun, Guojing

    2018-04-19

    The lymphocyte activation gene 3 (LAG-3) is a CD4 homolog with binding affinity to MHC class II molecules. It is thought that LAG-3 exerts a bimodal function, such that co-ligation of LAG-3 and CD3 could deliver an inhibitory signal in conventional T cells, whereas, on regulatory T cells, LAG-3 expression could promote their inhibitory function. In this study, we investigated the role of LAG-3 expression on CD4 + T cells in patients with long bone fracture. We found that LAG-3 + cells represented approximately 13% of peripheral blood CD4 + T cells on average. Compared to LAG-3 - CD4 + T cells, LAG-3 + CD4 + T cells presented significantly higher Foxp3 and CTLA-4 expression. Directly ex vivo or with TCR stimulation, LAG-3 + CD4 + T cells expressed significantly higher levels of IL-10 and TGF-β than LAG-3 - CD4 + T cells. Interestingly, blocking the LAG-3-MHC class II interaction actually increased the IL-10 expression by LAG-3 + CD4 + T cells. The frequency of LAG-3 + CD4 + T cell was positively correlated with restoration of healthy bone function in long bone fracture patients. These results together suggested that LAG-3 is a marker of CD4 + T cells with regulatory function; at the same time, LAG-3 might have limited the full suppressive potential of Treg cells.

  7. Pyrrole-Imidazole Polyamides: Manual Solid-Phase Synthesis.

    PubMed

    Pauff, Steven M; Fallows, Andrew J; Mackay, Simon P; Su, Wu; Cullis, Paul M; Burley, Glenn A

    2015-12-01

    Pyrrole-imidazole polyamides (PAs) are a family of DNA-binding peptides that bind in the minor groove of double-stranded DNA (dsDNA) in a sequence-selective, programmable fashion. This protocol describes a detailed manual procedure for the solid-phase synthesis of this family of compounds. The protocol entails solution-phase synthesis of the Boc-protected pyrrole (Py) and imidazole (Im) carboxylic acid building blocks. This unit also describes the importance of choosing the appropriate condensing agent to form the amide linkages between each building block. Finally, a monomeric coupling protocol and a fragment-based approach are described that delivers PAs in 13% to 30% yield in 8 days. Copyright © 2015 John Wiley & Sons, Inc.

  8. Increased Variability in Tacting under a Lag 3 Schedule of Reinforcement

    ERIC Educational Resources Information Center

    Heldt, Juliane; Schlinger, Henry D., Jr.

    2012-01-01

    Research has shown that variability may be an operant dimension of behavior. One method of reinforcing response variability is to use a lag schedule of reinforcement (Page & Neuringer, 1985). Several studies have shown that a Lag 1 schedule is effective in increasing variable responding with human participants (e.g., Esch, Esch, & Love, 2009; Lee,…

  9. The Lag-Luminosity Relation in the GRB Source-Frame: An Investigation with Swift BAT Bursts

    NASA Technical Reports Server (NTRS)

    Ukwatta, T. N.; Dhuga, K. S.; Stamatikos, M.; Dermer, C. D.; Sakamoto, T.; Sonbas, E.; Parke, W. C.; Maximon, L. C.; Linnemann, J. T.; Bhat, P. N.; hide

    2012-01-01

    Spectral lag, which is defined as the difference in time of arrival of high and low energy photons, is a common feature in Gamma-ray Bursts (GRBs). Previous investigations have shown a correlation between this lag and the isotropic peak luminosity for long duration bursts. However, most of the previous investigations used lags extracted in the observer-frame only. In this work (based on a sample of 43 Swift long GRBs with known redshifts), we present an analysis of the lag-luminosity relation in the GRB source-frame. Our analysis indicates a higher degree of correlation -0.82+/-0.05 (chance probability of approx 5.5 X 10(exp -5) between the spectral lag and the isotropic peak luminosity, L(sub iso), with a best-fit power-law index of -1.2 +/- 0.2, such that L(sub iso) varies as lag(exp -1.2). In addition, there is an anti-correlation between the source-frame spectral lag and the source-frame peak energy of the burst spectrum, E(sub pk)(1 + z).

  10. Yeast multistress resistance and lag-phase characterisation during wine fermentation.

    PubMed

    Ferreira, David; Galeote, Virginie; Sanchez, Isabelle; Legras, Jean-Luc; Ortiz-Julien, Anne; Dequin, Sylvie

    2017-09-01

    Saccharomyces cerevisiae has been used to perform wine fermentation for several millennia due to its endurance and unmatched qualities. Nevertheless, at the moment of inoculation, wine yeasts must cope with specific stress factors that still challenge wine makers by slowing down or compromising the fermentation process. To better assess the role of genetic and environmental factors that govern multistress resistance during the wine fermentation lag phase, we used a factorial plan to characterise the individual and combined impact of relevant stress factors on eight S. cerevisiae and two non-S. cerevisiae wine yeast strains that are currently commercialised. The S. cerevisiae strains are very genetically diverse, belonging to the wine and flor groups, and frequently contain a previously described XVIVIII translocation that confers increased resistance to sulphites. We found that low temperature and osmotic stress substantially affected all strains, promoting considerably extended lag phases. SO2 addition had a partially temperature-dependent effect, whereas low phytosterol and thiamine concentrations impacted the lag phase in a strain-dependent manner. No major synergic effects of multistress were detected. The diversity of lag-phase durations and stress responses observed among wine strains offer new insights to better control this critical step of fermentation. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  11. Lag and seasonality considerations in evaluating AVHRR NDVI response to precipitation

    USGS Publications Warehouse

    Ji, Lei; Peters, Albert J.

    2005-01-01

    Assessment of the relationship between the normalized difference vegetation index (NDVI) and precipitation is important in understanding vegetation and climate interaction at a large scale. NDVI response to precipitation, however, is difficult to quantify due to the lag and seasonality effects, which will vary due to vegetation cover type, soils and climate. A time series analysis was performed on biweekly NDVI and precipitation around weather stations in the northern and central U.S. Great Plains. Regression models that incorporate lag and seasonality effects were used to quantify the relationship between NDVI and lagged precipitation in grasslands and croplands. It was found that the time lag was shorter in the early growing season, but longer in the mid- to late-growing season for most locations. The regression models with seasonal adjustment indicate that the relationship between NDVI and precipitation over the entire growing season was strong, with R2 values of 0.69 and 0.72 for grasslands and croplands, respectively. We conclude that vegetation greenness can be predicted using current and antecedent precipitation, if seasonal effects are taken into account.

  12. A cascade autocatalytic strand displacement amplification and hybridization chain reaction event for label-free and ultrasensitive electrochemical nucleic acid biosensing.

    PubMed

    Chen, Zhiqiang; Liu, Ying; Xin, Chen; Zhao, Jikuan; Liu, Shufeng

    2018-08-15

    Herein, an autocatalytic strand displacement amplification (ASDA) strategy was proposed for the first time, which was further ingeniously coupled with hybridization chain reaction (HCR) event for the isothermal, label-free and multiple amplification toward nucleic acid detection. During the ASDA module, the target recognition opens the immobilized hairpin probe (IP) and initiates the annealing of the auxiliary DNA strand (AS) with the opened IP for the successive polymerization and nicking reaction in the presence of DNA polymerase and nicking endonuclease. This induces the target recycling and generation of a large amount of intermediate DNA sequences, which can be used as target analogy to execute the autocatalytic strand displacement amplification. Simultaneously, the introduced AS strand can propagate the HCR between two hairpins (H1 and H2) to form a linear DNA concatamer with cytosine (C)-rich loop region, which can facilitate the in-situ synthesis of silver nanoclusters (AgNCs) as electrochemical tags for further amplification toward target responses. With current cascade ASDA and HCR strategy, the detection of target DNA could be achieved with a low detection limit of about 0.16 fM and a good selectivity. The developed biosensor also exhibits the distinct advantages of flexibility and simplicity in probe design and biosensor fabrication, and label-free electrochemical detection, thus opens a promising avenue for the detection of nucleic acid with low abundance in bioanalysis and clinical biomedicine. Copyright © 2018 Elsevier B.V. All rights reserved.

  13. Hydrophilic excipients modulate the time lag of time-controlled disintegrating press-coated tablets.

    PubMed

    Lin, Shan-Yang; Li, Mei-Jane; Lin, Kung-Hsu

    2004-08-16

    An oral press-coated tablet was developed by means of direct compression to achieve the time-controlled disintegrating or rupturing function with a distinct predetermined lag time. This press-coated tablet containing sodium diclofenac in the inner core was formulated with an outer shell by different weight ratios of hydrophobic polymer of micronized ethylcellulose (EC) powder and hydrophilic excipients such as spray-dried lactose (SDL) or hydroxypropyl methylcellulose (HPMC). The effect of the formulation of an outer shell comprising both hydrophobic polymer and hydrophilic excipients on the time lag of drug release was investigated. The release profile of the press-coated tablet exhibited a time period without drug release (time lag) followed by a rapid and complete release phase, in which the outer shell ruptured or broke into 2 halves. The lag phase was markedly dependent on the weight ratios of EC/SDL or EC/HPMC in the outer shell. Different time lags of the press-coated tablets from 1.0 to 16.3 hours could be modulated by changing the type and amount of the excipients. A semilogarithmic plot of the time lag of the tablet against the weight ratios of EC/SDL or EC/HPMC in the outer shell demonstrated a good linear relationship, with r = 0.976 and r = 0.982, respectively. The predetermined time lag prior to the drug release from a press-coated tablet prepared by using a micronized EC as a retarding coating shell can be adequately scheduled with the addition of hydrophilic excipients according to the time or site requirements.

  14. Assessing protein conformational sampling methods based on bivariate lag-distributions of backbone angles

    PubMed Central

    Maadooliat, Mehdi; Huang, Jianhua Z.

    2013-01-01

    Despite considerable progress in the past decades, protein structure prediction remains one of the major unsolved problems in computational biology. Angular-sampling-based methods have been extensively studied recently due to their ability to capture the continuous conformational space of protein structures. The literature has focused on using a variety of parametric models of the sequential dependencies between angle pairs along the protein chains. In this article, we present a thorough review of angular-sampling-based methods by assessing three main questions: What is the best distribution type to model the protein angles? What is a reasonable number of components in a mixture model that should be considered to accurately parameterize the joint distribution of the angles? and What is the order of the local sequence–structure dependency that should be considered by a prediction method? We assess the model fits for different methods using bivariate lag-distributions of the dihedral/planar angles. Moreover, the main information across the lags can be extracted using a technique called Lag singular value decomposition (LagSVD), which considers the joint distribution of the dihedral/planar angles over different lags using a nonparametric approach and monitors the behavior of the lag-distribution of the angles using singular value decomposition. As a result, we developed graphical tools and numerical measurements to compare and evaluate the performance of different model fits. Furthermore, we developed a web-tool (http://www.stat.tamu.edu/∼madoliat/LagSVD) that can be used to produce informative animations. PMID:22926831

  15. Trans-activation of the 5' to 3' viral DNA strand transfer by nucleocapsid protein during reverse transcription of HIV1 RNA.

    PubMed

    Darlix, J L; Vincent, A; Gabus, C; de Rocquigny, H; Roques, B

    1993-08-01

    Two DNA strand transfer reactions take place during reverse transcription of the retroviral genome. The first transfer, that of the minus-strand strong stop DNA from the 5' end of the viral RNA to the 3' end, has been studied in vitro with two RNAs mimicking the 5' and 3' regions of the HIV1 genome and with nucleocapsid protein, NCp7, and reverse transcriptase. The results show that NCp7 strongly activates the 5' to 3' DNA strand transfer during reverse transcription while a basic peptide resembling NCp7 is inactive. Activation of the first transfer by several NCp7 derived peptides and the influence of the terminal redundancies (R) present at the 5' and 3' ends of HIV1 RNA were also examined. The first transfer is optimal in the presence of intact NCp7 and necessitates R on both the 5' and 3' RNAs. Sequencing of full length viral DNA products reveals approximately 40% misincorporations at the first nucleotide beyond the transfer point. If such base misincorporations occur during proviral DNA synthesis with possible homologous recombinations it may well contribute to the high level of genetic variability of HIV.

  16. Convergent strand array liquid pumping system

    NASA Technical Reports Server (NTRS)

    Collins, Earl R., Jr. (Inventor)

    1989-01-01

    A surface-tension liquid pumping system is provided by one or more arrays of converging solid monofilament fibers or metal wires (strands) spaced apart at an input end to gather liquid, and gathered close together at the opposite end where menisci forms between wetted strands to force liquid in the direction of convergence of the strands. The liquid pumping system is independent of gravity. It is illustrated as being used in a heat pump having a heating box to vaporize the liquid and a condensing chamber. Condensed liquid is returned by the pumping system to the heating box where it is again vaporized. A vapor tube carries the vapor to the condensing chamber. In that way, a closed system pumps heat from the heating box to the evaporating chamber and from there radiated to the atmosphere.

  17. Strand development and splice device : final report, February 3, 2009.

    DOT National Transportation Integrated Search

    2010-02-01

    "A new device for gripping prestressing strands was developed and tested. The device could provide a means of anchoring the terminal end of a strand in order to provide a mechanism for developing bonded strand at the service limit state, to provide t...

  18. DNA damage bypass operates in the S and G2 phases of the cell cycle and exhibits differential mutagenicity

    PubMed Central

    Diamant, Noam; Hendel, Ayal; Vered, Ilan; Carell, Thomas; Reißner, Thomas; de Wind, Niels; Geacinov, Nicholas; Livneh, Zvi

    2012-01-01

    Translesion DNA synthesis (TLS) employs low-fidelity DNA polymerases to bypass replication-blocking lesions, and being associated with chromosomal replication was presumed to occur in the S phase of the cell cycle. Using immunostaining with anti-replication protein A antibodies, we show that in UV-irradiated mammalian cells, chromosomal single-stranded gaps formed in S phase during replication persist into the G2 phase of the cell cycle, where their repair is completed depending on DNA polymerase ζ and Rev1. Analysis of TLS using a high-resolution gapped-plasmid assay system in cell populations enriched by centrifugal elutriation for specific cell cycle phases showed that TLS operates both in S and G2. Moreover, the mutagenic specificity of TLS in G2 was different from S, and in some cases overall mutation frequency was higher. These results suggest that TLS repair of single-stranded gaps caused by DNA lesions can lag behind chromosomal replication, is separable from it, and occurs both in the S and G2 phases of the cell cycle. Such a mechanism may function to maintain efficient replication, which can progress despite the presence of DNA lesions, with TLS lagging behind and patching regions of discontinuity. PMID:21908406

  19. Measuring attention using flash-lag effect.

    PubMed

    Shioiri, Satoshi; Yamamoto, Ken; Oshida, Hiroki; Matsubara, Kazuya; Yaguchi, Hirohisa

    2010-08-13

    We investigated the effect of attention on the flash-lag effect (FLE) in order to determine whether the FLE can be used to estimate the effect of visual attention. The FLE is the effect that a flash aligned with a moving object is perceived to lag the moving object, and several studies have shown that attention reduces its magnitude. We measured the FLE as a function of the number or speed of moving objects. The results showed that the effect of cueing, which we attributed the effect of attention, on the FLE increased monotonically with the number or the speed of the objects. This suggests that the amount of attention can be estimated by measuring the FLE, assuming that more amount of attention is required for a larger number or faster speed of objects to attend. On the basis of this presumption, we attempted to measure the spatial spread of visual attention by FLE measurements. The estimated spatial spreads were similar to those estimated by other experimental methods.

  20. Control of DNA strand displacement kinetics using toehold exchange.

    PubMed

    Zhang, David Yu; Winfree, Erik

    2009-12-02

    DNA is increasingly being used as the engineering material of choice for the construction of nanoscale circuits, structures, and motors. Many of these enzyme-free constructions function by DNA strand displacement reactions. The kinetics of strand displacement can be modulated by toeholds, short single-stranded segments of DNA that colocalize reactant DNA molecules. Recently, the toehold exchange process was introduced as a method for designing fast and reversible strand displacement reactions. Here, we characterize the kinetics of DNA toehold exchange and model it as a three-step process. This model is simple and quantitatively predicts the kinetics of 85 different strand displacement reactions from the DNA sequences. Furthermore, we use toehold exchange to construct a simple catalytic reaction. This work improves the understanding of the kinetics of nucleic acid reactions and will be useful in the rational design of dynamic DNA and RNA circuits and nanodevices.

  1. Melatonin for preventing and treating jet lag.

    PubMed

    Herxheimer, A; Petrie, K J

    2001-01-01

    Jet-lag commonly affects air travellers who cross several time zones. It results from the body's internal rhythms being out of step with the day-night cycle at the destination. Melatonin is a pineal hormone that plays a central part in regulating bodily rhythms and has been used as a drug to re-align them with the outside world. To assess the effectiveness of oral melatonin taken in different dosage regimens for alleviating jet-lag after air travel across several time zones. We searched the Cochrane Controlled Trials Register, MEDLINE, EMBASE, PsychLit and Science Citation Index electronically, and the journals 'Aviation, Space and Environmental Medicine' and 'Sleep' by hand. We searched citation lists of relevant studies for other relevant trials. We asked principal authors of relevant studies to tell us about unpublished trials. Reports of adverse events linked to melatonin use outside randomised trials were searched for systematically in 'Side Effects of Drugs' (SED) and SED Annuals, 'Reactions Weekly', MEDLINE, and the adverse drug reactions databases of the WHO Uppsala Monitoring Centre (UMC) and the US Food & Drug Administration. Randomised trials in airline passengers, airline staff or military personnel given oral melatonin, compared with placebo or other medication. Outcome measures should consist of subjective rating of jet-lag or related components, such as subjective wellbeing, daytime tiredness, onset and quality of sleep, psychological functioning, duration of return to normal, or indicators of circadian rhythms. : Ten trials met the inclusion criteria. All compared melatonin with placebo; one in addition compared it with a hypnotic, zolpidem. Nine of the trials were of adequate quality to contribute to the assessment, one had a design fault and could not be used in the assessment. Reports of adverse events outside trials were found through MEDLINE, 'Reactions Weekly', and in the WHO UMC database. : Nine of the ten trials found that melatonin, taken

  2. Strand-invading linear probe combined with unmodified PNA.

    PubMed

    Asanuma, Hiroyuki; Niwa, Rie; Akahane, Mariko; Murayama, Keiji; Kashida, Hiromu; Kamiya, Yukiko

    2016-09-15

    Efficient strand invasion by a linear probe to fluorescently label double-stranded DNA has been implemented by employing a probe and unmodified PNA. As a fluorophore, we utilized ethynylperylene. Multiple ethynylperylene residues were incorporated into the DNA probe via a d-threoninol scaffold. The ethynylperylene did not significantly disrupt hybridization with complementary DNA. The linear probe self-quenched in the absence of target DNA and did not hybridize with PNA. A gel-shift assay revealed that linear probe and PNA combination invaded the central region of double-stranded DNA upon heat-shock treatment to form a double duplex. To further suppress the background emission and increase the stability of the probe/DNA duplex, a probe containing anthraquinones as well as ethynylperylene was synthesized. This probe and PNA invader pair detected an internal sequence in a double-stranded DNA with high sensitivity when heat shock treatment was used. The probe and PNA pair was able to invade at the terminus of a long double-stranded DNA at 40°C at 100mM NaCl concentration. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Sulfolobus chromatin proteins modulate strand displacement by DNA polymerase B1

    PubMed Central

    Sun, Fei; Huang, Li

    2013-01-01

    Strand displacement by a DNA polymerase serves a key role in Okazaki fragment maturation, which involves displacement of the RNA primer of the preexisting Okazaki fragment into a flap structure, and subsequent flap removal and fragment ligation. We investigated the role of Sulfolobus chromatin proteins Sso7d and Cren7 in strand displacement by DNA polymerase B1 (PolB1) from the hyperthermophilic archaeon Sulfolobus solfataricus. PolB1 showed a robust strand displacement activity and was capable of synthesizing thousands of nucleotides on a DNA-primed 72-nt single-stranded circular DNA template. This activity was inhibited by both Sso7d and Cren7, which limited the flap length to 3–4 nt at saturating concentrations. However, neither protein inhibited RNA displacement on an RNA-primed single-stranded DNA minicircle by PolB1. Strand displacement remained sensitive to modulation by the chromatin proteins when PolB1 was in association with proliferating cell nuclear antigen. Inhibition of DNA instead of RNA strand displacement by the chromatin proteins is consistent with the finding that double-stranded DNA was more efficiently bound and stabilized than an RNA:DNA duplex by these proteins. Our results suggest that Sulfolobus chromatin proteins modulate strand displacement by PolB1, permitting efficient removal of the RNA primer while inhibiting excessive displacement of the newly synthesized DNA strand during Okazaki fragment maturation. PMID:23821667

  4. Programmable DNA Hydrogels Assembled from Multidomain DNA Strands.

    PubMed

    Jiang, Huiling; Pan, Victor; Vivek, Skanda; Weeks, Eric R; Ke, Yonggang

    2016-06-16

    Hydrogels are important in biological and medical applications, such as drug delivery and tissue engineering. DNA hydrogels have attracted significant attention due to the programmability and biocompatibility of the material. We developed a series of low-cost one-strand DNA hydrogels self-assembled from single-stranded DNA monomers containing multiple palindromic domains. This new hydrogel design is simple and programmable. Thermal stability, mechanical properties, and loading capacity of these one-strand DNA hydrogels can be readily regulated by simply adjusting the DNA domains. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. 75 FR 4104 - Prestressed Concrete Steel Wire Strand From China

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-01-26

    ... Concrete Steel Wire Strand From China AGENCY: United States International Trade Commission. ACTION... wire strand, provided for in subheading 7312.10.30 of the Harmonized Tariff Schedule of the United... merchandise as PC strand, produced from wire of nonstainless, non-galvanized steel, which is suitable for use...

  6. The Lag-Luminosity Relation in the GRB Source Frame: An Investigation with Swift BAT Bursts

    NASA Technical Reports Server (NTRS)

    Ukwatta, T. N.; Dhuga, K. S.; Stamatikos, M.; Dermer, C. D.; Sakamoto, T.; Sonbas, E.; Parke, W. C.; Maximon, L. C.; Linnemann, J. T.; Bhat, P. N.; hide

    2011-01-01

    Spectral lag. which is defined as the difference in time of arrival of high- and low-energy photons. is a common feature in gamma-ray bursts (GRBs). Previous investigations have shown a correlation between this lag and the isotropic peak luminosity for long duration bursts. However. most of the previous investigations used lags extracted in the observer frame only. In this work (based on a sample of 43 Swift long GRBs with known redshifts). we present an analysis of the lag-luminosity relation in the GRB source frame. Our analysis indicates a higher degree of correlation -0.82 +/- 0.05 (chance probability of approx. 5.5 x 10(exp -5) between the spectral lag and the isotropic peak luminosity, L(sub iso). with a best-fitting power-law index of -1.2 +/- 0.2. In addition, there is an anticorrelation between the source-frame spectral lag and the source-frame peak energy of the burst spectrum.

  7. Stranded investment, prices and privacy factor in FERC rulings

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

    O`Driscoll, M.

    The Federal Energy Regulatory Commission upheld its rejection of United Illuminating Co.`s bid to recover stranded investment costs. Since UI has no wholesale customers, it is a matter best left to state regulators, FERC said. UI`s stranded investment recovery plan was part of the company`s transmission access tariff, which provides for open access transmission service at cost-based rates. FERC ordered UI to delete the stranded investment provisions, saying UI was trying to recover in its wholesale transmission rates the costs of generation facility investments that were incurred to provide service to retail customers that leave its system, reasoning that UImore » was seeking protection from what may be legitimate retail franchise competition, which is a state matter. UI, however, said deleting the stranded investment provision would preclude it from arguing in an individual rate filling under the transmission tariff that stranded investment costs should be borne by the wheeling customer.« less

  8. Mass stranding of wedge-tailed shearwater chicks in Hawaii

    USGS Publications Warehouse

    Work, Thierry M.; Rameyer, Robert

    1999-01-01

    Unusual numbers of wedge-tailed shearwater (Puffinus pacificus) chicks stranded on Oahu (Hawaii, USA) in 1994. Compared to healthy wedge-tailed shearwater (WTSW) chicks, stranded chicks were underweight, dehydrated, leukopenic, lymphopenic, eosinopenic, and heterophilic; some birds were toxemic and septic. Stranded chicks also were hypoglycemic and had elevated aspartate amino transferase levels. Most chicks apparently died from emaciation, dehydration, or bacteremia. Because many birds with bacteremia also had severe necrosis of the gastrointestinal (GI) mucosa associated with bacteria, we suspect the GI tract to be the source of disseminated bacterial infection. The identity of the bacteria was not confirmed. The daily number of chicks stranded was significantly related to average wind speeds, and the mortality coincided with the fledging period for WTSW. Strong southeasterly winds were a distinguishing meteorologic factor in 1994 and contributed to the distribution of stranded chicks on Oahu. More objective data on WTSW demographics would enhance future efforts to determine predisposing causes of WTSW wrecks and their effects on seabird colonies.

  9. A generalized expression for lag-time in the gas-phase permeation of hollow tubes

    NASA Technical Reports Server (NTRS)

    Shah, K. K.; Nelson, H. G.; Johnson, D. L.; Hamaker, F. M.

    1975-01-01

    A generalized expression for the nonsteady-state parameter, lag-time, has been obtained from Fick's second law for gas-phase transport through hollow, cylindrical membranes. This generalized expression is simplified for three limiting cases of practical interest: (1) diffusion controlled transport, (2) phase boundary reaction control at the inlet surface, and (3) phase boundary reaction control at the outlet surface. In all three cases the lag-time expressions were found to be inversely proportional only to the diffusion coefficient and functionally dependent on the membrane radii. Finally, the lag-time expressions were applied to experimentally obtained lag-time data for alpha-phase titanium and alpha-phase iron.

  10. Electronic trigger for capacitive touchscreen and extension of ISO 15781 standard time lag measurements to smartphones

    NASA Astrophysics Data System (ADS)

    Bucher, François-Xavier; Cao, Frédéric; Viard, Clément; Guichard, Frédéric

    2014-03-01

    We present in this paper a novel capacitive device that stimulates the touchscreen interface of a smartphone (or of any imaging device equipped with a capacitive touchscreen) and synchronizes triggering with the DxO LED Universal Timer to measure shooting time lag and shutter lag according to ISO 15781:2013. The device and protocol extend the time lag measurement beyond the standard by including negative shutter lag, a phenomenon that is more and more commonly found in smartphones. The device is computer-controlled, and this feature, combined with measurement algorithms, makes it possible to automatize a large series of captures so as to provide more refined statistical analyses when, for example, the shutter lag of "zero shutter lag" devices is limited by the frame time as our measurements confirm.

  11. The Parameter of Preposition Stranding: A View from Child English

    ERIC Educational Resources Information Center

    Sugisaki, Koji; Snyder, William

    2006-01-01

    In this squib we examine the time course of children's acquisition of English to evaluate the basic insights of Kayne's (1981; 1984) proposals on preposition stranding. Kayne argued that the availability of preposition stranding (P-stranding) in English is parametrically linked to the availability of double object datives and the prepositional…

  12. SINGLE STRAND-CONTAINING REPLICATING MOLECULES OF CIRCULAR MITOCHONDRIAL DNA

    PubMed Central

    Wolstenholme, David R.; Koike, Katsuro; Cochran-Fouts, Patricia

    1973-01-01

    Mitochondrial DNAs (mtDNAs) from Chang rat solid hepatomas and Novikoff rat ascites hepatomas were examined in the electron microscope after preparation by the aqueous and by the formamide protein monolayer techniques. MtDNAs from both tumors were found to include double-forked circular molecules with a form and size suggesting they were replicative intermediates. These molecules were of two classes. In molecules of one class, all three segments were apparently totally double stranded. Molecules of the second class were distinguished by the fact that one of the segments spanning the region between the forks in which replication had occurred (the daughter segments) was either totally single stranded, or contained a single-stranded region associated with one of the forks. Daughter segments of both totally double-stranded and single strand-containing replicating molecules varied in length from about 3 to about 80% of the circular contour length of the molecule. Similar classes of replicating molecules were found in mtDNA from regenerating rat liver and chick embryos, indicating them to be normal intermediates in the replication of mtDNA All of the mtDNAs examined included partially single-stranded simple (nonforked) circular molecules. A possible scheme for the replication of mtDNA is presented, based on the different molecular forms observed PMID:4345165

  13. Equilibrious Strand Exchange Promoted by DNA Conformational Switching

    NASA Astrophysics Data System (ADS)

    Wu, Zhiguo; Xie, Xiao; Li, Puzhen; Zhao, Jiayi; Huang, Lili; Zhou, Xiang

    2013-01-01

    Most of DNA strand exchange reactions in vitro are based on toehold strategy which is generally nonequilibrium, and intracellular strand exchange mediated by proteins shows little sequence specificity. Herein, a new strand exchange promoted by equilibrious DNA conformational switching is verified. Duplexes containing c-myc sequence which is potentially converted into G-quadruplex are designed in this strategy. The dynamic equilibrium between duplex and G4-DNA is response to the specific exchange of homologous single-stranded DNA (ssDNA). The SER is enzyme free and sequence specific. No ATP is needed and the displaced ssDNAs are identical to the homologous ssDNAs. The SER products and exchange kenetics are analyzed by PAGE and the RecA mediated SER is performed as the contrast. This SER is a new feature of G4-DNAs and a novel strategy to utilize the dynamic equilibrium of DNA conformations.

  14. On the thermal runaway of combustible fluids in lagging material

    NASA Astrophysics Data System (ADS)

    McIntosh, A. C.; Griffiths, J. F.

    1995-01-01

    This paper presents the mathematical foundations for a simple theory for investigating the phenomenon of ignition of flammable fluids in lagging material that are used for insulation of hot pipework, for transport of heat transfer fluids, or other similar situations. Experiments with porous material impregnated with a flammable fluid have simulated the self-heating known to occur when combustible liquids leak from a hot pipe into lagging surrounding the pipe or are split from another source on to the lagging. A theory to explain these findings is presented which shows that there is a watershed temperature beyond which substantial self-heating will take place. Although the theory does not take account of diffusion, it simulates the main physics of the phenomenon-that is, combustible fluid, which normally in the open air would evaporate and not be a hazard, can, within a porous medium, remain dispersed within the porous structure long enough for the exothermic oxidation to develop into ignition.

  15. New Views on Strand Asymmetry in Insect Mitochondrial Genomes

    PubMed Central

    Wei, Shu-Jun; Shi, Min; Chen, Xue-Xin; Sharkey, Michael J.; van Achterberg, Cornelis; Ye, Gong-Yin; He, Jun-Hua

    2010-01-01

    Strand asymmetry in nucleotide composition is a remarkable feature of animal mitochondrial genomes. Understanding the mutation processes that shape strand asymmetry is essential for comprehensive knowledge of genome evolution, demographical population history and accurate phylogenetic inference. Previous studies found that the relative contributions of different substitution types to strand asymmetry are associated with replication alone or both replication and transcription. However, the relative contributions of replication and transcription to strand asymmetry remain unclear. Here we conducted a broad survey of strand asymmetry across 120 insect mitochondrial genomes, with special reference to the correlation between the signs of skew values and replication orientation/gene direction. The results show that the sign of GC skew on entire mitochondrial genomes is reversed in all species of three distantly related families of insects, Philopteridae (Phthiraptera), Aleyrodidae (Hemiptera) and Braconidae (Hymenoptera); the replication-related elements in the A+T-rich regions of these species are inverted, confirming that reversal of strand asymmetry (GC skew) was caused by inversion of replication origin; and finally, the sign of GC skew value is associated with replication orientation but not with gene direction, while that of AT skew value varies with gene direction, replication and codon positions used in analyses. These findings show that deaminations during replication and other mutations contribute more than selection on amino acid sequences to strand compositions of G and C, and that the replication process has a stronger affect on A and T content than does transcription. Our results may contribute to genome-wide studies of replication and transcription mechanisms. PMID:20856815

  16. Single-stranded DNA and RNA origami.

    PubMed

    Han, Dongran; Qi, Xiaodong; Myhrvold, Cameron; Wang, Bei; Dai, Mingjie; Jiang, Shuoxing; Bates, Maxwell; Liu, Yan; An, Byoungkwon; Zhang, Fei; Yan, Hao; Yin, Peng

    2017-12-15

    Self-folding of an information-carrying polymer into a defined structure is foundational to biology and offers attractive potential as a synthetic strategy. Although multicomponent self-assembly has produced complex synthetic nanostructures, unimolecular folding has seen limited progress. We describe a framework to design and synthesize a single DNA or RNA strand to self-fold into a complex yet unknotted structure that approximates an arbitrary user-prescribed shape. We experimentally construct diverse multikilobase single-stranded structures, including a ~10,000-nucleotide (nt) DNA structure and a ~6000-nt RNA structure. We demonstrate facile replication of the strand in vitro and in living cells. The work here thus establishes unimolecular folding as a general strategy for constructing complex and replicable nucleic acid nanostructures, and expands the design space and material scalability for bottom-up nanotechnology. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  17. Anticipated and zero-lag synchronization in motifs of delay-coupled systems

    NASA Astrophysics Data System (ADS)

    Mirasso, Claudio R.; Carelli, Pedro V.; Pereira, Tiago; Matias, Fernanda S.; Copelli, Mauro

    2017-11-01

    Anticipated and zero-lag synchronization have been observed in different scientific fields. In the brain, they might play a fundamental role in information processing, temporal coding and spatial attention. Recent numerical work on anticipated and zero-lag synchronization studied the role of delays. However, an analytical understanding of the conditions for these phenomena remains elusive. In this paper, we study both phenomena in systems with small delays. By performing a phase reduction and studying phase locked solutions, we uncover the functional relation between the delay, excitation and inhibition for the onset of anticipated synchronization in a sender-receiver-interneuron motif. In the case of zero-lag synchronization in a chain motif, we determine the stability conditions. These analytical solutions provide an excellent prediction of the phase-locked regimes of Hodgkin-Huxley models and Roessler oscillators.

  18. [Effects of orthokeratology lenses on the magnitude of accommodative lag and accommodativeconvergence/accommodation].

    PubMed

    Ren, Qiujin; Yue, Hui; Zhou, Qing

    2016-02-01

    To evaluate the change in accommodative lag and accommodation convergence/accommodation (AC/A) after patients with myopia wear orthokeratology lenses. 
 A total of 48 myopic subjects (a test group), who wore orthokeratology lenses regularly, and 48 myopic subjects (a control group), who wore spectacles regularly, were enrolled for this study from January 2011 to January 2013 in Optometric Center, the Forth Hospital of Changsha. Accommodative lag was measured by fused cross cylinder method, where the patients should gaze at the front optotypes 40 cm away. Gradient of the AC/A ratio was measured by Von Grafe method to check closer distance heterophoria. Accommodative lag and AC/A ratio were analyzed by statistics.
 After 1-year follow-up, accommodative lag and AC/A rate in patients with low or moderate myopia in the test group was decreased in 1, 3, 6 months or 1 year compared with that in the control group (P<0.05). 
 Compared with spectacles, orthokeratology lenses are able to decrease accommodative lag and high AC/A rate in patients with low or moderate myopia. The relationship between accommodation and convergence is improved by orthokeratology lenses. Orthokeratology is an effective way to control myopia.

  19. The Reverberation Lag in the Low-mass X-ray Binary H1743-322

    NASA Astrophysics Data System (ADS)

    De Marco, Barbara; Ponti, Gabriele

    2016-07-01

    The evolution of the inner accretion flow of a black hole X-ray binary during an outburst is still a matter of active research. X-ray reverberation lags are powerful tools for constraining disk-corona geometry. We present a study of X-ray lags in the black hole transient H1743-322. We compared the results obtained from analysis of all the publicly available XMM-Newton observations. These observations were carried out during two different outbursts that occurred in 2008 and 2014. During all the observations the source was caught in the hard state and at similar luminosities ({L}3-10{keV}/{L}{Edd}˜ 0.004). We detected a soft X-ray lag of ˜60 ms, most likely due to thermal reverberation. We did not detect any significant change of the lag amplitude among the different observations, indicating a similar disk-corona geometry at the same luminosity in the hard state. On the other hand, we observe significant differences between the reverberation lag detected in H1743-322 and in GX 339-4 (at similar luminosities in the hard state), which might indicate variations of the geometry from source to source.

  20. Role of Lymphocyte Activation Gene-3 (Lag-3) in Conventional and Regulatory T Cell Function in Allogeneic Transplantation

    PubMed Central

    Sega, Emanuela I.; Leveson-Gower, Dennis B.; Florek, Mareike; Schneidawind, Dominik; Luong, Richard H.; Negrin, Robert S.

    2014-01-01

    Lag-3 has emerged as an important molecule in T cell biology. We investigated the role of Lag-3 in conventional T cell (Tcon) and regulatory T cell (Treg) function in murine GVHD with the hypothesis that Lag-3 engagement diminishes alloreactive T cell responses after bone marrow transplantation. We demonstrate that Lag-3 deficient Tcon (Lag-3−/− Tcon) induce significantly more severe GVHD than wild type (WT) Tcon and that the absence of Lag-3 on CD4 but not CD8 T cells is responsible for exacerbating GVHD. Lag-3−/− Tcon exhibited increased activation and proliferation as indicated by CFSE and bioluminescence imaging analyses and higher levels of activation markers such as CD69, CD107a, granzyme B, and Ki-67 as well as production of IL-10 and IFN-g early after transplantation. Lag-3−/− Tcon were less responsive to suppression by WT Treg as compared to WT Tcon. The absence of Lag-3, however, did not impair Treg function as both Lag-3−/− and WT Treg equally suppress the proliferation of Tcon in vitro and in vivo and protect against GVHD. Further, we demonstrate that allogeneic Treg acquire recipient MHC class II molecules through a process termed trogocytosis. As MHC class II is a ligand for Lag-3, we propose a novel suppression mechanism employed by Treg involving the acquisition of host MHC-II followed by the engagement of Lag-3 on T cells. These studies demonstrate for the first time the biologic function of Lag-3 expression on conventional and regulatory T cells in GVHD and identify Lag-3 as an important regulatory molecule involved in alloreactive T cell proliferation and activation after bone marrow transplantation. PMID:24475140

  1. An investigation of the lag between the start of research and the development of new technology

    NASA Technical Reports Server (NTRS)

    Glass, S. E.

    1982-01-01

    The lag which occurs between the start of NASA-sponsored research and the development of new technology is addressed. A possible common gestation period is examined. The lags vary from one to zero years. The observed lag as it relates to patent applications is shorter than the lag as it relates to invention disclosures. The sequential hypothesis testing showed that invention disclosures correlated better to the measures of research effort used then did patent applications.

  2. Leading and lagging indicators of occupational health and safety: The moderating role of safety leadership.

    PubMed

    Sheehan, Cathy; Donohue, Ross; Shea, Tracey; Cooper, Brian; Cieri, Helen De

    2016-07-01

    In response to the call for empirical evidence of a connection between leading and lagging indicators of occupational health and safety (OHS), the first aim of the current research is to consider the association between leading and lagging indicators of OHS. Our second aim is to investigate the moderating effect of safety leadership on the association between leading and lagging indicators. Data were collected from 3578 employees nested within 66 workplaces. Multi-level modelling was used to test the two hypotheses. The results confirm an association between leading and lagging indicators of OHS as well as the moderating impact of middle management safety leadership on the direct association. The association between leading and lagging indicators provides OHS practitioners with useful information to substantiate efforts within organisations to move away from a traditional focus on lagging indicators towards a preventative focus on leading indicators. The research also highlights the important role played by middle managers and the value of OHS leadership development and investment at the middle management level. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Analysis and IbM simulation of the stages in bacterial lag phase: basis for an updated definition.

    PubMed

    Prats, Clara; Giró, Antoni; Ferrer, Jordi; López, Daniel; Vives-Rego, Josep

    2008-05-07

    The lag phase is the initial phase of a culture that precedes exponential growth and occurs when the conditions of the culture medium differ from the pre-inoculation conditions. It is usually defined by means of cell density because the number of individuals remains approximately constant or slowly increases, and it is quantified with the lag parameter lambda. The lag phase has been studied through mathematical modelling and by means of specific experiments. In recent years, Individual-based Modelling (IbM) has provided helpful insights into lag phase studies. In this paper, the definition of lag phase is thoroughly examined. Evolution of the total biomass and the total number of bacteria during lag phase is tackled separately. The lag phase lasts until the culture reaches a maximum growth rate both in biomass and cell density. Once in the exponential phase, both rates are constant over time and equal to each other. Both evolutions are split into an initial phase and a transition phase, according to their growth rates. A population-level mathematical model is presented to describe the transitional phase in cell density. INDividual DIScrete SIMulation (INDISIM) is used to check the outcomes of this analysis. Simulations allow the separate study of the evolution of cell density and total biomass in a batch culture, they provide a depiction of different observed cases in lag evolution at the individual-cell level, and are used to test the population-level model. The results show that the geometrical lag parameter lambda is not appropriate as a universal definition for the lag phase. Moreover, the lag phase cannot be characterized by a single parameter. For the studied cases, the lag phases of both the total biomass and the population are required to fully characterize the evolution of bacterial cultures. The results presented prove once more that the lag phase is a complex process that requires a more complete definition. This will be possible only after the phenomena

  4. A forward bias method for lag correction of an a-Si flat panel detector

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

    Starman, Jared; Tognina, Carlo; Partain, Larry

    2012-01-15

    Purpose: Digital a-Si flat panel (FP) x-ray detectors can exhibit detector lag, or residual signal, of several percent that can cause ghosting in projection images or severe shading artifacts, known as the radar artifact, in cone-beam computed tomography (CBCT) reconstructions. A major contributor to detector lag is believed to be defect states, or traps, in the a-Si layer of the FP. Software methods to characterize and correct for the detector lag exist, but they may make assumptions such as system linearity and time invariance, which may not be true. The purpose of this work is to investigate a new hardwaremore » based method to reduce lag in an a-Si FP and to evaluate its effectiveness at removing shading artifacts in CBCT reconstructions. The feasibility of a novel, partially hardware based solution is also examined. Methods: The proposed hardware solution for lag reduction requires only a minor change to the FP. For pulsed irradiation, the proposed method inserts a new operation step between the readout and data collection stages. During this new stage the photodiode is operated in a forward bias mode, which fills the defect states with charge. A Varian 4030CB panel was modified to allow for operation in the forward bias mode. The contrast of residual lag ghosts was measured for lag frames 2 and 100 after irradiation ceased for standard and forward bias modes. Detector step response, lag, SNR, modulation transfer function (MTF), and detective quantum efficiency (DQE) measurements were made with standard and forward bias firmware. CBCT data of pelvic and head phantoms were also collected. Results: Overall, the 2nd and 100th detector lag frame residual signals were reduced 70%-88% using the new method. SNR, MTF, and DQE measurements show a small decrease in collected signal and a small increase in noise. The forward bias hardware successfully reduced the radar artifact in the CBCT reconstruction of the pelvic and head phantoms by 48%-81%. Conclusions: Overall

  5. Solid phase sequencing of double-stranded nucleic acids

    DOEpatents

    Fu, Dong-Jing; Cantor, Charles R.; Koster, Hubert; Smith, Cassandra L.

    2002-01-01

    This invention relates to methods for detecting and sequencing of target double-stranded nucleic acid sequences, to nucleic acid probes and arrays of probes useful in these methods, and to kits and systems which contain these probes. Useful methods involve hybridizing the nucleic acids or nucleic acids which represent complementary or homologous sequences of the target to an array of nucleic acid probes. These probe comprise a single-stranded portion, an optional double-stranded portion and a variable sequence within the single-stranded portion. The molecular weights of the hybridized nucleic acids of the set can be determined by mass spectroscopy, and the sequence of the target determined from the molecular weights of the fragments. Nucleic acids whose sequences can be determined include nucleic acids in biological samples such as patient biopsies and environmental samples. Probes may be fixed to a solid support such as a hybridization chip to facilitate automated determination of molecular weights and identification of the target sequence.

  6. A Novel Computational Method to Reduce Leaky Reaction in DNA Strand Displacement.

    PubMed

    Li, Xin; Wang, Xun; Song, Tao; Lu, Wei; Chen, Zhihua; Shi, Xiaolong

    2015-01-01

    DNA strand displacement technique is widely used in DNA programming, DNA biosensors, and gene analysis. In DNA strand displacement, leaky reactions can cause DNA signals decay and detecting DNA signals fails. The mostly used method to avoid leakage is cleaning up after upstream leaky reactions, and it remains a challenge to develop reliable DNA strand displacement technique with low leakage. In this work, we address the challenge by experimentally evaluating the basic factors, including reaction time, ratio of reactants, and ion concentration to the leakage in DNA strand displacement. Specifically, fluorescent probes and a hairpin structure reporting DNA strand are designed to detect the output of DNA strand displacement, and thus can evaluate the leakage of DNA strand displacement reactions with different reaction time, ratios of reactants, and ion concentrations. From the obtained data, mathematical models for evaluating leakage are achieved by curve derivation. As a result, it is obtained that long time incubation, high concentration of fuel strand, and inappropriate amount of ion concentration can weaken leaky reactions. This contributes to a method to set proper reaction conditions to reduce leakage in DNA strand displacement.

  7. A Novel Computational Method to Reduce Leaky Reaction in DNA Strand Displacement

    PubMed Central

    Li, Xin; Wang, Xun; Song, Tao; Lu, Wei; Chen, Zhihua; Shi, Xiaolong

    2015-01-01

    DNA strand displacement technique is widely used in DNA programming, DNA biosensors, and gene analysis. In DNA strand displacement, leaky reactions can cause DNA signals decay and detecting DNA signals fails. The mostly used method to avoid leakage is cleaning up after upstream leaky reactions, and it remains a challenge to develop reliable DNA strand displacement technique with low leakage. In this work, we address the challenge by experimentally evaluating the basic factors, including reaction time, ratio of reactants, and ion concentration to the leakage in DNA strand displacement. Specifically, fluorescent probes and a hairpin structure reporting DNA strand are designed to detect the output of DNA strand displacement, and thus can evaluate the leakage of DNA strand displacement reactions with different reaction time, ratios of reactants, and ion concentrations. From the obtained data, mathematical models for evaluating leakage are achieved by curve derivation. As a result, it is obtained that long time incubation, high concentration of fuel strand, and inappropriate amount of ion concentration can weaken leaky reactions. This contributes to a method to set proper reaction conditions to reduce leakage in DNA strand displacement. PMID:26491602

  8. Double-Stranded RNA Is Detected by Immunofluorescence Analysis in RNA and DNA Virus Infections, Including Those by Negative-Stranded RNA Viruses.

    PubMed

    Son, Kyung-No; Liang, Zhiguo; Lipton, Howard L

    2015-09-01

    Early biochemical studies of viral replication suggested that most viruses produce double-stranded RNA (dsRNA), which is essential for the induction of the host immune response. However, it was reported in 2006 that dsRNA could be detected by immunofluorescence antibody staining in double-stranded DNA and positive-strand RNA virus infections but not in negative-strand RNA virus infections. Other reports in the literature seemed to support these observations. This suggested that negative-strand RNA viruses produce little, if any, dsRNA or that more efficient viral countermeasures to mask dsRNA are mounted. Because of our interest in the use of dsRNA antibodies for virus discovery, particularly in pathological specimens, we wanted to determine how universal immunostaining for dsRNA might be in animal virus infections. We have detected the in situ formation of dsRNA in cells infected with vesicular stomatitis virus, measles virus, influenza A virus, and Nyamanini virus, which represent viruses from different negative-strand RNA virus families. dsRNA was also detected in cells infected with lymphocytic choriomeningitis virus, an ambisense RNA virus, and minute virus of mice (MVM), a single-stranded DNA (ssDNA) parvovirus, but not hepatitis B virus. Although dsRNA staining was primarily observed in the cytoplasm, it was also seen in the nucleus of cells infected with influenza A virus, Nyamanini virus, and MVM. Thus, it is likely that most animal virus infections produce dsRNA species that can be detected by immunofluorescence staining. The apoptosis induced in several uninfected cell lines failed to upregulate dsRNA formation. An effective antiviral host immune response depends on recognition of viral invasion and an intact innate immune system as a first line of defense. Double-stranded RNA (dsRNA) is a viral product essential for the induction of innate immunity, leading to the production of type I interferons (IFNs) and the activation of hundreds of IFN

  9. Early biometric lag in the prediction of small for gestational age neonates and preeclampsia.

    PubMed

    Schwartz, Nadav; Pessel, Cara; Coletta, Jaclyn; Krieger, Abba M; Timor-Tritsch, Ilan E

    2011-01-01

    An early fetal growth lag may be a marker of future complications. We sought to determine the utility of early biometric variables in predicting adverse pregnancy outcomes. In this retrospective cohort study, the crown-rump length at 11 to 14 weeks and the head circumference, biparietal diameter, abdominal circumference, femur length, humerus length, transverse cerebellar diameter, and estimated fetal weight at 18 to 24 weeks were converted to an estimated gestational age using published regression formulas. Sonographic fetal growth (difference between each biometric gestational age and the crown-rump length gestational age) minus expected fetal growth (number of days elapsed between the two scans) yielded the biometric growth lag. These lags were tested as predictors of small for gestational age (SGA) neonates (≤10th percentile) and preeclampsia. A total of 245 patients were included. Thirty-two (13.1%) delivered an SGA neonate, and 43 (17.6%) had the composite outcome. The head circumference, biparietal diameter, abdominal circumference, and estimated fetal weight lags were identified as significant predictors of SGA neonates after adjusted analyses (P < .05). The addition of either the estimated fetal weight or abdominal circumference lag to maternal characteristics alone significantly improved the performance of the predictive model, achieving areas under the curve of 0.72 and 0.74, respectively. No significant association was found between the biometric lag variables and the development of preeclampsia. Routinely available biometric data can be used to improve the prediction of adverse outcomes such as SGA. These biometric lags should be considered in efforts to develop screening algorithms for adverse outcomes.

  10. Effects of timber harvesting on the lag time of Caspar Creek watershed

    Treesearch

    Karen Hardison Sendek

    1985-01-01

    Abstract - Hydrograph lag time was analyzed to determine changes after road construction and after selective, tractor-yarded logging in a Caspar Creek watershed, Mendocino County, California. The paired watershed technique was used. Hydrograph lag time for each storm was the time separation between the midpoint of precipitation and the time coordinate of the runoff...

  11. Efficient synthesis of supercoiled M13 DNA molecule containing a site specifically placed psoralen adduct and its use as a substrate for DNA replication

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

    Kodadek, T.; Gamper, H.

    The authors report a simple method for the in vitro synthesis of large quantities of site specifically modified DNA. The protocol involves extension of an oligonucleotide primer annealed to M13 single-stranded DNA using part of the T4 DNA polymerase holoenzyme. The resulting nicked double-stranded circles are ligated and supercoiled in the same tube, producing good yields of form I DNA. When the oligonucleotide primer is chemically modified, the resultant product contains a site-specific lesion. In this study, they report the synthesis of an M13 mp19 form I DNA which contains a psoralen monoadduct or cross-link at the KpnI site. Theymore » demonstrate the utility of these modified substrates by assessing the ability of the bacteriophage T4 DNA replication complex to bypass the damage and show that the psoralen monoadduct poses a severe block to the holoenzyme when attached to the template strand.« less

  12. Autonomous parvovirus LuIII encapsidates equal amounts of plus and minus DNA strands

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

    Bates, R.C.; Snyder, C.E.; Banerjee, P.T.

    1984-02-01

    Autonomous parvoviruses are thought to uniquely encapsidate single-stranded DNA of minus polarity. In contrast, the defective adeno-associated viruses separately encapsidate equal amounts of plus and minus DNA strands. The uniqueness of minus strand encapsidation is reexamined for the autonomous parvoviruses. Although it was found that Kilham rat virus and H-1 virus encapsidate varying but small amounts of complementary-strand DNA, it was unexpected to find that LuIII virus encapsidated equal amounts of plus and minus DNA. The extracted LuIII DNA possessed properties of double-stranded replicative-form DNA, including insensitivity to S1 endonuclease, cleavage by restriction enzymes, and conversion to unit-length, single-stranded DNAmore » when electrophoresed under denaturing conditions. However, the inability of this DNA to form single-stranded DNA circles when denatured and then renatured in the presence of formamide and the lack of double-stranded DNA circle formation after treatment with exonuclease III and reannealing shows a lack of sequence homology of the 3' and 5' termini of LuIII DNA, in contrast to adeno-associated virus DNA. Digestion of LuIII double-stranded DNA with EcoRI and HincII and separation of plus and minus DNA strands on composite agarose-acrylamide gels identified a heterogeneity present only in the plus DNA strand. These results suggest that strand specificity of viral DNA encapsidation is not a useful property for differentiation between the autonomous and defective parvoviruses. Furthermore, encapsidation by LuIII of equal amounts of complementary DNA strands in contrast to encapsidation of minus strands by H-1 virus, when propagated in the same host cell type, suggests that selection of strands for encapsidation is a virus-coded rather than host-controlled event.« less

  13. Exploring lag times between monthly atmospheric deposition and stream chemistry in Appalachian forests using cross-correlation

    NASA Astrophysics Data System (ADS)

    DeWalle, David R.; Boyer, Elizabeth W.; Buda, Anthony R.

    2016-12-01

    Forecasts of ecosystem changes due to variations in atmospheric emissions policies require a fundamental understanding of lag times between changes in chemical inputs and watershed response. Impacts of changes in atmospheric deposition in the United States have been documented using national and regional long-term environmental monitoring programs beginning several decades ago. Consequently, time series of weekly NADP atmospheric wet deposition and monthly EPA-Long Term Monitoring stream chemistry now exist for much of the Northeast which may provide insights into lag times. In this study of Appalachian forest basins, we estimated lag times for S, N and Cl by cross-correlating monthly data from four pairs of stream and deposition monitoring sites during the period from 1978 to 2012. A systems or impulse response function approach to cross-correlation was used to estimate lag times where the input deposition time series was pre-whitened using regression modeling and the stream response time series was filtered using the deposition regression model prior to cross-correlation. Cross-correlations for S were greatest at annual intervals over a relatively well-defined range of lags with the maximum correlations occurring at mean lags of 48 months. Chloride results were similar but more erratic with a mean lag of 57 months. Few high-correlation lags for N were indicated. Given the growing availability of atmospheric deposition and surface water chemistry monitoring data and our results for four Appalachian basins, further testing of cross-correlation as a method of estimating lag times on other basins appears justified.

  14. Minimization of diauxic growth lag-phase for high-efficiency biogas production.

    PubMed

    Kim, Min Jee; Kim, Sang Hun

    2017-02-01

    The objective of this study was to develop a minimization method of a diauxic growth lag-phase for the biogas production from agricultural by-products (ABPs). Specifically, the effects of proximate composition on the biogas production and degradation rates of the ABPs were investigated, and a new method based on proximate composition combinations was developed to minimize the diauxic growth lag-phase. Experiments were performed using biogas potential tests at a substrate loading of 2.5 g VS/L and feed to microorganism ratio (F/M) of 0.5 under the mesophilic condition. The ABPs were classified based on proximate composition (carbohydrate, protein, and fat etc.). The biogas production patterns, lag phase, and times taken for 90% biogas production (T90) were used for the evaluation of the biogas production with biochemical methane potential (BMP) test. The high- or medium-carbohydrate and low-fat ABPs (cheese whey, cabbage, and skim milk) showed a single step digestion process and low-carbohydrate and high-fat ABPs (bean curd and perilla seed) showed a two-step digestion process. The mixture of high-fat ABPs and high-carbohydrate ABPs reduced the lag-phase and increased the biogas yield more than that from single ABP by 35-46%. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. How to compare cross-lagged associations in a multilevel autoregressive model.

    PubMed

    Schuurman, Noémi K; Ferrer, Emilio; de Boer-Sonnenschein, Mieke; Hamaker, Ellen L

    2016-06-01

    By modeling variables over time it is possible to investigate the Granger-causal cross-lagged associations between variables. By comparing the standardized cross-lagged coefficients, the relative strength of these associations can be evaluated in order to determine important driving forces in the dynamic system. The aim of this study was twofold: first, to illustrate the added value of a multilevel multivariate autoregressive modeling approach for investigating these associations over more traditional techniques; and second, to discuss how the coefficients of the multilevel autoregressive model should be standardized for comparing the strength of the cross-lagged associations. The hierarchical structure of multilevel multivariate autoregressive models complicates standardization, because subject-based statistics or group-based statistics can be used to standardize the coefficients, and each method may result in different conclusions. We argue that in order to make a meaningful comparison of the strength of the cross-lagged associations, the coefficients should be standardized within persons. We further illustrate the bivariate multilevel autoregressive model and the standardization of the coefficients, and we show that disregarding individual differences in dynamics can prove misleading, by means of an empirical example on experienced competence and exhaustion in persons diagnosed with burnout. (PsycINFO Database Record (c) 2016 APA, all rights reserved).

  16. Stranding Events of Kogia Whales along the Brazilian Coast.

    PubMed

    Moura, Jailson F; Acevedo-Trejos, Esteban; Tavares, Davi C; Meirelles, Ana C O; Silva, Cristine P N; Oliveira, Larissa R; Santos, Roberta A; Wickert, Janaína C; Machado, Rodrigo; Siciliano, Salvatore; Merico, Agostino

    2016-01-01

    The genus Kogia, which comprises only two extant species, Kogia sima and Kogia breviceps, represents one of the least known groups of cetaceans in the global ocean. In some coastal regions, however, stranding events of these species have been relatively common over the last decades. Stranding provides the opportunity to investigate the biology of these cetaceans and to explore the epidemiological aspects associated with the mortality of the organisms found on the beach. A number of disturbances (including pelagic fisheries, chemical pollution, boat strikes, and noise pollution) have been confirmed to pose a particular threat to the Kogia species. However, no study has yet investigated potential relationships between environmental conditions and stranding events. Here we analyse how a collection of environmental, physical, and biological variables, such as wind, sea surface temperature (SST), water depth, and chlorophyll-a, correlate to Kogia stranding events along the Brazilian coast. The results of our statistical analyses suggest that K. sima is more likely found in warm tropical waters, which provide an explanation for the high frequency of stranding in northeastern Brazilian coast. In contrast, K. breviceps appears to have a preference for temperate and productive waters. Wind speed results to be also an important factor for predicting Kogia strandings in Brazilian coast. Additionally, literature information in combination with our own data and analyses of stomach contents confirms that oceanic cephalopods constitute the primary nutritional source of both Kogia species. By using the available information as a qualitative proxy for habitat preference and feeding ecology, our study provides a novel and comprehensive assessment of Kogia stranding data in relation to environmental conditions along the Brazilian coast.

  17. Evaluation of the limiting antigen avidity EIA (LAg) in people who inject drugs in Greece.

    PubMed

    Nikolopoulos, G K; Katsoulidou, A; Kantzanou, M; Rokka, C; Tsiara, C; Sypsa, V; Paraskevis, D; Psichogiou, M; Friedman, S; Hatzakis, A

    2017-01-01

    This analysis assessed the utility of the limiting antigen avidity assay (LAg). Samples of people who inject drugs (PWID) in Greece with documented duration of HIV-1 infection were tested by LAg. A LAg-normalized optical density (ODn) ⩽1·5 corresponds to a recency window period of 130 days. The proportion true recent (PTR) and proportion false recent (PFR) were estimated in 28 seroconverters and in 366 samples collected >6 months after HIV diagnosis, respectively. The association between LAg ODn and HIV RNA level was evaluated in 232 persons. The PTR was 85·7%. The PFR was 20·8% but fell to 5·9% in samples from treatment-naive individuals with long-standing infection (>1 year), and to 0 in samples with the circulating recombinant form CRF35 AD. A LAg-based algorithm with a PFR of 3·3% estimated a similar incidence trend to that calculated by analyses based on HIV-1 seroconversions. In recently infected persons indicated by LAg, the median log10 HIV RNA level was high (5·30, interquartile range 4·56-5·90). LAg can help identify highly infectious HIV(+) individuals as it accurately identifies recent infections and is correlated with the HIV RNA level. It can also produce reliable estimates of HIV-1 incidence.

  18. A principled approach to the stranded cost issue

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

    Maloney, M.T.; Sauer, R.D.

    1998-04-01

    The case against stranded cost recovery is a strong one, whether investments in generation were efficient ex ante or not. This is the unavoidable conclusion once the proper roles of the regulator, utilities, investors, and consumers in the regulatory system of the past century are clearly identified. It is improper and inefficient for government to sanction the mistakes of the private sector by taxing consumers in order to rescue producers. This is precisely what stranded cost recovery does. Denial of stranded cost recovery is consistent with the role of the regulator as a substitute for salutary market forces and, indeed,more » is required by it.« less

  19. Peripheral Design of Progressive Addition Lenses and the Lag of Accommodation in Myopes.

    PubMed

    Schilling, Tim; Ohlendorf, Arne; Varnas, Saulius R; Wahl, Siegfried

    2017-07-01

    Insufficient accommodative response is assumed to result in myopia progression. We have investigated if the accommodative lag in myopes is different between a single vision lens (SVL) and the progressive addition lens PAL 2, clinically trialled for its ability to reduce progression of myopia, and if there exist differences in accommodative lag between PAL 2 and other PALs with the same addition power (+1.50 D). The influence of spherical SVL and four different designs of PALs that differ in the near zone width (PAL 1) or that have different signs and magnitude of horizontal gradients of mean power adjacent to their near vision zones (PAL 3 and PAL 4) on the accommodative response was investigated for different near viewing distances (40, 33, and 25 cm) in 31 subjects, aged 18 to 25 years. The SVL correction resulted in insufficient accommodative response for the near object viewing distances tested. PAL 2 did significantly reduce accommodative lag for all near object distances tested. The PAL design with a more negative horizontal mean power gradient (PAL 4) provided a lower lag of accommodation when compared with PAL 2 at the shortest object distance of 25 cm (P = 0.03) and was able to reduce the lag of accommodation to a level below the depth of focus for the higher near working distances tested. Designs of PAL with more negative horizontal mean power gradients are the most effective in lowering the lag of accommodation in myopes. This could make them good test candidates for myopia control applications.

  20. DNA strand-exchange patterns associated with double-strand break-induced and spontaneous mitotic crossovers in Saccharomyces cerevisiae

    PubMed Central

    2018-01-01

    Mitotic recombination can result in loss of heterozygosity and chromosomal rearrangements that shape genome structure and initiate human disease. Engineered double-strand breaks (DSBs) are a potent initiator of recombination, but whether spontaneous events initiate with the breakage of one or both DNA strands remains unclear. In the current study, a crossover (CO)-specific assay was used to compare heteroduplex DNA (hetDNA) profiles, which reflect strand exchange intermediates, associated with DSB-induced versus spontaneous events in yeast. Most DSB-induced CO products had the two-sided hetDNA predicted by the canonical DSB repair model, with a switch in hetDNA position from one product to the other at the position of the break. Approximately 40% of COs, however, had hetDNA on only one side of the initiating break. This anomaly can be explained by a modified model in which there is frequent processing of an early invasion (D-loop) intermediate prior to extension of the invading end. Finally, hetDNA tracts exhibited complexities consistent with frequent expansion of the DSB into a gap, migration of strand-exchange junctions, and template switching during gap-filling reactions. hetDNA patterns in spontaneous COs isolated in either a wild-type background or in a background with elevated levels of reactive oxygen species (tsa1Δ mutant) were similar to those associated with the DSB-induced events, suggesting that DSBs are the major instigator of spontaneous mitotic recombination in yeast. PMID:29579095

  1. The lag effects and vulnerabilities of temperature effects on cardiovascular disease mortality in a subtropical climate zone in China.

    PubMed

    Huang, Jixia; Wang, Jinfeng; Yu, Weiwei

    2014-04-11

    This research quantifies the lag effects and vulnerabilities of temperature effects on cardiovascular disease in Changsha--a subtropical climate zone of China. A Poisson regression model within a distributed lag nonlinear models framework was used to examine the lag effects of cold- and heat-related CVD mortality. The lag effect for heat-related CVD mortality was just 0-3 days. In contrast, we observed a statistically significant association with 10-25 lag days for cold-related CVD mortality. Low temperatures with 0-2 lag days increased the mortality risk for those ≥65 years and females. For all ages, the cumulative effects of cold-related CVD mortality was 6.6% (95% CI: 5.2%-8.2%) for 30 lag days while that of heat-related CVD mortality was 4.9% (95% CI: 2.0%-7.9%) for 3 lag days. We found that in Changsha city, the lag effect of hot temperatures is short while the lag effect of cold temperatures is long. Females and older people were more sensitive to extreme hot and cold temperatures than males and younger people.

  2. A Comparison of Several Methods of Measuring Ignition Lag in a Compression-ignition Engine

    NASA Technical Reports Server (NTRS)

    Spanogle, J A

    1934-01-01

    The ignition lag of a fuel oil in the combustion chamber of a high speed compression-ignition engine was measured by three different methods. The start of injection of the fuel as observed with a Stoborama was taken as the start of the period of ignition lag in all cases. The end of the period of ignition lag was determined by observation of the appearance of incandescence in the combustion chamber, by inspection of a pressure-time card for evidence of pressure rise, and by analysis of the indicator card for evidence of the combustion of a small but definite quantity of fuel. A comparison of the values for ignition lags obtained by these three methods indicates that the appearance of incandescence is later than other evidences of the start of combustion, that visual inspection of a pressure-time diagram gives consistent and usable values with a minimum requirement of time and/or apparatus, and that analysis of the indicator card is not worth while for ignition lag alone.

  3. Estimation of watershed lag times and times of concentration for the Kansas City Area : technical summary.

    DOT National Transportation Integrated Search

    2016-04-01

    The new lag-time equation for the Kansas City area performs better than the 2001 KU-KDOT equation for urban lag time, which was developed from a study of 14 gaged watersheds in Johnson County (McEnroe & Zhao, 2001). The main advantage of the new lag-...

  4. Break-induced telomere synthesis underlies alternative telomere maintenance

    PubMed Central

    Dilley, Robert L.; Verma, Priyanka; Cho, Nam Woo; Winters, Harrison D.; Wondisford, Anne R.; Greenberg, Roger A.

    2017-01-01

    Homology-directed DNA repair is essential for genome maintenance through templated DNA synthesis. Alternative lengthening of telomeres (ALT) necessitates homology-directed DNA repair to maintain telomeres in about 10–15% of human cancers. How DNA damage induces assembly and execution of a DNA replication complex (break-induced replisome) at telomeres or elsewhere in the mammalian genome is poorly understood. Here we define break-induced telomere synthesis and demonstrate that it utilizes a specialized replisome, which underlies ALT telomere maintenance. DNA double-strand breaks enact nascent telomere synthesis by long-tract unidirectional replication. Proliferating cell nuclear antigen (PCNA) loading by replication factor C (RFC) acts as the initial sensor of telomere damage to establish predominance of DNA polymerase δ (Pol δ) through its POLD3 subunit. Break-induced telomere synthesis requires the RFC–PCNA–Pol δ axis, but is independent of other canonical replisome components, ATM and ATR, or the homologous recombination protein Rad51. Thus, the inception of telomere damage recognition by the break-induced replisome orchestrates homology-directed telomere maintenance. PMID:27760120

  5. Effect of Repetition Lag on Priming of Unfamiliar Visual Objects in Young and Older Adults

    PubMed Central

    Gordon, Leamarie T.; Soldan, Anja; Thomas, Ayanna K.; Stern, Yaakov

    2013-01-01

    Across three experiments, we examined the effect of repetition lag on priming of unfamiliar visual objects in healthy young and older adults. Multiple levels of lag were examined, ranging from short (one to four intervening stimuli) to long (50+ intervening stimuli). In each experiment, subjects viewed a series of new and repeated line drawings of objects and decided whether they depicted structurally possible or impossible figures. Experiment 1 and 2 found similar levels of priming in young and older adults at short and medium lags. At the longer repetition lags (∼20+ intervening stimuli), older adults showed less overall priming, as measured by reaction time facilitation, than young adults. This indicates that older adults can rapidly encode unfamiliar three-dimensional objects to support priming at shorter lags; however, they cannot maintain these representations over longer intervals. In addition to repetition lag, we also explored the relationship between priming and cognitive reserve, as measured by education and verbal intelligence. In the older adults, higher levels of cognitive reserve were associated with greater reaction time priming, suggesting that cognitive reserve may mediate the relationship between aging and priming. PMID:23276220

  6. Strand displacement activated peroxidase activity of hemin for fluorescent DNA sensing.

    PubMed

    Wang, Quanbo; Xu, Nan; Gui, Zhen; Lei, Jianping; Ju, Huangxian; Yan, Feng

    2015-10-07

    To efficiently regulate the catalytic activity of the peroxidase mimic hemin, this work designs a double-stranded DNA probe containing an intermolecular dimer of hemin, whose peroxidase activity can be activated by a DNA strand displacement reaction. The double-stranded probe is prepared by annealing two strands of hemin labelled DNA oligonucleotides. Using the fluorescent oxidation product of tyramine by H2O2 as a tracing molecule, the low peroxidase activity of the hemin dimer ensures a low fluorescence background. The strand displacement reaction of the target DNA dissociates the hemin dimer and thus significantly increases the catalytic activity of hemin to produce a large amount of dityramine for fluorescence signal readout. Based on the strand displacement regulated peroxidase activity, a simple and sensitive homogeneous fluorescent DNA sensing method is proposed. The detection can conveniently be carried out in a 96-well plate within 20 min with a detection limit of 0.18 nM. This method shows high specificity, which can effectively distinguish single-base mismatched DNA from perfectly matched target DNA. The DNA strand displacement regulated catalytic activity of hemin has promising application in the determination of various DNA analytes.

  7. The Sloan Digital Sky Survey Reverberation Mapping Project: Composite Lags at z ≤ 1

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

    Li, Jennifer; Shen, Yue; Horne, Keith

    We present composite broad-line region (BLR) reverberation mapping lag measurements for H α , H β , He ii λ 4686, and Mg ii for a sample of 144, z ≲ 1 quasars from the Sloan Digital Sky Survey Reverberation Mapping (SDSS-RM) project. Using only the 32-epoch spectroscopic light curves in the first six-month season of SDSS-RM observations, we compile correlation function measurements for individual objects and then coadd them to allow the measurement of the average lags for our sample at mean redshifts of 0.4 (for H α ) and ∼0.65 (for the other lines). At similar quasar luminositiesmore » and redshifts, the sample-averaged lag decreases in the order of Mg ii, H α , H β , and He ii. This decrease in lags is accompanied by an increase in the mean line width of the four lines, and is roughly consistent with the virialized motion for BLR gas in photoionization equilibrium. These are among the first RM measurements of stratified BLR structure at z > 0.3. Dividing our sample by luminosity, H α shows clear evidence of increasing lags with luminosity, consistent with the expectation from the measured BLR size–luminosity relation based on H β . The other three lines do not show a clear luminosity trend in their average lags due to the limited dynamic range of luminosity probed and the poor average correlation signals in the divided samples, a situation that will be improved with the incorporation of additional photometric and spectroscopic data from SDSS-RM. We discuss the utility and caveats of composite lag measurements for large statistical quasar samples with reverberation mapping data.« less

  8. The Sloan Digital Sky Survey Reverberation Mapping Project: Composite Lags at z ≤ 1

    NASA Astrophysics Data System (ADS)

    Li, Jennifer; Shen, Yue; Horne, Keith; Brandt, W. N.; Greene, Jenny E.; Grier, C. J.; Ho, Luis C.; Kochanek, Chris; Schneider, Donald P.; Trump, Jonathan R.; Dawson, Kyle S.; Pan, Kaike; Bizyaev, Dmitry; Oravetz, Daniel; Simmons, Audrey; Malanushenko, Elena

    2017-09-01

    We present composite broad-line region (BLR) reverberation mapping lag measurements for Hα, Hβ, He II λ4686, and Mg II for a sample of 144, z ≲ 1 quasars from the Sloan Digital Sky Survey Reverberation Mapping (SDSS-RM) project. Using only the 32-epoch spectroscopic light curves in the first six-month season of SDSS-RM observations, we compile correlation function measurements for individual objects and then coadd them to allow the measurement of the average lags for our sample at mean redshifts of 0.4 (for Hα) and ˜0.65 (for the other lines). At similar quasar luminosities and redshifts, the sample-averaged lag decreases in the order of Mg II, Hα, Hβ, and He II. This decrease in lags is accompanied by an increase in the mean line width of the four lines, and is roughly consistent with the virialized motion for BLR gas in photoionization equilibrium. These are among the first RM measurements of stratified BLR structure at z > 0.3. Dividing our sample by luminosity, Hα shows clear evidence of increasing lags with luminosity, consistent with the expectation from the measured BLR size-luminosity relation based on Hβ. The other three lines do not show a clear luminosity trend in their average lags due to the limited dynamic range of luminosity probed and the poor average correlation signals in the divided samples, a situation that will be improved with the incorporation of additional photometric and spectroscopic data from SDSS-RM. We discuss the utility and caveats of composite lag measurements for large statistical quasar samples with reverberation mapping data.

  9. Synthesis of Deoxyribonucleic Acid After Ultraviolet Irradiation of Sensitive and Resistant Haemophilus influenzae

    PubMed Central

    Modak, Sohan P.; Setlow, Jane K.

    1969-01-01

    Synthesis of deoxyribonucleic acid (DNA) has been measured as a function of ultraviolet (UV) radiation dose in wild-type and seven UV-sensitive strains of Haemophilus influenzae. At the UV doses used, all strains were able to resume DNA synthesis, even those which are unable to excise pyrimidine dimers from their DNA. These excisionless strains showed longer UV-induced delays in DNA synthesis than all but one of the other strains. The longest delay was shown by DB117, a strain which can excise dimers but which is recombination deficient and unable to rejoin X ray-induced single-strand breaks. All strains showed a progressive decrease in sensitivity as they approached the stationary phase. PMID:5305934

  10. Lagging adaptation to warming climate in Arabidopsis thaliana.

    PubMed

    Wilczek, Amity M; Cooper, Martha D; Korves, Tonia M; Schmitt, Johanna

    2014-06-03

    If climate change outpaces the rate of adaptive evolution within a site, populations previously well adapted to local conditions may decline or disappear, and banked seeds from those populations will be unsuitable for restoring them. However, if such adaptational lag has occurred, immigrants from historically warmer climates will outperform natives and may provide genetic potential for evolutionary rescue. We tested for lagging adaptation to warming climate using banked seeds of the annual weed Arabidopsis thaliana in common garden experiments in four sites across the species' native European range: Valencia, Spain; Norwich, United Kingdom; Halle, Germany; and Oulu, Finland. Genotypes originating from geographic regions near the planting site had high relative fitness in each site, direct evidence for broad-scale geographic adaptation in this model species. However, genotypes originating in sites historically warmer than the planting site had higher average relative fitness than local genotypes in every site, especially at the northern range limit in Finland. This result suggests that local adaptive optima have shifted rapidly with recent warming across the species' native range. Climatic optima also differed among seasonal germination cohorts within the Norwich site, suggesting that populations occurring where summer germination is common may have greater evolutionary potential to persist under future warming. If adaptational lag has occurred over just a few decades in banked seeds of an annual species, it may be an important consideration for managing longer-lived species, as well as for attempts to conserve threatened populations through ex situ preservation.

  11. Lagging adaptation to warming climate in Arabidopsis thaliana

    PubMed Central

    Wilczek, Amity M.; Cooper, Martha D.; Korves, Tonia M.; Schmitt, Johanna

    2014-01-01

    If climate change outpaces the rate of adaptive evolution within a site, populations previously well adapted to local conditions may decline or disappear, and banked seeds from those populations will be unsuitable for restoring them. However, if such adaptational lag has occurred, immigrants from historically warmer climates will outperform natives and may provide genetic potential for evolutionary rescue. We tested for lagging adaptation to warming climate using banked seeds of the annual weed Arabidopsis thaliana in common garden experiments in four sites across the species’ native European range: Valencia, Spain; Norwich, United Kingdom; Halle, Germany; and Oulu, Finland. Genotypes originating from geographic regions near the planting site had high relative fitness in each site, direct evidence for broad-scale geographic adaptation in this model species. However, genotypes originating in sites historically warmer than the planting site had higher average relative fitness than local genotypes in every site, especially at the northern range limit in Finland. This result suggests that local adaptive optima have shifted rapidly with recent warming across the species’ native range. Climatic optima also differed among seasonal germination cohorts within the Norwich site, suggesting that populations occurring where summer germination is common may have greater evolutionary potential to persist under future warming. If adaptational lag has occurred over just a few decades in banked seeds of an annual species, it may be an important consideration for managing longer-lived species, as well as for attempts to conserve threatened populations through ex situ preservation. PMID:24843140

  12. Shear lag sutures: Improved suture repair through the use of adhesives

    PubMed Central

    Linderman, Stephen W.; Kormpakis, Ioannis; Gelberman, Richard H.; Birman, Victor; Wegst, Ulrike G. K.; Genin, Guy M.; Thomopoulos, Stavros

    2015-01-01

    Suture materials and surgical knot tying techniques have improved dramatically since their first use over five millennia ago. However, the approach remains limited by the ability of the suture to transfer load to tissue at suture anchor points. Here, we predict that adhesive-coated sutures can improve mechanical load transfer beyond the range of performance of existing suture methods, thereby strengthening repairs and decreasing the risk of failure. The mechanical properties of suitable adhesives were identified using a shear lag model. Examination of the design space for an optimal adhesive demonstrated requirements for strong adhesion and low stiffness to maximize the strength of the adhesive-coated suture repair construct. To experimentally assess the model, we evaluated single strands of sutures coated with highly flexible cyanoacrylates (Loctite 4903 and 4902), cyanoacrylate (Loctite QuickTite Instant Adhesive Gel), rubber cement, rubber/gasket adhesive (1300 Scotch-Weld Neoprene High Performance Rubber & Gasket Adhesive), an albumin-glutaraldehyde adhesive (BioGlue), or poly(dopamine). As a clinically relevant proof-of-concept, cyanoacrylate-coated sutures were then used to perform a clinically relevant flexor digitorum tendon repair in cadaver tissue. The repair performed with adhesive-coated suture had significantly higher strength compared to the standard repair without adhesive. Notably, cyanoacrylate provides strong adhesion with high stiffness and brittle behavior, and is therefore not an ideal adhesive for enhancing suture repair. Nevertheless, the improvement in repair properties in a clinically relevant setting, even using a non-ideal adhesive, demonstrates the potential for the proposed approach to improve outcomes for treatments requiring suture fixation. Further study is necessary to develop a strongly adherent, compliant adhesive within the optimal design space described by the model. PMID:26022966

  13. Calibration of Watershed Lag Time Equation for Philippine Hydrology using RADARSAT Digital Elevation Models

    NASA Astrophysics Data System (ADS)

    Cipriano, F. R.; Lagmay, A. M. A.; Horritt, M.; Mendoza, J.; Sabio, G.; Punay, K. N.; Taniza, H. J.; Uichanco, C.

    2015-12-01

    Widespread flooding is a major problem in the Philippines. The country experiences heavy amount of rainfall throughout the year and several areas are prone to flood hazards because of its unique topography. Human casualties and destruction of infrastructure are just some of the damages caused by flooding and the Philippine government has undertaken various efforts to mitigate these hazards. One of the solutions was to create flood hazard maps of different floodplains and use them to predict the possible catastrophic results of different rain scenarios. To produce these maps with accurate output, different input parameters were needed and one of those is calculating hydrological components from topographical data. This paper presents how a calibrated lag time (TL) equation was obtained using measurable catchment parameters. Lag time is an essential input in flood mapping and is defined as the duration between the peak rainfall and peak discharge of the watershed. The lag time equation involves three measurable parameters, namely, watershed length (L), maximum potential retention (S) derived from the curve number, and watershed slope (Y), all of which were available from RADARSAT Digital Elevation Models (DEM). This approach was based on a similar method developed by CH2M Hill and Horritt for Taiwan, which has a similar set of meteorological and hydrological parameters with the Philippines. Rainfall data from fourteen water level sensors covering 67 storms from all the regions in the country were used to estimate the actual lag time. These sensors were chosen by using a screening process that considers the distance of the sensors from the sea, the availability of recorded data, and the catchment size. The actual lag time values were plotted against the values obtained from the Natural Resource Conservation Management handbook lag time equation. Regression analysis was used to obtain the final calibrated equation that would be used to calculate the lag time

  14. Synthesis of 4'-C-aminoalkyl-2'-O-methyl modified RNA and their biological properties.

    PubMed

    Koizumi, Kana; Maeda, Yusuke; Kano, Toshifumi; Yoshida, Hisae; Sakamoto, Taiichi; Yamagishi, Kenji; Ueno, Yoshihito

    2018-05-17

    In this paper, we describe the synthesis of 4'-C-aminoalkyl-2'-O-methylnucleosides and the properties of RNAs containing these analogs. Phosphoramidites of 4'-C-aminoethyl and 4'-C-aminopropyl-2'-O-methyluridines were prepared using glucose as starting material, and RNAs containing the analogs were synthesized using the phosphoramidites. Thermal denaturation studies revealed that these nucleoside analogs decreased the thermal stabilities of double-stranded RNAs (dsRNAs). Results of NMR, molecular modeling, and CD spectra measurements suggested that 4'-C-aminoalkyl-2'-O-methyluridine adopts an C2'-endo sugar puckering in dsRNA. The 4'-C-aminoalkyl modifications in the passenger strand and the guide strand outside the seed region were well tolerated for RNAi activity of siRNAs. Single-stranded RNAs (ssRNAs) and siRNAs containing the 4'-C-aminoethyl and 4'-C-aminopropyl analogs showed high stability in buffer containing bovine serum. Thus, siRNAs containing the 4'-C-aminoethyl and 4'-C-aminopropyl analogs are good candidates for the development of therapeutic siRNA molecules. Copyright © 2018 Elsevier Ltd. All rights reserved.

  15. Effect of Subelement Spacing in Rrp Nb3Sn Deformed Strands

    NASA Astrophysics Data System (ADS)

    Barzi, E.; Turrioni, D.; Alsharo'a, M.; Field, M.; Hong, S.; Parrell, J.; Yamada, R.; Zhang, Y.; Zlobin, A. V.

    2008-03-01

    The Restacked Rod Process (RRP) is the Nb3Sn strand technology presently producing the largest critical current densities at 4.2 K and 12 T. However, when subject to transverse plastic deformation, RRP subelements (SE) merge into each other, creating larger filaments with a somewhat continuous barrier. In this case, the strand sees a larger effective filament size and its instability can dramatically increase locally leading to a cable quench. To reduce and possibly eliminate this effect, Oxford Instruments Superconducting Technology (OST) developed for FNAL a modified RRP strand design with larger Cu spacing between SE's arranged in a 60/61 array. Strand samples of this design with sizes from 0.7 to 1 mm were first evaluated for transport current properties. A comparison study was then performed between the regular 54/61 and the modified 60/61 design using 0.7 mm round and deformed strands. Finite element modeling of the deformed strands was also performed with ANSYS.

  16. Role of stranded gas in increasing global gas supplies

    USGS Publications Warehouse

    Attanasi, E.D.; Freeman, P.A.

    2013-01-01

    This report synthesizes the findings of three regional studies in order to evaluate, at the global scale, the contribution that stranded gas resources can make to global natural gas supplies. Stranded gas, as defined for this study, is natural gas in discovered conventional gas and oil fields that is currently not commercially producible for either physical or economic reasons. The regional studies evaluated the cost of bringing the large volumes of undeveloped gas in stranded gas fields to selected markets. In particular, stranded gas fields of selected Atlantic Basin countries, north Africa, Russia, and central Asia are screened to determine whether the volumes are sufficient to meet Europe’s increasing demand for gas imports. Stranded gas fields in Russia, central Asia, Southeast Asia, and Australia are also screened to estimate development, production, and transport costs and corresponding gas volumes that could be supplied to Asian markets in China, India, Japan, and South Korea. The data and cost analysis presented here suggest that for the European market and the markets examined in Asia, the development of stranded gas provides a way to meet projected gas import demands for the 2020-to-2040 period. Although this is a reconnaissance-type appraisal, it is based on volumes of gas that are associated with individual identified fields. Individual field data were carefully examined. Some fields were not evaluated because current technology was insufficient or it appeared the gas was likely to be held off the export market. Most of the evaluated stranded gas can be produced and delivered to markets at costs comparable to historical prices. Moreover, the associated volumes of gas are sufficient to provide an interim supply while additional technologies are developed to unlock gas diffused in shale and hydrates or while countries transition to making a greater use of renewable energy sources.

  17. Photochemical Acceleration of DNA Strand Displacement by Using Ultrafast DNA Photo-crosslinking.

    PubMed

    Nakamura, Shigetaka; Hashimoto, Hirokazu; Kobayashi, Satoshi; Fujimoto, Kenzo

    2017-10-18

    DNA strand displacement is an essential reaction in genetic recombination, biological processes, and DNA nanotechnology. In particular, various DNA nanodevices enable complicated calculations. However, it takes time before the output is obtained, so acceleration of DNA strand displacement is required for a rapid-response DNA nanodevice. Herein, DNA strand displacement by using DNA photo-crosslinking to accelerate this displacement is evaluated. The DNA photo-crosslinking of 3-cyanovinylcarbazole ( CNV K) was accelerated at least 20 times, showing a faster DNA strand displacement. The rate of photo-crosslinking is a key factor and the rate of DNA strand displacement is accelerated through ultrafast photo-crosslinking. The rate of DNA strand displacement was regulated by photoirradiation energy. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Corrosion characteristics of unprotected post-tensioning strands under stress.

    DOT National Transportation Integrated Search

    2014-05-01

    An investigation was conducted to determine the effect of stress condition : and environmental exposure on corrosion of post-tensioned strands during ungrouted periods. : Exposures for periods of up to 4 weeks of stressed, as-received strand placed i...

  19. Temporal evolution of photon energy emitted from two-component advective flows: origin of time lag

    NASA Astrophysics Data System (ADS)

    Chatterjee, Arka; Chakrabarti, Sandip K.; Ghosh, Himadri

    2017-12-01

    X-ray time lag of black hole candidates contains important information regarding the emission geometry. Recently, study of time lags from observational data revealed very intriguing properties. To investigate the real cause of this lag behavior with energy and spectral states, we study photon paths inside a two-component advective flow (TCAF) which appears to be a satisfactory model to explain the spectral and timing properties. We employ the Monte Carlo simulation technique to carry out the Comptonization process. We use a relativistic thick disk in Schwarzschild geometry as the CENtrifugal pressure supported BOundary Layer (CENBOL) which is the Compton cloud. In TCAF, this is the post-shock region of the advective component. Keplerian disk on the equatorial plane which is truncated at the inner edge i.e. at the outer boundary of the CENBOL, acts as the soft photon source. Ray-tracing code is employed to track the photons to a distantly located observer. We compute the cumulative time taken by a photon during Comptonization, reflection and following the curved geometry on the way to the observer. Time lags between various hard and soft bands have been calculated. We study the variation of time lags with accretion rates, CENBOL size and inclination angle. Time lags for different energy channels are plotted for different inclination angles. The general trend of variation of time lag with QPO frequency and energy as observed in satellite data is reproduced.

  20. Identification of a broad-spectrum inhibitor of virus RNA synthesis: validation of a prototype virus-based approach

    PubMed Central

    Filone, Claire Marie; Hodges, Erin N.; Honeyman, Brian; Bushkin, G. Guy; Boyd, Karla; Platt, Andrew; Ni, Feng; Strom, Kyle; Hensley, Lisa; Snyder, John K.; Connor, John H.

    2013-01-01

    There are no approved therapeutics for the most deadly nonsegmented negative-strand (NNS) RNA viruses, including Ebola (EBOV). To identify new chemical scaffolds for development of broad-spectrum antivirals, we undertook a prototype-based lead identification screen. Using the prototype NNS virus, vesicular stomatitis virus (VSV), multiple inhibitory compounds were identified. Three compounds were investigated for broad-spectrum activity, and inhibited EBOV infection. The most potent, CMLDBU3402, was selected for further study. CMLDBU3402 did not show significant activity against segmented negative-strand RNA viruses suggesting proscribed broad-spectrum activity. Mechanistic analysis indicated that CMLDBU3402 blocked VSV viral RNA synthesis and inhibited EBOV RNA transcription, demonstrating a consistent mechanism of action against genetically distinct viruses. The identification of this chemical backbone as a broad-spectrum inhibitor of viral RNA synthesis offers significant potential for the development of new therapies for highly pathogenic viruses. PMID:23521799

  1. 75 FR 36678 - Prestressed Concrete Steel Wire Strand From China; Determinations

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-28

    ... Concrete Steel Wire Strand From China; Determinations On the basis of the record \\1\\ developed in the... prestressed concrete steel wire strand (PC strand), provided for in subheading 7312.10.30 of the Harmonized... Spring Wire Corp. (Bedford Heights, OH); Insteel Wire Products Co. (Mt. Airy, NC); and Sumiden Wire...

  2. A novel single fluorophore-labeled double-stranded oligonucleotide probe for fluorescence-enhanced nucleic acid detection based on the inherent quenching ability of deoxyguanosine bases and competitive strand-displacement reaction.

    PubMed

    Zhang, Yingwei; Tian, Jingqi; Li, Hailong; Wang, Lei; Sun, Xuping

    2012-01-01

    We develop a novel single fluorophore-labeled double-stranded oligonucleotide (OND) probe for rapid, nanostructure-free, fluorescence-enhanced nucleic acid detection for the first time. We further demonstrate such probe is able to well discriminate single-base mutation in nucleic acid. The design takes advantage of an inherent quenching ability of guanine bases. The short strand of the probe is designed with an end-labeled fluorophore that is placed adjacent to two guanines as the quencher located on the long opposite strand, resulting in great quenching of dye fluorescence. In the presence of a target complementary to the long strand of the probe, a competitive strand-displacement reaction occurs and the long strand forms a more stable duplex with the target, resulting in the two strands of the probe being separated from each other. As a consequence of this displacement, the fluorophore and the quencher are no longer in close proximity and dye fluorescence increases, signaling the presence of target.

  3. Analysis of Quasi-periodic Oscillations and Time Lag in Ultraluminous X-Ray Sources with XMM-Newton

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

    Li, Zi-Jian; Xiao, Guang-Cheng; Zhang, Shu

    We investigated the power density spectrum (PDS) and time lag of ultraluminous X-ray sources (ULXs) observed by XMM-Newton . We determined the PDSs for each ULX and found that five of them show intrinsic variability due to obvious quasi-periodic oscillations (QPOs) of mHz–1 Hz, consistent with previous reports. We further investigated these five ULXs to determine their possible time lag. The ULX QPOs exhibit a soft time lag that is linearly related to the QPO frequency. We discuss the likelihood of the ULX QPOs being type-C QPO analogs, and the time lag models. The ULXs might harbor intermediate-mass black holesmore » if their QPOs are type-C QPO analogs. We suggest that the soft lag and the linearity may be due to reverberation.« less

  4. Bioprinting Using Mechanically Robust Core-Shell Cell-Laden Hydrogel Strands.

    PubMed

    Mistry, Pritesh; Aied, Ahmed; Alexander, Morgan; Shakesheff, Kevin; Bennett, Andrew; Yang, Jing

    2017-06-01

    The strand material in extrusion-based bioprinting determines the microenvironments of the embedded cells and the initial mechanical properties of the constructs. One unmet challenge is the combination of optimal biological and mechanical properties in bioprinted constructs. Here, a novel bioprinting method that utilizes core-shell cell-laden strands with a mechanically robust shell and an extracellular matrix-like core has been developed. Cells encapsulated in the strands demonstrate high cell viability and tissue-like functions during cultivation. This process of bioprinting using core-shell strands with optimal biochemical and biomechanical properties represents a new strategy for fabricating functional human tissues and organs. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Mechanical behaviors of multi-filament twist superconducting strand under tensile and cyclic loading

    NASA Astrophysics Data System (ADS)

    Wang, Xu; Li, Yingxu; Gao, Yuanwen

    2016-01-01

    The superconducting strand, serving as the basic unit cell of the cable-in-conduit-conductors (CICCs), is a typical multi-filament twist composite which is always subjected to a cyclic loading under the operating condition. Meanwhile, the superconducting material Nb3Sn in the strand is sensitive to strain frequently relating to the performance degradation of the superconductivity. Therefore, a comprehensive study on the mechanical behavior of the strand helps understanding the superconducting performance of the strained Nb3Sn strands. To address this issue, taking the LMI (internal tin) strand as an example, a three-dimensional structural finite element model, named as the Multi-filament twist model, of the strand with the real configuration of the LMI strand is built to study the influences of the plasticity of the component materials, the twist of the filament bundle, the initial thermal residual stress and the breakage and its evolution of the filaments on the mechanical behaviors of the strand. The effective properties of superconducting filament bundle with random filament breakage and its evolution versus strain are obtained based on the damage theory of fiber-reinforced composite materials proposed by Curtin and Zhou. From the calculation results of this model, we find that the occurrence of the hysteresis loop in the cyclic loading curve is determined by the reverse yielding of the elastic-plastic materials in the strand. Both the initial thermal residual stress in the strand and the pitch length of the filaments have significant impacts on the axial and hysteretic behaviors of the strand. The damage of the filaments also affects the axial mechanical behavior of the strand remarkably at large axial strain. The critical current of the strand is calculated by the scaling law with the results of the Multi-filament twist model. The predicted results of the Multi-filament twist model show an acceptable agreement with the experiment.

  6. Molecular investigation of evaporation of biodroplets containing single-strand DNA on graphene surface.

    PubMed

    Akbari, Fahimeh; Foroutan, Masumeh

    2018-02-14

    In this study, the water droplet behaviour of four different types of single-strand DNA with homogeneous base sequence on a graphene substrate during evaporation of the droplet was investigated using molecular dynamics (MD) simulation. The simulation results indicated that the evaporation depended on the DNA sequence. The observed changes can be divided into four parts: (i) vaporization mode, (ii) evaporation flux, (iii) mechanism of single-strand placement on the surface, and (iv) consideration of remaining single strands after evaporation. Our simulation observations indicated different evaporation modes for thymine biodroplets as compared to those for other biodroplets. The evaporation of the thymine biodroplets occurred with an increase in the contact angle, while that of the other biodroplets occur in a constant contact angle mode. Moreover, thymine biodroplets generate the lowest contact line compared to other single strands, and it is always placed far away from the centre of the droplets during evaporation. Investigating variations in the evaporation flux shows that thymine has the highest evaporation flux and guanine has the lowest. Moreover, during initial evaporation, the flux of evaporation increases at the triple point of the biodroplets containing thymine single strands, while it decreases in the other biodroplets. The following observation was obtained from the study of the placement of single strands on the substrate: guanine and thymine interacted slower than other single strands during evaporation with graphene, adenine single strand had a higher folding during evaporation, and guanine single strand showed the lowest end-to-end distance. The investigation of single-strand DNA after evaporation shows that adenine produces the most stable structure at the end of evaporation. In addition, cytosine is the most stretched single-strand DNA due to its lack of internal π-π stacking and hydrogen bonding. Therefore, cytosine single strand is more

  7. ON THE ORIGIN OF THE {gamma}-RAY/OPTICAL LAGS IN LUMINOUS BLAZARS

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

    Janiak, Mateusz; Sikora, Marek; Moderski, Rafal

    2012-12-01

    Blazars are strongly variable sources that occasionally show spectacular flares visible in various energy bands. These flares are often, but not always, correlated. In a number of cases, the peaks of optical flares are found to be somewhat delayed with respect to the {gamma}-ray peaks. One notable example of such a delay was found in 3C 279 by Hayashida et al. and interpreted as a result of steeper drop with a distance of the energy density of an external radiation field than of the magnetic energy density. In this paper, we demonstrate that, in general, depending on the respective energymore » density profile along the jet, such lags can have both signs and that they can take place for any ratio of these energy densities. We study the dependence of such lags on the ratio of these energy densities at a distance of a maximal energy dissipation in a jet, on their gradients, as well as on the time profile of the relativistic electron injection within the moving source. We show how prominent such lags can be, and their expected timescales. We suggest that studies of such lags can provide a powerful tool to resolve the structure of relativistic jets and their radiative environment. As an example we model the lag observed in 3C 279, showing that in this object the flare is produced at a distance of a few parsecs from the central black hole, consistent with our previous inferences based on the spectra and optical polarization properties.« less

  8. Assessing delay and lag in sagittal trunk control using a tracking task.

    PubMed

    Reeves, N Peter; Luis, Abraham; Chan, Elizabeth C; Sal Y Rosas, Victor G; Tanaka, Martin L

    2018-05-17

    Slower trunk muscle responses are linked to back pain and injury. Unfortunately, clinical assessments of spine function do not objectively evaluate this important attribute, which reflects speed of trunk control. Speed of trunk control can be parsed into two components: (1) delay, the time it takes to initiate a movement, and (2) lag, the time it takes to execute a movement once initiated. The goal of this study is to demonstrate a new approach to assess delay and lag in trunk control using a simple tracking task. Ten healthy subjects performed four blocks of six trials of trunk tracking in the sagittal plane. Delay and lag were estimated by modeling trunk control for predictable and unpredictable (control mode) trunk movements in flexion and extension (control direction) at movement amplitudes of 2°, 4°, and 6° (control amplitude). The main effect of control mode, direction, and amplitude of movement were compared between trial blocks to assess secondary influencers (e.g., fatigue). Only control mode was consistent across trial blocks with predictable movements being faster than unpredictable for both delay and lag. Control direction and amplitude effects on delay and lag were consistent across the first two trial blocks and less consistent in later blocks. Given the heterogeneity in the presentation of back pain, clinical assessment of trunk control should include different control modes, directions, and amplitudes. To reduce testing time and the influence of fatigue, we recommend six trials to assess trunk control. Copyright © 2018 Elsevier Ltd. All rights reserved.

  9. Consequences of varied soil hydraulic and meteorological complexity on unsaturated zone time lag estimates.

    PubMed

    Vero, S E; Ibrahim, T G; Creamer, R E; Grant, J; Healy, M G; Henry, T; Kramers, G; Richards, K G; Fenton, O

    2014-12-01

    The true efficacy of a programme of agricultural mitigation measures within a catchment to improve water quality can be determined only after a certain hydrologic time lag period (subsequent to implementation) has elapsed. As the biophysical response to policy is not synchronous, accurate estimates of total time lag (unsaturated and saturated) become critical to manage the expectations of policy makers. The estimation of the vertical unsaturated zone component of time lag is vital as it indicates early trends (initial breakthrough), bulk (centre of mass) and total (Exit) travel times. Typically, estimation of time lag through the unsaturated zone is poor, due to the lack of site specific soil physical data, or by assuming saturated conditions. Numerical models (e.g. Hydrus 1D) enable estimates of time lag with varied levels of input data. The current study examines the consequences of varied soil hydraulic and meteorological complexity on unsaturated zone time lag estimates using simulated and actual soil profiles. Results indicated that: greater temporal resolution (from daily to hourly) of meteorological data was more critical as the saturated hydraulic conductivity of the soil decreased; high clay content soils failed to converge reflecting prevalence of lateral component as a contaminant pathway; elucidation of soil hydraulic properties was influenced by the complexity of soil physical data employed (textural menu, ROSETTA, full and partial soil water characteristic curves), which consequently affected time lag ranges; as the importance of the unsaturated zone increases with respect to total travel times the requirements for high complexity/resolution input data become greater. The methodology presented herein demonstrates that decisions made regarding input data and landscape position will have consequences for the estimated range of vertical travel times. Insufficiencies or inaccuracies regarding such input data can therefore mislead policy makers regarding

  10. TRACING THE REVERBERATION LAG IN THE HARD STATE OF BLACK HOLE X-RAY BINARIES

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

    De Marco, B.; Ponti, G.; Nandra, K.

    2015-11-20

    We report results obtained from a systematic analysis of X-ray lags in a sample of black hole X-ray binaries, with the aim of assessing the presence of reverberation lags and studying their evolution during outburst. We used XMM-Newton and simultaneous Rossi X-ray Timing Explorer (RXTE) observations to obtain broadband energy coverage of both the disk and the hard X-ray Comptonization components. In most cases the detection of reverberation lags is hampered by low levels of variability-power signal-to-noise ratio (typically when the source is in a soft state) and/or short exposure times. The most detailed study was possible for GX 339-4more » in the hard state, which allowed us to characterize the evolution of X-ray lags as a function of luminosity in a single source. Over all the sampled frequencies (∼0.05–9 Hz), we observe the hard lags intrinsic to the power-law component, already well known from previous RXTE studies. The XMM-Newton soft X-ray response allows us to detail the disk variability. At low frequencies (long timescales) the disk component always leads the power-law component. On the other hand, a soft reverberation lag (ascribable to thermal reprocessing) is always detected at high frequencies (short timescales). The intrinsic amplitude of the reverberation lag decreases as the source luminosity and the disk fraction increase. This suggests that the distance between the X-ray source and the region of the optically thick disk where reprocessing occurs gradually decreases as GX 339-4 rises in luminosity through the hard state, possibly as a consequence of reduced disk truncation.« less

  11. DNA polymerases eta and kappa exchange with the polymerase delta holoenzyme to complete common fragile site synthesis.

    PubMed

    Barnes, Ryan P; Hile, Suzanne E; Lee, Marietta Y; Eckert, Kristin A

    2017-09-01

    Common fragile sites (CFSs) are inherently unstable genomic loci that are recurrently altered in human tumor cells. Despite their instability, CFS are ubiquitous throughout the human genome and associated with large tumor suppressor genes or oncogenes. CFSs are enriched with repetitive DNA sequences, one feature postulated to explain why these loci are inherently difficult to replicate, and sensitive to replication stress. We have shown that specialized DNA polymerases (Pols) η and κ replicate CFS-derived sequences more efficiently than the replicative Pol δ. However, we lacked an understanding of how these enzymes cooperate to ensure efficient CFS replication. Here, we designed a model of lagging strand replication with RFC loaded PCNA that allows for maximal activity of the four-subunit human Pol δ holoenzyme, Pol η, and Pol κ in polymerase mixing assays. We discovered that Pol η and κ are both able to exchange with Pol δ stalled at repetitive CFS sequences, enhancing Normalized Replication Efficiency. We used this model to test the impact of PCNA mono-ubiquitination on polymerase exchange, and found no change in polymerase cooperativity in CFS replication compared with unmodified PCNA. Finally, we modeled replication stress in vitro using aphidicolin and found that Pol δ holoenzyme synthesis was significantly inhibited in a dose-dependent manner, preventing any replication past the CFS. Importantly, Pol η and κ were still proficient in rescuing this stalled Pol δ synthesis, which may explain, in part, the CFS instability phenotype of aphidicolin-treated Pol η and Pol κ-deficient cells. In total, our data support a model wherein Pol δ stalling at CFSs allows for free exchange with a specialized polymerase that is not driven by PCNA. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Separation of 1-23-kb complementary DNA strands by urea-agarose gel electrophoresis.

    PubMed

    Hegedüs, Eva; Kókai, Endre; Kotlyar, Alexander; Dombrádi, Viktor; Szabó, Gábor

    2009-09-01

    Double-stranded (ds), as well as denatured, single-stranded (ss) DNA samples can be analyzed on urea-agarose gels. Here we report that after denaturation by heat in the presence of 8 M urea, the two strands of the same ds DNA fragment of approximately 1-20-kb size migrate differently in 1 M urea containing agarose gels. The two strands are readily distinguished on Southern blots by ss-specific probes. The different migration of the two strands could be attributed to their different, base composition-dependent conformation impinging on the electrophoretic mobility of the ss molecules. This phenomenon can be exploited for the efficient preparation of strand-specific probes and for the separation of the complementary DNA strands for subsequent analysis, offering a new tool for various cell biological research areas.

  13. Accretion Disk Reverberation with Hubble Space Telescope Observations of NGC 4593: Evidence for Diffuse Continuum Lags

    NASA Astrophysics Data System (ADS)

    Cackett, Edward M.; Chiang, Chia-Ying; McHardy, Ian; Edelson, Rick; Goad, Michael R.; Horne, Keith; Korista, Kirk T.

    2018-04-01

    The Seyfert 1 galaxy NGC 4593 was monitored spectroscopically with the Hubble Space Telescope as part of a reverberation mapping campaign that also included Swift, Kepler, and ground-based photometric monitoring. During 2016 July 12–August 6, we obtained 26 spectra across a nearly continuous wavelength range of ∼1150–10000 Å. These were combined with Swift data to produce a UV/optical “lag spectrum,” which shows the interband lag relative to the Swift UVW2 band as a function of wavelength. The broad shape of the lag spectrum appears to follow the τ ∝ λ 4/3 relation seen previously in photometric interband lag measurements of other active galactic nuclei (AGNs). This shape is consistent with the standard thin disk model, but the magnitude of the lags implies a disk that is a factor of ∼3 larger than predicted, again consistent with what has been previously seen in other AGNs. In all cases these large disk sizes, which are also implied by independent gravitational microlensing of higher-mass AGNs, cannot be simply reconciled with the standard model. However, the most striking feature in this higher-resolution lag spectrum is a clear excess around the 3646 Å Balmer jump. This strongly suggests that diffuse emission from gas in the much larger broad-line region (BLR) must also contribute significantly to the interband lags. While the relative contributions of the disk and BLR cannot be uniquely determined in these initial measurements, it is clear that both will need to be considered to comprehensively model and understand AGN lag spectra.

  14. Does the benefit of testing depend on lag, and if so, why? Evaluating the elaborative retrieval hypothesis.

    PubMed

    Rawson, Katherine A; Vaughn, Kalif E; Carpenter, Shana K

    2015-05-01

    Despite the voluminous literatures on testing effects and lag effects, surprisingly few studies have examined whether testing and lag effects interact, and no prior research has directly investigated why this might be the case. To this end, in the present research we evaluated the elaborative retrieval hypothesis (ERH) as a possible explanation for why testing effects depend on lag. Elaborative retrieval involves the activation of cue-related information during the long-term memory search for the target. If the target is successfully retrieved, this additional information is encoded with the cue-target pair to yield a more elaborated memory trace that enhances target access on a later memory test. The ERH states that the degree of elaborative retrieval during practice is greater when testing takes place after a long rather than a short lag (whereas elaborative retrieval during restudy is minimal at either lag). Across two experiments, final-test performance was greater following practice testing than following restudy only, and this memorial advantage was greater with long-lag than with short-lag practice. The final test also included novel cue conditions used to diagnose the degree of elaborative retrieval during practice. The overall pattern of performance in these conditions provided consistent evidence for the ERH, with more extensive elaborative retrieval during long- than during short-lag practice testing.

  15. Proceedings of a Workshop on Antarctic Meteorite Stranding Surfaces

    NASA Technical Reports Server (NTRS)

    Cassidy, W. A. (Editor); Whillans, I. M. (Editor)

    1990-01-01

    The discovery of large numbers of meteorites on the Antarctic Ice Sheet is one of the most exciting developments in polar science in recent years. The meteorites are found on areas of ice called stranding surfaces. Because of the sudden availability of hundreds, and then thousands, of new meteorite specimens at these sites, the significance of the discovery of meteorite stranding surfaces in Antarctica had an immediate and profound impact on planetary science, but there is also in this discovery an enormous, largely unrealized potential to glaciology for records of climatic and ice sheet changes. The glaciological interest derives from the antiquity of the ice in meteorite stranding surfaces. This exposed ice covers a range of ages, probably between zero and more than 500,000 years. The Workshop on Antarctic Meteorite Stranding Surfaces was convened to explore this potential and to devise a course of action that could be recommended to granting agencies. The workshop recognized three prime functions of meteorite stranding surfaces. They provide: (1) A proxy record of climatic change (i.e., a long record of climatic change is probably preserved in the exposed ice stratigraphy); (2) A proxy record of ice volume change; and (3) A source of unique nonterrestrial material.

  16. Function and regulation of LAG3 on CD4+CD25- T cells in non-small cell lung cancer.

    PubMed

    Ma, Qin-Yun; Huang, Da-Yu; Zhang, Hui-Jun; Wang, Shaohua; Chen, Xiao-Feng

    2017-11-15

    LAG3 is a surface molecule found on a subset of immune cells. The precise function of LAG3 appears to be context-dependent. In this study, we investigated the effect of LAG3 on CD4 + CD25 - T cells from non-small cell lung cancer (NSCLC) patients. We found that in the peripheral blood mononuclear cells of NSCLC patients, LAG3 was significantly increased in CD4 + T cells directly ex vivo and primarily in the CD4 + CD25 - fraction, which was regulated by prolonged TCR stimulation and the presence of IL-27. TCR stimulation also increased CD25 expression, but not Foxp3 expression, in LAG3-expressing CD4 + CD25 - cells Compared to LAG3-nonexpressing CD4 + CD25 - cells, LAG3-expressing CD4 + CD25 - cells presented significantly higher levels of PD1 and TIM3, two inhibitory receptors best described in exhausted CD8 + T effector cells. LAG3-expressing CD4 + CD25 - cells also presented impaired proliferation compared with LAG3-nonexpressing CD4 + CD25 - cells but could be partially rescued by inhibiting both PD1 and TIM3. Interestingly, CD8 + T cells co-incubated with LAG3-expressing CD4 + CD25 - cells at equal cell numbers demonstrated significantly lower proliferation than CD8 + T cells incubated alone. Co-culture with CD8 + T cell and LAG3-expressing CD4 + CD25 - T cell also upregulated soluble IL-10 level in the supernatant, of which the concentration was positively correlated with the number of LAG3-expressing CD4 + CD25 - T cells. In addition, we found that LAG3-expressing CD4 + CD25 - T cells infiltrated the resected tumors and were present at higher frequencies of in metastases than in primary tumors. Taken together, these data suggest that LAG3-expressing CD4 + CD25 - T cells represent another regulatory immune cell type with potential to interfere with anti-tumor immunity. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Tensile and dimensional properties of wood strands made from plantation southern pine lumber

    Treesearch

    Qinglin Wu; Zhiyong Cai; Jong N. Lee

    2005-01-01

    Working stresses and performance of strand composite lumber largely depend upon the properties of each individual strand. Southern pine strands from plantation lumber grown in southern Louisiana were investigated in this study in order to understand strand behaviors. The effects of hot-pressing and resin application on tensile modulus, strength, and dimensional...

  18. Method for fabricating multi-strand superconducting cable

    DOEpatents

    Borden, A.R.

    1985-04-01

    Multi-strand superconducting cables adapted to be used, for example, to wind a magnet are fabricated by directing wire strands inwardly from spools disposed on the perimeter of a rotating disk and wrapping them diagonally around a tapered mandrel with a flattened cross-sectional shape with a core having a wedge-shaped channel. As the cable is pulled axially, flexibly coupled wedge-shaped pieces are continuously passed through the channel in the mandrel and inserted into the cable as an internal support therefor.

  19. The influence of interactions between accommodation and convergence on the lag of accommodation.

    PubMed

    Schor, C

    1999-03-01

    Several models of myopia predict that growth of axial length is stimulated by blur. Accommodative lag has been suggested as an important source of blur in the development of myopia and this study has modeled how cross-link interactions between accommodation and convergence might interact with uncorrected distance heterophoria and refractive error to influence accommodative lag. Accommodative lag was simulated with two models of interactions between accommodation and convergence (one with and one without adaptable tonic elements). Simulations of both models indicate that both uncorrected hyperopia and esophoria increase the lag of accommodative and uncorrected myopia and exophoria decrease the lag or introduce a lead of accommodation in response to the near (40 cm) stimulus. These effects were increased when gain of either cross-link, accommodative convergence (AC/A) or convergence accommodation (CA/C), was increased within a moderate range of values while the other was fixed at a normal value (clamped condition). These effects were exaggerated when both the AC/A and CA/C ratios were increased (covaried condition) and affects of cross-link gain were negated when an increase of one cross-link (e.g. AC/A) was accompanied by a reduction of the other cross-link (e.g. CA/C) (reciprocal condition). The inclusion of tonic adaptation in the model reduced steady state errors of accommodation for all conditions except when the AC/A ratio was very high (2 MA/D). Combinations of cross-link interactions between accommodation and convergence that resemble either clamped or reciprocal patterns occur naturally in clinical populations. Simulations suggest that these two patterns of abnormal cross-link interactions could affect the progression of myopia differently. Adaptable tonic accommodation and tonic vergence could potentially reduce the progression of myopia by reducing the lag of accommodation.

  20. Increasing Vocal Variability in Children with Autism Using a Lag Schedule of Reinforcement

    ERIC Educational Resources Information Center

    Esch, John W.; Esch, Barbara E.; Love, Jessa R.

    2009-01-01

    Variability has been demonstrated to be an operant dimension of behavior (Neuringer, 2002; Page & Neuringer, 1985). Recently, lag schedules have been used to demonstrate operant variability of verbal behavior in persons with a diagnosis of autism (e.g., Lee, McComas, & Jawor, 2002). The current study evaluated the effects of a Lag 1 schedule on…

  1. Improving strand quality of upland oaks for use in oriented strand board

    Treesearch

    David B. DeValliance; Jody D. Gray; Shawn T. Grushecky

    2013-01-01

    Past research estimates that more than 1 million tons of oak logging residues go unused in West Virginia each year. Much research has been done investigating potential products and markets for this underutilized resource. West Virginia is home to an oriented strand board (OSB) producer that consumes large volumes of small diameter, low quality round wood. However, the...

  2. Estimation of watershed lag times and times of concentration for the Kansas City Area.

    DOT National Transportation Integrated Search

    2016-04-01

    Lag time (TL) and time of concentration (TC) are two related measures of how quickly a stream responds to : runoff-producing rainfall over its watershed. In this report, a general relationship for lag time is derived from the : Manning equation for f...

  3. High-pressure liquid-monopropellant strand combustion.

    NASA Technical Reports Server (NTRS)

    Faeth, G. M.

    1972-01-01

    Examination of the influence of dissolved gases on the state of the liquid surface during high-pressure liquid-monopropellant combustion through the use of a strand burning experiment. Liquid surface temperatures were measured, using fine-wire thermocouples, during the strand combustion of ethyl nitrate, normal propyl nitrate, and propylene glycol dinitrate at pressures up to 81 atm. These measurements were compared with the predictions of a variable-property gas-phase analysis assuming an infinite activation energy for the decomposition reaction. The state of the liquid surface was estimated using a conventional low-pressure phase equilibrium model, as well as a high-pressure version that considered the presence of dissolved combustion-product gases in the liquid phase. The high-pressure model was found to give a superior prediction of measured liquid surface temperatures. Computed total pressures required for the surface to reach its critical mixing point during strand combustion were found to be in the range from 2.15 to 4.62 times the critical pressure of the pure propellant. Computed dissolved gas concentrations at the liquid surface were in the range from 35 to 50% near the critical combustion condition.

  4. Double-stranded RNA virus in the human pathogenic fungus Blastomyces dermatitidis.

    PubMed Central

    Kohno, S; Fujimura, T; Rulong, S; Kwon-Chung, K J

    1994-01-01

    Double-stranded RNA viruses were detected in a strain of Blastomyces dermatitidis isolated from a patient in Uganda. The viral particles are spherical (mostly 44 to 50 nm in diameter) and consist of about 25% double-stranded RNA (5 kb) and 75% protein (90 kDa). The virus contains transcriptional RNA polymerase activity; it synthesized single-stranded RNA in vitro in a conservative manner. The newly synthesized single-stranded RNA was a full-length strand, and the rate of chain elongation was approximately 170 nucleotides per min. The virus-containing strain shows no morphological difference from virus-free strains in the mycelial phase. Although the association with the presence of the virus is unclear, the virus-infected strain converts to the yeast form at 37 degrees C, but the yeast cells fail to multiply at that temperature. Images PMID:7933142

  5. High-order flux correction/finite difference schemes for strand grids

    NASA Astrophysics Data System (ADS)

    Katz, Aaron; Work, Dalon

    2015-02-01

    A novel high-order method combining unstructured flux correction along body surfaces and high-order finite differences normal to surfaces is formulated for unsteady viscous flows on strand grids. The flux correction algorithm is applied in each unstructured layer of the strand grid, and the layers are then coupled together via a source term containing derivatives in the strand direction. Strand-direction derivatives are approximated to high-order via summation-by-parts operators for first derivatives and second derivatives with variable coefficients. We show how this procedure allows for the proper truncation error canceling properties required for the flux correction scheme. The resulting scheme possesses third-order design accuracy, but often exhibits fourth-order accuracy when higher-order derivatives are employed in the strand direction, especially for highly viscous flows. We prove discrete conservation for the new scheme and time stability in the absence of the flux correction terms. Results in two dimensions are presented that demonstrate improvements in accuracy with minimal computational and algorithmic overhead over traditional second-order algorithms.

  6. Interpreting the spatio-temporal patterns of sea turtle strandings: Going with the flow

    USGS Publications Warehouse

    Hart, K.M.; Mooreside, P.; Crowder, L.B.

    2006-01-01

    Knowledge of the spatial and temporal distribution of specific mortality sources is crucial for management of species that are vulnerable to human interactions. Beachcast carcasses represent an unknown fraction of at-sea mortalities. While a variety of physical (e.g., water temperature) and biological (e.g., decomposition) factors as well as the distribution of animals and their mortality sources likely affect the probability of carcass stranding, physical oceanography plays a major role in where and when carcasses strand. Here, we evaluate the influence of nearshore physical oceanographic and wind regimes on sea turtle strandings to decipher seasonal trends and make qualitative predictions about stranding patterns along oceanfront beaches. We use results from oceanic drift-bottle experiments to check our predictions and provide an upper limit on stranding proportions. We compare predicted current regimes from a 3D physical oceanographic model to spatial and temporal locations of both sea turtle carcass strandings and drift bottle landfalls. Drift bottle return rates suggest an upper limit for the proportion of sea turtle carcasses that strand (about 20%). In the South Atlantic Bight, seasonal development of along-shelf flow coincides with increased numbers of strandings of both turtles and drift bottles in late spring and early summer. The model also predicts net offshore flow of surface waters during winter - the season with the fewest relative strandings. The drift bottle data provide a reasonable upper bound on how likely carcasses are to reach land from points offshore and bound the general timeframe for stranding post-mortem (< two weeks). Our findings suggest that marine turtle strandings follow a seasonal regime predictable from physical oceanography and mimicked by drift bottle experiments. Managers can use these findings to reevaluate incidental strandings limits and fishery takes for both nearshore and offshore mortality sources. ?? 2005 Elsevier Ltd

  7. Diphosphates at the 5' end of the positive strand of yeast L-A double-stranded RNA virus as a molecular self-identity tag.

    PubMed

    Fujimura, Tsutomu; Esteban, Rosa

    2016-10-01

    The 5'end of RNA conveys important information on self-identity. In mammalian cells, double-stranded RNA (dsRNA) with 5'di- or triphosphates generated during virus infection is recognized as foreign and elicits the host innate immune response. Here, we analyze the 5' ends of the dsRNA genome of the yeast L-A virus. The positive strand has largely diphosphates with a minor amount of triphosphates, while the negative strand has only diphosphates. Although the virus can produce capped transcripts by cap snatching, neither strand carried a cap structure, suggesting that only non-capped transcripts serve as genomic RNA for encapsidation. We also found that the 5' diphosphates of the positive but not the negative strand within the dsRNA genome are crucial for transcription in vitro. Furthermore, the presence of a cap structure in the dsRNA abrogated its template activity. Given that the 5' diphosphates of the transcripts are also essential for cap acquisition and that host cytosolic RNAs (mRNA, rRNA, and tRNA) are uniformly devoid of 5' pp-structures, the L-A virus takes advantage of its 5' terminal diphosphates, using them as a self-identity tag to propagate in the host cytoplasm. © 2016 John Wiley & Sons Ltd.

  8. Corrosion of post-tensioning strands in ungrouted ducts - unstressed condition

    NASA Astrophysics Data System (ADS)

    Hutchison, Michael

    Recent failures and severe corrosion distress of post-tensioned (PT) bridges in Florida have revealed corrosion of the 7-wire strands in tendons. Post-tensioned duct assemblies are fitted with multiple 7-wire steel strands and ducts are subsequently filled with grout. During construction, the length of time from the moment in which the strands have been inserted into the ducts, until the ducts are grouted, is referred to as the `ungrouted' period. During this phase, the steel strands are vulnerable to corrosion and consequently the length of this period is restricted (typically to 7 days) by construction guidelines. This investigation focuses on determining the extent of corrosion that may take place during that period, but limited to strands that were in the unstressed condition. Visual inspections and tensile testing were used to identify trends in corrosion development. Corrosion induced cracking mechanisms were also investigated via wire bending and metallographic cross section evaluation. Corrosion damage on unstressed strands during ungrouted periods of durations in the order of those otherwise currently prescribed did not appear to seriously degrade mechanical performance as measured by standardized tests. However the presence of stress in the ungrouted period, as is normally the case, may activate other mechanisms (e.g., EAC) that require further investigation. As expected in the unstressed condition, no evidence of transverse cracking was observed.

  9. A comparison of alternative variants of the lead and lag time TTO.

    PubMed

    Devlin, Nancy; Buckingham, Ken; Shah, Koonal; Tsuchiya, Aki; Tilling, Carl; Wilkinson, Grahame; van Hout, Ben

    2013-05-01

    'Lead Time' TTO improves upon conventional TTO by providing a uniform method for eliciting positive and negative values. This research investigates (i) the values generated from different combinations of time in poor health and in full health; and the order in which these appear (lead vs. lag); (ii) whether values concur with participants' views about states; (iii) methods for handling extreme preferences. n = 208 participants valued five EQ-5D states, using two of four variants. Combinations of lead time and health state duration were: 10 years and 20 years; 5 years and 1 year; 5 years and 10 years; and a health state duration of 5 years with a lag time of 10 years. Longer lead times capture more preferences, but may involve a framing effect. Lag time results in less non-trading for mild states, and less time being traded for severe states. Negative values broadly agree with participants' stated opinion that the state is worse than dead. The values are sensitive to the ratio of lead time to duration of poor health, and the order in which these appear (lead vs. lag). It is feasible to handle extreme preferences though challenges remain. Copyright © 2012 John Wiley & Sons, Ltd.

  10. Development of a 10 m quasi-isotropic strand assembled from 2G wires

    NASA Astrophysics Data System (ADS)

    Kan, Changtao; Wang, Yinshun; Hou, Yanbing; Li, Yan; Zhang, Han; Fu, Yu; Jiang, Zhe

    2018-03-01

    Quasi-isotropic strands made of second generation (2G) high temperature superconducting (HTS) wires are attractive to applications of high-field magnets at low temperatures and power transmission cables at liquid nitrogen temperature in virtue of their high current carrying capability and well mechanical property. In this contribution, a 10 m length quasi-isotropic strand is manufactured and successfully tested in liquid nitrogen to verify the feasibility of an industrial scale production of the strand by the existing cabling technologies. The strand with copper sheath consists of 72 symmetrically assembled 2G wires. The uniformity of critical properties of long quasi-isotropic strands, including critical current and n-value, is very important for their using. Critical currents as well as n-values of the strand are measured every 1 m respectively and compared with the simulation results. Critical current and n-value of the strand are calculated basing on the self-consistent model solved by the finite element method (FEM). Effects of self-field on the critical current and n-value distributions in wires of the strand are analyzed in detail. The simulation results show good agreement with the experimental data and the 10 m quasi-isotropic strand has good critical properties uniformity.

  11. Corrosion characteristics of post-tensioning strands in ungrouted ducts : summary.

    DOT National Transportation Integrated Search

    2011-01-01

    To prevent corrosion of post-tensioning strands, FDOT construction specifications currently require post-tensioning ducts to be grouted within seven calendar days of strand installation. This period challenges construction schedules on large projects...

  12. Lagging skills contribute to challenging behaviors in children with autism spectrum disorder without intellectual disability.

    PubMed

    Maddox, Brenna B; Cleary, Patrick; Kuschner, Emily S; Miller, Judith S; Armour, Anna Chelsea; Guy, Lisa; Kenworthy, Lauren; Schultz, Robert T; Yerys, Benjamin E

    2017-08-01

    Many children with autism spectrum disorder display challenging behaviors. These behaviors are not limited to those with cognitive and/or language impairments. The Collaborative and Proactive Solutions framework proposes that challenging behaviors result from an incompatibility between environmental demands and a child's "lagging skills." The primary Collaborative and Proactive Solutions lagging skills-executive function, emotion regulation, language, and social skills-are often areas of weakness for individuals with autism spectrum disorder. The purpose of this study was to evaluate whether these lagging skills are associated with challenging behaviors in youth with autism spectrum disorder without intellectual disability. Parents of 182 youth with autism spectrum disorder (6-15 years) completed measures of their children's challenging behaviors, executive function, language, emotion regulation, and social skills. We tested whether the Collaborative and Proactive Solutions lagging skills predicted challenging behaviors using multiple linear regression. The Collaborative and Proactive Solutions lagging skills explained significant variance in participants' challenging behaviors. The Depression (emotion regulation), Inhibit (executive function), and Sameness (executive function) scales emerged as significant predictors. Impairments in emotion regulation and executive function may contribute substantially to aggressive and oppositional behaviors in school-age youth with autism spectrum disorder without intellectual disability. Treatment for challenging behaviors in this group may consider targeting the incompatibility between environmental demands and a child's lagging skills.

  13. Current Status of Biomedical Book Reviewing: Part II. Time Lag in Biomedical Book Reviewing

    PubMed Central

    Chen, Ching-Chih

    1974-01-01

    This part of the study explores the effectiveness of the review media in terms of speed of reviewing, comprehensiveness of review treatment, and authority. The time lags for the fifty-four journals varied widely, the mean ranging from 5.8 months to forty-two months. The time lags for all 3,347 reviews varied even more widely, ranging from less than a month to 108 months after a book was off the press. The 3,347 reviews had a mean time lag of 10.43 months and a standard deviation of 6.63 months. PMID:4826480

  14. Recovery of stranded costs under electric deregulation: The Winstar doctrine

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

    Person, J.C.

    This paper explores the applicability of the Winstar doctrine to the recovery of stranded costs arising from the deregulation of the electric utility industry. Such stranded costs, which have been widely estimated to be in the $100--200 billion range, represent those utility assets whose book value exceed their market value. Not addressed in this paper are the ongoing state and federal legislative initiatives to allow for the recovery of some or all of a utility`s stranded costs, such as through the assessment of competitive transmission charges (CTCs) or through stranded cost securitization. Rather, this paper presents one of several ofmore » legal arguments that could be utilized in those situations where a legislative solution either does not exist or does not allow for full book value recovery.« less

  15. Statistical Assessment of Cetacean Stranding Events in Cape Cod Area

    NASA Technical Reports Server (NTRS)

    Zellar, R.; Pulkkinen, A.; Moore, K.; Reeb, D.; Karakoylu, E.; Uritskaya, O.

    2017-01-01

    Cetacean (whales, dolphins and porpoises) mass strandings are a longstanding mystery in the field of marine biology that continue to be recorded in coastal environments around the world. For each of these events, anywhere from a few to several hundred otherwise healthy animals strand in onshore environments, often for no apparent reason. While the causes of these events remain unclear, anthropogenic and naturogenic mechanisms have been suggested. We present results of an inter-disciplinary study that draws expertise from space weather, marine mammal biology and ecology, and marine mammal stranding response. This study assessed 16 years of cetacean stranding events in the Cape Cod (Massachusetts, USA) area concurrently with a large dataset of meteorological, geophysical, biological, oceanographic and space weather data to produce inferences about possible causes for these unexplained events.

  16. Stem cell identity and template DNA strand segregation.

    PubMed

    Tajbakhsh, Shahragim

    2008-12-01

    The quest for stem cell properties to distinguish their identity from that of committed daughters has led to a re-investigation of the notion that DNA strands are not equivalent, and 'immortal' DNA strands are retained in stem cells whereas newly replicated DNA strands segregate to the differentiating daughter cell during mitosis. Whether this process occurs only in stem cells, and also in all tissues, remains unclear. That individual chromosomes can be also partitioned non-randomly raises the question if this phenomenon is related to the immortal DNA hypothesis, and it underscores the need for high-resolution techniques to observe these events empirically. Although initially postulated as a mechanism to avoid DNA replication errors, alternative views including epigenetic regulation and sister chromatid silencing may provide insights into this process.

  17. Lag threads organize the brain’s intrinsic activity

    PubMed Central

    Mitra, Anish; Snyder, Abraham Z.; Blazey, Tyler; Raichle, Marcus E.

    2015-01-01

    It has been widely reported that intrinsic brain activity, in a variety of animals including humans, is spatiotemporally structured. Specifically, propagated slow activity has been repeatedly demonstrated in animals. In human resting-state fMRI, spontaneous activity has been understood predominantly in terms of zero-lag temporal synchrony within widely distributed functional systems (resting-state networks). Here, we use resting-state fMRI from 1,376 normal, young adults to demonstrate that multiple, highly reproducible, temporal sequences of propagated activity, which we term “lag threads,” are present in the brain. Moreover, this propagated activity is largely unidirectional within conventionally understood resting-state networks. Modeling experiments show that resting-state networks naturally emerge as a consequence of shared patterns of propagation. An implication of these results is that common physiologic mechanisms may underlie spontaneous activity as imaged with fMRI in humans and slowly propagated activity as studied in animals. PMID:25825720

  18. Sub-Ensemble Monitoring of DNA Strand Displacement Using Multiparameter Single-Molecule FRET.

    PubMed

    Baltierra-Jasso, Laura E; Morten, Michael J; Magennis, Steven W

    2018-03-05

    Non-enzymatic DNA strand displacement is an important mechanism in dynamic DNA nanotechnology. Here, we show that the large parameter space that is accessible by single-molecule FRET is ideal for the simultaneous monitoring of multiple reactants and products of DNA strand exchange reactions. We monitored the strand displacement from double-stranded DNA (dsDNA) by single-stranded DNA (ssDNA) at 37 °C; the data were modelled as a second-order reaction approaching equilibrium, with a rate constant of 10 m -1  s -1 . We also followed the displacement from a DNA three-way junction (3WJ) by ssDNA. The presence of three internal mismatched bases in the middle of the invading strand did not prevent displacement from the 3WJ, but reduced the second-order rate constant by about 50 %. We attribute strand exchange in the dsDNA and 3WJ to a zero-toehold pathway from the blunt-ended duplex arms. The single-molecule approach demonstrated here will be useful for studying complex DNA networks. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Reverse Flood Routing with the Lag-and-Route Storage Model

    NASA Astrophysics Data System (ADS)

    Mazi, K.; Koussis, A. D.

    2010-09-01

    This work presents a method for reverse routing of flood waves in open channels, which is an inverse problem of the signal identification type. Inflow determination from outflow measurements is useful in hydrologic forensics and in optimal reservoir control, but has been seldom studied. Such problems are ill posed and their solution is sensitive to small perturbations present in the data, or to any related uncertainty. Therefore the major difficulty in solving this inverse problem consists in controlling the amplification of errors that inevitably befall flow measurements, from which the inflow signal is to be determined. The lag-and-route model offers a convenient framework for reverse routing, because not only is formal deconvolution not required, but also reverse routing is through a single linear reservoir. In addition, this inversion degenerates to calculating the intermediate inflow (prior to the lag step) simply as the sum of the outflow and of its time derivative multiplied by the reservoir’s time constant. The remaining time shifting (lag) of the intermediate, reversed flow presents no complications, as pure translation causes no error amplification. Note that reverse routing with the inverted Muskingum scheme (Koussis et al., submitted to the 12th Plinius Conference) fails when that scheme is specialised to the Kalinin-Miljukov model (linear reservoirs in series). The principal functioning of the reverse routing procedure was verified first with perfect field data (outflow hydrograph generated by forward routing of a known inflow hydrograph). The field data were then seeded with random error. To smooth the oscillations caused by the imperfect (measured) outflow data, we applied a multipoint Savitzky-Golay low-pass filter. The combination of reverse routing and filtering achieved an effective recovery of the inflow signal extremely efficiently. Specifically, we compared the reverse routing results of the inverted lag-and-route model and of the inverted

  20. Development length of 0.6-inch prestressing strand in standard I-shaped pretensioned concrete beams

    NASA Astrophysics Data System (ADS)

    Barnes, Robert Wesley

    The use of 0.6 in prestressing strand at a center-to-center spacing of 2 in allows for the optimal implementation of High Strength Concrete (HSC) in precast, prestressed concrete bridge superstructures. For this strand configuration, partial debonding of strands is a desirable alternative to the more traditional method of draping strands to alleviate extreme concrete stresses after prestress release. Recent experimental evidence suggests that existing code provisions addressing the anchorage of pretensioned strands do not adequately describe the behavior of these strands. In addition, the anchorage behavior of partially debonded strands is not fully understood. These uncertainties have combined to hinder the full exploitation of HSC in pretensioned concrete construction. A research study was conducted to determine the anchorage behavior of 0.6 in strands at 2 in spacing in full-size bridge members. The experimental program consisted of assessing transfer and development lengths in plant-cast AASHTO Type I I-beams. The influence of concrete compressive strengths ranging from 5700 to 14,700 psi was examined. In order to consider the full range of strand surface conditions found in practice, the prestressing strand featured either a bright mill finish or a rusted surface condition. The anchorage behavior of partially debonded strands was investigated by using a variety of strand debonding configurations---including debonded strand percentages as high as 75 percent. A limited investigation of the effect of horizontal web reinforcement on anchorage behavior was performed. Pull-out tests were performed in an attempt to correlate results with the bond quality of the strands used in the study. The correlation between strand draw-in and the anchorage behavior of prestressing strands was also examined. A review of the evolution and shortcomings of existing code provisions for the anchorage of prestressing strands is presented. Results of the experimental program are reported

  1. The Effect of Basepair Mismatch on DNA Strand Displacement.

    PubMed

    Broadwater, D W Bo; Kim, Harold D

    2016-04-12

    DNA strand displacement is a key reaction in DNA homologous recombination and DNA mismatch repair and is also heavily utilized in DNA-based computation and locomotion. Despite its ubiquity in science and engineering, sequence-dependent effects of displacement kinetics have not been extensively characterized. Here, we measured toehold-mediated strand displacement kinetics using single-molecule fluorescence in the presence of a single basepair mismatch. The apparent displacement rate varied significantly when the mismatch was introduced in the invading DNA strand. The rate generally decreased as the mismatch in the invader was encountered earlier in displacement. Our data indicate that a single base pair mismatch in the invader stalls branch migration and displacement occurs via direct dissociation of the destabilized incumbent strand from the substrate strand. We combined both branch migration and direct dissociation into a model, which we term the concurrent displacement model, and used the first passage time approach to quantitatively explain the salient features of the observed relationship. We also introduce the concept of splitting probabilities to justify that the concurrent model can be simplified into a three-step sequential model in the presence of an invader mismatch. We expect our model to become a powerful tool to design DNA-based reaction schemes with broad functionality. Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  2. Diverse Responses of Global Vegetation to Climate Changes: Spatial Patterns and Time-lag Effects

    NASA Astrophysics Data System (ADS)

    Wu, D.; Zhao, X.; Zhou, T.; Huang, K.; Xu, W.

    2014-12-01

    Global climate changes have enormous influences on vegetation growth, meanwhile, response of vegetation to climate express space diversity and time-lag effects, which account for spatial-temporal disparities of climate change and spatial heterogeneity of ecosystem. Revelation of this phenomenon will help us further understanding the impact of climate change on vegetation. Assessment and forecast of global environmental change can be also improved under further climate change. Here we present space diversity and time-lag effects patterns of global vegetation respond to three climate factors (temperature, precipitation and solar radiation) based on quantitative analysis of satellite data (NDVI) and Climate data (Climate Research Unit). We assessed the time-lag effects of global vegetation to main climate factors based on the great correlation fitness between NDVI and the three climate factors respectively among 0-12 months' temporal lags. On this basis, integrated response model of NDVI and the three climate factors was built to analyze contribution of different climate factors to vegetation growth with multiple regression model and partial correlation model. In the result, different vegetation types have distinct temporal lags to the three climate factors. For the precipitation, temporal lags of grasslands are the shortest while the evergreen broad-leaf forests are the longest, which means that grasslands are more sensitive to precipitation than evergreen broad-leaf forests. Analysis of different climate factors' contribution to vegetation reveal that vegetation are dominated by temperature in the high northern latitudes; they are mainly restricted by precipitation in arid and semi-arid areas (Australia, Western America); in humid areas of low and intermediate latitudes (Amazon, Eastern America), vegetation are mainly influenced by solar radiation. Our results reveal the time-lag effects and major driving factors of global vegetation growth and explain the

  3. Transport and time lag of chlorofluorocarbon gases in the unsaturated zone, Rabis Creek, Denmark

    USGS Publications Warehouse

    Engesgaard, Peter; Højberg, Anker L.; Hinsby, Klaus; Jensen, Karsten H.; Laier, Troels; Larsen, Flemming; Busenberg, Eurybiades; Plummer, Niel

    2004-01-01

    Transport of chlorofluorocarbon (CFC) gases through the unsaturated zone to the water table is affected by gas diffusion, air–water exchange (solubility), sorption to the soil matrix, advective–dispersive transport in the water phase, and, in some cases, anaerobic degradation. In deep unsaturated zones, this may lead to a time lag between entry of gases at the land surface and recharge to groundwater. Data from a Danish field site were used to investigate how time lag is affected by variations in water content and to explore the use of simple analytical solutions to calculate time lag. Numerical simulations demonstrate that either degradation or sorption of CFC-11 takes place, whereas CFC-12 and CFC-113 are nonreactive. Water flow did not appreciably affect transport. An analytical solution for the period with a linear increase in atmospheric CFC concentrations (approximately early 1970s to early 1990s) was used to calculate CFC profiles and time lags. We compared the analytical results with numerical simulations. The time lags in the 15-m-deep unsaturated zone increase from 4.2 to between 5.2 and 6.1 yr and from 3.4 to 3.9 yr for CFC-11 and CFC-12, respectively, when simulations change from use of an exponential to a linear increase in atmospheric concentrations. The CFC concentrations at the water table before the early 1990s can be estimated by displacing the atmospheric input function by these fixed time lags. A sensitivity study demonstrates conditions under which a time lag in the unsaturated zone becomes important. The most critical parameter is the tortuosity coefficient. The analytical approach is valid for the low range of tortuosity coefficients (τ = 0.1–0.4) and unsaturated zones greater than approximately 20 m in thickness. In these cases the CFC distribution may still be from either the exponential or linear phase. In other cases, the use of numerical models, as described in our work and elsewhere, is an option.

  4. Factors affecting harp seal (Pagophilus groenlandicus) strandings in the Northwest Atlantic.

    PubMed

    Soulen, Brianne K; Cammen, Kristina; Schultz, Thomas F; Johnston, David W

    2013-01-01

    The effects of climate change on high latitude regions are becoming increasingly evident, particularly in the rapid decline of sea ice cover in the Arctic. Many high latitude species dependent on sea ice are being forced to adapt to changing habitats. Harp seals (Pagophilus groenlandicus) are an indicator species for changing high-latitude ecosystems. This study analyzed multiple factors including ice cover, demographics, and genetic diversity, which could affect harp seal stranding rates along the eastern coast of the United States. Ice cover assessments were conducted for the month of February in the Gulf of St. Lawrence whelping region from 1991-2010 using remote sensing data, and harp seal stranding data were collected over the same time period. Genetic diversity, which may affect how quickly species can adapt to changing climates, was assessed using ten microsatellite markers to determine mean d (2) in a subset of stranded and by-caught (presumably healthy) seals sampled along the northeast U.S. coast. Our study found a strong negative correlation (R (2) = 0.49) between ice cover in the Gulf of St. Lawrence and yearling harp seal strandings, but found no relationship between sea ice conditions and adult strandings. Our analysis revealed that male seals stranded more frequently than females during the study period and that this relationship was strongest during light ice years. In contrast, we found no significant difference in mean d (2) between stranded and by-caught harp seals. The results demonstrate that sea ice cover and demographic factors have a greater influence on harp seal stranding rates than genetic diversity, with only a little of the variance in mean d (2) among stranded seals explained by ice cover. Any changes in these factors could have major implications for harp seals, and these findings should be considered in the development of future management plans for the Arctic that incorporate climate variability.

  5. Factors Affecting Harp Seal (Pagophilus groenlandicus) Strandings in the Northwest Atlantic

    PubMed Central

    Schultz, Thomas F.; Johnston, David W.

    2013-01-01

    The effects of climate change on high latitude regions are becoming increasingly evident, particularly in the rapid decline of sea ice cover in the Arctic. Many high latitude species dependent on sea ice are being forced to adapt to changing habitats. Harp seals (Pagophilus groenlandicus) are an indicator species for changing high-latitude ecosystems. This study analyzed multiple factors including ice cover, demographics, and genetic diversity, which could affect harp seal stranding rates along the eastern coast of the United States. Ice cover assessments were conducted for the month of February in the Gulf of St. Lawrence whelping region from 1991–2010 using remote sensing data, and harp seal stranding data were collected over the same time period. Genetic diversity, which may affect how quickly species can adapt to changing climates, was assessed using ten microsatellite markers to determine mean d 2 in a subset of stranded and by-caught (presumably healthy) seals sampled along the northeast U.S. coast. Our study found a strong negative correlation (R 2 = 0.49) between ice cover in the Gulf of St. Lawrence and yearling harp seal strandings, but found no relationship between sea ice conditions and adult strandings. Our analysis revealed that male seals stranded more frequently than females during the study period and that this relationship was strongest during light ice years. In contrast, we found no significant difference in mean d 2 between stranded and by-caught harp seals. The results demonstrate that sea ice cover and demographic factors have a greater influence on harp seal stranding rates than genetic diversity, with only a little of the variance in mean d 2 among stranded seals explained by ice cover. Any changes in these factors could have major implications for harp seals, and these findings should be considered in the development of future management plans for the Arctic that incorporate climate variability. PMID:23874759

  6. Database documentation of marine mammal stranding and mortality: current status review and future prospects.

    PubMed

    Chan, Derek K P; Tsui, Henry C L; Kot, Brian C W

    2017-11-21

    Databases are systematic tools to archive and manage information related to marine mammal stranding and mortality events. Stranding response networks, governmental authorities and non-governmental organizations have established regional or national stranding networks and have developed unique standard stranding response and necropsy protocols to document and track stranded marine mammal demographics, signalment and health data. The objectives of this study were to (1) describe and review the current status of marine mammal stranding and mortality databases worldwide, including the year established, types of database and their goals; and (2) summarize the geographic range included in the database, the number of cases recorded, accessibility, filter and display methods. Peer-reviewed literature was searched, focussing on published databases of live and dead marine mammal strandings and mortality and information released from stranding response organizations (i.e. online updates, journal articles and annual stranding reports). Databases that were not published in the primary literature or recognized by government agencies were excluded. Based on these criteria, 10 marine mammal stranding and mortality databases were identified, and strandings and necropsy data found in these databases were evaluated. We discuss the results, limitations and future prospects of database development. Future prospects include the development and application of virtopsy, a new necropsy investigation tool. A centralized web-accessed database of all available postmortem multimedia from stranded marine mammals may eventually support marine conservation and policy decisions, which will allow the use of marine animals as sentinels of ecosystem health, working towards a 'One Ocean-One Health' ideal.

  7. Dynamic properties of unbonded, multi-strand beams subjected to flexural loading

    NASA Astrophysics Data System (ADS)

    Asker, Haval K.; Rongong, Jem A.; Lord, Charles E.

    2018-02-01

    Beam-like structures, constructed from many long strands that are constrained rather than bonded together, can provide appreciable levels of structural damping through friction between individual strands. This paper describes experimental and numerical studies, carried out on square-section metal beams, which are aimed at improving understanding of the relationship between construction and performance. A beam is formed from a pack of square-section strands that is held together at various compression loads with pre-calibrated clamps. Flexural deformation of the assembled beam is simulated using standard finite element analysis employing simple Coulomb friction at the interfaces. The validity of the assumptions used in the models is confirmed by comparison with three point bend tests on a regular nine strand construction at several different clamp loads. Dynamic loss factors for this beam are obtained by conducting forced vibration tests, which show that the damping is insensitive to frequency. Subsequent numerical studies are used to investigate the effects of increasing the number of strands whilst maintaining the overall cross-section geometry of the beam. It is found that the system stiffness drops and loss factor increases when more strands are used for a maintained beam cross-section. Interestingly, the energy dissipated by each beam construction is almost the same. These results provide a vital and necessary insight into the physics for stranded structures and materials that are largely prevalent in mechanical (e.g. cables) and electrical (e.g. wires) elements.

  8. Toxin MqsR Cleaves Single-Stranded mRNA with Various 5 Ends

    DTIC Science & Technology

    2016-08-24

    either protein ORIGINAL RESEARCH Toxin MqsR cleaves single- stranded mRNA with various 5’ ends Nityananda Chowdhury1,*, Brian W. Kwan1,*, Louise C...in which a single 5′- GCU site was predicted to be single- stranded (ssRNA), double- stranded (dsRNA), in the loop of a stem - loop (slRNA), or in a...single- stranded 5′- GCU sites since cleavage was approximately 20- fold higher than cleavage seen with the 5′- GCU site in the stem - loop and

  9. The Roles of Family B and D DNA Polymerases in Thermococcus Species 9°N Okazaki Fragment Maturation*

    PubMed Central

    Greenough, Lucia; Kelman, Zvi; Gardner, Andrew F.

    2015-01-01

    During replication, Okazaki fragment maturation is a fundamental process that joins discontinuously synthesized DNA fragments into a contiguous lagging strand. Efficient maturation prevents repeat sequence expansions, small duplications, and generation of double-stranded DNA breaks. To address the components required for the process in Thermococcus, Okazaki fragment maturation was reconstituted in vitro using purified proteins from Thermococcus species 9°N or cell extracts. A dual color fluorescence assay was developed to monitor reaction substrates, intermediates, and products. DNA polymerase D (polD) was proposed to function as the replicative polymerase in Thermococcus replicating both the leading and the lagging strands. It is shown here, however, that it stops before the previous Okazaki fragments, failing to rapidly process them. Instead, Family B DNA polymerase (polB) was observed to rapidly fill the gaps left by polD and displaces the downstream Okazaki fragment to create a flap structure. This flap structure was cleaved by flap endonuclease 1 (Fen1) and the resultant nick was ligated by DNA ligase to form a mature lagging strand. The similarities to both bacterial and eukaryotic systems and evolutionary implications of archaeal Okazaki fragment maturation are discussed. PMID:25814667

  10. Inter-strand current sharing and ac loss measurements in superconducting YBCO Roebel cables

    DOE PAGES

    Majoros, M.; Sumption, M. D.; Collings, E. W.; ...

    2015-04-08

    A Roebel cable, one twist pitch long, was modified from its as-received state by soldering copper strips between the strands to provide inter-strand connections enabling current sharing. Various DC transport currents (representing different percentages of its critical current) were applied to a single strand of such a modified cable at 77 K in a liquid nitrogen bath. Simultaneous monitoring of I–V curves in different parts of the strand as well as in its interconnections with other strands was made using a number of sensitive Keithley nanovoltmeters in combination with a multichannel high-speed data acquisition card, all controlled via LabView software.more » Current sharing onset was observed at about 1.02 of strand I c. At a strand current of 1.3I c about 5% of the current was shared through the copper strip interconnections. A finite element method modeling was performed to estimate the inter-strand resistivities required to enable different levels of current sharing. The relative contributions of coupling and hysteretic magnetization (and loss) were compared, and for our cable and tape geometry, and at dB/dt=1 T s -1, and our inter-strand resistance of 0.77 mΩ, (enabling a current sharing of 5% at 1.3I c) the coupling component was 0.32% of the hysteretic component. However, inter-strand contact resistance values of 100–1000 times smaller (close to those of NbTi and Nb 3Sn based accelerator cables) would make the coupling components comparable in size to the hysteretic components.« less

  11. Inter-strand current sharing and ac loss measurements in superconducting YBCO Roebel cables

    DOE PAGES

    sumption, Mike; Majoros, Milan; Collings, E. W.; ...

    2014-11-07

    A Roebel cable, one twist pitch long, was modified from its as-received state by soldering copper strips between the strands to provide inter-strand connections enabling current sharing. Various DC transport currents (representing different percentages of its critical current) were applied to a single strand of such a modified cable at 77 K in a liquid nitrogen bath. Simultaneous monitoring of I–V curves in different parts of the strand as well as in its interconnections with other strands was made using a number of sensitive Keithley nanovoltmeters in combination with a multichannel high-speed data acquisition card, all controlled via LabView software.more » Current sharing onset was observed at about 1.02 of strand I c. At a strand current of 1.3I c about 5% of the current was shared through the copper strip interconnections. A finite element method modeling was performed to estimate the inter-strand resistivities required to enable different levels of current sharing. The relative contributions of coupling and hysteretic magnetization (and loss) were compared, and for our cable and tape geometry, and at dB/dt=1 T s -1, and our inter-strand resistance of 0.77 mΩ, (enabling a current sharing of 5% at 1.3I c ) the coupling component was 0.32% of the hysteretic component. However, inter-strand contact resistance values of 100–1000 times smaller (close to those of NbTi and Nb 3Sn based accelerator cables) would make the coupling components comparable in size to the hysteretic components.« less

  12. Inter-strand current sharing and ac loss measurements in superconducting YBCO Roebel cables

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

    Majoros, M.; Sumption, M. D.; Collings, E. W.

    A Roebel cable, one twist pitch long, was modified from its as-received state by soldering copper strips between the strands to provide inter-strand connections enabling current sharing. Various DC transport currents (representing different percentages of its critical current) were applied to a single strand of such a modified cable at 77 K in a liquid nitrogen bath. Simultaneous monitoring of I–V curves in different parts of the strand as well as in its interconnections with other strands was made using a number of sensitive Keithley nanovoltmeters in combination with a multichannel high-speed data acquisition card, all controlled via LabView software.more » Current sharing onset was observed at about 1.02 of strand I c. At a strand current of 1.3I c about 5% of the current was shared through the copper strip interconnections. A finite element method modeling was performed to estimate the inter-strand resistivities required to enable different levels of current sharing. The relative contributions of coupling and hysteretic magnetization (and loss) were compared, and for our cable and tape geometry, and at dB/dt=1 T s -1, and our inter-strand resistance of 0.77 mΩ, (enabling a current sharing of 5% at 1.3I c) the coupling component was 0.32% of the hysteretic component. However, inter-strand contact resistance values of 100–1000 times smaller (close to those of NbTi and Nb 3Sn based accelerator cables) would make the coupling components comparable in size to the hysteretic components.« less

  13. Inter-strand current sharing and ac loss measurements in superconducting YBCO Roebel cables

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

    sumption, Mike; Majoros, Milan; Collings, E. W.

    A Roebel cable, one twist pitch long, was modified from its as-received state by soldering copper strips between the strands to provide inter-strand connections enabling current sharing. Various DC transport currents (representing different percentages of its critical current) were applied to a single strand of such a modified cable at 77 K in a liquid nitrogen bath. Simultaneous monitoring of I–V curves in different parts of the strand as well as in its interconnections with other strands was made using a number of sensitive Keithley nanovoltmeters in combination with a multichannel high-speed data acquisition card, all controlled via LabView software.more » Current sharing onset was observed at about 1.02 of strand I c. At a strand current of 1.3I c about 5% of the current was shared through the copper strip interconnections. A finite element method modeling was performed to estimate the inter-strand resistivities required to enable different levels of current sharing. The relative contributions of coupling and hysteretic magnetization (and loss) were compared, and for our cable and tape geometry, and at dB/dt=1 T s -1, and our inter-strand resistance of 0.77 mΩ, (enabling a current sharing of 5% at 1.3I c ) the coupling component was 0.32% of the hysteretic component. However, inter-strand contact resistance values of 100–1000 times smaller (close to those of NbTi and Nb 3Sn based accelerator cables) would make the coupling components comparable in size to the hysteretic components.« less

  14. I. The design, synthesis, and structure of antiparallel beta-sheet and beta-strand mimics. II. The design of a scripted chemistry outreach program to high schools

    NASA Astrophysics Data System (ADS)

    Waldman, Amy Sue

    I. Protein structure is not easily predicted from the linear sequence of amino acids. An increased ability to create protein structures would allow researchers to develop new peptide-based therapeutics and materials, and would provide insights into the mechanisms of protein folding. Toward this end, we have designed and synthesized two-stranded antiparallel beta-sheet mimics containing conformationally biased scaffolds and semicarbazide, urea, and hydrazide linker groups that attach peptide chains to the scaffold. The mimics exhibited populations of intramolecularly hydrogen-bonded beta-sheet-like conformers as determined by spectroscopic techniques such as FTIR, sp1H NMR, and ROESY studies. During our studies, we determined that a urea-hydrazide beta-strand mimic was able to tightly hydrogen bond to peptides in an antiparallel beta-sheet-like configuration. Several derivatives of the urea-hydrazide beta-strand mimic were synthesized. Preliminary data by electron microscopy indicate that the beta-strand mimics have an effect on the folding of Alzheimer's Abeta peptide. These data suggest that the urea-hydrazide beta-strand mimics and related compounds may be developed into therapeutics which effect the folding of the Abeta peptide into neurotoxic aggregates. II. In recent years, there has been concern about the low level of science literacy and science interest among Americans. A declining interest in science impacts the abilities of people to make informed decisions about technology. To increase the interest in science among secondary students, we have developed the UCI Chemistry Outreach Program to High Schools. The Program features demonstration shows and discussions about chemistry in everyday life. The development and use of show scripts has enabled large numbers of graduate and undergraduate student volunteers to demonstrate chemistry to more than 12,000 local high school students. Teachers, students, and volunteers have expressed their enjoyment of The UCI

  15. Lagged correlation networks

    NASA Astrophysics Data System (ADS)

    Curme, Chester

    Technological advances have provided scientists with large high-dimensional datasets that describe the behaviors of complex systems: from the statistics of energy levels in complex quantum systems, to the time-dependent transcription of genes, to price fluctuations among assets in a financial market. In this environment, where it may be difficult to infer the joint distribution of the data, network science has flourished as a way to gain insight into the structure and organization of such systems by focusing on pairwise interactions. This work focuses on a particular setting, in which a system is described by multivariate time series data. We consider time-lagged correlations among elements in this system, in such a way that the measured interactions among elements are asymmetric. Finally, we allow these interactions to be characteristically weak, so that statistical uncertainties may be important to consider when inferring the structure of the system. We introduce a methodology for constructing statistically validated networks to describe such a system, extend the methodology to accommodate interactions with a periodic component, and show how consideration of bipartite community structures in these networks can aid in the construction of robust statistical models. An example of such a system is a financial market, in which high frequency returns data may be used to describe contagion, or the spreading of shocks in price among assets. These data provide the experimental testing ground for our methodology. We study NYSE data from both the present day and one decade ago, examine the time scales over which the validated lagged correlation networks exist, and relate differences in the topological properties of the networks to an increasing economic efficiency. We uncover daily periodicities in the validated interactions, and relate our findings to explanations of the Epps Effect, an empirical phenomenon of financial time series. We also study bipartite community

  16. [Complexity and its integrative effects of the time lags of environment factors affecting Larix gmelinii stem sap flow].

    PubMed

    Wang, Hui-Mei; Sun, Wei; Zu, Yuan-Gang; Wang, Wen-Jie

    2011-12-01

    Based on the one-year (2005) observations with a frequency of half hour on the stem sap flow of Larix gmelinii plantation trees planted in 1969 and the related environmental factors air humidity (RH), air temperature (T(air)), photosynthetic components active radiation (PAR), soil temperature (T(soil)), and soil moisture (TDR), principal analysis (PCA) and correction analysis were made on the time lag effect of the stem flow in different seasons (26 days of each season) and in a year via dislocation analysis, with the complexity and its integrative effects of the time lags of environment factors affecting the stem sap flow approached. The results showed that in different seasons and for different environmental factors, the time lag effect varied obviously. In general, the time lag of PAR was 0.5-1 hour ahead of sap flow, that of T(air) and RH was 0-2 hours ahead of or behind the sap flow, and the time lags of T(soil) and TDR were much longer or sometimes undetectable. Because of the complexity of the time lags, no evident improvements were observed in the linear correlations (R2, slope, and intercept) when the time lags based on short-term (20 days) data were used to correct the time lags based on whole year data. However, obvious improvements were found in the standardized and non-standardized correlation coefficients in stepwise multiple regressions, i.e., the time lag corrections could improve the effects of RH, but decreased the effects of PAR, T(air), and T(soil). PCA could be used to simplify the complexity. The first and the second principal components could stand for over 75% information of all the environmental factors in different seasons and in whole year. The time lags of both the first and the second principal components were 1-1.5 hours in advance of the sap flow, except in winter (no time lag effect).

  17. 7 CFR 1755.370 - RUS specification for seven wire galvanized steel strand.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 11 2012-01-01 2012-01-01 false RUS specification for seven wire galvanized steel... steel strand. (a) RUS incorporates by reference ASTM A475-78, Standard Specification for Zinc-Coated Steel Wire Strand, issued May 1978. All seven wire galvanized steel strand purchased after April 1, 1990...

  18. 7 CFR 1755.370 - RUS specification for seven wire galvanized steel strand.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 11 2014-01-01 2014-01-01 false RUS specification for seven wire galvanized steel... steel strand. (a) RUS incorporates by reference ASTM A475-78, Standard Specification for Zinc-Coated Steel Wire Strand, issued May 1978. All seven wire galvanized steel strand purchased after April 1, 1990...

  19. 7 CFR 1755.370 - RUS specification for seven wire galvanized steel strand.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 11 2013-01-01 2013-01-01 false RUS specification for seven wire galvanized steel... steel strand. (a) RUS incorporates by reference ASTM A475-78, Standard Specification for Zinc-Coated Steel Wire Strand, issued May 1978. All seven wire galvanized steel strand purchased after April 1, 1990...

  20. Effect of phase lag on cyclic durability of laminated composite

    NASA Astrophysics Data System (ADS)

    Andersons, Janis; Limonov, V.; Tamuzs, Vitants

    1992-07-01

    Theoretical and experimental results on fatigue of laminated fiber reinforced composites under out-of-phase, biaxial cyclic loading are presented. Experiments were carried out on tubular filament wound samples of epoxy matrix/organic (Kevlar type) fiber composites. Fatigue strength under two different loading modes, namely cyclic torsion combined with axial tension or compression, was investigated for phase lags psi = 0, pi/2, and pi. Durability was shown to decrease with increasing phase shift both for axial tension (R = 0.1) and compression (R = 10). A matrix failure criterion was proposed for a unidirectionally reinforced ply, and the ply discount method was modified to account for phase lag. Calculated S-N curves agree reasonably well with experimental data.

  1. Time lags between crown and basal sap flows in tropical lianas and co-occurring trees.

    PubMed

    Chen, Ya-Jun; Bongers, Frans; Tomlinson, Kyle; Fan, Ze-Xin; Lin, Hua; Zhang, Shu-Bin; Zheng, Yu-Long; Li, Yang-Ping; Cao, Kun-Fang; Zhang, Jiao-Lin

    2016-06-01

    Water storage in the stems of woody plants contributes to their responses to short-term water shortages. To estimate the contribution of water storage to the daily water budget of trees, time lags of sap flow between different positions of trunk are used as a proxy of stem water storage. In lianas, another large group of woody species, it has rarely been studied whether stored water functions in their daily water use, despite their increasing roles in the carbon and water dynamics of tropical forests caused by their increasing abundance. We hypothesized that lianas would exhibit large time lags due to their extremely long stems, wide vessels and large volume of parenchyma in the stem. We examined time lags in sap flow, diel changes of stem volumetric water content (VWC) and biophysical properties of sapwood of 19 lianas and 26 co-occurring trees from 27 species in 4 forests (karst, tropical seasonal, flood plain and savanna) during a wet season. The plants varied in height/length from <5 to >60 m. The results showed that lianas had significantly higher saturated water content (SWC) and much lower wood density than trees. Seven of 19 liana individuals had no time lags; in contrast, only 3 of 26 tree individuals had no time lags. In general, lianas had shorter time lags than trees in our data set, but this difference was not significant for our most conservative analyses. Across trees and lianas, time lag duration increased with diurnal maximum changeable VWC but was independent of the body size, path length, wood density and SWC. The results suggest that in most lianas, internal stem water storage contributes little to daily water budget, while trees may rely more on stored water in the stem. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  2. Data centers as dispatchable loads to harness stranded power

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

    Kim, Kibaek; Yang, Fan; Zavala, Victor M.

    Here, we analyze how traditional data center placement and optimal placement of dispatchable data centers affect power grid efficiency. We use detailed network models, stochastic optimization formulations, and diverse renewable generation scenarios to perform our analysis. Our results reveal that significant spillage and stranded power will persist in power grids as wind power levels are increased. A counter-intuitive finding is that collocating data centers with inflexible loads next to wind farms has limited impacts on renewable portfolio standard (RPS) goals because it provides limited system-level flexibility. Such an approach can, in fact, increase stranded power and fossil-fueled generation. In contrast,more » optimally placing data centers that are dispatchable provides system-wide flexibility, reduces stranded power, and improves efficiency. In short, optimally placed dispatchable computing loads can enable better scaling to high RPS. In our case study, we find that these dispatchable computing loads are powered to 60-80% of their requested capacity, indicating that there are significant economic incentives provided by stranded power.« less

  3. Data centers as dispatchable loads to harness stranded power

    DOE PAGES

    Kim, Kibaek; Yang, Fan; Zavala, Victor M.; ...

    2016-07-20

    Here, we analyze how traditional data center placement and optimal placement of dispatchable data centers affect power grid efficiency. We use detailed network models, stochastic optimization formulations, and diverse renewable generation scenarios to perform our analysis. Our results reveal that significant spillage and stranded power will persist in power grids as wind power levels are increased. A counter-intuitive finding is that collocating data centers with inflexible loads next to wind farms has limited impacts on renewable portfolio standard (RPS) goals because it provides limited system-level flexibility. Such an approach can, in fact, increase stranded power and fossil-fueled generation. In contrast,more » optimally placing data centers that are dispatchable provides system-wide flexibility, reduces stranded power, and improves efficiency. In short, optimally placed dispatchable computing loads can enable better scaling to high RPS. In our case study, we find that these dispatchable computing loads are powered to 60-80% of their requested capacity, indicating that there are significant economic incentives provided by stranded power.« less

  4. X-linked Acrogigantism (X-LAG) Syndrome: Clinical Profile and Therapeutic Responses

    PubMed Central

    Beckers, Albert; Lodish, Maya Beth; Trivellin, Giampaolo; Rostomyan, Liliya; Lee, Misu; Faucz, Fabio R; Yuan, Bo; Choong, Catherine S; Caberg, Jean-Hubert; Verrua, Elisa; Naves, Luciana Ansaneli; Cheetham, Tim D; Young, Jacques; Lysy, Philippe A; Petrossians, Patrick; Cotterill, Andrew; Shah, Nalini Samir; Metzger, Daniel; Castermans, Emilie; Ambrosio, Maria Rosaria; Villa, Chiara; Strebkova, Natalia; Mazerkina, Nadia; Gaillard, Stéphan; Barra, Gustavo Barcelos; Casulari, Luis Augusto; Neggers, Sebastian J.; Salvatori, Roberto; Jaffrain-Rea, Marie-Lise; Zacharin, Margaret; Santamaria, Beatriz Lecumberri; Zacharieva, Sabina; Lim, Ee Mun; Mantovani, Giovanna; Zatelli, Maria Chaira; Collins, Michael T; Bonneville, Jean-François; Quezado, Martha; Chittiboina, Prashant; Oldfield, Edward H.; Bours, Vincent; Liu, Pengfei; De Herder, Wouter; Pellegata, Natalia; Lupski, James R.; Daly, Adrian F.; Stratakis, Constantine A.

    2015-01-01

    X-linked acro-gigantism (X-LAG) is a new syndrome of pituitary gigantism, caused by microduplications on chromosome Xq26.3, encompassing the gene GPR101, which is highly upregulated in pituitary tumors. We conducted this study to explore the clinical, radiological and hormonal phenotype and responses to therapy in patients with X-LAG syndrome. The study included 18 patients (13 sporadic) with X-LAG and a microduplication in chromosome Xq26.3. All sporadic cases had unique duplications and the inheritance pattern in 2 families was dominant with all Xq26.3 duplication carriers being affected. Patients began to grow rapidly as early as 2–3 months of age (median 12 months). At diagnosis (median delay 27 months), patients had a median height and weight SDS score of >+3.9 SDS. Apart from the increased overall body size, the children had acromegalic symptoms including acral enlargement and facial coarsening. More than a third of cases had increased appetite. Patients had marked hypersecretion of GH/IGF-1 and prolactin, usually due to a pituitary macroadenoma or hyperplasia. Primary neurosurgical control was achieved with extensive anterior pituitary resection but postoperative hypopituitarism was frequent. Control with somatostatin analogs was not readily achieved despite moderate to high somatostatin receptor subtype-2 expression in tumor tissue. Postoperative adjuvant pegvisomant achieved control of IGF-1 all 5 cases in which it was employed. X-LAG is a new infant-onset gigantism syndrome that has a severe clinical phenotype leading to challenging disease management. PMID:25712922

  5. Dependence of the Contact Resistance on the Design of Stranded Conductors

    PubMed Central

    Zeroukhi, Youcef; Napieralska-Juszczak, Ewa; Vega, Guillaume; Komeza, Krzysztof; Morganti, Fabrice; Wiak, Slawomir

    2014-01-01

    During the manufacturing process multi-strand conductors are subject to compressive force and rotation moments. The current distribution in the multi-strand conductors is not uniform and is controlled by the transverse resistivity. This is mainly determined by the contact resistance at the strand crossovers and inter-strand contact resistance. The surface layer properties, and in particular the crystalline structure and degree of oxidation, are key parameters in determining the transverse resistivity. The experimental set-ups made it possible to find the dependence of contact resistivity as a function of continuous working stresses and cable design. A study based on measurements and numerical simulation is made to identify the contact resistivity functions. PMID:25196112

  6. Co-opting the Fanconi Anemia Genomic Stability Pathway Enables Herpesvirus DNA Synthesis and Productive Growth

    PubMed Central

    Karttunen, Heidi; Savas, Jeffrey N.; McKinney, Caleb; Chen, Yu-Hung; Yates, John R.; Hukkanen, Veijo; Huang, Tony T.; Mohr, Ian

    2015-01-01

    SUMMARY DNA damage associated with viral DNA synthesis can result in double strand breaks that threaten genome integrity and must be repaired. Here, we establish that the cellular Fanconi Anemia (FA) genomic stability pathway is exploited by HSV1 to promote viral DNA synthesis and enable its productive growth. Potent FA pathway activation in HSV1-infected cells resulted in monoubiquitination of FA effector proteins, FANCI and FANCD2 (FANCI-D2) and required the viral DNA polymerase. FANCD2 relocalized to viral replication compartments and FANCI-D2 interacted with a multi-subunit complex containing the virus-encoded single-stranded DNA-binding protein ICP8. Significantly, while HSV1 productive growth was impaired in monoubiquitination-defective FA patient cells, this restriction was partially surmounted by antagonizing the DNA-dependent protein kinase (DNA-PK), a critical enzyme required for non-homologous end-joining (NHEJ). This identifies the FA-pathway as a new cellular factor required for herpesvirus productive growth and suggests that FA-mediated suppression of NHEJ is a fundamental step in the viral lifecycle. PMID:24954902

  7. On the biophysics and kinetics of toehold-mediated DNA strand displacement

    PubMed Central

    Srinivas, Niranjan; Ouldridge, Thomas E.; Šulc, Petr; Schaeffer, Joseph M.; Yurke, Bernard; Louis, Ard A.; Doye, Jonathan P. K.; Winfree, Erik

    2013-01-01

    Dynamic DNA nanotechnology often uses toehold-mediated strand displacement for controlling reaction kinetics. Although the dependence of strand displacement kinetics on toehold length has been experimentally characterized and phenomenologically modeled, detailed biophysical understanding has remained elusive. Here, we study strand displacement at multiple levels of detail, using an intuitive model of a random walk on a 1D energy landscape, a secondary structure kinetics model with single base-pair steps and a coarse-grained molecular model that incorporates 3D geometric and steric effects. Further, we experimentally investigate the thermodynamics of three-way branch migration. Two factors explain the dependence of strand displacement kinetics on toehold length: (i) the physical process by which a single step of branch migration occurs is significantly slower than the fraying of a single base pair and (ii) initiating branch migration incurs a thermodynamic penalty, not captured by state-of-the-art nearest neighbor models of DNA, due to the additional overhang it engenders at the junction. Our findings are consistent with previously measured or inferred rates for hybridization, fraying and branch migration, and they provide a biophysical explanation of strand displacement kinetics. Our work paves the way for accurate modeling of strand displacement cascades, which would facilitate the simulation and construction of more complex molecular systems. PMID:24019238

  8. On the biophysics and kinetics of toehold-mediated DNA strand displacement.

    PubMed

    Srinivas, Niranjan; Ouldridge, Thomas E; Sulc, Petr; Schaeffer, Joseph M; Yurke, Bernard; Louis, Ard A; Doye, Jonathan P K; Winfree, Erik

    2013-12-01

    Dynamic DNA nanotechnology often uses toehold-mediated strand displacement for controlling reaction kinetics. Although the dependence of strand displacement kinetics on toehold length has been experimentally characterized and phenomenologically modeled, detailed biophysical understanding has remained elusive. Here, we study strand displacement at multiple levels of detail, using an intuitive model of a random walk on a 1D energy landscape, a secondary structure kinetics model with single base-pair steps and a coarse-grained molecular model that incorporates 3D geometric and steric effects. Further, we experimentally investigate the thermodynamics of three-way branch migration. Two factors explain the dependence of strand displacement kinetics on toehold length: (i) the physical process by which a single step of branch migration occurs is significantly slower than the fraying of a single base pair and (ii) initiating branch migration incurs a thermodynamic penalty, not captured by state-of-the-art nearest neighbor models of DNA, due to the additional overhang it engenders at the junction. Our findings are consistent with previously measured or inferred rates for hybridization, fraying and branch migration, and they provide a biophysical explanation of strand displacement kinetics. Our work paves the way for accurate modeling of strand displacement cascades, which would facilitate the simulation and construction of more complex molecular systems.

  9. X-ray short-time lags in the Fe-K energy band produced by scattering clouds in active galactic nuclei

    NASA Astrophysics Data System (ADS)

    Mizumoto, Misaki; Done, Chris; Hagino, Kouichi; Ebisawa, Ken; Tsujimoto, Masahiro; Odaka, Hirokazu

    2018-07-01

    X-rays illuminating the accretion disc in active galactic nuclei give rise to an iron K line and its associated reflection spectrum that are lagged behind the continuum variability by the light-travel time from the source to the disc. The measured lag time-scales in the iron band can be as short as ˜Rg/c, where Rg is the gravitational radius, which is often interpreted as evidence for a very small continuum source close to the event horizon of a rapidly spinning black hole. However, the short lags can also be produced by reflection from more distant material, because the primary photons with no time-delay dilute the time-lags caused by the reprocessed photons. We perform a Monte Carlo simulation to calculate the dilution effect in the X-ray reverberation lags from a half-shell of neutral material placed at 100 Rg from the central source. This gives lags of ˜2 Rg/c, but the iron line is a distinctly narrow feature in the lag-energy plot, whereas the data often show a broader line. We show that both the short lag and the line broadening can be reproduced, if the scattering material is outflowing at ˜0.1c. The velocity structure in the wind can also give shifts in the line profile in the lag-energy plot calculated at different frequencies. Hence we propose that the observed broad iron reverberation lags and shifts in profile as a function of frequency of variability can arise from a disc wind at fairly large distances from the X-ray source.

  10. X-ray short-time lags in the Fe-K energy band produced by scattering clouds in active galactic nuclei

    NASA Astrophysics Data System (ADS)

    Mizumoto, Misaki; Done, Chris; Hagino, Kouichi; Ebisawa, Ken; Tsujimoto, Masahiro; Odaka, Hirokazu

    2018-05-01

    X-rays illuminating the accretion disc in active galactic nuclei give rise to an iron K line and its associated reflection spectrum which are lagged behind the continuum variability by the light-travel time from the source to the disc. The measured lag timescales in the iron band can be as short as ˜Rg/c, where Rg is the gravitational radius, which is often interpreted as evidence for a very small continuum source close to the event horizon of a rapidly spinning black hole. However, the short lags can also be produced by reflection from more distant material, because the primary photons with no time-delay dilute the time-lags caused by the reprocessed photons. We perform a Monte-Carlo simulation to calculate the dilution effect in the X-ray reverberation lags from a half-shell of neutral material placed at 100 Rg from the central source. This gives lags of ˜2 Rg/c, but the iron line is a distinctly narrow feature in the lag-energy plot, whereas the data often show a broader line. We show that both the short lag and the line broadening can be reproduced if the scattering material is outflowing at ˜0.1c. The velocity structure in the wind can also give shifts in the line profile in the lag-energy plot calculated at different frequencies. Hence we propose that the observed broad iron reverberation lags and shifts in profile as a function of frequency of variability can arise from a disc wind at fairly large distances from the X-ray source.

  11. Nonlinear flap-lag axial equations of a rotating beam

    NASA Technical Reports Server (NTRS)

    Kaza, K. R. V.; Kvaternik, R. G.

    1977-01-01

    It is possible to identify essentially four approaches by which analysts have established either the linear or nonlinear governing equations of motion for a particular problem related to the dynamics of rotating elastic bodies. The approaches include the effective applied load artifice in combination with a variational principle and the use of Newton's second law, written as D'Alembert's principle, applied to the deformed configuration. A third approach is a variational method in which nonlinear strain-displacement relations and a first-degree displacement field are used. The method introduced by Vigneron (1975) for deriving the linear flap-lag equations of a rotating beam constitutes the fourth approach. The reported investigation shows that all four approaches make use of the geometric nonlinear theory of elasticity. An alternative method for deriving the nonlinear coupled flap-lag-axial equations of motion is also discussed.

  12. Tyrosinase autoactivation and the problem of the lag period.

    PubMed

    Naish-Byfield, S; Riley, P A

    1998-06-01

    Evidence is presented for the binding of the quinone oxidation product of the monohydric phenol substrate, 4-hydroxyanisole, to mushroom tyrosinase. Column chromatography and SDS-PAGE separation showed labelling of the enzyme when incubated with 14C ring-labelled 4-hydroxyanisole. It is proposed that covalent binding to the enzyme and other proteins is through reaction of accessible nucleophilic groups, including thiols and amino groups, with the anisylquinone. This reductive addition enables the indirect generation of the catecholic substrate, which acts as an electron donor for the bicupric active site of met-tyrosinase and explains the lag kinetics of tyrosinase oxidation of non-cyclizing substrates. The effects of diluting the enzyme or the addition of amino acids on the lag period was consistent with a mechanism involving indirect generation of the dihydric phenol, which acts as the met-enzyme-recruiting substrate.

  13. The prescription pickup lag, an automatic prescription refill program, and community pharmacy operations.

    PubMed

    Lester, Corey A; Chui, Michelle A

    2016-01-01

    To determine the effect of an automatic prescription refill program on the prescription pickup lag in community pharmacy. A post-only quasi-experimental design comparing automatic and manual refill prescription cohorts for each of the 3 Centers for Medicare and Medicaid medication adherence metrics. A 29-store community pharmacy chain in the Midwest. Community-dwelling patients over the age of 65 years receiving prescription medications included in the statin, renin-angiotensin-aldosterone system antagonist, or non-insulin diabetes adherence metrics. An automatic prescription refill program that initiated prescription refills on a standardized, recurrent basis, eliminating the need for patients to phone in or drop off prescription refills. The prescription pickup lag, defined as the number of days between a prescription being adjudicated in the pharmacy and the prescription being picked up by the patient. A total of 37,207 prescription fills were examined. There were 20.5%, 22.4%, and 23.3% of patients enrolled in the automatic prescription refill program for the statin, renin-angiotensin-aldosterone system antagonist, and diabetes adherence metrics, respectively. Prescriptions in the automatic prescription refill cohorts experienced a median pickup lag of 7 days compared with 1 day for the manual refill prescriptions. 35.2% of all manual refill prescriptions had a pickup lag of 0 days compared with 13% for automatic refills. However, 15.4% of automatic prescription refills had a pickup lag of greater than 14 days, compared with 4.8% of manual refills. Prescriptions in the automatic prescription refill programs were associated with a significantly longer amount of time in the pharmacy before being picked up by the patient. This increased pickup lag may contribute positively by smoothing out workload demands of pharmacy staff, but may contribute negatively owing to an increased amount of rework and greater inventory requirements. Copyright © 2016 American

  14. ARE THE kHz QPO LAGS IN NEUTRON STAR 4U 1608–52 DUE TO REVERBERATION?

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

    Cackett, Edward M., E-mail: ecackett@wayne.edu

    2016-08-01

    X-ray reverberation lags have recently been discovered in both active galactic nuclei (AGNs) and black hole X-ray binaries. A recent study of the neutron star low-mass X-ray binary (LMXB) 4U 1608 52 has also shown significant lags, whose properties hint at a reverberation origin. Here, we adapt general relativistic ray tracing impulse response functions used to model X-ray reverberation in AGNs for neutron star LMXBs. Assuming that relativistic reflection forms the broad iron line and associated reflection continuum, we use reflection fits to the energy spectrum along with the impulse response functions to calculate the expected lags as a functionmore » of energy over the range of observed kHz quasi-periodic oscillation (QPO) frequencies in 4U 1608 52. We find that the lag energy spectrum is expected to increase with increasing energy above 8 keV, while the observed lags in 4U 1608 52 show the opposite behavior. This demonstrates that the lags in the lower kHz QPO of 4U 1608 52 are not solely due to reverberation. We do note, however, that the models appear to be more consistent with the much flatter lag energy spectrum observed in the upper kHz QPO of several neutron star LMXBs, suggesting that lower and upper kHz QPOs may have different origins.« less

  15. A Lagging Model for Describing Drawdown Induced by a Constant-Rate Pumping in a Leaky Confined Aquifer

    NASA Astrophysics Data System (ADS)

    Lin, Ye-Chen; Yeh, Hund-Der

    2017-10-01

    This study proposes a generalized Darcy's law with considering phase lags in both the water flux and drawdown gradient to develop a lagging flow model for describing drawdown induced by constant-rate pumping (CRP) in a leaky confined aquifer. The present model has a mathematical formulation similar to the dual-porosity model. The Laplace-domain solution of the model with the effect of wellbore storage is derived by the Laplace transform method. The time-domain solution for the case of neglecting the wellbore storage and well radius is developed by the use of Laplace transform and Weber transform. The results of sensitivity analysis based on the solution indicate that the drawdown is very sensitive to the change in each of the transmissivity and storativity. Also, a study for the lagging effect on the drawdown indicates that its influence is significant associated with the lag times. The present solution is also employed to analyze a data set taken from a CRP test conducted in a fractured aquifer in South Dakota, USA. The results show the prediction of this new solution with considering the phase lags has very good fit to the field data, especially at early pumping time. In addition, the phase lags seem to have a scale effect as indicated in the results. In other words, the lagging behavior is positively correlated with the observed distance in the Madison aquifer.

  16. The role of cytosine methylation on charge transport through a DNA strand

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

    Qi, Jianqing, E-mail: jqqi@uw.edu; Anantram, M. P., E-mail: anantmp@uw.edu; Govind, Niranjan, E-mail: niri.govind@pnnl.gov

    Cytosine methylation has been found to play a crucial role in various biological processes, including a number of human diseases. The detection of this small modification remains challenging. In this work, we computationally explore the possibility of detecting methylated DNA strands through direct electrical conductance measurements. Using density functional theory and the Landauer-Büttiker method, we study the electronic properties and charge transport through an eight base-pair methylated DNA strand and its native counterpart. We first analyze the effect of cytosine methylation on the tight-binding parameters of two DNA strands and then model the transmission of the electrons and conductance throughmore » the strands both with and without decoherence. We find that the main difference of the tight-binding parameters between the native DNA and the methylated DNA lies in the on-site energies of (methylated) cytosine bases. The intra- and inter-strand hopping integrals between two nearest neighboring guanine base and (methylated) cytosine base also change with the addition of the methyl groups. Our calculations show that in the phase-coherent limit, the transmission of the methylated strand is close to the native strand when the energy is nearby the highest occupied molecular orbital level and larger than the native strand by 5 times in the bandgap. The trend in transmission also holds in the presence of the decoherence with the same rate. The lower conductance for the methylated strand in the experiment is suggested to be caused by the more stable structure due to the introduction of the methyl groups. We also study the role of the exchange-correlation functional and the effect of contact coupling by choosing coupling strengths ranging from weak to strong coupling limit.« less

  17. The role of cytosine methylation on charge transport through a DNA strand

    NASA Astrophysics Data System (ADS)

    Qi, Jianqing; Govind, Niranjan; Anantram, M. P.

    2015-09-01

    Cytosine methylation has been found to play a crucial role in various biological processes, including a number of human diseases. The detection of this small modification remains challenging. In this work, we computationally explore the possibility of detecting methylated DNA strands through direct electrical conductance measurements. Using density functional theory and the Landauer-Büttiker method, we study the electronic properties and charge transport through an eight base-pair methylated DNA strand and its native counterpart. We first analyze the effect of cytosine methylation on the tight-binding parameters of two DNA strands and then model the transmission of the electrons and conductance through the strands both with and without decoherence. We find that the main difference of the tight-binding parameters between the native DNA and the methylated DNA lies in the on-site energies of (methylated) cytosine bases. The intra- and inter-strand hopping integrals between two nearest neighboring guanine base and (methylated) cytosine base also change with the addition of the methyl groups. Our calculations show that in the phase-coherent limit, the transmission of the methylated strand is close to the native strand when the energy is nearby the highest occupied molecular orbital level and larger than the native strand by 5 times in the bandgap. The trend in transmission also holds in the presence of the decoherence with the same rate. The lower conductance for the methylated strand in the experiment is suggested to be caused by the more stable structure due to the introduction of the methyl groups. We also study the role of the exchange-correlation functional and the effect of contact coupling by choosing coupling strengths ranging from weak to strong coupling limit.

  18. Host Pah1p phosphatidate phosphatase limits viral replication by regulating phospholipid synthesis

    PubMed Central

    Zhang, Zhenlu; He, Guijuan; Catanzaro, Nicholas; Wu, Zujian; Xie, Lianhui

    2018-01-01

    Replication of positive-strand RNA viruses [(+)RNA viruses] takes place in membrane-bound viral replication complexes (VRCs). Formation of VRCs requires virus-mediated manipulation of cellular lipid synthesis. Here, we report significantly enhanced brome mosaic virus (BMV) replication and much improved cell growth in yeast cells lacking PAH1 (pah1Δ), the sole yeast ortholog of human LIPIN genes. PAH1 encodes Pah1p (phosphatidic acid phosphohydrolase), which converts phosphatidate (PA) to diacylglycerol that is subsequently used for the synthesis of the storage lipid triacylglycerol. Inactivation of Pah1p leads to altered lipid composition, including high levels of PA, total phospholipids, ergosterol ester, and free fatty acids, as well as expansion of the nuclear membrane. In pah1Δ cells, BMV replication protein 1a and double-stranded RNA localized to the extended nuclear membrane, there was a significant increase in the number of VRCs formed, and BMV genomic replication increased by 2-fold compared to wild-type cells. In another yeast mutant that lacks both PAH1 and DGK1 (encodes diacylglycerol kinase converting diacylglycerol to PA), which has a normal nuclear membrane but maintains similar lipid compositional changes as in pah1Δ cells, BMV replicated as efficiently as in pah1Δ cells, suggesting that the altered lipid composition was responsible for the enhanced BMV replication. We further showed that increased levels of total phospholipids play an important role because the enhanced BMV replication required active synthesis of phosphatidylcholine, the major membrane phospholipid. Moreover, overexpression of a phosphatidylcholine synthesis gene (CHO2) promoted BMV replication. Conversely, overexpression of PAH1 or plant PAH1 orthologs inhibited BMV replication in yeast or Nicotiana benthamiana plants. Competing with its host for limited resources, BMV inhibited host growth, which was markedly alleviated in pah1Δ cells. Our work suggests that Pah1p promotes

  19. Click nucleic acid ligation: applications in biology and nanotechnology.

    PubMed

    El-Sagheer, Afaf H; Brown, Tom

    2012-08-21

    Biochemical strategies that use a combination of synthetic oligonucleotides, thermostable DNA polymerases, and DNA ligases can produce large DNA constructs up to 1 megabase in length. Although these ambitious targets are feasible biochemically, comparable technologies for the chemical synthesis of long DNA strands lag far behind. The best available chemical approach is the solid-phase phosphoramidite method, which can be used to assemble DNA strands up to 150 bases in length. Beyond this point, deficiencies in the chemistry make it impossible to produce pure DNA. A possible alternative approach to the chemical synthesis of large DNA strands is to join together carefully purified synthetic oligonucleotides by chemical methods. Click ligation by the copper-catalyzed azide-alkyne (CuAAC) reaction could facilitate this process. In this Account, we describe the synthesis, characterization, and applications of oligonucleotides prepared by click ligation. The alkyne and azide oligonucleotide strands can be prepared by standard protocols, and the ligation reaction is compatible with a wide range of chemical modifications to DNA and RNA. We have employed click ligation to synthesize DNA constructs up to 300 bases in length and much longer sequences are feasible. When the resulting triazole linkage is placed in a PCR template, various DNA polymerases correctly copy the entire base sequence. We have also successfully demonstrated both in vitro transcription and rolling circle amplification through the modified linkage. This linkage has shown in vivo biocompatibility: an antibiotic resistance gene containing triazole linkages functions in E. coli . Using click ligation, we have synthesized hairpin ribozymes up to 100 nucleotides in length and a hammerhead ribozyme with the triazole linkage located at the substrate cleavage site. At the opposite end of the length scale, click-ligated, cyclic mini-DNA duplexes have been used as models to study base pairing. Cyclic duplexes have

  20. Higher 5-hydroxymethylcytosine identifies immortal DNA strand chromosomes in asymmetrically self-renewing distributed stem cells.

    PubMed

    Huh, Yang Hoon; Cohen, Justin; Sherley, James L

    2013-10-15

    Immortal strands are the targeted chromosomal DNA strands of nonrandom sister chromatid segregation, a mitotic chromosome segregation pattern unique to asymmetrically self-renewing distributed stem cells (DSCs). By nonrandom segregation, immortal DNA strands become the oldest DNA strands in asymmetrically self-renewing DSCs. Nonrandom segregation of immortal DNA strands may limit DSC mutagenesis, preserve DSC fate, and contribute to DSC aging. The mechanisms responsible for specification and maintenance of immortal DNA strands are unknown. To discover clues to these mechanisms, we investigated the 5-methylcytosine and 5-hydroxymethylcytosine (5hmC) content on chromosomes in mouse hair follicle DSCs during nonrandom segregation. Although 5-methylcytosine content did not differ significantly, the relative content of 5hmC was significantly higher in chromosomes containing immortal DNA strands than in opposed mitotic chromosomes containing younger mortal DNA strands. The difference in relative 5hmC content was caused by the loss of 5hmC from mortal chromosomes. These findings implicate higher 5hmC as a specific molecular determinant of immortal DNA strand chromosomes. Because 5hmC is an intermediate during DNA demethylation, we propose a ten-eleven translocase enzyme mechanism for both the specification and maintenance of nonrandomly segregated immortal DNA strands. The proposed mechanism reveals a means by which DSCs "know" the generational age of immortal DNA strands. The mechanism is supported by molecular expression data and accounts for the selection of newly replicated DNA strands when nonrandom segregation is initiated. These mechanistic insights also provide a possible basis for another characteristic property of immortal DNA strands, their guanine ribonucleotide dependency.

  1. G-Strands on symmetric spaces

    PubMed Central

    2017-01-01

    We study the G-strand equations that are extensions of the classical chiral model of particle physics in the particular setting of broken symmetries described by symmetric spaces. These equations are simple field theory models whose configuration space is a Lie group, or in this case a symmetric space. In this class of systems, we derive several models that are completely integrable on finite dimensional Lie group G, and we treat in more detail examples with symmetric space SU(2)/S1 and SO(4)/SO(3). The latter model simplifies to an apparently new integrable nine-dimensional system. We also study the G-strands on the infinite dimensional group of diffeomorphisms, which gives, together with the Sobolev norm, systems of 1+2 Camassa–Holm equations. The solutions of these equations on the complementary space related to the Witt algebra decomposition are the odd function solutions. PMID:28413343

  2. Chimera and modulated drift states in a ring of nonlocally coupled oscillators with heterogeneous phase lags

    NASA Astrophysics Data System (ADS)

    Choe, Chol-Ung; Kim, Ryong-Son; Ri, Ji-Song

    2017-09-01

    We consider a ring of phase oscillators with nonlocal coupling strength and heterogeneous phase lags. We analyze the effects of heterogeneity in the phase lags on the existence and stability of a variety of steady states. A nonlocal coupling with heterogeneous phase lags that allows the system to be solved analytically is suggested and the stability of solutions along the Ott-Antonsen invariant manifold is explored. We present a complete bifurcation diagram for stationary patterns including the uniform drift and modulated drift states as well as chimera state, which reveals that the stable modulated drift state and a continuum of metastable drift states could occur due to the heterogeneity of the phase lags. We verify our theoretical results using the direct numerical simulations of the model system.

  3. Tolerance of DNA Mismatches in Dmc1 Recombinase-mediated DNA Strand Exchange*

    PubMed Central

    Borgogno, María V.; Monti, Mariela R.; Zhao, Weixing; Sung, Patrick; Argaraña, Carlos E.; Pezza, Roberto J.

    2016-01-01

    Recombination between homologous chromosomes is required for the faithful meiotic segregation of chromosomes and leads to the generation of genetic diversity. The conserved meiosis-specific Dmc1 recombinase catalyzes homologous recombination triggered by DNA double strand breaks through the exchange of parental DNA sequences. Although providing an efficient rate of DNA strand exchange between polymorphic alleles, Dmc1 must also guard against recombination between divergent sequences. How DNA mismatches affect Dmc1-mediated DNA strand exchange is not understood. We have used fluorescence resonance energy transfer to study the mechanism of Dmc1-mediated strand exchange between DNA oligonucleotides with different degrees of heterology. The efficiency of strand exchange is highly sensitive to the location, type, and distribution of mismatches. Mismatches near the 3′ end of the initiating DNA strand have a small effect, whereas most mismatches near the 5′ end impede strand exchange dramatically. The Hop2-Mnd1 protein complex stimulates Dmc1-catalyzed strand exchange on homologous DNA or containing a single mismatch. We observed that Dmc1 can reject divergent DNA sequences while bypassing a few mismatches in the DNA sequence. Our findings have important implications in understanding meiotic recombination. First, Dmc1 acts as an initial barrier for heterologous recombination, with the mismatch repair system providing a second level of proofreading, to ensure that ectopic sequences are not recombined. Second, Dmc1 stepping over infrequent mismatches is likely critical for allowing recombination between the polymorphic sequences of homologous chromosomes, thus contributing to gene conversion and genetic diversity. PMID:26709229

  4. Melatonin for the prevention and treatment of jet lag.

    PubMed

    Herxheimer, A; Petrie, K J

    2002-01-01

    : Jet-lag commonly affects air travellers who cross several time zones. It results from the body's internal rhythms being out of step with the day-night cycle at the destination. Melatonin is a pineal hormone that plays a central part in regulating bodily rhythms and has been used as a drug to re-align them with the outside world. : To assess the effectiveness of oral melatonin taken in different dosage regimens for alleviating jet-lag after air travel across several time zones. : We searched the Cochrane Controlled Trials Register, MEDLINE, EMBASE, PsychLit and Science Citation Index electronically, and the journals 'Aviation, Space and Environmental Medicine' and 'Sleep' by hand. We searched citation lists of relevant studies for other relevant trials. We asked principal authors of relevant studies to tell us about unpublished trials. Reports of adverse events linked to melatonin use outside randomised trials were searched for systematically in 'Side Effects of Drugs' (SED) and SED Annuals, 'Reactions Weekly', MEDLINE, and the adverse drug reactions databases of the WHO Uppsala Monitoring Centre (UMC) and the US Food & Drug Administration. : Randomised trials in airline passengers, airline staff or military personnel given oral melatonin, compared with placebo or other medication. Outcome measures should consist of subjective rating of jet-lag or related components, such as subjective wellbeing, daytime tiredness, onset and quality of sleep, psychological functioning, duration of return to normal, or indicators of circadian rhythms. : Ten trials met the inclusion criteria. All compared melatonin with placebo; one in addition compared it with a hypnotic, zolpidem. Nine of the trials were of adequate quality to contribute to the assessment, one had a design fault and could not be used in the assessment. Reports of adverse events outside trials were found through MEDLINE, 'Reactions Weekly', and in the WHO UMC database. : Nine of the ten trials found that melatonin

  5. Investigating effects of hydroxypropyl methylcellulose (HPMC) molecular weight grades on lag time of press-coated ethylcellulose tablets.

    PubMed

    Patadia, Riddhish; Vora, Chintan; Mittal, Karan; Mashru, Rajashree

    2016-11-01

    The research undertaken exemplifies the effects of hydroxypropyl methylcellulose (HPMC) molecular weight (MW) grades of on lag time of press-coated ethylcellulose (EC) tablets. The formulation comprised an immediate release core (containing prednisone as a model drug) surrounded by compression coating with variegated EC-HPMC blends. Five selected HPMC grades (E5, E15, E50, K100LV and K4M) were explored at three different concentrations (10% w/w, 20% w/w and 30% w/w in outer coat) to understand their effects on lag time and drug release. In vitro drug release testing demonstrated that, with increase in concentration of E5 and E15, up to 30% w/w, the mean lag time decreased progressively; whereas with remaining grades, the mean lag time initially decreased up to 20% w/w level and thereafter increased for 30% w/w level. Importantly, with increase in HPMC concentration in the outer coat, the variability in lag time (%RSD; n = 6) was decreased for each of E5, E15 and E50, whereas increased for K100LV and K4M. In general, the variability in lag time was increased with increase in HPMC MW at studied concentration levels. Markedly, tablets with 30% w/w K4M in outer coat exhibited slight premature release (before the rupture of outer coat) along with high variability in lag time. Overall, the study concluded that low MW HPMCs (E5, E15 and E50) were found rather efficient than higher MW HPMCs for developing robust EC-based press-coated pulsatile release formulations where precise lag time followed by sharp burst release is desired.

  6. A lagged variable model for characterizing temporally dynamic export of legacy anthropogenic nitrogen from watersheds to rivers.

    PubMed

    Chen, Dingjiang; Guo, Yi; Hu, Minpeng; Dahlgren, Randy A

    2015-08-01

    Legacy nitrogen (N) sources originating from anthropogenic N inputs (NANI) may be a major cause of increasing riverine N exports in many regions, despite a significant decline in NANI. However, little quantitative knowledge exists concerning the lag effect of NANI on riverine N export. As a result, the N leaching lag effect is not well represented in most current watershed models. This study developed a lagged variable model (LVM) to address temporally dynamic export of watershed NANI to rivers. Employing a Koyck transformation approach used in economic analyses, the LVM expresses the indefinite number of lag terms from previous years' NANI with a lag term that incorporates the previous year's riverine N flux, enabling us to inversely calibrate model parameters from measurable variables using Bayesian statistics. Applying the LVM to the upper Jiaojiang watershed in eastern China for 1980-2010 indicated that ~97% of riverine export of annual NANI occurred in the current year and succeeding 10 years (~11 years lag time) and ~72% of annual riverine N flux was derived from previous years' NANI. Existing NANI over the 1993-2010 period would have required a 22% reduction to attain the target TN level (1.0 mg N L(-1)), guiding watershed N source controls considering the lag effect. The LVM was developed with parsimony of model structure and parameters (only four parameters in this study); thus, it is easy to develop and apply in other watersheds. The LVM provides a simple and effective tool for quantifying the lag effect of anthropogenic N input on riverine export in support of efficient development and evaluation of watershed N control strategies.

  7. Role of Saccharomyces cerevisiae Msh2 and Msh3 repair proteins in double-strand break-induced recombination

    PubMed Central

    Sugawara, Neal; Pâques, Frédéric; Colaiácovo, Mónica; Haber, James E.

    1997-01-01

    When gene conversion is initiated by a double-strand break (DSB), any nonhomologous DNA that may be present at the ends must be removed before new DNA synthesis can be initiated. In Saccharomyces cerevisiae, removal of nonhomologous ends depends not only on the nucleotide excision repair endonuclease Rad1/Rad10 but also on Msh2 and Msh3, two proteins that are required to correct mismatched bp. These proteins have no effect when DSB ends are homologous to the donor, either in the kinetics of recombination or in the proportion of gene conversions associated with crossing-over. A second DSB repair pathway, single-strand annealing also requires Rad1/Rad10 and Msh2/Msh3, but reveals a difference in their roles. When the flanking homologous regions that anneal are 205 bp, the requirement for Msh2/Msh3 is as great as for Rad1/Rad10; but when the annealing partners are 1,170 bp, Msh2/Msh3 have little effect, while Rad1/Rad10 are still required. Mismatch repair proteins Msh6, Pms1, and Mlh1 are not required. We suggest Msh2 and Msh3 recognize not only heteroduplex loops and mismatched bp, but also branched DNA structures with a free 3′ tail. PMID:9256462

  8. Role of Saccharomyces cerevisiae Msh2 and Msh3 repair proteins in double-strand break-induced recombination.

    PubMed

    Sugawara, N; Pâques, F; Colaiácovo, M; Haber, J E

    1997-08-19

    When gene conversion is initiated by a double-strand break (DSB), any nonhomologous DNA that may be present at the ends must be removed before new DNA synthesis can be initiated. In Saccharomyces cerevisiae, removal of nonhomologous ends depends not only on the nucleotide excision repair endonuclease Rad1/Rad10 but also on Msh2 and Msh3, two proteins that are required to correct mismatched bp. These proteins have no effect when DSB ends are homologous to the donor, either in the kinetics of recombination or in the proportion of gene conversions associated with crossing-over. A second DSB repair pathway, single-strand annealing also requires Rad1/Rad10 and Msh2/Msh3, but reveals a difference in their roles. When the flanking homologous regions that anneal are 205 bp, the requirement for Msh2/Msh3 is as great as for Rad1/Rad10; but when the annealing partners are 1,170 bp, Msh2/Msh3 have little effect, while Rad1/Rad10 are still required. Mismatch repair proteins Msh6, Pms1, and Mlh1 are not required. We suggest Msh2 and Msh3 recognize not only heteroduplex loops and mismatched bp, but also branched DNA structures with a free 3' tail.

  9. Estimation of Prestress Force Distribution in the Multi-Strand System of Prestressed Concrete Structures

    PubMed Central

    Cho, Keunhee; Park, Sung Yong; Cho, Jeong-Rae; Kim, Sung Tae; Park, Young-Hwan

    2015-01-01

    Prestressed concrete (PSC) is one of the most reliable, durable and widely used construction materials, which overcomes the weakness of concrete in tension by the introduction of a prestress force. Smart strands enabling measurement of the prestress force have recently been developed to maintain PSC structures throughout their lifetime. However, the smart strand cannot give a representative indication of the whole prestress force when used in multi-strand systems since each strand sustains a different prestress force. In this paper, the actual distribution of the prestress force in a multi-strand system is examined using elastomagnetic (EM) sensors to develop a method for tracking representative indicators of the prestress force using smart strands. PMID:26083230

  10. An experimental and analytical investigation of stall effects on flap-lag stability in forward flight

    NASA Technical Reports Server (NTRS)

    Nagabhushanam, J.; Gaonkar, Gopal H.; Mcnulty, Michael J.

    1987-01-01

    Experiments have been performed with a 1.62 m diameter hingeless rotor in a wind tunnel to investigate flap-lag stability of isolated rotors in forward flight. The three-bladed rotor model closely approaches the simple theoretical concept of a hingeless rotor as a set of rigid, articulated flap-lag blades with offset and spring restrained flap and lag hinges. Lag regressing mode stability data was obtained for advance ratios as high as 0.55 for various combinations of collective pitch and shaft angle. The prediction includes quasi-steady stall effects on rotor trim and Floquet stability analyses. Correlation between data and prediction is presented and is compared with that of an earlier study based on a linear theory without stall effects. While the results with stall effects show marked differences from the linear theory results, the stall theory still falls short of adequate agreement with the experimental data.

  11. Expression of LAG-3 defines exhaustion of intratumoral PD-1+ T cells and correlates with poor outcome in follicular lymphoma

    PubMed Central

    Yang, Zhi-Zhang; Kim, Hyo Jin; Villasboas, Jose C.; Chen, Ya-Ping; Price-Troska, Tammy; Jalali, Shahrzad; Wilson, Mara; Novak, Anne J.; Ansell, Stephen M.

    2017-01-01

    Exhausted T-cells in follicular lymphoma (FL) typically express PD-1, but expression of PD-1 is not limited to exhausted cells. Although expected to be functionally suppressed, we found that the population of intratumoral PD-1+ T cells were predominantly responsible for production of cytokines and granules. This surprising finding prompted us to explore the involvement of LAG-3 to specifically identify functionally exhausted T cells. We found that LAG-3 was expressed on a subset of intratumoral T cells from FL and LAG-3+ T cells almost exclusively came from PD-1+ population. CyTOF analysis revealed that intratumoral LAG-3+ T cells were phenotypically heterogeneous as LAG-3 was expressed on a variety of T cell subsets. In contrast to PD-1+LAG-3- cells, intratumoral PD-1+LAG-3+ T cells exhibited reduced capacity to produce cytokines and granules. LAG-3 expression could be substantially upregulated on CD4+ or CD8+ T cells by IL-12, a cytokine that has been shown to induce T-cell exhaustion and be increased in the serum of lymphoma patients. Furthermore, we found that blockade of both PD-1 and LAG-3 signaling enhanced the function of intratumoral CD8+ T cells resulting in increased IFN-γ and IL-2 production. Clinically, LAG-3 expression on intratumoral T cells correlated with a poor outcome in FL patients. Taken together, we find that LAG-3 expression is necessary to identify the population of intratumoral PD-1+ T cells that are functionally exhausted and, in contrast, find that PD-1+LAG-3- T cells are simply activated cells that are immunologically functional. These findings may have important implications for immune checkpoint therapy in FL. PMID:28977875

  12. Review: the environmental status and implications of the nitrate time lag in Europe and North America

    NASA Astrophysics Data System (ADS)

    Vero, Sara E.; Basu, Nandita B.; Van Meter, Kimberly; Richards, Karl G.; Mellander, Per-Erik; Healy, Mark G.; Fenton, Owen

    2018-02-01

    The efficacy of water quality policies aiming to reduce or prevent nitrate contamination of waterbodies may be constrained by the inherent delay or "time lag" of water and solute transport through unsaturated (soil) and saturated (groundwater) pathways. These delays must be quantified in order to establish realistic deadlines, thresholds and policy expectations, and to design effective best management practices. The objective of this review is to synthesise the current state of research on nitrate-related time lags in both the European and North American environmental and legislative contexts. The durations of time lags have been found to differ according to climatic, pedological, landscape and management scenarios. Elucidation of these driving factors at a watershed scale is essential where water quality is impaired or at risk. Finally, the existence of time lags is increasingly being acknowledged at a policy level and incorporated into the development of environmental legislation. However, the full impact of these time lags is not yet fully understood or appreciated, and continued outreach and education in scientific, public and policy venues is still required.

  13. miRNA*: a passenger stranded in RNA-induced silencing complex?

    PubMed

    Mah, S M; Buske, C; Humphries, R K; Kuchenbauer, F

    2010-01-01

    Processing of the pre-microRNA (pre-miRNA) through Dicer1 generates a miRNA duplex, consisting of a miRNA and miRNA* strand (also termed guide strand and passenger strand, respectively). Despite the general consensus that miRNA*s have no regulatory activity, recent publications have provided evidence that the abundance, possible function, and physiological relevance of miRNA*s have been underestimated. This review provides an account of our current understanding of miRNA* origination and activity, mounting evidence for their unique functions and regulatory mechanisms, and examples of specific miRNA*s from the literature.

  14. Survey of stranded gas and delivered costs to Europe of selected gas resources

    USGS Publications Warehouse

    Attanasi, E.D.; Freeman, P.A.

    2011-01-01

    Two important trends affecting the expected growth of global gas markets are (1) the shift by many industrialized countries from coal-fired electricity generation to the use of natural gas to generate electricity and (2) the industrialization of the heavily populated Asian countries of India and China. This paper surveys discovered gas in stranded conventional gas accumulations and presents estimates of the cost of developing and producing stranded gas in selected countries. Stranded gas is natural gas in discovered or identified fields that is not currently commercially producible for either physical or economic reasons. Published reserves of gas at the global level do not distinguish between volumes of gas in producing fields and volumes in nonproducing fields. Data on stranded gas reported here-that is the volumes, geographical distribution, and size distributions of stranded gas fields at the country and regional level-are based on the examination of individual-field data and represent a significant improvement in information available to industry and government decision makers. Globally, stranded gas is pervasive, but large volumes in large accumulations are concentrated in only a few areas. The cost component of the paper focuses on stranded conventional gas accumulations in Africa and South America that have the potential to augment supplies to Europe. The methods described for the computation of extraction and transport costs are innovative in that they use information on the sizes and geographical distribution of the identified stranded gas fields. The costs are based on industry data specific to the country and geologic basin where the stranded gas is located. Gas supplies to Europe can be increased significantly at competitive costs by the development of stranded gas. Net extraction costs of producing the identified gas depend critically on the natural-gas-liquids (NGLs) content, the prevailing prices of liquids, the size of the gas accumulation, and the

  15. Detecting PM2.5's Correlations between Neighboring Cities Using a Time-Lagged Cross-Correlation Coefficient.

    PubMed

    Wang, Fang; Wang, Lin; Chen, Yuming

    2017-08-31

    In order to investigate the time-dependent cross-correlations of fine particulate (PM2.5) series among neighboring cities in Northern China, in this paper, we propose a new cross-correlation coefficient, the time-lagged q-L dependent height crosscorrelation coefficient (denoted by p q (τ, L)), which incorporates the time-lag factor and the fluctuation amplitude information into the analogous height cross-correlation analysis coefficient. Numerical tests are performed to illustrate that the newly proposed coefficient ρ q (τ, L) can be used to detect cross-correlations between two series with time lags and to identify different range of fluctuations at which two series possess cross-correlations. Applying the new coefficient to analyze the time-dependent cross-correlations of PM2.5 series between Beijing and the three neighboring cities of Tianjin, Zhangjiakou, and Baoding, we find that time lags between the PM2.5 series with larger fluctuations are longer than those between PM2.5 series withsmaller fluctuations. Our analysis also shows that cross-correlations between the PM2.5 series of two neighboring cities are significant and the time lags between two PM2.5 series of neighboring cities are significantly non-zero. These findings providenew scientific support on the view that air pollution in neighboring cities can affect one another not simultaneously but with a time lag.

  16. Higher 5-hydroxymethylcytosine identifies immortal DNA strand chromosomes in asymmetrically self-renewing distributed stem cells

    PubMed Central

    Huh, Yang Hoon; Cohen, Justin; Sherley, James L.

    2013-01-01

    Immortal strands are the targeted chromosomal DNA strands of nonrandom sister chromatid segregation, a mitotic chromosome segregation pattern unique to asymmetrically self-renewing distributed stem cells (DSCs). By nonrandom segregation, immortal DNA strands become the oldest DNA strands in asymmetrically self-renewing DSCs. Nonrandom segregation of immortal DNA strands may limit DSC mutagenesis, preserve DSC fate, and contribute to DSC aging. The mechanisms responsible for specification and maintenance of immortal DNA strands are unknown. To discover clues to these mechanisms, we investigated the 5-methylcytosine and 5-hydroxymethylcytosine (5hmC) content on chromosomes in mouse hair follicle DSCs during nonrandom segregation. Although 5-methylcytosine content did not differ significantly, the relative content of 5hmC was significantly higher in chromosomes containing immortal DNA strands than in opposed mitotic chromosomes containing younger mortal DNA strands. The difference in relative 5hmC content was caused by the loss of 5hmC from mortal chromosomes. These findings implicate higher 5hmC as a specific molecular determinant of immortal DNA strand chromosomes. Because 5hmC is an intermediate during DNA demethylation, we propose a ten-eleven translocase enzyme mechanism for both the specification and maintenance of nonrandomly segregated immortal DNA strands. The proposed mechanism reveals a means by which DSCs “know” the generational age of immortal DNA strands. The mechanism is supported by molecular expression data and accounts for the selection of newly replicated DNA strands when nonrandom segregation is initiated. These mechanistic insights also provide a possible basis for another characteristic property of immortal DNA strands, their guanine ribonucleotide dependency. PMID:24082118

  17. Oriented-strand-board- the wave of the future- for the building trade

    Treesearch

    Linda Ashton

    1984-01-01

    Move over, plywood. Oriented-strand board is here. It's less expensive. It's as durable. It has as many uses. And it is the wave of the future. "Oriented-strand board is a direct substitute for plywood" said Jerry Buckner, plant manager for the Martco oriented-strand board plant in Lemoyen. OSB, as it is commonly called, is a structural panel made...

  18. Investigating critical effects of variegated lubricants, glidants and hydrophilic additives on lag time of press coated ethylcellulose tablets.

    PubMed

    Patadia, Riddhish; Vora, Chintan; Mittal, Karan; Mashru, Rajashree

    2016-01-01

    The research envisaged focuses on vital impacts of variegated lubricants, glidants and hydrophilic additives on lag time of press coated ethylcellulose (EC) tablets using prednisone as a model drug. Several lubricants and glidants such as magnesium stearate, colloidal SiO2, sodium stearyl fumarate, talc, stearic acid, polyethylene glycol (6000) and glyceryl behenate were investigated to understand their effects on lag time by changing their concentrations in outer coat. Further, the effects of hydrophilic additives on lag time were examined for hydroxypropylmethylcellulose (E5), hydroxypropylcellulose (EF and SSL), povidone (K30), copovidone, polyethylene glycol (4000), lactose and mannitol. In vitro drug release testing revealed that each selected lubricant/glidant, if present even at concentration of 0.25% w/w, significantly reduced the lag time of press coated tablets. Specifically, colloidal SiO2 and/or magnesium stearate were detrimental while other lubricants/glidants were relatively less injurious. Among hydrophilic additives, freely water soluble fillers had utmost influence in lag time, whereas, comparatively less impact was observed with polymeric binders. Concisely, glidant and lubricant should be chosen to have minimal impact on lag time and further judicious selection of hydrophilic additives should be exercised for modulating lag time of pulsatile release formulations.

  19. Evidence that MEK1 positively promotes interhomologue double-strand break repair

    PubMed Central

    Terentyev, Yaroslav; Johnson, Rebecca; Neale, Matthew J.; Khisroon, Muhammad; Bishop-Bailey, Anna; Goldman, Alastair S. H.

    2010-01-01

    During meiosis there is an imperative to create sufficient crossovers for homologue segregation. This can be achieved during repair of programmed DNA double-strand breaks (DSBs), which are biased towards using a homologue rather than sister chromatid as a repair template. Various proteins contribute to this bias, one of which is a meiosis specific kinase Mek1. It has been proposed that Mek1 establishes the bias by creating a barrier to sister chromatid repair, as distinct from enforcing strand invasion with the homologue. We looked for evidence that Mek1 positively stimulates strand invasion of the homologue. This was done by analysing repair of DSBs induced by the VMA1-derived endonuclease (VDE) and flanked by directly repeated sequences that can be used for intrachromatid single-strand annealing (SSA). SSA competes with interhomologue strand invasion significantly more successfully when Mek1 function is lost. We suggest the increase in intrachromosomal SSA reflects an opportunistic default repair pathway due to loss of a MEK1 stimulated bias for strand invasion of the homologous chromosome. Making use of an inhibitor sensitive mek1-as1 allele, we found that Mek1 function influences the repair pathway throughout the first4–5 h of meiosis. Perhaps reflecting a particular need to create bias for successful interhomologue events before chromosome pairing is complete. PMID:20223769

  20. Reducing the lag of accommodation by auditory biofeedback: A pilot study.

    PubMed

    Wagner, Sandra; Ohlendorf, Arne; Schaeffel, Frank; Wahl, Siegfried

    2016-12-01

    The purpose of this study was to investigate whether a reduction of the accommodative lag is possible by training the accuracy of accommodation using auditory biofeedback. Accommodation responses were measured in thirty-one young adults with myopia for dioptric target distances of 2.0, 2.5, and 3.0D using an eccentric infrared photorefractor. For the biofeedback training, subjects were randomly assigned to an experimental (n=15) or a control group (n=16). Subjects of the experimental group were provided with two tones while fixating a target, one tone was related to their accommodative response and the second to the target distance. Their task was to match these tones. The control group did not receive any auditory biofeedback. Two different training methods were applied, a continuous training of 200s, and ten consecutive sessions of 20s each. The training effects on the lag of accommodation (change Δ) were highly variable. Regarding the entire study group, the observed change in the accommodative lag was greater at closer distances, while no difference between the two training methods was revealed. Nevertheless, seven experimental subjects reduced their lag by ⩾0.3D (3.0D target distance: Δ long =-0.29±0.20D, Δ short =-0.24±0.21D). This reduction was also seen in two control subjects. Remeasurement revealed that the average training effect cannot be preserved over a period of 5-7days. The current investigation has shown that the accuracy of accommodation can be trained in some subjects using auditory biofeedback for target distances of 2.5D or closer. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Internet and free press are associated with reduced lags in global outbreak reporting.

    PubMed

    McAlarnen, Lindsey; Smith, Katherine; Brownstein, John S; Jerde, Christopher

    2014-10-30

    Global outbreak detection and reporting have generally improved for a variety of infectious diseases and geographic regions in recent decades. Nevertheless, lags in outbreak reporting remain a threat to the global human health and economy. In the time between first occurrence of a novel disease incident and public notification of an outbreak, infected individuals have a greater possibility of traveling and spreading the pathogen to other nations. Shortening outbreak reporting lags has the potential to improve global health by preventing local outbreaks from escalating into global epidemics. Reporting lags between the first record and the first public report of an event were calculated for 318 outbreaks occurring 1996-2009. The influence of freedom of the press, Internet usage, per capita health expenditure, and cell phone subscriptions, on the timeliness of outbreak reporting was evaluated. Freer presses and increasing Internet usage correlate with reduced time between the first record of an outbreak and the public report. Increasing Internet usage reduced the expected reporting lag from more than one month in nations without Internet users to one day in those where 75 of 100 people use the Internet. Advances in technology and the emergence of more open and free governments are associated with to improved global infectious disease surveillance.

  2. A Fixed-Lag Kalman Smoother to Filter Power Line Interference in Electrocardiogram Recordings.

    PubMed

    Warmerdam, G J J; Vullings, R; Schmitt, L; Van Laar, J O E H; Bergmans, J W M

    2017-08-01

    Filtering power line interference (PLI) from electrocardiogram (ECG) recordings can lead to significant distortions of the ECG and mask clinically relevant features in ECG waveform morphology. The objective of this study is to filter PLI from ECG recordings with minimal distortion of the ECG waveform. In this paper, we propose a fixed-lag Kalman smoother with adaptive noise estimation. The performance of this Kalman smoother in filtering PLI is compared to that of a fixed-bandwidth notch filter and several adaptive PLI filters that have been proposed in the literature. To evaluate the performance, we corrupted clean neonatal ECG recordings with various simulated PLI. Furthermore, examples are shown of filtering real PLI from an adult and a fetal ECG recording. The fixed-lag Kalman smoother outperforms other PLI filters in terms of step response settling time (improvements that range from 0.1 to 1 s) and signal-to-noise ratio (improvements that range from 17 to 23 dB). Our fixed-lag Kalman smoother can be used for semi real-time applications with a limited delay of 0.4 s. The fixed-lag Kalman smoother presented in this study outperforms other methods for filtering PLI and leads to minimal distortion of the ECG waveform.

  3. Architecture and biogenesis of plus-strand RNA virus replication factories

    PubMed Central

    Paul, David; Bartenschlager, Ralf

    2013-01-01

    Plus-strand RNA virus replication occurs in tight association with cytoplasmic host cell membranes. Both, viral and cellular factors cooperatively generate distinct organelle-like structures, designated viral replication factories. This compartmentalization allows coordination of the different steps of the viral replication cycle, highly efficient genome replication and protection of the viral RNA from cellular defense mechanisms. Electron tomography studies conducted during the last couple of years revealed the three dimensional structure of numerous plus-strand RNA virus replication compartments and highlight morphological analogies between different virus families. Based on the morphology of virus-induced membrane rearrangements, we propose two separate subclasses: the invaginated vesicle/spherule type and the double membrane vesicle type. This review discusses common themes and distinct differences in the architecture of plus-strand RNA virus-induced membrane alterations and summarizes recent progress that has been made in understanding the complex interplay between viral and co-opted cellular factors in biogenesis and maintenance of plus-strand RNA virus replication factories. PMID:24175228

  4. Programmable energy landscapes for kinetic control of DNA strand displacement.

    PubMed

    Machinek, Robert R F; Ouldridge, Thomas E; Haley, Natalie E C; Bath, Jonathan; Turberfield, Andrew J

    2014-11-10

    DNA is used to construct synthetic systems that sense, actuate, move and compute. The operation of many dynamic DNA devices depends on toehold-mediated strand displacement, by which one DNA strand displaces another from a duplex. Kinetic control of strand displacement is particularly important in autonomous molecular machinery and molecular computation, in which non-equilibrium systems are controlled through rates of competing processes. Here, we introduce a new method based on the creation of mismatched base pairs as kinetic barriers to strand displacement. Reaction rate constants can be tuned across three orders of magnitude by altering the position of such a defect without significantly changing the stabilities of reactants or products. By modelling reaction free-energy landscapes, we explore the mechanistic basis of this control mechanism. We also demonstrate that oxDNA, a coarse-grained model of DNA, is capable of accurately predicting and explaining the impact of mismatches on displacement kinetics.

  5. Transformation of Saccharomyces cerevisiae with UV-irradiated single-stranded plasmid.

    PubMed

    Zgaga, Z

    1991-08-01

    UV-irradiated single-stranded replicative plasmids were used to transform different yeast strains. The low doses of UV used in this study (10-75 J/m2) caused a significant decrease in the transforming efficiency of plasmid DNA in the Rad+ strain, while they had no effect on transformation with double-stranded plasmids of comparable size. Neither the rev3 mutation, nor the rad18 or rad52 mutations influenced the efficiency of transformation with irradiated single-stranded plasmid. However, it was found to be decreased in the double rev3 rad52 mutant. Extracellular irradiation of plasmid that contains both URA3 and LEU2 genes (psLU) gave rise to up to 5% Leu- transformants among selected Ura+ ones in the repair-proficient strain. Induction of Leu- transformants was dose-dependent and only partially depressed in the rev3 mutant. These results suggest that both mutagenic and recombinational repair processes operate on UV-damaged single-stranded DNA in yeast.

  6. Lagged effects of family-supportive organization perceptions and supervision in relation to generalized work-related resources.

    PubMed

    Matthews, Russell A; Toumbeva, Tatiana H

    2015-07-01

    In the present study, grounded in organizational support and social exchange theory, the dynamic lagged interplay between family supportive supervision (FSS), family supportive organization perceptions (FSOP), perceived organizational support (POS), and leader-member exchange (LMX) was examined. Data were collected from 435 respondents over 3 time points with 6-week lags between assessments. Consistent with theory, FSS had a significant lagged effect on FSOP, whereas the reverse relationship was not supported. Interestingly, contrary to conservation of resources theory, we did not find significant lagged effects between POS and FSOP. Results further indicated that LMX and FSS were reciprocally related over time, suggesting the potential for a dynamic, mutually beneficial exchange relationship between subordinates and supervisors. Theoretical implications and considerations for research and practice are discussed. (PsycINFO Database Record (c) 2015 APA, all rights reserved).

  7. On the Nature of QPO Phase Lags in Black Hole Candidates

    NASA Technical Reports Server (NTRS)

    Shaposhnikov, Nikolai

    2012-01-01

    Observations of quasi-periodic oscillations (QPOs) in X-ray binaries hold a key to understanding many aspects of these enigmatic systems. Complex appearance of the Fourier phase lags related to QPOs is one of the most puzzling observational effects in accreting black holes. In this Letter we show that QPO properties, including phase lags, can be explained in a framework of a simple scenario, where the oscillating media provides a feedback on the emerging spectrum. We demonstrate that the QPO waveform is presented by the product of a perturbation and a time delayed response factors, where the response is energy dependent. The essential property of this effect is its non-linear and multiplicative nature. Our multiplicative reverberation model successfully describes the QPO components in energy dependent power spectra as well as the appearance of the phase lags between signal in different energy bands. We apply our model to QPOs observed by Rossi X-ray Timing Explorer in BH candidate XTE J1550-564. We briefly discuss the implications of the observed energy dependence of the QPO reverberation times and amplitudes to the nature of the power law spectral component and its variability.

  8. Stranded cost recovery: Reregulating the electricity markets in the United States

    NASA Astrophysics Data System (ADS)

    Wagle, Pushkar Ghanashyam

    2000-10-01

    For the past few years, Stranded Cost recovery has been one of the most contentious issues regarding the restructuring of electricity markets among the regulators, researchers, and the other interested parties. Among the states that have moved towards retail competition, some have already made decisions regarding the levels of the stranded cost recovery. So the question is: how have these states handled the "stranded cost problem"? Following the introduction and the historical perspective of the industry in the first chapter, the second chapter takes a broad view for understanding the overall process of deregulation. It attempts to analyze why some states have made a rapid transition to competition in the electric utility industry, while other states are just beginning to consider the issue. White (1996) and Ando & Palmer (1998) have conducted a similar exercise. We present a more comprehensive and theoretically informed econometric analysis that sheds light over some of the crucial issues involved in restructuring, such as, stranded cost recovery, regulation of transmission and distribution sectors, and establishment of Independent System Operator, etc. This chapter offers the rationale for alternative econometric techniques, and extends the political economy analysis to incorporate actual timings of retail competition. Once we have identified the role of stranded cost in restructuring and the theoretical foundations, we study empirically the political economy of states' decisions to grant stranded cost recovery. This constitutes the third chapter. Here, we concentrate on California and Pennsylvania, two states that are at the frontiers of deregulation, and compare their respective treatments of the stranded cost. We probe the reasons behind Pennsylvania's lead over California on the path towards deregulation.

  9. Late Quaternary slip history of the Mill Creek strand of the San Andreas fault in San Gorgonio Pass, southern California: The role of a subsidiary left-lateral fault in strand switching

    USGS Publications Warehouse

    Kendrick, Katherine J.; Matti, Jonathan; Mahan, Shannon

    2015-01-01

    The fault history of the Mill Creek strand of the San Andreas fault (SAF) in the San Gorgonio Pass region, along with the reconstructed geomorphology surrounding this fault strand, reveals the important role of the left-lateral Pinto Mountain fault in the regional fault strand switching. The Mill Creek strand has 7.1–8.7 km total slip. Following this displacement, the Pinto Mountain fault offset the Mill Creek strand 1–1.25 km, as SAF slip transferred to the San Bernardino, Banning, and Garnet Hill strands. An alluvial complex within the Mission Creek watershed can be linked to palinspastic reconstruction of drainage segments to constrain slip history of the Mill Creek strand. We investigated surface remnants through detailed geologic mapping, morphometric and stratigraphic analysis, geochronology, and pedogenic analysis. The degree of soil development constrains the duration of surface stability when correlated to other regional, independently dated pedons. This correlation indicates that the oldest surfaces are significantly older than 500 ka. Luminescence dates of 106 ka and 95 ka from (respectively) 5 and 4 m beneath a younger fan surface are consistent with age estimates based on soil-profile development. Offset of the Mill Creek strand by the Pinto Mountain fault suggests a short-term slip rate of ∼10–12.5 mm/yr for the Pinto Mountain fault, and a lower long-term slip rate. Uplift of the Yucaipa Ridge block during the period of Mill Creek strand activity is consistent with thermochronologic modeled uplift estimates.

  10. Tolerance of DNA Mismatches in Dmc1 Recombinase-mediated DNA Strand Exchange.

    PubMed

    Borgogno, María V; Monti, Mariela R; Zhao, Weixing; Sung, Patrick; Argaraña, Carlos E; Pezza, Roberto J

    2016-03-04

    Recombination between homologous chromosomes is required for the faithful meiotic segregation of chromosomes and leads to the generation of genetic diversity. The conserved meiosis-specific Dmc1 recombinase catalyzes homologous recombination triggered by DNA double strand breaks through the exchange of parental DNA sequences. Although providing an efficient rate of DNA strand exchange between polymorphic alleles, Dmc1 must also guard against recombination between divergent sequences. How DNA mismatches affect Dmc1-mediated DNA strand exchange is not understood. We have used fluorescence resonance energy transfer to study the mechanism of Dmc1-mediated strand exchange between DNA oligonucleotides with different degrees of heterology. The efficiency of strand exchange is highly sensitive to the location, type, and distribution of mismatches. Mismatches near the 3' end of the initiating DNA strand have a small effect, whereas most mismatches near the 5' end impede strand exchange dramatically. The Hop2-Mnd1 protein complex stimulates Dmc1-catalyzed strand exchange on homologous DNA or containing a single mismatch. We observed that Dmc1 can reject divergent DNA sequences while bypassing a few mismatches in the DNA sequence. Our findings have important implications in understanding meiotic recombination. First, Dmc1 acts as an initial barrier for heterologous recombination, with the mismatch repair system providing a second level of proofreading, to ensure that ectopic sequences are not recombined. Second, Dmc1 stepping over infrequent mismatches is likely critical for allowing recombination between the polymorphic sequences of homologous chromosomes, thus contributing to gene conversion and genetic diversity. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Replication of tobacco mosaic virus RNA.

    PubMed Central

    Buck, K W

    1999-01-01

    The replication of tobacco mosaic virus (TMV) RNA involves synthesis of a negative-strand RNA using the genomic positive-strand RNA as a template, followed by the synthesis of positive-strand RNA on the negative-strand RNA templates. Intermediates of replication isolated from infected cells include completely double-stranded RNA (replicative form) and partly double-stranded and partly single-stranded RNA (replicative intermediate), but it is not known whether these structures are double-stranded or largely single-stranded in vivo. The synthesis of negative strands ceases before that of positive strands, and positive and negative strands may be synthesized by two different polymerases. The genomic-length negative strand also serves as a template for the synthesis of subgenomic mRNAs for the virus movement and coat proteins. Both the virus-encoded 126-kDa protein, which has amino-acid sequence motifs typical of methyltransferases and helicases, and the 183-kDa protein, which has additional motifs characteristic of RNA-dependent RNA polymerases, are required for efficient TMV RNA replication. Purified TMV RNA polymerase also contains a host protein serologically related to the RNA-binding subunit of the yeast translational initiation factor, eIF3. Study of Arabidopsis mutants defective in RNA replication indicates that at least two host proteins are needed for TMV RNA replication. The tomato resistance gene Tm-1 may also encode a mutant form of a host protein component of the TMV replicase. TMV replicase complexes are located on the endoplasmic reticulum in close association with the cytoskeleton in cytoplasmic bodies called viroplasms, which mature to produce 'X bodies'. Viroplasms are sites of both RNA replication and protein synthesis, and may provide compartments in which the various stages of the virus mutiplication cycle (protein synthesis, RNA replication, virus movement, encapsidation) are localized and coordinated. Membranes may also be important for the

  12. Improved deoxyribozymes for synthesis of covalently branched DNA and RNA.

    PubMed

    Lee, Christine S; Mui, Timothy P; Silverman, Scott K

    2011-01-01

    A covalently branched nucleic acid can be synthesized by joining the 2'-hydroxyl of the branch-site ribonucleotide of a DNA or RNA strand to the activated 5'-phosphorus of a separate DNA or RNA strand. We have previously used deoxyribozymes to synthesize several types of branched nucleic acids for experiments in biotechnology and biochemistry. Here, we report in vitro selection experiments to identify improved deoxyribozymes for synthesis of branched DNA and RNA. Each of the new deoxyribozymes requires Mn²(+) as a cofactor, rather than Mg²(+) as used by our previous branch-forming deoxyribozymes, and each has an initially random region of 40 rather than 22 or fewer combined nucleotides. The deoxyribozymes all function by forming a three-helix-junction (3HJ) complex with their two oligonucleotide substrates. For synthesis of branched DNA, the best new deoxyribozyme, 8LV13, has k(obs) on the order of 0.1 min⁻¹, which is about two orders of magnitude faster than our previously identified 15HA9 deoxyribozyme. 8LV13 also functions at closer-to-neutral pH than does 15HA9 (pH 7.5 versus 9.0) and has useful tolerance for many DNA substrate sequences. For synthesis of branched RNA, two new deoxyribozymes, 8LX1 and 8LX6, were identified with broad sequence tolerances and substantial activity at pH 7.5, versus pH 9.0 for many of our previous deoxyribozymes that form branched RNA. These experiments provide new, and in key aspects improved, practical catalysts for preparation of synthetic branched DNA and RNA.

  13. Consequences of Fluid Lag in Three-Dimensional Hydraulic Fractures

    NASA Astrophysics Data System (ADS)

    Advani (Deceased), S. H.; Lee, T. S.; Dean, R. H.; Pak, C. K.; Avasthi, J. M.

    1997-04-01

    Research investigations on three-dimensional (3-D) rectangular hydraulic fracture configurations with varying degrees of fluid lag are reported. This paper demonstrates that a 3-D fracture model coupled with fluid lag (a small region of reduced pressure) at the fracture tip can predict very large excess pressure measurements for hydraulic fracture processes. Predictions of fracture propagation based on critical stress intensity factors are extremely sensitive to the pressure profile at the tip of a propagating fracture. This strong sensitivity to the pressure profile at the tip of a hydraulic fracture is more strongly pronounced in 3-D models versus 2-D models because 3-D fractures are clamped at the top and bottom, and pressures in the 3-D fractures that are far removed from the fracture tip have little effect on the stress intensity factor at the fracture tip. This rationale for the excess pressure mechanism is in marked contrast to the crack tip process damage zone assumptions and attendant high rock fracture toughness value hypotheses advanced in the literature. A comparison with field data is presented to illustrate the proposed fracture fluid pressure sensitivity phenomenon. This paper does not attempt to calculate the length of the fluid lag region in a propagating fracture but instead attempts to show that the pressure profile at the tip of the propagating fracture plays a major role in fracture propagation, and this role is magnified in 3-D models. Int. J. Numer. Anal. Meth. Geomech., vol. 21, 229-240 (1997).

  14. Reduce Nb3Sn Strand Deformation when Fabricating High Jc Rutherford Cables

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

    Peng, Xuan

    2012-12-17

    During Phase I, our efforts were to reduce subelements deformation when fabricating Nb3Sn Rutherford cables. Our first focus is on 217-sublement tube type strand. We successfully made a few billets in OD tube with different Cu spacing between subelements, and supplied the strands to Fermi Lab for cabling. Through the rolling test characterization, these types of strands did not have enough bonding between subelements to withstand the deformation. We saw copper cracking between subelements in the deformed strands. We scaled up the billet from OD to 1.5 OD, and made two billets. This greatly improves the bonding. There is nomore » copper cracking in the deformed strands when we scaled up the diameter of the billets. Fermi Lab successfully made cables using one of this improved strands. In their cables, no Cu cracking and no filament bridging occurred. We also successfully made a couple of billets with hex OD and round ID subelements for 61-subelement restack. Due to the lack of bonding, we could not judge its cabling property properly. But we know through this experiment, we could keep the Nb round, once we select the proper Cu spacing.« less

  15. 75 FR 37382 - Notice of Antidumping Duty Order: Prestressed Concrete Steel Wire Strand from the People's...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-29

    ... strand (``PC strand'') from the People's Republic of China (``PRC''). On June 22, 2010, the ITC notified... investigation of PC strand from the PRC. See Prestressed Concrete Steel Wire Strand From the People's Republic... Determination''). Scope of the Order The scope of this investigation consists of PC strand, produced from wire...

  16. State-dependent resource harvesting with lagged information about system states

    USGS Publications Warehouse

    Johnson, Fred A.; Fackler, Paul L.; Boomer, G Scott; Zimmerman, Guthrie S.; Williams, Byron K.; Nichols, James D.; Dorazio, Robert

    2016-01-01

    Markov decision processes (MDPs), which involve a temporal sequence of actions conditioned on the state of the managed system, are increasingly being applied in natural resource management. This study focuses on the modification of a traditional MDP to account for those cases in which an action must be chosen after a significant time lag in observing system state, but just prior to a new observation. In order to calculate an optimal decision policy under these conditions, possible actions must be conditioned on the previous observed system state and action taken. We show how to solve these problems when the state transition structure is known and when it is uncertain. Our focus is on the latter case, and we show how actions must be conditioned not only on the previous system state and action, but on the probabilities associated with alternative models of system dynamics. To demonstrate this framework, we calculated and simulated optimal, adaptive policies for MDPs with lagged states for the problem of deciding annual harvest regulations for mallards (Anas platyrhynchos) in the United States. In this particular example, changes in harvest policy induced by the use of lagged information about system state were sufficient to maintain expected management performance (e.g. population size, harvest) even in the face of an uncertain system state at the time of a decision.

  17. Extending the Distributed Lag Model framework to handle chemical mixtures.

    PubMed

    Bello, Ghalib A; Arora, Manish; Austin, Christine; Horton, Megan K; Wright, Robert O; Gennings, Chris

    2017-07-01

    Distributed Lag Models (DLMs) are used in environmental health studies to analyze the time-delayed effect of an exposure on an outcome of interest. Given the increasing need for analytical tools for evaluation of the effects of exposure to multi-pollutant mixtures, this study attempts to extend the classical DLM framework to accommodate and evaluate multiple longitudinally observed exposures. We introduce 2 techniques for quantifying the time-varying mixture effect of multiple exposures on an outcome of interest. Lagged WQS, the first technique, is based on Weighted Quantile Sum (WQS) regression, a penalized regression method that estimates mixture effects using a weighted index. We also introduce Tree-based DLMs, a nonparametric alternative for assessment of lagged mixture effects. This technique is based on the Random Forest (RF) algorithm, a nonparametric, tree-based estimation technique that has shown excellent performance in a wide variety of domains. In a simulation study, we tested the feasibility of these techniques and evaluated their performance in comparison to standard methodology. Both methods exhibited relatively robust performance, accurately capturing pre-defined non-linear functional relationships in different simulation settings. Further, we applied these techniques to data on perinatal exposure to environmental metal toxicants, with the goal of evaluating the effects of exposure on neurodevelopment. Our methods identified critical neurodevelopmental windows showing significant sensitivity to metal mixtures. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Radioresistance of GGG Sequences to Prompt Strand Break Formation from Direct-Type Radiation Damage

    PubMed Central

    Black, Paul J.; Miller, Adam S.; Hayes, Jeffrey J.

    2016-01-01

    Purpose As humans, we are constantly exposed to ionizing radiation from natural, man-made and cosmic sources which can damage DNA, leading to deleterious effects including cancer incidence. In this work we introduce a method to monitor strand breaks resulting from damage due to the direct effect of ionizing radiation and provide evidence for sequence-dependent effects leading to strand breaks. Materials and methods To analyze only DNA strand breaks caused by radiation damage due to the direct effect of ionizing radiation, we combined an established technique to generate dehydrated DNA samples with a technique to analyze single strand breaks on short oligonucleotide sequences via denaturing gel electrophoresis. Results We find that direct damage primarily results in a reduced number of strand breaks in guanine triplet regions (GGG) when compared to isolated guanine (G) bases with identical flanking base context. In addition, we observe strand break behavior possibly indicative of protection of guanine bases when flanked by pyrimidines, and sensitization of guanine to strand break when flanked by adenine (A) bases in both isolated G and GGG cases. Conclusions These observations provide insight into the strand break behavior in GGG regions damaged via the direct effect of ionizing radiation. In addition, this could be indicative of DNA sequences that are naturally more susceptible to strand break due to the direct effect of ionizing radiation. PMID:27349757

  19. Four-Year Cross-Lagged Associations between Physical and Mental Health in the Medical Outcomes Study.

    ERIC Educational Resources Information Center

    Hays, Ron D.; And Others

    1994-01-01

    Applied structural equation modeling to evaluation of cross-lagged panel models. Self-reports of physical and mental health at three time points spanning four-year interval were analyzed to illustrate cross-lagged analysis methodology. Data were analyzed from 856 patients with hypertension, diabetes, heart disease, or depression. Cross-lagged…

  20. Modeling the lag period and exponential growth of Listeria monocytogenes under conditions of fluctuating temperature and water activity values.

    PubMed

    Muñoz-Cuevas, Marina; Fernández, Pablo S; George, Susan; Pin, Carmen

    2010-05-01

    The dynamic model for the growth of a bacterial population described by Baranyi and Roberts (J. Baranyi and T. A. Roberts, Int. J. Food Microbiol. 23:277-294, 1994) was applied to model the lag period and exponential growth of Listeria monocytogenes under conditions of fluctuating temperature and water activity (a(w)) values. To model the duration of the lag phase, the dependence of the parameter h(0), which quantifies the amount of work done during the lag period, on the previous and current environmental conditions was determined experimentally. This parameter depended not only on the magnitude of the change between the previous and current environmental conditions but also on the current growth conditions. In an exponentially growing population, any change in the environment requiring a certain amount of work to adapt to the new conditions initiated a lag period that lasted until that work was finished. Observations for several scenarios in which exponential growth was halted by a sudden change in the temperature and/or a(w) were in good agreement with predictions. When a population already in a lag period was subjected to environmental fluctuations, the system was reset with a new lag phase. The work to be done during the new lag phase was estimated to be the workload due to the environmental change plus the unfinished workload from the uncompleted previous lag phase.