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Sample records for duplex dna effect

  1. Effect of Radiofrequency Radiation on DNA Duplex Stability and Replication.

    Science.gov (United States)

    1983-08-01

    R.W., J.B. Dodgson, I.F. Nes, and R.D. Wells. Duplex regions in single-stranded OX174 DNA are cleaved by a restriction endonuclease from Haemophilus ... aegyptius . J Biol Chem 252:7300-7306 (1977). *i Brown, R.F., S.V. Marshall, and C.W. Hughes. Effect of radiofrequency radi- ation (RFR) on excision

  2. Spermine Condenses DNA, but Not RNA Duplexes

    Energy Technology Data Exchange (ETDEWEB)

    Katz, Andrea M.; Tolokh, Igor S.; Pabit, Suzette A.; Baker, Nathan; Onufriev, Alexey V.; Pollack, Lois

    2017-01-01

    Interactions between the polyamine spermine and nucleic acids drive important cellular processes. Spermine condenses DNA, and some RNAs such as poly(rA):poly(rU). A large fraction of the spermine present in cells is bound to RNA, but apparently does not condense it. Here, we study the effect of spermine binding to short duplex RNA and DNA and compare our findings with predictions of molecular dynamics simulations. When small numbers of spermine are introduced, RNA with a designed sequence, containing a mixture of 14 GC pairs and 11 AU pairs, resists condensation relative to DNA of an equivalent sequence or to 25 base pair poly(rA):poly(rU) RNA. Comparison of wide-angle x-ray scattering profiles with simulation suggests that spermine is sequestered deep within the major groove of mixed sequence RNA, preventing condensation by limiting opportunities to bridge to other molecules as well as stabilizing the RNA by locking it into a particular conformation. In contrast, for DNA, simulations suggest that spermine binds external to the duplex, offering opportunities for intermolecular interaction. The goal of this study is to explain how RNA can remain soluble, and available for interaction with other molecules in the cell, despite the presence of spermine at concentrations high enough to precipitate DNA.

  3. DNA duplex membrane effect for the electrochemical detection of single-base DNA mutations

    Institute of Scientific and Technical Information of China (English)

    Luo Chunxiong; Mao Yongdong; Ouyang Qi

    2006-01-01

    Here we report a new method to detect DNA point mutations.The method is based on the formation and deformation of double-stranded DNA(dsDNA)membranes on a gold surface.It can encage reporter molecules between the gold surface and the double-stranded DNA or keep them away from the gold surface.In these systems,Fe(CN)63- was used as the reporter.As the temperature increases,a sharp electrochemical signal change in the melting curve of wild-type dsDNA appears.At a special temperature,the and single base mutation target.Thus,the system provides a simple and sensitive method to detect DNA point mutations without labeling targets.

  4. Overstretching of a 30 bp DNA duplex studied with steered molecular dynamics simulation: Effects of structural defects on structure and force-extension relation

    Science.gov (United States)

    Li, H.; Gisler, T.

    2009-11-01

    Single-molecule experiments on polymeric DNA show that the molecule can be overstretched at nearly constant force by about 70% beyond its relaxed contour length. In this publication we use steered molecular dynamics (MD) simulation to study the effect of structural defects on force-extension curves and structures at high elongation in a 30 base pair duplex pulled by its torsionally unconstrained 5' -5' ends. The defect-free duplex shows a plateau in the force-extension curve at 120pN in which large segments with inclined and paired bases (“S-DNA”) near both ends of the duplex coexist with a central B-type segment separated from the former by small denaturation bubbles. In the presence of a base mismatch or a nick, force-extension curves are very similar to the ones of the defect-free duplex. For the duplex with a base mismatch, S-type segments with highly inclined base pairs are not observed; rather, the overstretched duplex consists of B-type segments separated by denaturation bubbles. The nicked duplex evolves, via a two-step transition, into a two-domain structure characterized by a large S-type segment coexisting with several short S-type segments which are separated by short denaturation bubbles. Our results suggest that in the presence of nicks the force-extension curve of highly elongated duplex DNA might reflect locally highly inhomogeneous stretching. Supplementary material in the form of a PDF file available from the Journal web page at 10.1140/epje/i2009-10524-5 and is accessible for authorised users.

  5. Effect of C5-Methylation of Cytosine on the UV-Induced Reactivity of Duplex DNA: Conformational and Electronic Factors.

    Science.gov (United States)

    Banyasz, Akos; Esposito, Luciana; Douki, Thierry; Perron, Marion; Lepori, Clément; Improta, Roberto; Markovitsi, Dimitra

    2016-05-12

    C5-methylation of cytosines is strongly correlated with UV-induced mutations detected in skin cancers. Mutational hot-spots appearing at TCG sites are due to the formation of pyrimidine cyclobutane dimers (CPDs). The present study, performed for the model DNA duplex (TCGTA)3·(TACGA)3 and the constitutive single strands, examines the factors underlying the effect of C5-methylation on pyrimidine dimerization at TCG sites. This effect is quantified for the first time by quantum yields ϕ. They were determined following irradiation at 255, 267, and 282 nm and subsequent photoproduct analysis using HPLC coupled to mass spectrometry. C5-methylation leads to an increase of the CPD quantum yield up to 80% with concomitant decrease of that of pyrimidine(6-4) pyrimidone adducts (64PPs) by at least a factor of 3. The obtained ϕ values cannot be explained only by the change of the cytosine absorption spectrum upon C5-methylation. The conformational and electronic factors that may affect the dimerization reaction are discussed in light of results obtained by fluorescence spectroscopy, molecular dynamics simulations, and quantum mechanical calculations. Thus, it appears that the presence of an extra methyl on cytosine affects the sugar puckering, thereby enhancing conformations of the TC step that are prone to CPD formation but less favorable to 64PPs. In addition, C5-methylation diminishes the amplitude of conformational motions in duplexes; in the resulting stiffer structure, ππ* excitations may be transferred from initially populated exciton states to reactive pyrimidines giving rise to CPDs.

  6. Effect of initial ion positions on the interactions of monovalent and divalent ions with a DNA duplex as revealed with atomistic molecular dynamics simulations.

    Science.gov (United States)

    Robbins, Timothy J; Wang, Yongmei

    2013-01-01

    Monovalent (Na(+)) and divalent (Mg(2+)) ion distributions around the Dickerson-Drew dodecamer were studied by atomistic molecular dynamics (MD) simulations with AMBER molecular modeling software. Different initial placements of ions were tried and the resulting effects on the ion distributions around DNA were investigated. For monovalent ions, results were found to be nearly independent of initial cation coordinates. However, Mg(2+) ions demonstrated a strong initial coordinate dependent behavior. While some divalent ions initially placed near the DNA formed essentially permanent direct coordination complexes with electronegative DNA atoms, Mg(2+) ions initially placed further away from the duplex formed a full, nonexchanging, octahedral first solvation shell. These fully solvated cations were still capable of binding with DNA with events lasting up to 20 ns, and in comparison were bound much longer than Na(+) ions. Force field parameters were also investigated with modest and little differences arising from ion (ions94 and ions08) and nucleic acid description (ff99, ff99bsc0, and ff10), respectively. Based on known Mg(2+) ion solvation structure, we conclude that in most cases Mg(2+) ions retain their first solvation shell, making only solvent-mediated contacts with DNA duplex. The proper way to simulate Mg(2+) ions around DNA duplex, therefore, should begin with ions placed in the bulk water.

  7. DNA Duplex Engineering for Enantioselective Fluorescent Sensor.

    Science.gov (United States)

    Hu, Yuehua; Lin, Fan; Wu, Tao; Zhou, Yufeng; Li, Qiusha; Shao, Yong; Xu, Zhiai

    2017-02-21

    The rapid identification of biomacromolecule structure that has a specific association with chiral enantiomers especially from natural sources will be helpful in developing enantioselective sensor and in speeding up drug exploitation. Herein, owing to its existence also in living cells, apurinic/apyrimidinic site (AP site) was first engineered into ds-DNA duplex to explore its competence in enantiomer selectivity. An AP site-specific fluorophore was utilized as an enantioselective discrimination probe to develop a straightforward chiral sensor using natural tetrahydropalmatine (L- and D-THP) as enantiomer representatives. We found that only L-THP can efficiently replace the prebound fluorophore to cause a significant fluorescence increase due to its specific binding with the AP site (two orders magnitude higher in affinity than binding with D-THP). The AP site binding specificity of L-THP over D-THP was assessed via intrinsic fluorescence, isothermal titration calorimetry, and DNA stability. The enantioselective performance can be easily tuned by the sequences near the AP site and the number of AP sites. A single AP site provides a perfect binding pocket to differentiate the chiral atom-induced structure discrepancy. We expect that our work will inspire interest in engineering local structures into a ds-DNA duplex for developing novel enantioselective sensors.

  8. Kinetics and thermodynamics of DNA, RNA, and hybrid duplex formation.

    Science.gov (United States)

    Rauzan, Brittany; McMichael, Elizabeth; Cave, Rachel; Sevcik, Lesley R; Ostrosky, Kara; Whitman, Elisabeth; Stegemann, Rachel; Sinclair, Audra L; Serra, Martin J; Deckert, Alice A

    2013-02-05

    The rates of duplex formation for two octamers of DNA (5' d-CACGGCTC/5' d-GAGCCGTG and 5' d-CACAGCAC/5' d-GTGCTGTG), the homologous RNA, and both sets of hybrids in 1 M NaCl buffer have been measured using stopped-flow spectroscopy. In addition, the thermodynamic parameters, ΔH° and ΔS°, have been determined for the same sequences under the same buffer conditions using optical melting techniques. These data reveal a linear free energy relationship between the free energy of activation for denaturation and the change in free energy for formation of the duplexes. This relationship indicates that these duplex formation reactions occur through a common unstructured transition state that is more similar to the single strands in solution than to the ensuing duplex. In addition, these data confirm that the greater stability of RNA duplexes relative to that of homologous DNA and hybrid duplexes is controlled by the denaturation rate and not the duplex formation rate.

  9. Defined presentation of carbohydrates on a duplex DNA scaffold.

    Science.gov (United States)

    Schlegel, Mark K; Hütter, Julia; Eriksson, Magdalena; Lepenies, Bernd; Seeberger, Peter H

    2011-12-16

    A new method for the spatially defined alignment of carbohydrates on a duplex DNA scaffold is presented. The use of an N-hydroxysuccinimide (NHS)-ester phosphoramidite along with carbohydrates containing an alkylamine linker allows for on-column labeling during solid-phase oligonucleotide synthesis. This modification method during solid-phase synthesis only requires the use of minimal amounts of complex carbohydrates. The covalently attached carbohydrates are presented in the major groove of the B-form duplex DNA as potential substrates for murine type II C-type lectin receptors mMGL1 and mMGL2. CD spectroscopy and thermal melting revealed only minimal disturbance of the overall helical structure. Surface plasmon resonance and cellular uptake studies with bone-marrow-derived dendritic cells were used to assess the capability of these carbohydrate-modified duplexes to bind to mMGL receptors.

  10. Micromechanics of base pair unzipping in the DNA duplex.

    Science.gov (United States)

    Volkov, Sergey N; Paramonova, Ekaterina V; Yakubovich, Alexander V; Solov'yov, Andrey V

    2012-01-25

    All-atom molecular dynamics (MD) simulations of DNA duplex unzipping in a water environment were performed. The investigated DNA double helix consists of a Drew-Dickerson dodecamer sequence and a hairpin (AAG) attached to the end of the double-helix chain. The considered system is used to examine the process of DNA strand separation under the action of an external force. This process occurs in vivo and now is being intensively investigated in experiments with single molecules. The DNA dodecamer duplex is consequently unzipped pair by pair by means of the steered MD. The unzipping trajectories turn out to be similar for the duplex parts with G·C content and rather distinct for the parts with A·T content. It is shown that during the unzipping each pair experiences two types of motion: relatively quick rotation together with all the duplex and slower motion in the frame of the unzipping fork. In the course of opening, the complementary pair passes through several distinct states: (i) the closed state in the double helix, (ii) the metastable preopened state in the unzipping fork and (iii) the unbound state. The performed simulations show that water molecules participate in the stabilization of the metastable states of the preopened base pairs in the DNA unzipping fork.

  11. Free energy estimation of short DNA duplex hybridizations

    Directory of Open Access Journals (Sweden)

    Leger Serge

    2010-02-01

    Full Text Available Abstract Background Estimation of DNA duplex hybridization free energy is widely used for predicting cross-hybridizations in DNA computing and microarray experiments. A number of software programs based on different methods and parametrizations are available for the theoretical estimation of duplex free energies. However, significant differences in free energy values are sometimes observed among estimations obtained with various methods, thus being difficult to decide what value is the accurate one. Results We present in this study a quantitative comparison of the similarities and differences among four published DNA/DNA duplex free energy calculation methods and an extended Nearest-Neighbour Model for perfect matches based on triplet interactions. The comparison was performed on a benchmark data set with 695 pairs of short oligos that we collected and manually curated from 29 publications. Sequence lengths range from 4 to 30 nucleotides and span a large GC-content percentage range. For perfect matches, we propose an extension of the Nearest-Neighbour Model that matches or exceeds the performance of the existing ones, both in terms of correlations and root mean squared errors. The proposed model was trained on experimental data with temperature, sodium and sequence concentration characteristics that span a wide range of values, thus conferring the model a higher power of generalization when used for free energy estimations of DNA duplexes under non-standard experimental conditions. Conclusions Based on our preliminary results, we conclude that no statistically significant differences exist among free energy approximations obtained with 4 publicly available and widely used programs, when benchmarked against a collection of 695 pairs of short oligos collected and curated by the authors of this work based on 29 publications. The extended Nearest-Neighbour Model based on triplet interactions presented in this work is capable of performing accurate

  12. Smectic phase in suspensions of gapped DNA duplexes

    Science.gov (United States)

    Salamonczyk, Miroslaw; Zhang, Jing; Portale, Giuseppe; Zhu, Chenhui; Kentzinger, Emmanuel; Gleeson, James T.; Jakli, Antal; de Michele, Cristiano; Dhont, Jan K. G.; Sprunt, Samuel; Stiakakis, Emmanuel

    2016-11-01

    Smectic ordering in aqueous solutions of monodisperse stiff double-stranded DNA fragments is known not to occur, despite the fact that these systems exhibit both chiral nematic and columnar mesophases. Here, we show, unambiguously, that a smectic-A type of phase is formed by increasing the DNA's flexibility through the introduction of an unpaired single-stranded DNA spacer in the middle of each duplex. This is unusual for a lyotropic system, where flexibility typically destabilizes the smectic phase. We also report on simulations suggesting that the gapped duplexes (resembling chain-sticks) attain a folded conformation in the smectic layers, and argue that this layer structure, which we designate as smectic-fA phase, is thermodynamically stabilized by both entropic and energetic contributions to the system's free energy. Our results demonstrate that DNA as a building block offers an exquisitely tunable means to engineer a potentially rich assortment of lyotropic liquid crystals.

  13. Hydrogen bonding versus stacking stabilization by modified nucleobases incorporated in PNA. DNA duplexes

    DEFF Research Database (Denmark)

    Sen, Anjana; Nielsen, Peter E

    2009-01-01

    The effects of incorporation of the modified nucleobases, 2,6-diaminopurine (D) (substituting for adenine) and 7-chloro-1,8-naphthyridin-2-(1H)-one (bicyclic thymine, bT) (substituting for thymine), that stabilize PNA.DNA duplex formation by increasing hydrogen bonding and/or base pair stacking...

  14. Conserved guanine-guanine stacking in tetraplex and duplex DNA.

    Science.gov (United States)

    Kypr, J; Fialová, M; Chládková, J; Tůmová, M; Vorlícková, M

    2001-12-01

    Using a series of suitably chosen oligonucleotides, we demonstrate that the DNA duplex of d(CCCCGGGG) provides an almost identical CD spectrum as the parallel-stranded tetraplex of d(GGGG). The CD spectra are very sensitive to base stacking in DNA so that the above observation indicates that guanine-guanine stacking is essentially the same within the duplex of d(CCCCGGGG) and the tetraplex of d(GGGG). A very similar CD spectrum is also provided by the A-form of d(CCCCGGGG) induced by trifluoroethanol. These results reveal that guanine-guanine stacking is a structural invariant conserved in various nucleic acid conformers. The structural invariance is likely to cohere with evolution of the genetic molecules and be important for fundamental functions, e.g. initiation of transcription.

  15. Structural, Dynamical, and Electronic Transport Properties of Modified DNA Duplexes Containing Size-Expanded Nucleobases

    Energy Technology Data Exchange (ETDEWEB)

    Fuentes-Cabrera, Miguel A [ORNL; Orozco, Modesto [Institut de Recerca Biomedica, Parc Cientific de Barcelona, Barcelona, Spain; Luque, Javier [Universitat de Barcelona; Sumpter, Bobby G [ORNL; Blas, Jose [Universidad de Castilla-La Mancha; Ordejon, Pablo J [ORNL; Huertas, Oscar [Universitat de Barcelona; Tabares, Carolina [Universitat de Barcelona

    2011-01-01

    Among the distinct strategies proposed to expand the genetic alphabet, sizeexpanded nucleobases are promising for the development of modified DNA duplexes with improved biotechnological properties. In particular, duplexes built up by replacing canonical bases with the corresponding benzo-fused counterparts could be valuable as molecular nanowires. In this context, this study reports the results of classical molecular dynamics simulations carried out to examine the structural and dynamical features of size-expanded DNAs, including both hybrid duplexes containing mixed pairs of natural and benzo-fused bases (xDNA) and pure size-expanded (xxDNA) duplexes. Furthermore, the electronic structure of both natural and size-expanded duplexes is examined by means of density functional computations. The results confirm that the structural and flexibility properties of the canonical DNA are globally little affected by the presence of benzo-fused bases. Themost relevant differences are found in the enhanced size of the grooves, and the reduction in the twist. However, the analysis also reveals subtle structural effects related to the nature and sequence of benzo-fused bases in the duplex. On the other hand, electronic structure calculations performed for xxDNAs confirm the reduction in the HOMOLUMO gap predicted from the analysis of the natural bases and their size-expanded counterparts, which suggests that pure size-expanded DNAs can be good conductors. A more complex situation is found for xDNAs, where fluctuations in the electrostatic interaction between base pairs exerts a decisive influence on the modulation of the energy gap.

  16. The Effect of Small Cosolutes that Mimic Molecular Crowding Conditions on the Stability of Triplexes Involving Duplex DNA

    Directory of Open Access Journals (Sweden)

    Anna Aviñó

    2016-02-01

    Full Text Available Triplex stability is studied in crowding conditions using small cosolutes (ethanol, acetonitrile and dimethylsulfoxide by ultraviolet (UV, circular dichroism (CD and nuclear magnetic resonance (NMR spectroscopies. The results indicate that the triplex is formed preferentially when the triplex forming oligonucleotide (TFO is RNA. In addition, DNA triplexes (D:D·D are clearly less stable in cosolute solutions while the stability of the RNA triplexes (R:D·D is only slightly decreased. The kinetic of triplex formation with RNA-TFO is slower than with DNA-TFO and the thermal stability of the triplex is increased with the salt concentration in EtOH-water solutions. Accordingly, RNA could be considered a potential molecule to form a stable triplex for regulatory purposes in molecular crowding conditions.

  17. Hole Transport in A-form DNA/RNA Hybrid Duplexes

    Science.gov (United States)

    Wong, Jiun Ru; Shao, Fangwei

    2017-01-01

    DNA/RNA hybrid duplexes are prevalent in many cellular functions and are an attractive target form for electrochemical biosensing and electric nanodevice. However the electronic conductivities of DNA/RNA hybrid duplex remain relatively unexplored and limited further technological applications. Here cyclopropyl-modified deoxyribose- and ribose-adenosines were developed to explore hole transport (HT) in both DNA duplex and DNA/RNA hybrids by probing the transient hole occupancies on adenine tracts. HT yields through both B-form and A-form double helixes displayed similar shallow distance dependence, although the HT yields of DNA/RNA hybrid duplexes were lower than those of DNA duplexes. The lack of oscillatory periods and direction dependence in HT through both helixes implied efficient hole propagation can be achieved via the hole delocalization and coherent HT over adenine tracts, regardless of the structural variations.

  18. Minor Groove Binding between Norfloxacin and DNA Duplexes in Solution: A Molecular Dynamics Study

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Molecular dynamics were used to investigate the interaction between norfloxacin and DNA duplex. The results showed that norfloxacin was situated in the minor groove of DNA,binding to the TCGA region of d [ATATCGATAT] 2. Specific hydrogen bonds were formed between norfloxacin and guanine base of DNA during the 2 ns MD, which may be the reason for the preferentiality of quinolone antibacterial towards the guanine base of DNA duplex.

  19. Sequence-selective targeting of duplex DNA by peptide nucleic acids

    DEFF Research Database (Denmark)

    Nielsen, Peter E

    2010-01-01

    Sequence-selective gene targeting constitutes an attractive drug-discovery approach for genetic therapy, with the aim of reducing or enhancing the activity of specific genes at the transcriptional level, or as part of a methodology for targeted gene repair. The pseudopeptide DNA mimic peptide...... nucleic acid (PNA) can recognize duplex DNA with high sequence specificity and affinity in triplex, duplex and double-duplex invasive modes or non-invasive triplex modes. Novel PNA modification has improved the affinity for DNA recognition via duplex invasion, double-duplex invasion and triplex...... recognition considerably. Such modifications have also resulted in new approaches to targeted gene repair and sequence-selective double-strand cleavage of genomic DNA....

  20. Force measurements reveal how small binders perturb the dissociation mechanisms of DNA duplex sequences

    Science.gov (United States)

    Burmistrova, Anastasia; Fresch, Barbara; Sluysmans, Damien; de Pauw, Edwin; Remacle, Françoise; Duwez, Anne-Sophie

    2016-06-01

    The force-driven separation of double-stranded DNA is crucial to the accomplishment of cellular processes like genome transactions. Ligands binding to short DNA sequences can have a local stabilizing or destabilizing effect and thus severely affect these processes. Although the design of ligands that bind to specific sequences is a field of intense research with promising biomedical applications, so far, their effect on the force-induced strand separation has remained elusive. Here, by means of AFM-based single molecule force spectroscopy, we show the co-existence of two different mechanisms for the separation of a short DNA duplex and demonstrate how they are perturbed by small binders. With the support of Molecular Dynamics simulations, we evidence that above a critical pulling rate one of the dissociation pathways becomes dominant, with a dramatic effect on the rupture forces. Around the critical threshold, we observe a drop of the most probable rupture forces for ligand-stabilized duplexes. Our results offer a deep understanding of how a stable DNA-ligand complex behaves under force-driven strand separation.The force-driven separation of double-stranded DNA is crucial to the accomplishment of cellular processes like genome transactions. Ligands binding to short DNA sequences can have a local stabilizing or destabilizing effect and thus severely affect these processes. Although the design of ligands that bind to specific sequences is a field of intense research with promising biomedical applications, so far, their effect on the force-induced strand separation has remained elusive. Here, by means of AFM-based single molecule force spectroscopy, we show the co-existence of two different mechanisms for the separation of a short DNA duplex and demonstrate how they are perturbed by small binders. With the support of Molecular Dynamics simulations, we evidence that above a critical pulling rate one of the dissociation pathways becomes dominant, with a dramatic effect

  1. Simultaneous detection of bovine and porcine DNA in pharmaceutical gelatin capsules by duplex PCR assay for Halal authentication.

    Science.gov (United States)

    Nikzad, Jafar; Shahhosseini, Soraya; Tabarzad, Maryam; Nafissi-Varcheh, Nastaran; Torshabi, Maryam

    2017-02-14

    In the pharmaceutical industry, hard- and soft-shelled capsules are typically made from gelatin, commonly derived from bovine and porcine sources. To ensure that pharmaceutical products comply with halal regulations in Muslim countries (no porcine products allowed), development of a valid, reliable, quick, and most importantly, cost-effective tests are of utmost importance. We developed a species-specific duplex polymerase chain reaction (PCR) assay targeting 149 bp porcine and 271 bp bovine mitochondrial DNA (mtDNA) to simultaneously detect both porcine and bovine DNA (in one reaction at the same time) in gelatin. Some additional simplex PCR tests (targeting 126 bp bovine and 212 bp porcine mtDNA) and real-time PCR using a commercially available kit (for identification of porcine DNA) were used to verify the selectivity and sensitivity of our duplex PCR. After optimization of DNA extraction and PCR methods, hard/soft pharmaceutical gelatin capsules (containing drug) were tested for the presence of porcine and/or bovine DNA. Duplex PCR detected the presence of as little as 0.1% porcine DNA, which was more accurate than the commercially available kit. Of all gelatin capsules tested (n = 24), 50% contained porcine DNA (pure porcine gelatin alone or in combination with bovine gelatin). Duplex PCR presents an easy-to-follow, quick, low-cost and reliable method to simultaneously detect porcine and bovine DNAs (>100 bp) in minute amounts in highly processed gelatin-containing pharmaceutical products (with a 0.1% sensitivity for porcine DNA) which may be used for halal authentication. Simultaneous detection of porcine and bovine DNA in gelatin capsules by duplex PCR.

  2. Thermodynamics of HMGB1 interaction with duplex DNA.

    Science.gov (United States)

    Müller, S; Bianchi, M E; Knapp, S

    2001-08-28

    The high mobility group protein HMGB1 is a small, highly abundant protein that binds to DNA in a non-sequence-specific manner. HMGB1 consists of 2 DNA binding domains, the HMG boxes A and B, followed by a short basic region and a continuous stretch of 30 glutamate or aspartate residues. Isothermal titration calorimetry was used to characterize the binding of HMGB1 to the double-stranded model DNAs poly(dAdT).(dTdA) and poly(dGdC).(dCdG). To elucidate the contribution of the different structural motifs to DNA binding, calorimetric measurements were performed comparing the single boxes A and B, the two boxes plus or minus the basic sequence stretch (AB(bt) and AB), and the full-length HMGB1 protein. Thermodynamically, binding of HMGB1 and all truncated constructs to duplex DNA was characterized by a positive enthalpy change at 15 degrees C. From the slopes of the temperature dependence of the binding enthalpies, heat capacity changes of -0.129 +/- 0.02 and -0.105 +/- 0.05 kcal mol(-1) K(-1) were determined for box A and full-length HMGB1, respectively. Significant differences in the binding characteristics were observed using full-length HMGB1, suggesting an important role for the acid tail in modulating DNA binding. Moreover, full-length HMGB1 binds differently these two DNA templates: binding to poly(dAdT).(dTdA) was cooperative, had a larger apparent binding site size, and proceeded with a much larger unfavorable binding enthalpy than binding to poly(dGdC).(dCdG).

  3. DNA Duplexes with Hydrophobic Modifications Inhibit Fusion between HIV-1 and Cell Membranes

    OpenAIRE

    Xu, Liang; Cai, Lifeng; Chen, Xueliang; Jiang, Xifeng; Chong, Huihui; Zheng, Baohua; Wang, Kun; He, Junlin; Chen, Wei; ZHANG, Tao; Cheng, Maosheng; He, Yuxian; Liu, Keliang

    2013-01-01

    Discovery of new drugs for the treatment of AIDS typically possessing unique structures associated with novel mechanisms of action has been of great importance due to the quick drug-resistant mutations of HIV-1 strains. The work presented in this report describes a novel class of DNA duplex-based HIV-1 fusion inhibitors. Hydrophobic groups were introduced into a DNA duplex skeleton either at one end, at both ends, or in the middle. These modified DNA duplexes inhibited fusion between HIV-1 an...

  4. Enhanced H-bonding and pi-stacking in DNA: a potent duplex-stabilizing and mismatch sensing nucleobase analogue

    DEFF Research Database (Denmark)

    Lou, Chenguang; Dallmann, Andre; Marafini, Pietro;

    2014-01-01

    X-pyrene is a new nucleic acid duplex stabilizing cytosine analogue that combines enhanced pi-stacking, hydrogen bonding and electrostatic interactions to greatly increase the stability of bulged DNA duplexes and DNA/RNA hybrids. X-pyrene is highly selective for guanine as a partner and duplex...... analogue for use in a variety of biological applications....

  5. Force-Induced Rupture of a DNA Duplex: From Fundamentals to Force Sensors.

    Science.gov (United States)

    Mosayebi, Majid; Louis, Ard A; Doye, Jonathan P K; Ouldridge, Thomas E

    2015-12-22

    The rupture of double-stranded DNA under stress is a key process in biophysics and nanotechnology. In this article, we consider the shear-induced rupture of short DNA duplexes, a system that has been given new importance by recently designed force sensors and nanotechnological devices. We argue that rupture must be understood as an activated process, where the duplex state is metastable and the strands will separate in a finite time that depends on the duplex length and the force applied. Thus, the critical shearing force required to rupture a duplex depends strongly on the time scale of observation. We use simple models of DNA to show that this approach naturally captures the observed dependence of the force required to rupture a duplex within a given time on duplex length. In particular, this critical force is zero for the shortest duplexes, before rising sharply and then plateauing in the long length limit. The prevailing approach, based on identifying when the presence of each additional base pair within the duplex is thermodynamically unfavorable rather than allowing for metastability, does not predict a time-scale-dependent critical force and does not naturally incorporate a critical force of zero for the shortest duplexes. We demonstrate that our findings have important consequences for the behavior of a new force-sensing nanodevice, which operates in a mixed mode that interpolates between shearing and unzipping. At a fixed time scale and duplex length, the critical force exhibits a sigmoidal dependence on the fraction of the duplex that is subject to shearing.

  6. Hybridization accompanying FRET event in labeled natural nucleoside-unnatural nucleoside containing chimeric DNA duplexes.

    Science.gov (United States)

    Bag, Subhendu Sekhar; Das, Suman K; Pradhan, Manoj Kumar; Jana, Subhashis

    2016-09-01

    Förster resonance energy transfer (FRET) is a highly efficient strategy in illuminating the structures, structural changes and dynamics of DNA, proteins and other biomolecules and thus is being widely utilized in studying such phenomena, in designing molecular/biomolecular probes for monitoring the hybridization event of two single stranded DNA to form duplex, in gene detection and in many other sensory applications in chemistry, biology and material sciences. Moreover, FRET can give information about the positional status of chromophores within the associated biomolecules with much more accuracy than other methods can yield. Toward this end, we want to report here the ability of fluorescent unnatural nucleoside, triazolylphenanthrene ((TPhen)BDo) to show FRET interaction upon hybridization with fluorescently labeled natural nucleosides, (Per)U or (OxoPy)U or (Per)U, forming two stable chimeric DNA duplexes. The pairing selectivity and the thermal duplex stability of the chimeric duplexes are higher than any of the duplexes with natural nucleoside formed. The hybridization results in a Förster resonance energy transfer (FRET) from donor triazolylphenanthrene of (TPhen)BDo to acceptor oxopyrene of (OxoPy)U and/or to perylene chromophore of (Per)U, respectively, in two chimeric DNA duplexes. Therefore, we have established the FRET process in two chimeric DNA duplexes wherein a fluorescently labeled natural nucleoside ((OxoPy)U or (Per)U) paired against an unnatural nucleoside ((TPhen)BDo) without sacrificing the duplex stability and B-DNA conformation. The hybridization accompanying FRET event in these classes of interacting fluorophores is new. Moreover, there is no report of such designed system of chimeric DNA duplex. Our observed phenomenon and the design can potentially be exploited in designing more of such efficient FRET pairs for useful application in the detection and analysis of biomolecular interactions and in material science application. Copyright

  7. Influence of buffer species on the thermodynamics of short DNA duplex melting: sodium phosphate versus sodium cacodylate.

    Science.gov (United States)

    Alemayehu, Saba; Fish, Daniel J; Brewood, Greg P; Horne, M Todd; Manyanga, Fidelis; Dickman, Rebekah; Yates, Ian; Benight, Albert S

    2009-03-05

    Thermodynamic parameters of the melting transitions of 53 short duplex DNAs were experimentally evaluated by differential scanning calorimetry melting curve analysis. Solvents for the DNA solutions contained approximately 1 M Na+ and either 10 mM cacodylate or phosphate buffer. Thermodynamic parameters obtained in the two solvent environments were compared and quantitatively assessed. Thermodynamic stabilities (deltaG(o) (25 degrees C)) of the duplexes studied ranged from quite stable perfect match duplexes (approximately -30 kcal/mol) to relatively unstable mismatch duplexes (approximately -9 kcal/mol) and ranged in length from 18 to 22 basepairs. A significant difference in stability (average free energy difference of approximately 3 kcal/mol) was found for all duplexes melted in phosphate (greater stability) versus cacodylate buffers. Measured effects of buffer species appear to be relatively unaffected by duplex length or sequence content. The popular sets of published nearest-neighbor (n-n) stability parameters for Watson-Crick (w/c) and single-base mismatches were evaluated from melting studies performed in cacodylate buffer (SantaLucia and Hicks, Annu. Rev. Biophys. Biomol. Struct. 2004, 33, 415). Thus, when using these parameters to make predictions of sequence dependent stability of DNA oligomers in buffers other than cacodylate (e.g., phosphate) one should be mindful that in addition to sodium ion concentration, the type of buffer species also provides a minor but significant contribution to duplex stability. Such considerations could potentially influence results of sequence dependent analysis using published n-n parameters and impact results of thermodynamic calculations. Such calculations and analyses are typically employed in the design and interpretation of DNA multiplex hybridization experiments.

  8. Analysis of Structural Flexibility of Damaged DNA Using Thiol-Tethered Oligonucleotide Duplexes.

    Directory of Open Access Journals (Sweden)

    Masashi Fujita

    Full Text Available Bent structures are formed in DNA by the binding of small molecules or proteins. We developed a chemical method to detect bent DNA structures. Oligonucleotide duplexes in which two mercaptoalkyl groups were attached to the positions facing each other across the major groove were prepared. When the duplex contained the cisplatin adduct, which was proved to induce static helix bending, interstrand disulfide bond formation under an oxygen atmosphere was detected by HPLC analyses, but not in the non-adducted duplex, when the two thiol-tethered nucleosides were separated by six base pairs. When the insert was five and seven base pairs, the disulfide bond was formed and was not formed, respectively, regardless of the cisplatin adduct formation. The same reaction was observed in the duplexes containing an abasic site analog and the (6–4 photoproduct. Compared with the cisplatin case, the disulfide bond formation was slower in these duplexes, but the reaction rate was nearly independent of the linker length. These results indicate that dynamic structural changes of the abasic site- and (6–4 photoproduct-containing duplexes could be detected by our method. It is strongly suggested that the UV-damaged DNA-binding protein, which specifically binds these duplexes and functions at the first step of global-genome nucleotide excision repair, recognizes the easily bendable nature of damaged DNA.

  9. Reversed assembly of dyes in an RNA duplex compared with those in DNA.

    Science.gov (United States)

    Fujii, Taiga; Urushihara, Masaaki; Kashida, Hiromu; Ito, Hiroshi; Liang, Xingguo; Yagi-Utsumi, Maho; Kato, Koichi; Asanuma, Hiroyuki

    2012-10-15

    We prepared reversed dye clusters by hybridizing two RNA oligomers, each of which tethered dyes (Methyl Red, 4'-methylthioazobenzene, and thiazole orange) on D-threoninols (threoninol nucleotides) at the center of their strands. NMR spectroscopic analyses revealed that two dyes from each strand were axially stacked in an antiparallel manner to each other in the duplex, and were located adjacent to the 3'-side of a natural nucleobase. Interestingly, this positional relationship of the dyes was completely the opposite of that assembled in DNA that we reported previously: dyes in DNA were located adjacent to the 5'-side of a natural nucleobase. This observation was also consistent with the circular dichroism of dimerized dyes in which the Cotton effect of the dyes (i.e., the winding properties of two dyes) was inverted in RNA relative to that in DNA. Further spectroscopic analyses revealed that clustering of the dyes on RNA duplexes induced distinct hypsochromicity and narrowing of the band, thus demonstrating that the dyes were axially stacked (i.e., H-aggregates) even on an A-type helix. On the basis of these results, we also prepared heterodimers of a fluorophore (thiazole orange) and quencher (Methyl Red) in an RNA duplex. Fluorescence from thiazole orange was found to be strongly quenched by Methyl Red due to the excitonic interaction, so that the ratio of fluorescent intensities of the RNA-thiazole orange conjugate with and without its complementary strand carrying a quencher became as high as 27. We believe that these RNA-dye conjugates are potentially useful probes for real-time monitoring of RNA interference (RNAi) mechanisms.

  10. Detection of Hepatitis B Virus DNA by Duplex Scorpion Primer-based PCR Assay

    Institute of Scientific and Technical Information of China (English)

    KONG De-Ming孔德明; SHEN Han-Xi沈含熙; MI Huai-Feng宓怀风

    2004-01-01

    The application of a new fiuorogenic probe-based PCR assay (PCR duplex scorpion primer assay) to the detection of Hepatitis B virus (HBV) DNA in human sera was described. Duplex scorpion primer is a modified variant of duplex Amplifluor, and the incorporation of a PCR stopper between probe and primer sequences improve the detection specificity and sensitivity. Combined with PCR amplification, this probe can give unambiguous positive results for the reactions initiated with more than 20 HBV molecules. In addition, the particular unimolecular probing mechanism of this probe makes the use of short target-specific probe sequence possible, which will render this probe applicable in some specific systems.

  11. Ultrasensitive Detection of RNA and DNA Viruses Simultaneously Using Duplex UNDP-PCR Assay

    Science.gov (United States)

    Wang, Zengguo; Zhang, Xiujuan; Zhao, Xiaomin; Du, Qian; Chang, Lingling; Tong, Dewen

    2015-01-01

    Mixed infection of multiple viruses is common in modern intensive pig rearing. However, there are no methods available to detect DNA and RNA viruses in the same reaction system in preclinical level. In this study, we aimed to develop a duplex ultrasensitive nanoparticle DNA probe-based PCR assay (duplex UNDP-PCR) that was able to simultaneously detect DNA and RNA viruses in the same reaction system. PCV2 and TGEV are selected as representatives of the two different types of viruses. PCV2 DNA and TGEV RNA were simultaneously released from the serum sample by boiling with lysis buffer, then magnetic beads and gold nanoparticles coated with single and/or duplex specific probes for TGEV and PCV2 were added to form a sandwich-like complex with nucleic acids released from viruses. After magnetic separation, DNA barcodes specific for PCV2 and TGEV were eluted using DTT and characterized by specific PCR assay for specific DNA barcodes subsequently. The duplex UNDP-PCR showed similar sensitivity as that of single UNDP-PCR and was able to detect 20 copies each of PCV2 and TGEV in the serum, showing approximately 250-fold more sensitivity than conventional duplex PCR/RT-PCR assays. No cross-reaction was observed with other viruses. The positive detection rate of single MMPs- and duplex MMPs-based duplex UNDP-PCR was identical, with 29.6% for PCV2, 9.3% for TGEV and 3.7% for PCV2 and TGEV mixed infection. This duplex UNDP-PCR assay could detect TGEV (RNA virus) and PCV2 (DNA virus) from large-scale serum samples simultaneously without the need for DNA/RNA extraction, purification and reverse transcription of RNA, and showed a significantly increased positive detection rate for PCV2 (29%) and TGEV (11.7%) preclinical infection than conventional duplex PCR/RT-PCR. Therefore, the established duplex UNDP-PCR is a rapid and economical detection method, exhibiting high sensitivity, specificity and reproducibility. PMID:26544710

  12. Ultrasensitive Detection of RNA and DNA Viruses Simultaneously Using Duplex UNDP-PCR Assay.

    Directory of Open Access Journals (Sweden)

    Yong Huang

    Full Text Available Mixed infection of multiple viruses is common in modern intensive pig rearing. However, there are no methods available to detect DNA and RNA viruses in the same reaction system in preclinical level. In this study, we aimed to develop a duplex ultrasensitive nanoparticle DNA probe-based PCR assay (duplex UNDP-PCR that was able to simultaneously detect DNA and RNA viruses in the same reaction system. PCV2 and TGEV are selected as representatives of the two different types of viruses. PCV2 DNA and TGEV RNA were simultaneously released from the serum sample by boiling with lysis buffer, then magnetic beads and gold nanoparticles coated with single and/or duplex specific probes for TGEV and PCV2 were added to form a sandwich-like complex with nucleic acids released from viruses. After magnetic separation, DNA barcodes specific for PCV2 and TGEV were eluted using DTT and characterized by specific PCR assay for specific DNA barcodes subsequently. The duplex UNDP-PCR showed similar sensitivity as that of single UNDP-PCR and was able to detect 20 copies each of PCV2 and TGEV in the serum, showing approximately 250-fold more sensitivity than conventional duplex PCR/RT-PCR assays. No cross-reaction was observed with other viruses. The positive detection rate of single MMPs- and duplex MMPs-based duplex UNDP-PCR was identical, with 29.6% for PCV2, 9.3% for TGEV and 3.7% for PCV2 and TGEV mixed infection. This duplex UNDP-PCR assay could detect TGEV (RNA virus and PCV2 (DNA virus from large-scale serum samples simultaneously without the need for DNA/RNA extraction, purification and reverse transcription of RNA, and showed a significantly increased positive detection rate for PCV2 (29% and TGEV (11.7% preclinical infection than conventional duplex PCR/RT-PCR. Therefore, the established duplex UNDP-PCR is a rapid and economical detection method, exhibiting high sensitivity, specificity and reproducibility.

  13. Effect of LNA- and OMeN-modified oligonucleotide probes on the stability and discrimination of mismatched base pairs of duplexes

    Indian Academy of Sciences (India)

    Ying Yan; Jing Yan; Xianyu Piao; Tianbiao Zhang; Yifu Guan

    2012-06-01

    Locked nucleic acid (LNA) and 2′--methyl nucleotide (OMeN) are the most extensively studied nucleotide analogues. Although both LNA and OMeN are characterized by the C3′-endo sugar pucker conformation, which is dominant in A-form DNA and RNA nucleotides, they demonstrate different binding behaviours. Previous studies have focused attention on their properties of duplex stabilities, hybridization kinetics and resistance against nuclease digestion; however, their ability to discriminate mismatched hybridizations has been explored much less. In this study, LNA- and OMeN-modified oligonucleotide probes have been prepared and their effects on the DNA duplex stability have been examined: LNA modifications can enhance the duplex stability, whereas OMeN modifications reduce the duplex stability. Next, we studied how the LNA:DNA and OMeN:DNA mismatches reduced the duplex stability. Melting temperature measurement showed that different LNA:DNA or OMeN:DNA mismatches indeed influence the duplex stability differently. LNA purines can discriminate LNA:DNA mismatches more effectively than LNA pyrimidines as well as DNA nucleotides. Furthermore, we designed five LNA- and five OMeN-modified oligonucleotide probes to simulate realistic situations where target–probe duplexes contain a complementary LNA:DNA or OMeN:DNA base pairs and a DNA:DNA mismatch simultaneously. The measured collective effect showed that the duplex stability was enhanced by the complementary LNA:DNA base pair but decreased by the DNA:DNA mismatch in a position-dependent manner regardless of the chemical identity and position of the complementary LNA:DNA base pair. On the other hand, the OMeN-modified probes also showed that the duplex stability was reduced by both the OMeN modification and the OMeN:DNA mismatch in a position-dependent manner.

  14. A novel form of intercalation involving four DNA duplexes in an acridine-4-carboxamide complex of d(CGTACG)2

    Science.gov (United States)

    Adams, Adrienne; Guss, J. Mitchell; Collyer, Charles A.; Denny, William A.; Wakelin, Laurence P. G.

    2000-01-01

    The structures of the complexes formed between 9-amino-[N-(2-dimethyl-amino)butyl]acridine-4-carboxamide and d(CG5BrUACG)2 and d(CGTACG)2 have been solved by X-ray crystallography using MAD phasing methodology and refined to a resolution of 1.6 Å. The complexes crystallised in space group C222. An asymmetric unit in the brominated complex comprises two strands of DNA, one disordered drug molecule, two cobalt (II) ions and 19 water molecules (31 in the native complex). Asymmetric units in the native complex also contain a sodium ion. The structures exhibit novel features not previously observed in crystals of DNA/drug complexes. The DNA helices stack in continuous columns with their central 4 bp adopting a B-like motif. However, despite being a palindromic sequence, the terminal GC base pairs engage in quite different interactions. At one end of the duplex there is a CpG dinucleotide overlap modified by ligand intercalation and terminal cytosine exchange between symmetry-related duplexes. A novel intercalation complex is formed involving four DNA duplexes, four ligand molecules and two pairs of base tetrads. The other end of the DNA is frayed with the terminal guanine lying in the minor groove of the next duplex in the column. The structure is stabilised by guanine N7/cobalt (II) coordination. We discuss our findings with respect to the effects of packing forces on DNA crystal structure, and the potential effects of intercalating agents on biochemical processes involving DNA quadruplexes and strand exchanges. NDB accession numbers: DD0032 (brominated) and DD0033 (native). PMID:11058124

  15. DNA Duplexes with Hydrophobic Modifications Inhibit Fusion between HIV-1 and Cell Membranes

    Science.gov (United States)

    Xu, Liang; Cai, Lifeng; Chen, Xueliang; Jiang, Xifeng; Chong, Huihui; Zheng, Baohua; Wang, Kun; He, Junlin; Chen, Wei; Zhang, Tao; Cheng, Maosheng; He, Yuxian

    2013-01-01

    Discovery of new drugs for the treatment of AIDS typically possessing unique structures associated with novel mechanisms of action has been of great importance due to the quick drug-resistant mutations of HIV-1 strains. The work presented in this report describes a novel class of DNA duplex-based HIV-1 fusion inhibitors. Hydrophobic groups were introduced into a DNA duplex skeleton either at one end, at both ends, or in the middle. These modified DNA duplexes inhibited fusion between HIV-1 and human cell membranes at micro- or submicromolar concentrations. Respective inhibitors adopted an aptamer pattern instead of a base-pairing interaction pattern. Structure-activity relationship studies of the respective DNA duplexes showed that the rigid and negatively charged DNA skeletons, in addition to the presence of hydrophobic groups, were crucial to the anti-HIV-1 activity of these compounds. A fluorescent resonance energy transfer (FRET)-based inhibitory assay showed that these duplex inhibitors interacted with the primary pocket in the gp41 N-terminal heptad repeat (NHR) instead of interacting with the lipid bilayers. PMID:23896466

  16. Fluorescence studies on the interaction of ethidium bromide with duplex, triplex and quadruplex DNA structures

    Institute of Scientific and Technical Information of China (English)

    孙雪光; 曹恩华; 何裕建; 秦静芬

    1999-01-01

    Under different conditions, oligonucleotides can form several alternative DNA structures such as duplex, triplex and quadruplex. All these structures can interact with ethidium bromide (EB) and make its fluorescence intensity change. The fluorescence spectra and other related parameters provided by static fluorescence techniques showed that the interaction mechanisms between EB and these structures were not always the same. Among them, B type duplex and triplex DNA adopt an intercalative mode when binding to the EB, which has a relatively high efficiency of energy transfer and the fluorescence of EB cannot be quenched easily. While for the parallel duplex DNA, the interaction mode is an outside binding in which energy transfer can hardly happen and its fluorescence intensity as well as Stern-Volmer constant is almost the same to the free EB. For the quadruplex, the binding mechanism to EB is more complex. Results from the energy transfer and quenching studies indicate that the two interaction modes note

  17. DNA-duplex linker for AFM-SELEX of DNA aptamer against human serum albumin.

    Science.gov (United States)

    Takenaka, Musashi; Okumura, Yuzo; Amino, Tomokazu; Miyachi, Yusuke; Ogino, Chiaki; Kondo, Akihiko

    2017-02-15

    DNA-duplex interactions in thymines and adenins are used as a linker for the novel methodology of Atomic Force Microscope-Systematic Evolution of Ligands by EXpotential enrichment (AFM-SELEX). This study used the hydrogen bonds in 10 mer of both thymines (T10) and adenines (A10). Initially, the interactive force in T10-A10 was measured by AFM, which returned an average interactive force of approximately 350pN. Based on this result, DNA aptamers against human serum albumin could be selected in the 4th round, and 15 different clones could be sequenced. The lowest dissociation constant of the selected aptamer was identified via surface plasmon resonance, and it proved to be identical to that of the commercial aptamer. Therefore, specific hydrogen bonds in DNA can be useful linkers for AFM-SELEX. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Binding and NMR structural studies on indoloquinoline-oligonucleotide conjugates targeting duplex DNA.

    Science.gov (United States)

    Eick, Andrea; Riechert-Krause, Fanny; Weisz, Klaus

    2012-06-20

    An 11-phenyl-indolo[3,2-b]quinoline (PIQ) was tethered through an aminoalkyl linker to the 5'-end of four pyrimidine oligonucleotides with T/C scrambled sequences at their two 5'-terminal positions. Binding to different double-helical DNA targets formed parallel triple helices with a PIQ-mediated stabilization that strongly depends on pH and the terminal base triad at the 5'-triplex-duplex junction. The most effective stabilization was observed with a TAT triplet at the 5'-junction under low pH conditions, pointing to a protonated ligand with a high triplex binding affinity and unfavorable charge repulsions in the case of a terminal C(+)GC triplet at the junction. The latter preference of the PIQ ligand for TAT over CGC is alleviated yet still preserved at higher pH. Intercalation of PIQ at the 5'-triplex-duplex junction as suggested by the triplex melting experiments was confirmed by homonuclear and heteronuclear NMR structural studies on a specifically isotope-labeled triplex. The NMR analysis revealed two coexisting species that only differ by a 180° rotation of the indoloquinoline within the intercalation pocket. NOE-derived molecular models indicate extensive stacking interactions of the indoloquinoline moiety with the TAT base triplet and CG base pair at the junction and a phenyl substituent that is positioned in the major groove and oriented almost perpendicular to the plane of the indoloquinoline.

  19. Exploring the Interactions of the Dietary Plant Flavonoids Fisetin and Naringenin with G-Quadruplex and Duplex DNA, Showing Contrasting Binding Behavior: Spectroscopic and Molecular Modeling Approaches.

    Science.gov (United States)

    Bhattacharjee, Snehasish; Chakraborty, Sandipan; Sengupta, Pradeep K; Bhowmik, Sudipta

    2016-09-01

    Guanine-rich sequences have the propensity to fold into a four-stranded DNA structure known as a G-quadruplex (G4). G4 forming sequences are abundant in the promoter region of several oncogenes and become a key target for anticancer drug binding. Here we have studied the interactions of two structurally similar dietary plant flavonoids fisetin and naringenin with G4 as well as double stranded (duplex) DNA by using different spectroscopic and modeling techniques. Our study demonstrates the differential binding ability of the two flavonoids with G4 and duplex DNA. Fisetin more strongly interacts with parallel G4 structure than duplex DNA, whereas naringenin shows stronger binding affinity to duplex rather than G4 DNA. Molecular docking results also corroborate our spectroscopic results, and it was found that both of the ligands are stacked externally in the G4 DNA structure. C-ring planarity of the flavonoid structure appears to be a crucial factor for preferential G4 DNA recognition of flavonoids. The goal of this study is to explore the critical effects of small differences in the structure of closely similar chemical classes of such small molecules (flavonoids) which lead to the contrasting binding properties with the two different forms of DNA. The resulting insights may be expected to facilitate the designing of the highly selective G4 DNA binders based on flavonoid scaffolds.

  20. Syntheses of DNA Duplexes That Contain a N4C-alkyl-N4C Interstrand Cross-Link

    Science.gov (United States)

    Miller, Paul S.

    2011-01-01

    A simple procedure is described for preparing short DNA duplexes that contain a single N4C-alkyl-N4C interstrand cross-link. The synthesis is carried out on an automated DNA synthesizer using standard phosphoramidite chemistry. The cross-link is introduced during the synthesis of the duplex. The method can be used to prepare mg quantities of cross-linked duplexes suitable for physical studies and for the preparation of larger DNA molecules that can be used as substrates to study DNA repair in whole cell extracts and in living cells in culture. PMID:21400705

  1. Conformational influence of the ribose 2'-hydroxyl group: crystal structures of DNA-RNA chimeric duplexes

    Science.gov (United States)

    Egli, M.; Usman, N.; Rich, A.

    1993-01-01

    We have crystallized three double-helical DNA-RNA chimeric duplexes and determined their structures by X-ray crystallography at resolutions between 2 and 2.25 A. The two self-complementary duplexes [r(G)d(CGTATACGC)]2 and [d(GCGT)r(A)d(TACGC)]2, as well as the Okazaki fragment d(GGGTATACGC).r(GCG)d(TATACCC), were found to adopt A-type conformations. The crystal structures are non-isomorphous, and the crystallographic environments for the three chimeras are different. A number of intramolecular interactions of the ribose 2'-hydroxyl groups contribute to the stabilization of the A-conformation. Hydrogen bonds between 2'-hydroxyls and 5'-oxygens or phosphate oxygens, in addition to the previously observed hydrogen bonds to 1'-oxygens of adjacent riboses and deoxyriboses, are observed in the DNA-RNA chimeric duplexes. The crystalline chimeric duplexes do not show a transition between the DNA A- and B-conformations. CD spectra suggest that the Okazaki fragment assumes an A-conformation in solution as well. In this molecule the three RNA residues may therefore lock the complete decamer in the A-conformation. Crystals of an all-DNA strand with the same sequence as the self-complementary chimeras show a morphology which is different from those of the chimera crystals. Moreover, the oligonucleotide does not match any of the sequence characteristics of DNAs usually adopting the A-conformation in the crystalline state (e.g., octamers with short alternating stretches of purines and pyrimidines). In DNA-RNA chimeric duplexes, it is therefore possible that a single RNA residue can drive the conformational equilibrium toward the A-conformation.

  2. A simple Duplex-PCR to evaluate the DNA quality of anthropological and forensic samples prior short tandem repeat typing.

    Science.gov (United States)

    von Wurmb-Schwark, Nicole; Schwark, Thorsten; Harbeck, Michaela; Oehmichen, Manfred

    2004-04-01

    Typing of DNA from ancient or otherwise highly degraded material, e.g. formalin fixed tissues, can be difficult, time consuming and costly. Very often, genetic typing is not possible at all. We present an inexpensive and easy to use Duplex-PCR that amplifies a 164 bp fragment specific for nuclear DNA together with a 260 bp mitochondrial DNA fragment and that can be employed as a pretest prior to short tandem repeat (STR) typing. All together, we analyzed DNA from 20 ancient bones, 20 formalin fixed tissues and 20 other forensic samples in different concentrations. Each sample that failed in the presented Duplex-amplification was also negative for STR typing, while samples that showed strong and clear signals in the Duplex-PCR led to reproducible genetic profiles using the multiplex kits AmpFLSTR Identifiler and Powerplex ES. The Duplex-PCR worked as a reliable indicator of DNA quality in the sample.

  3. Thermal equivalence of DNA duplexes without calculation of melting temperature

    Science.gov (United States)

    Weber, Gerald; Haslam, Niall; Whiteford, Nava; Prügel-Bennett, Adam; Essex, Jonathan W.; Neylon, Cameron

    2006-01-01

    The common key to nearly all processes involving DNA is the hybridization and melting of the double helix: from transmission of genetic information and RNA transcription, to polymerase chain reaction and DNA microarray analysis, DNA mechanical nanodevices and DNA computing. Selecting DNA sequences with similar melting temperatures is essential for many applications in biotechnology. We show that instead of calculating these temperatures, a single parameter can be derived from a statistical-mechanics model that conveniently represents the thermodynamic equivalence of DNA sequences. This parameter is shown to order experimental melting temperatures correctly, is much more readily obtained than the melting temperature, and is easier to handle than the numerous parameters of empirical regression models.

  4. Microfluidic thermodynamics of the shift in thermal stability of DNA duplex in a microchannel laminar flow.

    Science.gov (United States)

    Yamashita, Kenichi; Miyazaki, Masaya; Yamaguchi, Yoshiko; Nakamura, Hiroyuki; Maeda, Hideaki

    2007-06-01

    This paper reports the shift in thermal stability of DNA duplex and its thermodynamics spectroscopically, caused by stretching and orientation of DNA strands in a microchannel laminar flow. For direct spectroscopic measurement of the microchannel, we prepared an in-house temperature-controllable microchannel-type flow cell. The melting curves of DNA oligomers in a microchannel laminar flow were measured. For DNA oligomers with more than 10 base pairs, the melting curve shifted to the high-temperature side with higher flow speed. However, for 8-base-pair DNA oligomers, a change in the melting profile was not observed in batchwise and microchannel flows. We undertook microfluidic thermodynamic analysis to elucidate details of the shift in thermal stability of the DNA duplex in a microchannel laminar flow. Enthalpy-entropy compensation is applicable to the microfluidic thermal stability shift. We studied the relationships between the enthalpy-entropy compensation and DNA strand length or flow speed. Results showed that the enthalpy-entropy compensation was influenced by both DNA strand length and flow speed, and the penalties of enthalpy were 2-12% greater than the benefits of entropy.

  5. Experimental mapping of DNA duplex shape enabled by global lineshape analyses of a nucleotide-independent nitroxide probe.

    Science.gov (United States)

    Ding, Yuan; Zhang, Xiaojun; Tham, Kenneth W; Qin, Peter Z

    2014-10-01

    Sequence-dependent variation in structure and dynamics of a DNA duplex, collectively referred to as 'DNA shape', critically impacts interactions between DNA and proteins. Here, a method based on the technique of site-directed spin labeling was developed to experimentally map shapes of two DNA duplexes that contain response elements of the p53 tumor suppressor. An R5a nitroxide spin label, which was covalently attached at a specific phosphate group, was scanned consecutively through the DNA duplex. X-band continuous-wave electron paramagnetic resonance spectroscopy was used to monitor rotational motions of R5a, which report on DNA structure and dynamics at the labeling site. An approach based on Pearson's coefficient analysis was developed to collectively examine the degree of similarity among the ensemble of R5a spectra. The resulting Pearson's coefficients were used to generate maps representing variation of R5a mobility along the DNA duplex. The R5a mobility maps were found to correlate with maps of certain DNA helical parameters, and were capable of revealing similarity and deviation in the shape of the two closely related DNA duplexes. Collectively, the R5a probe and the Pearson's coefficient-based lineshape analysis scheme yielded a generalizable method for examining sequence-dependent DNA shapes.

  6. Sequence-specific intercalating agents: intercalation at specific sequences on duplex DNA via major groove recognition by oligonucleotide-intercalator conjugates.

    Science.gov (United States)

    Sun, J S; François, J C; Montenay-Garestier, T; Saison-Behmoaras, T; Roig, V; Thuong, N T; Hélène, C

    1989-01-01

    An acridine derivative was covalently linked to the 5' end of a homopyrimidine oligonucleotide. Specific binding to a homopurine-homopyrimidine sequence of duplex DNA was demonstrated by spectroscopic studies (absorption and fluorescence) and by "footprinting" experiments with a copper phenanthroline chelate used as an artificial nuclease. A hypochromism and a red shift of the acridine absorption were observed. Triple-helix formation was also accompanied by a hypochromism in the ultraviolet range. The fluorescence of the acridine ring was quenched by a stacking interaction with a G.C base pair adjacent to the homopurine-homopyrimidine target sequence. The intercalating agent strongly stabilized the complex formed by the oligopyrimidine with its target duplex sequence. Cytosine methylation further increased the stability of the complexes. Footprinting studies revealed that the oligopyrimidine binds in a parallel orientation with respect to the homopurine-containing strand of the duplex. The intercalated acridine extended by 2 base pairs the region of the duplex protected by the oligopyrimidine against degradation by the nuclease activity of the copper phenanthroline chelate. Random intercalation of the acridine ring was lost due to the repulsive effect of the negatively charged oligonucleotide tail. Intercalation occurred only at those double-stranded sequences where the homopyrimidine oligonucleotide recognized the major groove of duplex DNA. Images PMID:2594761

  7. Effect of heat treatment on corrosion behavior of duplex coatings

    Directory of Open Access Journals (Sweden)

    K. Raghu Ram Mohan Reddy

    2017-01-01

    Full Text Available In the present paper, duplex WC-Co/NiCrAlY coating is coated onto Ti6Al4V substrate and vacuum heat treatment is employed to investigate the corrosion behavior of heat treated samples as well as Ti6Al4V substrate for comparison. In this duplex coating system, High Velocity Oxy Fuel (HVOF process is used to deposit NiCrAlY interlayer with a constant thickness of 200 μm and WC-Co ceramic top layer with varying thickness of 250 μm, 350 μm and 450 μm deposited by Detonation Spray (DS process. Different heat treatment temperatures (600–1150 °C were employed for the coated samples to study the microstructure and the effect on corrosion resistance of the duplex coatings. Potentiodynamic polarization tests were carried to investigate the corrosion performance of duplex coated heat treated samples and the substrate in Ringer’s solution at 37 °C and prepared the pH to 5.7. The microstructure upon corrosion after heat treatment was characterized by SEM analysis to understand the corrosion behavior. The results disclosed that at all heat treatment temperatures, all the coated samples exhibited better corrosion resistance than the base substrate. However, during 950 °C and 1150 °C heat treatment temperatures, it was observed highest corrosion potential than 600 °C and 800 °C. The 350 μm thickness, coated sample exhibited highest corrosion resistance compared to other two coated samples and the substrate at all heat treatment temperatures.

  8. Dynamical transition of water in the grooves of DNA duplex at low temperature.

    Science.gov (United States)

    Biswal, Debasmita; Jana, Biman; Pal, Subrata; Bagchi, Biman

    2009-04-02

    At low temperature (below its freezing/melting temperature), liquid water under confinement is known to exhibit anomalous dynamical features. Here we study structure and dynamics of water in the grooves of a long DNA duplex using molecular dynamics simulations with TIP5P potential at low temperature. We find signatures of a dynamical transition in both translational and orientational dynamics of water molecules in both the major and the minor grooves of a DNA duplex. The transition occurs at a slightly higher temperature (T(GL) approximately 255 K) than the temperature at which the bulk water is found to undergo a dynamical transition, which for the TIP5P potential is at 247 K. Groove water, however, exhibits markedly different temperature dependence of its properties from the bulk. Entropy calculations reveal that the minor groove water is ordered even at room temperature, and the transition at T approximately 255 K can be characterized as a strong-to-strong dynamical transition. Confinement of water in the grooves of DNA favors the formation of a low density four-coordinated state (as a consequence of enthalpy-entropy balance) that makes the liquid-liquid transition stronger. The low temperature water is characterized by pronounced tetrahedral order, as manifested in the sharp rise near 109 degrees in the O-O-O angle distribution. We find that the Adams-Gibbs relation between configurational entropy and translational diffusion holds quite well when the two quantities are plotted together in a master plot for different region of aqueous DNA duplex (bulk, major, and minor grooves) at different temperatures. The activation energy for the transfer of water molecules between different regions of DNA is found to be weakly dependent on temperature.

  9. Neomycin-neomycin dimer: an all-carbohydrate scaffold with high affinity for AT-rich DNA duplexes.

    Science.gov (United States)

    Kumar, Sunil; Xue, Liang; Arya, Dev P

    2011-05-18

    A dimeric neomycin-neomycin conjugate 3 with a flexible linker, 2,2'-(ethylenedioxy)bis(ethylamine), has been synthesized and characterized. Dimer 3 can selectively bind to AT-rich DNA duplexes with high affinity. Biophysical studies have been performed between 3 and different nucleic acids with varying base composition and conformation by using ITC (isothermal calorimetry), CD (circular dichroism), FID (fluorescent intercalator displacement), and UV (ultraviolet) thermal denaturation experiments. A few conclusions can be drawn from this study: (1) FID assay with 3 and polynucleotides demonstrates the preference of 3 toward AT-rich sequences over GC-rich sequences. (2) FID assay and UV thermal denaturation experiments show that 3 has a higher affinity for the poly(dA)·poly(dT) DNA duplex than for the poly(dA)·2poly(dT) DNA triplex. Contrary to neomycin, 3 destabilizes poly(dA)·2poly(dT) triplex but stabilizes poly(dA)·poly(dT) duplex, suggesting the major groove as the binding site. (3) UV thermal denaturation studies and ITC experiments show that 3 stabilizes continuous AT-tract DNA better than DNA duplexes with alternating AT bases. (4) CD and FID titration studies show a DNA binding site size of 10-12 base pairs/drug, depending upon the structure/sequence of the duplex for AT-rich DNA duplexes. (5) FID and ITC titration between 3 and an intramolecular DNA duplex [d(5'-A(12)-x-T(12)-3'), x = hexaethylene glycol linker] results in a binding stoichiometry of 1:1 with a binding constant ∼10(8) M(-1) at 100 mM KCl. (6) FID assay using 3 and 512 hairpin DNA sequences that vary in their AT base content and placement also show a higher binding selectivity of 3 toward continuous AT-rich than toward DNA duplexes with alternate AT base pairs. (7) Salt-dependent studies indicate the formation of three ion pairs during binding of the DNA duplex d[5'-A(12)-x-T(12)-3'] and 3. (8) ITC-derived binding constants between 3 and DNA duplexes have the following order: AT

  10. DUPLEX: A molecular mechanics program in torsion angle space for computing structures of DNA and RNA

    Energy Technology Data Exchange (ETDEWEB)

    Hingerty, B.E.

    1992-07-01

    DUPLEX produces energy minimized structures of DNA and RNA of any base sequence for single and double strands. The smallest subunits are deoxydinucleoside monophosphates, and up to 12 residues, single or double stranded can be treated. In addition, it can incorporate NMR derived interproton distances an constraints in the minimizations. Both upper and lower bounds for these distances can be specified. The program has been designed to run on a UNICOS Cray supercomputer, but should run, albeit slowly, on a laboratory computer such as a VAX or a workstation.

  11. Atomistic details of the molecular recognition of DNA-RNA hybrid duplex by ribonuclease H enzyme

    Indian Academy of Sciences (India)

    Gorle Suresh; U Deva Priyakumar

    2015-10-01

    Bacillus halodurans (ℎ) ribonuclease H (RNase H) belongs to the nucleotidyl-transferase (NT) superfamily and is a prototypical member of a large family of enzymes that use two-metal ion (Mg2+ or Mn2+) catalysis to cleave nucleic acids. Long timescale molecular dynamics simulations have been performed on the ℎRNase H-DNA-RNA hybrid complex and the respective monomers to understand the recognition mechanism, conformational preorganization, active site dynamics and energetics involved in the complex formation. Several structural and energetic analyses were performed and significant structural changes are observed in enzyme and hybrid duplex during complex formation. Hybrid molecule binding to RNase H enzyme leads to conformational changes in the DNA strand. The ability of the DNA strand in the hybrid duplex to sample conformations corresponding to typical A- and B-type nucleic acids and the characteristic minor groove width-seem to be crucial for efficient binding. Sugar moieties in certain positions interacting with the protein structure undergo notable conformational transitions. The water coordination and arrangement around the metal ions in active site region are quite stable, suggesting their important role in enzymatic catalysis. Details of key interactions found at the interface of enzyme-nucleic acid complex that are responsible for its stability are discussed.

  12. Targeting duplex DNA with chimeric α,β-triplex-forming oligonucleotides

    Science.gov (United States)

    Kolganova, N. A.; Shchyolkina, A. K.; Chudinov, A. V.; Zasedatelev, A. S.; Florentiev, V. L.; Timofeev, E. N.

    2012-01-01

    Triplex-directed DNA recognition is strictly limited by polypurine sequences. In an attempt to address this problem with synthetic biology tools, we designed a panel of short chimeric α,β-triplex-forming oligonucleotides (TFOs) and studied their interaction with fluorescently labelled duplex hairpins using various techniques. The hybridization of hairpin with an array of chimeric probes suggests that recognition of double-stranded DNA follows complicated rules combining reversed Hoogsteen and non-canonical homologous hydrogen bonding. In the presence of magnesium ions, chimeric TFOs are able to form highly stable α,β-triplexes, as indicated by native gel-electrophoresis, on-array thermal denaturation and fluorescence-quenching experiments. CD spectra of chimeric triplexes exhibited features typically observed for anti-parallel purine triplexes with a GA or GT third strand. The high potential of chimeric α,β-TFOs in targeting double-stranded DNA was demonstrated in the EcoRI endonuclease protection assay. In this paper, we report, for the first time, the recognition of base pair inversions in a duplex by chimeric TFOs containing α-thymidine and α-deoxyguanosine. PMID:22641847

  13. Specific DNA duplex formation at an artificial lipid bilayer: fluorescence microscopy after Sybr Green I staining

    Directory of Open Access Journals (Sweden)

    Emma Werz

    2014-10-01

    Full Text Available The article describes the immobilization of different probe oligonucleotides (4, 7, 10 carrying each a racemic mixture of 2,3-bis(hexadecyloxypropan-1-ol (1a at the 5’-terminus on a stable artificial lipid bilayer composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC. The bilayer separates two compartments (cis/trans channel of an optical transparent microfluidic sample carrier with perfusion capabilities. Injection of unlabeled target DNA sequences (6, 8, or 9, differing in sequence and length, leads in the case of complementarity to the formation of stable DNA duplexes at the bilayer surface. This could be verified by Sybr Green I double strand staining, followed by incubation periods and thorough perfusions, and was visualized by single molecule fluorescence spectroscopy and microscopy. The different bilayer-immobilized complexes consisting of various DNA duplexes and the fluorescent dye were studied with respect to the kinetics of their formation as well as to their stability against perfusion.

  14. Thermodynamic and kinetic characterization of duplex formation between 2'-O, 4'-C-methylene-modified oligoribonucleotides, DNA and RNA

    DEFF Research Database (Denmark)

    Christensen, Ulla

    2007-01-01

    2'-O,4'-C-methylene-linked ribonucleotide derivatives, named LNA (locked nucleic acid) and BNA (bridged nucleic acid) are nucleic acid analogoues that have shown high-affinity recognition of DNA and RNA, and the employment of LNA oligomers for antisense activity, gene regulation and nucleic acid...... the strength of duplexes formed with the complementary DNA and RNA....

  15. Brownian dynamics simulations of sequence-dependent duplex denaturation in dynamically superhelical DNA

    Science.gov (United States)

    Mielke, Steven P.; Grønbech-Jensen, Niels; Krishnan, V. V.; Fink, William H.; Benham, Craig J.

    2005-09-01

    The topological state of DNA in vivo is dynamically regulated by a number of processes that involve interactions with bound proteins. In one such process, the tracking of RNA polymerase along the double helix during transcription, restriction of rotational motion of the polymerase and associated structures, generates waves of overtwist downstream and undertwist upstream from the site of transcription. The resulting superhelical stress is often sufficient to drive double-stranded DNA into a denatured state at locations such as promoters and origins of replication, where sequence-specific duplex opening is a prerequisite for biological function. In this way, transcription and other events that actively supercoil the DNA provide a mechanism for dynamically coupling genetic activity with regulatory and other cellular processes. Although computer modeling has provided insight into the equilibrium dynamics of DNA supercoiling, to date no model has appeared for simulating sequence-dependent DNA strand separation under the nonequilibrium conditions imposed by the dynamic introduction of torsional stress. Here, we introduce such a model and present results from an initial set of computer simulations in which the sequences of dynamically superhelical, 147 base pair DNA circles were systematically altered in order to probe the accuracy with which the model can predict location, extent, and time of stress-induced duplex denaturation. The results agree both with well-tested statistical mechanical calculations and with available experimental information. Additionally, we find that sites susceptible to denaturation show a propensity for localizing to supercoil apices, suggesting that base sequence determines locations of strand separation not only through the energetics of interstrand interactions, but also by influencing the geometry of supercoiling.

  16. EFFECT OF INTERMETALLIC PHASES ON CORROSION BEHAVIOR AND MECHANICAL PROPERTIES OF DUPLEX STAINLESS STEEL AND SUPER-DUPLEX STAINLESS STEEL

    Directory of Open Access Journals (Sweden)

    Prabhu Paulraj

    2015-08-01

    Full Text Available Duplex Stainless Steels (DSS and Super Duplex Stainless Steel (SDSS have excellent integration of mechanical and corrosion properties. However, the formation of intermetallic phases is a major problem in their usage. The mechanical and corrosion properties are deteriorated due to the presence of intermetallic phases. These phases are induced during welding, prolonged exposure to high temperatures, and improper heat treatments. The main emphasis of this review article is on intermetallic phases and their effects on corrosion and mechanical properties. First the effect of various alloying elements on DSS and SDSS has been discussed followed by formation of various intermetallic phases. The intermetallic phases affect impact toughness and corrosion resistance significantly. Their deleterious effect on weldments has also been reviewed.

  17. Comparative analysis of inosine-substituted duplex DNA by circular dichroism and X-ray crystallography.

    Science.gov (United States)

    Peters, Justin P; Kowal, Ewa A; Pallan, Pradeep S; Egli, Martin; Maher, L James

    2017-09-04

    Leveraging structural biology tools, we report the results of experiments seeking to determine if the different mechanical properties of DNA polymers with base analog substitutions can be attributed, at least in part, to induced changes from classical B-form DNA. The underlying hypothesis is that different inherent bending and twisting flexibilities may characterize non-canonical B-DNA, so that it is inappropriate to interpret mechanical changes caused by base analog substitution as resulting simply from 'electrostatic' or 'base stacking' influences without considering the larger context of altered helical geometry. Circular dichroism spectra of inosine-substituted oligonucleotides and longer base-substituted DNAs in solution indicated non-canonical helical conformations, with the degree of deviation from a standard B-form geometry depending on the number of I⋅C pairs. X-ray diffraction of a highly inosine-substituted DNA decamer crystal (eight I⋅C and two A⋅T pairs) revealed an A-tract-like conformation with a uniformly narrow minor groove, reduced helical rise, and the majority of sugars adopting a C1'-exo (southeastern) conformation. This contrasts with the standard B-DNA geometry with C2'-endo sugar puckers (south conformation). In contrast, the crystal structure of a decamer with only four I⋅C pairs has a geometry similar to that of the reference duplex with eight G⋅C and two A⋅T pairs. The unique crystal geometry of the inosine-rich duplex is noteworthy given its unusual CD signature in solution and the altered mechanical properties of some inosine-containing DNAs.

  18. Enhanced thermal stability and mismatch discrimination of mutation-carrying DNA duplexes and their kinetic and thermodynamic properties in microchannel laminar flow.

    Science.gov (United States)

    Nagata, Maria Portia B; Yamashita, Kenichi; Miyazaki, Masaya; Nakamura, Hiroyuki; Maeda, Hideaki

    2009-07-01

    This article reports the enhancement of thermal stability involving normal duplex and mutation-carrying DNA duplexes in microchannel laminar flow. The application of an in-house temperature-controllable microchannel-type flow cell is demonstrated for improved discrimination of mismatch base pairs such as A-G and T-G that are difficult to distinguish due to the rather small thermal destabilizations. Enhancement in thermal stability is reflected by an increased thermal melting temperature achieved in microchannel laminar flow as compared with batch reactions. To examine the kinetics and thermodynamics of duplex-coil equilibrium of DNA oligomers, denaturation-renaturation hysteresis curves were measured. The influence of microchannel laminar flow on DNA base mismatch analysis was described from the kinetic and thermodynamic perspectives. An increasing trend was observed for association rate constant as flow rate increased. In contrast, an apparent decrease in dissociation rate constant was observed with increasing flow rate. The magnitudes of the activation energies of dissociation were nearly constant for both the batch and microchannel laminar flow systems at all flow rates. In contrast, the magnitudes of activation energies of association decreased as flow rate increased. These results clearly show how microchannel laminar flow induces change in reaction rate by effecting change in activation energy. We anticipate, therefore, that this approach based on microchannel laminar flow system holds great promise for improved mismatch discrimination in DNA analyses, particularly on single-base-pair mismatch, by pronouncedly enhancing thermal stability.

  19. Sequence-specific binding and cleavage of duplex DNA by a radioiodinated, intercalator-linked, triplex-forming oligonucleotide

    Energy Technology Data Exchange (ETDEWEB)

    Orson, Frank M.; McShan, W. Michael; Kinsey, Berma M

    1996-05-01

    Applications of oligodeoxynucleotides to modulate gene expression have been the subject of much recent research. We have sought to develop a method to permanently inactivate a gene, or potentially kill cells containing abnormal genes. In this report, we show that a DNA intercalator conjugated to a triplex-forming oligonucleotide can be labeled with an Auger electron emitting radioisotope, can cleave its duplex DNA target, and can specifically bind the target sequence contained in a total of 10 kilobases of irrelevant DNA.

  20. Visualizing Transient Watson-Crick Like Mispairs in DNA and RNA Duplexes

    Science.gov (United States)

    Kimsey, Isaac J.; Petzold, Katja; Sathyamoorthy, Bharathwaj; Stein, Zachary W.; Al-Hashimi, Hashim M.

    2015-01-01

    Rare tautomeric and anionic nucleobases are believed to play fundamental biological roles but their prevalence and functional importance has remained elusive because they exist transiently, in low-abundance, and involve subtle movements of protons that are difficult to visualize. Using NMR relaxation dispersion, we show that wobble dG•dT and rG•rU mispairs in DNA and RNA duplexes exist in dynamic equilibrium with short-lived, low-populated Watson-Crick like mispairs that are stabilized by rare enolic or anionic bases. These mispairs can evade Watson-Crick fidelity checkpoints and form with probabilities (10−3-10−5) that strongly imply a universal role in replication and translation errors. Our results indicate that rare tautomeric and anionic bases are widespread in nucleic acids, expanding their structural and functional complexity beyond that attainable with canonical bases. PMID:25762137

  1. Susceptibility to superhelically driven DNA duplex destabilization: a highly conserved property of yeast replication origins.

    Directory of Open Access Journals (Sweden)

    2005-06-01

    Full Text Available Strand separation is obligatory for several DNA functions, including replication. However, local DNA properties such as A+T content or thermodynamic stability alone do not determine the susceptibility to this transition in vivo. Rather, superhelical stresses provide long-range coupling among the transition behaviors of all base pairs within a topologically constrained domain. We have developed methods to analyze superhelically induced duplex destabilization (SIDD in genomic DNA that take into account both this long-range stress-induced coupling and sequence-dependent local thermodynamic stability. Here we apply this approach to examine the SIDD properties of 39 experimentally well-characterized autonomously replicating DNA sequences (ARS elements, which function as replication origins in the yeast Saccharomyces cerevisiae. We find that these ARS elements have a strikingly increased susceptibility to SIDD relative to their surrounding sequences. On average, these ARS elements require 4.78 kcal/mol less free energy to separate than do their immediately surrounding sequences, making them more than 2,000 times easier to open. Statistical analysis shows that the probability of this strong an association between SIDD sites and ARS elements arising by chance is approximately 4 x 10. This local enhancement of the propensity to separate to single strands under superhelical stress has obvious implications for origin function. SIDD properties also could be used, in conjunction with other known origin attributes, to identify putative replication origins in yeast, and possibly in other metazoan genomes.

  2. A population-wide applicable HLA-DQ2 and DQ8 genotyping using DNA from dried blood spots and duplex allele-specific qPCR amplification.

    Science.gov (United States)

    Aguayo-Patrón, Sandra; Beltrán-Sauceda, Lizbeth; Calderón de la Barca, Ana María

    2016-11-01

    Genotyping of HLA-DQ2 and DQ8 haplotypes is important for diagnosis or for screening of early risk detection of celiac disease or type 1 diabetes. Usually, venous blood DNA extraction and expensive and time consuming amplification are used, that hinder population-wide studies. We assayed a friendly HLA-DQ2 and DQ8 genotyping procedure using a combination of DNA from dried blood spot (DBS) and duplex allele-specific qPCR amplification using SYBR Green. DNA was extracted using home-made buffers and compared to an extraction commercial kit. Duplex reactions by qPCR were designed using each Tm allele amplicon for reference samples (positive HLA-DQ2 or DQ8) with allele-specific primers. DBS samples from 558 children (7.99 ± 2.47 y) were collected. The DNA final yield obtained by the home-made extractive procedure was higher than from the commercial kit (1.11 ± 0.56 vs 0.23 ± 0.14 μg), while the quality was similar for both DNA samples. There was concordance in the amplification profiles for DNA samples obtained with both methods. All of four alleles from DQ2 and DQ8 haplotypes were accurately identified in duplex reactions. By using DBS samples and DNA extraction home-made procedure, the costs were reduced by 60%. The whole procedure is cost-effective for HLA-DQ2 and DQ8 genotyping.

  3. Fluorescent C-linked C8-aryl-guanine probe for distinguishing syn from anti structures in duplex DNA.

    Science.gov (United States)

    Manderville, Richard A; Omumi, Alireza; Rankin née Schlitt, Katherine M; Wilson, Katie A; Millen, Andrea L; Wetmore, Stacey D

    2012-06-18

    The synthesis and optical properties of the carbon (C)-linked C(8)-(2"-benzo[b]thienyl)-2'-deoxyguanosine ((Bth)dG), which acts as a fluorescent reporter of syn versus anti glycosidic conformations in duplex DNA, are described. In the syn-conformation, the probe stabilizes a G:G mismatch, emits at ∼385 nm (excitation ∼285 nm), and shows an induced circular dichroism (ICD) signal at ∼320 nm. Molecular dynamics (MD) simulations predict a wedge (W)-conformation for the mismatched duplex with the C(8)-benzo[b]thienyl moiety residing in the minor groove. In contrast, the probe destabilizes the duplex when base paired with its normal pyrimidine partner C. With flanking purine bases, a major groove B-type duplex is favored with (Bth)dG present in the anti-conformation emitting at ∼413 nm (excitation ∼326 nm) and no ICD signal. However, with flanking pyrimidine bases, (Bth)dG adopts the syn-conformation when base paired with C, and MD simulations predict a base-displaced stacked (S)-conformation, with the opposing C flipped out of the helix. The different duplex (B-, S-, and W-) conformers formed upon incorporation of (Bth)dG are known to play a critical role in the biological activity of N-linked C8-dG adducts formed by arylamine carcinogens. Bulky environment-sensitive fluorescent C(8)-dG adducts that mimic the duplex structures formed by carcinogens may be useful in luminescence-based DNA polymerase assays.

  4. Identification of pork contamination in meatball using genetic marker mitochondrial DNA cytochrome b gene by duplex-PCR

    Science.gov (United States)

    Novianty, E.; Kartikasari, L. R.; Lee, J. H.; Cahyadi, M.

    2017-04-01

    Meat based food products have a big opportunity to mix and adulterated with other meats. Muslim communities are prohibited to consume pork-containing product or other pig derivatives in food. Therefore, the high sensitivity, fast, cheap and accurate approach is needed to detect pig contamination in raw meat and meat-processed product such as meatball. The aim of this study was to identify pork contamination in meatball using genetic marker of mitochondrial DNA cytochrome b gene by duplex-PCR. Samples were prepared and designed by following the proportions 0, 1, 5, 10, 25% of pork in meatballs, respectively. The DNA genome was extracted from meatballs and polymerase chain reaction (PCR) was performed using species specific primer to isolate mt-DNA cytochrome b gene. The results showed that the DNA genome was successfully isolated from pork, beef, and contaminated meatballs. Furthermore, 2% agarose gels was able to visualize of duplex-PCR to identify pork contamination in meatballs up to very small proportion (1%). It can be concluded that duplex-PCR of mt-DNA cytochrome b gene was very sensitive to identify pork contamination in meatball with the presence of specific 398 bp DNA band.

  5. Thioredoxin suppresses microscopic hopping of T7 DNA polymerase on duplex DNA

    NARCIS (Netherlands)

    Etson, Candice M.; Hamdan, Samir M.; Richardson, Charles C.; Oijen, Antoine M. van; Richardson, Charles C.

    2010-01-01

    The DNA polymerases involved in DNA replication achieve high processivity of nucleotide incorporation by forming a complex with processivity factors. A model system for replicative DNA polymerases, the bacteriophage T7 DNA polymerase (gp5), encoded by gene 5, forms a tight, 1:1 complex with

  6. Molecular design of synthetic benzimidazoles for the switchover of the duplex to G-quadruplex DNA recognition.

    Science.gov (United States)

    Maji, Basudeb; Bhattacharya, Santanu

    2013-01-01

    Benzimidazole derivatives are well known for their antibacterial, antiviral, anticonvulsant, antihistaminic, anthelmintic and antidepressant activities. Benzimidazole's unique base-selective DNA recognition property has been studied widely. However, most of the early benzimidazole systems have been targeted towards the binding of duplex DNA. Here we have shown the evolution and progress of the design and synthesis of new benzimidazole systems towards selective recognition of the double-stranded DNA first. Then in order to achieve selective recognition of the G-quadruplex DNA and utilize their potential as future anti-cancer drug candidates, we have demonstrated their selective cytotoxicity towards the cancer cells and potent telomerase inhibition ability.

  7. Affinity modification of EcoRII DNA methyltransferase by the dialdehyde-substituted DNA duplexes: mapping the enzyme region that interacts with DNA.

    Science.gov (United States)

    Gritsenko, Oksana M; Koudan, Elizaveta V; Mikhailov, Sergey N; Ermolinsky, Boris S; Van Aerschot, Arthur; Herdewijn, Piet; Gromova, Elizaveta S

    2002-01-01

    Affinity modification of EcoRII DNA methyltransferase (M x EcoRII) by DNA duplexes containing oxidized 2'-O-beta-D-ribofuranosylcytidine (Crib*) or 1-(beta-D-galactopyranosyl)thymine (Tgal*) residues was performed. Cross-linking yields do not change irrespective of whether active Crib* replaces an outer or an inner (target) deoxycytidine within the EcoRII recognition site. Chemical hydrolysis of M x EcoRII in the covalent cross-linked complex with the Tgal*-substituted DNA indicates the region Gly268-Met391 of the methylase that is likely to interact with the DNA sugar-phosphate backbone. Both specific and non-specific DNA interact with the same M x EcoRII region. Our results support the theoretically predicted DNA binding region of M x EcoRII.

  8. Synthesis of C-5, C-2' and C-4'-neomycin-conjugated triplex forming oligonucleotides and their affinity to DNA-duplexes.

    Science.gov (United States)

    Tähtinen, Ville; Granqvist, Lotta; Virta, Pasi

    2015-08-01

    Neomycin-conjugated homopyrimidine oligo 2'-deoxyribonucleotides have been synthesized on a solid phase and their potential as triplex forming oligonucleotides (TFOs) with DNA-duplexes has been studied. For the synthesis of the conjugates, C-5, C-2' and C-4'-tethered alkyne-modified nucleoside derivatives were used as an integral part of the standard automated oligonucleotide chain elongation. An azide-derived neomycin was then conjugated to the incorporated terminal alkynes by Cu(I)-catalyzed 1,3-dipolar cycloaddition (the click chemistry). Concentrated ammonia released the desired conjugates in acceptable purity and yields. The site of conjugation was expectedly important for the Hoogsteen-face recognition: C-5-conjugation showed a notable positive effect, whereas the influence of the C-2' and C-4'-modification remained marginal. In addition to conventional characterization methods (UV- and CD-spectroscopy), (19)F NMR spectroscopy was applied for the monitoring of triplex/duplex/single strand-conversions.

  9. Residual dipolar coupling constants and structure determination of large DNA duplexes

    Energy Technology Data Exchange (ETDEWEB)

    Mauffret, Olivier; Tevanian, Georges; Fermandjian, Serge [Institut Gustave-Roussy, Departement de Biologie et Pharmacologie Structurales (France)], E-mail: sfermand@igr.fr

    2002-12-15

    Several NMR works have shown that long-range information provided by residual dipolar couplings (RDCs) significantly improve the global structure definition of RNAs and DNAs. Most of these are based on the use of a large set of RDCs, the collect of which requires samples labeled with {sup 13}C, {sup 15}N, and sometimes, {sup 2}H. Here, we carried out torsion-angle dynamics simulations on a non-self complementary DNA fragment of 17 base-pairs, d(GGAAAATATCTAGCAGT).(ACTGCTAGAGATTTTCC). This reproduces the U5 LTR distal end of the HIV-1 cDNA that contains the enzyme integrase binding site. Simulations aimed at evaluating the impact of RDCs on the structure definition of long oligonucleotides, were performed in incorporating (i) nOe-distances at both < 4.5 A and < 5 A; (ii) a small set of {sup 13}C-{sup 1}H RDCs, easily detectable at the natural abundance, and (iii) a larger set of RDCs only accessible through the {sup 13}C labeling of DNAs. Agreement between a target structure and a simulated structure was measured in terms of precision and accuracy. Results allowed to define conditions in which accurate DNA structures can be determined. We confirmed the strong impact of RDCs on the structure determination, and, above all, we found that a small set of RDC constraints (ca. 50) detectable at the natural abundance is sufficient to accurately derive the global and local DNA duplex structures when used in conjunction with nOe-distances < 5 A.

  10. Robust IR-based detection of stable and fractionally populated G-C(+) and A-T Hoogsteen base pairs in duplex DNA.

    Science.gov (United States)

    Stelling, Allison L; Xu, Yu; Zhou, Huiqing; Choi, Seung H; Clay, Mary C; Merriman, Dawn K; Al-Hashimi, Hashim M

    2017-06-01

    Noncanonical G-C(+) and A-T Hoogsteen base pairs can form in duplex DNA and play roles in recognition, damage repair, and replication. Identifying Hoogsteen base pairs in DNA duplexes remains challenging due to difficulties in resolving syn versus antipurine bases with X-ray crystallography; and size limitations and line broadening can make them difficult to characterize by NMR spectroscopy. Here, we show how infrared (IR) spectroscopy can identify G-C(+) and A-T Hoogsteen base pairs in duplex DNA across a range of different structural contexts. The utility of IR-based detection of Hoogsteen base pairs is demonstrated by characterizing the first example of adjacent A-T and G-C(+) Hoogsteen base pairs in a DNA duplex where severe broadening complicates detection with NMR. © 2017 Federation of European Biochemical Societies.

  11. Electroporation and microinjection successfully deliver single-stranded and duplex DNA into live cells as detected by FRET measurements.

    Directory of Open Access Journals (Sweden)

    Rosemary A Bamford

    Full Text Available Förster resonance energy transfer (FRET technology relies on the close proximity of two compatible fluorophores for energy transfer. Tagged (Cy3 and Cy5 complementary DNA strands forming a stable duplex and a doubly-tagged single strand were shown to demonstrate FRET outside of a cellular environment. FRET was also observed after transfecting these DNA strands into fixed and live cells using methods such as microinjection and electroporation, but not when using lipid based transfection reagents, unless in the presence of the endosomal acidification inhibitor bafilomycin. Avoiding the endocytosis pathway is essential for efficient delivery of intact DNA probes into cells.

  12. DNA Binding and Recognition of a CC Mismatch in a DNA Duplex by Water-Soluble Peptidocalix[4]arenes: Synthesis and Applications.

    Science.gov (United States)

    Alavijeh, Nahid S; Zadmard, Reza; Balalaie, Saeed; Alavijeh, Mohammad S; Soltani, Nima

    2016-10-07

    Water-soluble peptidocalix[4]arenes were synthesized by the introduction of arginine-rich narrow groove-binding residues at lower rims through solid-phase synthesis. The study of binding of these water-soluble bidentate ligands to well-matched and mismatched DNA duplexes by fluorescent titrations, ethidium bromide (EB) displacement assays, DNA-melting experiments, and circular dichroism (CD) analysis revealed a sequence-dependent groove-binding mechanism.

  13. DNA duplex length and salt concentration dependence of enthalpy-entropy compensation parameters for DNA melting.

    Science.gov (United States)

    Starikov, E B; Nordén, Bengt

    2009-08-20

    Systematical differential calorimetry experiments on DNA oligomers with different lengths and placed in water solutions with various added salt concentrations may, in principle, unravel important information about the structure and dynamics of the DNA and their water-counterion surrounding. With this in mind, to reinterpret the most recent results of calorimetric experiments on DNA oligomers of such a kind, the recent enthalpy-entropy compensation theory has been used. It is demonstrated that the application of the latter could enable direct estimation of thermodynamic parameters of the microphase transitions connected to the changes in DNA dynamical regimes versus the length of the biopolymers and the ionic strengths of their water solutions, and this calls for much more systematical experimental and theoretical studies in this field.

  14. DNA Duplex Length and Salt Concentration Dependence of Enthalpy−Entropy Compensation Parameters for DNA Melting

    KAUST Repository

    Starikov, E. B.

    2009-08-20

    Systematical differential calorimetry experiments on DNA oligomers with different lengths and placed in water solutions with various added salt concentrations may, in principle, unravel important information about the structure and dynamics of the DNA and their water-counterion surrounding. With this in mind, to reinterpret the most recent results of calorimetric experiments on DNA oligomers of such a kind, the recent enthalpy-entropy compensation theory has been used. It is demonstrated that the application of the latter could enable direct estimation of thermodynamic parameters of the microphase transitions connected to the changes in DNA dynamical regimes versus the length of the biopolymers and the ionic strengths of their water solutions, and this calls for much more systematical experimental and theoretical studies in this field. © 2009 American Chemical Society.

  15. Direct measurement of sequence-dependent transition path times and conformational diffusion in DNA duplex formation.

    Science.gov (United States)

    Neupane, Krishna; Wang, Feng; Woodside, Michael T

    2017-02-07

    The conformational diffusion coefficient, D, sets the timescale for microscopic structural changes during folding transitions in biomolecules like nucleic acids and proteins. D encodes significant information about the folding dynamics such as the roughness of the energy landscape governing the folding and the level of internal friction in the molecule, but it is challenging to measure. The most sensitive measure of D is the time required to cross the energy barrier that dominates folding kinetics, known as the transition path time. To investigate the sequence dependence of D in DNA duplex formation, we measured individual transition paths from equilibrium folding trajectories of single DNA hairpins held under tension in high-resolution optical tweezers. Studying hairpins with the same helix length but with G:C base-pair content varying from 0 to 100%, we determined both the average time to cross the transition paths, τtp, and the distribution of individual transit times, PTP(t). We then estimated D from both τtp and PTP(t) from theories assuming one-dimensional diffusive motion over a harmonic barrier. τtp decreased roughly linearly with the G:C content of the hairpin helix, being 50% longer for hairpins with only A:T base pairs than for those with only G:C base pairs. Conversely, D increased linearly with helix G:C content, roughly doubling as the G:C content increased from 0 to 100%. These results reveal that G:C base pairs form faster than A:T base pairs because of faster conformational diffusion, possibly reflecting lower torsional barriers, and demonstrate the power of transition path measurements for elucidating the microscopic determinants of folding.

  16. Cloned plasmid DNA fragments as calibrators for controlling GMOs: different real-time duplex quantitative PCR methods.

    Science.gov (United States)

    Taverniers, Isabel; Van Bockstaele, Erik; De Loose, Marc

    2004-03-01

    Analytical real-time PCR technology is a powerful tool for implementation of the GMO labeling regulations enforced in the EU. The quality of analytical measurement data obtained by quantitative real-time PCR depends on the correct use of calibrator and reference materials (RMs). For GMO methods of analysis, the choice of appropriate RMs is currently under debate. So far, genomic DNA solutions from certified reference materials (CRMs) are most often used as calibrators for GMO quantification by means of real-time PCR. However, due to some intrinsic features of these CRMs, errors may be expected in the estimations of DNA sequence quantities. In this paper, two new real-time PCR methods are presented for Roundup Ready soybean, in which two types of plasmid DNA fragments are used as calibrators. Single-target plasmids (STPs) diluted in a background of genomic DNA were used in the first method. Multiple-target plasmids (MTPs) containing both sequences in one molecule were used as calibrators for the second method. Both methods simultaneously detect a promoter 35S sequence as GMO-specific target and a lectin gene sequence as endogenous reference target in a duplex PCR. For the estimation of relative GMO percentages both "delta C(T)" and "standard curve" approaches are tested. Delta C(T) methods are based on direct comparison of measured C(T) values of both the GMO-specific target and the endogenous target. Standard curve methods measure absolute amounts of target copies or haploid genome equivalents. A duplex delta C(T) method with STP calibrators performed at least as well as a similar method with genomic DNA calibrators from commercial CRMs. Besides this, high quality results were obtained with a standard curve method using MTP calibrators. This paper demonstrates that plasmid DNA molecules containing either one or multiple target sequences form perfect alternative calibrators for GMO quantification and are especially suitable for duplex PCR reactions.

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  18. Photo-controlled binding of MutS to photo-caged DNA duplexes incorporating 4-O-(2-nitrobenzyl) or 4-O-[2-(2-nitrophenyl)propyl]thymidine.

    Science.gov (United States)

    Seio, Kohji; Ohno, Yurie; Ohno, Kentaro; Takeshita, Leo; Kanamori, Takashi; Masaki, Yoshiaki; Sekine, Mitsuo

    2016-10-01

    Mismatch binding protein MutS binding to bulge structure in DNA duplexes was controlled by UV irradiation. 4-O-(2-Nitrobenzyl)thymidine or 4-O-[2-(2-nitrophenyl)propyl]thymidine was incorporated into DNA duplexes a bulged position. The MutS did not bind to the caged DNA duplexes but bound after removing the 2-nitrobenzyl or 2-(2-nitrophenyl)propyl group by photo-irradiation. By using photo-caged DNA duplex, we revealed that binding of MutS to the uncaged DNA downstream of the T7 RNA promoter weakly inhibited transcription by T7 RNA polymerase.

  19. Prototropic equilibria in DNA containing one-electron oxidized GC: intra-duplex vs. duplex to solvent deprotonation.

    Science.gov (United States)

    Adhikary, Amitava; Kumar, Anil; Munafo, Shawn A; Khanduri, Deepti; Sevilla, Michael D

    2010-01-01

    By use of ESR and UV-vis spectral studies, this work identifies the protonation states of one-electron oxidized G:C (viz. G˙+:C, G(N1–H)˙:C(+H+), G(N1–H)˙:C, and G(N2-H)˙:C) in a DNA oligomer d[TGCGCGCA]2. Benchmark ESR and UV-vis spectra from one electron oxidized 1-Me-dGuo are employed to analyze the spectral data obtained in one-electron oxidized d[TGCGCGCA]2 at various pHs. At pH ≥7, the initial site of deprotonation of one-electron oxidized d[TGCGCGCA]2 to the surrounding solvent is found to be at N1 forming G(N1–H)˙:C at 155 K. However, upon annealing to 175 K, the site of deprotonation to the solvent shifts to an equilibrium mixture of G(N1–H)˙:C and G(N2–H)˙:C. For the first time, the presence of G(N2–H)˙:C in a ds DNA-oligomer is shown to be easily distinguished from the other prototropic forms, owing to its readily observable nitrogen hyperfine coupling (Azz(N2) = 16 G). In addition, for the oligomer in H2O, an additional 8 G N2–H proton HFCC is found. This ESR identification is supported by a UV-vis absorption at 630 nm which is characteristic for G(N2–H)˙ in model compounds and oligomers. We find that the extent of photo-conversion to the C1′ sugar radical (C1′˙) in the one-electron oxidized d[TGCGCGCA]2 allows for a clear distinction among the various G:C protonation states which can not be easily distinguished by ESR or UV-vis spectroscopies with this order for the extent of photo-conversion: G˙+:C > G(N1–H)˙:C(+H+) ≫ G(N1–H)˙:C. We propose that it is the G˙+:C form that undergoes deprotonation at the sugar and this requires reprotonation of G within the lifetime of exited state

  20. Prototropic Equilibria in DNA Containing One-electron Oxidized GC: Intra-duplex vs. Duplex to Solvent Deprotonation

    Science.gov (United States)

    Adhikary, Amitava; Kumar, Anil; Munafo, Shawn A.; Khanduri, Deepti; Sevilla, Michael D.

    2015-01-01

    By use of ESR and UV-vis spectral studies, this work identifies the protonation states of one-electron oxidized G:C (viz. G•+:C, G(N1-H)•:C(+H+), G(N1-H)•:C, and G(N2-H)•:C) in a DNA oligomer d[TGCGCGCA]2. Benchmark ESR and UV-vis spectra from one electron oxidized 1-Me-dGuo are employed to analyze the spectral data obtained in one-electron oxidized d[TGCGCGCA]2 at various pHs. At pH ≥7, the initial site of deprotonation of one-electron oxidized d[TGCGCGCA]2 to the surrounding solvent is found to be at N1 forming G(N1-H)•:C at 155 K. However, upon annealing to 175 K, the site of deprotonation to the solvent shifts to an equilibrium mixture of G(N1-H)•:C and G(N2-H)•:C. For the first time, the presence of G(N2-H)•:C in a ds DNA-oligomer is shown to be easily distinguished from the other prototropic forms, owing to its readily observable nitrogen hyperfine coupling (Azz(N2)= 16 G). In addition, for the oligomer in H2O, an additional 8 G N2-H proton HFCC is found. This ESR identification is supported by a UV-vis absorption at 630 nm which is characteristic for G(N2-H)• in model compounds and oligomers. We find that the extent of photo-conversion to the C1′ sugar radical (C1′•) in the one-electron oxidized d[TGCGCGCA]2 allows for a clear distinction among the various G:C protonation states which can not be easily distinguished by ESR or UV-vis spectroscopies with this order for the extent of photo-conversion: G•+:C > G(N1-H)•:C(+H+) >> G(N1-H)•:C. We propose that it is the G•+:C form that undergoes deprotonation at the sugar and this requires reprotonation of G within the lifetime of exited state. PMID:21491657

  1. Calculating Distortions of Short DNA Duplexes with Base Pairing Between an Oxidatively Damaged Guanine and a Guanine

    Directory of Open Access Journals (Sweden)

    Masayo Suzuki

    2014-07-01

    Full Text Available DNA is constantly being oxidized, and oxidized DNA is prone to mutation; moreover, guanine is highly sensitive to several oxidative stressors. Several oxidatively damaged forms of guanine—including 2,2,4-triamino-5(2H-oxazolone (Oz, iminoallantoin (Ia, and spiroiminodihydantoin (Sp—can be paired with guanine, and cause G:C-C:G transversions. Previous findings indicate that guanine is incorporated more efficiently opposite Oz than opposite Ia or Sp, and that these differences in efficiency cannot be explained by differences in the stabilities of G:Oz, G:Ia, and G:Sp base pairs calculated ab initio. Here, to explain previous experimental result, we used a 3-base-pair model DNA duplex to calculate the difference in the stability and the distortion of DNA containing a G:Oz, G:Ia, or G:Sp base pair. We found that the stability of the structure containing 5ꞌ and 3ꞌ base pairs adjacent to G:Oz was more stable than that containing the respective base pairs adjacent to G:Ia or G:Sp. Moreover, the distortion of the structure in the DNA model duplex that contained a G:Oz was smaller than that containing a G:Ia or G:Sp. Therefore, our discussion can explain the previous results involving translesion synthesis past an oxidatively damaged guanine.

  2. Site-Directed Spin-Labeling of Nucleic Acids by Click Chemistry. Detection of Abasic Sites in Duplex DNA by EPR Spectroscopy

    DEFF Research Database (Denmark)

    Sigurdsson, Snorri; Vogel, Stefan; Shelke, Sandip

    2010-01-01

    This paper describes a spin label that can detect and identify local structural deformations in duplex DNA, in particular abasic sites. The spin label was incorporated into DNA by a new postsynthetic approach using click-chemistry on a solid support, which simplified both the synthesis and purifi......This paper describes a spin label that can detect and identify local structural deformations in duplex DNA, in particular abasic sites. The spin label was incorporated into DNA by a new postsynthetic approach using click-chemistry on a solid support, which simplified both the synthesis...

  3. Kinetics of DNA duplex formation: A-tracts versus AT-tracts.

    Science.gov (United States)

    Wyer, Jean Ann; Kristensen, Mads Bejder; Jones, Nykola C; Hoffmann, Søren Vrønning; Nielsen, Steen Brøndsted

    2014-09-21

    The hybridisation and melting of DNA strands are critical steps in many biological processes, but still a deeper understanding of the kinetics is lacking. This is evident from the absence of a clear correlation between rate constants for duplex formation and the number of bases in the strand or the sequence. Here we have probed differences between formation times of A-tracts and AT-tracts by studying complementary model strands mainly comprised of adenine (A) and thymine (T) in stopped-flow (SF) experiments. These strands are relevant as DNA replication begins in regions with a large number of AT base pairs. Interpretation of our results is aided by secondary-structure modelling where both the fractions of the different types of structures and the number of paired bases in the lowest-energy ones are determined. The model is based on calculation of free energies using fixed values for enthalpies and entropies associated with base pairing and a stochastic sampling of the possible structures. We find that the strand length affects rates: the activation energy for the formation of short (16-base pairs) A-tracts is larger than that for longer ones (20-base pairs). Activation energies for the formation of AT-tracts are an order of magnitude larger, and larger for shorter strands than for long ones. These higher activation energies are in agreement with the fact that the fraction of unpaired bases in the constituent AT-tract strands is less than in those which comprise the A-tracts. That the pre-structures of the single strands significantly affect rates is also used to rationalise the results obtained for two pairs of complementary 12-mer strands that have the same bases but in a different sequence; we report here similar activation energies as reported earlier and that these are strongly sequence dependent. Finally, we demonstrate that SF can be coupled with the measurement of circular dichroism (CD) in the vacuum ultraviolet (VUV) region, taking advantage of a

  4. Calcium phosphate composite materials including inorganic powders, BSA or duplex DNA prepared by W/O/W interfacial reaction method

    Energy Technology Data Exchange (ETDEWEB)

    Fujiwara, Masahiro [National Institute of Advanced Industrial Science and Technology (AIST), Kansai Center, 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577 (Japan)], E-mail: m-fujiwara@aist.go.jp; Shiokawa, Kumi; Morigaki, Kenichi; Tatsu, Yoshiro; Nakahara, Yoshiko [National Institute of Advanced Industrial Science and Technology (AIST), Kansai Center, 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577 (Japan)

    2008-03-10

    We reported before that inorganic reaction occurring at the interface of W/O/W emulsion is advantageous to produce hollow spheres (microcapsules) of inorganic matrices such as silica. This process enables us to include various materials into inorganic matrices directly. Calcium phosphates were also produced from NH{sub 4}H{sub 2}PO{sub 4} and Ca(OH){sub 2} by this interfacial reaction method. Various biomaterials are directly incorporated into crystalline calcium phosphate matrices, when the biomaterials are added to the inner water phase of the W/O/W emulsion. ZrO{sub 2} and Al{sub 2}O{sub 3} powders were effectively encapsulated in calcium phosphates such as hydroxyapatite (HAp). The images of backscattered electron of FE-SEM observations indicated that ZrO{sub 2} particles were included in HAp, while they adhered to the surface of HAp in the case of a simple precipitation method. Biomacromolecules such as BSA and duplex DNA were also included in HAp using the inner water phases dissolving them. Fluorescent microscopy observations revealed that biomacromolecules incorporated in HAp localized in some domains of the HAp matrices. Biomacromolecules thus included were scarcely liberated into deionized water, indicating their strong encapsulation in HAp. This general and simple methodology will provide various composite materials of calcium phosphates, which are applicable to regenerative medicine, DDS, GDS and more.

  5. Human RAD52 Captures and Holds DNA Strands, Increases DNA Flexibility, and Prevents Melting of Duplex DNA: Implications for DNA Recombination

    Directory of Open Access Journals (Sweden)

    Ineke Brouwer

    2017-03-01

    Full Text Available Human RAD52 promotes annealing of complementary single-stranded DNA (ssDNA. In-depth knowledge of RAD52-DNA interaction is required to understand how its activity is integrated in DNA repair processes. Here, we visualize individual fluorescent RAD52 complexes interacting with single DNA molecules. The interaction with ssDNA is rapid, static, and tight, where ssDNA appears to wrap around RAD52 complexes that promote intra-molecular bridging. With double-stranded DNA (dsDNA, interaction is slower, weaker, and often diffusive. Interestingly, force spectroscopy experiments show that RAD52 alters the mechanics dsDNA by enhancing DNA flexibility and increasing DNA contour length, suggesting intercalation. RAD52 binding changes the nature of the overstretching transition of dsDNA and prevents DNA melting, which is advantageous for strand clamping during or after annealing. DNA-bound RAD52 is efficient at capturing ssDNA in trans. Together, these effects may help key steps in DNA repair, such as second-end capture during homologous recombination or strand annealing during RAD51-independent recombination reactions.

  6. Evaluation of the Gibbs Free Energy Changes and Melting Temperatures of DNA/DNA Duplexes Using Hybridization Enthalpy Calculated by Molecular Dynamics Simulation.

    Science.gov (United States)

    Lomzov, Alexander A; Vorobjev, Yury N; Pyshnyi, Dmitrii V

    2015-12-10

    A molecular dynamics simulation approach was applied for the prediction of the thermal stability of oligonucleotide duplexes. It was shown that the enthalpy of the DNA/DNA complex formation could be calculated using this approach. We have studied the influence of various simulation parameters on the secondary structure and the hybridization enthalpy value of Dickerson-Drew dodecamer. The optimal simulation parameters for the most reliable prediction of the enthalpy values were determined. The thermodynamic parameters (enthalpy and entropy changes) of a duplex formation were obtained experimentally for 305 oligonucleotides of various lengths and GC-content. The resulting database was studied with molecular dynamics (MD) simulation using the optimized simulation parameters. Gibbs free energy changes and the melting temperatures were evaluated using the experimental correlation between enthalpy and entropy changes of the duplex formation and the enthalpy values calculated by the MD simulation. The average errors in the predictions of enthalpy, the Gibbs free energy change, and the melting temperature of oligonucleotide complexes were 11%, 10%, and 4.4 °C, respectively. We have shown that the molecular dynamics simulation gives a possibility to calculate the thermal stability of native DNA/DNA complexes a priori with an unexpectedly high accuracy.

  7. Effect of Deleterious Phases on Corrosion Resistance of Duplex Stainless Steel (2205

    Directory of Open Access Journals (Sweden)

    AbdulKadar M. Godil

    2013-07-01

    Full Text Available Duplex stainless steel is a Ferritic(BCC-Austenitic(FCC steel, covers the advantages of both Austenitic and Ferritic Stainless steels. They having good mechanical and corrosion resistance properties are widely used in many industries like chemical plants, refineries for critical equipments such as pressure vessels, heatexchangers, water heaters. Major problem occurs with duplex steels when they are worked or heated above about temperature of 280°C. Detrimental phases like Sigma, Chi, Laves and Alpha prime form when the Duplex steels are treated above this temperature and they retard the properties of Duplex stainless steels. They also cause embrittlement above temperature of 475°C called “475°C embrittlement”. During welding of duplex steels, Secondary austenite also forms, which is also one of the harmful phases in duplex steels. Among all of these phases, Sigma (σ is extremely harmful to the corrosion resistance of steel. Due to these limitations duplexgrades are not used above certain temperature ranges. In this experimental work a plate of duplex grade 2205 in hot worked condition was procured from TCR Advanced Engineering Pvt. Ltd., GIDC, Vadodara. Initially chemical composition of the plate was checked with emission spectrometer, tensile test and hardness tests werecarried out for comparing with the standard data. As there was no Sigma phase detected when tested with ASTM 930 in the received sample, Sigma phase was intentionally produced by giving heat treatment in the range of 700-850°C. Sigma phases were quantified with ASTM 930 practice A, by electrolytic etching with 40% NaOH. The effect of Sigma phase on corrosion resistance was measured by ASTM G48. The pitting corrosion resistance was evaluated in terms of average pit depth and overall corrosion rate.

  8. Human PSF concentrates DNA and stimulates duplex capture in DMC1-mediated homologous pairing

    Science.gov (United States)

    Morozumi, Yuichi; Ino, Ryohei; Takaku, Motoki; Hosokawa, Mihoko; Chuma, Shinichiro; Kurumizaka, Hitoshi

    2012-01-01

    PSF is considered to have multiple functions in RNA processing, transcription and DNA repair by mitotic recombination. In the present study, we found that PSF is produced in spermatogonia, spermatocytes and spermatids, suggesting that PSF may also function in meiotic recombination. We tested the effect of PSF on homologous pairing by the meiosis-specific recombinase DMC1, and found that human PSF robustly stimulated it. PSF synergistically enhanced the formation of a synaptic complex containing DMC1, ssDNA and dsDNA during homologous pairing. The PSF-mediated DMC1 stimulation may be promoted by its DNA aggregation activity, which increases the local concentrations of ssDNA and dsDNA for homologous pairing by DMC1. These results suggested that PSF may function as an activator for the meiosis-specific recombinase DMC1 in higher eukaryotes. PMID:22156371

  9. Enthalpy and heat capacity changes for formation of an oligomeric DNA duplex: interpretation in terms of coupled processes of formation and association of single-stranded helices.

    Science.gov (United States)

    Holbrook, J A; Capp, M W; Saecker, R M; Record, M T

    1999-06-29

    The thermodynamics of self-assembly of a 14 base pair DNA double helix from complementary strands have been investigated by titration (ITC) and differential scanning (DSC) calorimetry, in conjunction with van't Hoff analysis of UV thermal scans of individual strands. These studies demonstrate that thermodynamic characterization of the temperature-dependent contributions of coupled conformational equilibria in the individual "denatured" strands and in the duplex is essential to understand the origins of duplex stability and to derive stability prediction schemes of general applicability. ITC studies of strand association at 293 K and 120 mM Na+ yield an enthalpy change of -73 +/- 2 kcal (mol of duplex)-1. ITC studies between 282 and 312 K at 20, 50, and 120 mM Na+ show that the enthalpy of duplex formation is only weakly salt concentration-dependent but is very strongly temperature-dependent, decreasing approximately linearly with increasing temperature with a heat capacity change (282-312 K) of -1.3 +/- 0.1 kcal K-1 (mol of duplex)-1. From DSC denaturation studies in 120 mM Na+, we obtain an enthalpy of duplex formation of -120 +/- 5 kcal (mol of duplex)-1 and an estimate of the corresponding heat capacity change of -0.8 +/- 0.4 kcal K-1 (mol of duplex)-1 at the Tm of 339 K. van't Hoff analysis of UV thermal scans on the individual strands indicates that single helix formation is noncooperative with a temperature-independent enthalpy change of -5.5 +/- 0.5 kcal at 120 mM Na+. From these observed enthalpy and heat capacity changes, we obtain the corresponding thermodynamic quantities for two fundamental processes: (i) formation of single helices from disordered strands, involving only intrastrand (vertical) interactions between neighboring bases; and (ii) formation of double helices by association (docking) of single helical strands, involving interstrand (horizontal and vertical) interactions. At 293 K and 120 mM Na+, we calculate that the enthalpy change for

  10. Free energy landscape and transition pathways from Watson-Crick to Hoogsteen base pairing in free duplex DNA.

    Science.gov (United States)

    Yang, Changwon; Kim, Eunae; Pak, Youngshang

    2015-09-18

    Houghton (HG) base pairing plays a central role in the DNA binding of proteins and small ligands. Probing detailed transition mechanism from Watson-Crick (WC) to HG base pair (bp) formation in duplex DNAs is of fundamental importance in terms of revealing intrinsic functions of double helical DNAs beyond their sequence determined functions. We investigated a free energy landscape of a free B-DNA with an adenosine-thymine (A-T) rich sequence to probe its conformational transition pathways from WC to HG base pairing. The free energy landscape was computed with a state-of-art two-dimensional umbrella molecular dynamics simulation at the all-atom level. The present simulation showed that in an isolated duplex DNA, the spontaneous transition from WC to HG bp takes place via multiple pathways. Notably, base flipping into the major and minor grooves was found to play an important role in forming these multiple transition pathways. This finding suggests that naked B-DNA under normal conditions has an inherent ability to form HG bps via spontaneous base opening events.

  11. Detection of mRNA of the cyclin D1 breast cancer marker by a novel duplex-DNA probe.

    Science.gov (United States)

    Segal, Meirav; Yavin, Eylon; Kafri, Pinhas; Shav-Tal, Yaron; Fischer, Bilha

    2013-06-27

    Previously, we have described 5-((4-methoxy-phenyl)-trans-vinyl)-2'-deoxy-uridine, 6, as a fluorescent uridine analogue exhibiting a 3000-fold higher quantum yield (Φ 0.12) and maximum emission (478 nm) which is 170 nm red-shifted as compared to uridine. Here, we utilized 6 for preparation of labeled oligodeoxynucleotide (ODN) probes based on MS2 and cyclin D1 (a known breast cancer mRNA marker) sequences. Cyclin D1-derived labeled-ssODN showed a 9.5-fold decrease of quantum yield upon duplex formation. On the basis of this finding, we developed the ds-NIF (nucleoside with intrinsic fluorescence)-probe methodology for detection of cyclin D1 mRNA, by which the fluorescent probe is released upon recognition of target mRNA by the relatively dark NIF-duplex-probe. Indeed, we successfully detected, a ss-deoxynucleic acid (DNA) variant of cyclin D1 mRNA using a dark NIF-labeled duplex-probe, and monitoring the recognition process by fluorescence spectroscopy and gel electrophoresis. Furthermore, we successfully detected cyclin D1 mRNA in RNA extracted from cancerous human cells, using ds-NIF methodology.

  12. Solvation Effects on Hole Mobility in the Poly G/Poly C Duplex

    CERN Document Server

    Lakhno, V D

    2013-01-01

    Theoretical calculations of solvation contribution to hole energy in a polynucleotide chain give very low hole mobility values at zero temperature, \\mu < 10^{-3} cm^2/(V s). We calculated hole mobility at physiological temperature for the Poly G/Poly C DNA duplex, which gave substantially larger mobility values. Mobility over the temperature range 20-400 K was calculated. Taking stacking interaction into account substantially increased hole mobility.

  13. Chemical shifts assignments of the archaeal MC1 protein and a strongly bent 15 base pairs DNA duplex in complex.

    Science.gov (United States)

    Loth, Karine; Landon, Céline; Paquet, Françoise

    2015-04-01

    MC1 is the most abundant architectural protein present in Methanosarcina thermophila CHTI55 in laboratory growth conditions and is structurally unrelated to other DNA-binding proteins. MC1 functions are to shape and to protect DNA against thermal denaturation by binding to it. Therefore, MC1 has a strong affinity for any double-stranded DNA. However, it recognizes and preferentially binds to bent DNA, such as four-way junctions and negatively supercoiled DNA minicircles. Combining NMR data, electron microscopy data, biochemistry, molecular modelisation and docking approaches, we proposed recently a new type of DNA/protein complex, in which the monomeric protein MC1 binds on the concave side of a strongly bent 15 base pairs DNA. We present here the NMR chemical shifts assignments of each partner in the complex, (1)H (15)N MC1 protein and (1)H (13)C (15)N bent duplex DNA, as first step towards the first experimental 3D structure of this new type of DNA/protein complex.

  14. Human MUS81-EME2 can cleave a variety of DNA structures including intact Holliday junction and nicked duplex.

    Science.gov (United States)

    Amangyeld, Tamir; Shin, Yong-Keol; Lee, Miju; Kwon, Buki; Seo, Yeon-Soo

    2014-05-01

    MUS81 shares a high-degree homology with the catalytic XPF subunit of the XPF-ERCC1 endonuclease complex. It is catalytically active only when complexed with the regulatory subunits Mms4 or Eme1 in budding and fission yeasts, respectively, and EME1 or EME2 in humans. Although Mus81 complexes are implicated in the resolution of recombination intermediates in vivo, recombinant yeast Mus81-Mms4 and human MUS81-EME1 isolated from Escherichia coli fail to cleave intact Holliday junctions (HJs) in vitro. In this study, we show that human recombinant MUS81-EME2 isolated from E. coli cleaves HJs relatively efficiently, compared to MUS81-EME1. Furthermore, MUS81-EME2 catalyzed cleavage of nicked and gapped duplex deoxyribonucleic acids (DNAs), generating double-strand breaks. The presence of a 5' phosphate terminus at nicks and gaps rendered DNA significantly less susceptible to the cleavage by MUS81-EME2 than its absence, raising the possibility that this activity could play a role in channeling damaged DNA duplexes that are not readily repaired into the recombinational repair pathways. Significant differences in substrate specificity observed with unmodified forms of MUS81-EME1 and MUS81-EME2 suggest that they play related but non-overlapping roles in DNA transactions. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  15. Effect of Temperature and Cl-Concentration on Pitting of 2205 Duplex Stainless Steel

    Institute of Scientific and Technical Information of China (English)

    DONG Chaofang; LUO Hong; XIAO Kui; SUN Ting; LIU Qian; LI Xiaogang

    2011-01-01

    The electrochemical behaviors of 2205 duplex stainless steel in NaCl solution with different temperatures and concentrations were studied by gravimetric tests,potentiodynamic polarization,electrochemical impedance spectroscopy and scanning electron microscopy.The experimental results show that temperature and chloride concentration have a great influence on the pitting resistance of 2205 duplex stainless steels.They not only effect the corrosion rate of pitting,but also change the shape of the pits.When NaCl solution was in low concentration and temperature below the critical pitting temperature,pits were very small and scattered with hemisphere-like shape.On the contrary,the pits of 2205 duplex stainless steel were large and sometimes had a lacy cover when the NaCl concentration was higher and the temperature was 70℃.

  16. Chemical structure and properties of interstrand cross-links formed by reaction of guanine residues with abasic sites in duplex DNA.

    Science.gov (United States)

    Catalano, Michael J; Liu, Shuo; Andersen, Nisana; Yang, Zhiyu; Johnson, Kevin M; Price, Nathan E; Wang, Yinsheng; Gates, Kent S

    2015-03-25

    A new type of interstrand cross-link resulting from the reaction of a DNA abasic site with a guanine residue on the opposing strand of the double helix was recently identified, but the chemical connectivity of the cross-link was not rigorously established. The work described here was designed to characterize the chemical structure and properties of dG-AP cross-links generated in duplex DNA. The approach involved characterization of the nucleoside cross-link "remnant" released by enzymatic digestion of DNA duplexes containing the dG-AP cross-link. We first carried out a chemical synthesis and complete spectroscopic structure determination of the putative cross-link remnant 9b composed of a 2-deoxyribose adduct attached to the exocyclic N(2)-amino group of dG. A reduced analogue of the cross-link remnant was also prepared (11b). Liquid chromatography-tandem mass spectrometric (LC-MS/MS) analysis revealed that the retention times and mass spectral properties of synthetic standards 9b and 11b matched those of the authentic cross-link remnants released by enzymatic digestion of duplexes containing the native and reduced dG-AP cross-link, respectively. These results establish the chemical connectivity of the dG-AP cross-link released from duplex DNA and provide a foundation for detection of this lesion in biological samples. The dG-AP cross-link in duplex DNA was remarkably stable, decomposing with a half-life of 22 days at pH 7 and 23 °C. The intrinsic chemical stability of the dG-AP cross-link suggests that this lesion in duplex DNA may have the power to block DNA-processing enzymes involved in transcription and replication.

  17. Melting of duplex DNA in the absence of ATP by NS3 helicase domain through specific interaction with a single-strand/double-strand junction

    Science.gov (United States)

    Reynolds, Kimberly A.; Cameron, Craig E.; Raney, Kevin D.

    2016-01-01

    Helicases unwind double-stranded nucleic acids, remove secondary structures from single-stranded nucleic acids, and remove proteins bound to nucleic acids. For many helicases, the mechanisms for these different functions share the ability to translocate with a directional bias as a result of ATP binding and hydrolysis. The nonstructural protein 3 (NS3) is an essential enzyme expressed by the hepatitis C virus (HCV) and is known to catalyze the unwinding of both DNA and RNA substrates in a 3′-to-5′ direction. We investigated the role of nucleic acid binding in the unwinding mechanism by examining ATP-independent unwinding. We observed that even in the absence of ATP, NS3 helicase domain (NS3h) unwound duplexes only when they contained a 3′-tail (i.e., 3′-to-5′ directionality). Blunt-ended duplexes and 5′-tailed duplexes were not melted even in the presence of a large excess concentration of the protein. NS3h was found to diffuse rapidly along single-stranded DNA at a rate of 30 nt2·s−1. Upon encountering an appropriate single-strand/double-strand (ss/ds) junction, NS3h slowly melted the duplex under conditions with excess protein concentration relative to DNA concentration. When a biotin-streptavidin block was placed into the ssDNA region, no melting of DNA was observed, suggesting that NS3h must diffuse along the ssDNA, and that the streptavidin blocked the diffusion. We conclude that the specific interaction between NS3h and the ss/dsDNA junction, coupled with diffusion allows binding energy to melt duplex DNA with a directional bias. Alternatively, we found that the full-length NS3 protein did not exhibit strict directionality and was dependent on duplex DNA length. NS3 was able to unwind the duplex even in the presence of the biotin-streptavidin block. We propose a non-canonical model of unwinding for NS3 in which the enzyme binds directly to the duplex via protein-protein interactions to melt the substrate. PMID:26091150

  18. Calorimetric and Spectroscopic Analysis of the Thermal Stability of Short Duplex DNA-Containing Sugar and Base-Modified Nucleotides.

    Science.gov (United States)

    Fakhfakh, Kareem; Hughesman, Curtis B; Louise Creagh, A; Kao, Vincent; Haynes, Charles

    2016-01-01

    Base- and sugar-modified analogs of DNA and RNA are finding ever expanding use in medicine and biotechnology as tools to better tailor structured oligonucleotides by altering their thermal stability, nuclease resistance, base-pairing specificity, antisense activity, or cellular uptake. Proper deployment of these chemical modifications generally requires knowledge of how each affects base-pairing properties and thermal stabilities. Here, we describe in detail how differential scanning calorimetry and UV spectroscopy may be used to quantify the melting thermodynamics of short dsDNA containing chemically modified nucleosides in one or both strands. Insights are provided into why and how the presence of highly stable base pairs containing modified nucleosides can alter the nature of calorimetry or melting spectroscopy data, and how each experiment must therefore be conducted to ensure high-quality melting thermodynamics data are obtained. Strengths and weaknesses of the two methods when applied to chemically modified duplexes are also addressed.

  19. Duplex ultrasound

    Science.gov (United States)

    Vascular ultrasound; Peripheral vascular ultrasound ... A duplex ultrasound combines: Traditional ultrasound: This uses sound waves that bounce off blood vessels to create pictures. Doppler ultrasound: This ...

  20. DETECTION OF PORK CONTAMINATION IN FRESH AND COOKED BEEF USING GENETIC MARKER MITOCHONDRIAL-DNA CYTOCHROME B BY DUPLEX-PCR

    Directory of Open Access Journals (Sweden)

    A. Ni’mah

    2016-03-01

    Full Text Available By mixing with pork, beef adulteration is frequently found in the traditional  market that very disturbing Moeslem community in Indonesia. This study was conducted to detect pork contamination in fresh and cooked beef using genetic marker mitochondrial DNA cytochrome b (mt-DNA Cyt b by duplex-PCR. A total of twelve samples was used in this study consisting six fresh meat samples and six cooked meat samples, respectively. Those beef and pork were bought from animal slaughterhouse and a supermarket in Surakarta. Cooked samples were prepared by boiling the meats in hot water at 100oC for 30 minutes. We designed pork contamination in beef in the level of 0, 1, 5, 10, 25%, respectively. The DNA genome was extracted and polymerase chain reaction (PCR was performed using species specific primer to isolate mt-DNA Cyt b gene from the samples. The results showed that the DNA genome was successfully extracted from pork, beef, and contaminated meat samples. In addition, visualization of duplex-PCR on 1.5% agarose gel was able to detect pork contamination in both fresh and cooked beef up to very small proportion (1%. The existence of pork in beef was indicated with the presence of specific 398 bp DNA band. It can be concluded, duplex-PCR of mt-DNA Cyt b gene was very sensitive in detection of pork contamination in fresh and cooked beef.

  1. Can copper(II) mediate Hoogsteen base-pairing in a left-handed DNA duplex? A pulse EPR study.

    Science.gov (United States)

    Santangelo, Maria Grazia; Antoni, Philipp M; Spingler, Bernhard; Jeschke, Gunnar

    2010-02-22

    Pulse EPR spectroscopy is used to investigate possible structural features of the copper(II) ion coordinated to poly(dG-dC).poly(dG-dC) in a frozen aqueous solution, and the structural changes of the polynucleotide induced by the presence of the metal ion. Two different copper species were identified and their geometry explained by a molecular model. According to this model, one species is exclusively coordinated to a single guanine with the N7 nitrogen atom forming a coordinative bond with the copper. In the other species, a guanine and a cytosine form a ternary complex together with the copper ion. A copper crosslink between the N7 of guanine and N3 of cytosine is proposed as the most probable coordination site. Moreover, no evidence was found for an interaction of either copper species with a phosphate group or equatorial water molecules. In addition, circular dichroism (CD) spectroscopy showed that the DNA of the Cu(II)-poly(dG-dC).poly(dG-dC) adducts resembles the left-handed Z-form. These results suggest that metal-mediated Hoogsteen base pairing, as previously proposed for a right-handed DNA duplex, can also occur in a double-stranded left-handed DNA.

  2. Recognition of double-stranded DNA using energetically activated duplexes with interstrand zippers of 1-, 2-or 4-pyrenyl-functionalized O2 '-alkylated RNA monomers

    DEFF Research Database (Denmark)

    Karmakar, Saswata; Madsen, Andreas Stahl; Guenther, Dale C.

    2014-01-01

    Despite advances with triplex-forming oligonucleotides, peptide nucleic acids, polyamides and more recently engineered proteins, there remains an urgent need for synthetic ligands that enable specific recognition of double-stranded (ds) DNA to accelerate studies aiming at detecting, regulating...... and modifying genes. Invaders, i.e., energetically activated DNA duplexes with interstrand zipper arrangements of intercalator-functionalized nucleotides, are emerging as an attractive approach toward this goal. Here, we characterize and compare Invaders based on 1-, 2- and 4-pyrenyl-functionalized O2...... hairpins with single nucleotide fidelity. Intercalator-mediated unwinding and activation of the double-stranded probe, coupled with extraordinary stabilization of probe target duplexes (Delta T-m/modification up to +14.0 degrees C), provides the driving force for dsDNA recognition. In contrast, Z...

  3. ‘Squeezed’ interparticle properties for plasmonic coupling and SERS characteristics of duplex DNA conjugated/linked gold nanoparticles of homo/hetero-sizes

    Science.gov (United States)

    Skeete, Zakiya; Cheng, Han-Wen; Ngo, Quang Minh; Salazar, Christian; Sun, Winny; Luo, Jin; Zhong, Chuan-Jian

    2016-08-01

    The formation of interparticle duplex DNA conjugates with gold nanoparticles constitutes the basis for interparticle plasmonic coupling responsible for surface-enhanced Raman scattering signal amplification, but understanding of its correlation with interparticle spatial properties and particle sizes, especially in aqueous solutions, remains elusive. This report describes findings of an investigation of interparticle plasmonic coupling based on experimental measurements of localized surface plasmon resonance and surface enhanced Raman scattering characteristics for gold nanoparticles in aqueous solutions upon introduction of interparticle duplex DNA conjugates to define the interparticle spatial properties. Theoretical simulations of the interparticle optical properties and electric field enhancement based on a dimer model have also been performed to aid the understanding of the experimental results. The results have revealed a ‘squeezed’ interparticle spatial characteristic in which the duplex DNA-defined distance is close or shorter than A-form DNA conformation, which are discussed in terms of the interparticle interactions, providing fresh insight into the interparticle double-stranded DNA-defined interparticle spatial properties for the design of highly-sensitive nanoprobes in solutions for biomolecular detection.

  4. Evaluation of Fluorescent Analogs of Deoxycytidine for Monitoring DNA Transitions from Duplex to Functional Structures

    Directory of Open Access Journals (Sweden)

    Yogini P. Bhavsar

    2011-01-01

    Full Text Available Topological variants of single-strand DNA (ssDNA structures, referred to as “functional DNA,” have been detected in regulatory regions of many genes and are thought to affect gene expression. Two fluorescent analogs of deoxycytidine, Pyrrolo-dC (PdC and 1,3-diaza-2-oxophenoxazine (tC∘, can be incorporated into DNA. Here, we describe spectroscopic studies of both analogs to determine fluorescent properties that report on structural transitions from double-strand DNA (dsDNA to ssDNA, a common pathway in the transition to functional DNA structures. We obtained fluorescence-detected circular dichroism (FDCD spectra, steady-state fluorescence spectra, and fluorescence lifetimes of the fluorophores in DNA. Our results show that PdC is advantageous in fluorescence lifetime studies because of a distinct ~2 ns change between paired and unpaired bases. However, tC∘ is a better probe for FDCD experiments that report on the helical structure of DNA surrounding the fluorophore. Both fluorophores provide complementary data to measure DNA structural transitions.

  5. Target-controlled formation of silver nanoclusters in abasic site-incorporated duplex DNA for label-free fluorescence detection of theophylline

    Science.gov (United States)

    Park, Ki Soo; Oh, Seung Soo; Soh, H. Tom; Park, Hyun Gyu

    2014-08-01

    A novel, label-free, fluorescence based sensor for theophylline has been developed. In the new sensor system, an abasic site-incorporated duplex DNA probe serves as both a pocket for recognition of theophylline and a template for the preparation of fluorescent silver nanoclusters. The strategy relies on theophylline-controlled formation of fluorescent silver nanoclusters from abasic site-incorporated duplex DNA. When theophylline is not present, silver ions interact with the cytosine groups opposite to the abasic site in duplex DNA. This interaction leads to efficient formation of intensely red fluorescent silver nanoclusters. In contrast, when theophylline is bound at the abasic site through pseudo base-pairing with appropriately positioned cytosines, silver ion binding to the cytosine nucleobase is prevented. Consequently, fluorescent silver nanoclusters are not formed causing a significant reduction of the fluorescence signal. By employing this new sensor, theophylline can be highly selectively detected at a concentration as low as 1.8 μM. Finally, the diagnostic capability and practical application of this sensor were demonstrated by its use in detecting theophylline in human blood serum.A novel, label-free, fluorescence based sensor for theophylline has been developed. In the new sensor system, an abasic site-incorporated duplex DNA probe serves as both a pocket for recognition of theophylline and a template for the preparation of fluorescent silver nanoclusters. The strategy relies on theophylline-controlled formation of fluorescent silver nanoclusters from abasic site-incorporated duplex DNA. When theophylline is not present, silver ions interact with the cytosine groups opposite to the abasic site in duplex DNA. This interaction leads to efficient formation of intensely red fluorescent silver nanoclusters. In contrast, when theophylline is bound at the abasic site through pseudo base-pairing with appropriately positioned cytosines, silver ion binding to

  6. G4-Tetra DNA Duplex Induce Lung Cancer Cell Apoptosis in A549 Cells

    Science.gov (United States)

    Xu, Xiaobo; Zhao, YiZhuo; Lu, Hu; Fu, Cuiping; Li, Xiao; Jiang, Liyan; Li, Shanqun

    2016-10-01

    The specific DNA is typically impermeable to the plasma membrane due to its natural characters, but DNA tetra structures (DTNs) can be readily uptake by cells in the absence of transfection agents, providing a new strategy to deliver DNA drugs. In this research, the delivery efficiency of tetrahedral DNA nanostructures was measured on adenocarcinomic human alveolar basal epithelial (A549) cells via delivering AS1411 (G4). The DNA tetra-AS1411 complex was rapidly and abundantly uptake by A549 cells, and the induced apoptosis was enhanced. Furthermore, biodistribution in mouse proved the rapid clearance from non-targeted organs in vivo. This study improved the understanding of potential function in DNA-based drug delivery and proved that DTNs-AS1411 could be potentially useful for the treatment of lung cancer.

  7. Differential targeting of unpaired bases within duplex DNA by the natural compound clerocidin: a valuable tool to dissect DNA secondary structure.

    Directory of Open Access Journals (Sweden)

    Matteo Nadai

    Full Text Available Non-canonical DNA structures have been postulated to mediate protein-nucleic acid interactions and to function as intermediates in the generation of frame-shift mutations when errors in DNA replication occur, which result in a variety of diseases and cancers. Compounds capable of binding to non-canonical DNA conformations may thus have significant diagnostic and therapeutic potential. Clerocidin is a natural diterpenoid which has been shown to selectively react with single-stranded bases without targeting the double helix. Here we performed a comprehensive analysis on several non-canonical DNA secondary structures, namely mismatches, nicks, bulges, hairpins, with sequence variations in both the single-stranded region and the double-stranded flanking segment. By analysis of clerocidin reactivity, we were able to identify the exposed reactive residues which provided information on both the secondary structure and the accessibility of the non-paired sites. Mismatches longer than 1 base were necessary to be reached by clerocidin reactive groups, while 1-base nicks were promptly targeted by clerocidin; in hairpins, clerocidin reactivity increased with the length of the hairpin loop, while, interestingly, reactivity towards bulges reached a maximum in 3-base-long bulges and declined in longer bulges. Electrophoretic mobility shift analysis demonstrated that bulges longer than 3 bases (i.e. 5- and 7-bases folded or stacked on the duplex region therefore being less accessible by the compound. Clerocidin thus represents a new valuable diagnostic tool to dissect DNA secondary structures.

  8. DNA-mediated electron transfer in DNA duplexes tethered to gold electrodes via phosphorothioated dA tags.

    Science.gov (United States)

    Campos, Rui; Kotlyar, Alexander; Ferapontova, Elena E

    2014-10-14

    The efficiency of DNA-based bioelectronic devices strongly depends on the way DNA molecules are linked to the electronic component. Commonly, DNA is tethered to metal electrodes via an alkanethiol linker representing an additional barrier for electron transport. Here we demonstrate that the replacement of the alkanethiol linker for a phosphorothioated adenosine tag increases the rate of DNA-mediated electron transfer (ET) up to 259 s(-1), representing the highest hitherto reported rate of electrochemically-modulated ET, and improves the stability of DNA-electrode surface binding. Both results offer pronounced technological and scientific benefits for DNA-based electronics.

  9. Effect of laser shock processing on fatigue crack growth of duplex stainless steel

    Energy Technology Data Exchange (ETDEWEB)

    Rubio-Gonzalez, C., E-mail: crubio@cidesi.mx [Centro de Ingenieria y Desarrollo Industrial, Pie de la Cuesta, 702, Desarrollo San Pablo, Queretaro, Qro., 76130 (Mexico); Felix-Martinez, C. [Centro de Ingenieria y Desarrollo Industrial, Pie de la Cuesta, 702, Desarrollo San Pablo, Queretaro, Qro., 76130 (Mexico); Gomez-Rosas, G. [Universidad de Guadalajara, Guadalajara, Jal (Mexico); Ocana, J.L.; Morales, M.; Porro, J.A. [Departamento de Fisica Aplicada a la Ingenieria Industrial, E.T.S.I.I., Universidad Politecnica de Madrid (Spain)

    2011-01-25

    Research highlights: {yields} LSP is an effective surface treatment to improve fatigue properties of duplex stainless steel. {yields} Increasing pulse density, fatigue crack growth rate is reduced. {yields} Microstructure is not affected by LSP. {yields} Compressive residual stresses increases increasing pulse density. - Abstract: Duplex stainless steels have wide application in different fields like the ship, petrochemical and chemical industries that is due to their high strength and excellent toughness properties as well as their high corrosion resistance. In this work an investigation is performed to evaluate the effect of laser shock processing on some mechanical properties of 2205 duplex stainless steel. Laser shock processing (LSP) or laser shock peening is a new technique for strengthening metals. This process induces a compressive residual stress field which increases fatigue crack initiation life and reduces fatigue crack growth rate. A convergent lens is used to deliver 2.5 J, 8 ns laser pulses by a Q-switched Nd:YAG laser, operating at 10 Hz with infrared (1064 nm) radiation. The pulses are focused to a diameter of 1.5 mm. Effect of pulse density in the residual stress field is evaluated. Residual stress distribution as a function of depth is determined by the contour method. It is observed that the higher the pulse density the greater the compressive residual stress. Pulse densities of 900, 1600 and 2500 pul/cm{sup 2} are used. Pre-cracked compact tension specimens were subjected to LSP process and then tested under cyclic loading with R = 0.1. Fatigue crack growth rate is determined and the effect of LSP process parameters is evaluated. In addition fracture toughness is determined in specimens with and without LSP treatment. It is observed that LSP reduces fatigue crack growth and increases fracture toughness if this steel.

  10. Structure of the DNA duplex d(ATTAAT2 with Hoogsteen hydrogen bonds.

    Directory of Open Access Journals (Sweden)

    Francisco J Acosta-Reyes

    Full Text Available The traditional Watson-Crick base pairs in DNA may occasionally adopt a Hoogsteen conformation, with a different organization of hydrogen bonds. Previous crystal structures have shown that the Hoogsteen conformation is favored in alternating AT sequences of DNA. Here we present new data for a different sequence, d(ATTAAT2, which is also found in the Hoogsteen conformation. Thus we demonstrate that other all-AT sequences of DNA with a different sequence may be found in the Hoogsteen conformation. We conclude that any all-AT sequence might acquire this conformation under appropriate conditions. We also compare the detailed features of DNA in either the Hoogsteen or Watson-Crick conformations.

  11. A single molecule study of G-quadruplex and short duplex DNA structures

    Science.gov (United States)

    Roy, William A., Jr.

    Given that certain conditions are met, a single stranded DNA/RNA (ssDNA/RNA) structure called G-quadruplex (GQ) can form in regions throughout the genome, including at the telomeres and internal regions of the chromosomes. These structures serve various functions depending on the region in which they form which include protecting the chromosome ends, interfering with telomere elongation in cancer cells, and regulating transcription and translation level gene expression. Due to their high stability, various cellular mechanisms, such as GQ destabilizing proteins, are employed to unfold these structures during DNA replication or repair. Yet, their distinct layered structure has made GQs an attractive drug target in cancer treatment as GQ stabilizing molecules could inhibit telomerase dependent telomere elongation, a mechanism occurring in the majority of cancer cells to avoid senescence and apoptosis. However, proteins or small molecules interact with GQ that is under the influence of various cellular tension mechanisms, including the tension applied by other nearby molecules or the tension due to DNA structure within the chromatin context. Therefore, it is important to characterize the stability of various GQs and their response to interacting molecules when subjected to a tensile force. We employed a novel DNA-based nano tension generator that utilizes the elastic properties of circularized short double-stranded DNA (dsDNA) oligonucleotides to apply tension on the GQ. Since this is a completely new approach, the majority of this thesis was dedicated to proof-of-principle studies that demonstrated the feasibility and functionality of the method.

  12. Structure of an argonaute silencing complex with a seed-containing guide DNA and target RNA duplex

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yanli; Juranek, Stefan; Li, Haitao; Sheng, Gang; Tuschl, Thomas; Patel, Dinshaw J. (MSKCC); (HHMI)

    2009-01-08

    Here we report on a 3.0 {angstrom} crystal structure of a ternary complex of wild-type Thermus thermophilus argonaute bound to a 5'-phosphorylated 21-nucleotide guide DNA and a 20-nucleotide target RNA containing cleavage-preventing mismatches at the 10-11 step. The seed segment (positions 2 to 8) adopts an A-helical-like Watson-Crick paired duplex, with both ends of the guide strand anchored in the complex. An arginine, inserted between guide-strand bases 10 and 11 in the binary complex, locking it in an inactive conformation, is released on ternary complex formation. The nucleic-acid-binding channel between the PAZ- and PIWI-containing lobes of argonaute widens on formation of a more open ternary complex. The relationship of structure to function was established by determining cleavage activity of ternary complexes containing position-dependent base mismatch, bulge and 2'-O-methyl modifications. Consistent with the geometry of the ternary complex, bulges residing in the seed segments of the target, but not the guide strand, were better accommodated and their complexes were catalytically active.

  13. Cost-effectiveness of identifying aortoiliac and femoropopliteal arterial disease with angiography or duplex scanning

    Energy Technology Data Exchange (ETDEWEB)

    Coffi, S.B. [Department of Surgery, Academic Medical Center, Amsterdam (Netherlands); Ubbink, D.Th. [Department of Surgery, Academic Medical Center, Amsterdam (Netherlands); Department of Clinical Epidemiology, Biostatistics and Bioinformatics, Academic Medical Center, Amsterdam (Netherlands)], E-mail: D.Ubbink@amc.nl; Dijkgraaf, M.G.W. [Department of Clinical Epidemiology, Biostatistics and Bioinformatics, Academic Medical Center, Amsterdam (Netherlands); Reekers, J.A. [Department of Radiology, Academic Medical Center, Amsterdam (Netherlands); Legemate, D.A. [Department of Surgery, Academic Medical Center, Amsterdam (Netherlands)

    2008-04-15

    Objectives: Cost-effectiveness analysis of three diagnostic imaging strategies for the assessment of aortoiliac and femoropopliteal arteries in patients with peripheral arterial occlusive disease. The strategies were: angiography as the reference strategy, duplex scanning (DS) plus supplementary angiography (S1) and DS plus confirmative angiography (S2). Design, materials and methods: A decision model was built with sensitivity and specificity data from literature, supplemented with prospective hospital cost data in Euro ( Euro ). The probability of correctly identifying the status of a lesion was taken as the primary outcome. We compared strategies by assessing the extra costs per additional correctly identified case. Results: Assuming no false positive or false negative results, angiography is the most effective strategy if the prevalence of significant obstructive lesions in the aortoiliac and femoropopliteal tract exceeds 70%, or if the sensitivity of duplex scanning is lower than 83%. In case of lower prevalence, strategy S1 becomes equally or even more effective than angiography. At a prevalence of 75%, performing angiography costs Euro 8443 per extra correctly identified case compared with strategy S1. Conclusions: In most situations angiography is more effective than diagnostic strategy S1. However, if society is unwilling to pay more than Euro 8443 for knowing a patient's disease status, diagnostic strategy S1 is a cost-effective alternative to angiography, especially at lower prevalence values.

  14. Sequence-specific protection of duplex DNA against restriction and methylation enzymes by pseudocomplementary PNAs

    DEFF Research Database (Denmark)

    Izvolsky, K I; Demidov, V V; Nielsen, P E;

    2000-01-01

    of decameric pcPNAs block an access of RNA polymerase to the corresponding promoter. Here, we show that this type of PNAs protects selected DNA sites containing all four nucleobases from the action of restriction enzymes and DNA methyltransferases. We have found that pcPNAs as short as octamers form stable......I restriction endonuclease and dam methylase. The pcPNA-assisted protection against enzymatic methylation is more efficient when the PNA-binding site embodies the methylase-recognition site rather than overlaps it. We conclude that pcPNAs may provide the robust tools allowing to sequence-specifically manipulate...

  15. Using DNA duplex stability information for transcription factor binding site discovery.

    Science.gov (United States)

    Gordân, Raluca; Hartemink, Alexander J

    2008-01-01

    Transcription factor (TF) binding site discovery is an important step in understanding transcriptional regulation. Many computational tools have already been developed, but their success in detecting TF motifs is still limited. We believe one of the main reasons for the low accuracy of current methods is that they do not take into account the structural aspects of TF-DNA interaction. We have previously shown that knowledge about the structural class of the TF and information about nucleosome occupancy can be used to improve motif discovery. Here, we demonstrate the benefits of using information about the DNA double-helical stability for motif discovery. We notice that, in general, the energy needed to destabilize the DNA double helix is higher at TF binding sites than at random DNA sites. We use this information to derive informative positional priors that we incorporate into a motif finding algorithm. When applied to yeast ChIP-chip data, the new informative priors improve the performance of the motif finder significantly when compared to priors that do not use the energetic stability information.

  16. Effect of ultrafine grain on tensile behaviour and corrosion resistance of the duplex stainless steel.

    Science.gov (United States)

    Jinlong, Lv; Tongxiang, Liang; Chen, Wang; Limin, Dong

    2016-05-01

    The ultrafine grained 2205 duplex stainless steel was obtained by cold rolling and annealing. The tensile properties were investigated at room temperature. Comparing with coarse grained stainless steel, ultrafine grained sample showed higher strength and plasticity. In addition, grain size changed deformation orientation. The strain induced α'-martensite was observed in coarse grained 2205 duplex stainless steel with large strain. However, the grain refinement inhibited the transformation of α'-martensite;nevertheless, more deformation twins improved the strength and plasticity of ultrafine grained 2205 duplex stainless steel. In addition, the grain refinement improved corrosion resistance of the 2205 duplex stainless steel in sodium chloride solution.

  17. Insights into the Structures of DNA Damaged by Hydroxyl Radical: Crystal Structures of DNA Duplexes Containing 5-Formyluracil

    Directory of Open Access Journals (Sweden)

    Masaru Tsunoda

    2010-01-01

    Full Text Available Hydroxyl radicals are potent mutagens that attack DNA to form various base and ribose derivatives. One of the major damaged thymine derivatives is 5-formyluracil (fU, which induces pyrimidine transition during replication. In order to establish the structural basis for such mutagenesis, the crystal structures of two kinds of DNA d(CGCGRATfUCGCG with R = A/G have been determined by X-ray crystallography. The fU residues form a Watson-Crick-type pair with A and two types of pairs (wobble and reversed wobble with G, the latter being a new type of base pair between ionized thymine base and guanine base. In silico structural modeling suggests that the DNA polymerase can accept the reversed wobble pair with G, as well as the Watson-Crick pair with A.

  18. DNA Duplex-Based Photodynamic Molecular Beacon for Targeted Killing of Retinoblastoma Cell.

    Science.gov (United States)

    Wei, Yanchun; Lu, Cuixia; Chen, Qun; Xing, Da

    2016-11-01

    Retinoblastoma (RB) is the most common primary intraocular malignancy of infancy. An alternative RB treatment protocol is proposed and tested. It is based on a photodynamic therapy (PDT) with a designed molecular beacon that specifically targets the murine double minute x (MDMX) high-expressed RB cells. A MDMX mRNA triggered photodynamic molecular beacon is designed by binding a photosensitizer molecule (pyropheophorbide-a, or PPa) and a black hole quencher-3 (BHQ3) through a complementary oligonucleotide sequence. Cells with and without MDMX high-expression are incubated with the beacon and then irradiated with a laser. The fluorescence and reactive oxygen species are detected in solution to verify the specific activation of PPa by the perfectly matched DNA targets. The cell viabilities are evaluated with CCK-8 and flow cytometry assay. The fluorescence and photo-cytoxicity of PPa is recovered and significantly higher in the MDMX high-expressed Y79 and WERI-Rb1 cells, compared to that with the MDMX low-expressed cells. The synthesized beacon exhibits high PDT efficiency toward MDMX high-expressed RB cells. The data suggest that the designed beacon may provide a potential alternative for RB therapy and secures the ground for future investigation.

  19. Effect of Secondary Phase Precipitation on the Corrosion Behavior of Duplex Stainless Steels

    Directory of Open Access Journals (Sweden)

    Kai Wang Chan

    2014-07-01

    Full Text Available Duplex stainless steels (DSSs with austenitic and ferritic phases have been increasingly used for many industrial applications due to their good mechanical properties and corrosion resistance in acidic, caustic and marine environments. However, DSSs are susceptible to intergranular, pitting and stress corrosion in corrosive environments due to the formation of secondary phases. Such phases are induced in DSSs during the fabrication, improper heat treatment, welding process and prolonged exposure to high temperatures during their service lives. These include the precipitation of sigma and chi phases at 700–900 °C and spinodal decomposition of ferritic grains into Cr-rich and Cr-poor phases at 350–550 °C, respectively. This article gives the state-of the-art review on the microstructural evolution of secondary phase formation and their effects on the corrosion behavior of DSSs.

  20. Combination of 768-well microplate array diagonal gel electrophoresis with duplex PCR of X and Y chromosome markers for quality control of epidemiological DNA banks.

    Science.gov (United States)

    Huang, Shuwen; Chen, Xiao-he; Day, Ian N M

    2006-08-01

    Large DNA banks for human epidemiological studies have become an increasingly important research tool. The power of genotype-phenotype studies is dependent both on the quality of phenotyping and of genotyping and of correct linking of phenotypes to genotypes. Samples must be tracked through numerous steps between subject or patient and post-genotypic data. Only one phenotype, sex, has a perfect and binary correlation with genotype. In mixed sex studies, it may be advantageous for purposes of quality control to keep sexes mixed during the steps from acquisition to DNA bank, in order to be able to check later for sample swaps. We have designed a duplex PCR combining an amplicon from MAOA marking the X chromosome and an amplicon from DDX3Y marking the Y chromosome. We combined this with a simple economical palmtop sized 768-well microplate compatible electrophoresis system developed in-house for examination of duplex PCR products. We applied this quality control test in the validation of two DNA banks.

  1. Effect of QPQ nitriding time on microstructure and wear resistance of SAF2906 duplex stainless steel

    Science.gov (United States)

    Liu, D.; Wu, G. X.; Shen, L. X.

    2017-01-01

    QPQ salt bath treatment of SAF2906 duplex stainless steel was conducted at 570 °C for 60 min, 90 min,120 min,150 min and 180 min, followed by post-oxidation process with heating temperature of 400°C and holding duration of 30 min. The effect of QPQ nitriding time on microstructure and wear resistance of SAF2906 duplex stainless steel was investigated by means of OM, SEM, XRD, microhardness test, adhesion strength test and wear resistance test. Microstructure observation showed outer layer was composed of Fe3O4. The main phase of the intermediate layer was CrN, αN and Fe2-3N. The main phase of the inner layer was CrN and S. The adhesion strength test of the surface layer-substrate showed the QPQ treated samples have favorable adhesion strength of HF-1 level. With the increase of nitriding time, the growth rate of the compound layer gradually slowed down and the surface hardness first increased and then decreased, and the maximum hardness was 1283 HV0.2 at 150 min. The dry siliding results showed that the wear resistance of the QPQ treated samples was at least 20 times than that of the substrate, and the optimum nitriding time to obtain the best wear resistance is 150 min. The worn surface morphology observation showed the main wear mechanism of the substrate was plough wear, while micro-cutting is the main wear mechanism that causes the damage of the QPQ treated samples.

  2. MD and NMR analyses of choline and TMA binding to duplex DNA: on the origins of aberrant sequence-dependent stability by alkyl cations in aqueous and water-free solvents.

    Science.gov (United States)

    Portella, Guillem; Germann, Markus W; Hud, Nicholas V; Orozco, Modesto

    2014-02-26

    It has been known for decades that alkylammonium ions, such as tetramethyl ammonium (TMA), alter the usual correlation between DNA GC-content and duplex stability. In some cases it is even possible for an AT-rich duplex to be more stable than a GC-rich duplex of the same length. There has been much speculation regarding the origin of this aberration in sequence-dependent DNA duplex stability, but no clear resolution. Using a combination of molecular dynamics simulations and NMR spectroscopy we demonstrate that choline (2-hydroxy-N,N,N-trimethylethanaminium) and TMA are preferentially localized in the minor groove of DNA duplexes at A·T base pairs and these same ions show less pronounced localization in the major groove compared to what has been demonstrated for alkali and alkali earth metal ions. Furthermore, free energy calculations show that single-stranded GC-rich sequences exhibit more favorable solvation by choline than single-stranded AT-rich sequences. The sequence-specific nature of choline and TMA binding provides a rationale for the enhanced stability of AT-rich sequences when alkyl-ammonium ions are used as the counterions of DNA. Our combined theoretical and experimental study provides one of the most detailed pictures to date of cations localized along DNA in the solution state, and provides insights that go beyond understanding alkyl-ammonium ion binding to DNA. In particular, because choline and TMA bind to DNA in a manner that is found to be distinct from that previously reported for Na(+), K(+), Mg(2+), and Ca(2+), our results reveal the important but underappreciated role that most other cations play in sequence-specific duplex stability.

  3. Pitting Corrosion of Super Duplex Stainless Steel - Effect of Isothermal Heat Treament

    OpenAIRE

    Lauritsen, Christian Rene

    2016-01-01

    Super duplex stainless steels (SDSS), with a chromium content of 25 wt$\\%$, contain a duplex structure which consists of ferrite and austenite, and have a pitting resistance equivalent number (PREN) equal or higher than 40. SDSS are affected by the alloying elements, microstructure and fabrication processes. The high degree of alloying elements in SDSS can lead to formation of intermetallic precipitates and secondary phases during heat treatments. Detrimental phases, such as sigma ($\\sigma$) ...

  4. Structural perturbations induced by the alpha-anomer of the aflatoxin B(1) formamidopyrimidine adduct in duplex and single-strand DNA.

    Science.gov (United States)

    Brown, Kyle L; Voehler, Markus W; Magee, Shane M; Harris, Constance M; Harris, Thomas M; Stone, Michael P

    2009-11-11

    The guanine N7 adduct of aflatoxin B(1) exo-8,9-epoxide hydrolyzes to form the formamidopyrimidine (AFB-FAPY) adduct, which interconverts between alpha and beta anomers. The beta anomer is highly mutagenic in Escherichia coli, producing G --> T transversions; it thermally stabilizes the DNA duplex. The AFB-alpha-FAPY adduct blocks replication; it destabilizes the DNA duplex. Herein, the structure of the AFB-alpha-FAPY adduct has been elucidated in 5'-d(C(1)T(2)A(3)T(4)X(5)A(6)T(7)T(8)C(9)A(10))-3'.5'-d(T(11)G(12)A(13)A(14)T(15)C(16)A(17)T(18)A(19)G(20))-3' (X = AFB-alpha-FAPY) using molecular dynamics calculations restrained by NMR-derived distances and torsion angles. The AFB moiety intercalates on the 5' face of the pyrimidine moiety at the damaged nucleotide between base pairs T(4).A(17) and X(5).C(16), placing the FAPY C5-N(5) bond in the R(a) axial conformation. Large perturbations of the epsilon and zeta backbone torsion angles are observed, and the base stacking register of the duplex is perturbed. The deoxyribose orientation shifts to become parallel to the FAPY base and displaced toward the minor groove. Intrastrand stacking between the AFB moiety and the 5' neighbor thymine remains, but strong interstrand stacking is not observed. A hydrogen bond between the formyl group and the exocyclic amine of the 3'-neighbor adenine stabilizes the E conformation of the formamide moiety. NMR studies reveal a similar 5'-intercalation of the AFB moiety for the AFB-alpha-FAPY adduct in the tetramer 5'-d(C(1)T(2)X(3)A(4))-3', involving the R(a) axial conformation of the FAPY C5-N(5) bond and the E conformation of the formamide moiety. Since in duplex DNA the AFB moiety of the AFB-beta-FAPY adduct also intercalates on the 5' side of the pyrimidine moiety at the damaged nucleotide, we conclude that favorable 5'-stacking leads to the R(a) conformational preference about the C5-N(5) bond; the same conformational preference about this bond is also observed at the nucleoside

  5. Binding of Cationic Bis-porphyrins Linked with p- or m-Xylylenediamine and Their Zinc(II Complexes to Duplex DNA

    Directory of Open Access Journals (Sweden)

    Tadayuki Uno

    2008-12-01

    Full Text Available Spectroscopic, viscometric, and molecular docking analysis of binding of cationic bis-porphyrins linked with p- or m-xylylenediamine (H2pXy and H2mXy and their zinc(II complexes (ZnpXy and ZnmXy to duplex DNA are described. H2pXy and H2mXy bound to calf thymus DNA (CTDNA stronger than unichromophoric H2TMPyP, and showed exciton-type induced circular dichroism spectra of their Soret bands. The H2TMPyP-like units of the metal-free bis-porphyrins did not intercalate into CTDNA, and thus the binding mode is outside binding with intramolecular stacking. ZnpXy showed favorable binding to A·T over G·C region, and should lie in the major groove of A·T region.

  6. The sense strand pre-cleaved RNA duplex mediates an efficient RNA interference with less off-target and immune response effects.

    Science.gov (United States)

    Lu, Xiaozhao; Yang, Guodong; Zhang, Jie; Fu, Haiyan; Jin, Liang; Wei, Mengying; Wang, Li; Lu, Zifan

    2011-04-01

    RNA interference is an appealing and promising therapeutic approach in cancer and other diseases. Designing novel strategies aiming to increase the efficiency, duration, and reduce the off-target silencing by sense strand is of great significance for its future application clinically. Here, we report that RNA duplex with the sense strand pre-cleaved at the base between base 10 and 11 relative to the 5' end of the antisense strand induced a target-specific RNA silencing effectively. Furthermore, different from the canonical RNA duplex, this novel RNA duplex rarely inhibits the luciferase activity in the reporter, bearing the target sequence corresponding to the sense strand, suggesting a less off-target effects of this novel strategy. Furthermore, the immune response of the novel RNA duplex induced a much milder immune response as seen from the NFkappaB activity. In addition, our newly designed RNA duplex should be easier for preservation than the asymmetric RNA duplex. Our results establish a novel method to design a new class of RNA duplex for improved RNA interference.

  7. Achieving single channel, full duplex wireless communication

    KAUST Repository

    Choi, Jung Il

    2010-01-01

    This paper discusses the design of a single channel full-duplex wireless transceiver. The design uses a combination of RF and baseband techniques to achieve full-duplexing with minimal effect on link reliability. Experiments on real nodes show the full-duplex prototype achieves median performance that is within 8% of an ideal full-duplexing system. This paper presents Antenna Cancellation, a novel technique for self-interference cancellation. In conjunction with existing RF interference cancellation and digital baseband interference cancellation, antenna cancellation achieves the amount of self-interference cancellation required for full-duplex operation. The paper also discusses potential MAC and network gains with full-duplexing. It suggests ways in which a full-duplex system can solve some important problems with existing wireless systems including hidden terminals, loss of throughput due to congestion, and large end-to-end delays. Copyright 2010 ACM.

  8. Electrochemical Investigation of Interaction between a Bifunctional Probe and GG Mismatch Duplex.

    Science.gov (United States)

    Li, Jiao; He, Hanping; Peng, Xiaoqian; Huang, Min; Zhang, Xiuhua; Wang, Shengfu

    2015-01-01

    A bifunctional probe (FecNC), containing a recognition part and an electrochemical active center, was applied to electrochemical detection of GG mismatch duplexes. The preparation of gold electrodes modified by mismatch and complementatry duplexes was characterized by electrochemical impedance spectroscopy (EIS) and optimized for better detection in terms of self-assembly time, hybridization time, and incubation time. The interaction between FecNC and DNA duplexes modified on the surface of a gold electrode was explored by square wave voltammetry (SWV) and EIS. The results showed that the DNA duplexes with GG mismatch on the surface of a gold electrode was easily detected by the largest electrochemical signal of the bifunctional probe because of its selective binding to GG mismatches. The bifunctional probe could offer a simple, effective electrochemical detection of GG mismatches, and theoretical bases for development of electrochemical biosensors. Further, the method would be favorable for diagnosis of genetic diseases.

  9. DNA secondary structure of the released strand stimulates WRN helicase action on forked duplexes without coordinate action of WRN exonuclease

    Energy Technology Data Exchange (ETDEWEB)

    Ahn, Byungchan, E-mail: bbccahn@mail.ulsan.ac.kr [Department of Life Sciences, University of Ulsan, Ulsan (Korea, Republic of); Bohr, Vilhelm A. [Laboratory of Molecular Gerontology, Biomedical Research Center, National Institute on Aging, Baltimore, MD (United States)

    2011-08-12

    Highlights: {yields} In this study, we investigated the effect of a DNA secondary structure on the two WRN activities. {yields} We found that a DNA secondary structure of the displaced strand during unwinding stimulates WRN helicase without coordinate action of WRN exonuclease. {yields} These results imply that WRN helicase and exonuclease activities can act independently. -- Abstract: Werner syndrome (WS) is an autosomal recessive premature aging disorder characterized by aging-related phenotypes and genomic instability. WS is caused by mutations in a gene encoding a nuclear protein, Werner syndrome protein (WRN), a member of the RecQ helicase family, that interestingly possesses both helicase and exonuclease activities. Previous studies have shown that the two activities act in concert on a single substrate. We investigated the effect of a DNA secondary structure on the two WRN activities and found that a DNA secondary structure of the displaced strand during unwinding stimulates WRN helicase without coordinate action of WRN exonuclease. These results imply that WRN helicase and exonuclease activities can act independently, and we propose that the uncoordinated action may be relevant to the in vivo activity of WRN.

  10. Effect of silver on microstructure and antibacterial property of 2205 duplex stainless steel.

    Science.gov (United States)

    Yang, Sheng-Min; Chen, Yi-Chun; Pan, Yeong-Tsuen; Lin, Dong-Yih

    2016-06-01

    In this study, 2205 duplex stainless steel (DSS) was employed to enhance the antibacterial properties of material through silver doping. The results demonstrated that silver-doped 2205 DSS produces an excellent bacteria-inhibiting effect against Escherichia coli and Staphylococcus aureus. The antibacterial rates were 100% and 99.5%, respectively. Because the mutual solubility of silver and iron is very low in both the solid and liquid states, a silver-rich compound solidified and dispersed at the ferrite/austenite interface and the ferrite, austenite, and secondary austenite phases in silver-doped 2205 DSS. Doping 2205 DSS with silver caused the Creq/Nieq ratio of ferrite to decrease; however, the lower Creq/Nieq ratio promoted the rapid nucleation of γ2-austenite from primary α-ferrite. After 12h of homogenisation treatment at 1200 °C, the solubility of silver in the γ-austenite and α-ferrite phases can be increased by 0.10% and 0.09%, respectively. Moreover, silver doping was found to accelerate the dissolution of secondary austenite in a ferrite matrix during homogenisation.

  11. Duplex real-time PCR method for the detection of sesame (Sesamum indicum) and flaxseed (Linum usitatissimum) DNA in processed food products.

    Science.gov (United States)

    López-Calleja, Inés María; de la Cruz, Silvia; Martín, Rosario; González, Isabel; García, Teresa

    2015-01-01

    The development of a duplex real-time polymerase chain reaction (PCR) method allowing the simultaneous detection of sesame and flaxseed DNA in commercial food products is described. This duplex real-time PCR technique is based in the design of sesame- and flaxseed-specific primers based on the ITS1 region and two TaqMan fluorescent probes. The method was positive for sesame and flaxseed, and showed no cross-reactivity for all other heterologous plant and animal species tested. Sesame and flaxseed could be detected in a series of model samples with defined raw and heat-treated sesame in flaxseed, and flaxseed in sesame, respectively, with detection limits of 1.3 mg kg(-1) for sesame and 1.4 mg kg(-1) for flaxseed. The applicability of the assay for determining sesame and flaxseed in different food matrices was investigated by analysing a total of 238 commercial foodstuffs. This PCR method is useful for highly selective and sensitive detection of traces of sesame and flaxseed in commercial food products.

  12. Effects of Severe Plastic Deformation and Heat Treatment on Transformation Behavior of Explosively Welded Duplex TiNi-TiNi

    Institute of Scientific and Technical Information of China (English)

    Li Juntao; Zheng Yanjun; Cui Lishan

    2007-01-01

    The effects of severe plastic deformation and heat treatment on the transformation behavior of explosively welded duplex TiNi-TiNi shape memory alloys (SMAs) were investigated by differential scanning calorimeter (DSC) measurements. The explosively welded duplex TiNi-TiNi plate of 0.7 mm thickness was cold-rolled at room temperature to a 60% reduction in thickness and then annealed at different temperatures for different durations. The results showed that low temperature (623-723K) heat-treatment led to the crystallization of the amorphous region, and re-crystallization occurred in the specimens annealed at higher temperatures (over 873 K). Research indicated that the change of martensitic transformation temperature is due to the change of internal stresses with increasing heat treatment temperature. The change of annealing time also led to a change in martensitic transformation temperature, which was associated with the precipitation and decomposition of Ti3Ni4 in TiNi-1.

  13. An exploration of sequence specific DNA-duplex/pyrene interactions for intercalated and surface-associated pyrene species. Final report, May 1, 1993--December 31, 1996

    Energy Technology Data Exchange (ETDEWEB)

    Netzel, T.L.

    1997-03-01

    The broad objective of this DOE sponsored work on photoinduced electron transfer (ET) within covalently modified DNA was to learn about the rates of Et among various DNA bases and commonly used organic electron donor (D) and acceptor (A) molecules. This hypothesis driven, multidisciplinary project combined skills in modified nucleic acid synthesis and in continuous and time-resolved optical spectroscopies. Covalently modified DNA chemistry as investigated in this program had two specific long term goals. The first was to use experimental and theoretical insights into the mechanisms of electron transfer (ET) reactions to design supramolecular assemblies of redox-active chromophores that function as efficient vectorial ET engines. The second was to construct oligonucleotide probes for real-time monitoring of intracellular processes involving DNA and RNA such as m-RNA expression and translocation. This research project laid the groundwork for studying ET reactions within DNA duplexes by examining the photophysics of uridine nucleosides which are covalently labeled at the 5-position with 1-pyrenyl chromophores.

  14. Field Effect Transistor Using Carbon Nanotubes and DNA as Electrical Gate

    Science.gov (United States)

    Abdalla, S.; Al-Marzouki, F. M.; Al-Ghamdi, Ahmed A.

    2017-02-01

    We present an electronic sensor in the molecular scale, which is very sensitive for detection and sensing of DNA characteristics and DNA activities in particular activities between DNA duplex and any protein. Here, the device shows that DNA is electronically inserted to be on the same time as an electrical device transducer and as a biological target in a carbon nanotube-DNA-carbon nanotube electronic sensor. We have performed a DNA binding through an amide group by the electron transfer through amide group. The presented device has shown an efficient and rapid procedure to bind the electrical vulnerability of DNA with the detection of enzymatic effectiveness leading to high efficient biosensor.

  15. Protective effect of enzymatic extracts from microalgae against DNA damage induced by H2O2.

    Science.gov (United States)

    Karawita, Rohan; Senevirathne, Mahinda; Athukorala, Yasantha; Affan, Abu; Lee, Young-Jae; Kim, Se-Kwon; Lee, Joon-Baek; Jeon, You-Jin

    2007-01-01

    The enzymatic extracts from seven species of microalgae (Pediastrum duplex, Dactylococcopsis fascicularis, Halochlorococcum porphyrae, Oltmannsiellopsis unicellularis, Achnanthes longipes, Navicula sp. and Amphora coffeaeformis) collected from three habitats (freshwater, tidal pool, and coastal benthic) at Jeju Island in Korea were investigated for their antioxidant activity. Of the extracts tested, the AMG 300 L (an exo 1, 4-alpha-D-glucosidase) extract of P. duplex, the Viscozyme extract of Navicula sp., and the Celluclast extract of A. longipes provided the most potential as antioxidants. Meanwhile, the Termamyl extract of P. duplex in an H(2)O(2) scavenging assay exhibited an approximate 60% scavenging effect. In this study, we report that the DNA damage inhibitory effects of P. duplex (Termamyl extract) and D. fascicularis (Kojizyme extract) were nearly 80% and 69% respectively at a concentration of 100 microg/ml. Thus, it is suggested that the microalgae tested in this study yield promising DNA damage inhibitory properties on mouse lymphoma L 5178 cells that are treated with H(2)O(2). Therefore, microalgae such as P. duplex may be an excellent source of naturally occurring antioxidant compounds with potent DNA damage inhibition potential.

  16. Study of corrosive effect of oil in super duplex stainless steels; Estudo do efeito corrosivo do petroleo em acos super duplex

    Energy Technology Data Exchange (ETDEWEB)

    Gusmao, E.F.; Azambuja, V.M. [IFES, Coordenadoria de Metalurgia, Vitoria, ES (Brazil); Santos, D.S. [Universidade Federal do Rio de Janeiro (PEMM/COPPE/UFRJ), RJ (Brazil). Pos-Graduacao em Engenharia Metalurgica e de Materiais

    2010-07-01

    The super duplex stainless steel was exposed in an environment at 75 degree C with oil for days, weeks and month to observe the change in mass. The corrosion leads to loss of weight of material which could harm the economy of a company, as this will have to stop production to replace the corroded part. Hence the great importance of studies on ways to mitigate the corrosion. There was a chemical attack by the reagent Behara and testing to study the quality of the protective coating after the tests with oil by electrochemical impedance. (author)

  17. Effects of fabrication conditions on mechanical properties and microstructure of duplex β″-Al{sub 2}O{sub 3} solid electrolyte

    Energy Technology Data Exchange (ETDEWEB)

    Canfield, Nathan L. [Energy Processes & Materials Division, Pacific Northwest National Laboratory, Richland, WA 99352 (United States); Kim, Jin Y., E-mail: Jin.Kim@pnnl.gov [Energy Processes & Materials Division, Pacific Northwest National Laboratory, Richland, WA 99352 (United States); Bonnett, Jeff F.; Pearson, R.L.; Sprenkle, Vincent L. [Energy Processes & Materials Division, Pacific Northwest National Laboratory, Richland, WA 99352 (United States); Jung, Keeyoung [Energy Storage Materials Research Center, Research Institute of Industrial Science and Technology (RIST), Pohang (Korea, Republic of)

    2015-07-15

    Highlights: • The concept of duplex BASEs is presented as a method to lower the ASR for NBBs. • Duplex BASEs consist of thin dense electrolyte and porous support. • Strength of converted BASEs shows a different trend from as-sintered samples. • Cell orientation gives significant impact on strength of duplex BASEs. - Abstract: Na-beta batteries are an attractive technology as a large-scale electrical energy storage for grid applications. However, additional improvements in performance and cost are needed for wide market penetration. To improve cell performance by minimizing polarizations, reduction of electrolyte thickness was attempted using a duplex structure consisting of a thin dense electrolyte layer and a porous support layer. In this paper, the effects of sintering conditions, dense electrolyte thickness, and cell orientation on the flexural strength of duplex BASEs fabricated using a vapor phase approach were investigated. It is shown that sintering at temperatures between 1500 and 1550 °C results in fine grained microstructures and the highest flexural strength after conversion. Increasing thickness of the dense electrolyte has a small impact on flexural strength, while the orientation of load such that the dense electrolyte is in tension instead of compression has major effects on strength for samples with a well-sintered dense electrolyte.

  18. Effects of prior solution treatment on thermal aging behavior of duplex stainless steels

    Energy Technology Data Exchange (ETDEWEB)

    Li, Shilei; Wang, Yanli; Zhang, Hailong; Li, Shuxiao [State Key Laboratory for Advanced Metals and Materials, University of Science and Technology Beijing, Beijing 100083 (China); Wang, Genqi [Yantai Taihai Marnoir Nuclear Equipment Co. Ltd., Yantai 264003 (China); Wang, Xitao, E-mail: xtwang@ustb.edu.cn [State Key Laboratory for Advanced Metals and Materials, University of Science and Technology Beijing, Beijing 100083 (China)

    2013-10-15

    The influence of solution temperature on thermal aging behavior was studied in duplex stainless steels. With increasing solution temperature, the ferrite contents remarkably increase, Cr and Ni elements redistribute. During thermal aging, the impact properties of higher solution temperature treated materials suffer a serious degradation, which is not only related with ferrite content but also the alloy compositions in ferrite. Enrichment of Ni in ferrite can accelerate the spinodal decomposition kinetics. Thermal aging-inducing strain fields in ferrite cause the embrittlement of DSS.

  19. Postsynthetic modification of oligonucleotides with imidazophenazine dye and its effect on duplex stability.

    Science.gov (United States)

    Dubey, Larysa; Ryazanova, Olga; Zozulya, Victor; Fedoryak, Dmytro; Dubey, Igor

    2011-01-01

    Carboxyalkyl derivative of the intercalating agent imidazo[4,5-b]phenazine was used for the postsynthetic oligonucleotide modification. Model pentadecathymidylate-imidazophenazine conjugate was prepared from 5'-aminoalkyl-modified (dT)(15) by using phosphonium coupling reagent BOP in the presence of 1-hydroxybenzotriazole. Spectral-fluorescent properties of the conjugate were studied. The attachment of the dye was found to increase the thermal stability of (dT)(15) duplex with poly(dA) by more than 4°C, probably by the intercalation mechanism.

  20. Effect of Aging on Precipitation Behavior and Pitting Corrosion Resistance of SAF2906 Super Duplex Stainless Steel

    Science.gov (United States)

    Li, Jianchun; Li, Guoping; Liang, Wei; Han, Peide; Wang, Hongxia

    2017-09-01

    The effect of aging temperature and holding time on the precipitation of secondary phases and pitting corrosion resistance of SAF2906 super duplex stainless steel was examined. Chromium nitride and σ phase were observed to preferentially precipitate at the ferrite/austenite interface. An amount of nitrides was also observed within the ferrite grain. The precipitation of chromium nitride occurred before the σ phase. The increase in aging temperature and holding time did not affect the concentration of the nitrides but increased the area fraction of the σ phase at a faster rate. The Cr2N precipitation in SAF2906 is more evident than that of the other duplex stainless steels. The variation tendency of the precipitation concentrations is primarily consistent with the prediction results of Thermo-Calc software. The electrochemical results showed that Cr2N and σ phase significantly reduced the pitting potential. Scanning electron microscope observations revealed that pits appear mainly in regions adjacent to sigma phase and Cr2N.

  1. Effects of Cold Rolling and Strain-Induced Martensite Formation in a SAF 2205 Duplex Stainless Steel

    Science.gov (United States)

    Breda, Marco; Brunelli, Katya; Grazzi, Francesco; Scherillo, Antonella; Calliari, Irene

    2015-02-01

    Duplex stainless steels (DSSs) are biphasic steels having a ferritic-austenitic microstructure that allows them to combine good mechanical and corrosion-resistance properties. However, these steels are sensitive to microstructural modifications, such as ferrite decomposition at high temperatures and the possibility of strain-induced martensite (SIM) formation from cold-worked austenite, which can significantly alter their interesting features. In the present work, the effects of cold rolling on the developed microstructural features in a cold-rolled SAF 2205 DSS and the onset of martensitic transformation are discussed. The material was deformed at room temperature from 3 to 85 pct thickness reduction, and several characterization techniques (scanning and transmission electron microscopy, X-ray diffraction, hardness measurements, and time-of-flight-neutron diffraction) were employed in order to fully describe the microstructural behavior of the steel. Despite the low stacking fault energy of DSS austenite, which contributed to SIM formation, the steel was found to be more stable than other stainless steel grades, such as AISI 304L. Rolling textures were similar to those pertaining to single-phase materials, but the presence of the biphasic (Duplex) microstructure imposed deformation constraints that affected the developed microstructural features, owing to phases interactions. Moreover, even if an intensification of the strain field in austenite was revealed, retarded SIM transformation kinetics and lower martensite amounts with respect to AISI 304L were observed.

  2. Effect of electromagnetic interaction during fusion welding of AISI 2205 duplex stainless steel on the corrosion resistance

    Science.gov (United States)

    García-Rentería, M. A.; López-Morelos, V. H.; González-Sánchez, J.; García-Hernández, R.; Dzib-Pérez, L.; Curiel-López, F. F.

    2017-02-01

    The effect of electromagnetic interaction of low intensity (EMILI) applied during fusion welding of AISI 2205 duplex stainless steel on the resistance to localised corrosion in natural seawater was investigated. The heat affected zone (HAZ) of samples welded under EMILI showed a higher temperature for pitting initiation and lower dissolution under anodic polarisation in chloride containing solutions than samples welded without EMILI. The EMILI assisted welding process developed in the present work enhanced the resistance to localised corrosion due to a modification on the microstructural evolution in the HAZ and the fusion zone during the thermal cycle involved in fusion welding. The application of EMILI reduced the size of the HAZ, limited coarsening of the ferrite grains and promoted regeneration of austenite in this zone, inducing a homogeneous passive condition of the surface. EMILI can be applied during fusion welding of structural or functional components of diverse size manufactured with duplex stainless steel designed to withstand aggressive environments such as natural seawater or marine atmospheres.

  3. An in vitro DNA double-strand break repair assay based on end-joining of defined duplex oligonucleotides.

    Science.gov (United States)

    Datta, Kamal; Purkayastha, Shubhadeep; Neumann, Ronald D; Winters, Thomas A

    2012-01-01

    DNA double-strand breaks (DSBs) are caused by endogenous cellular processes such as oxidative metabolism, or by exogenous events like exposure to ionizing radiation or other genotoxic agents. Repair of these DSBs is essential for the maintenance of cellular genomic integrity. In human cells, and cells of other higher eukaryotes, DSBs are primarily repaired by the nonhomologous end-joining (NHEJ) DSB repair pathway. Most in vitro assays that have been designed to measure NHEJ activity employ linear plasmid DNA as end-joining substrates, and such assays have made significant contributions to our understanding of the biochemical mechanisms of NHEJ. Here we describe an in vitro end-joining assay employing linear oligonucleotides that has distinct advantages over plasmid-based assays for the study of structure-function relationships between the proteins of the NHEJ pathway and synthetic DNA end-joining substrates possessing predetermined DSB configurations and chemistries.

  4. Development of bis-locked nucleic acid (bisLNA) oligonucleotides for efficient invasion of supercoiled duplex DNA

    DEFF Research Database (Denmark)

    Moreno, Pedro M D; Geny, Sylvain; Pabon, Y Vladimir;

    2013-01-01

    In spite of the many developments in synthetic oligonucleotide (ON) chemistry and design, invasion into double-stranded DNA (DSI) under physiological salt and pH conditions remains a challenge. In this work, we provide a new ON tool based on locked nucleic acids (LNAs), designed for strand invasi...

  5. Effect of gold nanoparticle on stability of the DNA molecule: A study of molecular dynamics simulation.

    Science.gov (United States)

    Izanloo, Cobra

    2017-09-26

    An understanding of the mechanism of DNA interactions with gold nanoparticles is useful in today medicine applications. We have performed a molecular dynamics simulation on a B-DNA duplex (CCTCAGGCCTCC) in the vicinity of a gold nanoparticle with a truncated octahedron structure composed of 201 gold atoms (diameter ∼1.8 nm) to investigate gold nanoparticle (GNP) effects on the stability of DNA. During simulation, the nanoparticle is closed to DNA and phosphate groups direct the particles into the major grooves of the DNA molecule. Because of peeling and untwisting states that are occur at end of DNA, the nucleotide base lies flat on the surface of GNP. The configuration entropy is estimated using the covariance matrix of atom-positional fluctuations for different bases. The results show that when a gold nanoparticle has interaction with DNA, entropy increases. The results of conformational energy and the hydrogen bond numbers for DNA indicated that DNA becomes unstable in the vicinity of a gold nanoparticle. The radial distribution function was calculated for water hydrogen-phosphate oxygen pairs. Almost for all nucleotide, the presence of a nanoparticle around DNA caused water molecules to be released from the DNA duplex and cations were close to the DNA.

  6. Novel interstrand communication systems within DNA duplexes based on 1-, 2- and 4-(phenylethynyl)pyrenes attached to 2′-amino-LNA: high-affinity hybridization and fluorescence sensing

    DEFF Research Database (Denmark)

    Astakhova, Irina; Lindegaard, Dorthe; Korshun, Vladimir A.

    2010-01-01

    Functionalisation of 2′-amino-LNA oligonucleotides with 1-, 2- and 4-(phenylethynyl)pyrene fluorophores via a carbonyl linker (PEPyc) resulted in efficient interstrand communication systems in nucleic acid duplexes, providing effective tools for stabilization of nanostructures and fluorescence...

  7. Mechanical and Microstructural Effects of Thermal Aging on Cast Duplex Stainless Steels by Experiment and Finite Element Method

    Energy Technology Data Exchange (ETDEWEB)

    Schwarm, Samuel C.; Mburu, Sarah N.; Kolli, Ratna P.; Perea, Daniel E.; Liu, Jia; Ankem, Sreeramamurthy

    2017-02-05

    Cast duplex stainless steel piping in light water nuclear reactors expe- rience thermal aging embrittlement during operational service. Interest in extending the operational life to 80 years requires an increased understanding of the microstructural evolution and corresponding changes in mechanical behavior. We analyze the evolution of the microstructure during thermal aging of cast CF-3 and CF-8 stainless steels using electron microscopy and atom probe tomography. The evolution of the mechanical properties is measured concurrently by mechanical methods such as tensile tests, Charpy V-notch tests, and instrumented nanoinden- tation. A microstructure-based finite element method model is developed and uti- lized in conjunction with the characterization results in order to correlate the local stress-strain effects in the microstructure with the bulk measurements. This work is supported by the DOE Nuclear Energy University Programs (NEUP), contract number DE-NE0000724.

  8. Computing DNA duplex instability profiles efficiently with a two-state model: trends of promoters and binding sites

    Directory of Open Access Journals (Sweden)

    Rapti Zoi

    2010-12-01

    Full Text Available Abstract Background DNA instability profiles have been used recently for predicting the transcriptional start site and the location of core promoters, and to gain insight into promoter action. It was also shown that the use of these profiles can significantly improve the performance of motif finding programs. Results In this work we introduce a new method for computing DNA instability profiles. The model that we use is a modified Ising-type model and it is implemented via statistical mechanics. Our linear time algorithm computes the profile of a 10,000 base-pair long sequence in less than one second. The method we use also allows the computation of the probability that several consecutive bases are unpaired simultaneously. This is a feature that is not available in other linear-time algorithms. We use the model to compare the thermodynamic trends of promoter sequences of several genomes. In addition, we report results that associate the location of local extrema in the instability profiles with the presence of core promoter elements at these locations and with the location of the transcription start sites (TSS. We also analyzed the instability scores of binding sites of several human core promoter elements. We show that the instability scores of functional binding sites of a given core promoter element are significantly different than the scores of sites with the same motif occurring outside the functional range (relative to the TSS. Conclusions The time efficiency of the algorithm and its genome-wide applications makes this work of broad interest to scientists interested in transcriptional regulation, motif discovery, and comparative genomics.

  9. Effect of aging temperature on phase decomposition and mechanical properties in cast duplex stainless steels

    Energy Technology Data Exchange (ETDEWEB)

    Mburu, Sarah; Kolli, R. Prakash; Perea, Daniel E.; Schwarm, Samuel C.; Eaton, Arielle; Liu, Jia; Patel, Shiv; Bartrand, Jonah; Ankem, Sreeramamurthy

    2017-04-01

    The microstructure and mechanical properties in unaged and thermally aged (at 280 oC, 320 oC, 360 oC, and 400 oC to 4300 h) CF–3 and CF–8 cast duplex stainless steels (CDSS) are investigated. The unaged CF–8 steel has Cr-rich M23C6 carbides located at the δ–ferrite/γ– austenite heterophase interfaces that were not observed in the CF–3 steel and this corresponds to a difference in mechanical properties. Both unaged steels exhibit incipient spinodal decomposition into Fe-rich α–domains and Cr-rich α’–domains. During aging, spinodal decomposition progresses and the mean wavelength (MW) and mean amplitude (MA) of the compositional fluctuations increase as a function of aging temperature. Additionally, G–phase precipitates form between the spinodal decomposition domains in CF–3 at 360 oC and 400 oC and in CF–8 at 400 oC. The microstructural evolution is correlated to changes in mechanical properties.

  10. Effect of microstructure on mechanical properties of a thin-walled cast duplex steel

    Energy Technology Data Exchange (ETDEWEB)

    Petzold, L.; Kreschel, T.; Peisker, D. [TU Bergakademie Freiberg, Institut fuer Eisen- und Stahltechnologie, Freiberg (Germany); Minnich, D. [Evosteel GmbH, Leipzig (Germany)

    2010-06-15

    The paper investigates the microstructure and the resulting mechanical properties of a duplex steel cast in sand dead-moulds. The chemical composition and the cooling rate are the main parameters affecting the properties. The chemical composition influences the thermodynamics of the phase transformation, the cooling rate determines the kinetics of formation of the microstructure. The latter varies with changes in wall thickness (investigated from 2 to 7 mm) and the position of the material within the casting. Through heat treatment, the composition of the microstructural components can be changed selectively afterwards, thereby the properties are improved. The correlation between microstructure and mechanical properties is explained quantitatively. Additionally to the austenite-ferrite ratio, the dispersion of microstructure has a large influence on the mechanical properties. Ranges of chemical composition and heat treatment parameters are identified where third phases, such as carbides and {sigma}-phase tend to occur. These influence the properties very sensitively even in small amounts. Finally, structural parameters are recognized that will lead to optimal combinations of properties. With an appropriate heat treatment technique, in particular the ductility properties are further increased. (Abstract Copyright [2010], Wiley Periodicals, Inc.)

  11. Purification, crystallization and preliminary X-ray analysis of the DNA-binding domain of AdpA, the central transcription factor in the A-factor regulatory cascade in the filamentous bacterium Streptomyces griseus, in complex with a duplex DNA.

    Science.gov (United States)

    Yao, Ming Dong; Miyazono, Ken-ichi; Ohtsuka, Jun; Hirano, Setsu; Nagata, Koji; Horinouchi, Sueharu; Ohnishi, Yasuo; Tanokura, Masaru

    2012-08-01

    Streptomyces griseus AdpA is the central transcription factor in the A-factor regulatory cascade and activates a number of genes that are required for both secondary metabolism and morphological differentiation, leading to the onset of streptomycin biosynthesis as well as aerial mycelium formation and sporulation. The DNA-binding domain of AdpA consists of two helix-turn-helix DNA-binding motifs and shows low nucleotide-sequence specificity. To reveal the molecular basis of the low nucleotide-sequence specificity, an attempt was made to obtain cocrystals of the DNA-binding domain of AdpA and several kinds of duplex DNA. The best diffracting crystal was obtained using a 14-mer duplex DNA with two-nucleotide overhangs at the 5'-ends. The crystal diffracted X-rays to 2.8 Å resolution and belonged to space group C222(1), with unit-cell parameters a = 76.86, b = 100.96, c = 101.25 Å. The Matthews coefficient (V(M) = 3.71 Å(3) Da(-1)) indicated that the crystal was most likely to contain one DNA-binding domain of AdpA and one duplex DNA in the asymmetric unit, with a solvent content of 66.8%.

  12. Thermal treatments effect on the austenite-ferrite equilibrium in a duplex stainless steel weld beads; Effet des traitements thermiques sur l equilibre austenite - ferrite dans un cordon de soudure en acier inoxydable duplex

    Energy Technology Data Exchange (ETDEWEB)

    Belkessa, Brahim; Badji, Riad; Bettahar, Kheireddine; Maza, Halim [Division de la Metallurgie et Mecanique. Centre de Recherche Scientifique et Technique en Soudage et Controle. B.P 64, Route de Dely Ibrahim, Cheraga, Alger (Algeria)

    2006-07-01

    Heat treatments in the temperature range between 800 to 1200 C, with a keeping at high temperature of 60 min, followed by a water quenching at 20 C, have been carried out on austeno-ferritic stainless steel welds (of type SAF 2205-UNS S31803). The heat treatments carried out at temperatures below 1000 C have modified the structure of the duplex stainless steel 2205 in inducing the formation of precipitates, identified by X-ray diffraction as being the intermetallic compound {sigma} and the chromium carbides M{sub 23}C{sub 6}. The treatments applied to temperatures superior to 1000 C shift the {delta}-{gamma} equilibrium towards the {delta} phase. Indeed, the increase of the ferrite rate with the treatment temperature is approximately linear. The ferrite rates are higher in the heat-affected zone, which has been submitted to a ferritizing due to the welding thermal effects. (O.M.)

  13. Isolation, characterization and molecular cloning of Duplex-Specific Nuclease from the hepatopancreas of the Kamchatka crab

    Directory of Open Access Journals (Sweden)

    Vagner Laura L

    2008-05-01

    Full Text Available Abstract Background Nucleases, which are key components of biologically diverse processes such as DNA replication, repair and recombination, antiviral defense, apoptosis and digestion, have revolutionized the field of molecular biology. Indeed many standard molecular strategies, including molecular cloning, studies of DNA-protein interactions, and analysis of nucleic acid structures, would be virtually impossible without these versatile enzymes. The discovery of nucleases with unique properties has often served as the basis for the development of modern molecular biology methods. Thus, the search for novel nucleases with potentially exploitable functions remains an important scientific undertaking. Results Using degenerative primers and the rapid amplification of cDNA ends (RACE procedure, we cloned the Duplex-Specific Nuclease (DSN gene from the hepatopancreas of the Kamchatka crab and determined its full primary structure. We also developed an effective method for purifying functional DSN from the crab hepatopancreas. The isolated enzyme was highly thermostable, exhibited a broad pH optimum (5.5 – 7.5 and required divalent cations for activity, with manganese and cobalt being especially effective. The enzyme was highly specific, cleaving double-stranded DNA or DNA in DNA-RNA hybrids, but not single-stranded DNA or single- or double-stranded RNA. Moreover, only DNA duplexes containing at least 9 base pairs were effectively cleaved by DSN; shorter DNA duplexes were left intact. Conclusion We describe a new DSN from Kamchatka crab hepatopancreas, determining its primary structure and developing a preparative method for its purification. We found that DSN had unique substrate specificity, cleaving only DNA duplexes longer than 8 base pairs, or DNA in DNA-RNA hybrids. Interestingly, the DSN primary structure is homologous to well-known Serratia-like non-specific nucleases structures, but the properties of DSN are distinct. The unique substrate

  14. Effect of Annealing Temperature on the Mechanical and Corrosion Behavior of a Newly Developed Novel Lean Duplex Stainless Steel

    Directory of Open Access Journals (Sweden)

    Yanjun Guo

    2014-09-01

    Full Text Available The effect of annealing temperature (1000–1150 °C on the microstructure evolution, mechanical properties, and pitting corrosion behavior of a newly developed novel lean duplex stainless steel with 20.53Cr-3.45Mn-2.08Ni-0.17N-0.31Mo was studied by means of optical metallographic microscopy (OMM, scanning electron microscopy (SEM, magnetic force microscopy (MFM, scanning Kelvin probe force microscopy (SKPFM, energy dispersive X-ray spectroscopy (EDS, uniaxial tensile tests (UTT, and potentiostatic critical pitting temperature (CPT. The results showed that tensile and yield strength, as well as the pitting corrosion resistance, could be degraded with annealing temperature increasing from 1000 up to 1150 °C. Meanwhile, the elongation at break reached the maximum of 52.7% after annealing at 1050 °C due to the effect of martensite transformation induced plasticity (TRIP. The localized pitting attack preferentially occurred at ferrite phase, indicating that the ferrite phase had inferior pitting corrosion resistance as compared to the austenite phase. With increasing annealing temperature, the pitting resistance equivalent number (PREN of ferrite phase dropped, while that of the austenite phase rose. Additionally, it was found that ferrite possessed a lower Volta potential than austenite phase. Moreover, the Volta potential difference between ferrite and austenite increased with the annealing temperature, which was well consistent with the difference of PREN.

  15. Effect of Annealing Temperature on the Mechanical and Corrosion Behavior of a Newly Developed Novel Lean Duplex Stainless Steel.

    Science.gov (United States)

    Guo, Yanjun; Hu, Jincheng; Li, Jin; Jiang, Laizhu; Liu, Tianwei; Wu, Yanping

    2014-09-12

    The effect of annealing temperature (1000-1150 °C) on the microstructure evolution, mechanical properties, and pitting corrosion behavior of a newly developed novel lean duplex stainless steel with 20.53Cr-3.45Mn-2.08Ni-0.17N-0.31Mo was studied by means of optical metallographic microscopy (OMM), scanning electron microscopy (SEM), magnetic force microscopy (MFM), scanning Kelvin probe force microscopy (SKPFM), energy dispersive X-ray spectroscopy (EDS), uniaxial tensile tests (UTT), and potentiostatic critical pitting temperature (CPT). The results showed that tensile and yield strength, as well as the pitting corrosion resistance, could be degraded with annealing temperature increasing from 1000 up to 1150 °C. Meanwhile, the elongation at break reached the maximum of 52.7% after annealing at 1050 °C due to the effect of martensite transformation induced plasticity (TRIP). The localized pitting attack preferentially occurred at ferrite phase, indicating that the ferrite phase had inferior pitting corrosion resistance as compared to the austenite phase. With increasing annealing temperature, the pitting resistance equivalent number (PREN) of ferrite phase dropped, while that of the austenite phase rose. Additionally, it was found that ferrite possessed a lower Volta potential than austenite phase. Moreover, the Volta potential difference between ferrite and austenite increased with the annealing temperature, which was well consistent with the difference of PREN.

  16. Detection of HIV proviral DNA by a duplex fluorescence PCR for early diagnosis of HIV infection in infants%双重荧光PCR检测HIV前病毒DNA及其在婴幼儿HIV感染早期诊断中的应用

    Institute of Scientific and Technical Information of China (English)

    张佳峰; 郭志宏; 黄晶晶; 丁晓贝; 黄蓓

    2013-01-01

    Objective To establish a duplex fluorescence PCR for detection of HIV proviral DNA and to evaluate its application for early diagnosis of HIV infection in infants .Methods A duplex fluores-cence PCR system was set up based on TaqMan technology for detection of human ribonuclease P ( RNase P) gene and long terminal repeat ( LTR) region of HIV.A recombinant plasmid containing the targeted gene fragment , pTG19-T, was constructed by TA cloning technique and used as the template for evaluation of sen -sitivity of the assay .Blood samples from 11 healthy individuals and 98 HIV-infected patients were collected and detected to validate the assay specificity .The assay of duplex fluorescence PCR was then carried out to detect 96 infant blood samples collected from several maternal and child health hospitals in Zhejiang province from January 2011 to September 2012 for early diagnosis of HIV infection .The results were compared with those by using the Roche HIV DNA qualitative detection kit .Results The established duplex fluorescence PCR could specifically detect HIV proviral DNA with a specificity of 100%and a detection sensitivity of 100 cps per reaction .The coincidence rate between the established assay and the Roche HIV DNA qualitative de -tection kit was 100%in the detection of 96 blood samples .Conclusion The duplex fluorescence PCR as-say showed advantages of cost-effectiveness , convenience , good specificity and accuracy with high sensitivi-ty.It could be used for early diagnosis of HIV infection in infants and also as a general technical platform for the detection of HIV proviral DNA .%  目的建立双重荧光PCR检测HIV前病毒DNA的方法,并应用于婴幼儿HIV感染的早期诊断。方法采用TaqMan技术,组建针对人类核糖核酸酶P( RNase P)和HIV的长末端重复序列( LTR)基因的双重荧光PCR体系;采用TA克隆技术构建pTG19-T重组质粒作为模板进行该方法灵敏度的评价;采用11

  17. Effect of Austenitic and Austeno-Ferritic Electrodes on 2205 Duplex and 316L Austenitic Stainless Steel Dissimilar Welds

    Science.gov (United States)

    Verma, Jagesvar; Taiwade, Ravindra V.

    2016-09-01

    This study addresses the effect of different types of austenitic and austeno-ferritic electrodes (E309L, E309LMo and E2209) on the relationship between weldability, microstructure, mechanical properties and corrosion resistance of shielded metal arc welded duplex/austenitic (2205/316L) stainless steel dissimilar joints using the combined techniques of optical, scanning electron microscope, energy-dispersive spectrometer and electrochemical. The results indicated that the change in electrode composition led to microstructural variations in the welds with the development of different complex phases such as vermicular ferrite, lathy ferrite, widmanstatten and intragranular austenite. Mechanical properties of welded joints were diverged based on compositions and solidification modes; it was observed that ferritic mode solidified weld dominated property wise. However, the pitting corrosion resistance of all welds showed different behavior in chloride solution; moreover, weld with E2209 was superior, whereas E309L exhibited lower resistance. Higher degree of sensitization was observed in E2209 weld, while lesser in E309L weld. Optimum ferrite content was achieved in all welds.

  18. Effect of rolling deformation and solution treatment on microstructure and mechanical properties of a cast duplex stainless steel

    Indian Academy of Sciences (India)

    S K Ghosh; D Mahata; R Roychaudhuri; R Mondal

    2012-10-01

    The present study deals with the effect of rolling deformation and solution treatment on the microstructure and mechanical properties of a cast duplex stainless steel. Cast steel reveals acicular/Widmanstätten morphology as well as island of austenite within the -ferrite matrix. Hot rolled samples exhibit the presence of lower volume percent of elongated band of -ferrite (∼40%) and austenite phase which convert into finer and fragmented microstructural constituents after 30% cold deformation. By the solution treatment, the elongated and broken crystalline grains recrystallize which leads to the formation of finer grains (<10 m) of austenite. X-ray diffraction analysis has corroborated well with the above-mentioned microstructural investigation. Enhancement in hardness, yield strength and tensile strength values as well as drop in percent elongation with cold deformation increases its suitability for use in thinner sections. 30% cold rolled and solution treated sample reveals attractive combination of strength and ductility (25.22 GPa%). The examination of fracture surface also substantiates the tensile results. The sub-surface micrographs provide the potential sites for initiation of microvoids.

  19. Effect of Austenitic and Austeno-Ferritic Electrodes on 2205 Duplex and 316L Austenitic Stainless Steel Dissimilar Welds

    Science.gov (United States)

    Verma, Jagesvar; Taiwade, Ravindra V.

    2016-11-01

    This study addresses the effect of different types of austenitic and austeno-ferritic electrodes (E309L, E309LMo and E2209) on the relationship between weldability, microstructure, mechanical properties and corrosion resistance of shielded metal arc welded duplex/austenitic (2205/316L) stainless steel dissimilar joints using the combined techniques of optical, scanning electron microscope, energy-dispersive spectrometer and electrochemical. The results indicated that the change in electrode composition led to microstructural variations in the welds with the development of different complex phases such as vermicular ferrite, lathy ferrite, widmanstatten and intragranular austenite. Mechanical properties of welded joints were diverged based on compositions and solidification modes; it was observed that ferritic mode solidified weld dominated property wise. However, the pitting corrosion resistance of all welds showed different behavior in chloride solution; moreover, weld with E2209 was superior, whereas E309L exhibited lower resistance. Higher degree of sensitization was observed in E2209 weld, while lesser in E309L weld. Optimum ferrite content was achieved in all welds.

  20. The effect of chloride ions on the corroded surface layer of 00Cr22Ni5Mo3N duplex stainless steel under cavitation.

    Science.gov (United States)

    Wan, Tong; Xiao, Ning; Shen, Hanjie; Yong, Xingyue

    2016-11-01

    The effects of Cl(-) on the corroded surface layer of 00Cr22Ni5Mo3N duplex stainless steel under cavitation in chloride solutions were investigated using nanoindentation in conjunction with XRD and XPS. The results demonstrate that Cl(-) had a strong effect on the nano-mechanical properties of the corroded surface layer under cavitation, and there was a threshold Cl(-) concentration. Furthermore, a close relationship between the nano-mechanical properties and the cavitation corrosion resistance of 00Cr22Ni5Mo3N duplex stainless steel was observed. The degradation of the nano-mechanical properties of the corroded surface layer was accelerated by the synergistic effect between cavitation erosion and corrosion. A key factor was the adsorption of Cl(-), which caused a preferential dissolution of the ferrous oxides in the passive film layer on the corroded surface layer. Cavitation further promoted the preferential dissolution of the ferrous oxides in the passive film layer. Simultaneously, cavitation accelerated the erosion of the ferrite in the corroded surface layer, resulting in the degradation of the nano-mechanical properties of the corroded surface layer on 00Cr22Ni5Mo3N duplex stainless steel under cavitation.

  1. A Location-Based Duplex Scheme for Cost Effective Rural Broadband Connectivity Using IEEE 802.22 Cognitive Radio Based Wireless Regional Area Networks

    Science.gov (United States)

    Kalidoss, R.; Bhagyaveni, M. A.; Vishvaksenan, K. S.

    2014-08-01

    The search for a method of utilizing the scarce spectrum in an efficient manner is an active area of research in both academic and industrial communities. IEEE 802.22 is a standard for wireless regional area network (WRAN) based on cognitive radio (CR) that operates over underutilized portions of TV bands (54-862 MHz). Time division duplex (TDD)-based WRAN cells have such advantages as dynamic traffic allocation, traffic asymmetry to users and ease of spectrum allocation. However, these cells suffer from severe cross time slot (CTS) interference when the frames of the cells are not synchronized with adjacent WRAN cells. In this paper, we evaluate the location-based duplex (LBD) scheme for eliminating the CTS interference. The proposed LBD system is much more flexible and efficient in providing asymmetric data service and eliminating CTS interference by exploiting the advantages of both TDD and frequency division duplex (FDD) schemes. We also compare the performance of LBD systems with virtual cell concepts. Furthermore, our simulation results reveal that LBD-based systems outperform the virtual cell approach in terms of the low signal-to-interference (SIR) ratio requirement by mitigating the effects of CTS.

  2. Structural Properties of G,T-Parallel Duplexes

    Directory of Open Access Journals (Sweden)

    Anna Aviñó

    2010-01-01

    Full Text Available The structure of G,T-parallel-stranded duplexes of DNA carrying similar amounts of adenine and guanine residues is studied by means of molecular dynamics (MD simulations and UV- and CD spectroscopies. In addition the impact of the substitution of adenine by 8-aminoadenine and guanine by 8-aminoguanine is analyzed. The presence of 8-aminoadenine and 8-aminoguanine stabilizes the parallel duplex structure. Binding of these oligonucleotides to their target polypyrimidine sequences to form the corresponding G,T-parallel triplex was not observed. Instead, when unmodified parallel-stranded duplexes were mixed with their polypyrimidine target, an interstrand Watson-Crick duplex was formed. As predicted by theoretical calculations parallel-stranded duplexes carrying 8-aminopurines did not bind to their target. The preference for the parallel-duplex over the Watson-Crick antiparallel duplex is attributed to the strong stabilization of the parallel duplex produced by the 8-aminopurines. Theoretical studies show that the isomorphism of the triads is crucial for the stability of the parallel triplex.

  3. Full Duplex Emulation via Spatial Separation of Half Duplex Nodes in a Planar Cellular Network

    DEFF Research Database (Denmark)

    Thomsen, Henning; Kim, Dong Min; Popovski, Petar;

    2016-01-01

    A Full Duplex Base Station (FD-BS) can be used to serve simultaneously two Half-Duplex (HD) Mobile Stations (MSs), one working in the uplink and one in the downlink, respectively. The same functionality can be realized by having two interconnected and spatially separated Half Duplex Base Stations...... (HD-BSs), which is a scheme termed CoMPflex (CoMP for In-Band Wireless Full Duplex). A FD-BS can be seen as a special case of CoMPflex with separation distance zero. In this paper we study the performance of CoMPflex in a two-dimensional cellular scenario using stochastic geometry and compare...... of communication reliability. Following the trend of wireless network densification, CoMPflex can be regarded as a method with a great potential to effectively use the dense HD deployments....

  4. Effect of Multipass Friction Stir Processing on Mechanical and Corrosion Behavior of 2507 Super Duplex Stainless Steel

    Science.gov (United States)

    Mishra, M. K.; Gunasekaran, G.; Rao, A. G.; Kashyap, B. P.; Prabhu, N.

    2016-12-01

    The microstructure, mechanical properties, and corrosion behavior of 2507 super duplex stainless steel after multipass friction stir processing (FSP) were examined. A significant refinement in grain size of both ferrite and austenite was observed in stir zone resulting in improved yield and tensile strength. Electrochemical impedance spectroscopy and anodic polarization studies in 3.5 wt.% NaCl solution showed nobler corrosion characteristics with increasing number of FSP passes. This was evident from the decrease in corrosion current density, decrease in passive current density, and increase in polarization resistance. Also, the decrease in density of defects, based on Mott-Schottky analysis, further confirms the improvement in corrosion resistance of 2507 super duplex stainless steel after multipass FSP.

  5. Synthesis, interactions, molecular structure, biological properties and molecular docking studies on Mn, Co, Zn complexes containing acetylacetone and pyridine ligands with DNA duplex.

    Science.gov (United States)

    Thamilarasan, V; Sengottuvelan, N; Stalin, N; Srinivasan, P; Chakkaravarthi, G

    2016-07-01

    Three metal complexes (1-3) of the type [Mn(acac)2(py)·H2O] (1), [Co(acac)2(py)·H2O] (2) and [Zn(acac)2(py)·H2O] (3), [Where acac=acetylacetone, py=pyridine] were synthesized and characterized by spectral (UV-vis, FT-IR, ESI-mass) analysis. The structure of complex 2 has been determined by single crystal X-ray diffraction studies and the configuration of ligand-coordinated to metal(II) ion was well described as distorted octahedral coordination geometry. The interaction of the complexes with CT-DNA has been explored by absorption, fluorescence, circular dichromism spectroscopy, viscosity measurements and molecular docking studies. The intrinsic binding constant Kb of complexes 1-3 with CT-DNA obtained from UV-vis absorption spectral studies were 2.1×10(4), 2.1×10(5) and 1.98×10(4)M(-1), respectively, which revealed that the complexes could interact with CT-DNA through groove binding. The results indicated that the complexes (1-3) were able to bind to DNA with different binding affinity, in the order: 2>1>3. The interaction of the compounds with bovine serum albumins were also investigated using fluorescence methods and the gel electrophoresis assay demonstrates weak cleavage ability of the pBR322 plasmid DNA in the presence of the metal complexes (1-3) with various activators. Further, the in vitro cytotoxic effect of the complexes were examined on cancerous cell line, with human breast cancer cells MCF-7.

  6. Modified naphthalene diimide as a suitable tetraplex DNA ligand: application to cancer diagnosis and anti-cancer drug

    Science.gov (United States)

    Takenaka, Shigeori

    2017-07-01

    It is known that naphthalene diimide carrying two substituents binds to DNA duplex with threading intercalation. Naphthalene diimide carrying ferrocene moieties, ferrocenylnaphthalene diimide (FND), formed a stable complex with DNA duplex and an electrochemical gene detection was achieved with current signal generated from FND bound to the DNA duplex between target DNA and DNA probe immobilized electrode. FND couldn't bind to the mismatched and its surrounding region of DNA duplex and thus FND was applied to the precision detection of single nucleotide polymorphisms (SNPs) using the improved discrimination ability between fully matched and mismatched DNA hybrids and multi-electrode chip. Some of FND derivatives bound to telomere DNA tetraplex stronger than to DNA duplex and was applied to cancer diagnosis as a measure of the elongated telomere DNA with telomerase as a suitable maker of cancer. Furthermore, cyclic naphthalene diimides realized the extremely high preference for DNA tetraplex over DNA duplex. Such molecules will open an effective anti-cancer drug based on telomerase specific inhibitor.

  7. Increased Y-chromosome detection by SRY duplexing

    DEFF Research Database (Denmark)

    Hansen, Morten Høgh; Clausen, Frederik Banch; Dziegiel, Morten Hanefeld

    2012-01-01

    Determining fetal sex noninvasively is dependent of a robust assay. We designed a novel SRY assay and combined it with a SRY assay from literature forming a duplex assay with the same fluorescent dye to increase detection of Y-chromosome at low cell-free fetal DNA or chimeric DNA concentrations....

  8. Microstructural evolution during aging at 800 °C and its effect on the magnetic behavior of UNS S32304 lean duplex stainless steel

    Science.gov (United States)

    Dille, J.; Areiza, M. C. L.; Tavares, S. S. M.; Pereira, G. R.; De Almeida, L. H.; Rebello, J. M. A.

    2017-03-01

    Duplex stainless steels are high strength and corrosion resistant alloys extensively used in chemical and petrochemical industries. However, exposition to temperatures in the range 300-1000 °C leads to precipitation of different phases having a detrimental effect on the mechanical properties and on the corrosion resistance of the alloy. In this work, the microstructural evolution during aging of a UNS S32304 lean duplex stainless steel was investigated by scanning electron microscopy, transmission electron microscopy and magnetic force microscopy. Formation of secondary austenite as well as Cr2N and Cr23C6 precipitation and, consequently, a decrease of ferrite volume fraction were observed. EDX analysis indicated that secondary austenite is depleted in chromium which is detrimental to the corrosion resistance of the alloy. A variation of magnetic properties and Eddy current measurement parameters during aging was simultaneously detected and can be explained by the decrease of ferrite volume content. Therefore, Eddy current non-destructive testing can be successfully applied to detect the formation of deleterious phases during aging.

  9. Submolecular Structure and Orientation of Oligonucleotide Duplexes Tethered to Gold Electrodes Probed by Infrared Reflection Absorption Spectroscopy: Effect of the Electrode Potentials.

    Science.gov (United States)

    Kékedy-Nagy, László; Ferapontova, Elena E; Brand, Izabella

    2017-02-23

    Unique electronic and ligand recognition properties of the DNA double helix provide basis for DNA applications in biomolecular electronic and biosensor devices. However, the relation between the structure of DNA at electrified interfaces and its electronic properties is still not well understood. Here, potential-driven changes in the submolecular structure of DNA double helices composed of either adenine-thymine (dAdT)25 or cytosine-guanine (dGdC)20 base pairs tethered to the gold electrodes are for the first time analyzed by in situ polarization modulation infrared reflection absorption spectroscopy (PM IRRAS) performed under the electrochemical control. It is shown that the conformation of the DNA duplexes tethered to gold electrodes via the C6 alkanethiol linker strongly depends on the nucleic acid sequence composition. The tilt of purine and pyrimidine rings of the complementary base pairs (dAdT and dGdC) depends on the potential applied to the electrode. By contrast, neither the conformation nor orientation of the ionic in character phosphate-sugar backbone is affected by the electrode potentials. At potentials more positive than the potential of zero charge (pzc), a gradual tilting of the double helix is observed. In this tilted orientation, the planes of the complementary purine and pyrimidine rings lie ideally parallel to each other. These potentials do not affect the integral stability of the DNA double helix at the charged interface. At potentials more negative than the pzc, DNA helices adopt a vertical to the gold surface orientation. Tilt of the purine and pyrimidine rings depends on the composition of the double helix. In monolayers composed of (dAdT)25 molecules the rings of the complementary base pairs lie parallel to each other. By contrast, the tilt of purine and pyrimidine rings in (dGdC)20 helices depends on the potential applied to the electrode. Such potential-induced mobility of the complementary base pairs can destabilize the helix structure

  10. Investigation of the Hot Plasticity of Duplex Stainless Steel

    Institute of Scientific and Technical Information of China (English)

    LIN Gang; ZHANG Zhi-xia; SONG Hong-wei; TONG Jun; ZHOU Can-dong

    2008-01-01

    Hot plasticity of a nitrogen alloyed 25Cr-7Ni-4 Mo duplex stainless steel was investigated.The results indicate that thc main factors affecting the hot plasticity of duplex stainless steel are listed as follows:coalescent force of phase interface,microstructure,and the phase ratio and difference between the mechanicsl propertms of ferrite and austenite.The heat treatment and sulphur contents have a notable effect on the hot plasticity.The reasonable heat treatrnents and the irlcreased interfacial coalescent force will effectively enhance the hot plasticity of duplex stainless steel.

  11. Effect of Phosphate-Buffered Solution Corrosion on the Ratcheting Fatigue Behavior of a Duplex Mg-Li-Al Alloy

    Science.gov (United States)

    Yuan, Xin; Yu, Dunji; Gao, Li-Lan; Gao, Hong

    2016-05-01

    This work reports the uniaxial ratcheting and fatigue behavior of a duplex Mg-Li-Al alloy under the influence of phosphate-buffered solution corrosion. Microstructural observations reveal pitting and filament corrosion defects, which impair the load-bearing capacity of the alloy and cause stress concentration, thus leading to an accelerated accumulation of ratcheting strain and shortened fatigue life under the same nominal loading conditions. Comparing Smith model, Smith-Watson-Topper model, and Paul-Sivaprasad-Dhar model, a ratcheting fatigue life prediction model based on the Broberg damage rule and the Paul-Sivaprasad-Dhar model was proposed, and the model yielded a superior prediction for the studied magnesium alloy.

  12. Next-generation bis-locked nucleic acids with stacking linker and 2'-glycylamino-LNA show enhanced DNA invasion into supercoiled duplexes

    DEFF Research Database (Denmark)

    Geny, Sylvain; Moreno, Pedro M D; Krzywkowski, Tomasz

    2016-01-01

    Targeting and invading double-stranded DNA with synthetic oligonucleotides under physiological conditions remain a challenge. Bis-locked nucleic acids (bisLNAs) are clamp-forming oligonucleotides able to invade into supercoiled DNA via combined Hoogsteen and Watson-Crick binding. To improve the b...

  13. Characterization of recombinant malarial RecQ DNA helicase.

    Science.gov (United States)

    Suntornthiticharoen, Pattra; Srila, Witsanu; Chavalitshewinkoon-Petmitr, Porntip; Limudomporn, Paviga; Yamabhai, Montarop

    2014-08-01

    RecQ DNA gene of multi-drug resistant Plasmodium falciparum K1 (PfRecQ1) was cloned, and the recombinant C-terminal-decahistidine-tagged PfRecQ1 was expressed in Escherichia coli. The purified enzyme could efficiently unwind partial duplex DNA substrate in a 3' to 5' direction. The malarial RecQ1 could not unwind substrates with both 5' and 3' overhangs, those with a 5' overhang, or blunt-ended DNA duplexes. Unwinding of DNA helicase activity was driven by the hydrolysis of ATP. The drug inhibitory effects of six compounds indicated that only doxorubicin and daunorubicin could inhibit the unwinding activity.

  14. Joint duplex mode selection, channel allocation, and power control for full-duplex cognitive femtocell networks

    Directory of Open Access Journals (Sweden)

    Mingjie Feng

    2015-02-01

    Full Text Available In this paper, we aim to maximize the sum rate of a full-duplex cognitive femtocell network (FDCFN as well as guaranteeing the quality of service (QoS of users in the form of a required signal to interference plus noise ratios (SINR. We first consider the case of a pair of channels, and develop optimum-achieving power control solutions. Then, for the case of multiple channels, we formulate joint duplex model selection, power control, and channel allocation as a mixed integer nonlinear problem (MINLP, and propose an iterative framework to solve it. The proposed iterative framework consists of a duplex mode selection scheme, a near-optimal distributed power control algorithm, and a greedy channel allocation algorithm. We prove the convergence of the proposed iterative framework as well as a lower bound for the greedy channel allocation algorithm. Numerical results show that the proposed schemes effectively improve the sum rate of FDCFNs.

  15. Effects of Nitrogen and Tensile Direction on Stress Corrosion Cracking Susceptibility of Ni-Free FeCrMnC-Based Duplex Stainless Steels

    Science.gov (United States)

    Ha, Heon-Young; Lee, Chang-Hoon; Lee, Tae-Ho; Kim, Sangshik

    2017-01-01

    Stress corrosion cracking (SCC) behavior of Ni-free duplex stainless steels containing N and C (Febalance-19Cr-8Mn-0.25C-(0.03, 0.21)N, in wt %) was investigated by using a slow strain rate test (SSRT) in air and aqueous NaCl solution with different tensile directions, including parallel (longitudinal) and perpendicular (transverse) to the rolling direction. It was found that alloying N was effective in increasing the resistance to SCC, while it was higher along the longitudinal direction than the transverse direction. The SCC susceptibility of the two alloys was assessed based on the electrochemical resistance to pitting corrosion, the corrosion morphology, and the fractographic analysis. PMID:28772651

  16. Enantiospecific kinking of DNA by a partially intercalating metal complex

    KAUST Repository

    Reymer, Anna

    2012-01-01

    Opposite enantiomers of [Ru(phenanthroline) 3] 2+ affect the persistence length of DNA differently, a long speculated effect of helix kinking. Our molecular dynamics simulations confirm a substantial change of duplex secondary structure produced by wedge-intercalation of one but not the other enantiomer. This effect is exploited by several classes of DNA operative proteins. © The Royal Society of Chemistry 2012.

  17. Combined rpoB duplex PCR and hsp65 PCR restriction fragment length polymorphism with capillary electrophoresis as an effective algorithm for identification of Mycobacterial species from clinical isolates

    Directory of Open Access Journals (Sweden)

    Huang Chen-Cheng

    2012-07-01

    Full Text Available Abstract Background Mycobacteria can be quickly and simply identified by PCR restriction-enzyme analysis (PRA, but misidentification can occur because of similarities in band sizes that are critical for discriminating among species. Capillary electrophoresis can provide computer-aided band discrimination. The aim of this research was to develop an algorithm for identifying mycobacteria by combined rpoB duplex PRA (DPRA and hsp65 PRA with capillary electrophoresis. Results Three hundred and seventy-six acid-fast bacillus smear-positive BACTEC cultures, including 200 Mycobacterium tuberculosis complexes (MTC and 176 non-tuberculous mycobacteria (NTM were analyzed. With combined hsp65 and rpoB DPRA, the accuracy rate was 100% (200 isolates for the MTC and 91.4% (161 isolates for the NTM. Among the discordant results (8.6% for the NTM, one isolate of Mycobacterial species and an isolate of M. flavescens were found as new sub-types in hsp65 PRA. Conclusions This effective and novel identification algorithm using combined rpoB DPRA and hsp65 PRA with capillary electrophoresis can rapidly identify mycobacteria and find new sub-types in hsp65 PRA. In addition, it is complementary to 16 S rDNA sequencing.

  18. Effects of Annealing Treatment Prior to Cold Rolling on Delayed Fracture Properties in Ferrite-Austenite Duplex Lightweight Steels

    Science.gov (United States)

    Sohn, Seok Su; Song, Hyejin; Kim, Jung Gi; Kwak, Jai-Hyun; Kim, Hyoung Seop; Lee, Sunghak

    2016-02-01

    Tensile properties of recently developed automotive high-strength steels containing about 10 wt pct of Mn and Al are superior to other conventional steels, but the active commercialization has been postponed because they are often subjected to cracking during formation or to the delayed fracture after formation. Here, the delayed fracture behavior of a ferrite-austenite duplex lightweight steel whose microstructure was modified by a batch annealing treatment at 1023 K (750 °C) prior to cold rolling was examined by HCl immersion tests of cup specimens, and was compared with that of an unmodified steel. After the batch annealing, band structures were almost decomposed as strong textures of {100} α-fibers and {111} γ-fibers were considerably dissolved, while ferrite grains were refined. The steel cup specimen having this modified microstructure was not cracked when immersed in an HCl solution for 18 days, whereas the specimen having unmodified microstructure underwent the delayed fracture within 1 day. This time delayed fracture was more critically affected by difference in deformation characteristics such as martensitic transformation and deformation inhomogeneity induced from concentration of residual stress or plastic strain, rather than the difference in initial microstructures. The present work gives a promise for automotive applications requiring excellent mechanical and delayed fracture properties as well as reduced specific weight.

  19. [Carotid duplex ultrasonography for neurosurgeons].

    Science.gov (United States)

    Sadahiro, Hirokazu; Ishihara, Hideyuki; Oka, Fumiaki; Suzuki, Michiyasu

    2011-12-01

    Carotid duplex ultrasonography (CDU) is one of the most well-known imaging methods for arteriosclerosis and ischemic stroke. For neurosurgeons, it is very important for the details of carotid plaque to be thoroughly investigated by CDU. Symptomatic carotid plaque is very fragile and easily changes morphologically, and so requires frequent CDU examination. Furthermore, after carotid endarterectomy (CEA) and carotid artery stenting (CAS), restenosis is evaluated with CDU. CDU facilitates not only morphological imaging in the B mode, but also allows a flow study with color Doppler and duplex imaging. So, CDU can help assess the presence of proximal and intracranial artery lesions in spite of only having a cervical view, and the patency of the extracranial artery to intracranial artery bypass is revealed with CDU, which shows a rich velocity and low pulsatility index (PI) in duplex imaging. For the examiner, it is necessary to ponder on what duplex imaging means in examinations, and to summarize all imaging finding.

  20. Effect of shot peening treatment in the behavior of residual stress in duplex stainless steel during medium cycle fatigue; Efeito do tratamento de shot peening no comportamento das tensoes residuais em aco inoxidavel duplex durante fadiga de medio ciclo

    Energy Technology Data Exchange (ETDEWEB)

    Pedrosa, Peter D.S.; Rebello, Joao Marcos A. [Coordenacao dos Programas de Pos-Graduacao de Engenharia (PEMM/COPPE/UFRJ), Rio de Janeiro, RJ (Brazil). Programa de Engenharia Metalurgica e de Materiais; Fonseca, Maria P. Cindra, E-mail: mcindra@vm.uff.b [Universidade Federal Fluminense (PGMEC/UFF), Niteroi, RJ (Brazil). Escola de Engenharia. Programa de Pos-Graduacao em Engenharia Mecanica

    2010-07-01

    The lifetime of duplex stainless steel parts experiencing cyclic fatigue is directly influenced by the residual stresses present in the ferrite and austenite phases. The motivation for this work was to analyze the behaviour of the residual stresses fields introduced by shot peening treatment in both phases, in the sample surface as in the subsurface layers, in low fatigue cycles, using the X-rays diffraction technique. The results shows that the compressive residual stresses introduced by the shot peening treatment in both phases improved fatigue life of the material. However, the cyclical loads produce partial or total relief in these residual stresses fields. It was verified that the shot peening process induced the formation of microcracks only in the ferrite phase. The largest variations in the total compressive residual stresses fields also occurred in this phase. The samples surfaces were analyzed by scanning electron microscopy. (author)

  1. Time-resolved fluorescence of 2-aminopurine in DNA duplexes in the presence of the EcoP15I Type III restriction-modification enzyme.

    Science.gov (United States)

    Ma, Long; Wu, Xiaohua; Wilson, Geoffrey G; Jones, Anita C; Dryden, David T F

    2014-06-20

    EcoP15I is a Type III DNA restriction and modification enzyme of Escherichia coli. We show that it contains two modification (Mod) subunits for sequence-specific methylation of DNA and one copy of a restriction endonuclease (Res) subunit for cleavage of DNA containing unmethylated target sequences. Previously the Mod2 dimer in the presence of cofactors was shown to use nucleotide flipping to gain access to the adenine base targeted for methylation (Reddy and Rao, J. Mol. Biol. 298 (2000) 597-610.). Surprisingly the Mod2 enzyme also appeared to flip a second adenine in the target sequence, one which was not subject to methylation. We show using fluorescence lifetime measurements of the adenine analogue, 2-aminopurine, that only the methylatable adenine undergoes flipping by the complete Res1Mod2 enzyme and that this occurs even in the absence of cofactors. We suggest that this is due to activation of the Mod2 core by the Res subunit.

  2. Destabilizing DNA during Rejoining Enhances Fidelity of Repair.

    Directory of Open Access Journals (Sweden)

    Richard Robinson

    2015-08-01

    Full Text Available A new study shows that during repair of DNA, the effect of a single-strand annealing protein is to destabilize DNA duplex formation so that annealing only occurs between perfectly matched strands; the protein then clamps the strands together for repair. Read the Research Article.

  3. Detection of chicken contamination in beef meatball using duplex-PCR Cyt b gene

    Science.gov (United States)

    Sari, E. P.; Kartikasari, L. R.; Cahyadi, M.

    2017-04-01

    Beef is one of expensive animal protein sources compared to other meats, on the other hand, chicken is cheap animal protein source. Mixing of chicken into beef meatball is possibly performed to decrease production cost. The aim of this study was to detect chicken contamination in beef meatball using Cytochrome b (Cyt b) gene by duplex-PCR. Sample was designed and prepared as follows, 100% of chicken meatball, 100% of beef meatball and serial level of chicken contaminations in beef meatball (1, 5, 10 and 25%, respectively). Isolation of DNA genome from meatball was according to the guideline of gSYNCTM DNA Extraction Kit for animal tissue. The PCR reaction was carried out using KAPA2G Fast Multiplex Mix. This study found that the DNA genome was succesfully extracted. Moreover, chicken contamination in beef meatball was indicated by the presence of 227 bp DNA band on 2% of agarose gels. Current study revealed that duplex-PCR using Cyt b gene as a genetic marker was able to detect chicken contamination in beef meatball until 1% of chicken meat in the sample. It can be effectively used to identify contamination and also authenticate species origin in animal products to protect consumer from undesirable contents in the food.

  4. Modeling stopped-flow data for nucleic acid duplex formation reactions: the importance of off-path intermediates.

    Science.gov (United States)

    Sikora, Jacqueline R; Rauzan, Brittany; Stegemann, Rachel; Deckert, Alice

    2013-08-01

    Evidence for unexpected off-path intermediates to DNA duplex formation is presented. These off-path intermediates are shown to involve unimolecular and, in one case, bimolecular structure in one of the single strands of complementary DNA. Three models are developed to account for the observed single-stranded structures that are formed in parallel with duplex formation. These models are applied to the analysis of stopped-flow data for eight different nonself-complementary duplex formation reactions in order to extract the elementary rate constant for formation of the duplex from the complementary random coil single-stranded DNA. The free energy of activation (at 25 °C) for the denaturation of each duplex is calculated from these data and is shown to have a linear correlation to the overall standard free energy for duplex formation (also at 25 °C). Duplexes that contain mismatches obey a parallel linear free-energy (LFE) relationship with a y-intercept that is greater than that of duplexes without mismatches. Slopes near unity for the LFE relationships indicate that all duplexes go through an early, unstructured transition state.

  5. Effects of cooling time and alloying elements on the microstructure of the gleeble-simulated heat-affected zone of 22% Cr duplex stainless steels

    Science.gov (United States)

    Hsieh, Rong-Iuan; Liou, Horng-Yih; Pan, Yeong-Tsuen

    2001-10-01

    The effects of austenite stabilizers, such as nitrogen, nickel, and manganese, and cooling time on the microstructure of the Gleeble simulated heat-affected zone (HAZ) of 22% Cr duplex stainless steels were investigated. The submerged are welding was performed for comparison purposes. Optical microscopy (OM) and transmission electron microscopy (TEM) were used for microscopic studies. The amount of Cr2N precipitates in the simulated HAZ was determined using the potentiostatic electrolysis method. The experimental results indicate that an increase in the nitrogen and nickel contents raised the δ to transformation temperature and also markedly increased the amount of austenite in the HAZ. The lengthened cooling time promotes the reformation of austenite. An increase in the austenite content reduces the supersaturation of nitrogen in ferrite matrix as well as the precipitation tendency of Cr2N. The optimum cooling time from 800 to 500 °C (Δ t 8/5) obtained from the Gleeble simulation is between 30 and 60 s, which ensures the austenite content in HAZ not falling below 25% and superior pitting and stress corrosion cracking resistance for the steels. The effect of manganese on the formation of austenite can be negligible.

  6. Atmospheric-Induced Stress Corrosion Cracking of Grade 2205 Duplex Stainless Steel—Effects of 475 °C Embrittlement and Process Orientation

    Directory of Open Access Journals (Sweden)

    Cem Örnek

    2016-07-01

    Full Text Available The effect of 475 °C embrittlement and microstructure process orientation on atmospheric-induced stress corrosion cracking (AISCC of grade 2205 duplex stainless steel has been investigated. AISCC tests were carried out under salt-laden, chloride-containing deposits, on U-bend samples manufactured in rolling (RD and transverse directions (TD. The occurrence of selective corrosion and stress corrosion cracking was observed, with samples in TD displaying higher propensity towards AISCC. Strains and tensile stresses were observed in both ferrite and austenite, with similar magnitudes in TD, whereas, larger strains and stresses in austenite in RD. The occurrence of 475 °C embrittlement was related to microstructural changes in the ferrite. Exposure to 475 °C heat treatment for 5 to 10 h resulted in better AISCC resistance, with spinodal decomposition believed to enhance the corrosion properties of the ferrite. The austenite was more susceptible to ageing treatments up to 50 h, with the ferrite becoming more susceptible with ageing in excess of 50 h. Increased susceptibility of the ferrite may be related to the formation of additional precipitates, such as R-phase. The implications of heat treatment at 475 °C and the effect of process orientation are discussed in light of microstructure development and propensity to AISCC.

  7. Nonlinear effects in the torsional adjustment of interacting DNA.

    Science.gov (United States)

    Kornyshev, A A; Wynveen, A

    2004-04-01

    DNA molecules in solution, having negatively charged phosphates and countercations readsorbed on its surface, possess a distinct charge separation motif to interact electrostatically. If their double-helical structure were ideal, duplexes in parallel juxtaposition could choose azimuthal alignment providing attraction, or at least a reduction of repulsion, between them. But duplexes are not perfect staircases and the distortions of their helical structure correlate with their base pair texts. If the patterns of distortions on the opposing molecules are uncorrelated, the mismatch will accumulate as a random walk and attraction vanishes. Based on this idea, a model of recognition of homologous sequences has been proposed [A. A. Kornyshev and S. Leikin, Phys. Rev. Lett. 86, 3666 (2001)]. But DNA has torsional elasticity. How will this help to relax a mismatch between the charge distributions on two nonhomologous DNA's? In the same work, the solution of this problem has been mapped onto a frustrated sine Gordon equation in a nonlocal random field (where the latter represents a pattern of twist angle distortions on the opposing molecules), but the results had been obtained in the limit of torsionally rigid molecules. In the present paper, by solving this equation numerically, we find a strongly nonlinear relaxation mechanism which utilizes static kink-soliton modes triggered by the "random field." In the range of parameters where the solitons do not emerge, we find good agreement with the results of a variational study [A. G. Cherstvy, A. A. Kornyshev, and S. Leikin, J. Phys. Chem. B (to be published)]. We reproduce the first-order transitions in the interaxial separation dependence, but detect also second-order or weak first-order transitions for shorter duplexes. The recognition energy between two nonhomologous DNA sequences is calculated as a function of interaxial separation and the length of juxtaposition. The soliton-caused kinky length dependence is discussed in

  8. 线粒体基因种属鉴定复合扩增体系%Duplex Amplification of Mitochondrial DNA for Species Identification

    Institute of Scientific and Technical Information of China (English)

    方月琴; 顾准; 侯一平

    2012-01-01

    针对cyt b基因和ND6基因序列,设计两对引物,以14种常见动物(包括人)生物性检材为对象进行PCR扩增,产物经聚丙烯酰胺凝胶电泳(PAGE)、硝酸银染色进行分析。结果证明,两条电泳条带者为人样本,分别是cyt b基因片段(358 bp)和ND6基因片段(181 bp);一条电泳条带者为动物来源,是358 bp cyt b基因片段。在37.5μL PCR反应体系中最小检出模板量为0.25 pg基因组DNA。检材在4℃、室温和37℃温度下,经过4个月后均可获得正确的种属区分。高度潮湿环境中放置50 d的检材、形成于水泥地面、墙面、泥土等基质上的血痕,本方法可得到正确区分。由此建立的种属鉴定的线粒体基因复合扩增体系,其检测片段短、灵敏度高、操作简易,适合法医学上微量、降解、陈旧检材的种属判定。%Two pairs of primers for the cyt b gene and ND6 gene sequences were designed and biological sample of 14 species of animals(including humans) PCR were amplified,and the products were analyzed by poly-acrylamide gel electrophoresis(PAGE) and stained with silver nitrate.The results demonstrate that the two electrophoretic bands of the cyt b gene fragment(358 bp) and ND6 gene fragment(181 bp) are human samples;and the one electrophoresis strip of 358 bp Cytb gene fragments is animal sources.In 37.5 μl PCR reaction system,the minimum detectable template concentration is 0.25 pg genomic DNA.Samples can be distinguished correctly at 4°C,room temperature and 37 °C,after four months later.Samples placed in a high humidity environment for 50 days and bloodstains formed in the concrete floor or on the walls,and soil matrix,can be distinguished correctly by the methods.The species identification method of the mitochondrial genome multiplex PCR system with short detected fragments are high sensitivity and easy to operate and suitable to determine the trace or degradation or obsolete samples of

  9. Effect of intercalator substituent and nucleotide sequence on the stability of DNA- and RNA-naphthalimide complexes.

    Science.gov (United States)

    Johnson, Charles A; Hudson, Graham A; Hardebeck, Laura K E; Jolley, Elizabeth A; Ren, Yi; Lewis, Michael; Znosko, Brent M

    2015-07-01

    DNA intercalators are commonly used as anti-cancer and anti-tumor agents. As a result, it is imperative to understand how changes in intercalator structure affect binding affinity to DNA. Amonafide and mitonafide, two naphthalimide derivatives that are active against HeLa and KB cells in vitro, were previously shown to intercalate into DNA. Here, a systematic study was undertaken to change the 3-substituent on the aromatic intercalator 1,8-naphthalimide to determine how 11 different functional groups with a variety of physical and electronic properties affect binding of the naphthalimide to DNA and RNA duplexes of different sequence compositions and lengths. Wavelength scans, NMR titrations, and circular dichroism were used to investigate the binding mode of 1,8-naphthalimide derivatives to short synthetic DNA. Optical melting experiments were used to measure the change in melting temperature of the DNA and RNA duplexes due to intercalation, which ranged from 0 to 19.4°C. Thermal stabilities were affected by changing the substituent, and several patterns and idiosyncrasies were identified. By systematically varying the 3-substituent, the binding strength of the same derivative to various DNA and RNA duplexes was compared. The binding strength of different derivatives to the same DNA and RNA sequences was also compared. The results of these comparisons shed light on the complexities of site specificity and binding strength in DNA-intercalator complexes. For example, the consequences of adding a 5'-TpG-3' or 5'-GpT-3' step to a duplex is dependent on the sequence composition of the duplex. When added to a poly-AT duplex, naphthalimide binding was enhanced by 5.6-11.5°C, but when added to a poly-GC duplex, naphthalimide binding was diminished by 3.2-6.9°C. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Coiled-coil conformation of a pentamidine-DNA complex.

    Science.gov (United States)

    Moreno, Tadeo; Pous, Joan; Subirana, Juan A; Campos, J Lourdes

    2010-03-01

    The coiled-coil structure formed by the complex of the DNA duplex d(ATATATATAT)(2) with pentamidine is presented. The duplex was found to have a mixed structure containing Watson-Crick and Hoogsteen base pairs. The drug stabilizes the coiled coil through the formation of cross-links between neighbouring duplexes. The central part of the drug is found in the minor groove as expected, whereas the charged terminal amidine groups protrude and interact with phosphates from neighbouring molecules. The formation of cross-links may be related to the biological effects of pentamidine, which is used as an antiprotozoal agent in trypanosomiasis, leishmaniasis and pneumonias associated with AIDS. The DNA sequence that was used is highly abundant in most eukaryotic genomes. However, very few data are available on DNA sequences which only contain A.T base pairs.

  11. Nonlinear effects in the torsional adjustment of interacting DNA

    Science.gov (United States)

    Kornyshev, A. A.; Wynveen, A.

    2004-04-01

    DNA molecules in solution, having negatively charged phosphates and countercations readsorbed on its surface, possess a distinct charge separation motif to interact electrostatically. If their double-helical structure were ideal, duplexes in parallel juxtaposition could choose azimuthal alignment providing attraction, or at least a reduction of repulsion, between them. But duplexes are not perfect staircases and the distortions of their helical structure correlate with their base pair texts. If the patterns of distortions on the opposing molecules are uncorrelated, the mismatch will accumulate as a random walk and attraction vanishes. Based on this idea, a model of recognition of homologous sequences has been proposed [A. A. Kornyshev and S. Leikin, Phys. Rev. Lett. 86, 3666 (2001)]. But DNA has torsional elasticity. How will this help to relax a mismatch between the charge distributions on two nonhomologous DNA’s? In the same work, the solution of this problem has been mapped onto a frustrated sine Gordon equation in a nonlocal random field (where the latter represents a pattern of twist angle distortions on the opposing molecules), but the results had been obtained in the limit of torsionally rigid molecules. In the present paper, by solving this equation numerically, we find a strongly nonlinear relaxation mechanism which utilizes static kink-soliton modes triggered by the “random field.” In the range of parameters where the solitons do not emerge, we find good agreement with the results of a variational study [A. G. Cherstvy, A. A. Kornyshev, and S. Leikin, J. Phys. Chem. B (to be published)]. We reproduce the first-order transitions in the interaxial separation dependence, but detect also second-order or weak first-order transitions for shorter duplexes. The recognition energy between two nonhomologous DNA sequences is calculated as a function of interaxial separation and the length of juxtaposition. The soliton-caused kinky length dependence is

  12. Biophysical properties of DNA in hydrated ionic liquids

    Science.gov (United States)

    Jumbri, Khairulazhar; Ahmad, Haslina; Abdulmalek, Emilia; Rahman, Mohd Basyaruddin Abdul

    2016-11-01

    The biophysical properties and behavior of natural calf thymus DNA in hydrated 1-ethyl-3-butylimidazolium bromide ionic liquid ([C2bim]Br) have been studied using spectroscopy technique. The effect of ionic liquid concentration and temperature towards the duplex B-DNA conformation were determined. The presence of ionic liquid causes higher duplex DNA stability with the DNA melting temperature of ˜56°C without any addition of buffer solutions. The electrostatic attraction between ionic liquid's cation and DNA phosphates groups was found play a main role in stabilizing native DNA structure. Understanding of the biophysical properties of DNA in this ionic media could be used as a platform for future development of specific solvent for nucleic acid nanotechnology.

  13. The effect of σ-phase precipitation at 800°C on the corrosion resistance in sea-water of a high alloyed duplex stainless steel

    NARCIS (Netherlands)

    Wilms, M.E.; Gadgil, V.J.; Krougman, J.M.; Ijsseling, F.P.

    1994-01-01

    Super-duplex stainless steels are recently developed high alloyed stainless steels that combine good mechanical properties with excellent corrosion resistance. Because of a high content of chromium and molybdenum, these alloys are susceptible to σ-phase precipitation during short exposure to

  14. Incorporation of 3-Aminobenzanthrone into 2′-Deoxyoligonucleotides and Its Impact on Duplex Stability

    Directory of Open Access Journals (Sweden)

    Mark Lukin

    2011-01-01

    Full Text Available 3-Nitrobenzanthrone (3NBA, an environmental pollutant and potent mutagen, causes DNA damage via the reaction of its metabolically activated form with the exocyclic amino groups of purines and the C-8 position of guanine. The present work describes a synthetic approach to the preparation of oligomeric 2′-deoxyribonucleotides containing a 2-(2′-deoxyguanosin-N2-yl-3-aminobenzanthrone moiety, one of the major DNA adducts found in tissues of living organisms exposed to 3NBA. The NMR spectra indicate that the damaged oligodeoxyribonucleotide is capable of forming a regular double helical structure with the polyaromatic moiety assuming a single conformation at room temperature; the spectra suggest that the 3ABA moiety resides in the duplex minor groove pointing toward the 5′-end of the modified strand. Thermodynamic studies show that the dG(N2-3ABA lesion has a stabilizing effect on the damaged duplex, a fact that correlates well with the long persistence of this damage in living organisms.

  15. Temperature effect on DNA molecular wires

    Science.gov (United States)

    Bui, Christopher Minh

    The demand of technology and information today has further pushed the fabrication process of nanotechnology, yet there are limits and obstacles set by the primary laws of physics. Therefore, researchers are pursuing alternative technologies. Deoxyribonucleic acids (DNA) molecular wire is one advantageous option due to its unique characteristics including self-assembly and naturally small; size. This thesis reports the temperature effect on the electrical properties of a double-stranded ?-DNA molecular wire. The data will help expand the DNA wire application and functionality. Thus, the data supports the charge hopping theory on DNA electrical conductivity. Diverse amount of literatures has demonstrated that DNA experiences a biochemical alteration when exposed under different temperature conditions. This change will also cause a change in the electrical properties. In this research, DNA will hang between two gold covered microelectrodes with a distance of 10 to 12 microns. The microelectrodes are fabricated through negative lithography techniques. Then, the samples were exposed to a numerous range of temperature from 25°C to 180°C and went through varying cycles of heating and cooling. The experimental results revealed that the DNA experienced a hysteresis like behavior where the impedance differed between the heating and cooling phase. The impedance of the DNA molecular wire increased when exposed to higher temperature. Furthermore, the impedance stops increasing after a certain amount of heat cycles before the DNA structure failed. The biology and thermodynamics of the DNA wire was analyzed due to the temperature hysteresis effect. The melting temperature and the bond dissociation temperature were evaluated to determine the cause of the impedance trends. The studies and analysis of the temperature effect provided certain insights towards the charge hopping transport mechanism. The thesis concludes with possible applications relating to the temperature effect of

  16. Corrosion behavior of duplex coatings

    Directory of Open Access Journals (Sweden)

    K. Raghu Ram Mohan Reddy

    2016-07-01

    Full Text Available The titanium alloys are used in defense, aerospace, automobile, chemical plants and biomedical applications due to their very high strength and lightweight properties. However, corrosion is a life-limiting factor when Ti alloys are exposed to different chemical environments at high temperatures. In the present paper, duplex NiCrAlY/WC–Co coating is coated onto Ti6Al4V substrate to investigate the corrosion behavior of both coated samples and the substrate. The duplex coating was performed with NiCrAlY as the intermediate coat of 200 μm thickness deposited by HVOF process and WC–Co ceramic top coat with varying thicknesses of 250 μm, 350 μm and 450 μm deposited by DS process. Potentiodynamic polarization tests were employed to investigate the corrosion performance of duplex coated samples and substrate in Ringer’s solution at 37 °C and pH value was set to 5.7. Finally the results reveal that 350 μm thick coated samples showed highest corrosion resistance compared to 250 μm thick samples as well as bare substrate. However, the 450 μm thick coated sample showed poor corrosion resistance compared to the substrate. The scale formed on the samples upon corrosion was characterized by using SEM analysis to understand the degree of corrosion behavior.

  17. The solution structure of double helical arabino nucleic acids (ANA and 2'F-ANA): effect of arabinoses in duplex-hairpin interconversion.

    Science.gov (United States)

    Martín-Pintado, Nerea; Yahyaee-Anzahaee, Maryam; Campos-Olivas, Ramón; Noronha, Anne M; Wilds, Christopher J; Damha, Masad J; González, Carlos

    2012-10-01

    We report here the first structure of double helical arabino nucleic acid (ANA), the C2'-stereoisomer of RNA, and the 2'-fluoro-ANA analogue (2'F-ANA). A chimeric dodecamer based on the Dickerson sequence, containing a contiguous central segment of arabino nucleotides, flanked by two 2'-deoxy-2'F-ANA wings was studied. Our data show that this chimeric oligonucleotide can adopt two different structures of comparable thermal stabilities. One structure is a monomeric hairpin in which the stem is formed by base paired 2'F-ANA nucleotides and the loop by unpaired ANA nucleotides. The second structure is a bimolecular duplex, with all the nucleotides (2'F-ANA and ANA) forming Watson-Crick base pairs. The duplex structure is canonical B-form, with all arabinoses adopting a pure C2'-endo conformation. In the ANA:ANA segment, steric interactions involving the 2'-OH substituent provoke slight changes in the glycosidic angles and, therefore, in the ANA:ANA base pair geometry. These distortions are not present in the 2'F-ANA:2'F-ANA regions of the duplex, where the -OH substituent is replaced by a smaller fluorine atom. 2'F-ANA nucleotides adopt the C2'-endo sugar pucker and fit very well into the geometry of B-form duplex, allowing for favourable 2'F···H8 interactions. This interaction shares many features of pseudo-hydrogen bonds previously observed in 2'F-ANA:RNA hybrids and in single 2'F-ANA nucleotides.

  18. DNA glue

    DEFF Research Database (Denmark)

    Filichev, Vyacheslav V; Astakhova, Irina V.; Malakhov, Andrei D.

    2008-01-01

    Significant alterations in thermal stability of parallel DNA triplexes and antiparallel duplexes were observed upon changing the attachment of ethynylpyrenes from para to ortho in the structure of phenylmethylglycerol inserted as a bulge into DNA (TINA). Insertions of two ortho-TINAs as a pseudo...

  19. Homologous recombination in the archaeon Sulfolobus acidocaldarius: effects of DNA substrates and mechanistic implications.

    Science.gov (United States)

    Rockwood, Jananie; Mao, Dominic; Grogan, Dennis W

    2013-09-01

    Although homologous recombination (HR) is known to influence the structure, stability, and evolution of microbial genomes, few of its functional properties have been measured in cells of hyperthermophilic archaea. The present study manipulated various properties of the parental DNAs in high-resolution assays of Sulfolobus acidocaldarius transformation, and measured the impact on the efficiency and pattern of marker transfer to the recipient chromosome. The relative orientation of homologous sequences, the type and position of chromosomal mutation being replaced, and the length of DNA flanking the marked region all affected the efficiency, linkage, tract continuity, and other parameters of marker transfer. Effects predicted specifically by the classical reciprocal-exchange model of HR were not observed. One analysis observed only 90 % linkage between markers defined by adjacent bases; in another series of experiments, sequence divergence up to 4 % had no detectable impact on overall efficiency of HR or on the co-transfer of a distal non-selected marker. The effects of introducing DNA via conjugation, rather than transformation, were more difficult to assess, but appeared to increase co-transfer (i.e. linkage) of relatively distant non-selected markers. The results indicate that HR events between gene-sized duplex DNAs and the S. acidocaldarius chromosome typically involve neither crossing over nor interference from a mismatch-activated anti-recombination system. Instead, the donor DNA may anneal to a transient chromosomal gap, as in the mechanism proposed for oligonucleotide-mediated transformation of Sulfolobus and other micro-organisms.

  20. Duplex real-time PCR assay for rapid identification of Staphylococcus aureus isolates from dairy cow milk.

    Science.gov (United States)

    Pilla, Rachel; Snel, Gustavo G M; Malvisi, Michela; Piccinini, Renata

    2013-05-01

    Staphylococcus aureus isolates from dairy cow mastitis are not always consistent with the characteristic morphology described, and molecular investigation is often needed. The aim of the study was to develop a duplex real-time PCR assay for rapid identification of Staph. aureus isolates, targeting both nuc and Sa442. Overall, 140 isolates collected from dairy cow mastitis in 90 different herds, were tested. All strains had been identified using morphological and biochemical characteristics. DNA from each strain was amplified in real-time PCR assay, to detect nuc or Sa442. Thereafter, a duplex real-time PCR assay was performed, and specificity of the amplified products was assessed by high resolution melting curve analysis. Out of 124 Staph. aureus isolates, 33 did not show the typical morphology or enzymic activity; in 118 strains, the two melt-curve peaks consistent with nuc and Sa442 were revealed, while 2 isolates showed only the peak consistent with Sa442. Four isolates bacteriologically identified as Staph. aureus, were PCR-negative and were further identified as Staph. pseudintermedius by sequencing. Staph. pseudintermedius and coagulase-negative staphylococci did not carry nuc or Sa442. The results showed the correct identification of all isolates, comprehending also coagulase-or nuc-negative Staph. aureus, while other coagulase-positive Staphylococci were correctly identified as non-Staph. aureus. Both sensitivity and specificity were 100%. High resolution melting analysis allowed easy detection of unspecific products. Finally, the duplex real-time PCR was applied directly to 40 milk samples, to detect infected mammary quarters. The assay confirmed the results of bacteriological analysis, on Staph. aureus-positive or-negative samples. Therefore, the proposed duplex real-time PCR could be used in laboratory routine as a cost-effective and powerful tool for high-throughput identification of atypical Staph. aureus isolates causing dairy cow mastitis. Also, it

  1. The Contribution of the Activation Entropy to the Gas-Phase Stability of Modified Nucleic Acid Duplexes

    Science.gov (United States)

    Hari, Yvonne; Dugovič, Branislav; Istrate, Alena; Fignolé, Annabel; Leumann, Christian J.; Schürch, Stefan

    2016-07-01

    Tricyclo-DNA (tcDNA) is a sugar-modified analogue of DNA currently tested for the treatment of Duchenne muscular dystrophy in an antisense approach. Tandem mass spectrometry plays a key role in modern medical diagnostics and has become a widespread technique for the structure elucidation and quantification of antisense oligonucleotides. Herein, mechanistic aspects of the fragmentation of tcDNA are discussed, which lay the basis for reliable sequencing and quantification of the antisense oligonucleotide. Excellent selectivity of tcDNA for complementary RNA is demonstrated in direct competition experiments. Moreover, the kinetic stability and fragmentation pattern of matched and mismatched tcDNA heteroduplexes were investigated and compared with non-modified DNA and RNA duplexes. Although the separation of the constituting strands is the entropy-favored fragmentation pathway of all nucleic acid duplexes, it was found to be only a minor pathway of tcDNA duplexes. The modified hybrid duplexes preferentially undergo neutral base loss and backbone cleavage. This difference is due to the low activation entropy for the strand dissociation of modified duplexes that arises from the conformational constraint of the tc-sugar-moiety. The low activation entropy results in a relatively high free activation enthalpy for the dissociation comparable to the free activation enthalpy of the alternative reaction pathway, the release of a nucleobase. The gas-phase behavior of tcDNA duplexes illustrates the impact of the activation entropy on the fragmentation kinetics and suggests that tandem mass spectrometric experiments are not suited to determine the relative stability of different types of nucleic acid duplexes.

  2. Azobenzene C-Nucleosides for Photocontrolled Hybridization of DNA at Room Temperature.

    Science.gov (United States)

    Goldau, Thomas; Murayama, Keiji; Brieke, Clara; Asanuma, Hiroyuki; Heckel, Alexander

    2015-12-01

    Herein, we report the reversible light-regulated destabilization of DNA duplexes by using azobenzene C-nucleoside photoswitches. The incorporation of two different azobenzene residues into DNA and their photoswitching properties are described. These new residues demonstrate a photoinduced destabilization effect comparable to the widely applied D-threoninol-linked azobenzene switch, which is currently the benchmark. The photoswitches presented herein show excellent photoswitching efficiencies in DNA duplexes - even at room temperature - which are superior to commonly used azobenzene-based nucleic acid photoswitches. In addition, these photoswitching residues exhibit high thermal stability and excellent fatigue resistance, thus rendering them one of the most efficient candidates for the regulation of duplex stability with light.

  3. Influence of polyethylene glycol on the ligation reaction with calf thymus DNA ligases I and II.

    Science.gov (United States)

    Teraoka, H; Tsukada, K

    1987-01-01

    High concentrations of the nonspecific macromolecule polyethylene glycol 6000 (PEG 6000) enabled DNA ligases I and II from calf thymus to catalyze intermolecular blunt-end ligation of duplex DNA. Intermolecular cohesive-end ligation with these enzymes was markedly stimulated in the presence of 10-16% (w/v) PEG 6000. The effect of PEG 6000 (4-16%) on the sealing of single-stranded breaks in duplex DNA with DNA ligases I and II was not appreciably stimulatory but rather inhibitory. PEG 6000 (15%) enhanced more twofold the rate of DNA ligase II-AMP complex formation, but moderately suppressed the rate of formation of DNA ligase 1-AMP complex. Polyamines and KCl inhibited blunt-end and cohesive-end ligations with DNA ligases I and II in the presence of PEG 6000.

  4. Effect of a Dual Charge on the DNA-Conjugated Redox Probe on DNA Sensing by Short Hairpin Beacons Tethered to Gold Electrodes.

    Science.gov (United States)

    Kékedy-Nagy, László; Shipovskov, Stepan; Ferapontova, Elena E

    2016-08-16

    Charges of redox species can critically affect both the interfacial state of DNA and electrochemistry of DNA-conjugated redox labels and, as a result, the electroanalytical performance of those systems. Here, we show that the kinetics of electron transfer (ET) between the gold electrode and methylene blue (MB) label conjugated to a double-stranded (ds) DNA tethered to gold strongly depend on the charge of the MB molecule, and that affects the performance of genosensors exploiting MB-labeled hairpin DNA beacons. Positively charged MB binds to dsDNA via electrostatic and intercalative/groove binding, and this binding allows the DNA-mediated electrochemistry of MB intercalated into the duplex and, as a result, a complex mode of the electrochemical signal change upon hairpin hybridization to the target DNA, dominated by the "on-off" signal change mode at nanomolar levels of the analyzed DNA. When MB bears an additional carboxylic group, the negative charge provided by this group prevents intimate interactions between MB and DNA, and then the ET in duplexes is limited by the diffusion of the MB-conjugated dsDNA (the phenomenon first shown in Farjami , E. ; Clima , L. ; Gothelf , K. ; Ferapontova , E. E. Anal. Chem. 2011 , 83 , 1594 ) providing the robust "off-on" nanomolar DNA sensing. Those results can be extended to other intercalating redox probes and are of strategic importance for design and development of electrochemical hybridization sensors exploiting DNA nanoswitchable architectures.

  5. Effect of welding process on the microstructure and properties of dissimilar weld joints between low alloy steel and duplex stainless steel

    Science.gov (United States)

    Wang, Jing; Lu, Min-xu; Zhang, Lei; Chang, Wei; Xu, Li-ning; Hu, Li-hua

    2012-06-01

    To obtain high-quality dissimilar weld joints, the processes of metal inert gas (MIG) welding and tungsten inert gas (TIG) welding for duplex stainless steel (DSS) and low alloy steel were compared in this paper. The microstructure and corrosion morphology of dissimilar weld joints were observed by scanning electron microscopy (SEM); the chemical compositions in different zones were detected by energy-dispersive spectroscopy (EDS); the mechanical properties were measured by microhardness test, tensile test, and impact test; the corrosion behavior was evaluated by polarization curves. Obvious concentration gradients of Ni and Cr exist between the fusion boundary and the type II boundary, where the hardness is much higher. The impact toughness of weld metal by MIG welding is higher than that by TIG welding. The corrosion current density of TIG weld metal is higher than that of MIG weld metal in a 3.5wt% NaCl solution. Galvanic corrosion happens between low alloy steel and weld metal, revealing the weakness of low alloy steel in industrial service. The quality of joints produced by MIG welding is better than that by TIG welding in mechanical performance and corrosion resistance. MIG welding with the filler metal ER2009 is the suitable welding process for dissimilar metals jointing between UNS S31803 duplex stainless steel and low alloy steel in practical application.

  6. Screening of Modified RNA duplexes

    DEFF Research Database (Denmark)

    Schyth, Brian Dall; Bramsen, Jesper Bertram; Kjems, Jørgen

    Because of sequence specific gene targeting activity siRNAs are regarded as promising active compounds in gene medicine. But one serious problem with delivering siRNAs as treatment is the now well-established non-specific activities of some RNA duplexes. Cellular reactions towards double stranded...... RNAs include the 2´-5´ oligoadenylate synthetase system, the protein kinase R, RIG-I and Toll-like receptor activated pathways all resulting in antiviral defence mechanism. We have previously shown that antiviral innate immune reactions against double stranded RNAs could be detected in vivo as partial...... protection against a fish pathogenic virus. This protection corresponded with an interferon response in the fish. Here we use this fish model to screen siRNAs containing various chemical modifications of the RNA backbone for their antiviral activity, the overall aim being identification of an siRNA form...

  7. The Effect of Constant and Pulsed Current Gas Tungsten Arc Welding on Joint Properties of 2205 Duplex Stainless Steel to 316L Austenitic Stainless Steel

    Science.gov (United States)

    Neissi, R.; Shamanian, M.; Hajihashemi, M.

    2016-05-01

    In this study, dissimilar 316L austenitic stainless steel/2205 duplex stainless steel (DSS) joints were fabricated by constant and pulsed current gas tungsten arc welding process using ER2209 DSS as a filler metal. Microstructures and joint properties were characterized using optical and electron scanning microscopy, tensile, Charpy V-notch impact and micro-hardness tests, and cyclic polarization measurements. Microstructural observations confirmed the presence of chromium nitride and delta ferrite in the heat-affected zone of DSS and 316L, respectively. In addition, there was some deviation in the austenite/ferrite ratio of the surface welding pass in comparison to the root welding pass. Besides having lower pitting potential, welded joints produced by constant current gas tungsten arc welding process, consisted of some brittle sigma phase precipitates, which resulted in some impact energy reduction. The tensile tests showed high tensile strength for the weld joints in which all the specimens were broken in 316L base metal.

  8. Solvent reorganization energies in A-DNA, B-DNA, and rhodamine 6G-DNA complexes from molecular dynamics simulations with a polarizable force field.

    Science.gov (United States)

    Vladimirov, Egor; Ivanova, Anela; Rösch, Notker

    2009-04-02

    We estimate solvent reorganization energies lambda(s) of electron transfer (ET) in DNA stacks between positively charged guanine (acceptor) and neutral guanine (donor), as well as in rhodamine 6G (R6G)-DNA complexes between R6G (acceptor) and neutral guanine (donor) from molecular dynamics simulations that used a polarizable force field in combination with a polarizable water model. We compare results from the polarizable scheme with those from a common nonpolarizable analogue. We also discuss the influence of charge sets, separate contributions of solute and solvent electronic polarizations, and partial contributions of different molecular groups to changes of lambda(s) due to electronic polarization. Independent of donor-acceptor distances, solvent reorganization energies of ET processes in DNA duplexes from a polarizable force field are about 30% smaller than the corresponding results from a nonpolarizable force field. The effective optical dielectric constant epsilon(infinity) = 1.5, extracted from pertinent scaling factors, is also independent of the donor-acceptor separation over a wide range of distances, from 3.4 to 50.0 A. Reorganization energies calculated with the polarizable force field agree satisfactorily with experimental data for DNA duplexes. Comparison of results for A-DNA and B-DNA forms as well as for the conformational alignment of the dye relative to the duplex in R6G-DNA complexes demonstrates that the conformation of a duplex hardly affects lambda(s). Among these DNA-related systems, the effective parameter epsilon(infinity) is remarkably constant over a broad range of donor-acceptor distances.

  9. INTERCALATING TRIPLEXES AND DUPLEXES USING ARYL NAPHTHOIMIDAZOL AND PROCESS FOR THE PREPARATION THEREOF

    DEFF Research Database (Denmark)

    2009-01-01

    There is provided an intercalating oligonucleotide for stabilizing natural or modified DNA and RNA triplexes, duplexes and hybrids thereof having the general structure (I) triplex forming oligonucleotides of the invention are capable of binding specifically to double stranded target nucleic acids...

  10. The influence of sintering time on the properties of PM duplex stainless steel

    OpenAIRE

    Z. Brytan; L.A. Dobrzański; M. Actis Grande; Rosso, M.

    2009-01-01

    Purpose: The purpose of this paper is to analyse the effect of sintering time on the pore morphology, microstructural changes, tensile properties and corrosion resistance of vacuum sintered duplex stainless steel.Design/methodology/approach: In presented study PM duplex stainless steels were obtained through mixing base ferritic stainless steel powder with controlled addition of elemental alloying powders and then sintered in a vacuum furnace with argon backfilling at 1250°C for different tim...

  11. Observe Effect of the Bacillus Subtilis Duplex Living Bacterium Granule in the Treatment of Infantile Anorexia%观察枯草杆菌二联活菌颗粒对小儿厌食治疗效果

    Institute of Scientific and Technical Information of China (English)

    李宜海

    2015-01-01

    Objective To observe the bacillus subtilis duplex living bacterium effect of granule for the treatment of infantile anorexia. Methods 38 patients with infantile anorexia, adopt bacillus subtilis duplex living bacterium granule treatment, compared before and after treatment in patients with hemoglobin, red blood cells and so on many indicators of results. Results After treatment in patients with hemoglobin, red blood cells and plasma albumin results of three indicators were (119.49±12.20)g/L, (4.42±0.27) ×1012/L, (150.33±26.71) mg/L, Before and after treatment compared, it was with signiifcant difference (P<0.05). Conclusion Patients with infantile anorexia by bacillus subtilis duplex living bacterium particles after treatment, can improve patients with symptoms of bad appetite, promote the diet nutrition reasonable collocation.%目的:观察枯草杆菌二联活菌颗粒对小儿厌食治疗效果。方法38例小儿厌食患者,均采用枯草杆菌二联活菌颗粒治疗,对比患者治疗前后的血红蛋白、红细胞等多项指标结果。结果治疗后患者的血红蛋白、红细胞以及血浆前白蛋白三项指标结果分别是(119.49±12.20) g/L、(4.42±0.27)×1012/L以及(150.33±26.71)mg/L;治疗前后对比显著差异(P<0.05)。结论小儿厌食患者采用枯草杆菌二联活菌颗粒治疗后,可以改善患者的不良食欲症状,促进其饮食搭配营养合理。

  12. Effect of air conditioning installation position on indoor air distribution in duplex apartment%空调安装位置对复式住宅室内流场的影响

    Institute of Scientific and Technical Information of China (English)

    王晶; 史柏语; 戚译天; 吴国忠; 赵文凯; 李栋

    2012-01-01

    建立复式住宅的三维数理模型,利用CFD软件对复式住宅室内气流组织进行数值模拟,探讨室内温度场和速度场的变化关系,分析2种客厅内不同空调安装位置对复式住宅室内气流组织和热舒适性的影响.结果表明:客厅内空调送风只能同时保证客厅内主要活动区域,以及与其直接连接的相邻且无遮挡空间的空气温度满足要求;在送风时,位于客厅内上层卧室外墙上方空调比位于客厅内西墙墙壁上方的空调对室内气流的扰动作用更大,对室内风速及温度的影响也更大.这对于复式住宅内空调优化设计和节能具有指导意义.%Three-dimensional physical and mathematical models of duplex apartment were established in this paper. Indoor air distribution of the duplex apartment was simulated by using computational fluid dynamics software CFD, and the impact of the different installation position of air conditioning on indoor air distribution in summer were analyzed. By comparing the change relations of the indoor temperature field and velocity field in different circumstances, and the influence of two installation positions of air conditioning that meet the requirements of air distribution and thermal comfort in the duplex apartment were analyzed. Results show that air supply of air conditioning inside the drawing room only meet air temperature requirements of the main activity areas of drawing room and its' directly connected neighboring space without occlusion at the same time; and the disturbance effect of indoor air flow fluctuates more fiercely when air conditioning located in the indoor central position than the border, as well as the influence of indoor wind speed and temperature.

  13. T4 DNA ligase is more than an effective trap of cyclized dsDNA.

    Science.gov (United States)

    Yuan, Chongli; Lou, Xiong Wen; Rhoades, Elizabeth; Chen, Huimin; Archer, Lynden A

    2007-01-01

    T4 DNA ligase is used in standard cyclization assays to trap double-stranded DNA (dsDNA) in low-probability, cyclic or highly bent conformations. The cyclization probability, deduced from the relative yield of cyclized product, can be used in conjunction with statistical mechanical models to extract the bending stiffness of dsDNA. By inserting the base analog 2-aminopurine (2-AP) at designated positions in 89 bp and 94 bp dsDNA fragments, we find that T4 DNA ligase can have a previously unknown effect. Specifically, we observe that addition of T4 ligase to dsDNA in proportions comparable to what is used in the cyclization assay leads to a significant increase in fluorescence from 2-AP. This effect is believed to originate from stabilization of local base-pair opening by formation of transient DNA-ligase complexes. Non-specific binding of T4 ligase to dsDNA is also confirmed using fluorescence correlation spectroscopy (FCS) experiments, which reveal a systematic reduction of dsDNA diffusivity in the presence of ligase. ATP competes with regular DNA for non-covalent binding to the T4 ligase and is found to significantly reduce DNA-ligase complexation. For short dsDNA fragments, however, the population of DNA-ligase complexes at typical ATP concentrations used in DNA cyclization studies is determined to be large enough to dominate the cyclization reaction.

  14. Practical, real-time, full duplex wireless

    KAUST Repository

    Jain, Mayank

    2011-01-01

    This paper presents a full duplex radio design using signal inversion and adaptive cancellation. Signal inversion uses a simple design based on a balanced/unbalanced (Balun) transformer. This new design, unlike prior work, supports wideband and high power systems. In theory, this new design has no limitation on bandwidth or power. In practice, we find that the signal inversion technique alone can cancel at least 45dB across a 40MHz bandwidth. Further, combining signal inversion cancellation with cancellation in the digital domain can reduce self-interference by up to 73dB for a 10MHz OFDM signal. This paper also presents a full duplex medium access control (MAC) design and evaluates it using a testbed of 5 prototype full duplex nodes. Full duplex reduces packet losses due to hidden terminals by up to 88%. Full duplex also mitigates unfair channel allocation in AP-based networks, increasing fairness from 0.85 to 0.98 while improving downlink throughput by 110% and uplink throughput by 15%. These experimental results show that a re- design of the wireless network stack to exploit full duplex capability can result in significant improvements in network performance. © 2011 ACM.

  15. Full-Duplex Communications in Large-Scale Cellular Networks

    KAUST Repository

    AlAmmouri, Ahmad

    2016-04-01

    In-band full-duplex (FD) communications have been optimistically promoted to improve the spectrum utilization and efficiency. However, the penetration of FD communications to the cellular networks domain is challenging due to the imposed uplink/downlink interference. This thesis presents a tractable framework, based on stochastic geometry, to study FD communications in multi-tier cellular networks. Particularly, we assess the FD communications effect on the network performance and quantify the associated gains. The study proves the vulnerability of the uplink to the downlink interference and shows that the improved FD rate gains harvested in the downlink (up to 97%) comes at the expense of a significant degradation in the uplink rate (up to 94%). Therefore, we propose a novel fine-grained duplexing scheme, denoted as α-duplex scheme, which allows a partial overlap between the uplink and the downlink frequency bands. We derive the required conditions to harvest rate gains from the α-duplex scheme and show its superiority to both the FD and half-duplex (HD) schemes. In particular, we show that the α-duplex scheme provides a simultaneous improvement of 28% for the downlink rate and 56% for the uplink rate. We also show that the amount of the overlap can be optimized based on the network design objective. Moreover, backward compatibility is an essential ingredient for the success of new technologies. In the context of in-band FD communication, FD base stations (BSs) should support HD users\\' equipment (UEs) without sacrificing the foreseen FD gains. The results show that FD-UEs are not necessarily required to harvest rate gains from FD-BSs. In particular, the results show that adding FD-UEs to FD-BSs offers a maximum of 5% rate gain over FD-BSs and HD-UEs case, which is a marginal gain compared to the burden required to implement FD transceivers at the UEs\\' side. To this end, we shed light on practical scenarios where HD-UEs operation with FD-BSs outperforms the

  16. Normalized cDNA libraries

    Science.gov (United States)

    Soares, Marcelo B.; Efstratiadis, Argiris

    1997-01-01

    This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to moderate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library.

  17. Thermodynamics of sequence-specific binding of PNA to DNA

    DEFF Research Database (Denmark)

    Ratilainen, T; Holmén, A; Tuite, E

    2000-01-01

    For further characterization of the hybridization properties of peptide nucleic acids (PNAs), the thermodynamics of hybridization of mixed sequence PNA-DNA duplexes have been studied. We have characterized the binding of PNA to DNA in terms of binding affinity (perfectly matched duplexes) and seq......For further characterization of the hybridization properties of peptide nucleic acids (PNAs), the thermodynamics of hybridization of mixed sequence PNA-DNA duplexes have been studied. We have characterized the binding of PNA to DNA in terms of binding affinity (perfectly matched duplexes...

  18. Control of aggregation-induced emission by DNA hybridization

    OpenAIRE

    Li, Shaoguang; Langenegger, Simon Matthias; Häner, Robert

    2013-01-01

    Aggregation-induced emission (AIE) was studied by hybridization of dialkynyl-tetraphenylethylene (DATPE) modified DNA strands. Molecular aggregation and fluorescence of DATPEs are controlled by duplex formation.

  19. Dwell-Time Distribution, Long Pausing and Arrest of Single-Ribosome Translation through the mRNA Duplex

    Directory of Open Access Journals (Sweden)

    Ping Xie

    2015-10-01

    Full Text Available Proteins in the cell are synthesized by a ribosome translating the genetic information encoded on the single-stranded messenger RNA (mRNA. It has been shown that the ribosome can also translate through the duplex region of the mRNA by unwinding the duplex. Here, based on our proposed model of the ribosome translation through the mRNA duplex we study theoretically the distribution of dwell times of the ribosome translation through the mRNA duplex under the effect of a pulling force externally applied to the ends of the mRNA to unzip the duplex. We provide quantitative explanations of the available single molecule experimental data on the distribution of dwell times with both short and long durations, on rescuing of the long paused ribosomes by raising the pulling force to unzip the duplex, on translational arrests induced by the mRNA duplex and Shine-Dalgarno(SD-like sequence in the mRNA. The functional consequences of the pauses or arrests caused by the mRNA duplex and the SD sequence are discussed and compared with those obtained from other types of pausing, such as those induced by “hungry” codons or interactions of specific sequences in the nascent chain with the ribosomal exit tunnel.

  20. Sex determination in 6 bovid species by duplex PCR.

    Science.gov (United States)

    Prashant; Gour, Digpal S; Dubey, Prem P; Jain, Anubhav; Gupta, Subhash C; Joshi, Balwinder K; Kumar, Dinesh

    2008-01-01

    Sex determination in domestic animals is of potential value to livestock breeding programs. The aim of this study was to develop a simple and accurate PCR-based sex determination protocol, which can be applicable to 6 major domesticated species of the family Bovidae, viz. Bos frontalis, B. grunniens, B. indicus, Bubalus bubalis, Capra hircus, and Ovis aries. In silico analysis was done to identify conserved DNA sequence in the HMG box region of the sex-determining region of the Y-chromosome (SRY gene) across the bovids. Duplex PCR assay, including the SRY gene and the GAPDH housekeeping gene, was optimized by using genomic DNA extracted from blood samples of known sex. It was possible to identify the sex of animals by amplifying both gender-specific (SRY) and autosomal (GAPDH) genes simultaneously in the duplex reaction, with the male yielding two bands and the female one band. The protocol was subjected to a blind test that showed a 100 percent specificity and accuracy, thus it can be used in sex determination in livestock breeding programs.

  1. Use of a Combined Duplex PCR/Dot Blot Assay for more sensitive genetic characterization

    Directory of Open Access Journals (Sweden)

    Erin Curry

    2008-01-01

    Full Text Available A reliable and sensitive method of genetic analysis is necessary to detect multiple specific nucleic acid sequences from samples containing limited template. The most widely utilized method of specific gene detection, polymerase chain reaction (PCR, imparts inconsistent results when assessing samples with restricted template, especially in a multiplex reaction when copies of target genes are unequal. This study aimed to compare two methods of PCR product analysis, fluorescent detection following agarose gel electrophoresis or dot blot hybridization with chemiluminescent evaluation, in the detection of a single copy gene (SRY and a multicopy gene (β-actin. Bovine embryo sex determination was employed to exploit the limited DNA template available and the target genes of unequal copies. Primers were used either independently or together in a duplex reaction with purified bovine genomic DNA or DNA isolated from embryos. When used independently, SRY and β-actin products were detected on a gel at the equivalent of 4-cell or 1-cell of DNA, respectively; however, the duplex reaction produced visible SRY bands at the 256 cell DNA equivalent and β-actin products at the 64 cell DNA equivalent. Upon blotting and hybridization of the duplex PCR reaction, product was visible at the 1–4 cell DNA equivalent. Duplex PCR was also conducted on 186 bovine embryos and product was subjected to gel electrophoresis or dot-blot hybridization in duplicate. Using PCR alone, sex determination was not possible for 22.6% of the samples. Using PCR combined with dot blot hybridization, 100.0% of the samples exhibited either both the male specific and β-actin products or the β-actin signal alone, indicating that the reaction worked in all samples. This study demonstrated that PCR amplification followed by dot blot hybridization provided more conclusive results in the evaluation of samples with low DNA concentrations and target genes of unequal copies.

  2. Effect of Sintering Atmosphere and Solution Treatment on Density, Microstructure and Tensile Properties of Duplex Stainless Steels Developed from Pre-alloyed Powders

    Science.gov (United States)

    Murali, Arun Prasad; Mahendran, Sudhahar; Ramajayam, Mariappan; Ganesan, Dharmalingam; Chinnaraj, Raj Kumar

    2017-01-01

    In this research, Powder Metallurgy (P/M) of Duplex Stainless Steels (DSS) of different compositions were prepared through pre-alloyed powders and elemental powders with and without addition of copper. The powder mix was developed by pot mill for 12 h to obtain the homogeneous mixture of pre-alloyed powder with elemental compositions. Cylindrical green compacts with the dimensions of 30 mm diameter and 12 mm height were compacted through universal testing machine at a pressure level of 560 ± 10 MPa. These green compacts were sintered at 1350 °C for 2 h in hydrogen and argon atmospheres. Some of the sintered stainless steel preforms were solution treated at 1050 °C followed by water quenching. The sintered as well as solution treated samples were analysed by metallography examination, Scanning Electron Microscopy and evaluation of mechanical properties. Ferrite content of sintered and solution treated DSS were measured by Fischer Ferritoscope. It is inferred that the hydrogen sintered DSS depicted better density (94% theoretical density) and tensile strength (695 MPa) than the argon sintered steels. Similarly the microstructure of solution treated DSS revealed existence of more volume of ferrite grains than its sintered condition. Solution treated hydrogen sintered DSS A (50 wt% 316L + 50 wt% 430L) exhibited higher tensile strength of 716 MPa and elongation of 17%, which are 10-13% increment than the sintered stainless steels.

  3. Effect of Sintering Atmosphere and Solution Treatment on Density, Microstructure and Tensile Properties of Duplex Stainless Steels Developed from Pre-alloyed Powders

    Science.gov (United States)

    Murali, Arun Prasad; Mahendran, Sudhahar; Ramajayam, Mariappan; Ganesan, Dharmalingam; Chinnaraj, Raj Kumar

    2017-10-01

    In this research, Powder Metallurgy (P/M) of Duplex Stainless Steels (DSS) of different compositions were prepared through pre-alloyed powders and elemental powders with and without addition of copper. The powder mix was developed by pot mill for 12 h to obtain the homogeneous mixture of pre-alloyed powder with elemental compositions. Cylindrical green compacts with the dimensions of 30 mm diameter and 12 mm height were compacted through universal testing machine at a pressure level of 560 ± 10 MPa. These green compacts were sintered at 1350 °C for 2 h in hydrogen and argon atmospheres. Some of the sintered stainless steel preforms were solution treated at 1050 °C followed by water quenching. The sintered as well as solution treated samples were analysed by metallography examination, Scanning Electron Microscopy and evaluation of mechanical properties. Ferrite content of sintered and solution treated DSS were measured by Fischer Ferritoscope. It is inferred that the hydrogen sintered DSS depicted better density (94% theoretical density) and tensile strength (695 MPa) than the argon sintered steels. Similarly the microstructure of solution treated DSS revealed existence of more volume of ferrite grains than its sintered condition. Solution treated hydrogen sintered DSS A (50 wt% 316L + 50 wt% 430L) exhibited higher tensile strength of 716 MPa and elongation of 17%, which are 10-13% increment than the sintered stainless steels.

  4. Effects of Duplex Nitriding and TiN Coating Treatment on Wear Resistance, Corrosion Resistance and Biocompatibility of Ti6Al4V Alloy

    Science.gov (United States)

    Kao, W. H.; Su, Y. L.; Hsieh, Y. T.

    2017-08-01

    Ti6Al4V alloy substrates were nitrided at 900 °C. TiN coatings were then deposited on the nitrided substrates using a closed-field unbalanced magnetron sputtering system. The microstructure, hardness and adhesion properties of the TiN-N-Ti6Al4V substrates were evaluated and compared with those of an untreated Ti6Al4V sample, a nitrided Ti6Al4V sample and a TiN-coated Ti6Al4V sample, respectively. The tribological properties of the various samples were investigated by means of reciprocating sliding wear tests performed in 0.9 wt.% NaCl solution against 316L, Si3N4 and Ti6Al4V balls, respectively. In addition, the corrosion resistance was evaluated using potentiodynamic polarization tests. Finally, the biocompatibility of the samples was investigated by observing the attachment and growth of purified mouse leukemic monocyte/macrophage cells (Raw 264.7) on the sample surface after culturing periods of 24, 72 and 120 h, respectively. Overall, the results showed that the duplex nitriding/TiN coating treatment significantly improved the tribological, anti-corrosion and biocompatibility properties of the original Ti6Al4V alloy.

  5. Purification and characterization of Rad3 ATPase/DNA helicase from Saccharomyces cerevisiae.

    Science.gov (United States)

    Harosh, I; Naumovski, L; Friedberg, E C

    1989-12-05

    The Rad3 ATPase/DNA helicase was purified to physical homogeneity from extracts of yeast cells containing overexpressed Rad3 protein. The DNA helicase can unwind duplex regions as short as 11 base pairs in a partially duplex circular DNA substrate and does so by a strictly processive mechanism. On partially duplex linear substrates, the enzyme has a strict 5'----3' polarity with respect to the single strand to which it binds. Nicked circular DNA is not utilized as a substrate, and the enzyme requires single-stranded gaps between 5 and 21 nucleotides long to unwind oligonucleotide fragments from partially duplex linear molecules. The enzyme also requires duplex regions at least 11 base pairs long when these are present at the ends of linear molecules. Rad3 DNA helicase activity is inhibited by the presence of ultraviolet-induced photoproducts in duplex regions of partially duplex circular molecules.

  6. In-Band α-Duplex Scheme for Cellular Networks: A Stochastic Geometry Approach

    KAUST Repository

    Alammouri, Ahmad

    2016-07-13

    In-band full-duplex (FD) communications have been optimistically promoted to improve the spectrum utilization and efficiency. However, the penetration of FD communications to the cellular networks domain is challenging due to the imposed uplink/downlink interference. This paper presents a tractable framework, based on stochastic geometry, to study FD communications in cellular networks. Particularly, we assess the FD communications effect on the network performance and quantify the associated gains. The study proves the vulnerability of the uplink to the downlink interference and shows that FD rate gains harvested in the downlink (up to 97%) come at the expense of a significant degradation in the uplink rate (up to 94%). Therefore, we propose a novel fine-grained duplexing scheme, denoted as -duplex scheme, which allows a partial overlap between the uplink and the downlink frequency bands. We derive the required conditions to harvest rate gains from the -duplex scheme and show its superiority to both the FD and half-duplex (HD) schemes. In particular, we show that the -duplex scheme provides a simultaneous improvement of 28% for the downlink rate and 56% for the uplink rate. Finally, we show that the amount of the overlap can be optimized based on the network design objective.

  7. Phase Transformations in Cast Duplex Stainless Steels

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Yoon-Jun [Iowa State Univ., Ames, IA (United States)

    2004-01-01

    Duplex stainless steels (DSS) constitute both ferrite and austenite as a matrix. Such a microstructure confers a high corrosion resistance with favorable mechanical properties. However, intermetallic phases such as σ and χ can also form during casting or high-temperature processing and can degrade the properties of the DSS. This research was initiated to develop time-temperature-transformation (TTT) and continuous-cooling-transformation (CCT) diagrams of two types of cast duplex stainless steels, CD3MN (Fe-22Cr-5Ni-Mo-N) and CD3MWCuN (Fe-25Cr-7Ni-Mo-W-Cu-N), in order to understand the time and temperature ranges for intermetallic phase formation. The alloys were heat treated isothermally or under controlled cooling conditions and then characterized using conventional metallographic methods that included tint etching, and also using electron microscopy (SEM, TEM) and wavelength dispersive spectroscopy (WDS). The kinetics of intermetallic-phase (σ + χ) formation were analyzed using the Johnson-Mehl-Avrami (MA) equation in the case of isothermal transformations and a modified form of this equation in the case of continuous cooling transformations. The rate of intermetallic-phase formation was found to be much faster in CD3MWCuN than CD3MN due mainly to differences in the major alloying contents such as Cr, Ni and Mo. To examine in more detail the effects of these elements of the phase stabilities; a series of eight steel castings was designed with the Cr, Ni and Mo contents systematically varied with respect to the nominal composition of CD3MN. The effects of varying the contents of alloying additions on the formation of intermetallic phases were also studied computationally using the commercial thermodynamic software package, Thermo-Calc. In general, σ was stabilized with increasing Cr addition and χ by increasing Mo addition. However, a delicate balance among Ni and other minor elements such as N and Si also exists. Phase equilibria in DSS can be affected by

  8. Phase Transformations in Cast Duplex Stainless Steels

    Energy Technology Data Exchange (ETDEWEB)

    Yoon-Jun Kim

    2004-12-19

    Duplex stainless steels (DSS) constitute both ferrite and austenite as a matrix. Such a microstructure confers a high corrosion resistance with favorable mechanical properties. However, intermetallic phases such as {sigma} and {chi} can also form during casting or high-temperature processing and can degrade the properties of the DSS. This research was initiated to develop time-temperature-transformation (TTT) and continuous-cooling-transformation (CCT) diagrams of two types of cast duplex stainless steels, CD3MN (Fe-22Cr-5Ni-Mo-N) and CD3MWCuN (Fe-25Cr-7Ni-Mo-W-Cu-N), in order to understand the time and temperature ranges for intermetallic phase formation. The alloys were heat treated isothermally or under controlled cooling conditions and then characterized using conventional metallographic methods that included tint etching, and also using electron microscopy (SEM, TEM) and wavelength dispersive spectroscopy (WDS). The kinetics of intermetallic-phase ({sigma} + {chi}) formation were analyzed using the Johnson-Mehl-Avrami (MA) equation in the case of isothermal transformations and a modified form of this equation in the case of continuous cooling transformations. The rate of intermetallic-phase formation was found to be much faster in CD3MWCuN than CD3MN due mainly to differences in the major alloying contents such as Cr, Ni and Mo. To examine in more detail the effects of these elements of the phase stabilities; a series of eight steel castings was designed with the Cr, Ni and Mo contents systematically varied with respect to the nominal composition of CD3MN. The effects of varying the contents of alloying additions on the formation of intermetallic phases were also studied computationally using the commercial thermodynamic software package, Thermo-Calc. In general, {sigma} was stabilized with increasing Cr addition and {chi} by increasing Mo addition. However, a delicate balance among Ni and other minor elements such as N and Si also exists. Phase equilibria in

  9. Nucleic acid nanomaterials: Silver-wired DNA

    Science.gov (United States)

    Auffinger, Pascal; Ennifar, Eric

    2017-10-01

    DNA double helical structures are supramolecular assemblies that are typically held together by classical Watson-Crick pairing. Now, nucleotide chelation of silver ions supports an extended silver-DNA hybrid duplex featuring an uninterrupted silver array.

  10. Effective interactions of DNA-stars

    Science.gov (United States)

    Abaurrea Velasco, Clara; Likos, Christos N.; Kahl, Gerhard

    2015-09-01

    We put forward a model that allows the calculation of the effective potential of two interacting DNA-stars, i.e., three-armed, Y-shaped, charged macromolecules, built up by three intertwined single-stranded DNAs. These particles are assumed to float on a flat interface separating two media with different dielectric properties. As the only input, our model requires the charge density along the branches and the interaction between two infinitesimally short segments, along two interacting rods. With this effective interaction at hand, a detailed investigations of the self-assembly scenarios of these molecules either via computer simulations or via theoretical frameworks comes within reach.

  11. Induced effect of irradiated exogenous DNA on wheat

    Institute of Scientific and Technical Information of China (English)

    李忠杰; 孙光祖; 等

    1996-01-01

    Irradiated exogenous DNA introduced into wheat can give rise to break of DNA-chain and damage of part of alkali radicals.Introducing exogenous DNA irradiated by γ rays could increase Do fructification rate and decrease seed size and lumpness.These tendencies became obvious with dose increase.In comparison with control DNA,introducing DNA irradiated could raise evidently mutagenic effect of pollen tube pathway technique.

  12. On full duplex Gaussian relay channels with self-interference

    KAUST Repository

    Behboodi, Arash

    2016-08-15

    Self interference (SI) in full duplex (FD) systems is the interference caused by the transmission stream on the reception stream. Being one of the main restrictive factors for performance of practical full duplex systems, however, not too much is known about its effect on the fundamental limits of relaying systems. In this work, we consider the full duplex three-node relay channel with SI where SI is modeled as an additive Gaussian noise whose variance is dependent on instantaneous input power. The classical achievable rates and upper bounds for the single three-node relay channel no longer apply due to the structure of SI. Achievable rates for Decode-and-Forward (DF) and Compress-and-Forward (CF) and upper bounds on the capacity are derived assuming Gaussian inputs and SI. The deterministic model is also introduced and its capacity is characterized. The optimal joint source-relay distributions is discussed. Numerical results are provided comparing the achievable rates and upper bound. © 2016 IEEE.

  13. Dynamic Restoration Processes in a 23Cr-6Ni-3Mo Duplex Stainless Steel: Effect of Austenite Morphology and Interface Characteristics

    Science.gov (United States)

    Haghdadi, N.; Cizek, P.; Beladi, H.; Hodgson, P. D.

    2017-07-01

    The austenite and ferrite microstructure evolution and restoration mechanisms were studied during hot uniaxial compression of a 23Cr-6Ni-3Mo duplex stainless steel with two markedly different austenite morphologies (i.e., equiaxed and Widmanstätten). The deformation was performed at a temperature of 1273 K (1000 °C) at a strain rate of 0.1 s-1. The strain was preferentially partitioned in ferrite for both the microstructures studied. Both austenite morphologies displayed frequent splitting into complex-shaped deformation bands, containing dislocation cells and local stacking faults. Equiaxed austenite was favorable to the local development of microbands (MBs), while its Widmanstätten counterpart appeared to be completely resistant to their formation. This was attributed to the complexity of deformation inside the irregularly shaped Widmanstätten plates precluding the formation of self-screening MB arrays. The MB boundaries were typically aligned along highly stressed slip planes. The presence of discontinuous dynamic recrystallization (DDRX) within both the austenite morphologies was very limited. A slightly higher fraction of DDRX was detected in Widmanstätten austenite, compared to equiaxed austenite, which was ascribed to its higher contribution to the overall deformation and lower fraction of low-mobility coherent twin boundaries. Furthermore, it was demonstrated that continuous dynamic recrystallization (CDRX) was the main restoration mechanism within ferrite for both the microstructure types studied. The CDRX development within ferrite was accelerated in the microstructure with equiaxed austenite. This was related to the comparatively lower fraction of coherent interphases in this microstructure, which would hinder the slip transmission across the interphase and make the strain concentrate within ferrite.

  14. Effect of oxygen on weld shape and crystallographic orientation of duplex stainless steel weld using advanced A-TIG (AA-TIG) welding method

    Energy Technology Data Exchange (ETDEWEB)

    Zou, Ying, E-mail: yingzou@jwri.osaka-u.ac.jp; Ueji, Rintaro; Fujii, Hidetoshi

    2014-05-01

    The double-shielded advanced A-TIG (AA-TIG) welding method was adopted in this study for the welding of the SUS329J4L duplex stainless steel with the shielding gases of different oxygen content levels. The oxygen content in the shielding gas was controlled by altering the oxygen content in the outer layer gas, while the inner layer remained pure argon to suppress oxidation on the tungsten electrode. As a result, a deep weld penetration was obtained due to the dissolution of oxygen into the weld metals. Additionally, the microstructure of the weld metal was changed by the dissolution of oxygen. The austenite phase at the ferrite grain boundary followed a Kurdjumov–Sachs (K–S) orientation relationship with the ferrite matrix phase at any oxide content. On the other hand, the orientation relationship between the intragranular austenite phase and the ferrite matrix phase exhibited different patterns under different oxygen content levels. When there was little oxide in the fusion zone, only a limited part of the intragranular austenite phase and the ferrite matrix phase followed the K–S orientation relationship. With the increase of the oxide, the correspondence of the K–S relationship increased and fit very well in the 2.5% O{sub 2} shielded sample. The investigation of this phenomenon was carried out along with the nucleation mechanisms of the intragranular austenite phases. - Highlights: • Weld penetration increased with the increase of the oxygen content. • Average diameter and number density of oxide were changed by the oxygen content. • K-S relationship of Widmanstätten austenite/ferrite wasn’t varied by oxide. • Orientation relationship of intragranular austenite/ferrite was varied by oxide.

  15. Effect of Hecogenin on DNA instability

    Directory of Open Access Journals (Sweden)

    Marina Sampaio Cruz

    2016-01-01

    Full Text Available Hecogenin is a sapogenin found in Agave species in high quantities and is responsible for the many therapeutic effects of these medicinal plants. In addition, this compound is also widely used in the pharmaceutical industry as a precursor for the synthesis of steroidal hormones and anti-inflammatory drugs. Despite Hecogenin being widely used, little is known about its toxicological properties. Therefore, the present study aimed to investigate the cytotoxic, genotoxic and mutagenic effects of Hecogenin on HepG2 cells. Cytotoxicity was analyzed using the MTT test. Then, genotoxic and mutagenic potentials were assessed by comet assay and cytokinesis-block micronucleus assay, respectively. Cytotoxic effect was observed only when cells were exposed to concentrations of Hecogenin equal or higher than 100 μM. Although a lower concentration of Hecogenin caused DNA damage, a reduction on nuclear mutagenic markers in HepG2 cells was observed. The results indicated that Hecogenin treatment generated DNA damage, but in fact it would be repaired, avoiding dissemination of the damage throughout the cell division. Further studies need to be performed to confirm the observed protective effect of Hecogenin against genomic instability.

  16. Effects of oxaliplatin on DNA condensation

    Institute of Scientific and Technical Information of China (English)

    JU HaiPeng; ZHANG HongYan; LI Wei; WANG PengYe

    2014-01-01

    In this paper the interactions between DNA and anti-cancer drug oxaliplatin were investigated by using magnetic tweezers.The dynamics of DNA condensation due to oxaliplatin was traced under various forces.It is found that torsion constraint in DNA enhances the ability of oxaliplatin for shortening DNA.The transplatin helps oxaliplatin combine to DNA and increase the rate of DNA condensation.All these results are consistent to the previously proposed model and are helpful for further investigation of interaction between DNA and oxaliplatin.

  17. Molecular mechanisms of DNA photodamage

    Energy Technology Data Exchange (ETDEWEB)

    Starrs, S.M

    2000-05-01

    Photodamage in DNA, caused by ultraviolet (UV) light, can occur by direct excitation of the nucleobases or indirectly via the action of photosensitisers. Such, DNA photodamage can be potentially mutagenic or lethal. Among the methods available for detecting UV-induced DNA damage, gel sequencing protocols, utilising synthetic oligodeoxyribonucleotides as targets for UV radiation, allow photolesions to be mapped at nucleotide resolution. This approach has been applied to investigate both DNA damage mechanisms. Following a general overview of DNA photoreactivity, and a description of the main experimental procedures, Chapter 3 identifies the origin of an anomalous mobility shift observed in purine chemical sequence ladders that can confuse the interpretation of DNA cleavage results; measures to abolish this shift are also described. Chapters 4 and 5 examine the alkali-labile DNA damage photosensitised by representative nonsteroidal antiinflammatory drugs (NSAIDs) and the fluoroquinolone antibiotics. Suprofen was the most photoactive NSAID studied, producing different patterns of guanine-specific damage in single-stranded and duplex DNA. Uniform modification of guanine bases, typifying attack by singlet oxygen, was observed in single-stranded oligodeoxyribonucleotides. In duplex molecules, modification was limited to the 5'-G of GG doublets, which is indicative of an electron transfer. The effect of quenchers and photoproduct analysis substantiated these findings. The quinolone, nalidixic acid, behaves similarly. The random base cleavage photosensitised by the fluoroquinolones, has been attributed to free radicals produced during their photodecomposition. Chapter 6 addresses the photoreactivity of purines within unusual DNA structures formed by the repeat sequences (GGA){sub n} and (GA){sub n}, and a minihairpin. There was no definitive evidence for enhanced purine reactivity caused by direct excitation. Finally, Chapter 7 investigates the mutagenic potential of a

  18. Wavelength-tunable duplex integrated light source

    Science.gov (United States)

    Okamoto, Hiroshi; Yasaka, Hiroshi; Oe, Kunishige

    1996-04-01

    A monolithically integrated opto-electronic device is proposed as a fast wavelength-switching light source. This tunable duplex integrated light source comprises two wavelength-tunable distributed Bragg reflector (DBR) laser diodes (LDs), two MQW-electro-absorption optical switches, a Y-shaped waveguide coupler, a MQW-electro-absorption modulator, and two thermal drift compensators (TDCs). The wavelength-switching time of the optical switches was estimated to be 60 ps including a 50-ps rise time for the electrical-pulse generator. The wavelength of a 10-Gbit/s NRZ-modulated optical signal can be switched without bit loss. The function of the TDCs is to keep the device-chip temperature constant. Thermal-transient- induced wavelength drift with a millisecond-order time constant, which has been reported for DBR-LDs, and thermal crosstalk between the tuning regions of the integrated LDs, which causes wavelength fluctuation, are effectively suppressed by thermal-drift-compensation operation using the TDCs.

  19. Ferrite Quantification Methodologies for Duplex Stainless Steel

    Directory of Open Access Journals (Sweden)

    Arnaldo Forgas Júnior

    2016-07-01

    Full Text Available In order to quantify ferrite content, three techniques, XRD, ferritoscope and optical metallography, were applied to a duplex stainless steel UNS S31803 solution-treated for 30 min at 1,000, 1,100 and 1,200 °C, and then compared to equilibrium of phases predicted by ThermoCalc® simulation. As expected, the microstructure is composed only by austenite and ferrite phases, and ferrite content increases as the solution treatment temperature increases. The microstructure presents preferred grains orientation along the rolling directions even for a sample solution treated for 30 min at 1,200 °C. For all solution treatment temperatures, the ferrite volume fractions obtained by XRD measurements were higher than those achieved by the other two techniques and ThermoCalc® simulation, probably due to texturing effect of previous rolling process. Values obtained by quantitative metallography look more assertive as it is a direct measurement method but the ferritoscope technique should be considered mainly for in loco measurement.

  20. Full Duplex Communications in 5G Small Cells

    DEFF Research Database (Denmark)

    Mahmood, Nurul Huda; Gatnau, Marta; Berardinelli, Gilberto

    2017-01-01

    Full duplex communication promises system performance improvement over conventional half duplex communication by allowing simultaneous transmission and reception. However, such concurrent communication results in strong self interference and an increase in the overall network interference, and ca...

  1. LNA effects on DNA binding and conformation

    DEFF Research Database (Denmark)

    Pabon-Martinez, Y Vladimir; Xu, You; Villa, Alessandra

    2017-01-01

    hybridization analysis and electrophoretic mobility shift assay with molecular dynamics (MD) simulations to better understand the underlying structural features of modified ONs in stabilizing duplex- and triplex structures. Particularly, we investigated the role played by the position and number of locked...... nucleic acid (LNA) substitutions in the ON when targeting a c-MYC or FXN (Frataxin) sequence. We found that LNA-containing single strand TFOs are conformationally pre-organized for major groove binding. Reduced content of LNA at consecutive positions at the 3'-end of a TFO destabilizes the triplex...... structure, whereas the presence of Twisted Intercalating Nucleic Acid (TINA) at the 3'-end of the TFO increases the rate and extent of triplex formation. A triplex-specific intercalating benzoquinoquinoxaline (BQQ) compound highly stabilizes LNA-containing triplex structures. Moreover, LNA...

  2. The effects of 4-MEI on cell proliferation, DNA breaking and DNA fragmentation.

    Science.gov (United States)

    Tazehkand, M Norizadeh; Moridikia, A; Hajipour, O; Valipour, E; Timocin, T; Topaktas, M; Yilmaz, M B

    4-Methylimidazole (4-MEI) is a color widely found in cola drinks, roasted foods, grilled meats, coffee and other foods. This study was aimed to investigate the 4-MEI effects on the cell proliferation, purified circular DNA and DNA from cells of rats treated with the 4-MEI.In this study, mouse 3T3-L1 cell line was treated with 4-MEI at concentrations of 300, 450, 600 and 750 µg/mL for 24 hours and 48 hours periods, after that cytotoxic effect of the 4-MEI was studied by MTT test. Also, the effect of 4-MEI on purified circular DNA (pET22b) was investigated by treating of the DNA with 4-MEI concentrations of 300, 450, 600 and 750 µg/ml. DNA was extracted from liver cells of rats that have been treated with 4-MEI doses of 25 and 50 mg/kg for 10 week and it was subjected to agarose gel electrophoreses analyses.4-MEI significantly inhibited cell proliferation of 3T3-L1 cell line at highest concentration for 24 h and at all concentration for 48 h treatment time. DNA fragmentation assay showed that 4-MEI at 50 mg/kg concentration clearly produced characteristic DNA smear and no DNA laddering (200bp) was observed when mouse was exposed to 4-MEI. The results obtained from plasmid DNA damaging assay showed that 4-MEI has noeffect on the DNA, because the electrophoretic pattern of DNA treated with 4-MEI showed three bands on agarose gel electrophoresis as it was for untreated control. 4-MEI showed cytotoxic effect on 3T3-L1 cells but no effect on plasmid DNA breaking. According to DNA fragmentation assay 4-MEI has necrosis effects on mouse liver cells (Tab. 1, Fig. 4, Ref. 27).

  3. Plastomes of the green algae Hydrodictyon reticulatum and Pediastrum duplex (Sphaeropleales, Chlorophyceae

    Directory of Open Access Journals (Sweden)

    Hilary A. McManus

    2017-05-01

    Full Text Available Background Comparative studies of chloroplast genomes (plastomes across the Chlorophyceae are revealing dynamic patterns of size variation, gene content, and genome rearrangements. Phylogenomic analyses are improving resolution of relationships, and uncovering novel lineages as new plastomes continue to be characterized. To gain further insight into the evolution of the chlorophyte plastome and increase the number of representative plastomes for the Sphaeropleales, this study presents two fully sequenced plastomes from the green algal family Hydrodictyaceae (Sphaeropleales, Chlorophyceae, one from Hydrodictyon reticulatum and the other from Pediastrum duplex. Methods Genomic DNA from Hydrodictyon reticulatum and Pediastrum duplex was subjected to Illumina paired-end sequencing and the complete plastomes were assembled for each. Plastome size and gene content were characterized and compared with other plastomes from the Sphaeropleales. Homology searches using BLASTX were used to characterize introns and open reading frames (orfs ≥ 300 bp. A phylogenetic analysis of gene order across the Sphaeropleales was performed. Results The plastome of Hydrodictyon reticulatum is 225,641 bp and Pediastrum duplex is 232,554 bp. The plastome structure and gene order of H. reticulatum and P. duplex are more similar to each other than to other members of the Sphaeropleales. Numerous unique open reading frames are found in both plastomes and the plastome of P. duplex contains putative viral protein genes, not found in other Sphaeropleales plastomes. Gene order analyses support the monophyly of the Hydrodictyaceae and their sister relationship to the Neochloridaceae. Discussion The complete plastomes of Hydrodictyon reticulatum and Pediastrum duplex, representing the largest of the Sphaeropleales sequenced thus far, once again highlight the variability in size, architecture, gene order and content across the Chlorophyceae. Novel intron insertion sites and unique

  4. Photochemistry of psoralen-DNA adducts, biological effects of psoralen-DNA adducts, applications of psoralen-DNA photochemistry

    Energy Technology Data Exchange (ETDEWEB)

    Shi, Yun-bo

    1988-03-01

    This thesis consists of three main parts and totally eight chapters. In Part I, The author will present studies on the photochemistry of psoralen-DNA adducts, specifically, the wavelength dependencies for the photoreversals of thymidine-HMT (4'-hydroxymethyl-4, 5', 8-trimenthylpsoralen) monoadducts and diadduct and the same adducts incorporated in DNA helices and the wavelength dependecies for the photocrossslinking of thymidine-HMT monoadducts in double-stranded helices. In Part II, The author will report some biological effects of psoralen-DNA adducts, i.e., the effects on double-stranded DNA stability, DNA structure, and transcription by E. coli and T7 RNA polymerases. Finally, The author will focus on the applications of psoralen-DNA photochemistry to investigation of protein-DNA interaction during transcription, which includes the interaction of E. coli and T7 RNA polymerases with DNA in elongation complexes arrested at specific psoralen-DNA adduct sites as revealed by DNase I footprinting experiments. 123 refs., 52 figs., 12 tabs.

  5. Construction and deconstruction of PLL/DNA multilayered films for DNA delivery: effect of ionic strength.

    Science.gov (United States)

    Ren, Kefeng; Wang, Youxiang; Ji, Jian; Lin, Quankui; Shen, Jiacong

    2005-12-10

    Through the layer-by-layer (LbL) self-assembly technique, DNA was incorporated into the multilayered films with poly-l-lysine (PLL). The effect of ionic strength on the construction and deconstruction of the PLL/DNA films was investigated. It was found that the salt concentration of the deposition solution had a significant effect on the construction of the films, which might attribute to the effect of salt ions on the conformation of polyelectrolytes and interaction between PLL and DNA molecules. A salt-induced deconstruction of the PLL/DNA films was observed. The extent of the deconstruction increased with the salt concentration in the incubation solution. The mechanism of the deconstruction was discussed. Taking the advantages of the LbL technique, the erasable PLL/DNA films could deposit onto a variety of surfaces, such as vascular stent, intervention catheter and tissue engineering scaffold, to serve as a novel DNA delivery system.

  6. Rapid Non-Crosslinking Aggregation of DNA-Functionalized Gold Nanorods and Nanotriangles for Colorimetric Single-Nucleotide Discrimination.

    Science.gov (United States)

    Wang, Guoqing; Akiyama, Yoshitsugu; Takarada, Tohru; Maeda, Mizuo

    2016-01-01

    Gold nanoparticles modified with DNA duplexes are rapidly and spontaneously aggregated at high ionic strength. In contrast, this aggregation is greatly suppressed when the DNA duplex has a single-base mismatch or a single-nucleotide overhang located at the outermost surface of the particle. These colloidal features emerge irrespective of the size and composition of the particle core; however, the effects of the shape remain unexplored. Using gold nanorods and nanotriangles (nanoplatelets), we show herein that both remarkable rapidity in colloidal aggregation and extreme susceptibility to DNA structural perturbations are preserved, regardless of the shape and aspect ratio of the core. It is also demonstrated that the DNA-modified gold nanorods and nanotriangles are applicable to naked-eye detection of a single-base difference in a gene model. The current study corroborates the generality of the unique colloidal properties of DNA-functionalized nanoparticles, and thus enhances the feasibility of their practical use.

  7. The effect of chronic alcohol consumption on mitochondrial DNA mutagenesis in human blood.

    Science.gov (United States)

    von Wurmb-Schwark, N; Ringleb, A; Schwark, T; Broese, T; Weirich, S; Schlaefke, D; Wegener, R; Oehmichen, M

    2008-01-01

    The 4977bp deletion of mitochondrial DNA (mtDNA) is known to accumulate with increasing age in post mitotic tissues. Recently, studies came out detecting this specific alteration also in fast replicating cells, e.g. in blood or skin tissue, often in correlation to specific diseases or -- specifically in skin -- external stressors such as UV radiation. In this study, we investigated mitochondrial mutagenesis in 69 patients with a chronic alcoholic disease and 46 age matched controls with a moderate drinking behavior. Two different fragments, specific for total and for deleted mtDNA (dmtDNA) were amplified in a duplex-PCR. A subsequent fragment analysis was performed and for relative quantification, the quotient of the peak areas of amplification products specific for deleted and total mtDNA was determined. Additionally, a real time PCR was performed to quantify mtDNA copy number. The relative amount of 4977bp deleted mtDNA in alcoholics was significantly increased compared to controls. On the other hand, no difference regarding the mtDNA/nuclear DNA ratio in both investigated groups was detected. Additionally, no age dependence could be found nor in alcoholics, neither in the control group. These findings indicate that mtDNA mutagenesis in blood can be influenced by stressors such as alcohol. Ethanol seems to be a significant factor to alter mitochondrial DNA in blood and might be an additional contributor for the cellular aging process.

  8. The effect of chronic alcohol consumption on mitochondrial DNA mutagenesis in human blood

    Energy Technology Data Exchange (ETDEWEB)

    Wurmb-Schwark, N. von [Institute of Legal Medicine, Christian Albrecht University of Kiel, Arnold-Heller-Str. 12, 24105 Kiel (Germany)], E-mail: nvonwurmb@rechtsmedizin.uni-kiel.de; Ringleb, A.; Schwark, T. [Institute of Legal Medicine, Christian Albrecht University of Kiel, Arnold-Heller-Str. 12, 24105 Kiel (Germany); Broese, T.; Weirich, S.; Schlaefke, D. [Clinic of Psychiatry and Psychotherapy, University of Rostock, Gehlsheimer Str. 20, Rostock (Germany); Wegener, R. [Institute of Legal Medicine, St-Georg-Str. 108, University of Rostock, 18055 Rostock (Germany); Oehmichen, M. [Institute of Legal Medicine, Christian Albrecht University of Kiel, Arnold-Heller-Str. 12, 24105 Kiel (Germany)

    2008-01-01

    The 4977 bp deletion of mitochondrial DNA (mtDNA) is known to accumulate with increasing age in post mitotic tissues. Recently, studies came out detecting this specific alteration also in fast replicating cells, e.g. in blood or skin tissue, often in correlation to specific diseases or - specifically in skin - external stressors such as UV radiation. In this study, we investigated mitochondrial mutagenesis in 69 patients with a chronic alcoholic disease and 46 age matched controls with a moderate drinking behavior. Two different fragments, specific for total and for deleted mtDNA (dmtDNA) were amplified in a duplex-PCR. A subsequent fragment analysis was performed and for relative quantification, the quotient of the peak areas of amplification products specific for deleted and total mtDNA was determined. Additionally, a real time PCR was performed to quantify mtDNA copy number. The relative amount of 4977 bp deleted mtDNA in alcoholics was significantly increased compared to controls. On the other hand, no difference regarding the mtDNA/nuclear DNA ratio in both investigated groups was detected. Additionally, no age dependence could be found nor in alcoholics, neither in the control group. These findings indicate that mtDNA mutagenesis in blood can be influenced by stressors such as alcohol. Ethanol seems to be a significant factor to alter mitochondrial DNA in blood and might be an additional contributor for the cellular aging process.

  9. Study of MMLV RT- Binding with DNA using Surface Plasmon Resonance Biosensor

    Institute of Scientific and Technical Information of China (English)

    Lei WU; Ming-Hui HUANG; Jian-Long ZHAO; Meng-Su YANG

    2005-01-01

    Surface plasmon resonance biosensor technique was used to study the binding of Moloney murine leukemia virus reverse transcriptase without RNase H domain (MMLV RT-) with DNA in the absence and in the presence of inhibitors. Different DNA substrates, including single-stranded DNA (ssDNA),DNA template-primer (T-P) duplex and gapped DNA, were immobilized on the biosensor chip surface using streptavidin-biotin, and MMLV RT--DNA binding kinetics were analyzed by different models. MMLV RT-could bind with ssDNA and the binding was involved in conformation change. MMLV RT- binding DNA T-P duplex and gapped DNA could be analyzed using the simple 1:1 Langmuir model. The lack of RNase H domain reduced the affinity between MMLV RT- and T-P duplex. The effects of RT inhibitors, including efavirenz, nevirapine and quercetin, on the interaction between MMLV RT- and gapped DNA were analyzed according to recovered kinetics parameters. Efavirenz slightly interfered with the binding between RT and DNA and the affinity constant in the presence of the inhibitor (KA=1.21× 106 M-1) was lower than in the absence of the inhibitor (KA=4.61× 106 M-1). Nevirapine induced relatively tight binding between RT and DNA and the affinity constant in the presence of the inhibsitor (KA=l.47×107 M-1) was approximately three folds higher than without nevirapine, mainly due to rapid association and slow dissociation. Quercetin, a flavonoid originating from plant which has previously shown strong inhibition of the activity of RT, was found to have minimal effect on the RT-DNA binding.

  10. Evaluation of effects of bivalent cations on the formation of purine-rich triple-helix DNA by ESI-FT-MS.

    Science.gov (United States)

    Wan, Cuihong; Cui, Meng; Song, Fengrui; Liu, Zhiqiang; Liu, Shuying

    2009-07-01

    The GGA triplet repeats are widely dispersed throughout eukaryotic genomes. (GGA)n or (GGT)n oligonucleotides can interact with double-stranded DNA containing (GGA:CCT)n to form triple-stranded DNA. The effects of 8 divalent metal ions (3 alkaline-earth metals and 5 transition metals) on formation of these purine-rich triple-helix DNA were investigated by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FT-MS). In the absence of metal ions, no triplex but single-strand, duplex, and purine homodimer ions were observed in mass spectra. The triple-helix DNA complexes were observed only in the presence of certain divalent ions. The effects of different divalent cations on the formation of purine-rich triplexes were compared. Transition-metal ions, especially Co(2+) and Ni(2+), significantly boost the formation of triple-helix DNA, whereas alkaline-earth metal ions have no positive effects on triplex formation. In addition, Ba(2+) is notably beneficial to the formation of homodimer instead of triplex.

  11. Duplex ultrasound for identifying renal artery stenosis

    DEFF Research Database (Denmark)

    Zachrisson, Karin; Herlitz, Hans; Lönn, Lars

    2017-01-01

    Background Renal artery duplex ultrasound (RADUS) is an established method for diagnosis of renal artery stenosis (RAS), but there is no consensus regarding optimal RADUS criteria. Purpose To define optimal cutoff values for RADUS parameters when screening for RAS using intra-arterial trans...

  12. Ultra-short silicon MMI duplexer

    Science.gov (United States)

    Yi, Huaxiang; Huang, Yawen; Wang, Xingjun; Zhou, Zhiping

    2012-11-01

    The fiber-to-the-home (FTTH) systems are growing fast these days, where two different wavelengths are used for upstream and downstream traffic, typically 1310nm and 1490nm. The duplexers are the key elements to separate these wavelengths into different path in central offices (CO) and optical network unit (ONU) in passive optical network (PON). Multimode interference (MMI) has some benefits to be a duplexer including large fabrication tolerance, low-temperature dependence, and low-polarization dependence, but its size is too large to integrate in conventional case. Based on the silicon photonics platform, ultra-short silicon MMI duplexer was demonstrated to separate the 1310nm and 1490nm lights. By studying the theory of self-image phenomena in MMI, the first order images are adopted in order to keep the device short. A cascaded MMI structure was investigated to implement the wavelength splitting, where both the light of 1310nm and 1490nm was input from the same port, and the 1490nm light was coupling cross the first MMI and output at the cross-port in the device while the 1310nm light was coupling through the first and second MMI and output at the bar-port in the device. The experiment was carried on with the SOI wafer of 340nm top silicon. The cascaded MMI was investigated to fold the length of the duplexer as short as 117μm with the extinct ratio over 10dB.

  13. Effects of Captan on DNA and DNA metabolic processes in human diploid fibroblasts.

    Science.gov (United States)

    Snyder, R D

    1992-01-01

    The fungicide Captan has been examined for its effects on DNA and DNA processing in order to better understand the genotoxicity associated with this agent. Captan treatment resulted in production of DNA single strand breaks and DNA-protein cross-links and elicited an excision repair response in human diploid fibroblasts. Captan was also shown to inhibit cellular DNA synthesis and to form stable adducts in herring sperm and human cellular DNA. Misincorporation of nucleotides into Captan-treated synthetic DNA templates was significantly elevated in an in vitro assay using E. coli DNA polymerase I, suggesting that DNA adduct formation by Captan could have mutagenic consequences. In sum, these studies demonstrate that Captan is capable of interacting with DNA at a number of levels and that these interactions could provide the basis for Captan's genotoxicity. The extreme cytotoxicity of this fungicide, however, could be due to other cellular effects since at the IC50 for cell killing, approximately 0.8 microM, none of the above genotoxic events could be detected by the methods employed.

  14. On the Secrecy Degrees of Freedom with Full Duplex Communication

    DEFF Research Database (Denmark)

    Mahmood, Nurul Huda; Mogensen, Preben Elgaard

    2017-01-01

    and the delay reduction of full duplex communication are somewhat limited in realistic network settings, leading researchers to study other possible applications of full duplex communication which can provide significantly higher gains over half duplex communication. Physical layer security is an example...... of such an application. The potential of full duplex nodes in improving the physical layer security of a communication link is investigated in this contribution. We specifically derive the information theoretic secrecy degrees of freedom measure for a pair of nodes communicating in full duplex mode. Moreover, closed...

  15. On the Ergodic Secrecy Capacity with Full Duplex Communication

    DEFF Research Database (Denmark)

    Mahmood, Nurul Huda; Shafique Ansari, Imran; Mogensen, Preben Elgaard

    2017-01-01

    Full duplex communication promises performance gains in terms of the throughput and the delay. Generally, it has been shown that the throughput and delay gains of full duplex communication are somewhat limited in realistic network settings, leading researchers to study other possible applications...... capacity for a pair of nodes with full duplex communication. The ergodic secrecy rate with full duplex communication is found to grow linearly with the log of the direct channel SNR as opposed to the flattened out secrecy rate with conventional half duplex communication, irrespective of the eavesdropper...

  16. Structural insight into DNA-assembled oligochromophores: crystallographic analysis of pyrene- and phenanthrene-modified DNA in complex with BpuJI endonuclease.

    Science.gov (United States)

    Probst, Markus; Aeschimann, Walter; Chau, Thi T H; Langenegger, Simon M; Stocker, Achim; Häner, Robert

    2016-09-06

    The use of the DNA duplex as a supramolecular scaffold is an established approach for the assembly of chromophore aggregates. In the absence of detailed structural insight, the characterization of thus assembled oligochromophores is, today, largely based on solution-phase spectroscopy. Here, we describe the crystal structures of three DNA-organized chromophore aggregates. DNA hybrids containing non-nucleosidic pyrene and phenanthrene building blocks were co-crystallized with the recently described binding domain of the restriction enzyme BpuJI. Crystal structures of these complexes were determined at 2.7, 1.9 and 1.6 Å resolutions. The structures reveal aromatic stacking interactions between pyrene and/or phenanthrene units within the framework of the B-DNA duplex. In hybrids containing a single modification in each DNA strand near the end of the duplex, the two polyaromatic hydrocarbons are engaged in a face-to-face stacking orientation. Due to crystal packing and steric effects, the terminal GC base pair is disrupted in all three crystal structures, which results in a non-perfect stacking arrangement of the aromatic chromophores in two of the structures. In a hybrid containing a total of three pyrenes, crystal lattice induced end-to-end stacking of individual DNA duplexes leads to the formation of an extended aromatic π-stack containing four co-axially arranged pyrenes. The aromatic planes of the stacked pyrenes are oriented in a parallel way. The study demonstrates the value of co-crystallization of chemically modified DNA with the recombinant binding domain of the restriction enzyme BpuJI for obtaining detailed structural insight into DNA-assembled oligochromophores.

  17. Effect of DNA type on response of DNA biosensor for carcinogens

    Science.gov (United States)

    Sani, Nor Diyana bt. Md.; Heng, Lee Yook; Surif, Salmijah; Lazim, Azwani Mat

    2013-11-01

    Carcinogens are cancer causing chemicals that can bind to DNA and cause damage to the DNA. These chemicals are available everywhere including in water, air, soil and food. Therefore, a sensor that can detect the presence of these chemicals will be a very useful tool. Since carcinogens bind to DNA, DNA can be used as the biological element in a biosensor. This study has utilized different types of DNA in a biosensor for carcinogen detection. The DNAs include double stranded calf thymus DNA, single stranded calf thymus DNA and guanine rich single stranded DNA. The modified SPE was exposed to a carcinogen followed by interaction with methylene blue which acts as the electroactive indicator. The SPE was then analysed using differential pulse voltammetry (DPV). Optimization studies were conducted for MB concentration and accumulation time, DNA concentration, as well as effect of buffer concentration, buffer pH and ionic strength. The performance of the biosensor was tested on a group 1 carcinogen, formaldehyde. The results indicated that the usage of guanine rich single stranded DNA also gives higher response as carcinogens prefer to bind with guanine compared to other bases.

  18. The Terahertz Controlled Duplex Isolator: Physical Grounds and Numerical Experiment

    Directory of Open Access Journals (Sweden)

    Konstantin Vytovtov

    2016-01-01

    Full Text Available Electromagnetic properties of an anisotropic stratified slab with an arbitrary orientation of the anisotropy axis under an oblique incidence of a plane harmonic wave are studied. The dependence of the eigenwave wavenumbers and the reflection coefficient on an anisotropy axis orientation and frequency is investigated. For the first time, the expression for the translation matrix is obtained in the compact analytical form. The controlled two-way dual-frequency (duplex isolator based on the above described slab is presented for the first time. It is based on the properties of the anisotropic structure described here but not on the Faraday effect.

  19. Effect of DNA methylation on protein-DNA interaction of HL-60 cells

    Institute of Scientific and Technical Information of China (English)

    何忠效; 白坚石; 张昱

    1999-01-01

    HL-60 cells have been induced with differentiation index 16 % by S-adenosyl-L-rnethionine (SAM) as inducer in the presence of optimum conceptration of 10 μmol/L. The methylation level of genorne DNA determined by HPLC is increased during cell differentiation. When restriction endonuclease Hae Ⅲ, Sma I, Sal I, XhoI and Hind Ⅲ which are sensitive to 5-methylcytosine were used to cleave the genorne DNA, a resistance effect was found. The interaction between DNA and DNA binding proteins is changed by using gel retarding test.

  20. Numerical modeling and optimization of machining duplex stainless steels

    Directory of Open Access Journals (Sweden)

    Rastee D. Koyee

    2015-01-01

    Full Text Available The shortcomings of the machining analytical and empirical models in combination with the industry demands have to be fulfilled. A three-dimensional finite element modeling (FEM introduces an attractive alternative to bridge the gap between pure empirical and fundamental scientific quantities, and fulfill the industry needs. However, the challenging aspects which hinder the successful adoption of FEM in the machining sector of manufacturing industry have to be solved first. One of the greatest challenges is the identification of the correct set of machining simulation input parameters. This study presents a new methodology to inversely calculate the input parameters when simulating the machining of standard duplex EN 1.4462 and super duplex EN 1.4410 stainless steels. JMatPro software is first used to model elastic–viscoplastic and physical work material behavior. In order to effectively obtain an optimum set of inversely identified friction coefficients, thermal contact conductance, Cockcroft–Latham critical damage value, percentage reduction in flow stress, and Taylor–Quinney coefficient, Taguchi-VIKOR coupled with Firefly Algorithm Neural Network System is applied. The optimization procedure effectively minimizes the overall differences between the experimentally measured performances such as cutting forces, tool nose temperature and chip thickness, and the numerically obtained ones at any specified cutting condition. The optimum set of input parameter is verified and used for the next step of 3D-FEM application. In the next stage of the study, design of experiments, numerical simulations, and fuzzy rule modeling approaches are employed to optimize types of chip breaker, insert shapes, process conditions, cutting parameters, and tool orientation angles based on many important performances. Through this study, not only a new methodology in defining the optimal set of controllable parameters for turning simulations is introduced, but also

  1. Effects of humic substances on fluorometric DNA quantification and DNA hybridization

    NARCIS (Netherlands)

    Bachoon, DS; Otero, E; Hodson, RE

    2001-01-01

    DNA extracts from sediment and water samples are often contaminated with coextracted humic-like impurities, Estuarine humic substances and vascular plant extract were used to evaluate the effect of the presence of such impurities on DNA hybridization and quantification. The presence of humic

  2. Easily denaturing nucleic acids derived from intercalating nucleic acids: thermal stability studies, dual duplex invasion and inhibition of transcription start

    DEFF Research Database (Denmark)

    Filichev, Vyacheslav V; Vester, Birte; Hansen, Lykke Haastrup;

    2005-01-01

    The bulged insertions of (R)-1-O-(pyren-1-ylmethyl)glycerol (monomer P) in two complementary 8mer DNA strands (intercalating nucleic acids) opposite to each other resulted in the formation of an easily denaturing duplex, which had lower thermal stability (21.0 degrees C) than the wild-type double...

  3. Macromolecular crowding increases binding of DNA polymerase to DNA: an adaptive effect

    Energy Technology Data Exchange (ETDEWEB)

    Zimmerman, S.B.; Harrison, B.

    1987-05-01

    Macromolecular crowding extends the range of ionic conditions supporting high DNA polymerase reaction rates. Reactions tested were nick translation and gap-filling by DNA polymerase I of E. coli, nuclease and polymerase activities of the large fragment of that polymerase, and polymerization by the T4 DNA polymerase. For all of these reactions, high concentrations of nonspecific polymers increased enzymatic activity under otherwise inhibitory conditions resulting from relatively high ionic strength. The primary mechanism of the polymer effect seems to be to increase the binding of polymerase to DNA. They suggest that this effect of protein-DNA complexes is only one example of a general metabolic buffering action of crowded solutions on a variety of macromolecular interactions.

  4. In-Band Full-Duplex Communications for Cellular Networks with Partial Uplink/Downlink Overlap

    KAUST Repository

    AlAmmouri, Ahmad

    2015-12-06

    In-band full-duplex (FD) communications have been optimistically promoted to improve the spectrum utilization in cellular networks. However, the explicit impact of spatial interference, imposed by FD communications, on uplink and downlink transmissions has been overlooked in the literature. This paper presents an extensive study of the explicit effect of FD communications on the uplink and downlink performances. For the sake of rigorous analysis, we develop a tractable framework based on stochastic geometry toolset. The developed model accounts for uplink truncated channel inversion power control in FD cellular networks. The study shows that FD communications improve the downlink throughput at the expense of significant degradation in the uplink throughput. Therefore, we propose a novel fine-grained duplexing scheme, denoted as α-duplex scheme, which allows a partial overlap between uplink and downlink frequency bands. To this end, we show that the amount of the overlap can be optimized via adjusting α to achieve a certain design objective.

  5. A duplex polymerase chain reaction assay for the identification of goat cashmere and sheep wool.

    Science.gov (United States)

    Geng, Qing-Rong

    2016-05-01

    In this article attempts were made to establish one-step duplex PCR assay for the identification of goat cashmere and sheep wool. Primers were selected from published papers or designed in the well-conserved region of mitochondrial D-loop genes after alignment of the available sequences in the GenBank database. A fragment of 294 bp from cashmere goat was amplified and three PCR fragments including a bright main band of approximately 404 bp in length were obtained from sheep. The duplex PCR was found to be effective in detecting mixed samples precisely when sheep wool was mixed to goat cashmere with the relative proportion of over 9.09%. The duplex PCR could be considered as a simple and promising method in identification of goat cashmere and sheep wool.

  6. Studies of the B-Z transition of DNA: The temperature dependence of the free-energy difference, the composition of the counterion sheath in mixed salt, and the preparation of a sample of the 5'-d[T-(m(5) C-G)12 -T] duplex in pure B-DNA or Z-DNA form.

    Science.gov (United States)

    Guéron, Maurice; Plateau, Pierre; Filoche, Marcel

    2016-07-01

    It is often envisioned that cations might coordinate at specific sites of nucleic acids and play an important structural role, for instance in the transition between B-DNA and Z-DNA. However, nucleic acid models explicitly devoid of specific sites may also exhibit features previously considered as evidence for specific binding. Such is the case of the "composite cylinder" (or CC) model which spreads out localized features of DNA structure and charge by cylindrical averaging, while sustaining the main difference between the B and Z structures, namely the better immersion of the B-DNA phosphodiester charges in the solution. Here, we analyze the non-electrostatic component of the free-energy difference between B-DNA and Z-DNA. We also compute the composition of the counterion sheath in a wide range of mixed-salt solutions and of temperatures: in contrast with the large difference of composition between the B-DNA and Z-DNA forms, the temperature dependence of sheath composition, previously unknown, is very weak. In order to validate the model, the mixed-salt predictions should be compared to experiment. We design a procedure for future measurements of the sheath composition based on Anomalous Small-Angle X-ray Scattering and complemented by (31) P NMR. With due consideration for the kinetics of the B-Z transition and for the capacity of generating at will the B or Z form in a single sample, the 5'-d[T-(m(5) C-G)12 -T] 26-mer emerges as a most suitable oligonucleotide for this study. Finally, the application of the finite element method to the resolution of the Poisson-Boltzmann equation is described in detail. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 369-384, 2016.

  7. Protection of aluminium by duplex coatings

    Energy Technology Data Exchange (ETDEWEB)

    Musil, J. [Ceska Akademie Ved, Prague (Czech Republic). Fyzikalni Ustav; Vlcek, J. [West Bohemia Univ., Plzen (Czech Republic). Dept. of Phys.; Jezek, V. [West Bohemia Univ., Plzen (Czech Republic). Dept. of Phys.; Benda, M. [West Bohemia Univ., Plzen (Czech Republic). Dept. of Phys.

    1995-11-01

    The paper reports on a new way of producing duplex coatings consisting of two steps. First, the substrate is coated by a physically vapour-deposited coating. Then, this precoated substrate is plasma nitrided or vacuum heat treated. This method was tested in the protection of substrates made of aluminium with a sputtered Ti coating about 5 {mu}m thick. The as-deposited and then plasma-nitrided or vacuum-heat-treated (Ti coating)/(Al substrate) couple was characterized by elemental depth profiles measured by glow discharge optical spectroscopy. It was shown that both the plasma nitriding and vacuum heat treatment process can stimulate a strong interdiffusion between Ti and the substrate elements. It results not only in the formation of a very broad interfacial region with a dramatic redistribution of the substrate elements in the Ti film but also in a formation of intermetallic Ti-Al compounds. This new duplex coating technique is described in detail. (orig.)

  8. Renal duplex Doppler ultrasound findings in diabetics

    Energy Technology Data Exchange (ETDEWEB)

    Shim, Hyang Yee; Kim, Young Geun; Kook, Cheol Keu; Yoon, Chong Hyun; Lee, Shin Hyung; Lee, Chang Joon [National Medical Center, Seoul (Korea, Republic of)

    1993-12-15

    The correlation between clinical-laboratory findings and renal duplex Doppler ultrasound findings was studied in 45 patients with diabetes mellitus to see the role of duplex Doppler ultrasound in the detection of diabetic nephropathy. The resistive indices in patients with elevated serum creatinine, BUN, proteinuria, and systolic blood pressure levels were statistically significantly higher than those in patients with normal levels (p<0.05). Also resistive indics in patients with retinopathy were higher than that in patients without retinopathy (p<0.05). But the ultrasound morphologic changes of kidney such as renal length, cortical eye-catching, and corticomedullarycontrast were not well correlated with clinical-laboratory data and resistive index. The resistive index of the kidney in conjunction with clinical-laboratory data in diabetics may be helpful in the evaluation of diabetic nephropathy

  9. Effect of the 3-halo substitution of the 2'-deoxy aminopyridinyl-pseudocytidine derivatives on the selectivity and stability of antiparallel triplex DNA with a CG inversion site.

    Science.gov (United States)

    Wang, Lei; Taniguchi, Yosuke; Okamura, Hidenori; Sasaki, Shigeki

    2017-07-15

    Triplex formation against a target duplex DNA has the potential to become a tool for the genome research. However, there is an intrinsic restriction on the duplex DNA sequences capable of forming the triplex DNA. Recently, we demonstrated the selective formation of the stable antiparallel triplexes containing the CG inversion sites using the 2'-deoxy-1-methylpseudocytidine derivative (ΨdC), whose amino group was conjugated with the 2-aminopyridine at its 5-position as an additional hydrogen bonding unit (AP-ΨdC). The 1-N of 2-aminopyridine was supposed to be protonated to form the hydrogen bond with the guanine of the CG inversion site. In this study, to test the effect of the 3-substitution of the 2-aminopyridine unit of AP-ΨdC on the triplex stability, we synthesized the 3-halogenated 2-aminopyridine derivatives of AP-ΨdC. The pKa values 1-N of the 2-aminopyridine unit of AP-ΨdC as the monomer nucleoside were determined to be 6.3 for 3-CH3 ((Me)AP-ΨdC), 6.1 for 3-H (AP-ΨdC), 4.3 for 3-Cl ((Cl)AP-ΨdC), 4.4 for 3-Br ((Br)AP-ΨdC), and 4.7 for 3-I ((I)AP-ΨdC), suggesting that all the halogenated AP-ΨdCs are not protonated under neutral conditions. Interestingly, although the recognition selectivity depends on the sequence context, the TFO having the sequence of the 3'-G-((I)AP-ΨdC)-A-5' context showed the selective triplex formation with the CG inversion site. These results suggest that the protonation at the 1-N position plays an important role in the stable and selective triplex formation of AP-ΨdC derivatives in any sequences. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Problems in repair-welding of duplex-treated tool steels

    Directory of Open Access Journals (Sweden)

    T. Muhič

    2009-01-01

    Full Text Available The present paper addresses problems in laser welding of die-cast tools used for aluminum pressure die-castings and plastic moulds. To extend life cycle of tools various surface improvements are used. These surface improvements significantly reduce weldability of the material. This paper presents development of defects in repair welding of duplex-treated tool steel. The procedure is aimed at reduction of defects by the newly developed repair laser welding techniques. Effects of different repair welding process parameters and techniques are considered. A microstructural analysis is conducted to detect defect formation and reveal the best laser welding method for duplex-treated tools.

  11. Humidity Effects on Conductivity of DNA Molecules

    Institute of Scientific and Technical Information of China (English)

    YAN Xun-Ling; DONG Rui-Xin; LIN Qing-De

    2006-01-01

    We present a model related to the humidity to describe the conductivity of homogeneous DNA molecule,where the hydration of phosphate group and bases are taken into account. The calculated results show the oscillation feature of dⅠ/dⅤ-Ⅴ curves and the semiconductor behavior of DNA. With the relative humidity increasing, the voltage gap becomes narrow and the maximum of conductance increases nonlinearly. The conductivity of DNA approaches to stabilization when the relative humidity reaches a certain value. These results are in agreement with experimental measurements.

  12. Duplex Stainless Steels-An overview

    Directory of Open Access Journals (Sweden)

    Dr. Sunil D.Kahar

    2017-04-01

    Full Text Available Stainless steel is one of the most important materials in the engineering world. The material‟s wide applications in chemical, petrochemical, off-shore, and power generation plants prove that it is one of the most reliable materials. The Newest fast growing family of stainless steels is duplex alloys. The ferritic-austenitic grades have a ferrite matrix intermix with austenite and in other words island of austenite in a continuous matrix of highly alloyed ferrite commonly called „Duplex‟ stainless steel. Duplex stainless steel covers ferritic/austenitic Fe-Cr-Ni alloy with between 30% to 70 % Ferrite .Due to high level of Cr, Mo, and N steels shows high pitting & stress corrosion cracking resistance in chloride-containing environments. Hence it is frequently used in oilrefinery heat exchangers & typical applications where there is a risk for SCC and localized corrosion as a result of chloride-containing process streams, cooling waters or deposits. Modern duplex stainless steels have generally good Weldability. Due to a balanced composition, where nitrogen plays an important role, austenite formation in the heat affected zone (HAZ and weld metal is rapid. Under normal welding conditions a sufficient amount of austenite is formed to maintain good resistance to localized corrosion where as too rapid cooling may result in excessive amounts of ferrite, reducing the toughness. Therefore, welding with low heat input in thick walled materials should be avoided. Welding methods, such as resistance welding, laser welding and electron beam welding, which cause extremely rapid cooling should also be avoided or used with extreme caution. Too slow cooling can in the higher alloyed duplex grades cause formation of inter-metallic phases detrimental to corrosion resistance and toughness.

  13. The effect of DNA supercoiling on nucleosome structure and stability.

    Science.gov (United States)

    Elbel, Tabea; Langowski, Jörg

    2015-02-18

    Nucleosomes have to open to allow access to DNA in transcription, replication, and DNA damage repair. Changes in DNA torsional strain (e.g. during transcription elongation) influence the accessibility of nucleosomal DNA. Here we investigated the effect of DNA supercoiling-induced torsional strain on nucleosome structure and stability by scanning force microscopy (SFM) and fluorescence correlation spectroscopy (FCS). Nucleosomes were reconstituted onto 2.7 kb DNA plasmids with varying superhelical densities. The SFM results show a clear dependence of the amount of DNA wrapped around the nucleosome core on the strength and type of supercoiling. Negative supercoiling led to smaller nucleosome opening angles as compared to relaxed or positively supercoiled DNA. FCS experiments show that nucleosomes reconstituted on negatively superhelical DNA are more resistant to salt-induced destabilization, as seen by reduced H2A-H2B dimer eviction from the nucleosome. Our results show that changes in DNA topology, e.g. during transcription elongation, affect the accessibility of nucleosomal DNA.

  14. Full duplex communication using visible light

    CERN Document Server

    Yang, Yongchao; Li, Yuanhang; Gao, Xumin; Yuan, Jialei; Zhu, Hongbo; Wang, Yongjin

    2016-01-01

    In this work, we propose, fabricate and characterize a full duplex communication system using visible light on a single chip. Both the suspended p-n junction InGaN/GaN multiple quantum well (MQW) devices and the suspended waveguides are obtained on a GaN-on-silicon platform by wafer-level processing. Two suspended p-n junction InGaN/GaN MQW devices that can both emit and detect light simultaneously are connected using suspended waveguides to form an in-plane visible light communication (VLC) system. The light that is emitted from one suspended p-n junction InGaN/GaN MQW device can induce a current in the device located at the other end of the waveguide via in-plane light coupling, thus leading to full duplex communication using visible light. This proof-of-concept in-plane VLC system paves the way towards the implementation of a full duplex communications system operating at the same frequency using visible light on a single chip.

  15. How effective is graphene nanopore geometry on DNA sequencing?

    CERN Document Server

    Satarifard, Vahid; Ejtehadi, Mohammad Reza

    2015-01-01

    In this paper we investigate the effects of graphene nanopore geometry on homopolymer ssDNA pulling process through nanopore using steered molecular dynamic (SMD) simulations. Different graphene nanopores are examined including axially symmetric and asymmetric monolayer graphene nanopores as well as five layer graphene polyhedral crystals (GPC). The pulling force profile, moving fashion of ssDNA, work done in irreversible DNA pulling and orientations of DNA bases near the nanopore are assessed. Simulation results demonstrate the strong effect of the pore shape as well as geometrical symmetry on free energy barrier, orientations and dynamic of DNA translocation through graphene nanopore. Our study proposes that the symmetric circular geometry of monolayer graphene nanopore with high pulling velocity can be used for DNA sequencing.

  16. Role of minor groove width and hydration pattern on amsacrine interaction with DNA.

    Directory of Open Access Journals (Sweden)

    Deepak K Jangir

    Full Text Available Amsacrine is an anilinoacridine derivative anticancer drug, used to treat a wide variety of malignancies. In cells, amsacrine poisons topoisomerase 2 by stabilizing DNA-drug-enzyme ternary complex. Presence of amsacrine increases the steady-state concentration of these ternary complexes which in turn hampers DNA replication and results in subsequent cell death. Due to reversible binding and rapid slip-out of amsacrine from DNA duplex, structural data is not available on amsacrine-DNA complexes. In the present work, we designed five oligonucleotide duplexes, differing in their minor groove widths and hydration pattern, and examined their binding with amsacrine using attenuated total reflection Fourier transform infrared (ATR-FTIR spectroscopy. Complexes of amsacrine with calf thymus DNA were also evaluated for a comparison. Our results demonstrate for the first time that amsacrine is not a simple intercalator; rather mixed type of DNA binding (intercalation and minor groove takes place between amsacrine and DNA. Further, this binding is highly sensitive towards the geometries and hydration patterns of different minor grooves present in the DNA. This study shows that ligand binding to DNA could be very sensitive to DNA base composition and DNA groove structures. Results demonstrated here could have implication for understanding cytotoxic mechanism of aminoacridine based anticancer drugs and provide directions to modify these drugs for better efficacy and few side-effects.

  17. Structurally Diverse Polyamines: Solid-Phase Synthesis and Interaction with DNA.

    Science.gov (United States)

    Umezawa, Naoki; Horai, Yuhei; Imamura, Yuki; Kawakubo, Makoto; Nakahira, Mariko; Kato, Nobuki; Muramatsu, Akira; Yoshikawa, Yuko; Yoshikawa, Kenichi; Higuchi, Tsunehiko

    2015-08-17

    A versatile solid-phase approach based on peptide chemistry was used to construct four classes of structurally diverse polyamines with modified backbones: linear, partially constrained, branched, and cyclic. Their effects on DNA duplex stability and structure were examined. The polyamines showed distinct activities, thus highlighting the importance of polyamine backbone structure. Interestingly, the rank order of polyamine ability for DNA compaction was different to that for their effects on circular dichroism and melting temperature, thus indicating that these polyamines have distinct effects on secondary and higher-order structures of DNA.

  18. Enhanced immunostimulatory effects of DNA-encapsulated peptide hydrogels.

    Science.gov (United States)

    Medina, Scott H; Li, Sandra; Howard, O M Zack; Dunlap, Micah; Trivett, Anna; Schneider, Joel P; Oppenheim, Joost J

    2015-01-01

    DNA that encodes tumor-specific antigens represents potential immunostimulatory agents. However, rapid enzymatic degradation and fragmentation of DNA during administration can result in limited vector expression and, consequently, poor efficacy. These challenges have necessitated the use of novel strategies for DNA delivery. Herein, we study the ability of cationic self-assembling peptide hydrogels to encapsulate plasmid DNA, and enhance its immunostimulatory potential in vivo. The effect of network charge on the gel's ability to retain the DNA was assessed employing three gel-forming peptides that vary systematically in formal charge. The peptide HLT2, having a formal charge of +5 at neutral pH, was optimal in encapsulating microgram quantities of DNA with little effect on its rheological properties, allowing its effective syringe delivery in vivo. The plasmid, DNA(TA), encapsulated within these gels encodes for a melanoma-specific gp100 antigen fused to the alarmin protein adjuvant HMGN1. Implantation of DNA(TA)-loaded HLT2 gels into mice resulted in an acute inflammatory response with the presence of polymorphonuclear cells, which was followed by infiltrating macrophages. These cellular infiltrates aid in the processing of encapsulated DNA, promoting increased lymphoproliferation and producing an enhanced immune response mediated by CD4+/IFNγ+ expressing Th1 cells, and complemented by the formation of gp100-specific antibodies.

  19. The Quadraplex Tetraploids Hybrids and Duplex Tetraploids Hybrids Are Responsible for Heterosis and Inbreeding Depression in Maize

    Institute of Scientific and Technical Information of China (English)

    WANG Ze-li; James A Birchler; WANG Lu-xin; JIN Liang

    2005-01-01

    The maize quadraplex tetraploids and duplex tetraploids were developed using Kato's protocols. The phenotype of heterosis and inbreeding depression over generations in their parents and progenies of F1, F2 and F3 were investigated.The results indicated that different duplex tetraploids have different genetic backgrounds, but they acquire maximum heterosis at same traits, such as the leaf length, leaf width, culm circumference and days to flowering. P.N. rises much faster from the F2 to F3 segment than the A.W. does for the plant height in duplex tetraploids. In comparing duplex and quadraplex over a generation the quadriplex is showing the greatest heterosis in plant height, leaf height, leaf width and days to flowering. Most of the examples achieve the maximum heterosis at Qu F2, with the exception of culm circumference,which achieves greatest heterosis at PNAW F1. Meanwhile, this experiment shows that quadraplex tetraploids has distinct additional favorable alleles that are not contained in duplex tetraploid, this is demonstrated by the heterosis found in crosses between the two duplex tetraploid. This finding helps explain quadraploids superiority and unique breeding behavior, in which, the progressive heterosis and inbreeding depression in maize are due mainly to linkage disequilibrium.The severe inbreeding depression in duplex tetraploids is due mainly to the rapid loss of complementary chromosomes or genes interactions in the first few generation of inbreeding. Correspondingly, the progressive heterosis in quadraplex tetraploids is due mainly to a progressive increase in complementarities of homologous chromosomes or genes interactions.Greater complementarities of homologous chromosomes or genes interactions in tetraploids maize alse helps explain recent molecular biology research indicating that some of traits in quadraplex tetraploids are more responsive to genetic diversity than in duplex tetraploids. In addition, the dosage effect of polyploid in relation

  20. Thermal Aging Phenomena in Cast Duplex Stainless Steels

    Energy Technology Data Exchange (ETDEWEB)

    Byun, T. S.; Yang, Y.; Overman, N. R.; Busby, J. T.

    2016-02-28

    Cast stainless steels (CASSs) have been extensively used for the large components of light water reactor (LWR) power plants such as primary coolant piping and pump casing. The thermal embrittlement of CASS components is one of the most serious concerns related to the extended-term operation of nuclear power plants. Many past researches have concluded that the formation of Cr–rich α'-phase by Spinodal decomposition of δ-ferrite phase is the primary mechanism for the thermal embrittlement. Cracking mechanism in the thermally-embrittled duplex stainless steels consists of the formation of cleavage at ferrite and its propagation via separation of ferrite-austenite interphase. This article intends to provide an introductory overview on the thermal aging phenomena in LWR relevant conditions. Firstly, the thermal aging effect on toughness is discussed in terms of the cause of embrittlement and influential parameters. An approximate analysis of thermal reaction using Arrhenius equation was carried out to scope the aging temperatures for the accelerated aging experiments to simulate the 60 and 80 years of services. Further, equilibrium precipitation calculation was performed for model CASS alloys using the CALPHAD program and the results are used to describe the precipitation behaviors in duplex stainless steels. These results are also to be used to guide an on-going research aiming to provide knowledge-based conclusive prediction for the integrity of the CASS components of LWR power plants during the service life extended up to and beyond 60 years.

  1. Molecular crowding effects on stability of DNA double helix

    Science.gov (United States)

    Singh, Amar; Singh, Navin

    2016-05-01

    Cellular environmental conditions critically affect the structure and stability of double stranded DNA (dsDNA) molecule. It is known that 20-30% of the total volume of the cell is occupied by the molecular crowders. The presence of these crowders, reduces the free space available to the base pairs of a DNA molecule, hence the movement of base pair is restricted. Here, we study the thermal opening of dsDNA molecule using Peyrard Bishop Dauxois (PBD) model. The presence of crowders in the model, that mimic those found in the cell nucleus, is realized through the potential term. Using the equilibrium statistical calculations, we find melting profile and melting probabilities of the chain. The opening of DNA molecule in the presence of these crowders is shown through the density plots. This study reveals that the stability of dsDNA molecule is influenced by entropic as well as enthalpic effects and is more stable in the crowded environment.

  2. Effect of knots on binding of intercalators to DNA

    Science.gov (United States)

    Medalion, Shlomi; Rabin, Yitzhak

    2014-05-01

    We study the effect of knots in circular dsDNA molecules on the binding of intercalating ligands. Using Monte Carlo simulations we show that depending on their handedness, the presence of knots can either suppress or enhance intercalation in supercoiled DNA. When the occupancy of intercalators on DNA is low, the effect of knots on intercalation can be captured by introducing a shift in the mean writhe of the chain that accounts for the writhe of the corresponding ideal knot. In the limit of high intercalator occupancy, the writhe distribution of different knots is strongly affected by excluded volume effects and therefore by salt concentration. Based on the finding that different knots yield well-separated probability distributions of bound intercalators, we propose a new experimental approach to determine DNA topology by monitoring the intensity of fluorescence emitted by dye molecules intercalated into knotted DNA molecules.

  3. Effect of DNA Hairpin Loops on the Twist of Planar DNA Origami Tiles

    Science.gov (United States)

    Li, Zhe; Wang, Lei; Yan, Hao; Liu, Yan

    2012-01-01

    The development of scaffolded DNA origami, a technique in which a long single-stranded viral genome is folded into arbitrary shapes by hundreds of short synthetic oligonucleotides, represents an important milestone in DNA nanotechnology. Recent findings have revealed that two-dimensional (2D)DNA origami structures based on the original design parameters adopt a global twist with respect to the tile plane, which may be because the conformation of the constituent DNA (10.67 bp/turn) deviates from the natural B-type helical twist (10.4 bp/turn). Here we aim to characterize the effects of DNA hairpin loops on the overall curvature of the tile and explore their ability to control, and ultimately eliminate any unwanted curvature. A series of dumbbell-shaped DNA loops were selectively displayed on the surface of DNA origami tiles with the expectation that repulsive interactions among the neighboring dumbbell loops and between the loops and the DNA origami tile would influence the structural features of the underlying tiles. A systematic, atomic force microscopy (AFM) study of how the number and position of the DNA loops influenced the global twist of the structure was performed, and several structural models to explain the results were proposed. The observations unambiguously revealed that the first generation of rectangular shaped origami tiles adopt a conformation in which the upper right (corner 2) and bottom left (corner 4) corners bend upward out of the plane, causing linear superstructures attached by these corners to form twisted ribbons. Our experimental observations are consistent with the twist model predicted by the DNA mechanical property simulation software CanDo. Through the systematic design and organization of various numbers of dumbbell loops on both surfaces of the tile, a nearly planar rectangular origami tile was achieved. PMID:22126326

  4. Development of duplex PCR for simultaneous detection of Theileria spp. and Anaplasma spp. in sheep and goats.

    Science.gov (United States)

    Cui, Yanyan; Zhang, Yan; Jian, Fuchun; Zhang, Longxian; Wang, Rongjun; Cao, Shuxuan; Wang, Xiaoxing; Yan, Yaqun; Ning, Changshen

    2017-05-01

    Theileria spp. and Anaplasma spp., which are important tick-borne pathogens (TBPs), impact the health of humans and animals in tropical and subtropical areas. Theileria and Anaplasma co-infections are common in sheep and goats. Following alignment of the relevant DNA sequences, two primer sets were designed to specifically target the Theileria spp. 18S rRNA and Anaplasma spp. 16S rRNA gene sequences. Genomic DNA from the two genera was serially diluted tenfold for testing the sensitivities of detection of the primer sets. The specificities of the primer sets were confirmed when DNA from Anaplasma and Theileria (positive controls), other related hematoparasites (negative controls) and ddH2O were used as templates. Fifty field samples were also used to evaluate the utility of single PCR and duplex PCR assays, and the detection results were compared with those of the PCR methods previously published. An optimized duplex PCR assay was established from the two primer sets based on the relevant genes from the two TBPs, and this assay generated products of 298-bp (Theileria spp.) and 139-bp (Anaplasma spp.). The detection limit of the assay was 29.4 × 10(-3) ng per μl, and there was no cross-reaction with the DNA from other hematoparasites. The results showed that the newly developed duplex PCR assay had an efficiency of detection (P > 0.05) similar to other published PCR methods. In this study, a duplex PCR assay was developed that can simultaneously identify Theileria spp. and Anaplasma spp. in sheep and goats. This duplex PCR is a potentially valuable assay for epidemiological studies of TBPs in that it can detect cases of mixed infections of the pathogens. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Profiling oxidative DNA damage: effects of antioxidants.

    Science.gov (United States)

    Box, Harold C; Patrzyc, Helen B; Budzinski, Edwin E; Dawidzik, Jean B; Freund, Harold G; Zeitouni, Nathalie C; Mahoney, Martin C

    2012-11-01

    The goal of this research was to determine whether antioxidant usage could be correlated with changes in DNA damage levels. Liquid Chromatography-tandem Mass Spectrometry (LC-MS/MS) was used to simultaneously measure five different oxidatively-induced base modifications in the DNA of WBC. Measurements of the five modifications were made before and after an 8-week trial during which participants took the SU.VI.MAX supplement. Levels of the five DNA modifications were compared among different groupings: users versus non-users of antioxidant supplements, before versus after the supplement intervention and men versus women. The statistical significance of differences between groups was most significant for pyrimidine base modifications and the observed trends reflect trends reported in epidemiological studies of antioxidant usage. A combination of modifications derived from pyrimidine bases is suggested as a superior indicator of oxidative stress.

  6. A duplex PCR for rapid and simultaneous detection of Brucella spp. in human blood samples.

    Science.gov (United States)

    Mirnejad, Reza; Mohamadi, Mozafar; Piranfar, Vahbeh; Mortazavi, Seied Mojtaba; Kachuei, Reza

    2013-06-01

    To design a duplex PCR for rapid and simultaneous detection of Brucella species. in human blood samples. Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis. Following DNA extraction, PCR assay were performed, using three primers that could simultaneously identify and differentiate three major species of pathogenic Brucella in humans and animals. Of the 52 peripheral bloods samples tested, 25 sample (48%) showed positive reactions in PCR. Twelve samples were positive for Brucella abortus 39 (B. abortus 39) (23%), 13 for Brucella melitensis 39 (B. melitensis 39) (25%) and 0 for Brucella ovis 39 (B. ovis 39) (0%). This work demonstrates that in case where specific primers were utilized, duplex PCR has proved to be a simple, fast, and relatively inexpensive method for simultaneous detection of important species of Brucella in clinical samples. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  7. A duplex PCR for the rapid and simultaneous detection of Brucella spp. in human blood samples

    Institute of Scientific and Technical Information of China (English)

    Reza Mirnejad; Mozafar mohamadi; Vahbeh Piranfar; Seied Mojtaba Mortazavi; Reza Kachuei

    2013-01-01

    Objective: To design a duplex PCR for rapid and simultaneous detection of Brucella species. in human blood samples. Methods: Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis. Following DNA extraction, PCR assay were performed, using three primers that could simultaneously identify and differentiate three major species of pathogenic Brucella in humans and animals. Results: Of the 52 peripheral bloods samples tested, 25 sample (48%) showed positive reactions in PCR. Twelve samples were positive for Brucella abortus (B. abortus) (23%), 13 for Brucella melitensis (B. melitensis) (25%) and 0 for Brucella ovis (B. ovis) (0%). Conclusions: This work de=monstrates that in case where specific primers were utilized, duplex PCR has proved to be a simple, fast, and relatively inexpensive method for simultaneous detection of important species of Brucella in clinical samples.

  8. Effect of the dnaN mutation on the growth of small DNA phages.

    Science.gov (United States)

    Taketo, A

    1981-01-01

    The effect of the dnaN mutation on the growth of single-stranded DNA phages was studied by burst experiments. In HC138 dnaN cells exposed to 42.5 degrees C at 5 min before infection, growth of spherical (microvirid or isometric) phages such as alpha 3, phi Kh-1 and phi X174 was partially reduced at the nonpermissive temperature. When infection was performed at 30 min after temperature shift-up, viral replication was completely inhibited at 42.5 degrees C in the dnaN strain but not in a dna+ revertant. At 41 degrees C, multiplication of filamentous (inovirid) phages M13 and fd was restricted specifically in HC138 F+ dnaN bacteria. When dnaN cells lysogenic for lambda i21 were grown at 42.5 degrees C for 60 min and then shifted down to 33 degrees C, a burst of lambda i21 occurred with concomitant cellular lysis, manifesting induction of the prophage development.

  9. Effects of incense smoke on human lymphocyte DNA.

    Science.gov (United States)

    Szeto, Yim Tong; Sok Wa Leong, Kosca; Keong Lam, Kason; Min Min Hong, Cynthia; Kai Mui Lee, Daphne; Teng Fun Chan, Yui; Benzie, Iris F F

    2009-01-01

    Incense burning is common in Southeast Asia, where it is a traditional and ceremonial practice in deity worship and paying respect to ancestors. However, incense emissions are an important source of indoor air pollution in Asia, and may induce health problems to those exposed. In this in vitro study the effects of incense emissions on human DNA were investigated using the comet assay. Particulates in smoke from six kinds of incense were trapped in saline or ethanol and human lymphocytes were exposed under controlled conditions. Results showed that DNA damage, including strand breaks, was induced by both aqueous and ethanolic extracts of two samples. The ethanolic extract of one sample induced DNA damage, while no significant DNA damage was found in the remaining three samples. The mechanisms underlying DNA damage induced by incense emissions were also investigated. Catalase (CAT), sodium azide, and superoxide dismutase (SOD) were co-incubated with extract, which exerted significant DNA damaging effects. Results showed that CAT with or without SOD diminished DNA damage, whereas sodium azide did not seem able to reduce DNA damage. Data indicate there are potential adverse health effects of such exposure, particularly for temple workers.

  10. q-Ising model on a duplex and a partially duplex clique

    CERN Document Server

    Chmiel, Anna; Sznajd-Weron, Katarzyna

    2016-01-01

    We analyze a modified kinetic Ising model, so called $q$- neighbor Ising model, with Metropolis dynamics,[Phys. Rev. E {\\bf92} 052105] on a duplex clique and a partially duplex clique. In the $q$-Ising model each spin interacts only with $q$ spins randomly chosen from the whole neighborhood. In the case of a duplex clique the change of a spin is allowed only if both levels simultaneously induce this change. Due to the mean-field like nature of the model we are able to derive the analytic form of transition probabilities and solve the corresponding master equation. The existence of the second level changes dramatically the character of the phase transition. In the case of the monoplex clique, the $q$-neighbor Ising model exhibits continuous phase transition for $q=3$, discontinuous phase transition for $q \\ge 4$ and for $q=1$ and $q=2$ the phase transition is not observed. On the other hand, in the case of the duplex clique continuous phase transitions are observed for all values of $q$, even for $q=1$ and $q=...

  11. Estudo dos efeitos da restrição na microestrutura, microdureza e tenacidade em juntas soldadas em aço inoxidável duplex Study of restriction effects on mMicrostructure, microhardness and toughness in welded joints of duplex stainless steel

    Directory of Open Access Journals (Sweden)

    Everton Barbosa Nunes

    2011-06-01

    Full Text Available Os aços inoxidáveis duplex (AID apresentam em sua microestrutura ferrita e austenita, de modo a aliar boas propriedades mecânicas e resistência à corrosão. Estes materiais possuem grande aplicação na indústria petroquímica, sendo o UNS S31803 um dos mais conhecidos. Existem poucos trabalhos referentes ao estudo da influência da restrição na soldagem da junta na formação de Widmanstätten. Logo, este trabalho tem como objetivo analisar a influência da restrição no balanceamento de fases, na morfologia da austenita, na microdureza e na tenacidade da Zona Afetada pelo Calor (ZAC e da Zona Fundida (ZF. Foram realizadas soldagens em juntas tipo V utilizando eletrodo revestido AWS E2209-17com dois níveis de energia: 15 kJ/cm e 20 kJ/cm, com e sem restrição. De maneira geral, houve maior quantidade de ferrita nos passes de acabamento em relação aos passes de enchimento e de raiz. Foram observadas maiores quantidades de Widmanstätten nas condições soldadas com restrição. A ZAC apresentou uma microestrutura mais grosseira nos passes de acabamento. No entanto, não foram observadas diferenças significativas nas microestruturas quando comparadas as condições com e sem restrição. As condições que apresentaram maior quantidade de Widmanstätten obtiveram menores níveis de microdureza. Não foram observadas diferenças na energia absorvida na ZAC e ZF.Duplex stainless steels present ferrite and austenite in their microstructure in order to join good mechanical properties and corrosion resistance. These materials are very used in petrochemical industry, being the DSS UNS S31803 one of the most commercially known. There were not much published literature to study the influence of the precipitation of Widmanstätten austenite in joins with some restriction. This work has as objective to analyze the influence of the restriction in the phase balance of austenite and ferrite, in the morphology of the austenite, in the

  12. Duplex tube steam reformer development program

    Energy Technology Data Exchange (ETDEWEB)

    Lewe, C K; Nieto, J M; Papadopoulos, A

    1978-09-01

    Work done in partial fulfillment of Task 7 of the Duplex Steam Reformer Development Program is described. The DSR concept acts as a double barrier between a process heat high temperature reactor plant (PNP) and a closed loop chemical heat pipe (CHP) for the long distance transport of chemical energy to a remote industrial user. The current state of the DSR design is described as well as related systems and equipment. The PNP concept presented is based upon work currently underway in the Federal Republic of Germany.

  13. On the Performance of In-Band Full-Duplex Cooperative Communications

    KAUST Repository

    Khafagy, Mohammad Galal

    2016-06-01

    In-band full-duplex, by which radios may simultaneously transmit and receive over the same channel, has been always considered practically-unfeasible due to the prohibitively strong self-interference. Indeed, a freshly-generated transmit signal power is typically ten orders of magnitude higher than that of a naturally-attenuated received signal. While unable to manage such an overwhelming interference, wireless communications resorted to half-duplex operation, transmitting and receiving over orthogonal channel resources. Recent research has demonstrated the practical feasibility of full-duplexing via successive sophisticated stages of signal suppression/cancellation, bringing this long-held assumption down and reviving the promising full-duplex potentials. Full-duplex relaying (FDR), where intermediate nodes may now support source-destination communication via simultaneous listening/forwarding, represents one of two full-duplex settings currently recommended for deployment in future fifth-generation (5G) systems. Theoretically, it has been widely accepted that FDR potentially doubles the channel capacity when compared to its half-duplex counterpart. Although FDR doubles the multiplexing gain, the effective signal-to-noise ratio (SNR) can be significantly degraded due to the residual self-interference (RSI) if not properly handled. In this work, efficient protocols are devised for different FDR settings. Selective cooperation is proposed for the canonical three-terminal FDR channel with RSI, which exploits the cooperative diversity offered by the independently fading source/relay message replicas arriving at the destination. Closed-form expressions are derived for the end-to-end SNR cumulative distribution function (CDF) under Rayleigh and Nakagami-m fading. Further, the offered diversity gain is presented as a function of the RSI scaling trend with the relay power. We show that the existing diversity problem in simple FDR protocols can be considerably fixed via

  14. Effects of Ionic Dependence of DNA Persistence Length on the DNA Condensation at Room Temperature

    Science.gov (United States)

    Mao, Wei; Liu, Yan-Hui; Hu, Lin; Xu, Hou-Qiang

    2016-05-01

    DNA persistence length is a key parameter for quantitative interpretation of the conformational properties of DNA and related to the bending rigidity of DNA. A series of experiments pointed out that, in the DNA condensation process by multivalent cations, the condensed DNA takes elongated coil or compact globule states and the population of the compact globule states increases with an increase in ionic concentration. At the same time, single molecule experiments carried out in solution with multivalent cations (such as spermidine, spermine) indicated that DNA persistence length strongly depends on the ionic concentration. In order to revolve the effects of ionic concentration dependence of persistence length on DNA condensation, a model including the ionic concentration dependence of persistence length and strong correlation of multivalent cation on DNA is provided. The autocorrelation function of the tangent vectors is found as an effective way to detect the ionic concentration dependence of toroidal conformations. With an increase in ion concentration, the first periodic oscillation contained in the autocorrelation function shifts, the number of segment contained in the first periodic oscillation decreases gradually. According to the experiments, the average long-axis length is defined to estimate the ionic concentration dependence of condensation process further. The relation between long-axis length and ionic concentration matches the experimental results qualitatively. Supported by National Natural Science Foundation of China under Grant Nos. 11047022, 11204045, 11464004 and 31360215; The Research Foundation from Ministry of Education of China (212152), Guizhou Provincial Tracking Key Program of Social Development (SY20123089, SZ20113069); The General Financial Grant from the China Postdoctoral Science Foundation (2014M562341); The Research Foundation for Young University Teachers from Guizhou University (201311); The West Light Foundation (2015) and College

  15. Collaborating functions of BLM and DNA topoisomerase I in regulating human rDNA transcription

    Energy Technology Data Exchange (ETDEWEB)

    Grierson, Patrick M. [Department of Microbiology, Immunology and Medical Genetics, The Ohio State University College of Medicine, Columbus, OH 43210 (United States); Acharya, Samir, E-mail: samir.acharya@osumc.edu [Department of Microbiology, Immunology and Medical Genetics, The Ohio State University College of Medicine, Columbus, OH 43210 (United States); Groden, Joanna [Department of Microbiology, Immunology and Medical Genetics, The Ohio State University College of Medicine, Columbus, OH 43210 (United States)

    2013-03-15

    Bloom's syndrome (BS) is an inherited disorder caused by loss of function of the recQ-like BLM helicase. It is characterized clinically by severe growth retardation and cancer predisposition. BLM localizes to PML nuclear bodies and to the nucleolus; its deficiency results in increased intra- and inter-chromosomal recombination, including hyper-recombination of rDNA repeats. Our previous work has shown that BLM facilitates RNA polymerase I-mediated rRNA transcription in the nucleolus (Grierson et al., 2012 [18]). This study uses protein co-immunoprecipitation and in vitro transcription/translation (IVTT) to identify a direct interaction of DNA topoisomerase I with the C-terminus of BLM in the nucleolus. In vitro helicase assays demonstrate that DNA topoisomerase I stimulates BLM helicase activity on a nucleolar-relevant RNA:DNA hybrid, but has an insignificant effect on BLM helicase activity on a control DNA:DNA duplex substrate. Reciprocally, BLM enhances the DNA relaxation activity of DNA topoisomerase I on supercoiled DNA substrates. Our study suggests that BLM and DNA topoisomerase I function coordinately to modulate RNA:DNA hybrid formation as well as relaxation of DNA supercoils in the context of nucleolar transcription.

  16. Effect of Welding Processes on Tensile and Impact Properties, Hardness and Microstructure of AISI 409M Ferritic Stainless Joints Fabricated by Duplex Stainless Steel Filler Metal

    Institute of Scientific and Technical Information of China (English)

    A K Lakshminarayanan; K Shanmugam; V Balasubramanian

    2009-01-01

    The effect of welding processes such as shielded metal arc welding, gas metal arc welding and gas tungsten arc welding on tensile and impact properties of the ferritic stainless steel conforming to AISI 409M grade is studied. Rolled plates of 4 mm thickness were used as the base material for preparing single pass butt welded joints. Tensile and impact properties, microhardness, microstructure and fracture surface morphology of the welded joints have been evaluated and the results are compared. From this investigatio.n, it is found that gas tungsten arc welded joints of ferritic stainless steel have superior tensile and impact properties compared with shielded metal are and gas metal arc welded joints and this is mainly due to the presence of finer grains in fusion zone and heat affected zone.

  17. Detection of Toxoplasma gondii and Epstein-Barr virus in HIV patients with clinical symptoms of suspected central nervous system infection using duplex real-time polymerase chain reaction

    Science.gov (United States)

    Rahmawati, E.; Ibrahim, F.; Imran, D.; Sudarmono, P.

    2017-08-01

    Focal brain lesion is a neurological complication in HIV, which is marked as a space occupying lesion (SOL) and needs rapid and effective treatment. This lesion is mainly caused by encephalitis toxoplasma and primary central nervous system lymphoma related to the Epstein-Barr virus (EBV) infection, which is difficult to distinguish using CT scan or magnetic resonance imaging (MRI). The gold standard of diagnosing focal brain lesion has been brain biopsy, but this examination is an invasive procedure that causes complications. The objective of this study is to obtain the rapid laboratory diagnosis of Toxoplasma gondii (T. gondii) and EBV infection. In this experimental study, blood and cerebrospinal fluid were obtained from HIV patients who were admitted to the Neurology Department of Cipto Mangunkusumo Hospital. The samples were examined using duplex real-time polymerase chain reaction (PCR) to detect T. gondii and EBV. The first step was the optimization of duplex real-time PCR, including the annealing temperature, primer and probe concentration, elution volume, and template volume. Minimal DNA detection was used to measure minimal T. gondii and EBV. Cross reactions were determined for technical specificity using the bacteria and viruses Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Mycobacterium tuberculosis H37Rv, Candida spp, cytomegalovirus, herpes zoster virus, and varicella zoster virus. Duplex real-time PCR was applied optimally to patients. In the optimization of duplex real-time PCR, the annealing temperature of T. gondii and EBV were 58 °C, the concentration of primer forward and reverse for T. gondii and EBV were 0.2 μM, the concentration of probe for T. gondii and EBV were 0.4μM and 0.2 μM, respectively. Minimal DNA detection of T. gondii and EBV were 5.68 copy/ml and 1.31 copy/ml, respectively. There was no cross reaction between another bacteria and virus that were used as the primer and probe for T. gondii and EBV. The

  18. Roles of the Amino Group of Purine Bases in the Thermodynamic Stability of DNA Base Pairing

    Directory of Open Access Journals (Sweden)

    Shu-ichi Nakano

    2014-08-01

    Full Text Available The energetic aspects of hydrogen-bonded base-pair interactions are important for the design of functional nucleotide analogs and for practical applications of oligonucleotides. The present study investigated the contribution of the 2-amino group of DNA purine bases to the thermodynamic stability of oligonucleotide duplexes under different salt and solvent conditions, using 2'-deoxyriboinosine (I and 2'-deoxyribo-2,6-diaminopurine (D as non-canonical nucleotides. The stability of DNA duplexes was changed by substitution of a single base pair in the following order: G•C > D•T ≈ I•C > A•T > G•T > I•T. The apparent stabilization energy due to the presence of the 2-amino group of G and D varied depending on the salt concentration, and decreased in the water-ethanol mixed solvent. The effects of salt concentration on the thermodynamics of DNA duplexes were found to be partially sequence-dependent, and the 2-amino group of the purine bases might have an influence on the binding of ions to DNA through the formation of a stable base-paired structure. Our results also showed that physiological salt conditions were energetically favorable for complementary base recognition, and conversely, low salt concentration media and ethanol-containing solvents were effective for low stringency oligonucleotide hybridization, in the context of conditions employed in this study.

  19. The effect of base pair mismatch on DNA strand displacement

    CERN Document Server

    Broadwater, Bo

    2016-01-01

    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 base pair 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 t...

  20. RF Self-Interference cancellation for Full-Duplex

    NARCIS (Netherlands)

    van Liempd, B.; Debaillie, B.; Craninckx, J.; Lavin, C.; Palacios, C.; Malotaux, S.; Long, J.R.; van den Broek, Dirk-Jan; Klumperink, Eric A.M.

    2014-01-01

    This paper proposes two RF self-interference cancellation techniques. Their small form-factor enables full-duplex communication links for small-to-medium size portable devices and hence promotes the adoption of full-duplex in mass-market applications and next-generation standards, e.g. IEEE802.11 an

  1. Scintigraphic features of duplex kidneys on DMSA renal cortical scans.

    Science.gov (United States)

    Kwatra, Neha; Shalaby-Rana, Eglal; Majd, Massoud

    2013-09-01

    The spectrum of manifestations of duplex kidneys on (99m)Tc-dimercaptosuccinic acid (DMSA) renal cortical scans and correlating findings on other imaging modalities are presented. Relevant embryology of the duplex systems and technical aspects of DMSA scintigraphy are reviewed.

  2. On the Ergodic Secrecy Capacity with Full Duplex Communication

    DEFF Research Database (Denmark)

    Mahmood, Nurul Huda; Shafique Ansari, Imran; Mogensen, Preben Elgaard;

    2017-01-01

    Full duplex communication promises performance gains in terms of the throughput and the delay. Generally, it has been shown that the throughput and delay gains of full duplex communication are somewhat limited in realistic network settings, leading researchers to study other possible applications...

  3. Autoregulation of the dnaA-dnaN operon and effects of DnaA protein levels on replication initiation in Bacillus subtilis.

    Science.gov (United States)

    Ogura, Y; Imai, Y; Ogasawara, N; Moriya, S

    2001-07-01

    In Escherichia coli, the DnaA protein level appears to play a pivotal role in determining the timing of replication initiation. To examine the effects on replication initiation in B. subtilis, we constructed a strain in which a copy of the dnaA gene was integrated at the purA locus on the chromosome under the control of an isopropyl-beta-D-thiogalactopyranoside (IPTG)-inducible promoter. However, increasing the DnaA level resulted in cell elongation and inhibition of cell growth by induction of the SOS response. Transcription of the native dnaA-dnaN operon was greatly reduced at high DnaA levels, but it was increased in a dnaA-null mutant, indicating autoregulation of the operon by DnaA. When a copy of the dnaN gene was added downstream of the additional dnaA gene at purA, the cells grew at high DnaA levels, suggesting that depletion of DnaN (beta subunit of DNA polymerase III) within the cell by repression of the native dnaA-dnaN operon at high DnaA levels was the cause of the SOS induction. Flow cytometry of the cells revealed that the cell mass at initiation of replication increased at a lower DnaA level and decreased at DnaA levels higher than those of the wild type. Proper timing of replication initiation was observed at DnaA levels nearly comparable to the wild-type level. These results suggest that if the DnaA level increases with progression of the replication cycle, it could act as a rate-limiting factor of replication initiation in B. subtilis.

  4. HPLC-an Effective Method of DNA Fragment Determination

    Institute of Scientific and Technical Information of China (English)

    HOU JingGuo; HE TianXi; MAO XueFeng; LIU HuiLing; DU XinZheng; NA PengJun; DENG HuaLing; GAO JingZhang

    2001-01-01

    @@ The role played by DNA in molecular biology is clearly recognized to be vital to life on this planet. 8-oxo-7,8-dihydro-2deoxyguanosine(=8-OHdG), is probably the most important product of "oxidative stress” in DNA. Its concentration in DNA is, in fact. a quantitative analysis of the degree of DNA damage that an organism has undergone. Due to the importance of 8-OHdG of nucleic acidg in mutagenesis, carcinogenesis and aging, numerous chemical and biological investigations have been made on this subject in the past time. Kuchino and co-workers have found that 8-OHdG residue in DNA is misreading during the process of DNA replication. Recently, some reports have been presented on high 8-OHdG levels in patients suffering from various diseases such as chronic hepatitis, Fanconi s anemia, diabetes mellitus and Helicobacter pylori infections. As a result, 8-OHdG is a useful marker for the study of DNA damage arising from reactive oxygen species and is of great significance for cancer research. The 8-OHdG levels in DNA can help understand the mechanism of carcinogens and lead to more effective treatments for many different types of cancer. For these reasons, an analysis of 8-OHdG with quickness, low cost and high accuracy is required.

  5. Effects of sequence on DNA wrapping around histones

    Science.gov (United States)

    Ortiz, Vanessa

    2011-03-01

    A central question in biophysics is whether the sequence of a DNA strand affects its mechanical properties. In epigenetics, these are thought to influence nucleosome positioning and gene expression. Theoretical and experimental attempts to answer this question have been hindered by an inability to directly resolve DNA structure and dynamics at the base-pair level. In our previous studies we used a detailed model of DNA to measure the effects of sequence on the stability of naked DNA under bending. Sequence was shown to influence DNA's ability to form kinks, which arise when certain motifs slide past others to form non-native contacts. Here, we have now included histone-DNA interactions to see if the results obtained for naked DNA are transferable to the problem of nucleosome positioning. Different DNA sequences interacting with the histone protein complex are studied, and their equilibrium and mechanical properties are compared among themselves and with the naked case. NLM training grant to the Computation and Informatics in Biology and Medicine Training Program (NLM T15LM007359).

  6. HPLC-an Effective Method of DNA Fragment Determination

    Institute of Scientific and Technical Information of China (English)

    HOU; JingGuo

    2001-01-01

    The role played by DNA in molecular biology is clearly recognized to be vital to life on this planet. 8-oxo-7,8-dihydro-2deoxyguanosine(=8-OHdG), is probably the most important product of "oxidative stress” in DNA. Its concentration in DNA is, in fact. a quantitative analysis of the degree of DNA damage that an organism has undergone. Due to the importance of 8-OHdG of nucleic acidg in mutagenesis, carcinogenesis and aging, numerous chemical and biological investigations have been made on this subject in the past time. Kuchino and co-workers have found that 8-OHdG residue in DNA is misreading during the process of DNA replication. Recently, some reports have been presented on high 8-OHdG levels in patients suffering from various diseases such as chronic hepatitis, Fanconi s anemia, diabetes mellitus and Helicobacter pylori infections. As a result, 8-OHdG is a useful marker for the study of DNA damage arising from reactive oxygen species and is of great significance for cancer research. The 8-OHdG levels in DNA can help understand the mechanism of carcinogens and lead to more effective treatments for many different types of cancer. For these reasons, an analysis of 8-OHdG with quickness, low cost and high accuracy is required.  ……

  7. Dynamics of nucleosome assembly and effects of DNA methylation.

    Science.gov (United States)

    Lee, Ju Yeon; Lee, Jaehyoun; Yue, Hongjun; Lee, Tae-Hee

    2015-02-13

    The nucleosome is the fundamental packing unit of the eukaryotic genome, and CpG methylation is an epigenetic modification associated with gene repression and silencing. We investigated nucleosome assembly mediated by histone chaperone Nap1 and the effects of CpG methylation based on three-color single molecule FRET measurements, which enabled direct monitoring of histone binding in the context of DNA wrapping. According to our observation, (H3-H4)2 tetramer incorporation must precede H2A-H2B dimer binding, which is independent of DNA termini wrapping. Upon CpG methylation, (H3-H4)2 tetramer incorporation and DNA termini wrapping are facilitated, whereas proper incorporation of H2A-H2B dimers is inhibited. We suggest that these changes are due to rigidified DNA and increased random binding of histones to DNA. According to the results, CpG methylation expedites nucleosome assembly in the presence of abundant DNA and histones, which may help facilitate gene packaging in chromatin. The results also indicate that the slowest steps in nucleosome assembly are DNA termini wrapping and tetramer positioning, both of which are affected heavily by changes in the physical properties of DNA.

  8. Duplex scanning using sparse data sequences

    DEFF Research Database (Denmark)

    Møllenbach, S. K.; Jensen, Jørgen Arendt

    2008-01-01

    The velocity distribution in vessels can be displayed using duplex scanning where B-mode acquisitions are interspaced with the velocity data. This gives an image for orientation, but lowers the maximum detectable velocity by a factor of two. Other pulse sequences either omits the B-mode image...... or leaves gaps in the velocity data, which makes it difficult to output audio data. The near full velocity range can be maintained and B-mode images shown by using a sparse data sequence with velocity and B-mode samples intermixed. The B-mode samples are placed in a (sparse) periodical pattern, which makes...... is scaled by the factor A/T. The approach has been investigated using in vivo RF data from the Hepatic vein, Carotid artery and Aorta from a 33 year old healthy male. A B-K Medical 3535 ultrasound scanner has been used in Duplex mode with a BK 8556, 3.2 MHz linear array probe. The sampling frequency...

  9. The effect of volume exclusion on the formation of DNA minicircle networks: implications to kinetoplast DNA

    Science.gov (United States)

    Diao, Y.; Hinson, K.; Sun, Y.; Arsuaga, J.

    2015-10-01

    Kinetoplast DNA (kDNA) is the mitochondrial of DNA of disease causing organisms such as Trypanosoma Brucei (T. Brucei) and Trypanosoma Cruzi (T. Cruzi). In most organisms, KDNA is made of thousands of small circular DNA molecules that are highly condensed and topologically linked forming a gigantic planar network. In our previous work we have developed mathematical and computational models to test the confinement hypothesis, that is that the formation of kDNA minicircle networks is a product of the high DNA condensation achieved in the mitochondrion of these organisms. In these studies we studied three parameters that characterize the growth of the network topology upon confinement: the critical percolation density, the mean saturation density and the mean valence (i.e. the number of mini circles topologically linked to any chosen minicircle). Experimental results on insect-infecting organisms showed that the mean valence is equal to three, forming a structure similar to those found in medieval chain-mails. These same studies hypothesized that this value of the mean valence was driven by the DNA excluded volume. Here we extend our previous work on kDNA by characterizing the effects of DNA excluded volume on the three descriptive parameters. Using computer simulations of polymer swelling we found that (1) in agreement with previous studies the linking probability of two minicircles does not decrease linearly with the distance between the two minicircles, (2) the mean valence grows linearly with the density of minicircles and decreases with the thickness of the excluded volume, (3) the critical percolation and mean saturation densities grow linearly with the thickness of the excluded volume. Our results therefore suggest that the swelling of the DNA molecule, due to electrostatic interactions, has relatively mild implications on the overall topology of the network. Our results also validate our topological descriptors since they appear to reflect the changes in the

  10. Development and Evaluation of a Single-Step Duplex PCR for Simultaneous Detection of Fasciola hepatica and Fasciola gigantica (Family Fasciolidae, Class Trematoda, Phylum Platyhelminthes)

    Science.gov (United States)

    Nguyen, Khue Thi; Nguyen, Nga Thi Bich; Doan, Huong Thi Thanh; Le, Xuyen Thi Kim; Hoang, Chau Thi Minh; De, Nguyen Van

    2012-01-01

    A single-step multiplex PCR (here referred to as a duplex PCR) has been developed for simultaneous detection and diagnosis of Fasciola hepatica and F. gigantica. These species overlap in distribution in many countries of North and East Africa and Central and Southeast Asia and are similar in egg morphology, making identification from fecal samples difficult. Based on a comparative alignment of mitochondrial DNA (mtDNA) spanning the region of cox1-trnT-rrnL, two species-specific forward primers were designed, FHF (for F. hepatica) and FGF (for F. gigantica), and a single reverse primer, FHGR (common for both species). Conventional PCR followed by sequencing was applied using species-specific primer pairs to verify the specificity of primers and the identity of Fasciola DNA templates. Duplex PCR (using three primers) was used for testing with the DNA extracted from adult worms, miracidia, and eggs, producing amplicons of 1,031 bp for F. hepatica and 615 bp for F. gigantica. The duplex PCR failed to amplify from DNA of other common liver and intestinal trematodes, including two opisthorchiids, three heterophyids, an echinostomid, another fasciolid, and a taeniid cestode. The sensitivity assay showed that the duplex PCR limit of detection for each Fasciola species was between 0.012 ng and 0.006 ng DNA. Evaluation using DNA templates from 32 Fasciola samples (28 adults and 4 eggs) and from 25 field-collected stools of ruminants and humans revealed specific bands of the correct size and the presence of Fasciola species. This novel mtDNA duplex PCR is a sensitive and fast tool for accurate identification of Fasciola species in areas of distributional and zonal overlap. PMID:22692744

  11. Development and evaluation of a single-step duplex PCR for simultaneous detection of Fasciola hepatica and Fasciola gigantica (family Fasciolidae, class Trematoda, phylum Platyhelminthes).

    Science.gov (United States)

    Le, Thanh Hoa; Nguyen, Khue Thi; Nguyen, Nga Thi Bich; Doan, Huong Thi Thanh; Le, Xuyen Thi Kim; Hoang, Chau Thi Minh; De, Nguyen Van

    2012-08-01

    A single-step multiplex PCR (here referred to as a duplex PCR) has been developed for simultaneous detection and diagnosis of Fasciola hepatica and F. gigantica. These species overlap in distribution in many countries of North and East Africa and Central and Southeast Asia and are similar in egg morphology, making identification from fecal samples difficult. Based on a comparative alignment of mitochondrial DNA (mtDNA) spanning the region of cox1-trnT-rrnL, two species-specific forward primers were designed, FHF (for F. hepatica) and FGF (for F. gigantica), and a single reverse primer, FHGR (common for both species). Conventional PCR followed by sequencing was applied using species-specific primer pairs to verify the specificity of primers and the identity of Fasciola DNA templates. Duplex PCR (using three primers) was used for testing with the DNA extracted from adult worms, miracidia, and eggs, producing amplicons of 1,031 bp for F. hepatica and 615 bp for F. gigantica. The duplex PCR failed to amplify from DNA of other common liver and intestinal trematodes, including two opisthorchiids, three heterophyids, an echinostomid, another fasciolid, and a taeniid cestode. The sensitivity assay showed that the duplex PCR limit of detection for each Fasciola species was between 0.012 ng and 0.006 ng DNA. Evaluation using DNA templates from 32 Fasciola samples (28 adults and 4 eggs) and from 25 field-collected stools of ruminants and humans revealed specific bands of the correct size and the presence of Fasciola species. This novel mtDNA duplex PCR is a sensitive and fast tool for accurate identification of Fasciola species in areas of distributional and zonal overlap.

  12. Effect of Bonding Temperature on Phase Transformation of Diffusion-Bonded Joints of Duplex Stainless Steel and Ti-6Al-4V Using Nickel and Copper as Composite Intermediate Metals

    Science.gov (United States)

    Kundu, Sukumar; Thirunavukarasu, Gopinath; Chatterjee, Subrata; Mishra, Brajendra

    2015-12-01

    In the present study, the effect of bonding temperature on phase transformation of diffusion-bonded joints of duplex stainless steel (DSS) and Ti-6Al-4V (Ti64) using simultaneously both nickel (Ni) and copper (Cu) interlayers was investigated in the temperature range of 1148 K to 1223 K (875 °C to 950 °C) insteps of 25 K (25 °C) for 60 minutes under 4 MPa uniaxial pressure in vacuum. Interfaces were characterized by scanning electron microscopy and interdiffusion of the chemical species across the diffusion interfaces were witnessed by electron probe microanalysis. At 1148 K (875 °C), layer-wise Cu4Ti, Cu2Ti, Cu4Ti3, CuTi, and CuTi2 phases were observed at the Cu-Ti64 interface; however, DSS-Ni and Ni-Cu interfaces were free from any intermetallic. At 1173 K and 1198 K (900 °C and 925 °C), Cu interlayer could not restrict the diffusion of atoms from Ti64 to Ni, and vice versa; and Ni-Ti-based intermetallics were formed at the Ni-Cu interface and throughout the Cu zone as well; however, at 1223 K (950 °C), both Ni and Cu interlayers could not inhibit the diffusion of atoms from Ti64 to DSS, and vice versa. The maximum shear strength of ~377 MPa was obtained for the diffusion couple processed at 1148 K (875 °C) and strength of the bonded joints gradually decreased with the increasing bonding temperature due to the widening of brittle intermetallics at the diffusion zone. Fracture path indicated that failure took place through the Cu4Ti intermetallic at the Cu-Ti64 interface when bonding was processed at 1148 K (875 °C). When bonding was processed at 1173 K and 1198 K (900 °C and 925 °C), fracture took place through the Ni3Ti intermetallic at the Ni-(Ni + Cu + Ti64 diffusion reaction) interface; however, at 1223 K (950 °C), fracture morphology indicated the brittle nature and the fracture took place apparently through the σ phase at the DSS-(DSS + Ni + Cu + Ti64 diffusion reaction) interface.

  13. Elasticity of DNA and the effect of Dendrimer Binding

    CERN Document Server

    Mogurampelly, Santosh; Netz, Roland R; Maiti, Prabal K

    2013-01-01

    Negatively charged DNA can be compacted by positively charged dendrimers and the degree of compaction is a delicate balance between the strength of the electrostatic interaction and the elasticity of DNA. We report various elastic properties of short double stranded DNA (dsDNA) and the effect of dendrimer binding using fully atomistic molecular dynamics and numerical simulations. In equilibrium at room temperature, the contour length distribution P(L) and end-to-end distance distribution P(R) are nearly Gaussian, the former gives an estimate of the stretch modulus {\\gamma}_1 of dsDNA in quantitative agreement with the literature value. The bend angle distribution P({\\theta}) of the dsDNA also has a Gaussian form and allows to extract a persistence length, L_p of 43 nm. When the dsDNA is compacted by positively charged dendrimer, the stretch modulus stays invariant but the effective bending rigidity estimated from the end-to-end distance distribution decreases dramatically due to backbone charge neutralization...

  14. The N(2)-Furfuryl-deoxyguanosine Adduct Does Not Alter the Structure of B-DNA.

    Science.gov (United States)

    Ghodke, Pratibha P; Gore, Kiran R; Harikrishna, S; Samanta, Biswajit; Kottur, Jithesh; Nair, Deepak T; Pradeepkumar, P I

    2016-01-15

    N(2)-Furfuryl-deoxyguanosine (fdG) is carcinogenic DNA adduct that originates from furfuryl alcohol. It is also a stable structural mimic of the damage induced by the nitrofurazone family of antibiotics. For the structural and functional studies of this model N(2)-dG adduct, reliable and rapid access to fdG-modified DNAs are warranted. Toward this end, here we report the synthesis of fdG-modified DNAs using phosphoramidite chemistry involving only three steps. The functional integrity of the modified DNA has been verified by primer extension studies with DNA polymerases I and IV from E. coli. Introduction of fdG into a DNA duplex decreases the Tm by ∼1.6 °C/modification. Molecular dynamics simulations of a DNA duplex bearing the fdG adduct revealed that though the overall B-DNA structure is maintained, this lesion can disrupt W-C H-bonding, stacking interactions, and minor groove hydrations to some extent at the modified site, and these effects lead to slight variations in the local base pair parameters. Overall, our studies show that fdG is tolerated at the minor groove of the DNA to a better extent compared with other bulky DNA damages, and this property will make it difficult for the DNA repair pathways to detect this adduct.

  15. Binding and interaction of di- and tri-substituted organometallic triptycene palladium complexes with DNA.

    Science.gov (United States)

    Kumari, Rina; Bhowmick, Sourav; Das, Neeladri; Das, Prolay

    2014-10-01

    Two triptycene-based ligands with pendant bromophenyl units have been prepared. These triptycene derivatives have been used as synthons for the synthesis of di and tri nuclear palladium complexes. The organic molecules and their corresponding organometallic complexes have been fully characterized using nuclear magnetic resonance (NMR), infrared (IR) spectroscopy and mass spectrometry. The mode of binding and effect of the complexes on pUC19 plasmid, calf thymus DNA and oligomer duplex DNA have been investigated by a host of analytical methods. The complexes brought about unwinding of supercoiled plasmid and the unwinding angle was found to be related to the binding affinity of the complexes with DNA, where both these parameters were guided by the structure of the complexes. Concentration-dependent inhibition of endonuclease activity of SspI and BamHI by the complexes indicates preference for G/C sequence for binding to DNA. However, neither the complexes did not introduce any cleavage at abasic site in oligomer duplex DNA, nor they created linear form of the plasmid upon co-incubation with the DNA samples. The interactions of the complexes with DNA were found to be strongly guided by the structure of the complexes, where intercalation as well as groove binding was observed, without inflicting any damage to the DNA. The mode of interaction of the complexes with DNA was further confirmed by isothermal calorimetry.

  16. [Analysis of Conformational Features of Watson-Crick Duplex Fragments by Molecular Mechanics and Quantum Mechanics Methods].

    Science.gov (United States)

    Poltev, V I; Anisimov, V M; Sanchez, C; Deriabina, A; Gonzalez, E; Garcia, D; Rivas, F; Polteva, N A

    2016-01-01

    It is generally accepted that the important characteristic features of the Watson-Crick duplex originate from the molecular structure of its subunits. However, it still remains to elucidate what properties of each subunit are responsible for the significant characteristic features of the DNA structure. The computations of desoxydinucleoside monophosphates complexes with Na-ions using density functional theory revealed a pivotal role of DNA conformational properties of single-chain minimal fragments in the development of unique features of the Watson-Crick duplex. We found that directionality of the sugar-phosphate backbone and the preferable ranges of its torsion angles, combined with the difference between purines and pyrimidines. in ring bases, define the dependence of three-dimensional structure of the Watson-Crick duplex on nucleotide base sequence. In this work, we extended these density functional theory computations to the minimal' fragments of DNA duplex, complementary desoxydinucleoside monophosphates complexes with Na-ions. Using several computational methods and various functionals, we performed a search for energy minima of BI-conformation for complementary desoxydinucleoside monophosphates complexes with different nucleoside sequences. Two sequences are optimized using ab initio method at the MP2/6-31++G** level of theory. The analysis of torsion angles, sugar ring puckering and mutual base positions of optimized structures demonstrates that the conformational characteristic features of complementary desoxydinucleoside monophosphates complexes with Na-ions remain within BI ranges and become closer to the corresponding characteristic features of the Watson-Crick duplex crystals. Qualitatively, the main characteristic features of each studied complementary desoxydinucleoside monophosphates complex remain invariant when different computational methods are used, although the quantitative values of some conformational parameters could vary lying within the

  17. Enhancement Effect of CpG DNA on the Somatostatin DNA Vaccine in Mice

    Institute of Scientific and Technical Information of China (English)

    XUE Chun-lin; MAO Da-gan; YANG Li-guo; CHENG Bao

    2007-01-01

    To study the immune effect of CpGDNA on somatostatin (SS) DNA vaccine, the 20-day-old experimental mice were immunized with 20 μg SS eukaryotic expression plasmid pES/2SS with different adjuvants in equal dose, such as the synthetic CpG-ODN, the pE-CpG plasmid, E. coli DNA and the crude liposome. A booster was given two weeks later. The results showed that the body weight gain of female mice in the SS immunized group was higher than that of the control (P<0.05). The levels of antibodies against SS, IgG2a/IgG1, spleen lymphocyte proliferation activity and the concentrations of GH and IGF- Ⅰ in the DNA vaccine groups combined with CpGDNA were significantly increased compared to that of the group immunized with DNA vaccine alone. All these suggested the recombinant SS expression plasmid can stimulate animals to produce antibodies against SS, and CpGDNA adjuvant can enhance the immune effect of DNA vaccine against SS and influence the concentration of GH and IGF- Ⅰ.

  18. Effect of fluoride ion on the stability of DNA hairpin

    Science.gov (United States)

    Liu, Chao; Zhai, Weili; Gong, Hongling; Liu, Yanhui; Chen, Hu

    2017-06-01

    Fluoride prevents tooth decay as an additive in oral hygiene products, while high dose intake of fluoride from contaminated drinking water leads to fluorosis. Here we studied the effect of fluoride ion on the stability of DNA double helix using magnetic tweezers. The equilibrium critical force decreases with increasing concentration of fluoride in the range from 1 mM to 100 mM. Our results give the first quantitative measurement of DNA stability in the presence of fluoride ion, which might disturb DNA-related biological processes to cause fluorosis.

  19. Effect of clustered peptide binding on DNA condensation.

    Science.gov (United States)

    Haley, Jennifer; Kabiru, Paul; Geng, Yan

    2010-01-01

    DNA condensation in-vitro has been studied as a model system to reveal common principles underlying gene packaging in biology, and as the critical first step towards the development of non-viral gene delivery vectors. In this study, we use a bio-inspired approach, where small DNA-binding peptides are controllably clustered by an amphiphilic block copolymer scaffold, to reveal the effect of clustered peptide binding on the energetics, size, shape and physical properties of DNA condensation in-vitro. This provides insights into the general architectural effect of gene-binding proteins on DNA condensation process. Moreover, the versatility afforded by regulating the clustering density and composition of peptides may provide a novel design platform for gene delivery applications in the future.

  20. Assessment of okadaic acid effects on cytotoxicity, DNA damage and DNA repair in human cells.

    Science.gov (United States)

    Valdiglesias, Vanessa; Méndez, Josefina; Pásaro, Eduardo; Cemeli, Eduardo; Anderson, Diana; Laffon, Blanca

    2010-07-07

    Okadaic acid (OA) is a phycotoxin produced by several types of dinoflagellates causing diarrheic shellfish poisoning (DSP) in humans. Symptoms induced by DSP toxins are mainly gastrointestinal, but the intoxication does not appear to be fatal. Despite this, this toxin presents a potential threat to human health even at concentrations too low to induce acute toxicity, since previous animal studies have shown that OA has very potent tumour promoting activity. However, its concrete action mechanism has not been described yet and the results reported with regard to OA cytotoxicity and genotoxicity are often contradictory. In the present study, the genotoxic and cytotoxic effects of OA on three different types of human cells (peripheral blood leukocytes, HepG2 hepatoma cells, and SHSY5Y neuroblastoma cells) were evaluated. Cells were treated with a range of OA concentrations in the presence and absence of S9 fraction, and MTT test and Comet assay were performed in order to evaluate cytotoxicity and genotoxicity, respectively. The possible effects of OA on DNA repair were also studied by means of the DNA repair competence assay, using bleomycin as DNA damage inductor. Treatment with OA in absence of S9 fraction induced not statistically significant decrease in cell viability and significant increase in DNA damage in all cell types at the highest concentrations investigated. However, only SHSY5Y cells showed OA induced genotoxic and cytotoxic effects in presence of S9 fraction. Furthermore, we found that OA can induce modulations in DNA repair processes when exposure was performed prior to BLM treatment, in co-exposure, or during the subsequent DNA repair process. Copyright 2010 Elsevier B.V. All rights reserved.

  1. Binding specificity and stability of duplexes formed by modified oligonucleotides with a 4096-hexanucleotide microarray

    Science.gov (United States)

    Timofeev, Edward; Mirzabekov, Andrei

    2001-01-01

    The binding of oligodeoxynucleotides modified with adenine 2′-O-methyl riboside, 2,6-diaminopurine 2′-O-methyl riboside, cytosine 2′-O-methyl riboside, 2,6-diaminopurine deoxyriboside or 5-bromodeoxyuridine was studied with a microarray containing all possible (4096) polyacrylamide-bound hexadeoxynucleotides (a generic microchip). The generic microchip was manufactured by using reductive immobilization of aminooligonucleotides in the activated copolymer of acrylamide, bis-acrylamide and N-(2,2-dimethoxyethyl) acrylamide. The binding of the fluorescently labeled modified octanucleotides to the array was analyzed with the use of both melting profiles and the fluorescence distribution at selected temperatures. Up to three substitutions of adenosines in the octamer sequence by adenine 2′-O-methyl ribosides (Am), 2,6-diaminopurine 2′-O-methyl ribosides (Dm) or 2,6-diaminopurine deoxyribosides (D) resulted in increased mismatch discrimination measured at the melting temperature of the corresponding perfect duplex. The stability of complexes formed by 2′-O-methyl-adenosine-modified oligodeoxynucleotides was slightly decreased with every additional substitution, yielding ∼4°C of total loss in melting temperature for three modifications, as followed from microchip thermal denaturation experiments. 2,6-Diaminopurine 2′-O-methyl riboside modifications led to considerable duplex stabilization. The cytosine 2′-O-methyl riboside and 5-bromodeoxyuridine modifications generally did not change either duplex stability or mismatch resolution. Denaturation experiments conducted with selected perfect duplexes on microchips and in solution showed similar results on thermal stabilities. Some hybridization artifacts were observed that might indicate the formation of parallel DNA. PMID:11410672

  2. Effect of sample storage time on detection of hybridization signals in Checkerboard DNA-DNA hybridization.

    Science.gov (United States)

    do Nascimento, Cássio; Muller, Katia; Sato, Sandra; Albuquerque Junior, Rubens Ferreira

    2012-04-01

    Long-term sample storage can affect the intensity of the hybridization signals provided by molecular diagnostic methods that use chemiluminescent detection. The aim of this study was to evaluate the effect of different storage times on the hybridization signals of 13 bacterial species detected by the Checkerboard DNA-DNA hybridization method using whole-genomic DNA probes. Ninety-six subgingival biofilm samples were collected from 36 healthy subjects, and the intensity of hybridization signals was evaluated at 4 different time periods: (1) immediately after collecting (n = 24) and (2) after storage at -20 °C for 6 months (n = 24), (3) for 12 months (n = 24), and (4) for 24 months (n = 24). The intensity of hybridization signals obtained from groups 1 and 2 were significantly higher than in the other groups (p  0.05). The Checkerboard DNA-DNA hybridization method was suitable to detect hybridization signals from all groups evaluated, and the intensity of signals decreased significantly after long periods of sample storage.

  3. Magnetic fields facilitate DNA-mediated charge transport

    CERN Document Server

    Wong, Jiun Ru; Shu, Jian-Jun; Shao, Fangwei

    2015-01-01

    Exaggerate radical-induced DNA damage under magnetic fields is of great concerns to medical biosafety and to bio-molecular device based upon DNA electronic conductivity. In this report, the effect of applying an external magnetic field (MF) on DNA-mediated charge transport (CT) was investigated by studying guanine oxidation by a kinetics trap (8CPG) via photoirradiation of anthraquinone (AQ) in the presence of an external MF. Positive enhancement in CT efficiencies was observed in both the proximal and distal 8CPG after applying a static MF of 300 mT. MF assisted CT has shown sensitivities to magnetic field strength, duplex structures, and the integrity of base pair stacking. MF effects on spin evolution of charge injection upon AQ irradiation and alignment of base pairs to CT-active conformation during radical propagation were proposed to be the two major factors that MF attributed to facilitate DNA-mediated CT. Herein, our results suggested that the electronic conductivity of duplex DNA can be enhanced by a...

  4. Buffer management in wireless full-duplex systems

    KAUST Repository

    Bouacida, Nader

    2015-10-19

    Wireless full-duplex radios can simultaneously transmit and receive using the same frequency. In theory, this can double the throughput. In fact, there is only little work addressing aspects other than throughput gains in full-duplex systems. Over-buffering in today\\'s networks or the so-called “bufferbloat” phenomenon creates excessive end-to-end delays resulting in network performance degradation. Our analysis shows that full-duplex systems may suffer from high latency caused by bloated buffers. In this paper, we address the problem of buffer management in full-duplex networks by using Wireless Queue Management (WQM), which is an active queue management technique for wireless networks. Our solution is based on Relay Full-Duplex MAC (RFD-MAC), an asynchronous media access control protocol designed for relay full-duplexing. We compare the performance of WQM in full-duplex environment to Drop Tail mechanism over various scenarios. Our solution reduces the end-to-end delay by two orders of magnitude while achieving similar throughput in most of the cases.

  5. Detection of DNA damage by using hairpin molecular beacon probes and graphene oxide.

    Science.gov (United States)

    Zhou, Jie; Lu, Qian; Tong, Ying; Wei, Wei; Liu, Songqin

    2012-09-15

    A hairpin molecular beacon tagged with carboxyfluorescein in combination with graphene oxide as a quencher reagent was used to detect the DNA damage by chemical reagents. The fluorescence of molecular beacon was quenched sharply by graphene oxide; while in the presence of its complementary DNA the quenching efficiency decreased because their hybridization prevented the strong adsorbability of molecular beacon on graphene oxide. If the complementary DNA was damaged by a chemical reagent and could not form intact duplex structure with molecular beacon, more molecular beacon would adsorb on graphene oxide increasing the quenching efficiency. Thus, damaged DNA could be detected based on different quenching efficiencies afforded by damaged and intact complementary DNA. The damage effects of chlorpyrifos-methyl and three metabolites of styrene such as mandelieaeids, phenylglyoxylieaeids and epoxystyrene on DNA were studied as models. The method for detection of DNA damage was reliable, rapid and simple compared to the biological methods. Copyright © 2012 Elsevier B.V. All rights reserved.

  6. Viral reverse transcriptases show selective high affinity binding to DNA-DNA primer-templates that resemble the polypurine tract.

    Directory of Open Access Journals (Sweden)

    Gauri R Nair

    Full Text Available Previous results using a SELEX (Systematic Evolution of Ligands by Exponential Enrichment-based approach that selected DNA primer-template duplexes binding with high affinity to HIV reverse transcriptase (RT showed that primers mimicking the 3' end, and in particular the six nt terminal G tract, of the RNA polypurine tract (PPT; HIV PPT: 5'-AAAAGAAAAGGGGGG-3' were preferentially selected. In this report, two viral (Moloney murine leukemia virus (MuLV and avian myeloblastosis virus (AMV and one retrotransposon (Ty3 RTs were used for selection. Like HIV RT, both viral RTs selected duplexes with primer strands mimicking the G tract at the PPT 3' end (AMV PPT: 5'-AGGGAGGGGGA-3'; MuLV PPT: 5'-AGAAAAAGGGGGG-3'. In contrast, Ty3, whose PPT lacks a G tract (5'-GAGAGAGAGGAA-3' showed no selective binding to any duplex sequences. Experiments were also conducted with DNA duplexes (termed DNA PPTs mimicking the RNA PPT-DNA duplex of each virus and a control duplex with a random DNA sequence. Retroviral RTs bound with high affinity to all viral DNA PPT constructs, with HIV and MuLV RTs showing comparable binding to the counterpart DNA PPT duplexes and reduced affinity to the AMV DNA PPT. AMV RT showed similar behavior with a modest preference for its own DNA PPT. Ty3 RT showed no preferential binding for its own or any other DNA PPT and viral RTs bound the Ty3 DNA PPT with relatively low affinity. In contrast, binding affinity of HIV RT to duplexes containing the HIV RNA PPT was less dependent on the G tract, which is known to be pivotal for efficient extension. We hypothesize that the G tract on the RNA PPT helps shift the binding orientation of RT to the 3' end of the PPT where extension can occur.

  7. Racemic DNA crystallography.

    Science.gov (United States)

    Mandal, Pradeep K; Collie, Gavin W; Kauffmann, Brice; Huc, Ivan

    2014-12-22

    Racemates increase the chances of crystallization by allowing molecular contacts to be formed in a greater number of ways. With the advent of protein synthesis, the production of protein racemates and racemic-protein crystallography are now possible. Curiously, racemic DNA crystallography had not been investigated despite the commercial availability of L- and D-deoxyribo-oligonucleotides. Here, we report a study into racemic DNA crystallography showing the strong propensity of racemic DNA mixtures to form racemic crystals. We describe racemic crystal structures of various DNA sequences and folded conformations, including duplexes, quadruplexes, and a four-way junction, showing that the advantages of racemic crystallography should extend to DNA.

  8. Competitive Effects of 2+ and 3+ Cations on DNA Compaction

    CERN Document Server

    Tongu, C; Yoshikawa, Y; Zinchenko, A A; Chen, N; Yoshikawa, K

    2016-01-01

    By using single-DNA observation with fluorescence microscopy, we observed the effects of divalent and trivalent cations on the higher-order structure of giant DNA (T4 DNA with 166 kbp). It was found that divalent cations, such as Mg(2+) and Ca(2+), inhibit DNA compaction induced by a trivalent cation, spermidine (SPD(3+)). On the other hand, in the absence of SPD(3+), divalent cations cause the shrinkage of DNA. These experimental observations are inconsistent with the well-established Debye-Huckel scheme regarding the shielding effect of counter ions, which is given as the additivity of contributions of cations with different valences. We interpreted the competition between 2+ and 3+ cations in terms of the change in the translational entropy of the counter ions before and after the folding transition of DNA. For the compaction with SPD(3+), we considered the increase in translational entropy due to the ion-exchange of the intrinsic monovalent cations condensing on a highly-charged polyelectrolyte, double-st...

  9. Duplex 2209 Weld Overlay by ESSC Process

    Directory of Open Access Journals (Sweden)

    Er. Manoj Kumar

    2017-03-01

    Full Text Available In the modern world of industrialization the wear is eating metal assets worth millions of dollars per year. The wear is in the form of corrosion, erosion, abrasion etc. which occur in the process industries like oil & gas, refineries, cement plants, steel plants, shipping and offshore working structures. The equipments like pressure vessels, heat exchangers, hydro processing reactors which very often work at elevated temperatures face corrosion in the internal diameter. Duplex 2209 weld overlay on ferrous material is developed for high corrosion resistance properties and having high productivity by Electroslag strip cladding process due to its less dilution ~10% as compared to SMAW , GTAW or FCAW process. Because of Low Dilution ~10% undiluted chemistry can be achieved with single layer as compared to other weld overlay processes. The facility was developed inhouse to carry out weld overlay by ESSC and Testing

  10. MICROSCOPIC CORROSION STUDIES OF DUPLEX STAINLESS STEELS

    Institute of Scientific and Technical Information of China (English)

    C.Leygraf; J.Pan; M.Femenia

    2004-01-01

    Electrochemical scanning tunneling microscopy and scanning electrochemical microscopy have been used for in situ monitoring of localized corrosion processes of different Duplex stainless steels (DSS) in acidic chloride solutions. The techniques allow imaging of local dissolution events with micrometer resolution, as opposed to conventional electrochemical techniques, which only give an overall view of the corrosion behavior. In addition, combined scanning Kelvin probe force microscopy and magnetic force microscopy were used for mapping the Volta potential variation over the surface of DSSs. A significant difference in Volta potential between the austenite and ferrite phases suggests galvanic interaction between the phases. A compositional gradient appears within 2 micrometers across the phase boundary, as seen with scanning Auger microscopy (SAM). In all, the studies suggest that higher alloyed DSS exhibit a more homogeneous dissolution behavior than lower alloyed DSS, due to higher and more similar corrosion resistance of the two phases, and enhanced resistance of the ferrite/austenite phase boundary regions.

  11. Rietveld and impedance analysis of cold and hot rolled duplex and lean duplex steels for application in paper and pulp industry

    Energy Technology Data Exchange (ETDEWEB)

    Esteves, Luiza; Lins, Vanessa de Freitas Cunha, E-mail: luizaeq@yahoo.com.br [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Departamento de Engenharia Quimica; Paiva, Paulo Renato Perdigao [Centro Federal de Educacao Tecnologica de Minas Gerais (CEFET), Belo Horizonte, MG (Brazil); Viana, Adolfo Kalergis do Nascimento [APERAM South America, Timoteo, MG (Brazil)

    2017-01-15

    In this study, X-Ray Diffraction (XRD) and Rietveld Refinement were performed to identify and quantify the ferrite and austenite phase of cold and hot rolled duplex stainless steels (UNS S31803) and lean duplex stainless steels (UNS S32304). Electrochemical impedance spectroscopy (EIS) was applied to evaluate the chemical behavior of duplex and lean duplex stainless steels in white, green, and black liquors of paper and pulp industry. Rietveld analysis results showed a higher austenite content than the standard limit for duplex steels in the hot rolled condition. The hot rolling condition plays a major role in improving corrosion resistance in white liquor mainly for the lean duplex steel. (author)

  12. Artificial, parallel, left-handed DNA helices.

    Science.gov (United States)

    Tian, Cheng; Zhang, Chuan; Li, Xiang; Li, Yingmei; Wang, Guansong; Mao, Chengde

    2012-12-19

    This communication reports an engineered DNA architecture. It contains multiple domains of half-turn-long, standard B-DNA duplexes. While each helical domain is right-handed and its two component strands are antiparallel, the global architecture is left-handed and the two component DNA strands are oriented parallel to each other.

  13. Duplex scanning on admission prevents unnecessary carotid endarterectomies.

    Science.gov (United States)

    Dalainas, I; Nano, G; Casana, R; Bianchi, P; Stegher, S; Malacrida, G; Tealdi, D G

    2006-06-01

    This retrospective single Institution study, aims to evaluate the performance of duplex scanning on admission of patients with carotid artery disease to avoid unnecessary carotid endarterectomies. From 1 January 1997 until 31 Decem-ber 2004, 1 504 patients were admitted to our Institution to undergo carotid endarterectomy. A duplex scan on admission was performed in all of them. A total of 1 369 from these patients (91%) underwent surgery, while 135 (9%) were dismissed because there was no indication for surgical treatment. They were put in conservative treatment and periodic duplex control appointments. In 9% of the patients, unnecessary carotid endarterectomy was avoided.

  14. Analysing Self Interference Cancellation in Full Duplex Radios

    DEFF Research Database (Denmark)

    Mahmood, Nurul Huda; Shafique Ansari, Imran; Berardinelli, Gilberto;

    2016-01-01

    Full duplex communication promises a theoretical $100\\%$ throughput gain by doubling the number of simultaneous transmissions. Such compelling gains are conditioned on perfect cancellation of the self interference power resulting from simultaneous transmission and reception. Generally, self...... cancellation. In this study, we derive an analytical model for the residual self interference power, and demonstrate various applications of the derived model in analysing the performance of a Full Duplex radio. In general, full duplex communication is found to provide only modest throughput gains over half...

  15. Duplex ultrasound and computed tomography angiography in the follow-up of endovascular abdominal aortic aneurysm repair: a comparative study*

    Science.gov (United States)

    Cantador, Alex Aparecido; Siqueira, Daniel Emílio Dalledone; Jacobsen, Octavio Barcellos; Baracat, Jamal; Pereira, Ines Minniti Rodrigues; Menezes, Fábio Hüsemann; Guillaumon, Ana Terezinha

    2016-01-01

    Objective To compare duplex ultrasound and computed tomography (CT) angiography in terms of their performance in detecting endoleaks, as well as in determining the diameter of the aneurysm sac, in the postoperative follow-up of endovascular abdominal aortic aneurysm repair. Materials and Methods This was a prospective study involving 30 patients who had undergone endovascular repair of infrarenal aortoiliac aneurysms. Duplex ultrasound and CT angiography were performed simultaneously by independent radiologists. Measurements of the aneurysm sac diameter were assessed, and the presence or absence of endoleaks was determined. Results The average diameter of the aneurysm sac, as determined by duplex ultrasound and CT angiography was 6.09 ± 1.95 and 6.27 ± 2.16 cm, respectively. Pearson's correlation coefficient showing a statistically significant correlation (R = 0.88; p < 0.01). Comparing the duplex ultrasound and CT angiography results regarding the detection of endoleaks, we found that the former had a negative predictive value of 92.59% and a specificity of 96.15%. Conclusion Our results show that there is little variation between the two methods evaluated, and that the choice between the two would have no significant effect on clinical management. Duplex ultrasound could replace CT angiography in the postoperative follow-up of endovascular aneurysm repair of the infrarenal aorta, because it is a low-cost procedure without the potential clinical complications related to the use of iodinated contrast and exposure to radiation. PMID:27777476

  16. Duplex ultrasound and computed tomography angiography in the follow-up of endovascular abdominal aortic aneurysm repair: a comparative study

    Energy Technology Data Exchange (ETDEWEB)

    Cantador, Alex Aparecido; Siqueira, Daniel Emilio Dalledone; Jacobsen, Octavio Barcellos; Baracat, Jamal; Pereira, Ines Minniti Rodrigues; Menezes, Fabio Hüsemann; Guillaumon, Ana Terezinha, E-mail: alex_cantador@yahoo.com.br [Universidade Estadual de Campinas (FCM/UNICAMP), Campinas, SP (Brazil). Faculdade de Ciencias Medicas

    2016-07-15

    Objective: To compare duplex ultrasound and computed tomography (CT) angiography in terms of their performance in detecting endoleaks, as well as in determining the diameter of the aneurysm sac, in the postoperative follow-up of endovascular abdominal aortic aneurysm repair. Materials and Methods: This was a prospective study involving 30 patients who had undergone endovascular repair of infrarenal aortoiliac aneurysms. Duplex ultrasound and CT angiography were performed simultaneously by independent radiologists. Measurements of the aneurysm sac diameter were assessed, and the presence or absence of endoleaks was determined. Results: The average diameter of the aneurysm sac, as determined by duplex ultrasound and CT angiography was 6.09 ± 1.95 and 6.27 ± 2.16 cm, respectively. Pearson's correlation coefficient showing a statistically significant correlation (R = 0.88; p < 0.01). Comparing the duplex ultrasound and CT angiography results regarding the detection of endoleaks, we found that the former had a negative predictive value of 92.59% and a specificity of 96.15%. Conclusion: Our results show that there is little variation between the two methods evaluated, and that the choice between the two would have no significant effect on clinical management. Duplex ultrasound could replace CT angiography in the postoperative follow-up of endovascular aneurysm repair of the infrarenal aorta, because it is a low-cost procedure without the potential clinical complications related to the use of iodinated contrast and exposure to radiation. (author)

  17. Duplex ultrasound and computed tomography angiography in the follow-up of endovascular abdominal aortic aneurysm repair: a comparative study

    Directory of Open Access Journals (Sweden)

    Alex Aparecido Cantador

    Full Text Available Abstract Objective: To compare duplex ultrasound and computed tomography (CT angiography in terms of their performance in detecting endoleaks, as well as in determining the diameter of the aneurysm sac, in the postoperative follow-up of endovascular abdominal aortic aneurysm repair. Materials and Methods: This was a prospective study involving 30 patients who had undergone endovascular repair of infrarenal aortoiliac aneurysms. Duplex ultrasound and CT angiography were performed simultaneously by independent radiologists. Measurements of the aneurysm sac diameter were assessed, and the presence or absence of endoleaks was determined. Results: The average diameter of the aneurysm sac, as determined by duplex ultrasound and CT angiography was 6.09 ± 1.95 and 6.27 ± 2.16 cm, respectively. Pearson's correlation coefficient showing a statistically significant correlation (R = 0.88; p < 0.01. Comparing the duplex ultrasound and CT angiography results regarding the detection of endoleaks, we found that the former had a negative predictive value of 92.59% and a specificity of 96.15%. Conclusion: Our results show that there is little variation between the two methods evaluated, and that the choice between the two would have no significant effect on clinical management. Duplex ultrasound could replace CT angiography in the postoperative follow-up of endovascular aneurysm repair of the infrarenal aorta, because it is a low-cost procedure without the potential clinical complications related to the use of iodinated contrast and exposure to radiation.

  18. A model of H-NS mediated compaction of bacterial DNA

    CERN Document Server

    Joyeux, Marc; 10.1016/j.bpj.2013.02.043

    2013-01-01

    The Histone-like Nucleoid Structuring protein (H-NS) is a nucleoid-associated protein, which is involved in both gene regulation and DNA compaction. H-NS can bind to DNA in two different ways: in trans, by binding to two separate DNA duplexes, or in cis, by binding to different sites on the same duplex. Based on scanning force microscopy imaging and optical trap-driven unzipping assays, it has recently been suggested that DNA compaction may result from the antagonistic effects of H-NS binding to DNA in trans and cis configurations. In order to get more insight into the compaction mechanism, we constructed a coarse-grained model description of the compaction of bacterial DNA by H-NS. These simulations highlight the fact that DNA compaction indeed results from the subtle equilibrium between several competing factors, which include the deformation dynamics of the plasmid and the several binding modes of protein dimers to DNA, i.e. dangling configurations, cis- and trans-binding. In particular, the degree of comp...

  19. Protein-DNA chimeras: synthesis of two-arm chimeras and non-mechanical effects of the DNA spring

    Energy Technology Data Exchange (ETDEWEB)

    Wang Yong; Wang, Andrew; Qu Hao; Zocchi, Giovanni, E-mail: zocchi@physics.ucla.ed [Department of Physics and Astronomy, University of California Los Angeles, Los Angeles, CA 90095-1547 (United States)

    2009-08-19

    DNA molecular springs have recently been used to control the activity of enzymes and ribozymes. In this approach, the mechanical stress exerted by the molecular spring alters the enzyme's conformation and thus the enzymatic activity. Here we describe a method alternative to our previous one to attach DNA molecular springs to proteins, where two separate DNA 'arms' are coupled to the protein and subsequently ligated. We report certain non-mechanical effects associated with the DNA spring observed in some chimeras with specific DNA sequences and the nucleotide binding enzyme guanylate kinase. If a ssDNA 'arm' is attached to the protein by one end only, we find that in some cases (depending on the DNA sequence and attachment point on the protein's surface) the unhybridized DNA arm inhibits the enzyme, while hybridization of the DNA arm leads to an apparent activation of the enzyme. One interpretation is that, in these cases, hybridization of the DNA arm removes it from the vicinity of the active site of the enzyme. We show how mechanical and non-mechanical effects of the DNA spring can be distinguished. This is important if one wants to use the protein-DNA chimeras to quantitatively study the response of the enzyme to mechanical perturbations.

  20. Effect of DNA groove binder distamycin A upon chromatin structure.

    Directory of Open Access Journals (Sweden)

    Parijat Majumder

    Full Text Available BACKGROUND: Distamycin A is a prototype minor groove binder, which binds to B-form DNA, preferentially at A/T rich sites. Extensive work in the past few decades has characterized the binding at the level of double stranded DNA. However, effect of the same on physiological DNA, i.e. DNA complexed in chromatin, has not been well studied. Here we elucidate from a structural perspective, the interaction of distamycin with soluble chromatin, isolated from Sprague-Dawley rat. METHODOLOGY/PRINCIPAL FINDINGS: Chromatin is a hierarchical assemblage of DNA and protein. Therefore, in order to characterize the interaction of the same with distamycin, we have classified the system into various levels, according to the requirements of the method adopted, and the information to be obtained. Isothermal titration calorimetry has been employed to characterize the binding at the levels of chromatin, chromatosome and chromosomal DNA. Thermodynamic parameters obtained thereof, identify enthalpy as the driving force for the association, with comparable binding affinity and free energy for chromatin and chromosomal DNA. Reaction enthalpies at different temperatures were utilized to evaluate the change in specific heat capacity (ΔCp, which, in turn, indicated a possible binding associated structural change. Ligand induced structural alterations have been monitored by two complementary methods--dynamic light scattering, and transmission electron microscopy. They indicate compaction of chromatin. Using transmission electron microscopy, we have visualized the effect of distamycin upon chromatin architecture at di- and trinucleosome levels. Our results elucidate the simultaneous involvement of linker bending and internucleosomal angle contraction in compaction process induced by distamycin. CONCLUSIONS/SIGNIFICANCE: We summarize here, for the first time, the thermodynamic parameters for the interaction of distamycin with soluble chromatin, and elucidate its effect on

  1. The Role of Entropic Effects on DNA Loop Formation

    Science.gov (United States)

    Wilson, David; Tkachenko, Alexei; Lillian, Todd; Perkins, Noel; Meiners, Jens Christian

    2009-03-01

    The formation of protein mediated DNA loops often regulates gene expression. Typically, a protein is simultaneously bound to two DNA operator sites. An example is the lactose repressor which binds to the Lac operon of E. coli. We characterize the mechanics of this system by calculating the free energy cost of loop formation. We construct a Hamiltonian that describes the change in DNA bending energy due to linear perturbations about the looped and open states, starting from a non-linear mechanical rod model that determines the shape and bending energy of the inter-operator DNA loop while capturing the intrinsic curvature and sequence-dependent elasticity of the DNA. The crystal structure of the LacI protein provides the boundary conditions for the DNA. We then calculate normal modes of the open and closed loops to account for the thermal fluctuations. The ratio of determinants of the two Hamiltonians yields the partition function, and the enthalphic and entropic cost of looping. This calculation goes beyond standard elastic energy models because it fully accounts for the substantial entropic differences between the two states. It also includes effects of sequence dependent curvature and stiffness and allows anisotropic variations in persistence length. From the free energy we then calculate the J-factor and ratio of loop lifetimes.

  2. Effects of N[superscript 2],N[superscript 2]-dimethylguanosine on RNA structure and stability: Crystal structure of an RNA duplex with tandem m[superscript 2 subscript 2]G:A pairs

    Energy Technology Data Exchange (ETDEWEB)

    Pallan, Pradeep S.; Kreutz, Christoph; Bosio, Silvia; Micura, Ronald; Egli, Martin (Innsbruck University); (Vanderbilt)

    2008-12-15

    Methylation of the exocyclic amino group of guanine is a relatively common modification in rRNA and tRNA. Single methylation (N(2)-methylguanosine, m(2)G) is the second most frequently encountered nucleoside analog in Escherichia coli rRNAs. The most prominent case of dual methylation (N(2),N(2)-dimethylguanosine, m(2) (2)G) is found in the majority of eukaryotic tRNAs at base pair m(2) (2)G26:A44. The latter modification eliminates the ability of the N(2) function to donate in hydrogen bonds and alters its pairing behavior, notably vis-a-vis C. Perhaps a less obvious consequence of the N(2),N(2)-dimethyl modification is its role in controlling the pairing modes between G and A. We have determined the crystal structure of a 13-mer RNA duplex with central tandem m(2) (2)G:A pairs. In the structure both pairs adopt an imino-hydrogen bonded, pseudo-Watson-Crick conformation. Thus, the sheared conformation frequently seen in tandem G:A pairs is avoided due to a potential steric clash between an N(2)-methyl group and the major groove edge of A. Additionally, for a series of G:A containing self-complementary RNAs we investigated how methylation affects competitive hairpin versus duplex formation based on UV melting profile analysis.

  3. A single-tube duplex and multiplex PCR for simultaneous detection of four cassava mosaic begomovirus species in cassava plants.

    Science.gov (United States)

    Aloyce, R C; Tairo, F; Sseruwagi, P; Rey, M E C; Ndunguru, J

    2013-04-01

    A single-tube duplex and multiplex PCR was developed for the simultaneous detection of African cassava mosaic virus (ACMV), East African cassava mosaic Cameroon virus (EACMCV), East African cassava mosaic Malawi virus (EACMMV) and East African cassava mosaic Zanzibar virus (EACMZV), four cassava mosaic begomoviruses (CMBs) affecting cassava in sub-Saharan Africa. Co-occurrence of the CMBs in cassava synergistically enhances disease symptoms and complicates their detection and diagnostics. Four primer pairs were designed to target DNA-A component sequences of cassava begomoviruses in a single tube PCR amplification using DNA extracted from dry-stored cassava leaves. Duplex and multiplex PCR enabled the simultaneous detection and differentiation of the four CMBs, namely ACMV (940bp), EACMCV (435bp), EACMMV (504bp) and EACMZV (260bp) in single and mixed infections, and sequencing results confirmed virus identities according to the respective published sequences of begomovirus species. In addition, we report here a modified Dellapotra et al. (1983) protocol, which was used to extract DNA from dry and fresh cassava leaves with comparable results. Using the duplex and multiplex techniques, time was saved and amount of reagents used were reduced, which translated into reduced cost of the diagnostics. This tool can be used by cassava breeders screening for disease resistance; scientists doing virus diagnostic studies; phytosanitary officers checking movement of diseased planting materials, and seed certification and multipliers for virus indexing.

  4. Aminomethylene peptide nucleic acid (am-PNA): synthesis, regio-/stereospecific DNA binding, and differential cell uptake of (α/γ,R/S)am-PNA analogues.

    Science.gov (United States)

    Mitra, Roopa; Ganesh, Krishna N

    2012-07-06

    Inherently chiral, cationic am-PNAs having pendant aminomethylene groups at α(R/S) or γ(S) sites on PNA backbone have been synthesized. The modified PNAs are shown to stabilize duplexes with complementary cDNA in a regio- and stereo-preferred manner with γ(S)-am PNA superior to α(R/S)-am PNAs and α(R)-am PNA better than the α(S) isomer. The enhanced stabilization of am-PNA:DNA duplexes is accompanied by a greater discrimination of mismatched bases. This seems to be a combined result of both electrostatic interactions and conformational preorganization of backbone favoring the cDNA binding. The am-PNAs are demonstrated to effectively traverse the cell membrane, localize in the nucleus of HeLa cells, and exhibit low toxicity to cells.

  5. Antiproliferative effects of DNA methyltransferase 3B depletion are not associated with DNA demethylation.

    Directory of Open Access Journals (Sweden)

    Sabine Hagemann

    Full Text Available Silencing of genes by hypermethylation contributes to cancer progression and has been shown to occur with increased frequency at specific genomic loci. However, the precise mechanisms underlying the establishment and maintenance of aberrant methylation marks are still elusive. The de novo DNA methyltransferase 3B (DNMT3B has been suggested to play an important role in the generation of cancer-specific methylation patterns. Previous studies have shown that a reduction of DNMT3B protein levels induces antiproliferative effects in cancer cells that were attributed to the demethylation and reactivation of tumor suppressor genes. However, methylation changes have not been analyzed in detail yet. Using RNA interference we reduced DNMT3B protein levels in colon cancer cell lines. Our results confirm that depletion of DNMT3B specifically reduced the proliferation rate of DNMT3B-overexpressing colon cancer cell lines. However, genome-scale DNA methylation profiling failed to reveal methylation changes at putative DNMT3B target genes, even in the complete absence of DNMT3B. These results show that DNMT3B is dispensable for the maintenance of aberrant DNA methylation patterns in human colon cancer cells and they have important implications for the development of targeted DNA methyltransferase inhibitors as epigenetic cancer drugs.

  6. Application of differential scanning calorimetry to measure the differential binding of ions, water and protons in the unfolding of DNA molecules.

    Science.gov (United States)

    Olsen, Chris M; Shikiya, Ronald; Ganugula, Rajkumar; Reiling-Steffensmeier, Calliste; Khutsishvili, Irine; Johnson, Sarah E; Marky, Luis A

    2016-05-01

    The overall stability of DNA molecules globally depends on base-pair stacking, base-pairing, polyelectrolyte effect and hydration contributions. In order to understand how they carry out their biological roles, it is essential to have a complete physical description of how the folding of nucleic acids takes place, including their ion and water binding. To investigate the role of ions, water and protons in the stability and melting behavior of DNA structures, we report here an experimental approach i.e., mainly differential scanning calorimetry (DSC), to determine linking numbers: the differential binding of ions (Δnion), water (ΔnW) and protons (ΔnH(+)) in the helix-coil transition of DNA molecules. We use DSC and temperature-dependent UV spectroscopic techniques to measure the differential binding of ions, water, and protons for the unfolding of a variety of DNA molecules: salmon testes DNA (ST-DNA), one dodecamer, one undecamer and one decamer duplexes, nine hairpin loops, and two triplexes. These methods can be applied to any conformational transition of a biomolecule. We determined complete thermodynamic profiles, including all three linking numbers, for the unfolding of each molecule. The favorable folding of a DNA helix results from a favorable enthalpy-unfavorable entropy compensation. DSC thermograms and UV melts as a function of salt, osmolyte and proton concentrations yielded releases of ions and water. Therefore, the favorable folding of each DNA molecule results from the formation of base-pair stacks and uptake of both counterions and water molecules. In addition, the triplex with C(+)GC base triplets yielded an uptake of protons. Furthermore, the folding of a DNA duplex is accompanied by a lower uptake of ions and a similar uptake of four water molecules as the DNA helix gets shorter. In addition, the oligomer duplexes and hairpin thermodynamic data suggest ion and water binding depends on the DNA sequence rather than DNA composition. Copyright

  7. Highly stable triple helix formation by homopyrimidine (l)-acyclic threoninol nucleic acids with single stranded DNA and RNA

    DEFF Research Database (Denmark)

    Kumar, Vipin; Kesavan, Venkitasamy; Gothelf, Kurt Vesterager

    2015-01-01

    or RNA, and these triplexes are significantly stronger than the corresponding DNA or RNA duplexes as shown in competition experiments. As a unique property the (l)-aTNAs exclusively form triplex structures with DNA and RNA and no duplex structures are observed by gel electrophoresis. The results were...

  8. Experimental Study of the Thermal Diffusivity and Heat Capacity Concerning Some Duplex Stainless Steel

    Directory of Open Access Journals (Sweden)

    Riad Harwill Abdul Abas

    2015-06-01

    Full Text Available In the present work, thermal diffusivity and heat capacity measurements have been investigated in temperature range between RT and 1473 K for different duplex stainless steel supplied by Outokumpu Stainless AB, Sweden. The purpose of this study is to get a reliable thermophysical data of these alloys and to study the effect of microstructure on the thermal diffusivity and heat capacity value. Results show the ferrite content in the duplex stainless steel increased with temperature at equilibrium state. On the other hand, ferrite content increased with increasing Cr/Ni ratio and there is no significant effect of ferrite content on the thermal diffusivity value at room temperature. Furthermore, the heat capacity of all samples increases with temperature from room temperature to 473 K, while it decreases with increasing temperature until 1073 K. Then it increases with temperature at higher temperature. Curie temperature and sigma phase formation temperature can be detected by heat capacity-temperature curves.

  9. Helix-coil transition of the self-complementary dG-dG-dA-dA-dT-dT-dC-dC duplex.

    Science.gov (United States)

    Patel, D J; Canuel, L L

    1979-05-15

    The helix-coil transition of the octanucleotide self-complementary duplex dG-dG-dA-dA-dT-dT-dC-dC has been monitored at the Watson-Crick protons, the base and sugar nonexchangeable protons and the backbone phosphates by high-resolution nuclear magnetic resonance (NMR) spectroscopy. The melting transition of the octanucleotide monitored by ultraviolet absorbance spectroscopy is characterized by the thermodynamic parameters delta H degree = -216.7 kJ/mol and delta S degree (25 degrees C) = -0.632 KJ mol-1 K-1 in 0.1 M NaCl, 10 mM phosphate solution. Correlation of the transition midpoint values monitored by the ultraviolet absorbance studies at strand concentrations below 0.2 mM and by NMR studies at 5.3 mM suggest that both methods are monitoring the octanucleotide duplex-to-strand transition. The NMR spectra of the Watson-Crick ring NH protons of the octanucleotide duplex have been followed as a function of temperature. The resonance from the terminal dG.dC base pairs broadens out at room temperature while the resonances from the other base pairs broaden simultaneously with the onset of the melting transition. The nonexchangeable base and sugar H-1' protons are resolved in the duplex and strand states and shift as average peaks through the melting transition. The experimental shifts on duplex formation have been compared with calculated values based on ring-current and atomic diamagnetic anisotropy contributions for a B-DNA base-pair-overlap geometry in solution. Several nonexchangeable proton resonances broaden in the fast-exchange region during the duplex-to-strand transition and the excess widths yield a duplex dissociation rate constant for the octanucleotide of 1.9 x 10(3) s-1 at 32 degrees C (fraction of duplex = 0.86) in 0.1 M NaCl, 10 mM phosphate buffer. The 31P resonances of the seven internucleotide phosphates are distributed over 0.6 ppm in the duplex state, shift downfield during the duplex-to-strand transition and undergo additional downfield shifts

  10. Magnetic resonance urography in duplex kidney with ectopic ureteral insertion

    Science.gov (United States)

    Thambidorai, Conjeevaram Rajendrarao; Anuar, Zulfiqar

    2011-01-01

    This is a report on the use of magnetic resonance urography (MRU) in a 6-year-old girl who presented with urinary incontinence. She had a left duplex kidney with poorly functioning upper moiety and ectopic insertion of the dilated upper pole ureter. MRU has been shown to be superior to conventional imaging techniques in delineating poorly functioning moieties of duplex kidneys and ectopic ureters. PMID:21897576

  11. Magnetic resonance urography in duplex kidney with ectopic ureteral insertion

    Directory of Open Access Journals (Sweden)

    Conjeevaram Rajendrarao Thambidorai

    2011-01-01

    Full Text Available This is a report on the use of magnetic resonance urography (MRU in a 6-year-old girl who presented with urinary incontinence. She had a left duplex kidney with poorly functioning upper moiety and ectopic insertion of the dilated upper pole ureter. MRU has been shown to be superior to conventional imaging techniques in delineating poorly functioning moieties of duplex kidneys and ectopic ureters.

  12. Properties of duplex stainless steels made by powder metallurgy

    OpenAIRE

    Rosso, M.; M. Actis Grande; Z. Brytan; L.A. Dobrzański

    2007-01-01

    Purpose: of this paper was to examine the mechanical properties of duplex stainless steels.Design/methodology/approach: In presented study duplex stainless steels were obtained through powder metallurgy starting from austenitic, martensitic base powders by controlled addition of alloying elements, such as Cr, Ni, Mo and Cu. In the studies behind the preparation of mixes, Schaeffler’s diagram was taken into consideration. Prepared mixes have been sintered in a vacuu...

  13. Rapid assessment of the effect of ciprofloxacin on chromosomal DNA from Escherichia coli using an in situ DNA fragmentation assay

    Directory of Open Access Journals (Sweden)

    Gosalvez Jaime

    2009-04-01

    Full Text Available Abstract Background Fluoroquinolones are extensively used antibiotics that induce DNA double-strand breaks (DSBs by trapping DNA gyrase and topoisomerase IV on DNA. This effect is usually evaluated using biochemical or molecular procedures, but these are not effective at the single-cell level. We assessed ciprofloxacin (CIP-induced chromosomal DNA breakage in single-cell Escherichia coli by direct visualization of the DNA fragments that diffused from the nucleoid obtained after bacterial lysis in an agarose microgel on a slide. Results Exposing the E. coli strain TG1 to CIP starting at a minimum inhibitory concentration (MIC of 0.012 μg/ml and at increasing doses for 40 min increased the DNA fragmentation progressively. DNA damage started to be detectable at the MIC dose. At a dose of 1 μg/ml of CIP, DNA damage was visualized clearly immediately after processing, and the DNA fragmentation increased progressively with the antibiotic incubation time. The level of DNA damage was much higher when the bacteria were taken from liquid LB broth than from solid LB agar. CIP treatment produced a progressively slower rate of DNA damage in bacteria in the stationary phase than in the exponentially growing phase. Removing the antibiotic after the 40 min incubation resulted in progressive DSB repair activity with time. The magnitude of DNA repair was inversely related to CIP dose and was noticeable after incubation with CIP at 0.1 μg/ml but scarce after 10 μg/ml. The repair activity was not strictly related to viability. Four E. coli strains with identified mechanisms of reduced sensitivity to CIP were assessed using this procedure and produced DNA fragmentation levels that were inversely related to MIC dose, except those with very high MIC dose. Conclusion This procedure for determining DNA fragmentation is a simple and rapid test for studying and evaluating the effect of quinolones.

  14. Hammerhead ribozyme activity and oligonucleotide duplex stability in mixed solutions of water and organic compounds

    Directory of Open Access Journals (Sweden)

    Shu-ichi Nakano

    2014-01-01

    Full Text Available Nucleic acids are useful for biomedical targeting and sensing applications in which the molecular environment is different from that of a dilute aqueous solution. In this study, the influence of various types of mixed solutions of water and water-soluble organic compounds on RNA was investigated by measuring the catalytic activity of the hammerhead ribozyme and the thermodynamic stability of an oligonucleotide duplex. The compounds with a net neutral charge, such as poly(ethylene glycol, small primary alcohols, amide compounds, and aprotic solvent molecules, added at high concentrations changed the ribozyme-catalyzed RNA cleavage rate, with the magnitude of the effect dependent on the NaCl concentration. These compounds also changed the thermodynamic stability of RNA base pairs of an oligonucleotide duplex and its dependence on the NaCl concentration. Specific interactions with RNA molecules and reduced water activity could account for the inhibiting effects on the ribozyme catalysis and destabilizing effects on the duplex stability. The salt concentration dependence data correlated with the dielectric constant, but not with water activity, viscosity, and the size of organic compounds. This observation suggests the significance of the dielectric constant effects on the RNA reactions under molecular crowding conditions created by organic compounds.

  15. Superparamagnetic nanoparticles for effective delivery of malaria DNA vaccine.

    Science.gov (United States)

    Al-Deen, Fatin Nawwab; Ho, Jenny; Selomulya, Cordelia; Ma, Charles; Coppel, Ross

    2011-04-05

    Low efficiency is often observed in the delivery of DNA vaccines. The use of superparamagnetic nanoparticles (SPIONs) to deliver genes via magnetofection could improve transfection efficiency and target the vector to its desired locality. Here, magnetofection was used to enhance the delivery of a malaria DNA vaccine encoding Plasmodium yoelii merozoite surface protein MSP1(19) (VR1020-PyMSP1(19)) that plays a critical role in Plasmodium immunity. The plasmid DNA (pDNA) containing membrane associated 19-kDa carboxyl-terminal fragment of merozoite surface protein 1 (PyMSP1(19)) was conjugated with superparamagnetic nanoparticles coated with polyethyleneimine (PEI) polymer, with different molar ratio of PEI nitrogen to DNA phosphate. We reported the effects of SPIONs-PEI complexation pH values on the properties of the resulting particles, including their ability to condense DNA and the gene expression in vitro. By initially lowering the pH value of SPIONs-PEI complexes to 2.0, the size of the complexes decreased since PEI contained a large number of amino groups that became increasingly protonated under acidic condition, with the electrostatic repulsion inducing less aggregation. Further reaggregation was prevented when the pHs of the complexes were increased to 4.0 and 7.0, respectively, before DNA addition. SPIONs/PEI complexes at pH 4.0 showed better binding capability with PyMSP1(19) gene-containing pDNA than those at neutral pH, despite the negligible differences in the size and surface charge of the complexes. This study indicated that the ability to protect DNA molecules due to the structure of the polymer at acidic pH could help improve the transfection efficiency. The transfection efficiency of magnetic nanoparticle as carrier for malaria DNA vaccine in vitro into eukaryotic cells, as indicated via PyMSP1(19) expression, was significantly enhanced under the application of external magnetic field, while the cytotoxicity was comparable to the benchmark nonviral

  16. Effective DNA Inhibitors of Cathepsin G by In Vitro Selection

    Directory of Open Access Journals (Sweden)

    Manlio Palumbo

    2008-06-01

    Full Text Available Cathepsin G (CatG is a chymotrypsin-like protease released upon degranulation of neutrophils. In several inflammatory and ischaemic diseases the impaired balance between CatG and its physiological inhibitors leads to tissue destruction and platelet aggregation. Inhibitors of CatG are suitable for the treatment of inflammatory diseases and procoagulant conditions. DNA released upon the death of neutrophils at injury sites binds CatG. Moreover, short DNA fragments are more inhibitory than genomic DNA. Defibrotide, a single stranded polydeoxyribonucleotide with antithrombotic effect is also a potent CatG inhibitor. Given the above experimental evidences we employed a selection protocol to assess whether DNA inhibition of CatG may be ascribed to specific sequences present in defibrotide DNA. A Selex protocol was applied to identify the single-stranded DNA sequences exhibiting the highest affinity for CatG, the diversity of a combinatorial pool of oligodeoxyribonucleotides being a good representation of the complexity found in defibrotide. Biophysical and biochemical studies confirmed that the selected sequences bind tightly to the target enzyme and also efficiently inhibit its catalytic activity. Sequence analysis carried out to unveil a motif responsible for CatG recognition showed a recurrence of alternating TG repeats in the selected CatG binders, adopting an extended conformation that grants maximal interaction with the highly charged protein surface. This unprecedented finding is validated by our results showing high affinity and inhibition of CatG by specific DNA sequences of variable length designed to maximally reduce pairing/folding interactions.

  17. Carbon Nanotube Field-Effect Transistor for DNA Sensing

    Science.gov (United States)

    Xuan, Chu T.; Thuy, Nguyen T.; Luyen, Tran T.; Huyen, Tran T. T.; Tuan, Mai A.

    2017-01-01

    A field-effect transistor (FET) using carbon nanotubes (CNTs) as the conducting channel (CNTFET) has been developed, designed such that the CNT conducting channel (15 μm long, 700 μm wide) is directly exposed to medium containing target deoxyribonucleic acid (DNA). The CNTFET operates at high ON-current of 1.91 μA, ON/OFF-current ratio of 1.2 × 105, conductance of 4.3 μS, and leakage current of 16.4 pA. We present initial trials showing the response of the CNTFET to injection of target DNA into aqueous medium.

  18. A duplex real-time PCR assay for the quantitative detection of Naegleria fowleri in water samples.

    Science.gov (United States)

    Behets, Jonas; Declerck, Priscilla; Delaedt, Yasmine; Verelst, Lieve; Ollevier, Frans

    2007-01-01

    A fast and accurate duplex real-time PCR (qPCR) was developed to detect and quantify the human pathogenic amoeba Naegleria fowleri in water samples. In this study, primers and probe based on the Mp2Cl5 gene were designed to amplify and quantify N. fowleri DNA in a single duplex reaction. The qPCR detection limit (DL) corresponds to the minimum DNA quantity showing significant fluorescence with at least 90% of the positive controls in a duplex reaction. Using fluorescent Taqman technology the qPCR was found to be 100% specific for N. fowleri with a DL of 3 N. fowleri cell equivalents and a PCR efficiency of 99%. The quantification limit (QL) was 16 N. fowleri cell equivalents (corresponded with 320 N. fowleri cell equivalents l(-1) water sample) in a duplex qPCR reaction and corresponds to the lowest DNA quantity amplifiable with a coefficient of variation less than 25%. To detect inhibition an exogenous internal positive control (IPC) was included in each PCR reaction preventing false negative results. Comparison of qPCR and most probable number (MPN) culture results confirms that the developed qPCR is well suited for rapid and quantitative detection of this human pathogen in real water samples. Nevertheless 'low contamination levels' of water samples (fowleri cells l(-1)) still require culture method analyses. When other thermophilic Naegleria are very dominant, the MPN culture method could result in an underestimation in the real number of N. fowleri and some caution is necessary to interpret the data. The N. fowleri qPCR could be a useful tool to study further competitive phenomena between thermophilic Naegleria strains.

  19. DNA Polymer Brush Patterning through Photocontrollable Surface-Initiated DNA Hybridization Chain Reaction.

    Science.gov (United States)

    Huang, Fujian; Zhou, Xiang; Yao, Dongbao; Xiao, Shiyan; Liang, Haojun

    2015-11-18

    The fabrication of DNA polymer brushes with spatial resolution onto a solid surface is a crucial step for biochip research and related applications, cell-free gene expression study, and even artificial cell fabrication. Here, for the first time, a DNA polymer brush patterning method is reported based on the photoactivation of an ortho-nitrobenzyl linker-embedded DNA hairpin structure and a subsequent surface-initiated DNA hybridization chain reaction (HCR). Inert DNA hairpins are exposed to ultraviolet light irradiation to generate DNA duplexes with two active sticky ends (toeholds) in a programmable manner. These activated DNA duplexes can initiate DNA HCR to generate multifunctional patterned DNA polymer brushes with complex geometrical shapes. Different multifunctional DNA polymer brush patterns can be fabricated on certain areas of the same solid surface using this method. Moreover, the patterned DNA brush surface can be used to capture target molecules in a desired manner.

  20. Conformational Effects of UV Light on DNA Origami.

    Science.gov (United States)

    Chen, Haorong; Li, Ruixin; Li, Shiming; Andréasson, Joakim; Choi, Jong Hyun

    2017-02-01

    The responses of DNA origami conformation to UV radiation of different wavelengths and doses are investigated. Short- and medium-wavelength UV light can cause photo-lesions in DNA origami. At moderate doses, the lesions do not cause any visible defects in the origami, nor do they significantly affect the hybridization capability. Instead, they help relieve the internal stress in the origami structure and restore it to the designed conformation. At high doses, staple dissociation increases which causes structural disintegration. Long-wavelength UV does not show any effect on origami conformation by itself. We show that this UV range can be used in conjunction with photoactive molecules for photo-reconfiguration, while avoiding any damage to the DNA structures.

  1. Binding Isotherms and Cooperative Effects for Metal-DNA Complexes

    CERN Document Server

    Gelagutashvili, Eteri

    2008-01-01

    The stoichiometric binding constants of Nickel(II), Cobalt(II), Manganese(II), Silver(I), Zinc(II) ions with DNA, from Spirulina platensis were determined from their binding isotherms by equilibrium dialysis and atomic absorption spectroscopy. It was shown, that the nature of these ions interaction with DNA, from S .platensis is different. For Cobalt(II), Zinc(II) ions were observed cooperative effects and existence of two different types of the binding sites. Nickel(II)_, Silver(I) -DNA complexes shows independent and identical binding sites and Manganese(II)_ negative cooperative interaction. The logarithm of binding constants for Cobalt (II)_, Nickel (II)_, Manganese (II)_, Zinc (II)_, Silver (I) - DNA, from S. platensis in 3 mM Na(I) are 5.11; 5.18; 4.77; 5.05; 5.42; respectively. The linear correlation of logarithm of binding constants (for complexes of metal-DNA from S. platensis) and the covalent index of Pauling are observed.

  2. DNA

    Science.gov (United States)

    Stent, Gunther S.

    1970-01-01

    This history for molecular genetics and its explanation of DNA begins with an analysis of the Golden Jubilee essay papers, 1955. The paper ends stating that the higher nervous system is the one major frontier of biological inquiry which still offers some romance of research. (Author/VW)

  3. A silicon carbide nanowire field effect transistor for DNA detection.

    Science.gov (United States)

    Fradetal, L; Bano, E; Attolini, G; Rossi, F; Stambouli, V

    2016-06-10

    This work reports on the label-free electrical detection of DNA molecules for the first time, using silicon carbide (SiC) as a novel material for the realization of nanowire field effect transistors (NWFETs). SiC is a promising semiconductor for this application due to its specific characteristics such as chemical inertness and biocompatibility. Non-intentionally n-doped SiC NWs are first grown using a bottom-up vapor-liquid-solid (VLS) mechanism, leading to the NWs exhibiting needle-shaped morphology, with a length of approximately 2 μm and a diameter ranging from 25 to 60 nm. Then, the SiC NWFETs are fabricated and functionalized with DNA molecule probes via covalent coupling using an amino-terminated organosilane. The drain current versus drain voltage (I d-V d) characteristics obtained after the DNA grafting and hybridization are reported from the comparative and simultaneous measurements carried out on the SiC NWFETs, used either as sensors or references. As a representative result, the current of the sensor is lowered by 22% after probe DNA grafting and by 7% after target DNA hybridization, while the current of the reference does not vary by more than ±0.6%. The current decrease confirms the field effect induced by the negative charges of the DNA molecules. Moreover, the selectivity, reproducibility, reversibility and stability of the studied devices are emphasized by de-hybridization, non-complementary hybridization and re-hybridization experiments. This first proof of concept opens the way for future developments using SiC-NW-based sensors.

  4. Effect of dephasing on DNA sequencing via transverse electronic transport

    Energy Technology Data Exchange (ETDEWEB)

    Zwolak, Michael [Los Alamos National Laboratory; Krems, Matt [NON LANL; Pershin, Yuriy V [NON LANL; Di Ventra, Massimiliano [NON LANL

    2009-01-01

    We study theoretically the effects of dephasing on DNA sequencing in a nanopore via transverse electronic transport. To do this, we couple classical molecular dynamics simulations with transport calculations using scattering theory. Previous studies, which did not include dephasing, have shown that by measuring the transverse current of a particular base multiple times, one can get distributions of currents for each base that are distinguishable. We introduce a dephasing parameter into transport calculations to simulate the effects of the ions and other fluctuations. These effects lower the overall magnitude of the current, but have little effect on the current distributions themselves. The results of this work further implicate that distinguishing DNA bases via transverse electronic transport has potential as a sequencing tool.

  5. Corrosion behaviour of sintered duplex stainless steels

    Energy Technology Data Exchange (ETDEWEB)

    Utrilla, M. Victoria; Urena, Alejandro; Otero, Enrique; Munez, Claudio Jose [Escuela Superior de Ciencias Experimentales y Tecnologia, Universidad Rey Juan Carlos, C/ Tulipan s/n, 28933 Mostoles, Madrid (Spain)

    2004-07-01

    Duplex austenite-ferrite stainless steels were prepared by mixing austenitic (316L) and ferritic (434L) atomized powders. Although different 316L/434L ratios were prepared, present work centred its study on 50% ferrite - 50% austenite sintered steel. The powders were mixed and pressed at 700 MPa and sintered at 1250 deg. C for 30 min in vacuum. The cooling rate was 5 deg. C/min. Solution treatment was carried out to homogenize the microstructure at 1100 deg. C during 20 min. A microstructural study of the material in solution was performed, evaluating the microstructure, proportion and shape of porosity, and ferrite percentage. This last was measured by two methods, quantitative metallography and Fischer ferrito-metry. The materials were heat treated in the range of 700 to 1000 deg. C, for 10, 30 and 60 min and water quenched, to study the microstructural changes and the influence on the intergranular corrosion resistance. The method used to evaluate the sensitization to the intergranular corrosion was the electrochemical potentio-kinetic reactivation procedure (EPR). The test solution was 0.5 M H{sub 2}SO{sub 4} + 0,01 M KSCN at 30 deg. C. The criterion used to evaluate the sensitization was the ratio between the maximum reactivation density (Ir) and the maximum activation density (Ia). The results of the electrochemical tests were discussed in relation with the microstructures observed at the different heat treatments. (authors)

  6. Simplex and duplex event-specific analytical methods for functional biotech maize.

    Science.gov (United States)

    Lee, Seong-Hun; Kim, Su-Jeong; Yi, Bu-Young

    2009-08-26

    Analytical methods are very important in the control of genetically modified organism (GMO) labeling systems or living modified organism (LMO) management for biotech crops. Event-specific primers and probes were developed for qualitative and quantitative analysis for biotech maize event 3272 and LY 038 on the basis of the 3' flanking regions, respectively. The qualitative primers confirmed the specificity by a single PCR product and sensitivity to 0.05% as a limit of detection (LOD). Simplex and duplex quantitative methods were also developed using TaqMan real-time PCR. One synthetic plasmid was constructed from two taxon-specific DNA sequences of maize and two event-specific 3' flanking DNA sequences of event 3272 and LY 038 as reference molecules. In-house validation of the quantitative methods was performed using six levels of mixing samples, from 0.1 to 10.0%. As a result, the biases from the true value and the relative deviations were all within the range of +/-30%. Limits of quantitation (LOQs) of the quantitative methods were all 0.1% for simplex real-time PCRs of event 3272 and LY 038 and 0.5% for duplex real-time PCR of LY 038. This study reports that event-specific analytical methods were applicable for qualitative and quantitative analysis for biotech maize event 3272 and LY 038.

  7. Space Radiation Effects on Human Cells: Modeling DNA Breakage, DNA Damage Foci Distribution, Chromosomal Aberrations and Tissue Effects

    Science.gov (United States)

    Ponomarev, A. L.; Huff, J. L.; Cucinotta, F. A.

    2011-01-01

    Future long-tem space travel will face challenges from radiation concerns as the space environment poses health risk to humans in space from radiations with high biological efficiency and adverse post-flight long-term effects. Solar particles events may dramatically affect the crew performance, while Galactic Cosmic Rays will induce a chronic exposure to high-linear-energy-transfer (LET) particles. These types of radiation, not present on the ground level, can increase the probability of a fatal cancer later in astronaut life. No feasible shielding is possible from radiation in space, especially for the heavy ion component, as suggested solutions will require a dramatic increase in the mass of the mission. Our research group focuses on fundamental research and strategic analysis leading to better shielding design and to better understanding of the biological mechanisms of radiation damage. We present our recent effort to model DNA damage and tissue damage using computational models based on the physics of heavy ion radiation, DNA structure and DNA damage and repair in human cells. Our particular area of expertise include the clustered DNA damage from high-LET radiation, the visualization of DSBs (DNA double strand breaks) via DNA damage foci, image analysis and the statistics of the foci for different experimental situations, chromosomal aberration formation through DSB misrepair, the kinetics of DSB repair leading to a model-derived spectrum of chromosomal aberrations, and, finally, the simulation of human tissue and the pattern of apoptotic cell damage. This compendium of theoretical and experimental data sheds light on the complex nature of radiation interacting with human DNA, cells and tissues, which can lead to mutagenesis and carcinogenesis later in human life after the space mission.

  8. Cytotoxic and DNA-topoisomerase effects of lapachol amine derivatives and interactions with DNA

    Directory of Open Access Journals (Sweden)

    A. Esteves-Souza

    2007-10-01

    Full Text Available The cytotoxic activity of amino (3a-e, aza-1-antraquinone (4a-e lapachol derivatives against Ehrlich carcinoma and human K562 leukemia cells was investigated. Cell viability was determined using MTT assay, after 48 (Ehrlich or 96 h (K562 of culture, and vincristine (for K562 leukemia and quercetin (for Ehrlich carcinoma were used as positive controls. The results showed dose-dependent growth-inhibiting activities and that the amino derivatives were active against the assayed cells, whereas the 4a-e derivatives were not. The allylamine derivative 3a was the most active against Ehrlich carcinoma, with IC50 = 16.94 ± 1.25 µM, and against K562 leukemia, with IC50 = 14.11 ± 1.39 µM. The analogous lawsone derivative, 5a, was also active against Ehrlich carcinoma (IC50 = 23.89 ± 2.3 µM, although the 5d and 5e derivatives showed lower activity. The interaction between 3a-d and calf thymus DNA was investigated by fluorimetric titration and the results showed a hyperchromic effect indicating binding to DNA as presented of ethidium bromide, used as positive control. The inhibitory action on DNA-topoisomerase II-a was also evaluated by a relaxation assay of supercoiled DNA plasmid, and the etoposide (200 µM was used as positive control. Significant inhibitory activities were observed for 3a-d at 200 µM and a partial inhibitory action was observed for lapachol and methoxylapachol.

  9. Studies on single and duplex aging of metastable beta titanium alloy Ti–15V–3Cr–3Al–3Sn

    Energy Technology Data Exchange (ETDEWEB)

    Santhosh, R. [School of Advanced Sciences, VIT University, Vellore 632 014 (India); Geetha, M. [School of Mechanical and Building Sciences, VIT University, Vellore 632 014 (India); Saxena, V.K. [Defence Metallurgical Research Laboratory, Hyderabad 500 058 (India); Nageswararao, M., E-mail: m.nageswararao@vit.ac.in [School of Mechanical and Building Sciences, VIT University, Vellore 632 014 (India)

    2014-08-25

    Highlights: • Preaging at 300 °C for 10 h led to an accelerating effect on α precipitation in Ti–15–3 alloy. • Preaging at 250 °C for 24 h led to a finer size and higher density of α precipitates. • Precipitate free zones were seen in the microstructure on aging at 400 and 450 °C. • Duplex aging results in four to five fold higher HCF life compared to single aging. - Abstract: This paper presents the age hardening behavior of metastable beta titanium alloy Ti–15 V–3Cr–3Al–3Sn subjected to various single and duplex aging treatments. Single aging was carried out in the range 200–550 °C for times up to 150 h (h). For duplex aging first step was performed in two different ways: (i) 250 °C for 24 h (ii) 300 °C for 10 h; second step was carried in the range 350–500 °C, for times up to 150 h. The decomposition of alpha (α) phase during aging was monitored by hardness measurements, tensile testing, different microscopic techniques and X-ray diffraction. Duplex aging treatments resulted in higher hardness and strength compared to single aging. There was a good balance of tensile strength and ductility after two duplex aging treatments 300 °C/10 h + 500 °C/10 h and 250 °C/24 h + 500 °C/8 h. Precipitate free zones (PFZs) were observed when aged at ⩽450 °C in the microstructure; aging at ⩾500 °C gave freedom from such zones. Duplex aging resulted in a finer size and higher density of α precipitates compared to single aging. Duplex aging 250 °C/24 h + 500 °C/8 h led to a four to five fold improvement in high cycle fatigue (HCF) life compared to single aging at 500 °C/10 h. The observed improvement in HCF life could thus be related to smaller size and larger density of α precipitates resulting from duplex aging. Optical microscopy and X-ray diffraction studies clearly revealed that preaging for 10 h at 300 °C accelerated the α-phase precipitation reaction during aging.

  10. Direct Electrical Detection of DNA Hybridization Based on Electrolyte-Gated Graphene Field-Effect Transistor

    Science.gov (United States)

    Ohno, Yasuhide; Okamoto, Shogo; Maehashi, Kenzo; Matsumoto, Kazuhiko

    2013-11-01

    DNA hybridization was electrically detected by graphene field-effect transistors. Probe DNA was modified on the graphene channel by a pyrene-based linker material. The transfer characteristic was shifted by the negative charges on the probe DNA, and the drain current was changed by the full-complementary DNA while no current change was observed after adding noncomplementary DNA, indicating that the graphene field-effect transistor detected the DNA hybridization. In addition, the number of DNAs was estimated by the simple plate capacitor model. As a result, one probe DNA was attached on the graphene channel per 10×10 nm2, indicating their high density functionalization. We estimated that 30% of probe DNA on the graphene channel was hybridized with 200 nM full-complementary DNA while only 5% of probe DNA was bound to the noncomplementary DNA. These results will help to pave the way for future biosensing applications based on graphene FETs.

  11. Duplex and triplex formation of mixed pyrimidine oligonucleotides with stacking of phenyl-triazole moieties in the major groove

    DEFF Research Database (Denmark)

    Andersen, Nicolai Krog; Døssing, Holger Balslev; Jensen, Frank;

    2011-01-01

    5-(1-Phenyl-1,2,3-triazol-4-yl)-2'-deoxycytidine was synthesized from a modified CuAAC protocol and incorporated into mixed pyrimidine oligonucleotide sequences together with the corresponding 5-(1-phenyl-1,2,3-triazol-4-yl)-2'-deoxyuridine. With consecutive incorporations of the two modified...... nucleosides, improved duplex formation with a complementary RNA and improved triplex formation with a complementary DNA duplex were observed. The improvement is due to π-π stacking of the phenyl-triazole moieties in the major groove. The strongest stacking and most pronounced positive influence on thermal...... stability was found in between the uridine analogues or with the cytidine analogue placed in the 3' direction to the uridine analogue. Modeling indicated a different orientation of the phenyl-triazole moieties in the major groove to account for the difference between the two nucleotides. The modified...

  12. Duplex real-time PCR for rapid simultaneous detection of Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans in Amphibian samples.

    Science.gov (United States)

    Blooi, M; Pasmans, F; Longcore, J E; Spitzen-van der Sluijs, A; Vercammen, F; Martel, A

    2013-12-01

    Chytridiomycosis is a lethal fungal disease contributing to declines and extinctions of amphibian species worldwide. The currently used molecular screening tests for chytridiomycosis fail to detect the recently described species Batrachochytrium salamandrivorans. In this study, we present a duplex real-time PCR that allows the simultaneous detection of B. salamandrivorans and Batrachochytrium dendrobatidis. With B. dendrobatidis- and B. salamandrivorans-specific primers and probes, detection of the two pathogens in amphibian samples is possible, with a detection limit of 0.1 genomic equivalent of zoospores of both pathogens per PCR. The developed real-time PCR shows high degrees of specificity and sensitivity, high linear correlations (r(2) > 0.995), and high amplification efficiencies (>94%) for B. dendrobatidis and B. salamandrivorans. In conclusion, the described duplex real-time PCR can be used to detect DNA of B. dendrobatidis and B. salamandrivorans with highly reproducible and reliable results.

  13. EPIGENETIC EFFECTS OF SHIFTWORK ON BLOOD DNA METHYLATION

    Science.gov (United States)

    Bollati, Valentina; Baccarelli, Andrea; Sartori, Samantha; Tarantini, Letizia; Motta, Valeria; Rota, Federica; Costa, Giovanni

    2012-01-01

    In the present study, the authors investigated the effects of shiftwork exposure on DNA methylation using peripheral blood DNA from subjects working in two chemical plants in Northern Italy. The investigation was designed to evaluate (a) DNA methyl- ation changes in Alu and long interspersed nuclear element-1 (LINE-1) repetitive elements as a surrogate of global methylation and (b) promoter methylation of gluco- corticoid receptor (GCR), tumor necrosis factor alpha (TNF-α), and interferon- gamma (IFN-γ). One hundred and fifty white male workers (mean ± SD: 41.0 ± 9 yrs of age) were examined: 100 3 × 8 rotating shiftworkers (40.4 ± 8.7 yrs of age) and 50 day workers (42.2 ± 9.4 yrs of age). The authors used bisulfite-pyrosequencing to esti- mate repetitive elements and gene-specific methylation. Multiple regression analysis, adjusted for age, body mass index (BMI), and job seniority, did not show any signifi- cant association between the five DNA methylation markers and shiftwork. However, job seniority, in all subjects, was significantly associated with Alu (β = −0.019, p = .033) and IFN-γ (β = −0.224, p 15 yrs) was associated with significantly lower Alu (β = −0.86, p = .006) and IFN-γ methylation (β = −6.50, p = .007) after adjust- ment for age, BMI, and morningness/eveningness. In addition, GCR significantly increased with length of shiftwork (β = 3.33, p = .05). The data showed alterations in blood DNA methylation in a group of shiftworkers, including changes in Alu repeti- tive elements methylation and gene-specific methylation of IFN-γ and TNF-α promoters. Further studies are required to determine the role of such alterations in mediating the effects of shiftwork on human health. PMID:20636218

  14. Slowing DNA Translocation in a Nanofluidic Field-Effect Transistor.

    Science.gov (United States)

    Liu, Yifan; Yobas, Levent

    2016-04-26

    Here, we present an experimental demonstration of slowing DNA translocation across a nanochannel by modulating the channel surface charge through an externally applied gate bias. The experiments were performed on a nanofluidic field-effect transistor, which is a monolithic integrated platform featuring a 50 nm-diameter in-plane alumina nanocapillary whose entire length is surrounded by a gate electrode. The field-effect transistor behavior was validated on the gating of ionic conductance and protein transport. The gating of DNA translocation was subsequently studied by measuring discrete current dips associated with single λ-DNA translocation events under a source-to-drain bias of 1 V. The translocation speeds under various gate bias conditions were extracted by fitting event histograms of the measured translocation time to the first passage time distributions obtained from a simple 1D biased diffusion model. A positive gate bias was observed to slow the translocation of single λ-DNA chains markedly; the translocation speed was reduced by an order of magnitude from 18.4 mm/s obtained under a floating gate down to 1.33 mm/s under a positive gate bias of 9 V. Therefore, a dynamic and flexible regulation of the DNA translocation speed, which is vital for single-molecule sequencing, can be achieved on this device by simply tuning the gate bias. The device is realized in a conventional semiconductor microfabrication process without the requirement of advanced lithography, and can be potentially further developed into a compact electronic single-molecule sequencer.

  15. An Innovative Algorithm to Implement Flow Control in Full Duplex Ethernet

    Institute of Scientific and Technical Information of China (English)

    WANGTao; GENing; FENGChongxi

    2003-01-01

    Nowadays Ethernet is a widely used LAN technology.It is based on a Media access control (MAC) method called Carrier sense multiple access with Collision detection(CAMS/CD).With CSMA/CD,hosts on Ethernet must operate in half duplex mode;with collision detection and random backoff,Ethernet is a self-contained LAN in the aspect of flow control.After the appearance of switching Ethernet,applications of full duplex Ethernet are booming,and how to manage network bandwidth and switch ports without CSMA/CD is a major issue.In this way,IEEE specifled and explicit flow control scheme,called "PAUSE operation" in Standard 802.3.In order to implement IEEE PAUSE operation and flow control effectively,we propose an innovative algorithm called Queue Length based PAUSE operation-QLPAUSE.It will set the pause time parameter according to the instant input port queue length and local condition checking.With it the hosts can make better use of the full duplex link.With the simulation results,it is proved that QLPAUSE is an active flow control,and it is an easy-to-implement and high efflciency scheme.Finally,the application of this algorithm is provided,with more and more Switching Ethernet,and PAUSE operation is accepted and supported by more and more manufacturers.Within many implementation methods,QLPAUSE is a best candidate.

  16. Relationship between microstructure and fracture types in a UNS S32205 duplex stainless steel

    Directory of Open Access Journals (Sweden)

    Maria Victoria Biezma

    2013-01-01

    Full Text Available Duplex stainless steels are susceptible to the formation of sigma phase at high temperature which could potentially be responsible for catastrophic service failure of components. Thermal treatments were applied to duplex stainless steels in order to promote the precipitation of different fractions of sigma phase into a ferrite-austenite microstructure. Quantitative image analysis was employed to characterize the microstructure and Charpy impact tests were used in order to evaluate the mechanical degradation caused by sigma phase presence. The fracture morphology of the Charpy test specimens were thoroughly observed in SEM, looking for a correlation between the microstructure and the fracture types in UNS S32205 duplex stainless steel. The main conclusion is the strong embrittlement effect of sigma phase since it is possible to observe a transition from transgranular fracture to intergranular fracture as increases the percentage of sigma phase. Thus, the mixed modes of fracture are predominant in the present study with high dependence on sigma phase percentages obtained by different thermal treatments.

  17. Hardfacing of duplex stainless steel using melting and diffusion processes

    Science.gov (United States)

    Lailatul, H.; Maleque, M. A.

    2017-03-01

    Duplex stainless steel (DSS) is a material with high potential successes in many new applications such as rail car manufacturing, automotive and chemical industries. Although DSS is widely used in various industries, this material has faced wear and hardness problems which obstruct a wider capability of this material and causes problems in current application. Therefore, development of surface modification has been introduced to produce hard protective layer or coating on DSS. The main aim of this work is to brief review on hard surface layer formation on DSS using melting and diffusion processes. Melting technique using tungsten inert gas (TIG) torch and diffusion technique using gas nitriding are the effective process to meet this requirement. The processing route plays a significant role in developing the hard surface layer for any application with effective cost and environmental factors. The good understanding and careful selection of processing route to form products are very important factors to decide the suitable techniques for surface engineering treatment. In this paper, an attempt is also made to consolidate the important research works done on melting and diffusion techniques of DSS in the past. The advantages and disadvantages between melting and diffusion technique are presented for better understanding on the feasibility of hard surface formation on DSS. Finally, it can be concluded that this work will open an avenue for further research on the application of suitable process for hard surface formation on DSS.

  18. Effect of DNA polymerase inhibitors on DNA repair in intact and permeable human fibroblasts: Evidence that DNA polymerases. delta. and. beta. are involved in DNA repair synthesis induced by N-methyl-N prime -nitro-N-nitrosoguanidine

    Energy Technology Data Exchange (ETDEWEB)

    Hammond, R.A.; Miller, M.R. (West Virginia Univ. Health Sciences Center, Morgantown (USA)); McClung, J.K. (Samuel Roberts Noble Foundation, Inc., East Ardmore, OK (USA))

    1990-01-09

    The involvement of DNA polymerases {alpha}, {beta}, and {delta} in DNA repair synthesis induced by N-methyl-N{prime}-nitro-N-nitrosoguanidine (MNNG) was investigated in human fibroblasts (HF). The effects of anti-(DNA polymerase {alpha}) monoclonal antibody, (p-n-butylphenyl)deoxyguanosine triphosphate (BuPdGTP), dideoxythymidine triphosphate (ddTTP), and aphidicolin on MNNG-induced DNA repair synthesis were investigated to dissect the roles of the different DNA polymerases. A subcellular system (permeable cells), in which DNA repair synthesis and DNA replication were differentiated by CsCl gradient centrifugation of BrdUMP density-labeled DNA, was used to examine the effects of the polymerase inhibitors. Another approach investigated the effects of several of these inhibitors of MNNG-induced DNA repair synthesis in intact cells by measuring the amount of ({sup 3}H)thymidine incorporated into repair DNA as determined by autoradiography and quantitation with an automated video image analysis system. In permeable cells, MNNG-induced DNA repair synthesis was inhibited 56% by 50 {mu}g of aphidicolin/mL, 6% by 10 {mu}M BuPdGTP, 13% by anti-(DNA polymerse {alpha}) monoclonal antibodies, and 29% by ddTTP. In intact cells, MNNG-induced DNA repair synthesis was inhibited 57% by 50 {mu}g of aphidicolin/mL and was not significantly inhibited by microinjecting anti-(DNA polymerase {alpha}) antibodies into HF nuclei. These results indicate that both DNA polymerase {delta} and {beta} are involved in repairing DNA damage caused by MNNG.

  19. Microvesicles Contribute to the Bystander Effect of DNA Damage.

    Science.gov (United States)

    Lin, Xiaozeng; Wei, Fengxiang; Major, Pierre; Al-Nedawi, Khalid; Al Saleh, Hassan A; Tang, Damu

    2017-04-07

    Genotoxic treatments elicit DNA damage response (DDR) not only in cells that are directly exposed but also in cells that are not in the field of treatment (bystander cells), a phenomenon that is commonly referred to as the bystander effect (BE). However, mechanisms underlying the BE remain elusive. We report here that etoposide and ultraviolet (UV) exposure stimulate the production of microvesicles (MVs) in DU145 prostate cancer cells. MVs isolated from UV-treated DU145 and A431 epidermoid carcinoma cells as well as etoposide-treated DU145 cells induced phosphorylation of ataxia-telangiectasia mutated (ATM) at serine 1981 (indicative of ATM activation) and phosphorylation of histone H2AX at serine 139 (γH2AX) in naïve DU145 cells. Importantly, neutralization of MVs derived from UV-treated cells with annexin V significantly reduced the MV-associated BE activities. Etoposide and UV are known to induce DDR primarily through the ATM and ATM- and Rad3-related (ATR) pathways, respectively. In this regard, MV is likely a common source for the DNA damage-induced bystander effect. However, pre-treatment of DU145 naïve cells with an ATM (KU55933) inhibitor does not affect the BE elicited by MVs isolated from etoposide-treated cells, indicating that the BE is induced upstream of ATM actions. Taken together, we provide evidence supporting that MVs are a source of the DNA damage-induced bystander effect.

  20. Detection of invasive infection caused by Fusarium solani and non-Fusarium solani species using a duplex quantitative PCR-based assay in a murine model of fusariosis.

    Science.gov (United States)

    Bernal-Martínez, Leticia; Buitrago, Maria J; Castelli, Maria V; Rodríguez-Tudela, Juan L; Cuenca-Estrella, Manuel

    2012-04-01

    A duplex Real Time PCR (RT-PCR) assay for detecting DNA of members of the genus Fusarium has been developed and validated by using two mouse models of invasive infection. The duplex RT-PCR technique employed two specific molecular beacon probes targeting a highly conserved region of the fungal rDNA gene. This technique showed a detection limit of 10 fg DNA per μl of sample and a specificity of 100%. The sensitivity in a total of 48 samples from a murine model of Fusarium solani infection was 93.9% for lung tissues and 86.7% for serum samples. In comparison, the sensitivity in a total of 45 samples of a F. oxysporum murine model infection was 87% for lung tissues and 42.8% for serum samples. This molecular technique could be a reliable method for the quantification and the evaluation of the disease in animal models and for the clinical diagnosis of fusariosis.

  1. Role of routine pre-operative screening venous duplex ultrasound in morbidly obese patients undergoing bariatric surgery

    Directory of Open Access Journals (Sweden)

    P Praveen Raj

    2017-01-01

    Full Text Available Background/Aims: It is well established that obesity is a strongly associated risk factor for post-operative deep vein thrombosis (DVT. Physical effects and pro-thrombotic, pro-inflammatory and hypofibrinolytic effects of severe obesity may predispose to idiopathic DVT (pre-operatively because of which bariatric patients are routinely screened before surgery. The aim of this study was to audit the use of routine screening venous duplex ultrasound in morbidly obese patients before undergoing bariatric surgery. Methods: We retrospectively reviewed 180 patients who underwent bariatric surgery from August 2013 to August 2014 who had undergone pre-operative screening bilateral lower-extremity venous duplex ultrasound for DVT. Data were collected on patient's demographics, history of venous thromboembolism, prior surgeries and duplex ultrasound details of the status of the deep veins and superficial veins of the lower limbs. Results: No patients had symptoms or signs of DVT pre-operatively. No patient gave history of DVT. No patient was found to have iliac, femoral or popliteal vein thrombosis. Superficial venous disease was found in 17 (8%. One patient had a right lower limb venous ulcer. Conclusion: Thromboembolic problems in the morbidly obese before bariatric surgery are infrequent, and screening venous duplex ultrasound can be done in high-risk patients only.

  2. DNA methylation in an engineered heart tissue model of cardiac hypertrophy: common signatures and effects of DNA methylation inhibitors.

    Science.gov (United States)

    Stenzig, Justus; Hirt, Marc N; Löser, Alexandra; Bartholdt, Lena M; Hensel, Jan-Tobias; Werner, Tessa R; Riemenschneider, Mona; Indenbirken, Daniela; Guenther, Thomas; Müller, Christian; Hübner, Norbert; Stoll, Monika; Eschenhagen, Thomas

    2016-01-01

    DNA methylation affects transcriptional regulation and constitutes a drug target in cancer biology. In cardiac hypertrophy, DNA methylation may control the fetal gene program. We therefore investigated DNA methylation signatures and their dynamics in an in vitro model of cardiac hypertrophy based on engineered heart tissue (EHT). We exposed EHTs from neonatal rat cardiomyocytes to a 12-fold increased afterload (AE) or to phenylephrine (PE 20 µM) and compared DNA methylation signatures to control EHT by pull-down assay and DNA methylation microarray. A 7-day intervention sufficed to induce contractile dysfunction and significantly decrease promoter methylation of hypertrophy-associated upregulated genes such as Nppa (encoding ANP) and Acta1 (α-skeletal actin) in both intervention groups. To evaluate whether pathological consequences of AE are affected by inhibiting de novo DNA methylation we applied AE in the absence and presence of DNA methyltransferase (DNMT) inhibitors: 5-aza-2'-deoxycytidine (aza, 100 µM, nucleosidic inhibitor), RG108 (60 µM, non-nucleosidic) or methylene disalicylic acid (MDSA, 25 µM, non-nucleosidic). Aza had no effect on EHT function, but RG108 and MDSA partially prevented the detrimental consequences of AE on force, contraction and relaxation velocity. RG108 reduced AE-induced Atp2a2 (SERCA2a) promoter methylation. The results provide evidence for dynamic DNA methylation in cardiac hypertrophy and warrant further investigation of the potential of DNA methylation in the treatment of cardiac hypertrophy.

  3. Hot Forging of Nitrogen Alloyed Duplex Stainless Steels

    Institute of Scientific and Technical Information of China (English)

    P.Chandramohan; S.S. Mohamed Nazirudeen; S.S. Ramakrishnan

    2007-01-01

    Duplex stainless steels are gaining global importance because of the need for a high strength corrosion resistant material. Three compositions of this group were selected with three different nitrogen contents viz, 0.15 wt pct (alloy 1), 0.23 wt pct (alloy 2) and 0.32 wt pct (alloy 3). The steels were melted in a high frequency induction furnace and hot forged to various reductions from 16% to 62%. In this work, the effect of hot forging on the ferrite content, hardness, yield strength, impact strength and grain orientation (texture) were studied. Fracture analysis on all the forged specimens using SEM reveals that a size reduction of 48% results in maximum ductility and impact strength as well as minimal ferrite content and grain size. Thus the mechanical properties are found to have a direct correlation to ferrite content and grain size. The highest impact strength was observed in specimens with the smallest grain size, which was observed in specimens forged to 48% reduction in size.

  4. JAEA Fatigue Analysis of EBR-II Duplex Tubing

    Energy Technology Data Exchange (ETDEWEB)

    J. H. Jackson; D. L. Porter; W. R. Lloyd

    2009-07-01

    This work addresses questions brought up concerning the mechanisms associated with fatigue crack growth retardation and/or arrest within the nickel bond layer in duplex 2¼ Cr-1Mo steel superheater tubes. Previous work performed at the Idaho National Laboratory (INL) indicated that the nickel bond layer did not function as a crack arrestor during fatigue crack propagation with the exception of one, isolated case involving an exceptionally low fatigue load and a high temperature (400 0C) environment. Since it is atypical for a fatigue crack to propagate from a relatively soft material (the nickel bond layer) to a harder material (the 2¼ Cr-1Mo steel) there has been speculation that the nickel bond layer was hardened in service. Additionally, there are questions surrounding the nature of the fatigue crack propagation within the nickel bond layer; specifically with regard to the presence of voids seen on micrographs of the bond layer and oxidation within the steel along the edge of the nickel bond layer. There is uncertainty as to the effect of these voids and/or oxide barriers with respect to potential fatigue crack arrest.

  5. Labeling-free fluorescent detection of DNA hybridization through FRET from pyrene excimer to DNA intercalator SYBR green I.

    Science.gov (United States)

    Zhou, Ruyi; Xu, Chen; Dong, Jie; Wang, Guojie

    2015-03-15

    A novel labeling-free fluorescence complex probe has been developed for DNA hybridization detection based on fluorescence resonance energy transfer (FRET) mechanism from pyrene excimer of pyrene-functionalized poly [2-(N, N-dimethylamino) ethyl methacrylate] (PFP) to SYBR Green I (SG, a specific intercalator of double-stranded DNA) in a cost-effective, rapid and simple manner. The complex probe consists of the positively charged PFP, SG and negatively charged single-stranded DNA (ssDNA). Upon adding a complementary strand to the complex probe solution, double-stranded DNA (dsDNA) was formed, followed by the intercalation of SG into dsDNA. The pyrene excimer emission was overlapped with the absorption of SG very well and the electrostatic interactions between PFP and dsDNA kept them in close proximity, enabling efficient FRET from pyrene excimer to SG. The fluorescence of SG in the duplex DNA resulting from FRET can be successfully applied to detect DNA hybridization with high sensitivity for a very low detection limit of 10nM and excellent selectivity for detection of single base pair mismatch. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. The beta subunit sliding DNA clamp is responsible for unassisted mutagenic translesion replication by DNA polymerase III holoenzyme.

    Science.gov (United States)

    Tomer, G; Reuven, N B; Livneh, Z

    1998-11-24

    The replication of damaged nucleotides that have escaped DNA repair leads to the formation of mutations caused by misincorporation opposite the lesion. In Escherichia coli, this process is under tight regulation of the SOS stress response and is carried out by DNA polymerase III in a process that involves also the RecA, UmuD' and UmuC proteins. We have shown that DNA polymerase III holoenzyme is able to replicate, unassisted, through a synthetic abasic site in a gapped duplex plasmid. Here, we show that DNA polymerase III*, a subassembly of DNA polymerase III holoenzyme lacking the beta subunit, is blocked very effectively by the synthetic abasic site in the same DNA substrate. Addition of the beta subunit caused a dramatic increase of at least 28-fold in the ability of the polymerase to perform translesion replication, reaching 52% bypass in 5 min. When the ssDNA region in the gapped plasmid was extended from 22 nucleotides to 350 nucleotides, translesion replication still depended on the beta subunit, but it was reduced by 80%. DNA sequence analysis of translesion replication products revealed mostly -1 frameshifts. This mutation type is changed to base substitution by the addition of UmuD', UmuC, and RecA, as demonstrated in a reconstituted SOS translesion replication reaction. These results indicate that the beta subunit sliding DNA clamp is the major determinant in the ability of DNA polymerase III holoenzyme to perform unassisted translesion replication and that this unassisted bypass produces primarily frameshifts.

  7. Role of the heat capacity change in understanding and modeling melting thermodynamics of complementary duplexes containing standard and nucleobase-modified LNA.

    Science.gov (United States)

    Hughesman, Curtis B; Turner, Robin F B; Haynes, Charles A

    2011-06-14

    Melting thermodynamic data obtained by differential scanning calorimetry (DSC) are reported for 43 duplexed oligonucleotides containing one or more locked nucleic acid (LNA) substitutions. The measured heat capacity change (ΔC(p)) for the helix-to-coil transition is used to compute the changes in enthalpy and entropy for melting of an LNA-bearing duplex at the T(m) of its corresponding isosequential unmodified DNA duplex to allow rigorous thermodynamic analysis of the stability enhancements provided by LNA substitutions. Contrary to previous studies, our analysis shows that the origin of the improved stability is almost exclusively a net reduction (ΔΔS° thermodynamics and the increased melting temperature (ΔT(m)) of heteroduplexes formed between an unmodified DNA strand and a complementary strand containing any number and configuration of standard LNA nucleotides A, T, C, and G. This single-base thermodynamic (SBT) model requires only four entropy-related parameters in addition to ΔC(p). Finally, DSC data for 20 duplexes containing the nucleobase-modified LNAs 2-aminoadenine (D) and 2-thiothymine (H) are reported and used to determine SBT model parameters for D and H. The data and model suggest that along with the greater stability enhancement provided by D and H bases relative to their corresponding A and T analogues, the unique pseudocomplementary properties of D-H base pairs may make their use appealing for in vitro and in vivo applications.

  8. Sarcomatoid carcinoma of the renal pelvis in duplex kidney

    Institute of Scientific and Technical Information of China (English)

    CHEN Ge-ming; CHEN Shan-wen; XIA Dan; LI Jun; YAN Sheng; JIN Bai-ye

    2011-01-01

    Sarcomatoid transitional cell carcinoma of the renal pelvis is a rare neoplasm with only 14 well-illustrated examples reported previously. Duplex kidney is the most common congenital abnormality of the urinary tract, with an incidence of around 2%. Neoplasia of the renal pelvis in duplex kidney is rare. We reported a case whose sarcomatoid carcinoma originated from the upper portion of the duplicated renal pelvis with hydronephrosis, and total nephroureterectomy with bladder cuff excision surgery of both renal units was carried out. Because of the rare nature of renal pelvic sarcomatoid carcinoma and its apparent lack of response to adjuvant therapy, it is essential to do early diagnosis and early radical surgery to improve survival. It is important to stress the need for frequent and diligent monitoring or treating complex duplex kidney with hydronephrosis of either moiety in case of a risk of having neoplasias.

  9. Quantitative analysis of TALE-DNA interactions suggests polarity effects.

    Science.gov (United States)

    Meckler, Joshua F; Bhakta, Mital S; Kim, Moon-Soo; Ovadia, Robert; Habrian, Chris H; Zykovich, Artem; Yu, Abigail; Lockwood, Sarah H; Morbitzer, Robert; Elsäesser, Janett; Lahaye, Thomas; Segal, David J; Baldwin, Enoch P

    2013-04-01

    Transcription activator-like effectors (TALEs) have revolutionized the field of genome engineering. We present here a systematic assessment of TALE DNA recognition, using quantitative electrophoretic mobility shift assays and reporter gene activation assays. Within TALE proteins, tandem 34-amino acid repeats recognize one base pair each and direct sequence-specific DNA binding through repeat variable di-residues (RVDs). We found that RVD choice can affect affinity by four orders of magnitude, with the relative RVD contribution in the order NG > HD ≈ NN > NI > NK. The NN repeat preferred the base G over A, whereas the NK repeat bound G with 10(3)-fold lower affinity. We compared AvrBs3, a naturally occurring TALE that recognizes its target using some atypical RVD-base combinations, with a designed TALE that precisely matches 'standard' RVDs with the target bases. This comparison revealed unexpected differences in sensitivity to substitutions of the invariant 5'-T. Another surprising observation was that base mismatches at the 5' end of the target site had more disruptive effects on affinity than those at the 3' end, particularly in designed TALEs. These results provide evidence that TALE-DNA recognition exhibits a hitherto un-described polarity effect, in which the N-terminal repeats contribute more to affinity than C-terminal ones.

  10. Effect of Aperiodicity on the Charge Transfer Through DNA Molecules

    Science.gov (United States)

    Ghosh, Angsula; Chaudhuri, Puspitapallab

    The effect of aperiodicity on the charge transfer process through DNA molecules is investigated using a tight-binding model. Single-stranded aperiodic Fibonacci polyGC and polyAT sequences along with aperiodic Rudin-Shapiro poly(GCAT) sequences are used in the study. Based on the tight-binding model, molecular orbital calculations of the DNA chains are performed and ionization potentials compared, as this might be relevant to understanding the charge transfer process. Charges migrate through the sequences in a multistep hopping process. Results for current conduction through aperiodic sequences are compared with those for the corresponding periodic sequences. We find that dinucleotide aperiodic Fibonacci sequences decrease the current while tetranucleotide aperiodic Rudin-Shapiro sequences increase the current when compared with the corresponding periodic sequences. The conductance in all cases decays exponentially as the sequence length increases.

  11. Programming a topologically constrained DNA nanostructure into a sensor

    Science.gov (United States)

    Liu, Meng; Zhang, Qiang; Li, Zhongping; Gu, Jimmy; Brennan, John D.; Li, Yingfu

    2016-01-01

    Many rationally engineered DNA nanostructures use mechanically interlocked topologies to connect individual DNA components, and their physical connectivity is achieved through the formation of a strong linking duplex. The existence of such a structural element also poses a significant topological constraint on functions of component rings. Herein, we hypothesize and confirm that DNA catenanes with a strong linking duplex prevent component rings from acting as the template for rolling circle amplification (RCA). However, by using an RNA-containing DNA [2] catenane with a strong linking duplex, we show that a stimuli-responsive RNA-cleaving DNAzyme can linearize one component ring, and thus enable RCA, producing an ultra-sensitive biosensing system. As an example, a DNA catenane biosensor is engineered to detect the model bacterial pathogen Escherichia coli through binding of a secreted protein, with a detection limit of 10 cells ml−1, thus establishing a new platform for further applications of mechanically interlocked DNA nanostructures. PMID:27337657

  12. Combining a Ru(II) "Building Block" and Rapid Screening Approach to Identify DNA Structure-Selective "Light Switch" Compounds.

    Science.gov (United States)

    Wachter, Erin; Moyá, Diego; Glazer, Edith C

    2017-02-13

    A chemically reactive Ru(II) "building block", able to undergo condensation reactions with substituted diamines, was utilized to create a small library of luminescent "light switch" dipyrido-[3,2-a:2',3'-c] phenazine (dppz) complexes. The impact of substituent identity, position, and the number of substituents on the light switch effect was investigated. An unbiased, parallel screening approach was used to evaluate the selectivity of the compounds for a variety of different biomolecules, including protein, nucleosides, single stranded DNA, duplex DNA, triplex DNA, and G-quadruplex DNA. Combining these two approaches allowed for the identification of hit molecules that showed different selectivities for biologically relevant DNA structures, particularly triplex and quadruplex DNA.

  13. Development of duplex-PCR for identification of Aeromonas species

    Directory of Open Access Journals (Sweden)

    Carina Lucena Mendes-Marques

    2013-10-01

    Full Text Available Introduction The number of reports of intestinal infections caused by Aeromonas spp. has increased significantly in recent years. In most clinical laboratories, identification of these bacteria is carried out by general phenotypic tests that sometimes do not accurately differentiate Aeromonas and Vibrio. Methods A duplex-polymerase chain reaction (PCR was developed directed to 2 targets identifying Aeromonas spp. pathogenic to humans. Results The duplex-PCR results were reproducible and specific for Aeromonas spp. pathogenic to humans. Conclusions This method will allow differentiation between Vibrio and Aeromonas spp. in patients with in cholera-like symptoms and can also be used in water quality monitoring.

  14. Comparison of simulated and measured quantities of a duplex reactor

    Energy Technology Data Exchange (ETDEWEB)

    Koskela, M.; Kajava, M. [ABB Marine, Helsinki (Finland)

    1997-12-31

    The purpose of this article is to illustrate the use of an analog simulator as a design tool when designing new power electric equipment. The purpose of simulation is to predict the functionality of electrical equipment to be constructed. Duplex reactor is an electromagnetic device designed to reduce voltage harmonics and short circuit currents in the ship electrical network. In this report a comparison between simulated and measured electrical quantities of a duplex reactor has been made. The purpose of the measurements was to show the correct functioning of the reactor. The simulation results and the measured waveforms corresponds well to each other. (orig.) 4 refs.

  15. Development of duplex-PCR for identification of Aeromonas species

    Directory of Open Access Journals (Sweden)

    Carina Lucena Mendes-Marques

    2013-05-01

    Full Text Available Introduction The number of reports of intestinal infections caused by Aeromonas spp. has increased significantly in recent years. In most clinical laboratories, identification of these bacteria is carried out by general phenotypic tests that sometimes do not accurately differentiate Aeromonas and Vibrio. Methods A duplex-polymerase chain reaction (PCR was developed directed to 2 targets identifying Aeromonas spp. pathogenic to humans. Results The duplex-PCR results were reproducible and specific for Aeromonas spp. pathogenic to humans. Conclusions This method will allow differentiation between Vibrio and Aeromonas spp. in patients with in cholera-like symptoms and can also be used in water quality monitoring.

  16. Charge conduction properties of a parallel-stranded DNA G-quadruplex: implications for chromosomal oxidative damage.

    Science.gov (United States)

    Huang, Yu Chuan; Cheng, Alan K H; Yu, Hua-Zhong; Sen, Dipankar

    2009-07-28

    The charge-flow properties and concomitant guanine damage patterns of a number of intermolecular and wholly parallel-stranded DNA G-quadruplexes were investigated. The DNA constructs were structurally well-defined and consisted of the G-quadruplex sandwiched and stacked between two Watson-Crick base-paired duplexes. Such duplex-quadruplex-duplex constructs were designed to minimize torsional stress as well as steric crowding at the duplex-quadruplex junctions. When anthraquinone was used to induce charge flow within the constructs, it was found that the quadruplex served both as a sink and as a moderately good conductor of electron holes, relative to DNA duplexes. Most strikingly, the quadruplex suffered very little charge-flow generated oxidative damage relative to guanines in the duplex regions and, indeed, to guanines in antiparallel quadruplexes reported in prior studies. It is likely that these differences result from a combination of steric and electronic factors. A biological conclusion that may be drawn from these data is that if, as anticipated, G-quadruplex structures form in vivo at the telomeres and other loci in eukaryotic chromosomes, their ability to serve as protective sinks against chromosomal oxidative damage may depend on their specific character and topology. From a separate perspective, our results on the conduction properties of duplex-quadruplex-duplex DNA composites suggest the utility of G-quadruplexes as junction modules in the construction of DNA-based biosensors and nanocircuitry.

  17. Spermine attenuates the action of the DNA intercalator, actinomycin D, on DNA binding and the inhibition of transcription and DNA replication.

    Directory of Open Access Journals (Sweden)

    Sheng-Yu Wang

    Full Text Available The anticancer activity of DNA intercalators is related to their ability to intercalate into the DNA duplex with high affinity, thereby interfering with DNA replication and transcription. Polyamines (spermine in particular are almost exclusively bound to nucleic acids and are involved in many cellular processes that require nucleic acids. Until now, the effects of polyamines on DNA intercalator activities have remained unclear because intercalation is the most important mechanism employed by DNA-binding drugs. Herein, using actinomycin D (ACTD as a model, we have attempted to elucidate the effects of spermine on the action of ACTD, including its DNA-binding ability, RNA and DNA polymerase interference, and its role in the transcription and replication inhibition of ACTD within cells. We found that spermine interfered with the binding and stabilization of ACTD to DNA. The presence of increasing concentrations of spermine enhanced the transcriptional and replication activities of RNA and DNA polymerases, respectively, in vitro treated with ActD. Moreover, a decrease in intracellular polyamine concentrations stimulated by methylglyoxal-bis(guanylhydrazone (MGBG enhanced the ACTD-induced inhibition of c-myc transcription and DNA replication in several cancer cell lines. The results indicated that spermine attenuates ACTD binding to DNA and its inhibition of transcription and DNA replication both in vitro and within cells. Finally, a synergistic antiproliferative effect of MGBG and ACTD was observed in a cell viability assay. Our findings will be of significant relevance to future developments in combination with cancer therapy by enhancing the anticancer activity of DNA interactors through polyamine depletion.

  18. Spermine attenuates the action of the DNA intercalator, actinomycin D, on DNA binding and the inhibition of transcription and DNA replication.

    Science.gov (United States)

    Wang, Sheng-Yu; Lee, Alan Yueh-Luen; Lee, Yueh-Luen; Lai, Yi-Hua; Chen, Jeremy J W; Wu, Wen-Lin; Yuann, Jeu-Ming P; Su, Wang-Lin; Chuang, Show-Mei; Hou, Ming-Hon

    2012-01-01

    The anticancer activity of DNA intercalators is related to their ability to intercalate into the DNA duplex with high affinity, thereby interfering with DNA replication and transcription. Polyamines (spermine in particular) are almost exclusively bound to nucleic acids and are involved in many cellular processes that require nucleic acids. Until now, the effects of polyamines on DNA intercalator activities have remained unclear because intercalation is the most important mechanism employed by DNA-binding drugs. Herein, using actinomycin D (ACTD) as a model, we have attempted to elucidate the effects of spermine on the action of ACTD, including its DNA-binding ability, RNA and DNA polymerase interference, and its role in the transcription and replication inhibition of ACTD within cells. We found that spermine interfered with the binding and stabilization of ACTD to DNA. The presence of increasing concentrations of spermine enhanced the transcriptional and replication activities of RNA and DNA polymerases, respectively, in vitro treated with ActD. Moreover, a decrease in intracellular polyamine concentrations stimulated by methylglyoxal-bis(guanylhydrazone) (MGBG) enhanced the ACTD-induced inhibition of c-myc transcription and DNA replication in several cancer cell lines. The results indicated that spermine attenuates ACTD binding to DNA and its inhibition of transcription and DNA replication both in vitro and within cells. Finally, a synergistic antiproliferative effect of MGBG and ACTD was observed in a cell viability assay. Our findings will be of significant relevance to future developments in combination with cancer therapy by enhancing the anticancer activity of DNA interactors through polyamine depletion.

  19. DNA circles with cruciforms from Isospora (Toxoplasma) gondii

    NARCIS (Netherlands)

    Weijers, P.J.; Borst, P.; Overdulve, J.P.; Fase-Fowler, F.; Berg, Marlene van den

    1984-01-01

    We have isolated a closed circular duplex DNA fraction from the unicellular parasite Isospora (Toxoplasma) gondii and examined the purified DNA by electron microscopy. A major part of this circular DNA consists of 12-μm circles containing a cruciform with 0.5-μm tails. We also found 23-μm circles wi

  20. Breaking the Transmitter-Receiver Isolation Barrier in Mobile Handsets with Spatial Duplexing

    DEFF Research Database (Denmark)

    Alrabadi, Osama; Tatomirescu, Alexandru; Knudsen, Mikael;

    2013-01-01

    In full-duplex radio communication systems like eUTRAN, CDMA-2000, the radio transmitter (Tx) is active at the same time as the radio receiver (Rx). The Tx and the Rx will be using separate dedicated frequency bands and the Tx-Rx isolation is ensured by duplex filters. However, agile duplexers req...

  1. On the Potential of Full Duplex Performance in 5G Ultra-Dense Small Cell Networks

    DEFF Research Database (Denmark)

    Gatnau, Marta; Fleischer, Marko; Berardinelli, Gilberto;

    2016-01-01

    Full duplex allows a device to transmit and receive simultaneously in the same frequency band, theoretically doubling the throughput compared to traditional half duplex systems. However, several limitations restrict the promised full duplex gain: non-ideal self-interference cancellation, increase...

  2. Survivin minigene DNA vaccination is effective against neuroblastoma.

    Science.gov (United States)

    Fest, Stefan; Huebener, Nicole; Bleeke, Matthias; Durmus, Tahir; Stermann, Alexander; Woehler, Anja; Baykan, Bianca; Zenclussen, Ana C; Michalsky, Elke; Jaeger, Ines S; Preissner, Robert; Hohn, Oliver; Weixler, Silke; Gaedicke, Gerhard; Lode, Holger N

    2009-07-01

    The inhibitor of apoptosis protein survivin is highly expressed in neuroblastoma (NB) and survivin-specific T cells were identified in Stage 4 patients. Therefore, we generated a novel survivin minigene DNA vaccine (pUS-high) encoding exclusively for survivin-derived peptides with superior MHC class I (H2-K(k)) binding affinities and tested its efficacy to suppress tumor growth and metastases in a syngeneic NB mouse model. Vaccination was performed by oral gavage of attenuated Salmonella typhimurium SL7207 carrying pUS-high. Mice receiving the pUS-high in the prophylactic setting presented a 48-52% reduction in s.c. tumor volume, weight and liver metastasis level in contrast to empty vector controls. This response was as effective as a survivin full-length vaccine and was associated with an increased target cell lysis, increased presence of CD8(+) T-cells at the primary tumor site and enhanced production of proinflammatory cytokines by systemic CD8(+) T cells. Furthermore, depletion of CD8(+) but not CD4(+) T-cells completely abrogated the pUS-high mediated primary tumor growth suppression, demonstrating a CD8(+) T-cell mediated effect. Therapeutic vaccination with pUS-high led to complete NB eradication in over 50% of immunized mice and surviving mice showed an over 80% reduction in primary tumor growth upon rechallenge in contrast to controls. In summary, survivin-based DNA vaccination is effective against NB and the rational minigene design provides a promising approach to circumvent potentially hazardous effects of using full length antiapoptotic genes as DNA vaccines.

  3. A novel universal real-time PCR system using the attached universal duplex probes for quantitative analysis of nucleic acids

    Directory of Open Access Journals (Sweden)

    Wilson Zoe A

    2008-06-01

    Full Text Available Abstract Background Real-time PCR techniques are being widely used for nucleic acids analysis, but one limitation of current frequently employed real-time PCR is the high cost of the labeled probe for each target molecule. Results We describe a real-time PCR technique employing attached universal duplex probes (AUDP, which has the advantage of generating fluorescence by probe hydrolysis and strand displacement over current real-time PCR methods. AUDP involves one set of universal duplex probes in which the 5' end of the fluorescent probe (FP and a complementary quenching probe (QP lie in close proximity so that fluorescence can be quenched. The PCR primer pair with attached universal template (UT and the FP are identical to the UT sequence. We have shown that the AUDP technique can be used for detecting multiple target DNA sequences in both simplex and duplex real-time PCR assays for gene expression analysis, genotype identification, and genetically modified organism (GMO quantification with comparable sensitivity, reproducibility, and repeatability with other real-time PCR methods. Conclusion The results from GMO quantification, gene expression analysis, genotype identification, and GMO quantification using AUDP real-time PCR assays indicate that the AUDP real-time PCR technique has been successfully applied in nucleic acids analysis, and the developed AUDP real-time PCR technique will offer an alternative way for nucleic acid analysis with high efficiency, reliability, and flexibility at low cost.

  4. Radiation damage to DNA: the effect of LET

    Energy Technology Data Exchange (ETDEWEB)

    Ward, J.F.; Milligan, J.R. [California Univ., San Diego, La Jolla, CA (United States). School of Medicine

    1997-03-01

    Mechanisms whereby ionizing radiation induced damage are introduced into cellular DNA are discussed. The types of lesions induced are summarized and the rationale is presented which supports the statement that radiation induced singly damaged sites are biologically unimportant. The conclusion that multiply damaged sites are critical is discussed and the mechanisms whereby such lesions are formed are presented. Structures of multiply damaged sites are summarized and problems which they present to cellular repair systems are discussed. Lastly the effects of linear energy transfer on the complexity of multiply damaged sites are surveyed and the consequences of this increased complexity are considered in terms of cell survival and mutation. (author)

  5. Effects of 8-halo-7-deaza-2'-deoxyguanosine triphosphate on DNA synthesis by DNA polymerases and cell proliferation.

    Science.gov (United States)

    Yin, Yizhen; Sasaki, Shigeki; Taniguchi, Yosuke

    2016-08-15

    8-OxodG (8-oxo-2'-deoxyguanosine) is representative of nucleoside damage and shows a genotoxicity. To significantly reveal the contributions of 7-NH and C8-oxygen to the mutagenic effect of 8-oxodG by DNA polymerases, we evaluated the effects of the 8-halo-7-deaza-dG (8-halogenated 7-deaza-2'-deoxyguanosine) derivatives by DNA polymerases. 8-Halo-7-deaza-dGTPs were poorly incorporated by both KF(exo(-)) and human DNA polymerase β opposite dC or dA into the template DNA. Furthermore, it was found that KF(exo(-)) was very sensitive to the introduction of the C8-halogen, while polymerase β can accommodate the C8-halogen resulting in an efficient dCTP insertion opposite the 8-halo-7-deaza-dG in the template DNA. These results indicate that strong hydrogen bonding between 7-NH in the 8-oxo-G nucleobase and 1-N in the adenine at the active site of the DNA polymerase is required for the mutagenic effects. Whereas, I-deaza-dGTP shows an antiproliferative effect for the HeLa cells, suggesting that it could become a candidate as a new antitumor agent.

  6. Cytotoxicity of new duplex drugs linking 3'-C-ethynylcytidine and 5-fluor-2'-deoxyuridine against human melanoma cells.

    Science.gov (United States)

    Schott, Sarah; Niessner, Heike; Sinnberg, Tobias; Venturelli, Sascha; Berger, Alexander; Ikenberg, Kristian; Villanueva, Jessie; Meier, Friedegund; Garbe, Claus; Busch, Christian

    2012-11-01

    Melanoma is an increasingly common and potentially fatal malignancy of the skin and some mucous membranes. As no cure exists for metastatic disease, there is an urgent need for novel drugs. 2'-Deoxy-5-fluorouridylyl-(3'-5')-3'-C-ethynylcytidine [5-FdU(3'-5')ECyd] and 3'-C-ethynylcytidinylyl-(5' → 1-O)-2-O-octadecyl-sn-glycerylyl-(3-O → 5')-2'-deoxy-5-fluorouridine [ECyd-lipid-5-FdU] represent cytostatic active duplex drugs, which can be metabolized into various active antimetabolites. We evaluated the cytotoxicity of these heterodinucleoside phosphate analogs, their corresponding monomers ECyd and 5-FdU and combinations thereof on six metastatic melanoma cell lines and six ex vivo patient-derived melanoma cells in comparison to current standard cytostatic agents and the BRAF V600E inhibitor Vemurafenib. In vitro (real-time)-proliferation assays demonstrated that 5-FdU(3'-5')ECyd and ECyd-lipid-5-FdU had a high cytotoxic efficacy causing 75% melanoma cell death at concentrations in the nanomolar and micromolar range. Cytotoxicity was conducted by induction of DNA cleavage indicating apoptotic cells. Chicken embryotoxicity demonstrated that the duplex drugs were less toxic than 5-FdU at 0.01 μM. In vivo the duplex drug 5-FdU(3'-5')ECyd was efficacious in the murine LOX IMVI melanoma xenograph model on administration of 11.2 mg/kg/injection every fourth day. Both duplex drugs are promising novel cytostatic agents for the treatment of malignant melanoma meriting clinical evaluation. Copyright © 2012 UICC.

  7. Analysis of full-duplex relay networks with opportunistic scheduling

    Directory of Open Access Journals (Sweden)

    Lei Shao

    2015-05-01

    Full Text Available This Letter addresses a two-hop decode-and-forward relay system with full-duplex relaying and opportunistic scheduling. Exact expressions for outage probability, average capacity and symbol error rate are presented in an independent identically distributed Rayleigh fading environment. Numerical and simulated results show the validity of the analytical results.

  8. Reliability Capacity of Half-Duplex Channels with Strict Deadlines

    DEFF Research Database (Denmark)

    Costa, Rui; Roetter, Daniel Enrique Lucani; Vinhoza, Tiago

    2015-01-01

    A fundamental characterization of a half-duplex wireless system with packet losses under traffic with hard deadlines is instrumental to understanding and developing efficient, coding aware policies for real-time applications. We set forth the concept of reliability capacity with a limited number...

  9. Full-duplex wireless communications systems self-interference cancellation

    CERN Document Server

    Le-Ngoc, Tho

    2017-01-01

    This book introduces the development of self-interference (SI)-cancellation techniques for full-duplex wireless communication systems. The authors rely on estimation theory and signal processing to develop SI-cancellation algorithms by generating an estimate of the received SI and subtracting it from the received signal. The authors also cover two new SI-cancellation methods using the new concept of active signal injection (ASI) for full-duplex MIMO-OFDM systems. The ASI approach adds an appropriate cancelling signal to each transmitted signal such that the combined signals from transmit antennas attenuate the SI at the receive antennas. The authors illustrate that the SI-pre-cancelling signal does not affect the data-bearing signal. This book is for researchers and professionals working in wireless communications and engineers willing to understand the challenges of deploying full-duplex and practical solutions to implement a full-duplex system. Advanced-level students in electrical engineering and computer ...

  10. Capacity of the Degraded Half-Duplex Relay Channel

    CERN Document Server

    Vijayakumaran, Saravanan; Lok, Tat M

    2007-01-01

    A discrete memoryless half-duplex relay channel is constructed from a broadcast channel from the source to the relay and destination and a multiple access channel from the source and relay to the destination. When the relay listens, the channel operates in the broadcast mode. The channel switches to the multiple access mode when the relay transmits. If the broadcast component channel is physically degraded, the half-duplex relay channel will also be referred to as physically degraded. The capacity of this degraded half-duplex relay channel is examined. It is shown that the block Markov coding suggested in the seminal paper by Cover and El Gamal can be modified to achieve capacity for the degraded half-duplex relay channel. In the code construction, the listen-transmit schedule of the relay is made to depend on the message to be sent and hence the schedule carries information itself. If the schedule is restricted to be deterministic, it is shown that the capacity can be achieved by a simple management of infor...

  11. Full-duplex MIMO system based on antenna cancellation technique

    DEFF Research Database (Denmark)

    Foroozanfard, Ehsan; Franek, Ondrej; Tatomirescu, Alexandru

    2014-01-01

    The performance of an antenna cancellation technique for a multiple-input– multiple-output (MIMO) full-duplex system that is based on null-steering beamforming and antenna polarization diversity is investigated. A practical implementation of a symmetric antenna topology comprising three dual-pola...

  12. DNA strand transfer catalyzed by vaccinia topoisomerase: ligation of DNAs containing a 3' mononucleotide overhang.

    Science.gov (United States)

    Cheng, C; Shuman, S

    2000-05-01

    The specificity of vaccinia topoisomerase for transesterification to DNA at the sequence 5'-CCCTT and its versatility in strand transfer have illuminated the recombinogenic properties of type IB topoisomerases and spawned topoisomerase-based strategies for DNA cloning. Here we characterize a pathway of topoisomerase-mediated DNA ligation in which enzyme bound covalently to a CCCTT end with an unpaired +1T nucleotide rapidly and efficiently joins the CCCTT strand to a duplex DNA containing a 3' A overhang. The joining reaction occurs with high efficiency, albeit slowly, to duplex DNAs containing 3' G, T or C overhangs. Strand transfer can be restricted to the correctly paired 3' A overhang by including 0.5 M NaCl in the ligation reaction mixture. The effects of base mismatches and increased ionic strength on the rates of 3' overhang ligation provide a quantitative picture of the relative contributions of +1 T:A base pairing and electrostatic interactions downstream of the scissile phosphate to the productive binding of an unlinked acceptor DNA to the active site. The results clarify the biochemistry underlying topoisomerase-cloning of PCR products with non-templated 3' overhangs.

  13. Site-Selective Monitoring of the Interaction of the SRA Domain of UHRF1 with Target DNA Sequences Labeled with 2-Aminopurine.

    Science.gov (United States)

    Greiner, Vanille J; Kovalenko, Lesia; Humbert, Nicolas; Richert, Ludovic; Birck, Catherine; Ruff, Marc; Zaporozhets, Olga A; Dhe-Paganon, Sirano; Bronner, Christian; Mély, Yves

    2015-10-06

    UHRF1 plays a central role in the maintenance and transmission of epigenetic modifications by recruiting DNMT1 to hemimethylated CpG sites via its SET and RING-associated (SRA) domain, ensuring error-free duplication of methylation profiles. To characterize SRA-induced changes in the conformation and dynamics of a target 12 bp DNA duplex as a function of the methylation status, we labeled duplexes by the environment-sensitive probe 2-aminopurine (2-Ap) at various positions near or far from the central CpG recognition site containing either a nonmodified cytosine (NM duplex), a methylated cytosine (HM duplex), or methylated cytosines on both strands (BM duplex). Steady-state and time-resolved fluorescence indicated that binding of SRA induced modest conformational and dynamical changes in NM, HM, and BM duplexes, with only slight destabilization of base pairs, restriction of global duplex flexibility, and diminution of local nucleobase mobility. Moreover, significant restriction of the local motion of residues flanking the methylcytosine in the HM duplex suggested that these residues are more rigidly bound to SRA, in line with a slightly higher affinity of the HM duplex as compared to that of the NM or BM duplex. Our results are consistent with a "reader" role, in which the SRA domain scans DNA sequences for hemimethylated CpG sites without perturbation of the structure of contacted nucleotides.

  14. Downlink Error Rates of Half-duplex Users in Full-duplex Networks over a Laplacian Inter-User Interference Limited and EGK fading

    KAUST Repository

    Soury, Hamza

    2017-03-14

    This paper develops a mathematical framework to study downlink error rates and throughput for half-duplex (HD) terminals served by a full-duplex (FD) base station (BS). The developed model is used to motivate long term pairing for users that have non-line of sight (NLOS) interfering link. Consequently, we study the interferer limited problem that appears between NLOS HD users-pair that are scheduled on the same FD channel. The distribution of the interference is first characterized via its distribution function, which is derived in closed form. Then, a comprehensive performance assessment for the proposed pairing scheme is provided by assuming Extended Generalized- $cal{K}$ (EGK) fading for the downlink and studying different modulation schemes. To this end, a unified closed form expression for the average symbol error rate is derived. Furthermore, we show the effective downlink throughput gain harvested by the pairing NLOS users as a function of the average signal-to-interferenceratio when compared to an idealized HD scenario with neither interference nor noise. Finally, we show the minimum required channel gain pairing threshold to harvest downlink throughput via the FD operation when compared to the HD case for each modulation scheme.

  15. The effect of DNA on mechanical properties of nanofiber hydrogels

    Science.gov (United States)

    Shin, Min Kyoon; Kim, Shi Hyeong; Jung, Sung-il; Kim, Sun I.; Kim, Seon Jeong; Kim, Byung Joo; So, Insuk

    2008-10-01

    Uniform poly(vinyl alcohol) (PVA) nanofiber gels incorporating double-stranded deoxyribonucleic acid (DNA) were fabricated without the aid of cross-linkers employing electrospinning. Creep and tensile tests of the DNA/PVA nanofiber gels were carried out in an aqueous medium to analyze interactions between the DNA and PVA. The DNA/PVA gels had a higher elastic modulus than the PVA gel. The viscosity with increasing external load was calculated by applying the Burger model to the creep curves. We conclude that DNA is relatively weakly bound with the PVA chains, although the elastic modulus of the DNA/PVA gels was higher than that of PVA gel.

  16. PolyA-Mediated DNA Assembly on Gold Nanoparticles for Thermodynamically Favorable and Rapid Hybridization Analysis.

    Science.gov (United States)

    Zhu, Dan; Song, Ping; Shen, Juwen; Su, Shao; Chao, Jie; Aldalbahi, Ali; Zhou, Ziang; Song, Shiping; Fan, Chunhai; Zuo, Xiaolei; Tian, Yang; Wang, Lianhui; Pei, Hao

    2016-05-03

    Understanding the behavior of biomolecules on nanointerface is critical in bioanalysis, which is great challenge due to the instability and the difficulty to control the orientation and loading density of biomolecules. Here, we investigated the thermodynamics and kinetics of DNA hybridization on gold nanoparticle, with the aim to improve the efficiency and speed of DNA analysis. We achieved precise and quantitative surface control by applying a recently developed poly adenines (polyA)-based assembly strategy on gold nanoparticles (DNA-AuNPs). PolyA served as an effective anchoring block based on the preferential binding with the AuNP surface and the appended recognition block adopted an upright conformation that favors DNA hybridization. The lateral spacing and surface density of DNA on AuNPs can be systematically modulated by adjusting the length of polyA block. We found the stability of duplex on AuNP was enhanced with the increasing length of polyA block. When the length of polyA block reached to 40 bases, the thermodynamic properties were more similar to that of duplex in solution. Fast hybridization rate was observed on the diblock DNA-AuNPs and was increased along with the length of polyA block. We consider the high stability and excellent hybridization performance come from the minimization of the DNA-DNA and DNA-AuNP interactions with the use of polyA block. This study provides better understanding of the behavior of biomolecules on the nanointerface and opens new opportunities to construct high-efficiency and high-speed biosensors for DNA analysis.

  17. The impact of the C-terminal domain on the interaction of human DNA topoisomerase II α and β with DNA.

    Directory of Open Access Journals (Sweden)

    Kathryn L Gilroy

    Full Text Available BACKGROUND: Type II DNA topoisomerases are essential, ubiquitous enzymes that act to relieve topological problems arising in DNA from normal cellular activity. Their mechanism of action involves the ATP-dependent transport of one DNA duplex through a transient break in a second DNA duplex; metal ions are essential for strand passage. Humans have two isoforms, topoisomerase IIα and topoisomerase IIβ, that have distinct roles in the cell. The C-terminal domain has been linked to isoform specific differences in activity and DNA interaction. METHODOLOGY/PRINCIPAL FINDINGS: We have investigated the role of the C-terminal domain in the binding of human topoisomerase IIα and topoisomerase IIβ to DNA in fluorescence anisotropy assays using full length and C-terminally truncated enzymes. We find that the C-terminal domain of topoisomerase IIβ but not topoisomerase IIα affects the binding of the enzyme to the DNA. The presence of metal ions has no effect on DNA binding. Additionally, we have examined strand passage of the full length and truncated enzymes in the presence of a number of supporting metal ions and find that there is no difference in relative decatenation between isoforms. We find that calcium and manganese, in addition to magnesium, can support strand passage by the human topoisomerase II enzymes. CONCLUSIONS/SIGNIFICANCE: The C-terminal domain of topoisomerase IIβ, but not that of topoisomerase IIα, alters the enzyme's K(D for DNA binding. This is consistent with previous data and may be related to the differential modes of action of the two isoforms in vivo. We also show strand passage with different supporting metal ions for human topoisomerase IIα or topoisomerase IIβ, either full length or C-terminally truncated. They all show the same preferences, whereby Mg > Ca > Mn.

  18. DMT of Multi-hop Cooperative Networks - Part II: Half-Duplex Networks with Full-Duplex Performance

    CERN Document Server

    Sreeram, K; Kumar, P Vijay

    2008-01-01

    We consider single-source single-sink (ss-ss) multi-hop relay networks, with slow-fading links and single-antenna half-duplex relay nodes. In a companion paper, we established some basic results which laid the foundation for the results presented here. In the present paper, we consider two families of networks of half-duplex networks. KPP networks may be viewed as the union of K node-disjoint parallel relaying paths. Generalizations of these networks include KPP(I) networks, which permit interference between paths and KPP(D) networks, which possess a direct link between source and sink. We characterize the DMT of these families of networks completely and show that they can achieve the cut-set bound, thus proving that full-duplex performance can be obtained even in the presence of the half-duplex constraint. We then consider layered networks, and prove that a linear DMT between maximum diversity and maximum multiplexing gain is achievable. All protocols in this paper are explicit and use only amplify-and-forwa...

  19. Raman study of the effects of polyamines on DNA:spermine and histamine

    Science.gov (United States)

    Ruiz-Chica, J.; Medina, M. A.; Sánchez-Jiménez, F.; Ramírez, F. J.

    1999-05-01

    Fourier transform Raman spectroscopy was used to investigate the interaction of spermine and histamine with calf-thymus DNA. Polyamine-DNA solutions at different polyamine concentrations ranging from 5 mM to 75 mM were prepared. For spermine, solutions no higher than 15 mM were prepared because this molecule induces condensation and aggregation on DNA at upper concentrations. Possible sites of bindings for polyamine-DNA complexes were discussed on the basis of the spectral changes observed with respect to the Raman spectra of DNA. The results seem to indicate that one spermine molecule induces on DNA a similar effect to two or more histamine molecules.

  20. Single-stranded oligonucleotide adducts formed by Pt complexes favoring left-handed base canting: steric effect of flanking residues and relevance to DNA adducts formed by Pt anticancer drugs.

    Science.gov (United States)

    Saad, Jamil S; Marzilli, Patricia A; Intini, Francesco P; Natile, Giovanni; Marzilli, Luigi G

    2011-09-01

    Platinum anticancer drug binding to DNA creates large distortions in the cross-link (G*G*) and the adjacent XG* base pair (bp) steps (G* = N7-platinated G). These distortions, which are responsible for anticancer activity, depend on features of the duplex (e.g., base pairing) and of the cross-link moiety (e.g., the position and canting of the G* bases). The duplex structure stabilizes the head-to-head (HH) over the head-to-tail (HT) orientation and right-handed (R) over left-handed (L) canting of the G* bases. To provide fundamental chemical information relevant to the assessment of such duplex effects, we examine (S,R,R,S)-BipPt(oligo) adducts (Bip = 2,2'-bipiperidine with S,R,R,S chiral centers at the N, C, C, and N chelate ring atoms, respectively; oligo = d(G*pG*) with 3'- and/or 5'-substituents). The moderately bulky (S,R,R,S)-Bip ligand favors L canting and slows rotation about the Pt-G* bonds, and the (S,R,R,S)-BipPt(oligo) models provide more useful data than do dynamic models derived from active Pt drugs. All 5'-substituents in (S,R,R,S)-BipPt(oligo) adducts favor the normal HH conformer (∼97%) by destabilizing the HT conformer through clashes with the 3'-G* residue rather than through favorable H-bonding interactions with the carrier ligand in the HH conformer. For all (S,R,R,S)-BipPt(oligo) adducts, the S pucker of the 5'-X residue is retained. For these adducts, a 5'-substituent had only modest effects on the degree of L canting for the (S,R,R,S)-BipPt(oligo) HH conformer. This small flanking 5'-substituent effect on an L-canted HH conformer contrasts with the significant decrease in the degree of R canting previously observed for flanking 5'-substituents in the R-canted (R,S,S,R)-BipPt(oligo) analogues. The present data support our earlier hypothesis that the distortion distinctive to the XG* bp step (S to N pucker change and movement of the X residue) is required for normal stacking and X·X' WC H bonding and to prevent XG* residue clashes.

  1. Crystal structure of the human FOXO3a-DBD/DNA complex suggests the effects of post-translational modification.

    Science.gov (United States)

    Tsai, Kuang-Lei; Sun, Yuh-Ju; Huang, Cheng-Yang; Yang, Jer-Yen; Hung, Mien-Chie; Hsiao, Chwan-Deng

    2007-01-01

    FOXO3a is a transcription factor of the FOXO family. The FOXO proteins participate in multiple signaling pathways, and their transcriptional activity is regulated by several post-translational mechanisms, including phosphorylation, acetylation and ubiquitination. Because these post-translational modification sites are located within the C-terminal basic region of the FOXO DNA-binding domain (FOXO-DBD), it is possible that these post-translational modifications could alter the DNA-binding characteristics. To understand how FOXO mediate transcriptional activity, we report here the 2.7 A crystal structure of the DNA-binding domain of FOXO3a (FOXO3a-DBD) bound to a 13-bp DNA duplex containing a FOXO consensus binding sequence (GTAAACA). Based on a unique structural feature in the C-terminal region and results from biochemical and mutational studies, our studies may explain how FOXO-DBD C-terminal phosphorylation by protein kinase B (PKB) or acetylation by cAMP-response element binding protein (CBP) can attenuate the DNA-binding activity and thereby reduce transcriptional activity of FOXO proteins. In addition, we demonstrate that the methyl groups of specific thymine bases within the consensus sequence are important for FOXO3a-DBD recognition of the consensus binding site.

  2. Rapid PCR amplification of DNA utilizing Coriolis effects.

    Science.gov (United States)

    Mårtensson, Gustaf; Skote, Martin; Malmqvist, Mats; Falk, Mats; Asp, Allan; Svanvik, Nicke; Johansson, Arne

    2006-08-01

    A novel polymerase chain reaction (PCR) method is presented that utilizes Coriolis and centrifugal effects, produced by rotation of the sample disc, in order to increase internal circulatory rates, and with them temperature homogenization and mixing speeds. A proof of concept has been presented by testing a rapid 45-cycle PCR DNA amplification protocol. During the repeated heating and cooling that constitutes a PCR process, the 100 microL samples were rotated at a speed equivalent to an effective acceleration of gravity of 7,000 g. A cycle time of 20.5 s gave a total process time of 15 min to complete the 45 cycles. A theoretical and numerical analysis of the resulting flow, which describes the increased mixing and temperature homogenization, is presented. The device gives excellent reaction speed efficiency, which is beneficial for rapid PCR.

  3. EFFECTS OF METAL IONS ON THE CONFORMATIONAL CHANGES OF DNA

    Institute of Scientific and Technical Information of China (English)

    G. Q. Liu; Y.Y. Meng; S.H. Liu; Y.H. Hu

    2005-01-01

    DNA takes on multi-different conformations such as A-, B-, C-, D- and Z-form. These conformations can transit to one another when DNA deposited in some metal ions solutions or when changing the concentrations of the same metal ions. Here, several major conformational transitions of DNA induced by metal ions under different environment were introduced and the mechanism of the interaction of metal ions with DNA was discuss in detail.

  4. The Formation of Martensitic Austenite During Nitridation of Martensitic and Duplex Stainless Steels

    Science.gov (United States)

    Zangiabadi, Amirali; Dalton, John C.; Wang, Danqi; Ernst, Frank; Heuer, Arthur H.

    2017-01-01

    Isothermal martensite/ferrite-to-austenite phase transformations have been observed after low-temperature nitridation in the martensite and δ-ferrite phases in 15-5 PH (precipitation hardening), 17-7 PH, and 2205 (duplex) stainless steels. These transformations, in the region with nitrogen concentrations of 8 to 16 at. pct, are consistent with the notion that nitrogen is a strong austenite stabilizer and substitutional diffusion is effectively frozen at the paraequilibrium temperatures of our experiments. Our microstructural and diffraction analyses provide conclusive evidence for the martensitic nature of these phase transformations.

  5. The effect of ancient DNA damage on inferences of demographic histories

    DEFF Research Database (Denmark)

    Axelsson, Erik; Willerslev, Eske; Gilbert, Marcus Thomas Pius

    2008-01-01

    The field of ancient DNA (aDNA) is casting new light on many evolutionary questions. However, problems associated with the postmortem instability of DNA may complicate the interpretation of aDNA data. For example, in population genetic studies, the inclusion of damaged DNA may inflate estimates...... of diversity. In this paper, we examine the effect of DNA damage on population genetic estimates of ancestral population size. We simulate data using standard coalescent simulations that include postmortem damage and show that estimates of effective population sizes are inflated around, or right after......, the sampling time of the ancestral DNA sequences. This bias leads to estimates of increasing, and then decreasing, population sizes, as observed in several recently published studies. We reanalyze a recently published data set of DNA sequences from the Bison (Bison bison/Bison priscus) and show that the signal...

  6. DNA adsorption measured with ultra-thin film organic field effect transistors

    NARCIS (Netherlands)

    Stoliar, P.; Bystrenova, E.; Quiroga, S.D.; Annibale, P.; Facchini, M.; Spijkman, M.; Setayesh, S.; Leeuw, D. de; Biscarini, F.

    2009-01-01

    Organic ultra-thin film field effect transistors (FET) are operated as label-free sensors of deoxyribonucleic acid (DNA) adsorption. Linearized plasmid DNA molecules (4361 base pairs) are deposited froma solution on two monolayers thick pentacene FET. The amount of adsorbed DNA is measured by AFM an

  7. Synthesis and structural characterization of piperazino-modified DNA that favours hybridization towards DNA over RNA

    DEFF Research Database (Denmark)

    Skov, Joan; Bryld, Torsten; Lindegaard, Dorthe

    2011-01-01

    We report the synthesis of two C4'-modified DNA analogues and characterize their structural impact on dsDNA duplexes. The 4'-C-piperazinomethyl modification stabilizes dsDNA by up to 5°C per incorporation. Extension of the modification with a butanoyl-linked pyrene increases the dsDNA stabilization...... modifications are tolerated in DNA:RNA hybrids but leave their melting temperatures virtually unaffected. Fluorescence data indicate that the pyrene moiety is residing outside the helix. The available data suggest that the DNA discrimination is due to (i) the positive charge of the piperazino ring having...... to a maximum of 9°C per incorporation. Using fluorescence, ultraviolet and nuclear magnetic resonance (NMR) spectroscopy, we show that the stabilization is achieved by pyrene intercalation in the dsDNA duplex. The pyrene moiety is not restricted to one intercalation site but rather switches between multiple...

  8. Effects of DNA replication on mRNA noise.

    Science.gov (United States)

    Peterson, Joseph R; Cole, John A; Fei, Jingyi; Ha, Taekjip; Luthey-Schulten, Zaida A

    2015-12-29

    There are several sources of fluctuations in gene expression. Here we study the effects of time-dependent DNA replication, itself a tightly controlled process, on noise in mRNA levels. Stochastic simulations of constitutive and regulated gene expression are used to analyze the time-averaged mean and variation in each case. The simulations demonstrate that to capture mRNA distributions correctly, chromosome replication must be realistically modeled. Slow relaxation of mRNA from the low copy number steady state before gene replication to the high steady state after replication is set by the transcript's half-life and contributes significantly to the shape of the mRNA distribution. Consequently both the intrinsic kinetics and the gene location play an important role in accounting for the mRNA average and variance. Exact analytic expressions for moments of the mRNA distributions that depend on the DNA copy number, gene location, cell doubling time, and the rates of transcription and degradation are derived for the case of constitutive expression and subsequently extended to provide approximate corrections for regulated expression and RNA polymerase variability. Comparisons of the simulated models and analytical expressions to experimentally measured mRNA distributions show that they better capture the physics of the system than previous theories.

  9. Label-Free Potentiometry for Detecting DNA Hybridization Using Peptide Nucleic Acid and DNA Probes

    Directory of Open Access Journals (Sweden)

    Yuji Miyahara

    2013-02-01

    Full Text Available Peptide nucleic acid (PNA has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry.

  10. Label-free potentiometry for detecting DNA hybridization using peptide nucleic acid and DNA probes.

    Science.gov (United States)

    Goda, Tatsuro; Singi, Ankit Balram; Maeda, Yasuhiro; Matsumoto, Akira; Torimura, Masaki; Aoki, Hiroshi; Miyahara, Yuji

    2013-02-07

    Peptide nucleic acid (PNA) has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM)-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry.

  11. The Inhibitory Effect of Non-Substrate and Substrate DNA on the Ligation and Self-Adenylylation Reactions Catalyzed by T4 DNA Ligase.

    Directory of Open Access Journals (Sweden)

    Robert J Bauer

    Full Text Available DNA ligases are essential both to in vivo replication, repair and recombination processes, and in vitro molecular biology protocols. Prior characterization of DNA ligases through gel shift assays has shown the presence of a nick site to be essential for tight binding between the enzyme and its dsDNA substrate, with no interaction evident on dsDNA lacking a nick. In the current study, we observed a significant substrate inhibition effect, as well as the inhibition of both the self-adenylylation and nick-sealing steps of T4 DNA ligase by non-nicked, non-substrate dsDNA. Inhibition by non-substrate DNA was dependent only on the total DNA concentration rather than the structure; with 1 μg/mL of 40-mers, 75-mers, or circular plasmid DNA all inhibiting ligation equally. A >15-fold reduction in T4 DNA ligase self-adenylylation rate when in the presence of high non-nicked dsDNA concentrations was observed. Finally, EMSAs were utilized to demonstrate that non-substrate dsDNA can compete with nicked dsDNA substrates for enzyme binding. Based upon these data, we hypothesize the inhibition of T4 DNA ligase by non-nicked dsDNA is direct evidence for a two-step nick-binding mechanism, with an initial, nick-independent, transient dsDNA-binding event preceding a transition to a stable binding complex in the presence of a nick site.

  12. The Inhibitory Effect of Non-Substrate and Substrate DNA on the Ligation and Self-Adenylylation Reactions Catalyzed by T4 DNA Ligase.

    Science.gov (United States)

    Bauer, Robert J; Evans, Thomas C; Lohman, Gregory J S

    2016-01-01

    DNA ligases are essential both to in vivo replication, repair and recombination processes, and in vitro molecular biology protocols. Prior characterization of DNA ligases through gel shift assays has shown the presence of a nick site to be essential for tight binding between the enzyme and its dsDNA substrate, with no interaction evident on dsDNA lacking a nick. In the current study, we observed a significant substrate inhibition effect, as well as the inhibition of both the self-adenylylation and nick-sealing steps of T4 DNA ligase by non-nicked, non-substrate dsDNA. Inhibition by non-substrate DNA was dependent only on the total DNA concentration rather than the structure; with 1 μg/mL of 40-mers, 75-mers, or circular plasmid DNA all inhibiting ligation equally. A >15-fold reduction in T4 DNA ligase self-adenylylation rate when in the presence of high non-nicked dsDNA concentrations was observed. Finally, EMSAs were utilized to demonstrate that non-substrate dsDNA can compete with nicked dsDNA substrates for enzyme binding. Based upon these data, we hypothesize the inhibition of T4 DNA ligase by non-nicked dsDNA is direct evidence for a two-step nick-binding mechanism, with an initial, nick-independent, transient dsDNA-binding event preceding a transition to a stable binding complex in the presence of a nick site.

  13. Theoretical Study on Effects of Salt and Temperature on Denaturation Transition of Double-stranded DNA

    Institute of Scientific and Technical Information of China (English)

    DONG Rui-Xin; YAN Xun-Ling; PANG Xiao-Feng; JIANG Shan; LIU Sheng-Gang

    2004-01-01

    We investigate the statistical mechanics properties of a nonlinear dynamics model of the denaturation of the DNA double-helix and study the effects of salt concentration and temperature on denaturation transition of DNA. The specific heat, entropy, and denaturation temperature of the system versus salt concentration are obtained. These results show that the denaturation of DNA not only depends on the temperature but also is influenced by the salt concentration in the solution of DNA, which are in agreement with experimental measurement.

  14. A model for parallel triple helix formation by RecA: single-single association with a homologous duplex via the minor groove.

    Science.gov (United States)

    Bertucat, G; Lavery, R; Prévost, C

    1998-12-01

    The nucleoproteic filaments of RecA polymerized on single stranded DNA are able to integrate double stranded DNA in a coaxial arrangement (with DNA stretched by a factor 1.5), to recognize homologous sequences in the duplex and to perform strand exchange between the single stranded and double stranded molecules. While experimental results favor the hypothesis of an invasion of the minor groove of the duplex by the single strand, parallel minor groove triple helices have never been isolated or even modeled, the minor groove offering little space for a third strand to interact. Based on an internal coordinate modeling study, we show here that such a structure is perfectly conceivable when the two interacting oligomers are stretched by a factor 1.5, in order to open the minor groove of the duplex. The model helix presents characteristics that coincide with known experimental data on unwinding, base pair inclination and inter-proton distances. Moreover, we show that extension and unwinding stabilize the triple helix. New patterns of triplet interaction via the minor groove are presented.

  15. Hole polarons in poly(G)-poly(C) and poly(A)-poly(T) DNA molecules

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The polaron might play an important role in the process of charge migration through duplex DNA stack. In the present work, we investigate properties of hole polarons in DNA molecules containing identical base pairs, such as poly(G)-poly(C) and poly(A)-poly(T), An extended tight-binding model (extended Su-Schrieffer-Heeger model), which involves the effect of an electric field in the direction of DNA stack, will be introduced. The transfer integral and electron-phonon coupling parameters in this model are obtained according to ab initio calculation for different base pair dimers. Calculations reveal that the polaron in poly(A)-poly(T) has a wider shape and a higher mobility under a specific electric field than that in poly(G)-poly(C) DNA stack.

  16. Investigation Into the Effects of Nucleotide Content on the Electrical Characteristics of DNA Plasmid Molecular Wires.

    Science.gov (United States)

    Goshi, Noah; Narenji, Alaleh; Bui, Chris; Mokili, John L; Kassegne, Sam

    2016-09-01

    In this study, we investigate the effect of nucleotide content on the conductivity of plasmid length DNA molecular wires covalently bound to high aspect-ratio gold electrodes. The DNA wires were all between [Formula: see text] in length (>6000bp), and contained either 39%, 53%, or 64% GC base-pairs. We compared the current-voltage (I-V) and frequency-impedance characteristics of the DNA wires with varying GC content, and observed statistically significantly higher conductivity in DNA wires containing higher GC content in both AC and DC measurement methods. Additionally, we noted that the conductivity decreased as a function of time for all DNA wires, with the impedance at 100 Hz nearly doubling over a period of seven days. All readings were taken in humidity and temperature controlled environments on DNA wires suspended above an insulative substrate, thus minimizing the effect of experimental and environmental factors as well as potential for nonlinear alternate DNA confirmations. While other groups have studied the effect of GC content on the conductivity of nanoscale DNA molecules (DNA wires at scales that may be required during the fabrication of DNA-based electronics. Furthermore, our results provide further evidence that many of the charge transfer theories developed from experiments using nanoscale DNA molecules may still be applicable for DNA wires at the micro scale.

  17. Investigation of Effects of Nucleotide Content on Electrical Characteristics of DNA Plasmid Molecular Wires.

    Science.gov (United States)

    Goshi, Noah; Narenji, Alaleh; Bui, Chris; Mokili, John L; Kassegne, Sam

    2016-07-28

    In this study, we investigate the effect of nucleotide content on the conductivity of plasmid length DNA molecular wires covalently bound to high aspect-ratio gold electrodes. The DNA wires were all between 2.20-2.35μm in length (>6000bp), and contained either 39%, 53%, or 64% GC base-pairs. We compared the current-voltage (I-V) and frequency-impedance characteristics of the DNA wires with varying GC content, and observed statistically significantly higher conductivity in DNA wires containing higher GC content in both AC and DC measurement methods. Additionally, we noted that the conductivity decreased as a function of time for all DNA wires, with the impedance at 100Hz nearly doubling over a period of seven days. All readings were taken in humidity and temperature controlled environments on DNA wires suspended above an insulative substrate, thus minimizing the effect of experimental and environmental factors as well as potential for nonlinear alternate DNA confirmations. While other groups have studied the effect of GC content on the conductivity of nano-scale DNA molecules (DNA wires at scales that may be required during the fabrication of DNA-based electronics. Furthermore, our results provide further evidence that many of the charge transfer theories developed from experiments using nano-scale DNA molecules may still be applicable for DNA wires at the micro-scale.

  18. Expression, purification and biochemical characterization of Methanocaldococcus jannaschii DNA ligase.

    Science.gov (United States)

    Wang, You; Xie, Juan-Juan; Han, Zhong; Liu, Jian-Hua; Liu, Xi-Peng

    2013-02-01

    We describe the biochemical characterization of Methanocaldococcus jannaschii (M. jannaschii) DNA ligase and its potential application in single nucleotide polymorphism (SNP) genotyping. The recombinant M. jannaschii DNA ligase is an ATP-dependent ligase. The ligase activity was dependent on metal ions of Mg(2+) and Mn(2+). The optimal concentrations of ATP cofactor and Mg(2+) ion were 0.01-2 and 10 mM, respectively. The optimal pH value for DNA ligation was 8.5. High concentrations of NaCl inhibited DNA ligation. The effects of mismatches on joining short oligonucleotides by M. jannaschii DNA ligase were fully characterized. The mismatches at the first position 5' to the nick inhibited ligation more than those at the first position 3' to the nick. The mismatches at other positions 5' to the nick (3rd to 7th sites) exhibited less inhibition on ligation. However, the introduction of a C/C mismatch at the third position 5' to the nick could completely inhibit the ligation of the terminal-mismatched nick of an oligonucleotide duplex by M. jannaschii DNA ligase. Therefore, introducing an additional mismatch at the third position 5' to the SNP site is a more effective approach in genotyping by M. jannaschii DNA ligase.

  19. Massive parallel analysis of the binding specificity of histone-like protein HU to single- and double-stranded DNA with generic oligodeoxyribonucleotide microchips.

    Energy Technology Data Exchange (ETDEWEB)

    Krylov, A. S.; Zasedateleva, O. A.; Prokopenko, D. V.; Rouviere-Yaniv, J.; Mirzabekov, A. D.; Biochip Technology Center; Engelhardt Inst. of Molecular Biology; Inst. de Biologie Physico-Chimique

    2001-06-15

    A generic hexadeoxyribonucleotide microchip has been applied to test the DNA-binding properties of HU histone-like bacterial protein, which is known to have a low sequence specificity. All 4096 hexamers flanked within 8mers by degenerate bases at both the 3'- and 5'-ends were immobilized within the 100 x 100 x 20 mm polyacrylamide gel pads of the microchip. Single-stranded immobilized oligonucleotides were converted in some experiments to the double-stranded form by hybridization with a specified mixture of 8mers. The DNA interaction with HU was characterized by three type of measurements: (i) binding of FITC-labeled HU to microchip oligonucleotides; (ii) melting curves of complexes of labeled HU with single-stranded microchip oligonucleotides; (iii) the effect of HU binding on melting curves of microchip double-stranded DNA labeled with another fluorescent dye, Texas Red. Large numbers of measurements of these parameters were carried out in parallel for all or many generic microchip elements in real time with a multi-wavelength fluorescence microscope. Statistical analysis of these data suggests some preference for HU binding to G/C-rich single-stranded oligonucleotides. HU complexes with double-stranded microchip 8mers can be divided into two groups in which HU binding either increased the melting temperature (T{sub m}) of duplexes or decreased it. The stabilized duplexes showed some preference for presence of the sequence motifs AAG, AGA and AAGA. In the second type of complex, enriched with A/T base pairs, the destabilization effect was higher for longer stretches of A/T duplexes. Binding of HU to labeled duplexes in the second type of complex caused some decrease in fluorescence. This decrease also correlates with the higher A/T content and lower T{sub m}. The results demonstrate that generic microchips could be an efficient approach in analysis of sequence specificity of proteins.

  20. Short sequence effect of ancient DNA on mammoth phylogenetic analyses

    Institute of Scientific and Technical Information of China (English)

    Guilian SHENG; Lianjuan WU; Xindong HOU; Junxia YUAN; Shenghong CHENG; Bojian ZHONG; Xulong LAI

    2009-01-01

    The evolution of Elephantidae has been intensively studied in the past few years, especially after 2006. The molecular approaches have made great contribution to the assumption that the extinct woolly mammoth has a close relationship with the Asian elephant instead of the African elephant. In this study, partial ancient DNA sequences of cytochrome b (cyt b) gene in mitochondrial genome were successfully retrieved from Late Pleistocene Mammuthus primigenius bones collected from Heilongjiang Province in Northeast China. Both the partial and complete homologous cyt b gene sequences and the whole mitochondrial genome sequences extracted from GenBank were aligned and used as datasets for phylogenetic analyses. All of the phylogenetic trees, based on either the partial or the complete cyt b gene, reject the relationship constructed by the whole mitochondrial genome, showing the occurrence of an effect of sequence length of cyt b gene on mammoth phylogenetic analyses.

  1. Investigation of corrosion and wear mechanisms in hard material-reinforced duplex steel coatings; Untersuchungen zum Korrosions- und Verschleissverhalten von hartstoffverstaerkten `Duplex`-Schutzschichten. Schlussbericht

    Energy Technology Data Exchange (ETDEWEB)

    Bouaifi, B. [Technische Univ. Clausthal, Clausthal-Zellerfeld (Germany). Inst. fuer Schweisstechnik und Trennende Fertigungsverfahren; Goellner, J. [Technische Univ. Magdeburg (Germany). Inst. fuer Werkstofftechnik und Werkstoffpruefung

    1998-09-30

    The hard-material reinforced duplex steel coatings were deposited by plasma arc two-powder surfacing. By varying the angle of the hard materials feeding process, the deposition of the carbides was optimized so that they are deposited into the matrix in the trailing zone of the welding torch under conditions of very low thermal effects near the freezing point of the weld pool. Microstructural studies revealed that the deposition of the hard materials prevents devlopment of the typical, ferritic-austenitic microstructure of duplex steels. Due to a dissolution and diffusion process, the microstructure of the matrix takes up carbon and chromium or tungsten, depending on the carbide, thus enhancing the austenitic material in the microstructure. The wear behaviour of the surface deposits was found to be very good, wear being reduced by a factor of 6, irrespective of the type of carbide. The friction-affected surfaces showed no dissolving effects or cracking. The corrosion behaviour in sulfuric acid is also good. The welded deposits exhibited the typical behaviour of a passive material. Wear mechanisms slightly reduce the resistance. The behaviour of various specimens in artificial seawater could be distinctly assessed. Small grain fractions and pre-heating temperatures of 100 C have a beneficial effect on the corrosion resistance. The technique recommends itself for applications such as coatings for baffle plates used in flue gas desulfurisation, pipes, pump components, flanges or nozzles, or for recurrent coating of system components affected by abrasive corrosion. (orig./CB) [Deutsch] Zur Herstellung hartstoffverstaerkter Duplex-Beschichtungen wurde das Plasma-Zwei-Pulver-Auftragschweissverfahren verwendet. Hierbei wurde durch Variation des Zufuhrwinkels der Hartstoffe der Einbringungsort der Karbide optimiert, so dass diese im Nachlauf des Schweissbrenners mit sehr geringer thermischer Beeinflussung im erstarrungsnahen Schmelzbadbereich in die Matrix eingelagert

  2. Left-handed DNA crossovers. Implications for DNA-DNA recognition and structural alterations.

    Science.gov (United States)

    Timsit, Y; Shatzky-Schwartz, M; Shakked, Z

    1999-02-01

    The close approach of DNA segments participates in many biological functions including DNA condensation and DNA processing. Previous crystallographic studies have shown that B-DNA self-fitting by mutual groove-backbone interaction produces right-handed DNA crossovers. These structures have opened new perspectives on the role of close DNA-DNA interactions in the architecture and activity the DNA molecule. In the present study, the analysis of the crystal packing of two B-DNA decamer duplexes d(CCIIICCCGG) and d(CCGCCGGCGG) reveals the existence of new modes of DNA crossing. Symmetric left-handed crossovers are produced by mutual fitting of DNA grooves at the crossing point. New sequence patterns contribute to stabilize longitudinal fitting of the sugar-phosphate backbone into the major groove. In addition, the close approach of DNA segments greatly influences the DNA conformation in a sequence dependent manner. This study provides new insights into the role of DNA sequence and structure in DNA-DNA recognition. In providing detailed molecular views of DNA crossovers of opposite chirality, this study can also help to elucidate the role of symmetry and chirality in the recognition of complex DNA structures by protein dimers or tetramers, such as topoisomerase II and recombinase enzymes. These results are discussed in the context of the possible relationships between DNA condensation and DNA processing.

  3. Effect of Cisplatin on the Flexibility of Linear DNA

    Institute of Scientific and Technical Information of China (English)

    JI Chao; ZHANG Ling-Yun; HOU Xi-Miao; DOU Shuo-Xing; WANG Peng-Ye

    2011-01-01

    With the aid of an atomic force microscope (AFM), we study the interaction between linear DNA fragment and cisplatin. For different cisplatin concentrations, the AFM used to observe the conformation of DNA has a gradual change. The contour length, the end-to-end distance and the local bend angles of the linear DNA fragment can be accurately measured. The persistence length of DNA interacting with cisplatin is decreased with the increasing cisplatin concentration. Furthermore, it is demonstrated that the local bend angles of DNA chains are increased by the binding interaction of cisplatin.%@@ With the aid of an atomic force microscope (AFM), we study the interaction between linear DNA fragment and cisplatin.For different cisplatin concentrations, the AFM used to observe the conformation of DNA has a gradual change.The contour length, the end-to-end distance and the local bend angles of the linear DNA fragment can be accurately measured.The persistence length of DNA interacting with cisplatin is decreased with the increasing cisplatin concentration.Furthermore, it is demonstrated that the local bend angles of DNA chains are increased by the binding interaction of cisplatin.

  4. Detection of influenza A virus using carbon nanotubes field effect transistor based DNA sensor

    Science.gov (United States)

    Tran, Thi Luyen; Nguyen, Thi Thuy; Huyen Tran, Thi Thu; Chu, Van Tuan; Thinh Tran, Quang; Tuan Mai, Anh

    2017-09-01

    The carbon nanotubes field effect transistor (CNTFET) based DNA sensor was developed, in this paper, for detection of influenza A virus DNA. Number of factors that influence the output signal and analytical results were investigated. The initial probe DNA, decides the available DNA strands on CNTs, was 10 μM. The hybridization time for defined single helix was 120 min. The hybridization temperature was set at 30 °C to get a net change in drain current of the DNA sensor without altering properties of any biological compounds. The response time of the DNA sensor was less than one minute with a high reproducibility. In addition, the DNA sensor has a wide linear detection range from 1 pM to 10 nM, and a very low detection limit of 1 pM. Finally, after 7-month storage in 7.4 pH buffer, the output signal of DNA sensor recovered 97%.

  5. Effect of food processing on plant DNA degradation and PCR-based GMO analysis: a review.

    Science.gov (United States)

    Gryson, Nicolas

    2010-03-01

    The applicability of a DNA-based method for GMO detection and quantification depends on the quality and quantity of the DNA. Important food-processing conditions, for example temperature and pH, may lead to degradation of the DNA, rendering PCR analysis impossible or GMO quantification unreliable. This review discusses the effect of several food processes on DNA degradation and subsequent GMO detection and quantification. The data show that, although many of these processes do indeed lead to the fragmentation of DNA, amplification of the DNA may still be possible. Length and composition of the amplicon may, however, affect the result, as also may the method of extraction used. Also, many techniques are used to describe the behaviour of DNA in food processing, which occasionally makes it difficult to compare research results. Further research should be aimed at defining ingredients in terms of their DNA quality and PCR amplification ability, and elaboration of matrix-specific certified reference materials.

  6. Non destructive method to follow the phase sigma in a duplex stainless steel; Metodologia nao destrutiva para acompanhamento da fase sigma, em um aco inoxidavel duplex

    Energy Technology Data Exchange (ETDEWEB)

    Silva, E.M.; Andrade, A.L.S. Souza; Fialho, W.M.L.; Araujo, B.R., E-mail: edgard@ifpb.edu.br [Instituto Federal de Educacao Ciencia e Tecnologia da Paraiba (IFPB), Joao Pessoa, PB (Brazil); Silva, J.H.R.; Leite, Josinaldo P.; Silva, Eloy M. [Instituto Federal de Educacao Ciencia e Tecnologia do Ceara (IFCE), CE (Brazil); Leite, Joao P. [Universidade Federal da Paraiba (UFPB), PB (Brazil)

    2014-07-01

    Duplex stainless steels are subject to embrittlement due to the formation of sigma phase, which is one with the greatest effect of weakening because they are rich in chromium and deplete the matrix of this element. In this paper, a non-destructive methodology based on measurements of Hall voltage, is presented for monitoring the formation of sigma phase at temperatures of 800 deg C and 900 deg C. Different field intensities are generated by an electromagnet and the flow of field lines is detected by a Hall effect sensor. Hall voltage measurements are proportional to the formation of sigma phase generated by different times of aging methods. The results are correlated with results of microscopic, hardness and X-ray diffraction. It was showed that exist a correlation between the Hall voltage and the amount of sigma phase. The formation of this phase influences the signal voltage by reducing the voltage. (author)

  7. Method for construction of normalized cDNA libraries

    Science.gov (United States)

    Soares, Marcelo B.; Efstratiadis, Argiris

    1998-01-01

    This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to appropriate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library. This invention also provides normalized cDNA libraries generated by the above-described method and uses of the generated libraries.

  8. Development of a one-step duplex RT-qPCR for the quantification of phocine distemper virus.

    Science.gov (United States)

    Bogomolni, Andrea L; Frasca, Salvatore; Matassa, Keith A; Nielsen, Ole; Rogers, Kara; De Guise, Sylvain

    2015-04-01

    Worldwide, stranded marine mammals and the network personnel who respond to marine mammal mortality have provided much of the information regarding marine morbillivirus infections. An assay to determine the amount of virus present in tissue samples would be useful to assist in routine surveying of animal health and for monitoring large-scale die-off events. False negatives from poor-quality samples prevent determination of the true extent of infection, while only small amounts of tissue samples or archived RNA may be available at the time of collection for future retrospective analysis. We developed a one-step duplex real-time reverse transcriptase-quantitative-PCR assay (RT-qPCR) based on Taqman probe technology to quantify phocine distemper virus (PDV) isolated from an outbreak in harbor (Phoca vitulina concolor) and gray seals (Halichoerus grypus) along the northeast US coast in 2006. The glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) gene was selected to assess RNA quality. This duplex assay is specific for PDV and sensitive through a range of 10(0) to 10(9) copies ds-plasmid DNA. For the GAPDH target, the reaction in duplex amplified 10(0) to 10(9) copies of ds-plasmid DNA and was detectable in multiple seal species. This assay reduced the likelihood of false negative results due to degradation of tissues and well-to-well variability while providing sensitive and specific detection of PDV, which would be applicable in molecular epidemiologic studies and pathogen detection in field and laboratory investigations involving a variety of seal species.

  9. CHARACTERIZATION AND PERFORMANCE OF DUPLEX-COATINGS ON Cr-V COLD WORK TOOL STEEL

    Directory of Open Access Journals (Sweden)

    Peter Jurči

    2015-09-01

    Full Text Available Specimens made of Vanadis 6 steel were heat treated, plasma nitrided and coated with Cr2N. The microstructure, phase constitution and mechanical properties of plasma nitrided areas and duplex-coatings have been investigated using the light microscopy, scanning electron microscopy, X-ray diffraction and microhardness measurements. The adhesion of the coatings and the wear performance were studied using the scratch test and ring-on-plate tribological testing. Worn surfaces were examined by scanning electron microscopy. Nitrided areas formed at lower temperature were free of compound “white” layer while hose developed at higher temperatures contained as the white layer so the nitrided network. Significant increase in substrate hardness was detected due to the nitriding. Beneficial effect of the nitriding on the adhesion of Cr2N coatings was clearly determined whereas the extent in improvement of the adhesion depends on the presence/no presence of “white” layer on the surface. The extent of beneficial effect of plasma nitriding on the wear performance follows the impact of the constitution of nitrided areas on the adhesion. The amelioration of wear performance of Cr2N coatings can be attributed to the supporting effect of hard nitrided intermediate region, which provides excellent resistance of the substrate against plastic deformation, under heavy loading in particular. Practical testing demonstrated many times prolonged service-time of duplex-treated tools for sheet metal working.

  10. Robust and accurate visual echo cancellation in a full-duplex projector-camera system.

    Science.gov (United States)

    Liao, Miao; Yang, Ruigang; Zhang, Zhengyou

    2008-10-01

    In this paper we study the problem of "visual echo" in a full-duplex projector-camera system for telecollaboration applications. Visual echo is defined as the appearance of projected contents observed by the camera. It can potentially saturate the projected contents, similar to audio echo in telephone conversation. Our approach to visual echo cancellation includes an offline calibration procedure that records the geometric and photometric transfer between the projector and the camera in a look-up table. During run-time, projected contents in the captured video are identified using the calibration information and suppressed, therefore achieving the goal of cancelling visual echo. Our approach can accurately handle full-color images under arbitrary reflectance of display surfaces and photometric response of the projector or camera. It is robust to geometric registration errors and quantization effects and is therefore particularly effective for high-frequency contents such as texts and hand drawings. We demonstrate the effectiveness of our approach with a variety of real images in a full-duplex projector-camera system.

  11. Comments on process of duplex coatings on aluminum alloys

    Institute of Scientific and Technical Information of China (English)

    Samir H.A.; QIAN Han-cheng(钱翰城); XIA Bo-cai(夏伯才); WU Shi-ming(吴仕明)

    2004-01-01

    Despite the great achievements made in improvement of wear resistance properties of aluminum alloys,their applications in heavy surface load-bearing are limited. Single coating is insufficient to produce the desired combination of surface properties. These problems can be solved through the duplex coatings. The aim of the present study is to overview the research advances on processes of duplex coatings on aluminum alloys combined with micro plasma oxidation process and with other modern processes such as physical vapour deposition and plasma assisted chemical vapour deposition and also to evaluate the performance of micro plasma oxidation coatings in improving the load-bearing, friction and wear resistance properties of aluminum alloys in comparison with other coatings. Wherein, a more detailed presentation of the processes and their performances and disadvantages are given as well.

  12. Herpes Zoster Duplex Unilateralis: Two Cases and Brief Literature Review

    Science.gov (United States)

    Son, Jee Hee; Chung, Bo Young; Kim, Hye One; Cho, Hee Jin

    2016-01-01

    Cases involving dermatomal herpes zoster in two or more locations are rare, especially in immunocompetent patients. When two noncontiguous dermatomes are involved, if affected unilaterally, it is called herpes zoster duplex unilateralis; if bilaterally, bilateralis. Here, we report two cases of herpes zoster duplex unilateralis. A 66-year-old man presented with painful erythematous grouped vesicles on his left scalp, forehead, trunk, and back (left [Lt.] V1, Lt. T8). Histologic findings were consistent with herpetic infection. A 33-year-old woman presented with painful erythematous grouped vesicles and crust on her left forehead and neck (Lt. V1, Lt. C5). Both patients were treated with oral administration of famcyclovir 750 mg/day for seven days. PMID:27904277

  13. Listen and talk full-duplex cognitive radio networks

    CERN Document Server

    Liao, Yun; Han, Zhu

    2016-01-01

    This brief focuses on the use of full-duplex radio in cognitive radio networks, presenting a novel spectrum sharing protocol that allows the secondary users to simultaneously sense and access the vacant spectrum. This protocol, called “Listen-and-talk” (LAT), is evaluated by both mathematical analysis and computer simulations in comparison with other existing protocols, including the listen-before-talk protocol. In addition to LAT-based signal processing and resource allocation, the brief discusses techniques such as spectrum sensing and dynamic spectrum access. The brief proposes LAT as a suitable access scheme for cognitive radio networks, which can support the quality-of-service requirements of these high priority applications. Fundamental theories and key techniques of cognitive radio networks are also covered. Listen and Talk: Full-duplex Cognitive Radio Networks is designed for researchers, developers, and professionals involved in cognitive radio networks. Advanced-level students studying signal pr...

  14. A 2-order MIMO Full-Duplex Antenna System

    DEFF Research Database (Denmark)

    Tsakalaki, Elpiniki; Foroozanfard, Ehsan; De Carvalho, Elisabeth

    2014-01-01

    The paper presents an antenna system with combined full-duplex and 2-order multiple-input-multiple-output (MIMO) functionalities, i.e., a system capable of spatially multiplexing and spatially demultiplexing 2 datastreams in the same frequency and in the same time. By exploiting symmetries....... On the other hand, the 2 MIMO ports (either at the Tx or at the Rx) are sufficiently decoupled thanks to polarization diversity. The proposed antenna system exhibits a remarkable level of fullduplex isolation over a wide bandwidth while maintaining low coupling between its MIMO ports and can serve...... as a concrete implementation of an antenna system equipped with both MIMO as well as full-duplex capabilities....

  15. The ABCs of molecular dynamics simulations on B-DNA, circa 2012

    Indian Academy of Sciences (India)

    David L Beveridge; Thomas E Cheatham III; Mihaly Mezei

    2012-07-01

    This article provides a retrospective on the ABC initiative in the area of all-atom molecular dynamics (MD) simulations including explicit solvent on all tetranucleotide steps of duplex B-form DNA duplex, ca. 2012. The ABC consortium has completed two phases of simulations, the most current being a set of 50–100 trajectories based on the AMBER ff99 force field together with the parmbsc0 modification. Some general perspectives on the field of MD on DNA and sequence effects on DNA structure are provided, followed by an overview our MD results, including a detailed comparison of the ff99/parmbsc0 results with crystal and NMR structures available for d(CGCGAATTCGCG). Some projects inspired by or related to the ABC initiative and database are also reviewed, including methods for the trajectory analyses, informatics of dealing with the large database of results, compressions of trajectories for efficacy of distribution, DNA solvation by water and ions, parameterization of coarse-grained models with applications and gene finding and genome annotation

  16. Quantitative analysis of the ion-dependent folding stability of DNA triplexes.

    Science.gov (United States)

    Chen, Gengsheng; Chen, Shi-Jie

    2011-12-01

    A DNA triplex is formed through binding of a third strand to the major groove of a duplex. Due to the high charge density of a DNA triplex, metal ions are critical for its stability. We recently developed the tightly bound ion (TBI) model for ion-nucleic acids interactions. The model accounts for the potential correlation and fluctuations of the ion distribution. We now apply the TBI model to analyze the ion dependence of the thermodynamic stability for DNA triplexes. We focus on two experimentally studied systems: a 24-base DNA triplex and a pair of interacting 14-base triplexes. Our theoretical calculations for the number of bound ions indicate that the TBI model provides improved predictions for the number of bound ions than the classical Poisson-Boltzmann (PB) equation. The improvement is more significant for a triplex, which has a higher charge density than a duplex. This is possibly due to the higher ion concentration around the triplex and hence a stronger ion correlation effect for a triplex. In addition, our analysis for the free energy landscape for a pair of 14-mer triplexes immersed in an ionic solution shows that divalent ions could induce an attractive force between the triplexes. Furthermore, we investigate how the protonated cytosines in the triplexes affect the stability of the triplex helices.

  17. Influence of heat input on weld bead geometry using duplex stainless steel wire electrode on low alloy steel specimens

    OpenAIRE

    Ajit Mondal; Manas Kumar Saha; Ritesh Hazra; Santanu Das

    2016-01-01

    Gas metal arc welding cladding becomes a popular surfacing technique in many modern industries as it enhances effectively corrosion resistance property and wear resistance property of structural members. Quality of weld cladding may be enhanced by controlling process parameters. If bead formation is found acceptable, cladding is also expected to be good. Weld bead characteristics are often assessed by bead geometry, and it is mainly influenced by heat input. In this paper, duplex stainless st...

  18. DNA Ministrings: Highly Safe and Effective Gene Delivery Vectors

    Directory of Open Access Journals (Sweden)

    Nafiseh Nafissi

    2014-01-01

    Full Text Available Conventional plasmid DNA vectors play a significant role in gene therapy, but they also have considerable limitations: they can elicit adverse immune responses because of bacterial sequences they contain for maintenance and amplification in prokaryotes, their bioavailability is compromised because of their large molecular size, and they may be genotoxic. We constructed an in vivo platform to produce ministring DNA—mini linear covalently closed DNA vectors—that are devoid of unwanted bacterial sequences and encode only the gene(s of interest and necessary eukaryotic expression elements. Transfection of rapidly and slowly dividing human cells with ministring DNA coding for enhanced green fluorescent protein resulted in significantly improved transfection, bioavailability, and cytoplasmic kinetics compared with parental plasmid precursors and isogenic circular covalently closed DNA counterparts. Ministring DNA that integrated into the genome of human cells caused chromosomal disruption and apoptotic death of possibly oncogenic vector integrants; thus, they may be safer than plasmid and circular DNA vectors.

  19. Human DNA ligase III bridges two DNA ends to promote specific intermolecular DNA end joining.

    Science.gov (United States)

    Kukshal, Vandna; Kim, In-Kwon; Hura, Gregory L; Tomkinson, Alan E; Tainer, John A; Ellenberger, Tom

    2015-08-18

    Mammalian DNA ligase III (LigIII) functions in both nuclear and mitochondrial DNA metabolism. In the nucleus, LigIII has functional redundancy with DNA ligase I whereas LigIII is the only mitochondrial DNA ligase and is essential for the survival of cells dependent upon oxidative respiration. The unique LigIII zinc finger (ZnF) domain is not required for catalytic activity but senses DNA strand breaks and stimulates intermolecular ligation of two DNAs by an unknown mechanism. Consistent with this activity, LigIII acts in an alternative pathway of DNA double strand break repair that buttresses canonical non-homologous end joining (NHEJ) and is manifest in NHEJ-defective cancer cells, but how LigIII acts in joining intermolecular DNA ends versus nick ligation is unclear. To investigate how LigIII efficiently joins two DNAs, we developed a real-time, fluorescence-based assay of DNA bridging suitable for high-throughput screening. On a nicked duplex DNA substrate, the results reveal binding competition between the ZnF and the oligonucleotide/oligosaccharide-binding domain, one of three domains constituting the LigIII catalytic core. In contrast, these domains collaborate and are essential for formation of a DNA-bridging intermediate by adenylated LigIII that positions a pair of blunt-ended duplex DNAs for efficient and specific intermolecular ligation.

  20. Human DNA ligase III bridges two DNA ends to promote specific intermolecular DNA end joining

    Science.gov (United States)

    Kukshal, Vandna; Kim, In-Kwon; Hura, Gregory L.; Tomkinson, Alan E.; Tainer, John A.; Ellenberger, Tom

    2015-01-01

    Mammalian DNA ligase III (LigIII) functions in both nuclear and mitochondrial DNA metabolism. In the nucleus, LigIII has functional redundancy with DNA ligase I whereas LigIII is the only mitochondrial DNA ligase and is essential for the survival of cells dependent upon oxidative respiration. The unique LigIII zinc finger (ZnF) domain is not required for catalytic activity but senses DNA strand breaks and stimulates intermolecular ligation of two DNAs by an unknown mechanism. Consistent with this activity, LigIII acts in an alternative pathway of DNA double strand break repair that buttresses canonical non-homologous end joining (NHEJ) and is manifest in NHEJ-defective cancer cells, but how LigIII acts in joining intermolecular DNA ends versus nick ligation is unclear. To investigate how LigIII efficiently joins two DNAs, we developed a real-time, fluorescence-based assay of DNA bridging suitable for high-throughput screening. On a nicked duplex DNA substrate, the results reveal binding competition between the ZnF and the oligonucleotide/oligosaccharide-binding domain, one of three domains constituting the LigIII catalytic core. In contrast, these domains collaborate and are essential for formation of a DNA-bridging intermediate by adenylated LigIII that positions a pair of blunt-ended duplex DNAs for efficient and specific intermolecular ligation. PMID:26130724

  1. Quantification of human mitochondrial DNA using synthesized DNA standards.

    Science.gov (United States)

    Kavlick, Mark F; Lawrence, Helen S; Merritt, R Travis; Fisher, Constance; Isenberg, Alice; Robertson, James M; Budowle, Bruce

    2011-11-01

    Successful mitochondrial DNA (mtDNA) forensic analysis depends on sufficient quantity and quality of mtDNA. A real-time quantitative PCR assay was developed to assess such characteristics in a DNA sample, which utilizes a duplex, synthetic DNA to ensure optimal quality assurance and quality control. The assay's 105-base pair target sequence facilitates amplification of degraded DNA and is minimally homologous to nonhuman mtDNA. The primers and probe hybridize to a region that has relatively few sequence polymorphisms. The assay can also identify the presence of PCR inhibitors and thus indicate the need for sample repurification. The results show that the assay provides information down to 10 copies and provides a dynamic range spanning seven orders of magnitude. Additional experiments demonstrated that as few as 300 mtDNA copies resulted in successful hypervariable region amplification, information that permits sample conservation and optimized downstream PCR testing. The assay described is rapid, reliable, and robust.

  2. Implementation of real-time duplex synthetic aperture ultrasonography

    DEFF Research Database (Denmark)

    Hemmsen, Martin Christian; Larsen, Lee; Kjeldsen, Thomas;

    2015-01-01

    This paper presents a real-time duplex synthetic aperture imaging system, implemented on a commercially available tablet. This includes real-time wireless reception of ultrasound signals and GPU processing for B-mode and Color Flow Imaging (CFM). The objective of the work is to investigate the im...... and that the required bandwidth between the probe and processing unit is within the current Wi-Fi standards....

  3. Evolution of microstresses in plastically deformed duplex steel

    Energy Technology Data Exchange (ETDEWEB)

    Baczmanski, A.; Wierzbanowski, K. [Akademia Gorniczo-Hutnicza, Krakow (Poland). WFTJ; Braham, C. [LMMM, URA-CNRS 1219, Ecole Nationale Superieure d' Arts et Metiers, Paris (France); Lodini, A. [IFTS, Univ. de Reims Charleville-Mezieres (France)

    2000-07-01

    The X-ray diffraction method has been applied to determine the internal stresses in two phases austeno-ferritic steel during uniaxial tensile test. The elasto-plastic deformation model was used to predict internal stresses and critical resolved shear stress for the both phases. The model calculations were successfully compared with the information obtained from the shift and broadening of diffraction peak. Finally, the parameters characterising elasto-plastic deformation for duplex steel were determined. (orig.)

  4. Tensile properties of duplex UNS S32205 and lean duplex UNS S32304 steels and the influence of short duration 475 ºC aging

    Directory of Open Access Journals (Sweden)

    Sérgio Souto Maior Tavares

    2012-12-01

    Full Text Available Duplex stainless steels are high strength and corrosion resistant steels extensively used in the petrochemical and chemical industries. The aging at 475 ºC for long periods of time provokes embrittlement and deterioration of corrosion resistance. However, short duration aging at 475 ºC may be used as heat treatment to improve mechanical resistance with small decrease in the other properties. In this work the flow stress curves of lean duplex UNS S32304 and duplex UNS S32205 steels were modeled with Hollomon's equation and work hardening exponents (n were determined. The analyses were conducted in specimens annealed and heat treated at 475 ºC for short periods of time. The aging at 475 ºC for 4 hours, 8 hours and 12 hours promoted significant hardening with small decrease of ductility. The work hardening exponents of both steels were compared, being higher in the duplex steel than in the lean duplex grade.

  5. Effects of Sequence on Transmission Properties of DNA Molecules

    Institute of Scientific and Technical Information of China (English)

    DONG Rui-Xin; YAN Xun-Ling; YANG Bing

    2008-01-01

    A double helix model of charge transport in DNA molecule is given and the transmission spectra of four DNA sequences are obtained. The calculated results show that the transmission characteristics of DNA are not only related to the longitudinal transport but also to the transverse transport of molecule. The periodic sequence with the same composition has stronger conduction ability. With the increasing of bases composition, the conductive ability reduces, but the weight of θ direction rises in charge transfer.

  6. Duplex Schemes in Multiple Antenna Two-Hop Relaying

    Directory of Open Access Journals (Sweden)

    Anja Klein

    2008-04-01

    Full Text Available A novel scheme for two-hop relaying defined as space division duplex (SDD relaying is proposed. In SDD relaying, multiple antenna beamforming techniques are applied at the intermediate relay station (RS in order to separate downlink and uplink signals of a bi-directional two-hop communication between two nodes, namely, S1 and S2. For conventional amplify-and-forward two-hop relaying, there appears a loss in spectral efficiency due to the fact that the RS cannot receive and transmit simultaneously on the same channel resource. In SDD relaying, this loss in spectral efficiency is circumvented by giving up the strict separation of downlink and uplink signals by either time division duplex or frequency division duplex. Two novel concepts for the derivation of the linear beamforming filters at the RS are proposed; they can be designed either by a three-step or a one-step concept. In SDD relaying, receive signals at S1 are interfered by transmit signals of S1, and receive signals at S2 are interfered by transmit signals of S2. An efficient method in order to combat this kind of interference is proposed in this paper. Furthermore, it is shown how the overall spectral efficiency of SDD relaying can be improved if the channels from S1 and S2 to the RS have different qualities.

  7. Sensitivity of the green alga Pediastrum duplex Meyen to allelochemicals is strain-specific and not related to co-occurrence with allelopathic macrophytes.

    Science.gov (United States)

    Eigemann, Falk; Vanormelingen, Pieter; Hilt, Sabine

    2013-01-01

    Interspecific differences in the response of microalgae to stress have numerous ecological implications. However, little is known of intraspecific sensitivities and the potential role of local genetic adaptation of populations. We compared the allelochemical sensitivity of 23 Pediastrum duplex Meyen strains, a common component of the freshwater phytoplankton. In order to test for local genetic adaptation, strains were isolated from water bodies with and without the allelopathically-active submerged macrophyte Myriophyllum. Strains were assigned to P. duplex on the basis of cell shape and colony morphology and only P. duplex strains that belonged to the same lineage in an ITS rDNA phylogeny were used. Inhibition of strain growth rates and maximum quantum yields of photosystem II were measured after exposure to tannic acid (TA) and co-culture with Myriophyllum spicatum. Growth rate inhibition varied over one order of magnitude between the P. duplex strains. There was no correlation between the presence of Myriophyllum in the source location and the sensitivity of the strains to TA or the presence of Myriophyllum, suggesting that at least strong unidirectional local adaptation to Myriophyllum had not taken place in the studied water bodies. The maximum quantum yield of photosystem II of TA exposed algae decreased, whereas the yield of algae exposed to M. spicatum was slightly higher than that of the controls. The ranking of P. duplex strain sensitivities differed between the types of exposure (single additions of TA versus co-existence with M. spicatum) and the parameter measured (growth rate versus maximum quantum yield), emphasizing the importance of measuring multiple traits when analysing strain-specific sensitivities towards allelochemicals. The observation that sensitivities to allelochemicals vary widely among strains of a single freshwater algal species should be taken into account if evaluating ecological consequences of allelopathic interactions.

  8. Sensitivity of the green alga Pediastrum duplex Meyen to allelochemicals is strain-specific and not related to co-occurrence with allelopathic macrophytes.

    Directory of Open Access Journals (Sweden)

    Falk Eigemann

    Full Text Available Interspecific differences in the response of microalgae to stress have numerous ecological implications. However, little is known of intraspecific sensitivities and the potential role of local genetic adaptation of populations. We compared the allelochemical sensitivity of 23 Pediastrum duplex Meyen strains, a common component of the freshwater phytoplankton. In order to test for local genetic adaptation, strains were isolated from water bodies with and without the allelopathically-active submerged macrophyte Myriophyllum. Strains were assigned to P. duplex on the basis of cell shape and colony morphology and only P. duplex strains that belonged to the same lineage in an ITS rDNA phylogeny were used. Inhibition of strain growth rates and maximum quantum yields of photosystem II were measured after exposure to tannic acid (TA and co-culture with Myriophyllum spicatum. Growth rate inhibition varied over one order of magnitude between the P. duplex strains. There was no correlation between the presence of Myriophyllum in the source location and the sensitivity of the strains to TA or the presence of Myriophyllum, suggesting that at least strong unidirectional local adaptation to Myriophyllum had not taken place in the studied water bodies. The maximum quantum yield of photosystem II of TA exposed algae decreased, whereas the yield of algae exposed to M. spicatum was slightly higher than that of the controls. The ranking of P. duplex strain sensitivities differed between the types of exposure (single additions of TA versus co-existence with M. spicatum and the parameter measured (growth rate versus maximum quantum yield, emphasizing the importance of measuring multiple traits when analysing strain-specific sensitivities towards allelochemicals. The observation that sensitivities to allelochemicals vary widely among strains of a single freshwater algal species should be taken into account if evaluating ecological consequences of allelopathic

  9. Effect of DNA binding protein Ssh12 from hyperthermophilic archaeon Sulfolobus shibatae on DNA supercoiling

    Institute of Scientific and Technical Information of China (English)

    楼慧强; 黄力; VietQ.Mai

    1999-01-01

    An 11.5-ku DNA binding protein, designated as Sshl2, was purified from the hyperthermophilic archaeon Sulfolobus shibatae by column chromatography in SP Sepharose, DNA cellulose and phosphocellulose. Sshl2 accounts for about 4 % of the total cellular protein. The protein is capable of binding to both negatively supercoiled and relaxed DNAs. Nick closure analysis revealed that Sshl2 constrains negative supercoils upon binding to DNA. While the ability of the protein to constrain supercoils is weak at 22℃ , it is enhanced substantially at temperatures higher than 37℃ . Both the cellular content and supercoil-constraining ability of Sshl2 suggest that the protein may play an important role in the organization and stabilization of the chromosome of S. shibatae.

  10. Effects of DNA damage on oocyte meiotic maturation and early embryonic development

    Directory of Open Access Journals (Sweden)

    Shen YIN,Junyu MA,Wei SHEN

    2014-09-01

    Full Text Available DNA damage is one of the most common threats to meiotic cells. It has the potential to induce infertility and genetic abnormalities that may be passed to the embryo. Here, we reviewed exogenous factors which could induce DNA damage. Specially, we addressed the different effects of DNA damage on mouse oocytes and embryonic development. Complex DNA damage, double-strand breaks, represents a more difficult repair process and involves various repair pathways. Understanding the mechanisms involved in DNA damage responses may improve therapeutic strategies for ovarian cancer and fertility preservation.

  11. Different effects of ppGpp on Escherichia coli DNA replication in vivo and in vitro.

    Science.gov (United States)

    Maciąg-Dorszyńska, Monika; Szalewska-Pałasz, Agnieszka; Węgrzyn, Grzegorz

    2013-01-01

    Inhibition of Escherichia coli DNA replication by guanosine tetraphosphate (ppGpp) is demonstrated in vitro. This finding is compatible with impairment of the DnaG primase activity by this nucleotide. However, in agreement to previous reports, we were not able to detect a rapid inhibition of DNA synthesis in E. coli cells under the stringent control conditions, when intracellular ppGpp levels increase dramatically. We suggest that the process of ppGpp-mediated inhibition of DnaG activity may be masked in E. coli cells, which could provide a rationale for explanation of differences between ppGpp effects on DNA replication in E. coli and Bacillus subtilis.

  12. Hardware Impairments Aware Transceiver for Full-Duplex Massive MIMO Relaying

    Science.gov (United States)

    Xia, Xiaochen; Zhang, Dongmei; Xu, Kui; Ma, Wenfeng; Xu, Youyun

    2015-12-01

    This paper studies the massive MIMO full-duplex relaying (MM-FDR), where multiple source-destination pairs communicate simultaneously with the help of a common full-duplex relay equipped with very large antenna arrays. Different from the traditional MM-FDR protocol, a general model where sources/destinations are allowed to equip with multiple antennas is considered. In contrast to the conventional MIMO system, massive MIMO must be built with low-cost components which are prone to hardware impairments. In this paper, the effect of hardware impairments is taken into consideration, and is modeled using transmit/receive distortion noises. We propose a low complexity hardware impairments aware transceiver scheme (named as HIA scheme) to mitigate the distortion noises by exploiting the statistical knowledge of channels and antenna arrays at sources and destinations. A joint degree of freedom and power optimization algorithm is presented to further optimize the spectral efficiency of HIA based MM-FDR. The results show that the HIA scheme can mitigate the "ceiling effect" appears in traditional MM-FDR protocol, if the numbers of antennas at sources and destinations can scale with that at the relay.

  13. A novel duplex WDM-PON with DPSK modulated downstream and re-modulation of the downlink signal for OOKupstream

    Institute of Scientific and Technical Information of China (English)

    Aftab Hussain; Idress Afridi; YU Chong-xit; XIN Xiang-jun; YUAN Quan-xin; LIU Bo; Ashiq Hussain,; Abdul Latif; Abid Munir; Yousaf Khan

    2012-01-01

    We experimentally demonstrate and analyze a 10 Gbit/s full duplex wavelength division multiplexing passive optical network (WDM-PON) system.A non-return-to-zero differential phase shift keying (NRZ-DPSK) modulation technique is first utilized for downlink direction,and then the downlink signal is re-modulated for the uplink direction using intensity modulation technique of on-offkeying (OOK) with a data rate of 10 Gbit/s per channel.An effective colorless WDM-PON full duplex transmission system is achieved for the data rate of 10 Gbit/s per channel with a channel spacing of 60 GHz over the distance of 25 km with low power penalty.

  14. A comparison of colour duplex ultrasonography after transurethral alprostadil and intracavernous alprostadil in the assessment of erectile dysfunction.

    Science.gov (United States)

    Ahn, H S; Lee, S W; Yoon, S J; Hann, H J; Hong, J M

    2004-01-01

    This study aimed to investigate whether transurethral alprostadil could be used for the diagnosis of erectile dysfunction using colour duplex ultrasound. The ultrasonography results were compared after transurethral and intracavernous alprostadil administration in 20 patients with erectile dysfunction. There were no significant differences in the mean peak systolic velocities (PSVs) between the two routes of administration, but the mean end diastolic velocities (EDVs) showed significant differences, with patients treated with transurethral alprostadil having higher EDVs. Linear regression analysis of the PSVs reached following the two routes of administration showed a moderate relationship, but linear regression analysis of the EDVs showed no relationship. We concluded that transurethral alprostadil was an inappropriate vasoactive drug to use with colour duplex ultrasonography for the evaluation of patients with erectile dysfunction because it required a longer scan time and it was less effective and less reliable than intracavernous alprostadil at stimulating complete corporeal smooth muscle relaxation.

  15. Sole and stable RNA duplexes of G-rich sequences located in the 5'-untranslated region of protooncogenes.

    Science.gov (United States)

    Saxena, Sarika; Miyoshi, Daisuke; Sugimoto, Naoki

    2010-08-24

    Guanine- (G-) rich nucleic acid sequences can form four-stranded structures called G-quadruplexes. It is widely held that the formation of a G-quadruplex in RNA is more feasible than in DNA because of the lack of a complementary strand in mRNA. Here, we analyzed sequences of 5'-untranslated regions of protooncogenes and surprisingly found that these regions showed an enrichment of not only guanine (G) but also cytosine (C) nucleotides. Since neighboring cytosine- (C-) rich regions can affect the formation and stability of a G-quadruplex structure, we further investigated the properties of DNA and RNA structures of G-rich and GC-rich regions. We selected typical GC-rich RNA sequences from protooncogenes and corresponding DNA sequences and investigated their structures. It was found that the GC-rich RNA sequences formed stable A-form duplexes as their major structure independent of the surrounding conditions, including the presence of different cations (Na(+), K(+), or Li(+)) or molecular crowding with 40 wt % poly(ethylene glycol) with an average molecular mass of 200 Da although there are a few exceptions in which only a combination of K(+) and molecular crowding induced a G-quadruplex structure of an extremely G-rich RNA sequence. In contrast, structural polymorphisms involving duplexes, G-quadruplexes, and i-motifs were observed for GC-rich DNA sequences depending on the surrounding factors. These results demonstrate the considerable structural and functional differences in GC-rich sequences of the genome (DNA) and transcriptosome (mRNA) with respect to the nucleic acid backbone. Moreover, it was suggested that structural study for a G-rich RNA sequence should be carried out under cell-mimicking condition where K(+) and crowding cosolutes exist.

  16. Sigma-phase formation in weldments of cast super duplex stainless steel; Formacion de fase sigma en uniones soldadas de acero inoxidable super duplex fundido

    Energy Technology Data Exchange (ETDEWEB)

    Garin, J. L.; Mannheim, R. L.; Camus, M. A.

    2011-07-01

    This paper describes the microstructural characteristics of weldments of cast super duplex stainless steel (J93404), being subjected to annealing processes to induce formation of sigma-phase at high temperatures. The influence of heating time at 1073 K, 1123 K and 1173 K upon precipitation of sigma in the heat affected zone, base metal and fusion zone of the weldments was analyzed. The experimental results revealed the formation of this intermetallic compound throughout decomposition of the ferritic phase into austenite and sigma. At earlier stages of the transformation the phase rapidly nucleates and growth along the ferrite-austenite grain boundaries, and then massively advances towards the bulk of the ferritic zone with greater effectiveness as temperature increases. The formation of sigma-phase in all weldments resembles the Johnson-Mehl-Avramis mechanism stated for nucleation and growth. (Author) 27 refs.

  17. Characterization of microstructure and texture across dissimilar super duplex/austenitic stainless steel weldment joint by super duplex filler metal

    Energy Technology Data Exchange (ETDEWEB)

    Eghlimi, Abbas, E-mail: a.eghlimi@ma.iut.ac.ir [Department of Materials Engineering, Isfahan University of Technology, Isfahan 84156-83111 (Iran, Islamic Republic of); Shamanian, Morteza [Department of Materials Engineering, Isfahan University of Technology, Isfahan 84156-83111 (Iran, Islamic Republic of); Eskandarian, Masoomeh [Department of Materials Engineering, Shiraz University, Shiraz 71348-51154 (Iran, Islamic Republic of); Zabolian, Azam [Department of Natural Resources, Isfahan University of Technology, Isfahan 84156-83111 (Iran, Islamic Republic of); Szpunar, Jerzy A. [Department of Mechanical Engineering, University of Saskatchewan, Saskatoon SK S7N 5A9 (Canada)

    2015-08-15

    In the present paper, microstructural changes across an as-welded dissimilar austenitic/duplex stainless steel couple welded by a super duplex stainless steel filler metal using gas tungsten arc welding process is characterized with optical microscopy and electron back-scattered diffraction techniques. Accordingly, variations of microstructure, texture, and grain boundary character distribution of base metals, heat affected zones, and weld metal were investigated. The results showed that the weld metal, which was composed of Widmanstätten austenite side-plates and allotriomorphic grain boundary austenite morphologies, had the weakest texture and was dominated by low angle boundaries. The welding process increased the ferrite content but decreased the texture intensity at the heat affected zone of the super duplex stainless steel base metal. In addition, through partial ferritization, it changed the morphology of elongated grains of the rolled microstructure to twinned partially transformed austenite plateaus scattered between ferrite textured colonies. However, the texture of the austenitic stainless steel heat affected zone was strengthened via encouraging recrystallization and formation of annealing twins. At both interfaces, an increase in the special character coincident site lattice boundaries of the primary phase as well as a strong texture with <100> orientation, mainly of Goss component, was observed. - Graphical abstract: Display Omitted - Highlights: • Weld metal showed local orientation at microscale but random texture at macroscale. • Intensification of <100> orientated grains was observed adjacent to the fusion lines. • The austenite texture was weaker than that of the ferrite in all duplex regions. • Welding caused twinned partially transformed austenites to form at SDSS HAZ. • At both interfaces, the ratio of special CSL boundaries of the primary phase increased.

  18. Development of a One-Step Duplex RT-PCR Method for the Simultaneous Detection of VP3/VP1 and VP1/P2B Regions of the Hepatitis A Virus.

    Science.gov (United States)

    Kim, Mi-Ju; Lee, Shin-Young; Kim, Hyun-Joong; Lee, Jeong Su; Joo, In Sun; Kwak, Hyo Sun; Kim, Hae-Yeong

    2016-08-28

    The simultaneous detection and accurate identification of hepatitis A virus (HAV) is critical in food safety and epidemiological studies to prevent the spread of HAV outbreaks. Towards this goal, a one-step duplex reverse-transcription (RT)-PCR method was developed targeting the VP1/P2B and VP3/VP1 regions of the HAV genome for the qualitative detection of HAV. An HAV RT-qPCR standard curve was produced for the quantification of HAV RNA. The detection limit of the duplex RT-PCR method was 2.8 × 10(1) copies of HAV. The PCR products enabled HAV genotyping analysis through DNA sequencing, which can be applied for epidemiological investigations. The ability of this duplex RT-PCR method to detect HAV was evaluated with HAV-spiked samples of fresh lettuce, frozen strawberries, and oysters. The limit of detection of the one-step duplex RT-PCR for each food model was 9.4 × 10(2) copies/20 g fresh lettuce, 9.7 × 10(3) copies/20 g frozen strawberries, and 4.1 × 10(3) copies/1.5 g oysters. Use of a one-step duplex RT-PCR method has advantages such as shorter time, decreased cost, and decreased labor owing to the single amplification reaction instead of four amplifications necessary for nested RT-PCR.

  19. Residual stresses and fatigue in a duplex stainless steel

    Energy Technology Data Exchange (ETDEWEB)

    Johansson, Johan

    1999-05-01

    Duplex stainless steels, consisting of approximately equal amounts of austenite and ferrite, often combine the best features of austenitic and ferritic stainless steels. They generally have good mechanical properties, including high strength and ductility, and the corrosion resistance is often better than conventional austenitic grades. This has lead to a growing use of duplex stainless steels as a material in mechanically loaded constructions. However, detailed knowledge regarding its mechanical properties and deformation mechanisms are still lacking. In this thesis special emphasis has been placed on the residual stresses and their influence on mechanical behaviour of duplex stainless steels. Due to the difference in coefficient of thermal expansion between the two phases, tensile microstresses are found in the austenitic phase and balancing compressive microstresses in the ferritic phase. The first part of this thesis is a literature survey, which will give an introduction to duplex stainless steels and review the fatigue properties of duplex stainless steels and the influence of residual stresses in two-phase material. The second part concerns the evolution of the residual stress state during uniaxial loading. Initial residual stresses were found to be almost two times higher in the transverse direction compared to the rolling direction. During loading the absolute value of the microstresses increased in the macroscopic elastic regime but started to decrease with increasing load in the macroscopic plastic regime. A significant increase of the microstresses was also found to occur during unloading. Finite element simulations also show stress variation within one phase and a strong influence of both the elastic and plastic anisotropy of the individual phases on the simulated stress state. In the third part, the load sharing between the phases during cyclic loading is studied. X-ray diffraction stress analysis and transmission electron microscopy show that even if

  20. Interaction of Proliferating Cell Nuclear Antigen With DNA at the Single Molecule Level

    KAUST Repository

    Raducanu, Vlad-Stefan

    2016-05-01

    Proliferating cell nuclear antigen (PCNA) is a key factor involved in Eukaryotic DNA replication and repair, as well as other cellular pathways. Its importance comes mainly from two aspects: the large numbers of interacting partners and the mechanism of facilitated diffusion along the DNA. The large numbers of interacting partners makes PCNA a necessary factor to consider when studying DNA replication, either in vitro or in vivo. The mechanism of facilitated diffusion along the DNA, i.e. sliding along the duplex, reduces the six degrees of freedom of the molecule, three degrees of freedom of translation and three degrees of freedom of rotation, to only two, translation along the duplex and rotational tracking of the helix. Through this mechanism PCNA can recruit its partner proteins and localize them to the right spot on the DNA, maybe in the right spatial orientation, more effectively and in coordination with other proteins. Passive loading of the closed PCNA ring on the DNA without free ends is a topologically forbidden process. Replication factor C (RFC) uses energy of ATP hydrolysis to mechanically open the PCNA ring and load it on the dsDNA. The first half of the introduction gives overview of PCNA and RFC and the loading mechanism of PCNA on dsDNA. The second half is dedicated to a diffusion model and to an algorithm for analyzing PCNA sliding. PCNA and RFC were successfully purified, simulations and a mean squared displacement analysis algorithm were run and showed good stability and experimental PCNA sliding data was analyzed and led to parameters similar to the ones in literature.

  1. The effect of polyamines on the binding of anti-DNA antibodies from patients with SLE and normal human subjects.

    Science.gov (United States)

    Wang, Xiao; Stearns, Nancy A; Li, Xingfu; Pisetsky, David S

    2014-07-01

    Antibodies to DNA (anti-DNA) are the serological hallmark of systemic lupus erythematosus (SLE). To elucidate specificity further, the effect of polyamines on the binding of anti-DNA antibodies from patients with lupus was tested by ELISA to calf thymus (CT) DNA; we also assessed the binding of plasmas of patients and normal human subjects (NHS) to Micrococcus luteus (MC) DNA. As these studies showed, spermine can dose-dependently inhibit SLE anti-DNA binding to CT DNA and can promote dissociation of preformed immune complexes. With MC DNA as antigen, spermine failed to inhibit the NHS anti-DNA binding. Studies using plasmas adsorbed to a CT DNA cellulose affinity indicated that SLE plasmas are mixtures of anti-DNA that differ in inhibition by spermine and binding to conserved and non-conserved determinants. Together, these studies demonstrate that spermine can influence the binding of anti-DNA autoantibodies and may contribute to the antigenicity of DNA.

  2. Effect of the Spiroiminodihydantoin Lesion on Nucleosome Stability and Positioning.

    Science.gov (United States)

    Norabuena, Erika M; Barnes Williams, Sara; Klureza, Margaret A; Goehring, Liana J; Gruessner, Brian; Radhakrishnan, Mala L; Jamieson, Elizabeth R; Núñez, Megan E

    2016-04-26

    DNA is constantly under attack by oxidants, generating a variety of potentially mutagenic covalently modified species, including oxidized guanine base products. One such product is spiroiminodihydantoin (Sp), a chiral, propeller-shaped lesion that strongly destabilizes the DNA helix in its vicinity. Despite its unusual shape and thermodynamic effect on double-stranded DNA structure, DNA duplexes containing the Sp lesion form stable nucleosomes upon being incubated with histone octamers. Indeed, among six different combinations of lesion location and stereochemistry, only two duplexes display a diminished ability to form nucleosomes, and these only by ∼25%; the other four are statistically indistinguishable from the control. Nonetheless, kinetic factors also play a role: when the histone proteins have less time during assembly of the core particle to sample both lesion-containing and normal DNA strands, they are more likely to bind the Sp lesion DNA than during slower assembly processes that better approximate thermodynamic equilibrium. Using DNase I footprinting and molecular modeling, we discovered that the Sp lesion causes only a small perturbation (±1-2 bp) on the translational position of the DNA within the nucleosome. Each diastereomeric pair of lesions has the same effect on nucleosome positioning, but lesions placed at different locations behave differently, illustrating that the location of the lesion and not its shape serves as the primary determinant of the most stable DNA orientation.

  3. Tooth surface geometry optimization of spiral bevel and hypoid gears generated by duplex helical method with circular profile blade

    Institute of Scientific and Technical Information of China (English)

    ZHANG Yu; YAN Hong-zhi; ZENG Tao; ZENG Yi-yu

    2016-01-01

    In order to effectively improve meshing performance of spiral bevel and hypoid gears generated by the duplex helical method, the effects of straight lined and circular cutting edges profile on meshing and contact of spiral bevel and hypoid gears were investigated analytically. Firstly, a mathematical model of spiral bevel and hypoid gears with circular blade profile was established according to the cutting characteristics of the duplex helical method. Based on a hypoid gear drive, the tooth bearings and the functions of transmission errors of four design cases were analyzed respectively by the use of the tooth contact analysis (TCA), and the contact stresses of the four design cases were analyzed and compared using simulation software. Finally, the curvature radius of the circular profile blade was optimized. The results show that the contact stresses are availably reduced, and the areas of edge contact and severe contact stresses can be avoided by selecting appropriate circular blade profile. In addition, the convex and concave sides are separately modified by the use of different curvature radii of inside and outside blades, which can increase the flexibility of the duplex helical method.

  4. The Effect of Geographical Scale of Sampling on DNA Barcoding

    Science.gov (United States)

    Bergsten, Johannes; Bilton, David T.; Fujisawa, Tomochika; Elliott, Miranda; Monaghan, Michael T.; Balke, Michael; Hendrich, Lars; Geijer, Joja; Herrmann, Jan; Foster, Garth N.; Ribera, Ignacio; Nilsson, Anders N.; Barraclough, Timothy G.; Vogler, Alfried P.

    2012-01-01

    Eight years after DNA barcoding was formally proposed on a large scale, CO1 sequences are rapidly accumulating from around the world. While studies to date have mostly targeted local or regional species assemblages, the recent launch of the global iBOL project (International Barcode of Life), highlights the need to understand the effects of geographical scale on Barcoding's goals. Sampling has been central in the debate on DNA Barcoding, but the effect of the geographical scale of sampling has not yet been thoroughly and explicitly tested with empirical data. Here, we present a CO1 data set of aquatic predaceous diving beetles of the tribe Agabini, sampled throughout Europe, and use it to investigate how the geographic scale of sampling affects 1) the estimated intraspecific variation of species, 2) the genetic distance to the most closely related heterospecific, 3) the ratio of intraspecific and interspecific variation, 4) the frequency of taxonomically recognized species found to be monophyletic, and 5) query identification performance based on 6 different species assignment methods. Intraspecific variation was significantly correlated with the geographical scale of sampling (R-square = 0.7), and more than half of the species with 10 or more sampled individuals (N = 29) showed higher intraspecific variation than 1% sequence divergence. In contrast, the distance to the closest heterospecific showed a significant decrease with increasing geographical scale of sampling. The average genetic distance dropped from > 7% for samples within 1 km, to 6000 km apart. Over a third of the species were not monophyletic, and the proportion increased through locally, nationally, regionally, and continentally restricted subsets of the data. The success of identifying queries decreased with increasing spatial scale of sampling; liberal methods declined from 100% to around 90%, whereas strict methods dropped to below 50% at continental scales. The proportion of query identifications

  5. Targeting Ongoing DNA Damage in Multiple Myeloma: Effects of DNA Damage Response Inhibitors on Plasma Cell Survival

    Directory of Open Access Journals (Sweden)

    Ana Belén Herrero

    2017-05-01

    Full Text Available Human myeloma cell lines (HMCLs and a subset of myeloma patients with poor prognosis exhibit high levels of replication stress (RS, leading to DNA damage. In this study, we confirmed the presence of DNA double-strand breaks (DSBs in several HMCLs by measuring γH2AX and RAD51 foci and analyzed the effect of various inhibitors of the DNA damage response on MM cell survival. Inhibition of ataxia telangiectasia and Rad3-related protein (ATR, the main kinase mediating the response to RS, using the specific inhibitor VE-821 induced more cell death in HMCLs than in control lymphoblastoid cells and U266, an HMCL with a low level of DNA damage. The absence of ATR was partially compensated by ataxia telangiectasia-mutated protein (ATM, since chemical inhibition of both kinases using VE-821 and KU-55933 significantly increased the death of MM cells with DNA damage. We found that ATM and ATR are involved in DSB repair by homologous recombination (HR in MM. Inhibition of both kinases resulted in a stronger inhibition that may underlie cell death induction, since abolition of HR using two different inhibitors severely reduced survival of HMCLs that exhibit DNA damage. On the other hand, inhibition of the other route involved in DSB repair, non-homologous end joining (NHEJ, using the DNA-PK inhibitor NU7441, did not affect MM cell viability. Interestingly, we found that NHEJ inhibition did not increase cell death when HR was simultaneously inhibited with the RAD51 inhibitor B02, but it clearly increased the level of cell death when HR was inhibited with the MRE11 inhibitor mirin, which interferes with recombination before DNA resection takes place. Taken together, our results demonstrate for the first time that MM cells with ongoing DNA damage rely on an intact HR pathway, which thereby suggests therapeutic opportunities. We also show that inhibition of HR after the initial step of end resection might be more appropriate for inducing MM cell death, since it

  6. Characterization of a novel DNA glycosylase from S. sahachiroi involved in the reduction and repair of azinomycin B induced DNA damage.

    Science.gov (United States)

    Wang, Shan; Liu, Kai; Xiao, Le; Yang, LiYuan; Li, Hong; Zhang, FeiXue; Lei, Lei; Li, ShengQing; Feng, Xu; Li, AiYing; He, Jing

    2016-01-01

    Azinomycin B is a hybrid polyketide/nonribosomal peptide natural product and possesses antitumor activity by interacting covalently with duplex DNA and inducing interstrand crosslinks. In the biosynthetic study of azinomycin B, a gene (orf1) adjacent to the azinomycin B gene cluster was found to be essential for the survival of the producer, Streptomyces sahachiroi ATCC33158. Sequence analyses revealed that Orf1 belongs to the HTH_42 superfamily of conserved bacterial proteins which are widely distributed in pathogenic and antibiotic-producing bacteria with unknown functions. The protein exhibits a protective effect against azinomycin B when heterologously expressed in azinomycin-sensitive strains. EMSA assays showed its sequence nonspecific binding to DNA and structure-specific binding to azinomycin B-adducted sites, and ChIP assays revealed extensive association of Orf1 with chromatin in vivo. Interestingly, Orf1 not only protects target sites by protein-DNA interaction but is also capable of repairing azinomycin B-mediated DNA cross-linking. It possesses the DNA glycosylase-like activity and specifically repairs DNA damage induced by azinomycin B through removal of both adducted nitrogenous bases in the cross-link. This bifunctional protein massively binds to genomic DNA to reduce drug attack risk as a novel DNA binding protein and triggers the base excision repair system as a novel DNA glycosylase.

  7. Microtwin formation in the {alpha} phase of duplex titanium alloys affected by strain rate

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Yi-Hsiang; Wu, Shu-Ming [Department of Mechanical and Mechatronic Engineering, National Taiwan Ocean University, No. 2 Pei Ning Road, Keelung 20224, Taiwan (China); Kao, Fang-Hsin [Department of Materials Science and Engineering, National Taiwan University, Taipei 10617, Taiwan (China); Wang, Shing-Hoa, E-mail: shwang@ntou.edu.tw [Department of Mechanical and Mechatronic Engineering, National Taiwan Ocean University, No. 2 Pei Ning Road, Keelung 20224, Taiwan (China); Yang, Jer-Ren [Department of Materials Science and Engineering, National Taiwan University, Taipei 10617, Taiwan (China); Yang, Chia-Chih [Department of Mechanical and Mechatronic Engineering, National Taiwan Ocean University, No. 2 Pei Ning Road, Keelung 20224, Taiwan (China); Chiou, Chuan-Sheng [Department of Mechanical Engineering, Yuan Ze University, Taoyuan 32003, Taiwan (China)

    2011-03-15

    Research highlights: {yields} The long and dense twins in {alpha} phase of SP700 alloy occurring at lower strain rates promote a good ductility. {yields} The deformation in SP700 alloy changed to micro twins-controlled mechanism in {alpha} as the strain rate decreases. {yields} The material has time to redistribute the deformed strain between {alpha} and {beta} as the strain rate decreases. - Abstract: The effect of tensile strain rate on deformation microstructure was investigated in Ti-6-4 (Ti-6Al-4V) and SP700 (Ti-4.5Al-3V-2Mo-2Fe) of the duplex titanium alloys. Below a strain rate of 10{sup -2} s{sup -1}, Ti-6-4 alloy had a higher ultimate tensile strength than SP700 alloy. However, the yield strength of SP700 was consistently greater than Ti-6-4 at different strain rates. The ductility of SP700 alloy associated with twin formation (especially at the slow strain rate of 10{sup -4} s{sup -1}), always exceeded that of Ti-6-4 alloy at different strain rates. It is caused by a large quantity of deformation twins took place in the {alpha} phase of SP700 due to the lower stacking fault energy by the {beta} stabilizer of molybdenum alloying. In addition, the local deformation more was imposed on the {alpha} grains from the surrounding {beta}-rich grains by redistributing strain as the strain rate decreased in SP700 duplex alloy.

  8. Improper Gaussian signaling in full-duplex relay channels with residual self-interference