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Sample records for high dna melting

  1. Force induced DNA melting

    International Nuclear Information System (INIS)

    Santosh, Mogurampelly; Maiti, Prabal K

    2009-01-01

    When pulled along the axis, double-strand DNA undergoes a large conformational change and elongates by roughly twice its initial contour length at a pulling force of about 70 pN. The transition to this highly overstretched form of DNA is very cooperative. Applying a force perpendicular to the DNA axis (unzipping), double-strand DNA can also be separated into two single-stranded DNA, this being a fundamental process in DNA replication. We study the DNA overstretching and unzipping transition using fully atomistic molecular dynamics (MD) simulations and argue that the conformational changes of double-strand DNA associated with either of the above mentioned processes can be viewed as force induced DNA melting. As the force at one end of the DNA is increased the DNA starts melting abruptly/smoothly above a critical force depending on the pulling direction. The critical force f m , at which DNA melts completely decreases as the temperature of the system is increased. The melting force in the case of unzipping is smaller compared to the melting force when the DNA is pulled along the helical axis. In the case of melting through unzipping, the double-strand separation has jumps which correspond to the different energy minima arising due to sequence of different base pairs. The fraction of Watson-Crick base pair hydrogen bond breaking as a function of force does not show smooth and continuous behavior and consists of plateaus followed by sharp jumps.

  2. High-Resolution Melting (HRM) of Hypervariable Mitochondrial DNA Regions for Forensic Science.

    Science.gov (United States)

    Dos Santos Rocha, Alípio; de Amorim, Isis Salviano Soares; Simão, Tatiana de Almeida; da Fonseca, Adenilson de Souza; Garrido, Rodrigo Grazinoli; Mencalha, Andre Luiz

    2018-03-01

    Forensic strategies commonly are proceeding by analysis of short tandem repeats (STRs); however, new additional strategies have been proposed for forensic science. Thus, this article standardized the high-resolution melting (HRM) of DNA for forensic analyzes. For HRM, mitochondrial DNA (mtDNA) from eight individuals were extracted from mucosa swabs by DNAzol reagent, samples were amplified by PCR and submitted to HRM analysis to identify differences in hypervariable (HV) regions I and II. To confirm HRM, all PCR products were DNA sequencing. The data suggest that is possible discriminate DNA from different samples by HRM curves. Also, uncommon dual-dissociation was identified in a single PCR product, increasing HRM analyzes by evaluation of melting peaks. Thus, HRM is accurate and useful to screening small differences in HVI and HVII regions from mtDNA and increase the efficiency of laboratory routines based on forensic genetics. © 2017 American Academy of Forensic Sciences.

  3. High resolution melting (HRM) analysis of DNA--its role and potential in food analysis.

    Science.gov (United States)

    Druml, Barbara; Cichna-Markl, Margit

    2014-09-01

    DNA based methods play an increasing role in food safety control and food adulteration detection. Recent papers show that high resolution melting (HRM) analysis is an interesting approach. It involves amplification of the target of interest in the presence of a saturation dye by the polymerase chain reaction (PCR) and subsequent melting of the amplicons by gradually increasing the temperature. Since the melting profile depends on the GC content, length, sequence and strand complementarity of the product, HRM analysis is highly suitable for the detection of single-base variants and small insertions or deletions. The review gives an introduction into HRM analysis, covers important aspects in the development of an HRM analysis method and describes how HRM data are analysed and interpreted. Then we discuss the potential of HRM analysis based methods in food analysis, i.e. for the identification of closely related species and cultivars and the identification of pathogenic microorganisms. Copyright © 2014 Elsevier Ltd. All rights reserved.

  4. ‘‘Blind'' mapping of genic DNA sequence polymorphisms in Lolium perenne L. by high resolution melting curve analysis

    DEFF Research Database (Denmark)

    Studer, Bruno; Jensen, Louise Bach; Fiil, Alice

    2009-01-01

    High resolution melting curve analysis (HRM) measures dissociation of double stranded DNA of a PCR product amplified in the presence of a saturating fluorescence dye. Recently, HRM proved successful to genotype DNA sequence polymorphisms such as SSRs and SNPs based on the shape of the melting...... curves. In this study, HRM was used for simultaneous screening and genotyping of genic DNA sequence polymorphisms identified in the Lolium perenne F2 mapping population VrnA. Melting profiles of PCR products amplified from previously published gene loci and from a novel gene putatively involved...

  5. Reliable discrimination of 10 ungulate species using high resolution melting analysis of faecal DNA.

    Directory of Open Access Journals (Sweden)

    Ana Ramón-Laca

    Full Text Available Identifying species occupying an area is essential for many ecological and conservation studies. Faecal DNA is a potentially powerful method for identifying cryptic mammalian species. In New Zealand, 10 species of ungulate (Order: Artiodactyla have established wild populations and are managed as pests because of their impacts on native ecosystems. However, identifying the ungulate species present within a management area based on pellet morphology is unreliable. We present a method that enables reliable identification of 10 ungulate species (red deer, sika deer, rusa deer, fallow deer, sambar deer, white-tailed deer, Himalayan tahr, Alpine chamois, feral sheep, and feral goat from swabs of faecal pellets. A high resolution melting (HRM assay, targeting a fragment of the 12S rRNA gene, was developed. Species-specific primers were designed and combined in a multiplex PCR resulting in fragments of different length and therefore different melting behaviour for each species. The method was developed using tissue from each of the 10 species, and was validated in blind trials. Our protocol enabled species to be determined for 94% of faecal pellet swabs collected during routine monitoring by the New Zealand Department of Conservation. Our HRM method enables high-throughput and cost-effective species identification from low DNA template samples, and could readily be adapted to discriminate other mammalian species from faecal DNA.

  6. Melting curves of gammairradiated DNA

    International Nuclear Information System (INIS)

    Hofer, H.; Altmann, H.; Kehrer, M.

    1978-08-01

    Melting curves of gammairradiated DNA and data derived of them, are reported. The diminished stability is explained by basedestruction. DNA denatures completely at room temperature, if at least every fifth basepair is broken or weakened by irradiation. (author)

  7. Refining DNA Barcoding Coupled High Resolution Melting for Discrimination of 12 Closely Related Croton Species.

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    Maslin Osathanunkul

    Full Text Available DNA barcoding coupled high resolution melting (Bar-HRM is an emerging method for species discrimination based on DNA dissociation kinetics. The aim of this work was to evaluate the suitability of different primer sets, derived from selected DNA regions, for Bar-HRM analysis of species in Croton (Euphorbiaceae, one of the largest genera of plants with over 1,200 species. Seven primer pairs were evaluated (matK, rbcL1, rbcL2, rbcL3, rpoC, trnL and ITS1 from four plastid regions, matK, rbcL, rpoC, and trnL, and the nuclear ribosomal marker ITS1. The primer pair derived from the ITS1 region was the single most effective region for the identification of the tested species, whereas the rbcL1 primer pair gave the lowest resolution. It was observed that the ITS1 barcode was the most useful DNA barcoding region overall for species discrimination out of all of the regions and primers assessed. Our Bar-HRM results here also provide further support for the hypothesis that both sequence and base composition affect DNA duplex stability.

  8. Detection of somatic mutations by high-resolution DNA melting (HRM) analysis in multiple cancers.

    Science.gov (United States)

    Gonzalez-Bosquet, Jesus; Calcei, Jacob; Wei, Jun S; Garcia-Closas, Montserrat; Sherman, Mark E; Hewitt, Stephen; Vockley, Joseph; Lissowska, Jolanta; Yang, Hannah P; Khan, Javed; Chanock, Stephen

    2011-01-17

    Identification of somatic mutations in cancer is a major goal for understanding and monitoring the events related to cancer initiation and progression. High resolution melting (HRM) curve analysis represents a fast, post-PCR high-throughput method for scanning somatic sequence alterations in target genes. The aim of this study was to assess the sensitivity and specificity of HRM analysis for tumor mutation screening in a range of tumor samples, which included 216 frozen pediatric small rounded blue-cell tumors as well as 180 paraffin-embedded tumors from breast, endometrial and ovarian cancers (60 of each). HRM analysis was performed in exons of the following candidate genes known to harbor established commonly observed mutations: PIK3CA, ERBB2, KRAS, TP53, EGFR, BRAF, GATA3, and FGFR3. Bi-directional sequencing analysis was used to determine the accuracy of the HRM analysis. For the 39 mutations observed in frozen samples, the sensitivity and specificity of HRM analysis were 97% and 87%, respectively. There were 67 mutation/variants in the paraffin-embedded samples, and the sensitivity and specificity for the HRM analysis were 88% and 80%, respectively. Paraffin-embedded samples require higher quantity of purified DNA for high performance. In summary, HRM analysis is a promising moderate-throughput screening test for mutations among known candidate genomic regions. Although the overall accuracy appears to be better in frozen specimens, somatic alterations were detected in DNA extracted from paraffin-embedded samples.

  9. Detection of somatic mutations by high-resolution DNA melting (HRM analysis in multiple cancers.

    Directory of Open Access Journals (Sweden)

    Jesus Gonzalez-Bosquet

    Full Text Available Identification of somatic mutations in cancer is a major goal for understanding and monitoring the events related to cancer initiation and progression. High resolution melting (HRM curve analysis represents a fast, post-PCR high-throughput method for scanning somatic sequence alterations in target genes. The aim of this study was to assess the sensitivity and specificity of HRM analysis for tumor mutation screening in a range of tumor samples, which included 216 frozen pediatric small rounded blue-cell tumors as well as 180 paraffin-embedded tumors from breast, endometrial and ovarian cancers (60 of each. HRM analysis was performed in exons of the following candidate genes known to harbor established commonly observed mutations: PIK3CA, ERBB2, KRAS, TP53, EGFR, BRAF, GATA3, and FGFR3. Bi-directional sequencing analysis was used to determine the accuracy of the HRM analysis. For the 39 mutations observed in frozen samples, the sensitivity and specificity of HRM analysis were 97% and 87%, respectively. There were 67 mutation/variants in the paraffin-embedded samples, and the sensitivity and specificity for the HRM analysis were 88% and 80%, respectively. Paraffin-embedded samples require higher quantity of purified DNA for high performance. In summary, HRM analysis is a promising moderate-throughput screening test for mutations among known candidate genomic regions. Although the overall accuracy appears to be better in frozen specimens, somatic alterations were detected in DNA extracted from paraffin-embedded samples.

  10. High DNA melting temperature predicts transcription start site location in human and mouse.

    LENUS (Irish Health Repository)

    Dineen, David G

    2009-12-01

    The accurate computational prediction of transcription start sites (TSS) in vertebrate genomes is a difficult problem. The physicochemical properties of DNA can be computed in various ways and a many combinations of DNA features have been tested in the past for use as predictors of transcription. We looked in detail at melting temperature, which measures the temperature, at which two strands of DNA separate, considering the cooperative nature of this process. We find that peaks in melting temperature correspond closely to experimentally determined transcription start sites in human and mouse chromosomes. Using melting temperature alone, and with simple thresholding, we can predict TSS with accuracy that is competitive with the most accurate state-of-the-art TSS prediction methods. Accuracy is measured using both experimentally and manually determined TSS. The method works especially well with CpG island containing promoters, but also works when CpG islands are absent. This result is clear evidence of the important role of the physical properties of DNA in the process of transcription. It also points to the importance for TSS prediction methods to include melting temperature as prior information.

  11. Rapid discrimination of Isaria javanica and Isaria poprawskii from Isaria spp. using high resolution DNA melting assays

    Science.gov (United States)

    The current study evaluates the potential of using high resolution DNA melting assays to discriminate species in the genus, Isaria. The study utilizes a previously identified 103 base pair PCR amplicon, which was reported to be selective for Isaria fumosorosea. Our study finds the amplicon selective...

  12. Supercoil Formation During DNA Melting

    Science.gov (United States)

    Sayar, Mehmet; Avsaroglu, Baris; Kabakcioglu, Alkan

    2009-03-01

    Supercoil formation plays a key role in determining the structure-function relationship in DNA. Biological and technological processes, such as protein synthesis, polymerase chain reaction, and microarrays relys on separation of the two strands in DNA, which is coupled to the unwinding of the supercoiled structure. This problem has been studied theoretically via Peyrard-Bishop and Poland-Scheraga type models, which include a simple representation of the DNA structural properties. In recent years, computational models, which provide a more realtistic representaion of DNA molecule, have been used to study the melting behavior of short DNA chains. Here, we will present a new coarse-grained model of DNA which is capable of simulating sufficiently long DNA chains for studying the supercoil formation during melting, without sacrificing the local structural properties. Our coarse-grained model successfully reproduces the local geometry of the DNA molecule, such as the 3'-5' directionality, major-minor groove structure, and the helical pitch. We will present our initial results on the dynamics of supercoiling during DNA melting.

  13. High-resolution melt-curve analysis of random-amplified-polymorphic-DNA markers, for the characterisation of pathogenic Leptospira

    DEFF Research Database (Denmark)

    Tulsiani, Suhella; Craig, S B; Graham, G C

    2010-01-01

    A new test for pathogenic Leptospira isolates, based on RAPD-PCR and high-resolution melt (HRM) analysis (which measures the melting temperature of amplicons in real time, using a fluorescent DNA-binding dye), has recently been developed. A characteristic profile of the amplicons can be used...... typed against 13 previously published RAPD primers, using a real-time cycler (the Corbett Life Science RotorGene 6000) and the optimised reagents from a commercial kit (Quantace SensiMix). RAPD-HRM at specific temperatures generated defining amplicon melt profiles for each of the tested serovars....... These profiles were evaluated as difference-curve graphs generated using the RotorGene software package, with a cut-off of at least 8 'U' (plus or minus). The results demonstrated that RAPD-HRM can be used to measure serovar diversity and establish identity, with a high degree of stability. The characterisation...

  14. Application of high-resolution DNA melting for genotyping in lepidopteran non-model species: Ostrinia furnacalis (Crambidae.

    Directory of Open Access Journals (Sweden)

    FengBo Li

    Full Text Available Development of an ideal marker system facilitates a better understanding of the genetic diversity in lepidopteran non-model organisms, which have abundant species, but relatively limited genomic resources. Single nucleotide polymorphisms (SNPs discovered within single-copy genes have proved to be desired markers, but SNP genotyping by current techniques remain laborious and expensive. High resolution melting (HRM curve analysis represents a simple, rapid and inexpensive genotyping method that is primarily confined to clinical and diagnostic studies. In this study, we evaluated the potential of HRM analysis for SNP genotyping in the lepidopteran non-model species Ostrinia furnacalis (Crambidae. Small amplicon and unlabeled probe assays were developed for the SNPs, which were identified in 30 females of O. furnacalis from 3 different populations by our direct sequencing. Both assays were then applied to genotype 90 unknown female DNA by prior mixing with known wild-type DNA. The genotyping results were compared with those that were obtained using bi-directional sequencing analysis. Our results demonstrated the efficiency and reliability of the HRM assays. HRM has the potential to provide simple, cost-effective genotyping assays and facilitates genotyping studies in any non-model lepidopteran species of interest.

  15. Validation of high-resolution DNA melting analysis for mutation scanning of the CDKL5 gene: identification of novel mutations.

    Science.gov (United States)

    Raymond, Laure; Diebold, Bertrand; Leroux, Céline; Maurey, Hélène; Drouin-Garraud, Valérie; Delahaye, Andre; Dulac, Olivier; Metreau, Julia; Melikishvili, Gia; Toutain, Annick; Rivier, François; Bahi-Buisson, Nadia; Bienvenu, Thierry

    2013-01-01

    Mutations in the cyclin-dependent kinase-like 5 gene (CDKL5) have been predominantly described in epileptic encephalopathies of female, including infantile spasms with Rett-like features. Up to now, detection of mutations in this gene was made by laborious, expensive and/or time consuming methods. Here, we decided to validate high-resolution melting analysis (HRMA) for mutation scanning of the CDKL5 gene. Firstly, using a large DNA bank consisting to 34 samples carrying different mutations and polymorphisms, we validated our analytical conditions to analyse the different exons and flanking intronic sequences of the CDKL5 gene by HRMA. Secondly, we screened CDKL5 by both HRMA and denaturing high performance liquid chromatography (dHPLC) in a cohort of 135 patients with early-onset seizures. Our results showed that point mutations and small insertions and deletions can be reliably detected by HRMA. Compared to dHPLC, HRMA profiles are more discriminated, thereby decreasing unnecessary sequencing. In this study, we identified eleven novel sequence variations including four pathogenic mutations (2.96% prevalence). HRMA appears cost-effective, easy to set up, highly sensitive, non-toxic and rapid for mutation screening, ideally suited for large genes with heterogeneous mutations located along the whole coding sequence, such as the CDKL5 gene. Copyright © 2012 Elsevier B.V. All rights reserved.

  16. Detection of plant oil DNA using high resolution melting (HRM) post PCR analysis: a tool for disclosure of olive oil adulteration.

    Science.gov (United States)

    Vietina, Michelangelo; Agrimonti, Caterina; Marmiroli, Nelson

    2013-12-15

    Extra virgin olive oil is frequently subjected to adulterations with addition of oils obtained from plants other than olive. DNA analysis is a fast and economic tool to identify plant components in oils. Extraction and amplification of DNA by PCR was tested in olives, in milled seeds and in oils, to investigate its use in olive oil traceability. DNA was extracted from different oils made of hazelnut, maize, sunflower, peanut, sesame, soybean, rice and pumpkin. Comparing the DNA melting profiles in reference plant materials and in the oils, it was possible to identify any plant components in oils and mixtures of oils. Real-Time PCR (RT-PCR) platform has been added of the new methodology of high resolution melting (HRM), both were used to analyse olive oils mixed with different percentage of other oils. Results showed HRM a cost effective method for efficient detection of adulterations in olive oils. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Rapid analysis of heterogeneously methylated DNA using digital methylation-sensitive high resolution melting: application to the CDKN2B (p15) gene

    DEFF Research Database (Denmark)

    Candiloro, Ida Lm; Mikeska, Thomas; Hokland, Peter

    2008-01-01

    ABSTRACT: BACKGROUND: Methylation-sensitive high resolution melting (MS-HRM) methodology is able to recognise heterogeneously methylated sequences by their characteristic melting profiles. To further analyse heterogeneously methylated sequences, we adopted a digital approach to MS-HRM (dMS-HRM) t......ABSTRACT: BACKGROUND: Methylation-sensitive high resolution melting (MS-HRM) methodology is able to recognise heterogeneously methylated sequences by their characteristic melting profiles. To further analyse heterogeneously methylated sequences, we adopted a digital approach to MS-HRM (d......MS-HRM) that involves the amplification of single templates after limiting dilution to quantify and to determine the degree of methylation. We used this approach to study methylation of the CDKN2B (p15) cell cycle progression inhibitor gene which is inactivated by DNA methylation in haematological malignancies...... the methylated alleles and assess the degree of methylation. Direct sequencing of selected dMS-HRM products was used to determine the exact DNA methylation pattern and confirmed the degree of methylation estimated by dMS-HRM. CONCLUSION: dMS-HRM is a powerful technique for the analysis of methylation in CDKN2B...

  18. High-resolution melting of 12S rRNA and cytochrome b DNA sequences for discrimination of species within distinct European animal families.

    Directory of Open Access Journals (Sweden)

    Jana Naue

    Full Text Available The cheap and easy identification of species is necessary within multiple fields of molecular biology. The use of high-resolution melting (HRM of DNA provides a fast closed-tube method for analysis of the sequence composition of the mitochondrial genes 12S rRNA and cytochrome b. We investigated the potential use of HRM for species identification within eleven different animal groups commonly found in Europe by animal-group-specific DNA amplification followed by DNA melting. Influence factors as DNA amount, additional single base alterations, and the existence of mixed samples were taken into consideration. Visual inspection combined with mathematical evaluation of the curve shapes did resolve nearly all species within an animal group. The assay can therefore not only be used for identification of animal groups and mixture analysis but also for species identification within the respective groups. The use of a universal 12S rRNA system additionally revealed a possible approach for species discrimination, mostly by exclusion. The use of the HRM assay showed to be a reliable, fast, and cheap method for species discrimination within a broad range of different animal species and can be used in a flexible "modular" manner depending on the question to be solved.

  19. Force-dependent melting of supercoiled DNA at thermophilic temperatures.

    Science.gov (United States)

    Galburt, E A; Tomko, E J; Stump, W T; Ruiz Manzano, A

    2014-01-01

    Local DNA opening plays an important role in DNA metabolism as the double-helix must be melted before the information contained within may be accessed. Cells finely tune the torsional state of their genomes to strike a balance between stability and accessibility. For example, while mesophilic life forms maintain negatively superhelical genomes, thermophilic life forms use unique mechanisms to maintain relaxed or even positively supercoiled genomes. Here, we use a single-molecule magnetic tweezers approach to quantify the force-dependent equilibrium between DNA melting and supercoiling at high temperatures populated by Thermophiles. We show that negatively supercoiled DNA denatures at 0.5 pN lower tension at thermophilic vs. mesophilic temperatures. This work demonstrates the ability to monitor DNA supercoiling at high temperature and opens the possibility to perform magnetic tweezers assays on thermophilic systems. The data allow for an estimation of the relative energies of base-pairing and DNA bending as a function of temperature and support speculation as to different general mechanisms of DNA opening in different environments. Lastly, our results imply that average in vivo DNA tensions range between 0.3 and 1.1 pN. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Whole-genome amplified DNA from stored dried blood spots is reliable in high resolution melting curve and sequencing analysis

    DEFF Research Database (Denmark)

    Winkel, Bo G; Hollegaard, Mads V; Olesen, Morten S

    2011-01-01

    BACKGROUND: The use of dried blood spots (DBS) samples in genomic workup has been limited by the relative low amounts of genomic DNA (gDNA) they contain. It remains to be proven that whole genome amplified DNA (wgaDNA) from stored DBS samples, constitutes a reliable alternative to gDNA.We wanted...

  1. Candidate DNA Barcode Tags Combined With High Resolution Melting (Bar-HRM Curve Analysis for Authentication of Senna alexandrina Mill. With Validation in Crude Drugs

    Directory of Open Access Journals (Sweden)

    Priyanka Mishra

    2018-03-01

    Full Text Available Senna alexandrina (Fabaceae is a globally recognized medicinal plant for its laxative properties as well as the only source of sennosides, and is highly exported bulk herb from India. Its major procurement is exclusively from limited cultivation, which leads to risks of deliberate or unintended adulteration. The market raw materials are in powdered or finished product form, which lead to difficulties in authentication. Here, DNA barcode tags based on chloroplast genes (rbcL and matK and intergenic spacers (psbA-trnH and ITS were developed for S. alexandrina along with the allied species. The ability and performance of the ITS1 region to discriminate among the Senna species resulted in the present proposal of the ITS1 tags as successful barcode. Further, these tags were coupled with high-resolution melting (HRM curve analysis in a real-time PCR genotyping method to derive Bar-HRM (Barcoding-HRM assays. Suitable HRM primer sets were designed through SNP detection and mutation scanning in genomic signatures of Senna species. The melting profiles of S. alexandrina and S. italica subsp. micrantha were almost identical and the remaining five species were clearly separated so that they can be differentiated by HRM method. The sensitivity of the method was utilized to authenticate market samples [Herbal Sample Assays (HSAs]. HSA01 (S. alexandrina crude drug sample from Bangalore and HSA06 (S. alexandrina crude drug sample from Tuticorin, Tamil Nadu, India were found to be highly contaminated with S. italica subsp. micrantha. Species admixture samples mixed in varying percentage was identified sensitively with detection of contamination as low as 1%. The melting profiles of PCR amplicons are clearly distinct, which enables the authentic differentiation of species by the HRM method. This study reveals that DNA barcoding coupled with HRM is an efficient molecular tool to authenticate Senna herbal products in the market for quality control in the drug

  2. Candidate DNA Barcode Tags Combined With High Resolution Melting (Bar-HRM) Curve Analysis for Authentication of Senna alexandrina Mill. With Validation in Crude Drugs.

    Science.gov (United States)

    Mishra, Priyanka; Shukla, Ashutosh K; Sundaresan, Velusamy

    2018-01-01

    Senna alexandrina (Fabaceae) is a globally recognized medicinal plant for its laxative properties as well as the only source of sennosides, and is highly exported bulk herb from India. Its major procurement is exclusively from limited cultivation, which leads to risks of deliberate or unintended adulteration. The market raw materials are in powdered or finished product form, which lead to difficulties in authentication. Here, DNA barcode tags based on chloroplast genes ( rbcL and matK ) and intergenic spacers ( psbA-trnH and ITS ) were developed for S. alexandrina along with the allied species. The ability and performance of the ITS1 region to discriminate among the Senna species resulted in the present proposal of the ITS1 tags as successful barcode. Further, these tags were coupled with high-resolution melting (HRM) curve analysis in a real-time PCR genotyping method to derive Bar-HRM (Barcoding-HRM) assays. Suitable HRM primer sets were designed through SNP detection and mutation scanning in genomic signatures of Senna species. The melting profiles of S. alexandrina and S . italica subsp. micrantha were almost identical and the remaining five species were clearly separated so that they can be differentiated by HRM method. The sensitivity of the method was utilized to authenticate market samples [Herbal Sample Assays (HSAs)]. HSA01 ( S. alexandrina crude drug sample from Bangalore) and HSA06 ( S. alexandrina crude drug sample from Tuticorin, Tamil Nadu, India) were found to be highly contaminated with S . italica subsp. micrantha . Species admixture samples mixed in varying percentage was identified sensitively with detection of contamination as low as 1%. The melting profiles of PCR amplicons are clearly distinct, which enables the authentic differentiation of species by the HRM method. This study reveals that DNA barcoding coupled with HRM is an efficient molecular tool to authenticate Senna herbal products in the market for quality control in the drug supply

  3. ITS2 barcoding DNA region combined with high resolution melting (HRM) analysis of Hyoscyami Semen, the mature seed of Hyoscyamus niger.

    Science.gov (United States)

    Xiong, Chao; Hu, Zhi-Gang; Tu, Yuan; Liu, He-Gang; Wang, Ping; Zhao, Ming-Ming; SHIi, Yu-Hua; Wu, Lan; Sun, Wei; Chen, Shi-Lin

    2016-12-01

    Hyoscyami Semen, the mature dried seed of Hyoscyamus niger L., has long been used as a traditional Chinese medicine to treat human diseases. Hyoscyami Semen is found in local markets in China. In markets, sellers and buyers commonly inadvertently mix the seeds of H. niger with the seeds of related species such as Hygrophila salicifolia (Vahl) Nees, Astragalus complanatus R. Br., Cuscuta australis R. Br., Cuscuta chinensis Lam., and Impatiens balsamina L. because of their similar morphologies or similar names. Thus, developing a reliable method for discriminating H. niger seeds from its adulterants is necessary to reduce confusion and ensure the safe use of Hyoscyami Semen. The present study was designed to evaluate the efficiency of high-resolution melting analysis combined with DNA barcoding (Bar-HRM) with internal transcribed spacer 2 to discriminate H. niger. Our results show that Bar-HRM successfully identified the adulterants and detected the proportion of H. niger DNA extract within an admixture. In particular, HRM detected H. niger DNA extract in A. complanatus DNA extract at concentrations as low as 1%. In conclusion, the Bar-HRM method developed in the present study for authenticating H. niger is rapid and cost-effective. It can be used in the future to guarantee the purity of Hyoscyami Semen for the clinical use. Copyright © 2016 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

  4. Detection and characterization of Leishmania (Leishmania and Leishmania (Viannia by SYBR green-based real-time PCR and high resolution melt analysis targeting kinetoplast minicircle DNA.

    Directory of Open Access Journals (Sweden)

    Marcello Ceccarelli

    Full Text Available Leishmaniasis is a neglected disease with a broad clinical spectrum which includes asymptomatic infection. A thorough diagnosis, able to distinguish and quantify Leishmania parasites in a clinical sample, constitutes a key step in choosing an appropriate therapy, making an accurate prognosis and performing epidemiological studies. Several molecular techniques have been shown to be effective in the diagnosis of leishmaniasis. In particular, a number of PCR methods have been developed on various target DNA sequences including kinetoplast minicircle constant regions. The first aim of this study was to develop a SYBR green-based qPCR assay for Leishmania (Leishmania infantum detection and quantification, using kinetoplast minicircle constant region as target. To this end, two assays were compared: the first used previously published primer pairs (qPCR1, whereas the second used a nested primer pairs generating a shorter PCR product (qPCR2. The second aim of this study was to evaluate the possibility to discriminate among subgenera Leishmania (Leishmania and Leishmania (Viannia using the qPCR2 assay followed by melting or High Resolution Melt (HRM analysis. Both assays used in this study showed good sensitivity and specificity, and a good correlation with standard IFAT methods in 62 canine clinical samples. However, the qPCR2 assay allowed to discriminate between Leishmania (Leishmania and Leishmania (Viannia subgenera through melting or HRM analysis. In addition to developing assays, we investigated the number and genetic variability of kinetoplast minicircles in the Leishmania (L. infantum WHO international reference strain (MHOM/TN/80/IPT1, highlighting the presence of minicircle subclasses and sequence heterogeneity. Specifically, the kinetoplast minicircle number per cell was estimated to be 26,566±1,192, while the subclass of minicircles amplifiable by qPCR2 was estimated to be 1,263±115. This heterogeneity, also observed in canine clinical

  5. Validation of methylation-sensitive high-resolution melting (MS-HRM) for the detection of stool DNA methylation in colorectal neoplasms.

    Science.gov (United States)

    Xiao, Zhujun; Li, Bingsheng; Wang, Guozhen; Zhu, Weisi; Wang, Zhongqiu; Lin, Jinfeng; Xu, Angao; Wang, Xinying

    2014-04-20

    Methylation-sensitive high-resolution melting (MS-HRM) is a new technique for assaying DNA methylation, but its feasibility for assaying stool in patients with colorectal cancer (CRC) is unknown. First, the MS-HRM and methylation-specific PCR (MSP) detection limits were tested. Second, the methylation statuses of SFRP2 and VIM were analyzed in stool samples by MS-HRM, and in matching tumor and normal colon tissues via bisulfite sequencing PCR (BSP). Third, a case-control study evaluated the diagnostic sensitivity and specificity of MS-HRM relative to results obtained with MSP and the fecal immunochemical test (FIT). Finally, the linearity and reproducibility of MS-HRM were assessed. The detection limits of MS-HRM and MSP were 1% and 5%, respectively. The diagnostic sensitivities of MS-HRM (87.3%, 55/63) in stool and BSP in matching tumor tissue (92.1%, 58/63) were highly consistent (κ=0.744). The MS-HRM assay detected 92.5% (37/40) methylation in CRCs, 94.4% (34/36) in advanced adenomas, and 8.8% (5/57) in normal controls. The results of MS-HRM analysis were stable and reliable and showed fairly good linearity for both SFRP2 (PHRM shows potential for CRC screening. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Quantifying DNA melting transitions using single-molecule force spectroscopy

    International Nuclear Information System (INIS)

    Calderon, Christopher P; Chen, W-H; Harris, Nolan C; Kiang, C-H; Lin, K-J

    2009-01-01

    We stretched a DNA molecule using an atomic force microscope (AFM) and quantified the mechanical properties associated with B and S forms of double-stranded DNA (dsDNA), molten DNA, and single-stranded DNA. We also fit overdamped diffusion models to the AFM time series and used these models to extract additional kinetic information about the system. Our analysis provides additional evidence supporting the view that S-DNA is a stable intermediate encountered during dsDNA melting by mechanical force. In addition, we demonstrated that the estimated diffusion models can detect dynamical signatures of conformational degrees of freedom not directly observed in experiments.

  7. Quantifying DNA melting transitions using single-molecule force spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Calderon, Christopher P [Department of Computational and Applied Mathematics, Rice University, Houston, TX (United States); Chen, W-H; Harris, Nolan C; Kiang, C-H [Department of Physics and Astronomy, Rice University, Houston, TX (United States); Lin, K-J [Department of Chemistry, National Chung Hsing University, Taichung, Taiwan (China)], E-mail: chkiang@rice.edu

    2009-01-21

    We stretched a DNA molecule using an atomic force microscope (AFM) and quantified the mechanical properties associated with B and S forms of double-stranded DNA (dsDNA), molten DNA, and single-stranded DNA. We also fit overdamped diffusion models to the AFM time series and used these models to extract additional kinetic information about the system. Our analysis provides additional evidence supporting the view that S-DNA is a stable intermediate encountered during dsDNA melting by mechanical force. In addition, we demonstrated that the estimated diffusion models can detect dynamical signatures of conformational degrees of freedom not directly observed in experiments.

  8. Ribo HRM--detection of inter- and intra-species polymorphisms within ribosomal DNA by high resolution melting analysis supported by application of artificial allelic standards.

    Science.gov (United States)

    Masny, Aleksander; Jagiełło, Agata; Płucienniczak, Grażyna; Golab, Elzbieta

    2012-09-01

    Ribo HRM, a single-tube PCR and high resolution melting (HRM) assay for detection of polymorphisms in the large subunit ribosomal DNA expansion segment V, was developed on a Trichinella model. Four Trichinella species: T. spiralis (isolates ISS3 and ISS160), T. nativa (isolates ISS10 and ISS70), T. britovi (isolates ISS2 and ISS392) and T. pseudospiralis (isolates ISS13 and ISS1348) were genotyped. Cloned allelic variants of the expansion segment V were used as standards to prepare reference HRM curves characteristic for single sequences and mixtures of several cloned sequences imitating allelic composition detected in Trichinella isolates. Using the primer pair Tsr1 and Trich1bi, it was possible to amplify a fragment of the ESV and detect PCR products obtained from the genomic DNA of pools of larvae belonging to the four investigated species: T. pseudospiralis, T. spiralis, T. britovi and T. nativa, in a single tube Real-Time PCR reaction. Differences in the shape of the HRM curves of Trichinella isolates suggested the presence of differences between examined isolates of T. nativa, T. britovi and T. pseudospiralis species. No differences were observed between T. spiralis isolates. The presence of polymorphisms within the amplified ESV sequence fragment of T. nativa T. britovi and T. pseudospiralis was confirmed by sequencing of the cloned PCR products. Novel sequences were discovered and deposited in GenBank (GenBank IDs: JN971020-JN971027, JN120902.1, JN120903.1, JN120904.1, JN120906.1, JN120905.1). Screening the ESV region of Trichinella for polymorphism is possible using the genotyping assay Ribo HRM at the current state of its development. The Ribo HRM assay could be useful in phylogenetic studies of the Trichinella genus. Copyright © 2012 Elsevier B.V. All rights reserved.

  9. On high-pressure melting of tantalum

    Science.gov (United States)

    Luo, Sheng-Nian; Swift, Damian C.

    2007-01-01

    The issues related to high-pressure melting of Ta are discussed within the context of diamond-anvil cell (DAC) and shock wave experiments, theoretical calculations and common melting models. The discrepancies between the extrapolations of the DAC melting curve and the melting point inferred from shock wave experiments, cannot be reconciled either by superheating or solid-solid phase transition. The failure to reproduce low-pressure DAC melting curve by melting models such as dislocation-mediated melting and the Lindemann law, and molecular dynamics and quantum mechanics-based calculations, undermines their predictions at moderate and high pressures. Despite claims to the contrary, the melting curve of Ta (as well as Mo and W) remains inconclusive at high pressures.

  10. Layered graphene-mica substrates induce melting of DNA origami

    Science.gov (United States)

    Green, Nathaniel S.; Pham, Phi H. Q.; Crow, Daniel T.; Burke, Peter J.; Norton, Michael L.

    2018-04-01

    Monolayer graphene supported on mica substrates induce melting of cross-shaped DNA origami. This behavior can be contrasted with the case of origami on graphene on graphite, where an expansion or partially re-organized structure is observed. On mica, only well-formed structures are observed. Comparison of the morphological differences observed for these probes after adsorption on these substrates provides insights into the sensitivity of DNA based nanostructures to the properties of the graphene monolayer, as modified by its substrate.

  11. A DNA Melting Exercise for a Large Laboratory Class

    Science.gov (United States)

    Levine, Lauren A.; Junker, Matthew; Stark, Myranda; Greenleaf, Dustin

    2015-01-01

    A simple and economical experimental setup is described that enables multiple individuals or groups within a laboratory class to measure the thermal melting of double stranded DNA simultaneously. The setup utilizes a basic spectrophotometer capable of measuring absorbance at 260 nm, UV plastic cuvettes, and a stirring hot plate. Students measure…

  12. Melt processed high-temperature superconductors

    CERN Document Server

    1993-01-01

    The achievement of large critical currents is critical to the applications of high-temperature superconductors. Recent developments have shown that melt processing is suitable for producing high J c oxide superconductors. Using magnetic forces between such high J c oxide superconductors and magnets, a person could be levitated.This book has grown largely out of research works on melt processing of high-temperature superconductors conducted at ISTEC Superconductivity Research Laboratory. The chapters build on melt processing, microstructural characterization, fundamentals of flux pinning, criti

  13. Decoding DNA labels by melting curve analysis using real-time PCR.

    Science.gov (United States)

    Balog, József A; Fehér, Liliána Z; Puskás, László G

    2017-12-01

    Synthetic DNA has been used as an authentication code for a diverse number of applications. However, existing decoding approaches are based on either DNA sequencing or the determination of DNA length variations. Here, we present a simple alternative protocol for labeling different objects using a small number of short DNA sequences that differ in their melting points. Code amplification and decoding can be done in two steps using quantitative PCR (qPCR). To obtain a DNA barcode with high complexity, we defined 8 template groups, each having 4 different DNA templates, yielding 158 (>2.5 billion) combinations of different individual melting temperature (Tm) values and corresponding ID codes. The reproducibility and specificity of the decoding was confirmed by using the most complex template mixture, which had 32 different products in 8 groups with different Tm values. The industrial applicability of our protocol was also demonstrated by labeling a drone with an oil-based paint containing a predefined DNA code, which was then successfully decoded. The method presented here consists of a simple code system based on a small number of synthetic DNA sequences and a cost-effective, rapid decoding protocol using a few qPCR reactions, enabling a wide range of authentication applications.

  14. CtGEM typing: Discrimination of Chlamydia trachomatis ocular and urogenital strains and major evolutionary lineages by high resolution melting analysis of two amplified DNA fragments.

    Science.gov (United States)

    Giffard, Philip M; Andersson, Patiyan; Wilson, Judith; Buckley, Cameron; Lilliebridge, Rachael; Harris, Tegan M; Kleinecke, Mariana; O'Grady, Kerry-Ann F; Huston, Wilhelmina M; Lambert, Stephen B; Whiley, David M; Holt, Deborah C

    2018-01-01

    Chlamydia trachomatis infects the urogenital tract (UGT) and eyes. Anatomical tropism is correlated with variation in the major outer membrane protein encoded by ompA. Strains possessing the ocular ompA variants A, B, Ba and C are typically found within the phylogenetically coherent "classical ocular lineage". However, variants B, Ba and C have also been found within three distinct strains in Australia, all associated with ocular disease in children and outside the classical ocular lineage. CtGEM genotyping is a method for detecting and discriminating ocular strains and also the major phylogenetic lineages. The rationale was facilitation of surveillance to inform responses to C. trachomatis detection in UGT specimens from young children. CtGEM typing is based on high resolution melting analysis (HRMA) of two PCR amplified fragments with high combinatorial resolving power, as defined by computerised comparison of 65 whole genomes. One fragment is from the hypothetical gene defined by Jali-1891 in the C. trachomatis B_Jali20 genome, while the other is from ompA. Twenty combinatorial CtGEM types have been shown to exist, and these encompass unique genotypes for all known ocular strains, and also delineate the TI and T2 major phylogenetic lineages, identify LGV strains and provide additional resolution beyond this. CtGEM typing and Sanger sequencing were compared with 42 C. trachomatis positive clinical specimens, and there were no disjunctions. CtGEM typing is a highly efficient method designed and tested using large scale comparative genomics. It divides C. trachomatis into clinically and biologically meaningful groups, and may have broad application in surveillance.

  15. High Resolution Melting Analysis for fast and cheap polymorphism screening of marine populations

    OpenAIRE

    sprotocols

    2015-01-01

    Authors: Anne-Leila Meistertzheim, Isabelle Calves, Sébastien Artigaud, Carolyn S. Friedman, Christine Paillard, Jean Laroche & Claude Ferec ### Abstract This protocol permits the mutation scanning of PCR products by high-resolution DNA melting analysis requiring the inclusion of a saturating intercalating dye in the PCR mix without labelled probe. During a scanning process, fluorescent melting curves of PCR amplicons are analyzed. Mutations modifying melting curve shapes, are allowed...

  16. Methylation-Sensitive High Resolution Melting (MS-HRM).

    Science.gov (United States)

    Hussmann, Dianna; Hansen, Lise Lotte

    2018-01-01

    Methylation-Sensitive High Resolution Melting (MS-HRM) is an in-tube, PCR-based method to detect methylation levels at specific loci of interest. A unique primer design facilitates a high sensitivity of the assays enabling detection of down to 0.1-1% methylated alleles in an unmethylated background.Primers for MS-HRM assays are designed to be complementary to the methylated allele, and a specific annealing temperature enables these primers to anneal both to the methylated and the unmethylated alleles thereby increasing the sensitivity of the assays. Bisulfite treatment of the DNA prior to performing MS-HRM ensures a different base composition between methylated and unmethylated DNA, which is used to separate the resulting amplicons by high resolution melting.The high sensitivity of MS-HRM has proven useful for detecting cancer biomarkers in a noninvasive manner in urine from bladder cancer patients, in stool from colorectal cancer patients, and in buccal mucosa from breast cancer patients. MS-HRM is a fast method to diagnose imprinted diseases and to clinically validate results from whole-epigenome studies. The ability to detect few copies of methylated DNA makes MS-HRM a key player in the quest for establishing links between environmental exposure, epigenetic changes, and disease.

  17. Product differentiation by analysis of DNA melting curves during the polymerase chain reaction.

    Science.gov (United States)

    Ririe, K M; Rasmussen, R P; Wittwer, C T

    1997-02-15

    A microvolume fluorometer integrated with a thermal cycler was used to acquire DNA melting curves during polymerase chain reaction by fluorescence monitoring of the double-stranded DNA specific dye SYBR Green I. Plotting fluorescence as a function of temperature as the thermal cycler heats through the dissociation temperature of the product gives a DNA melting curve. The shape and position of this DNA melting curve are functions of the GC/AT ratio, length, and sequence and can be used to differentiate amplification products separated by less than 2 degrees C in melting temperature. Desired products can be distinguished from undesirable products, in many cases eliminating the need for gel electrophoresis. Analysis of melting curves can extend the dynamic range of initial template quantification when amplification is monitored with double-stranded DNA specific dyes. Complete amplification and analysis of products can be performed in less than 15 min.

  18. Permeability and 3-D melt geometry in shear-induced high melt fraction conduits

    Science.gov (United States)

    Zhu, W.; Cordonnier, B.; Qi, C.; Kohlstedt, D. L.

    2017-12-01

    Observations of dunite channels in ophiolites and uranium-series disequilibria in mid-ocean ridge basalt suggest that melt transport in the upper mantle beneath mid-ocean ridges is strongly channelized. Formation of high melt fraction conduits could result from mechanical shear, pyroxene dissolution, and lithological partitioning. Deformation experiments (e.g. Holtzman et al., 2003) demonstrate that shear stress causes initially homogeneously distributed melt to segregate into an array of melt-rich bands, flanked by melt-depleted regions. At the same average melt fraction, the permeability of high melt fraction conduits could be orders of magnitude higher than that of their homogenous counterparts. However, it is difficult to determine the permeability of melt-rich bands. Using X-ray synchrotron microtomography, we obtained high-resolution images of 3-dimensional (3-D) melt distribution in a partially molten rock containing shear-induced high melt fraction conduits. Sample CQ0705, an olivine-alkali basalt aggregate with a nominal melt fraction of 4%, was deformed in torsion at a temperature of 1473 K and a confining pressure of 300 MPa to a shear strain of 13.3. A sub-volume of CQ0705 encompassing 3-4 melt-rich bands was imaged. Microtomography data were reduced to binary form so that solid olivine is distinguishable from basalt glass. At a spatial resolution of 160 nm, the 3-D images reveal the shape and connectedness of melt pockets in the melt-rich bands. Thin melt channels formed at grain edges are connected at large melt nodes at grain corners. Initial data analysis shows a clear preferred orientation of melt pockets alignment subparallel to the melt-rich band. We use the experimentally determined geometrical parameters of melt topology to create a digital rock with identical 3-D microstructures. Stokes flow simulations are conducted on the digital rock to obtain the permeability tensor. Using this digital rock physics approach, we determine how deformation

  19. High-resolution melt-curve analysis of random amplified polymorphic DNA (RAPD-HRM) for the characterisation of pathogenic leptospires: intra-serovar divergence, inter-serovar convergence, and evidence of attenuation in Leptospira reference collections.

    Science.gov (United States)

    Tulsiani, S M; Craig, S B; Graham, G C; Cobbold, R C; Dohnt, M F; Burns, M-A; Jansen, C C; Leung, L K-P; Field, H E; Smythe, L D

    2010-07-01

    High-resolution melt-curve analysis of random amplified polymorphic DNA (RAPD-HRM) is a novel technology that has emerged as a possible method to characterise leptospires to serovar level. RAPD-HRM has recently been used to measure intra-serovar convergence between strains of the same serovar as well as inter-serovar divergence between strains of different serovars. The results indicate that intra-serovar heterogeneity and inter-serovar homogeneity may limit the application of RAPD-HRM in routine diagnostics. They also indicate that genetic attenuation of aged, high-passage-number isolates could undermine the use of RAPD-HRM or any other molecular technology. Such genetic attenuation may account for a general decrease seen in titres of rabbit hyperimmune antibodies over time. Before RAPD-HRM can be further advanced as a routine diagnostic tool, strains more representative of the wild-type serovars of a given region need to be identified. Further, RAPD-HRM analysis of reference strains indicates that the routine renewal of reference collections, with new isolates, may be needed to maintain the genetic integrity of the collections.

  20. Comparative melting and healing of B-DNA and Z-DNA by an infrared laser pulse

    Energy Technology Data Exchange (ETDEWEB)

    Man, Viet Hoang; Pan, Feng; Sagui, Celeste, E-mail: sagui@ncsu.edu; Roland, Christopher, E-mail: cmroland@ncsu.edu [Department of Physics, North Carolina State University, Raleigh, North Carolina 27695-8202 (United States)

    2016-04-14

    We explore the use of a fast laser melting simulation approach combined with atomistic molecular dynamics simulations in order to determine the melting and healing responses of B-DNA and Z-DNA dodecamers with the same d(5′-CGCGCGCGCGCG-3′){sub 2} sequence. The frequency of the laser pulse is specifically tuned to disrupt Watson-Crick hydrogen bonds, thus inducing melting of the DNA duplexes. Subsequently, the structures relax and partially refold, depending on the field strength. In addition to the inherent interest of the nonequilibrium melting process, we propose that fast melting by an infrared laser pulse could be used as a technique for a fast comparison of relative stabilities of same-sequence oligonucleotides with different secondary structures with full atomistic detail of the structures and solvent. This could be particularly useful for nonstandard secondary structures involving non-canonical base pairs, mismatches, etc.

  1. Comparative melting and healing of B-DNA and Z-DNA by an infrared laser pulse

    International Nuclear Information System (INIS)

    Man, Viet Hoang; Pan, Feng; Sagui, Celeste; Roland, Christopher

    2016-01-01

    We explore the use of a fast laser melting simulation approach combined with atomistic molecular dynamics simulations in order to determine the melting and healing responses of B-DNA and Z-DNA dodecamers with the same d(5′-CGCGCGCGCGCG-3′) 2 sequence. The frequency of the laser pulse is specifically tuned to disrupt Watson-Crick hydrogen bonds, thus inducing melting of the DNA duplexes. Subsequently, the structures relax and partially refold, depending on the field strength. In addition to the inherent interest of the nonequilibrium melting process, we propose that fast melting by an infrared laser pulse could be used as a technique for a fast comparison of relative stabilities of same-sequence oligonucleotides with different secondary structures with full atomistic detail of the structures and solvent. This could be particularly useful for nonstandard secondary structures involving non-canonical base pairs, mismatches, etc.

  2. Assembly and melting of DNA nanotubes from single-sequence tiles

    International Nuclear Information System (INIS)

    Sobey, T L; Renner, S; Simmel, F C

    2009-01-01

    DNA melting and renaturation studies are an extremely valuable tool to study the kinetics and thermodynamics of duplex dissociation and reassociation reactions. These are important not only in a biological or biotechnological context, but also for DNA nanotechnology which aims at the construction of molecular materials by DNA self-assembly. We here study experimentally the formation and melting of a DNA nanotube structure, which is composed of many copies of an oligonucleotide containing several palindromic sequences. This is done using temperature-controlled UV absorption measurements correlated with atomic force microscopy, fluorescence microscopy and transmission electron microscopy techniques. In the melting studies, important factors such as DNA strand concentration, hierarchy of assembly and annealing protocol are investigated. Assembly and melting of the nanotubes are shown to proceed via different pathways. Whereas assembly occurs in several hierarchical steps related to the formation of tiles, lattices and tubes, melting of DNA nanotubes appears to occur in a single step. This is proposed to relate to fundamental differences between closed, three-dimensional tube-like structures and open, two-dimensional lattices. DNA melting studies can lead to a better understanding of the many factors that affect the assembly process which will be essential for the assembly of increasingly complex DNA nanostructures.

  3. High Resolution Melting (HRM) applied to wine authenticity.

    Science.gov (United States)

    Pereira, Leonor; Gomes, Sónia; Castro, Cláudia; Eiras-Dias, José Eduardo; Brazão, João; Graça, António; Fernandes, José R; Martins-Lopes, Paula

    2017-02-01

    Wine authenticity methods are in increasing demand mainly in Denomination of Origin designations. The DNA-based methodologies are a reliable means of tracking food/wine varietal composition. The main aim of this work was the study of High Resolution Melting (HRM) application as a screening method for must and wine authenticity. Three sample types (leaf, must and wine) were used to validate the three developed HRM assays (Vv1-705bp; Vv2-375bp; and Vv3-119bp). The Vv1 HRM assay was only successful when applied to leaf and must samples. The Vv2 HRM assay successfully amplified all sample types, allowing genotype discrimination based on melting temperature values. The smallest amplicon, Vv3, produced a coincident melting curve shape in all sample types (leaf and wine) with corresponding genotypes. This study presents sensitive, rapid and efficient HRM assays applied for the first time to wine samples suitable for wine authenticity purposes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Simultaneous Profiling of DNA Mutation and Methylation by Melting Analysis Using Magnetoresistive Biosensor Array

    DEFF Research Database (Denmark)

    Rizzi, Giovanni; Lee, Jung-Rok; Dahl, Christina

    2017-01-01

    specificity. Genomic (mutation) or bisulphite-treated (methylation) DNA is amplified using nondiscriminatory primers, and the amplicons are then hybridized to a giant magnetoresistive (GMR) biosensor array followed by melting curve measurements. The GMR biosensor platform offers scalable multiplexed detection...

  5. High-resolution melt-curve analysis of random amplified polymorphic DNA (RAPD-HRM) for the characterisation of pathogenic leptospires

    DEFF Research Database (Denmark)

    Tulsiani, Suhella; Craig, S B; Graham, G C

    2010-01-01

    as well as inter-serovar divergence between strains of different serovars. The results indicate that intra-serovar heterogeneity and inter-serovar homogeneity may limit the application of RAPD-HRM in routine diagnostics. They also indicate that genetic attenuation of aged, high-passage-number isolates...... could undermine the use of RAPD-HRM or any other molecular technology. Such genetic attenuation may account for a general decrease seen in titres of rabbit hyperimmune antibodies over time. Before RAPD-HRM can be further advanced as a routine diagnostic tool, strains more representative of the wild...

  6. Evaluation of errors in determination of DNA melting curve registered with differential scanning calorimetry

    International Nuclear Information System (INIS)

    Lando, D.Y.; Fridman, A.S.; Galyuk, E.N.; Dalyan, Y.B.; Grigoryan, I.E.; Haroutiunian, S.G.

    2013-01-01

    The differential scanning calorimetry (DSC) is more sensitive than UV absorption spectrophotometry as a tool for the measurement of DNA melting curves. The advantage of DSC is a direct determination of differential melting curves (DMC) obtained without numerical differentiation. However, the difference between the helix-coil transition enthalpies of AT and GC base pairs can cause distortions in the shape of melting curve. Up to date, the errors caused by those distortions were not evaluated. In this study, a simple procedure of recalculation of a calorimetric DMC into a real DMC is developed. It demonstrates that the 'real' melting curve and differential melting curve deviate very slightly from the same two curves calculated from DSC data. The melting temperature and the temperature melting range are usually the same even if the difference in the enthalpies is several times higher than a real one

  7. Microstructures define melting of molybdenum at high pressures

    Science.gov (United States)

    Hrubiak, Rostislav; Meng, Yue; Shen, Guoyin

    2017-03-01

    High-pressure melting anchors the phase diagram of a material, revealing the effect of pressure on the breakdown of the ordering of atoms in the solid. An important case is molybdenum, which has long been speculated to undergo an exceptionally steep increase in melting temperature when compressed. On the other hand, previous experiments showed nearly constant melting temperature as a function of pressure, in large discrepancy with theoretical expectations. Here we report a high-slope melting curve in molybdenum by synchrotron X-ray diffraction analysis of crystalline microstructures, generated by heating and subsequently rapidly quenching samples in a laser-heated diamond anvil cell. Distinct microstructural changes, observed at pressures up to 130 gigapascals, appear exclusively after melting, thus offering a reliable melting criterion. In addition, our study reveals a previously unsuspected transition in molybdenum at high pressure and high temperature, which yields highly textured body-centred cubic nanograins above a transition temperature.

  8. UV-induced influence of N-nitrosoamines on melting parameters of DNA in vitro

    International Nuclear Information System (INIS)

    Yamshanov, V.A.

    1979-01-01

    The results of studies have shown the UV-induced decrease of melting temperatures of the DNA of E. coli and chick erythrocytes under the influence of simple N-nitrosoamines (NDMA, NDEA, NDPA). Either UV or nitrosoamines separately failed to effect the DNA or their action was insignificant. It is suggested that this effect may be partly due to the action of UV on DNA

  9. UV-induced influence of N-nitrosoamines on melting parameters of DNA in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Yamshanov, V A [Nauchno-Issledovatel' skij Inst. Onkologii, Leningrad (USSR)

    1979-07-01

    The results of studies have shown the UV-induced decrease of melting temperatures of the DNA of E. coli and chick erythrocytes under the influence of simple N-nitrosoamines (NDMA, NDEA, NDPA). Either UV or nitrosoamines separately failed to effect the DNA or their action was insignificant. It is suggested that this effect may be partly due to the action of UV on DNA.

  10. On-chip magnetic bead-based DNA melting curve analysis using a magnetoresistive sensor

    International Nuclear Information System (INIS)

    Rizzi, Giovanni; Østerberg, Frederik W.; Henriksen, Anders D.; Dufva, Martin; Hansen, Mikkel F.

    2015-01-01

    We present real-time measurements of DNA melting curves in a chip-based system that detects the amount of surface-bound magnetic beads using magnetoresistive magnetic field sensors. The sensors detect the difference between the amount of beads bound to the top and bottom sensor branches of the differential sensor geometry. The sensor surfaces are functionalized with wild type (WT) and mutant type (MT) capture probes, differing by a single base insertion (a single nucleotide polymorphism, SNP). Complementary biotinylated targets in suspension couple streptavidin magnetic beads to the sensor surface. The beads are magnetized by the field arising from the bias current passed through the sensors. We demonstrate the first on-chip measurements of the melting of DNA hybrids upon a ramping of the temperature. This overcomes the limitation of using a single washing condition at constant temperature. Moreover, we demonstrate that a single sensor bridge can be used to genotype a SNP. - Highlights: • We apply magnetoresistive sensors to study solid-surface hybridization kinetics of DNA. • We measure DNA melting profiles for perfectly matching DNA duplexes and for a single base mismatch. • We present a procedure to correct for temperature dependencies of the sensor output. • We reliably extract melting temperatures for the DNA hybrids. • We demonstrate direct measurement of differential binding signal for two probes on a single sensor

  11. Study of force induced melting of dsDNA as a function of length and conformation

    International Nuclear Information System (INIS)

    Danilowicz, Claudia; Hatch, Kristi; Conover, Alyson; Gunaratne, Ruwan; Coljee, Vincent; Prentiss, Mara; Ducas, Theodore

    2010-01-01

    We measure the constant force required to melt double-stranded (ds) DNA as a function of length for lengths from 12 to 100 000 base pairs, where the force is applied to the 3'3' or 5'5' ends of the dsDNA. Molecules with 32 base pairs or fewer melt before overstretching. For these short molecules, the melting force is independent of the ends to which the force is applied and the shear force as a function of length is well described by de Gennes theory with a de Gennes length of less than 10 bp. Molecules with lengths of 500 base pairs or more overstretch before melting. For these long molecules, the melting force depends on the ends to which the force is applied. The melting force as a function of length increases even when the length exceeds 1000 bp, where the length dependence is inconsistent with de Gennes theory. Finally, we expand de Gennes melting theory to 3'5' pulling and compare the predictions with experimental results.

  12. On-chip magnetic bead-based DNA melting curve analysis using a magnetoresistive sensor

    DEFF Research Database (Denmark)

    Rizzi, Giovanni; Østerberg, Frederik Westergaard; Henriksen, Anders Dahl

    2014-01-01

    We present real-time measurements of DNA melting curves in a chip-based system that detects the amount of surface-bound magnetic beads using magnetoresistive magnetic field sensors. The sensors detect the difference between the amount of beads bound to the top and bottom sensor branches....... The beads are magnetized by the field arising from the bias current passed through the sensors. We demonstrate the first on-chip measurements of the melting of DNA hybrids upon a ramping of the temperature. This overcomes the limitation of using a single washing condition at constant temperature. Moreover...

  13. High resolution melting for mutation scanning of TP53 exons 5–8

    International Nuclear Information System (INIS)

    Krypuy, Michael; Dobrovic, Alexander; Ahmed, Ahmed Ashour; Etemadmoghadam, Dariush; Hyland, Sarah J; Australian Ovarian Cancer Study Group; Fazio, Anna de; Fox, Stephen B; Brenton, James D; Bowtell, David D

    2007-01-01

    p53 is commonly inactivated by mutations in the DNA-binding domain in a wide range of cancers. As mutant p53 often influences response to therapy, effective and rapid methods to scan for mutations in TP53 are likely to be of clinical value. We therefore evaluated the use of high resolution melting (HRM) as a rapid mutation scanning tool for TP53 in tumour samples. We designed PCR amplicons for HRM mutation scanning of TP53 exons 5 to 8 and tested them with DNA from cell lines hemizygous or homozygous for known mutations. We assessed the sensitivity of each PCR amplicon using dilutions of cell line DNA in normal wild-type DNA. We then performed a blinded assessment on ovarian tumour DNA samples that had been previously sequenced for mutations in TP53 to assess the sensitivity and positive predictive value of the HRM technique. We also performed HRM analysis on breast tumour DNA samples with unknown TP53 mutation status. One cell line mutation was not readily observed when exon 5 was amplified. As exon 5 contained multiple melting domains, we divided the exon into two amplicons for further screening. Sequence changes were also introduced into some of the primers to improve the melting characteristics of the amplicon. Aberrant HRM curves indicative of TP53 mutations were observed for each of the samples in the ovarian tumour DNA panel. Comparison of the HRM results with the sequencing results revealed that each mutation was detected by HRM in the correct exon. For the breast tumour panel, we detected seven aberrant melt profiles by HRM and subsequent sequencing confirmed the presence of these and no other mutations in the predicted exons. HRM is an effective technique for simple and rapid scanning of TP53 mutations that can markedly reduce the amount of sequencing required in mutational studies of TP53

  14. Determination of melting curves of irradiated DNA preparations and of preparations isolated from irradiated calf lymph nodes

    International Nuclear Information System (INIS)

    Grabowska, B.

    1977-01-01

    Measurements of melting curves enabled to establish differences of melting temperature, hyperchromic effect and breadth of the helix - coil phase transition dependent on dose of the ionizing radiation applied and on kind of the irradiated object. Changes of the investigated parameters of DNA irradiated after isolation were detectably more pronounced that of DNA from irradiated lymph nodes. The obtained results suggest a protective role of tissue to the secondary structure of DNA. (author)

  15. Melted flux liquids in high-Tc superconductors

    International Nuclear Information System (INIS)

    Nelson, D.R.

    1989-01-01

    A theory of the entangles flux liquids which arise in the new high-T c superconductors is reviewed. New physics appears because of the weak interplanar couplings and high critical temperatures in these materials. Flux line wandering melts the conventional Abrikosov flux lattice over large portions of the phase diagram and leads to a novel entangled vortex state. The authors suggest that a heavily entangled flux liquid could exhibit glassy behavior on experimental time scales, in analogy with viscoelastic behavior in dense polymer melts

  16. Flux lattice melting in high-Tc superconductors

    International Nuclear Information System (INIS)

    Houghton, A.; Pelcovits, R.A.; Sudbo, A.

    1989-01-01

    We derive the wave-vector-dependent elastic moduli for a flux line lattice in compounds with underlying tetragonal crystalline symmetry. We find that it is essential to retain wave-vector dependence of the moduli when dealing with compounds where κ is large, as it is in the high-T c materials. We use our results to establish a Lindemann criterion for flux lattice melting, which we then compare with experimental data on two materials, and find excellent agreement. The melting curves are suppressed well below the mean-field superconducting-normal transition line and are linear in temperature over a wide range of magnetic fields. The point H=0, T=T c is approached as 1-T/T c ∼H 1/2 . The degree of suppression of the melting curves among the different compounds is accounted for in the main by differences in mass anisotropy

  17. Analysis of HIV using a high resolution melting (HRM) diversity assay: automation of HRM data analysis enhances the utility of the assay for analysis of HIV incidence.

    Science.gov (United States)

    Cousins, Matthew M; Swan, David; Magaret, Craig A; Hoover, Donald R; Eshleman, Susan H

    2012-01-01

    HIV diversity may be a useful biomarker for discriminating between recent and non-recent HIV infection. The high resolution melting (HRM) diversity assay was developed to quantify HIV diversity in viral populations without sequencing. In this assay, HIV diversity is expressed as a single numeric HRM score that represents the width of a melting peak. HRM scores are highly associated with diversity measures obtained with next generation sequencing. In this report, a software package, the HRM Diversity Assay Analysis Tool (DivMelt), was developed to automate calculation of HRM scores from melting curve data. DivMelt uses computational algorithms to calculate HRM scores by identifying the start (T1) and end (T2) melting temperatures for a DNA sample and subtracting them (T2 - T1 =  HRM score). DivMelt contains many user-supplied analysis parameters to allow analyses to be tailored to different contexts. DivMelt analysis options were optimized to discriminate between recent and non-recent HIV infection and to maximize HRM score reproducibility. HRM scores calculated using DivMelt were compared to HRM scores obtained using a manual method that is based on visual inspection of DNA melting curves. HRM scores generated with DivMelt agreed with manually generated HRM scores obtained from the same DNA melting data. Optimal parameters for discriminating between recent and non-recent HIV infection were identified. DivMelt provided greater discrimination between recent and non-recent HIV infection than the manual method. DivMelt provides a rapid, accurate method of determining HRM scores from melting curve data, facilitating use of the HRM diversity assay for large-scale studies.

  18. Duchenne muscular dystrophy: High-resolution melting curve ...

    African Journals Online (AJOL)

    Duchenne muscular dystrophy: High-resolution melting curve analysis as an affordable diagnostic mutation scanning tool in a South African cohort. ... Genetic screening for D/BMD in South Africa currently includes multiple ligase-dependent probe amplification (MLPA) for exonic deletions and duplications and linkage ...

  19. Rheology of High-Melt-Strength Polypropylene for Additive Manufacturing

    DEFF Research Database (Denmark)

    Hofstätter, Thomas; Kamleitner, Florian; Jagenteufel, Ralf

    Acrylonitrile butadiene styrene (ABS) is a widely used material for additive manufacturing (AM) fused deposition modeling (FDM). The rheological properties of high-melt-strength polypropylene (HMS-PP) were compared to commercially available ABS 250 filament to study the possibility of using...

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

    KAUST Repository

    Starikov, E. B.; Nordén, Bengt

    2009-01-01

    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

  1. High-pressure melting curve of KCl: Evidence against lattice-instability theories of melting

    International Nuclear Information System (INIS)

    Ross, M.; Wolf, G.

    1986-01-01

    We show that the large curvature in the T-P melting curve of KCl is the result of a reordering of the liquid to a more densely packed arrangement. As a result theories of melting, such as the instability model, which do not take into account the structure of the liquid fail to predict the correct pressure dependence of the melting curve

  2. 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.

  3. Difference in melting profiles of gamma irradiated DNA from chicken erythrocytes and from Escherichia coli B/r

    International Nuclear Information System (INIS)

    Kopff, J.; Miller, G.; Leyko, W.

    1977-01-01

    Effects of gamma irradiation on melting curves of DNA from chicken erythrocytes and Escherichia coli B/r were compared. Considerable changes, following gamma irradiation in the case of chicken erythrocytes DNA and no changes in the case of DNA from Escherichia coli B/r were observed. To explain the lack of changes in gamma irradiated samples of DNA from Escherichia coli B/r it was assumed that the original effects of irradiation were obscured by the process of renaturation of DNA. To exclude the above mentioned effect, examination of gamma irradiated DNA from Escherichia coli B/r was carried out with the addition of formaldehyde immediately after irradiation of the sample. Using this procedure changes of melting profiles of DNA from Escherichia coli B/r were demonstrated. (author)

  4. High resolution melt curve analysis based on methylation status for human semen identification.

    Science.gov (United States)

    Fachet, Caitlyn; Quarino, Lawrence; Karnas, K Joy

    2017-03-01

    A high resolution melt curve assay to differentiate semen from blood, saliva, urine, and vaginal fluid based on methylation status at the Dapper Isoform 1 (DACT1) gene was developed. Stains made from blood, saliva, urine, semen, and vaginal fluid were obtained from volunteers and DNA was isolated using either organic extraction (saliva, urine, and vaginal fluid) or Chelex ® 100 extraction (blood and semen). Extracts were then subjected to bisulfite modification in order to convert unmethylated cytosines to uracil, consequently creating sequences whose amplicons have melt curves that vary depending on their initial methylation status. When primers designed to amplify the promoter region of the DACT1 gene were used, DNA from semen samples was distinguishable from other fluids by a having a statistically significant lower melting temperature. The assay was found to be sperm-significant since semen from a vasectomized man produced a melting temperature similar to the non-semen body fluids. Blood and semen stains stored up to 5 months and tested at various intervals showed little variation in melt temperature indicating the methylation status was stable during the course of the study. The assay is a more viable method for forensic science practice than most molecular-based methods for body fluid stain identification since it is time efficient and utilizes instrumentation common to forensic biology laboratories. In addition, the assay is advantageous over traditional presumptive chemical methods for body fluid identification since results are confirmatory and the assay offers the possibility of multiplexing which may test for multiple body fluids simultaneously.

  5. Automated Microfluidic Platform for Serial Polymerase Chain Reaction and High-Resolution Melting Analysis.

    Science.gov (United States)

    Cao, Weidong; Bean, Brian; Corey, Scott; Coursey, Johnathan S; Hasson, Kenton C; Inoue, Hiroshi; Isano, Taisuke; Kanderian, Sami; Lane, Ben; Liang, Hongye; Murphy, Brian; Owen, Greg; Shinoda, Nobuhiko; Zeng, Shulin; Knight, Ivor T

    2016-06-01

    We report the development of an automated genetic analyzer for human sample testing based on microfluidic rapid polymerase chain reaction (PCR) with high-resolution melting analysis (HRMA). The integrated DNA microfluidic cartridge was used on a platform designed with a robotic pipettor system that works by sequentially picking up different test solutions from a 384-well plate, mixing them in the tips, and delivering mixed fluids to the DNA cartridge. A novel image feedback flow control system based on a Canon 5D Mark II digital camera was developed for controlling fluid movement through a complex microfluidic branching network without the use of valves. The same camera was used for measuring the high-resolution melt curve of DNA amplicons that were generated in the microfluidic chip. Owing to fast heating and cooling as well as sensitive temperature measurement in the microfluidic channels, the time frame for PCR and HRMA was dramatically reduced from hours to minutes. Preliminary testing results demonstrated that rapid serial PCR and HRMA are possible while still achieving high data quality that is suitable for human sample testing. © 2015 Society for Laboratory Automation and Screening.

  6. Innovative electron-beam welding of high-melting metals

    International Nuclear Information System (INIS)

    Behr, W.; Reisgen, U.

    2007-01-01

    Since its establishment as nuclear research plant Juelich in the year 1956, the research centre Juelich (FZJ) is concerned with the material processing of special metals. Among those are, above all, the high-melting refractory metals niobium, molybdenum and tungsten. Electron beam welding has always been considered to be an innovative special welding method; in the FZJ, electron beam welding has, moreover, always been adapted to the increasing demands made by research partners and involved manufacturing and design sectors. From the manual equipment technology right up to highly modern multi-beam technique, the technically feasible for fundamental research has, this way, always been realised. (Abstract Copyright [2007], Wiley Periodicals, Inc.) [de

  7. High resolution optical DNA mapping

    Science.gov (United States)

    Baday, Murat

    Many types of diseases including cancer and autism are associated with copy-number variations in the genome. Most of these variations could not be identified with existing sequencing and optical DNA mapping methods. We have developed Multi-color Super-resolution technique, with potential for high throughput and low cost, which can allow us to recognize more of these variations. Our technique has made 10--fold improvement in the resolution of optical DNA mapping. Using a 180 kb BAC clone as a model system, we resolved dense patterns from 108 fluorescent labels of two different colors representing two different sequence-motifs. Overall, a detailed DNA map with 100 bp resolution was achieved, which has the potential to reveal detailed information about genetic variance and to facilitate medical diagnosis of genetic disease.

  8. High-Melt Carbon-Carbon Coating for Nozzle Extensions

    Science.gov (United States)

    Thompson, James

    2015-01-01

    Carbon-Carbon Advanced Technologies, Inc. (C-CAT), has developed a high-melt coating for use in nozzle extensions in next-generation spacecraft. The coating is composed primarily of carbon-carbon, a carbon-fiber and carbon-matrix composite material that has gained a spaceworthy reputation due to its ability to withstand ultrahigh temperatures. C-CAT's high-melt coating embeds hafnium carbide (HfC) and zirconium diboride (ZrB2) within the outer layers of a carbon-carbon structure. The coating demonstrated enhanced high-temperature durability and suffered no erosion during a test in NASA's Arc Jet Complex. (Test parameters: stagnation heat flux=198 BTD/sq ft-sec; pressure=.265 atm; temperature=3,100 F; four cycles totaling 28 minutes) In Phase I of the project, C-CAT successfully demonstrated large-scale manufacturability with a 40-inch cylinder representing the end of a nozzle extension and a 16-inch flanged cylinder representing the attach flange of a nozzle extension. These demonstrators were manufactured without spalling or delaminations. In Phase II, C-CAT worked with engine designers to develop a nozzle extension stub skirt interfaced with an Aerojet Rocketdyne RL10 engine. All objectives for Phase II were successfully met. Additional nonengine applications for the coating include thermal protection systems (TPS) for next-generation spacecraft and hypersonic aircraft.

  9. A slow atomic diffusion process in high-entropy glass-forming metallic melts

    Science.gov (United States)

    Chen, Changjiu; Wong, Kaikin; Krishnan, Rithin P.; Embs, Jan P.; Chathoth, Suresh M.

    2018-04-01

    Quasi-elastic neutron scattering has been used to study atomic relaxation processes in high-entropy glass-forming metallic melts with different glass-forming ability (GFA). The momentum transfer dependence of mean relaxation time shows a highly collective atomic transport process in the alloy melts with the highest and lowest GFA. However, a jump diffusion process is the long-range atomic transport process in the intermediate GFA alloy melt. Nevertheless, atomic mobility close to the melting temperature of these alloy melts is quite similar, and the temperature dependence of the diffusion coefficient exhibits a non-Arrhenius behavior. The atomic mobility in these high-entropy melts is much slower than that of the best glass-forming melts at their respective melting temperatures.

  10. Rapid Detection and Differentiation of Clonorchis sinensis and Opisthorchis viverrini Using Real-Time PCR and High Resolution Melting Analysis

    OpenAIRE

    Cai, Xian-Quan; Yu, Hai-Qiong; Li, Rong; Yue, Qiao-Yun; Liu, Guo-Hua; Bai, Jian-Shan; Deng, Yan; Qiu, De-Yi; Zhu, Xing-Quan

    2014-01-01

    Clonorchis sinensis and Opisthorchis viverrini are both important fish-borne pathogens, causing serious public health problem in Asia. The present study developed an assay integrating real-time PCR and high resolution melting (HRM) analysis for the specific detection and rapid identification of C. sinensis and O. viverrini. Primers targeting COX1 gene were highly specific for these liver flukes, as evidenced by the negative amplification of closely related trematodes. Assays using genomic DNA...

  11. Low melting high lithia glass compositions and methods

    Science.gov (United States)

    Jantzen, Carol M.; Pickett, John B.; Cicero-Herman, Connie A.; Marra, James C.

    2000-01-01

    The invention relates to methods of vitrifying waste and for lowering the melting point of glass forming systems by including lithia formers in the glass forming composition in significant amounts, typically from about 0.16 wt % to about 11 wt %, based on the total glass forming oxides. The lithia is typically included as a replacement for alkali oxide glass formers that would normally be present in a particular glass forming system. Replacement can occur on a mole percent or weight percent basis, and typically results in a composition wherein lithia forms about 10 wt % to about 100 wt % of the alkali oxide glass formers present in the composition. The present invention also relates to the high lithia glass compositions formed by these methods. The invention is useful for stabilization of numerous types of waste materials, including aqueous waste streams, sludge solids, mixtures of aqueous supernate and sludge solids, combinations of spent filter aids from waste water treatment and waste sludges, supernate alone, incinerator ash, incinerator offgas blowdown, or combinations thereof, geological mine tailings and sludges, asbestos, inorganic filter media, cement waste forms in need of remediation, spent or partially spent ion exchange resins or zeolites, contaminated soils, lead paint, etc. The decrease in melting point achieved by the present invention desirably prevents volatilization of hazardous or radioactive species during vitrification.

  12. Rapid detection and identification of four major Schistosoma species by high-resolution melt (HRM) analysis.

    Science.gov (United States)

    Li, Juan; Zhao, Guang-Hui; Lin, RuiQing; Blair, David; Sugiyama, Hiromu; Zhu, Xing-Quan

    2015-11-01

    Schistosomiasis, caused by blood flukes belonging to several species of the genus Schistosoma, is a serious and widespread parasitic disease. Accurate and rapid differentiation of these etiological agents of animal and human schistosomiasis to species level can be difficult. We report a real-time PCR assay coupled with a high-resolution melt (HRM) assay targeting a portion of the nuclear 18S rDNA to detect, identify, and distinguish between four major blood fluke species (Schistosoma japonicum, Schistosoma mansoni, Schistosoma haematobium, and Schistosoma mekongi). Using this system, the Schistosoma spp. was accurately identified and could also be distinguished from all other trematode species with which they were compared. As little as 10(-5) ng genomic DNA from a Schistosoma sp. could be detected. This process is inexpensive, easy, and can be completed within 3 h. Examination of 21 representative Schistosoma samples from 15 geographical localities in seven endemic countries validated the value of the HRM detection assay and proved its reliability. The melting curves were characterized by peaks of 83.65 °C for S. japonicum and S. mekongi, 85.65 °C for S. mansoni, and 85.85 °C for S. haematobium. The present study developed a real-time PCR coupled with HRM analysis assay for detection and differential identification of S. mansoni, S. haematobium, S. japonicum, and S. mekongi. This method is rapid, sensitive, and inexpensive. It has important implications for epidemiological studies of Schistosoma.

  13. Massively parallel digital high resolution melt for rapid and absolutely quantitative sequence profiling

    Science.gov (United States)

    Velez, Daniel Ortiz; Mack, Hannah; Jupe, Julietta; Hawker, Sinead; Kulkarni, Ninad; Hedayatnia, Behnam; Zhang, Yang; Lawrence, Shelley; Fraley, Stephanie I.

    2017-02-01

    In clinical diagnostics and pathogen detection, profiling of complex samples for low-level genotypes represents a significant challenge. Advances in speed, sensitivity, and extent of multiplexing of molecular pathogen detection assays are needed to improve patient care. We report the development of an integrated platform enabling the identification of bacterial pathogen DNA sequences in complex samples in less than four hours. The system incorporates a microfluidic chip and instrumentation to accomplish universal PCR amplification, High Resolution Melting (HRM), and machine learning within 20,000 picoliter scale reactions, simultaneously. Clinically relevant concentrations of bacterial DNA molecules are separated by digitization across 20,000 reactions and amplified with universal primers targeting the bacterial 16S gene. Amplification is followed by HRM sequence fingerprinting in all reactions, simultaneously. The resulting bacteria-specific melt curves are identified by Support Vector Machine learning, and individual pathogen loads are quantified. The platform reduces reaction volumes by 99.995% and achieves a greater than 200-fold increase in dynamic range of detection compared to traditional PCR HRM approaches. Type I and II error rates are reduced by 99% and 100% respectively, compared to intercalating dye-based digital PCR (dPCR) methods. This technology could impact a number of quantitative profiling applications, especially infectious disease diagnostics.

  14. High Resolution Melting (HRM) for High-Throughput Genotyping—Limitations and Caveats in Practical Case Studies

    Science.gov (United States)

    Słomka, Marcin; Sobalska-Kwapis, Marta; Wachulec, Monika; Bartosz, Grzegorz

    2017-01-01

    High resolution melting (HRM) is a convenient method for gene scanning as well as genotyping of individual and multiple single nucleotide polymorphisms (SNPs). This rapid, simple, closed-tube, homogenous, and cost-efficient approach has the capacity for high specificity and sensitivity, while allowing easy transition to high-throughput scale. In this paper, we provide examples from our laboratory practice of some problematic issues which can affect the performance and data analysis of HRM results, especially with regard to reference curve-based targeted genotyping. We present those examples in order of the typical experimental workflow, and discuss the crucial significance of the respective experimental errors and limitations for the quality and analysis of results. The experimental details which have a decisive impact on correct execution of a HRM genotyping experiment include type and quality of DNA source material, reproducibility of isolation method and template DNA preparation, primer and amplicon design, automation-derived preparation and pipetting inconsistencies, as well as physical limitations in melting curve distinction for alternative variants and careful selection of samples for validation by sequencing. We provide a case-by-case analysis and discussion of actual problems we encountered and solutions that should be taken into account by researchers newly attempting HRM genotyping, especially in a high-throughput setup. PMID:29099791

  15. High Resolution Melting (HRM) for High-Throughput Genotyping-Limitations and Caveats in Practical Case Studies.

    Science.gov (United States)

    Słomka, Marcin; Sobalska-Kwapis, Marta; Wachulec, Monika; Bartosz, Grzegorz; Strapagiel, Dominik

    2017-11-03

    High resolution melting (HRM) is a convenient method for gene scanning as well as genotyping of individual and multiple single nucleotide polymorphisms (SNPs). This rapid, simple, closed-tube, homogenous, and cost-efficient approach has the capacity for high specificity and sensitivity, while allowing easy transition to high-throughput scale. In this paper, we provide examples from our laboratory practice of some problematic issues which can affect the performance and data analysis of HRM results, especially with regard to reference curve-based targeted genotyping. We present those examples in order of the typical experimental workflow, and discuss the crucial significance of the respective experimental errors and limitations for the quality and analysis of results. The experimental details which have a decisive impact on correct execution of a HRM genotyping experiment include type and quality of DNA source material, reproducibility of isolation method and template DNA preparation, primer and amplicon design, automation-derived preparation and pipetting inconsistencies, as well as physical limitations in melting curve distinction for alternative variants and careful selection of samples for validation by sequencing. We provide a case-by-case analysis and discussion of actual problems we encountered and solutions that should be taken into account by researchers newly attempting HRM genotyping, especially in a high-throughput setup.

  16. High Resolution Melting (HRM for High-Throughput Genotyping—Limitations and Caveats in Practical Case Studies

    Directory of Open Access Journals (Sweden)

    Marcin Słomka

    2017-11-01

    Full Text Available High resolution melting (HRM is a convenient method for gene scanning as well as genotyping of individual and multiple single nucleotide polymorphisms (SNPs. This rapid, simple, closed-tube, homogenous, and cost-efficient approach has the capacity for high specificity and sensitivity, while allowing easy transition to high-throughput scale. In this paper, we provide examples from our laboratory practice of some problematic issues which can affect the performance and data analysis of HRM results, especially with regard to reference curve-based targeted genotyping. We present those examples in order of the typical experimental workflow, and discuss the crucial significance of the respective experimental errors and limitations for the quality and analysis of results. The experimental details which have a decisive impact on correct execution of a HRM genotyping experiment include type and quality of DNA source material, reproducibility of isolation method and template DNA preparation, primer and amplicon design, automation-derived preparation and pipetting inconsistencies, as well as physical limitations in melting curve distinction for alternative variants and careful selection of samples for validation by sequencing. We provide a case-by-case analysis and discussion of actual problems we encountered and solutions that should be taken into account by researchers newly attempting HRM genotyping, especially in a high-throughput setup.

  17. Vacancies und melting curves of metals at high pressure

    International Nuclear Information System (INIS)

    Gorecki, T.

    1977-01-01

    The vacancy mechanism of the melting process is utilized as a starting point in derivation of the pressure dependence of melting temperature for metals. The results obtained for the initial slope of the melting curve are compared with experimental data for 45 metals (including U, Np, Pu, rare earths) and in most cases the agreement is very good. An on-linearity of the fusion curve and appearence of the maximum on the melting curve at a pressure approximately equal to the bulk modulus is also predicted with qualitative agreement with existing experimental data. (orig./GSC) [de

  18. Rapid detection of Echinococcus species by a high-resolution melting (HRM) approach.

    Science.gov (United States)

    Santos, Guilherme Brzoskowski; Espínola, Sergio Martín; Ferreira, Henrique Bunselmeyer; Margis, Rogerio; Zaha, Arnaldo

    2013-11-14

    High-resolution melting (HRM) provides a low-cost, fast and sensitive scanning method that allows the detection of DNA sequence variations in a single step, which makes it appropriate for application in parasite identification and genotyping. The aim of this work was to implement an HRM-PCR assay targeting part of the mitochondrial cox1 gene to achieve an accurate and fast method for Echinococcus spp. differentiation. For melting analysis, a total of 107 samples from seven species were used in this study. The species analyzed included Echinococcus granulosus (n = 41) and Echinococcus ortleppi (n = 50) from bovine, Echinococcus vogeli (n = 2) from paca, Echinococcus oligarthra (n = 3) from agouti, Echinococcus multilocularis (n = 6) from monkey and Echinococcus canadensis (n = 2) and Taenia hydatigena (n = 3) from pig. DNA extraction was performed, and a 444-bp fragment of the cox1 gene was amplified. Two approaches were used, one based on HRM analysis, and a second using SYBR Green Tm-based. In the HRM analysis, a specific profile for each species was observed. Although some species exhibited almost the same melting temperature (Tm) value, the HRM profiles could be clearly discriminated. The SYBR Green Tm-based analysis showed differences between E. granulosus and E. ortleppi and between E. vogeli and E. oligarthra. In this work, we report the implementation of HRM analysis to differentiate species of the genus Echinococcus using part of the mitochondrial gene cox1. This method may be also potentially applied to identify other species belonging to the Taeniidae family.

  19. High-resolution melt PCR analysis for genotyping of Ureaplasma parvum isolates directly from clinical samples.

    Science.gov (United States)

    Payne, Matthew S; Tabone, Tania; Kemp, Matthew W; Keelan, Jeffrey A; Spiller, O Brad; Newnham, John P

    2014-02-01

    Ureaplasma sp. infection in neonates and adults underlies a variety of disease pathologies. Of the two human Ureaplasma spp., Ureaplasma parvum is clinically the most common. We have developed a high-resolution melt (HRM) PCR assay for the differentiation of the four serovars of U. parvum in a single step. Currently U. parvum strains are separated into four serovars by sequencing the promoter and coding region of the multiple-banded antigen (MBA) gene. We designed primers to conserved sequences within this region for PCR amplification and HRM analysis to generate reproducible and distinct melt profiles that distinguish clonal representatives of serovars 1, 3, 6, and 14. Furthermore, our HRM PCR assay could classify DNA extracted from 74 known (MBA-sequenced) test strains with 100% accuracy. Importantly, HRM PCR was also able to identify U. parvum serovars directly from 16 clinical swabs. HRM PCR performed with DNA consisting of mixtures of combined known serovars yielded profiles that were easily distinguished from those for single-serovar controls. These profiles mirrored clinical samples that contained mixed serovars. Unfortunately, melt curve analysis software is not yet robust enough to identify the composition of mixed serovar samples, only that more than one serovar is present. HRM PCR provides a single-step, rapid, cost-effective means to differentiate the four serovars of U. parvum that did not amplify any of the known 10 serovars of Ureaplasma urealyticum tested in parallel. Choice of reaction reagents was found to be crucial to allow sufficient sensitivity to differentiate U. parvum serovars directly from clinical swabs rather than requiring cell enrichment using microbial culture techniques.

  20. Optimization of PCR Condition: The First Study of High Resolution Melting Technique for Screening of APOA1 Variance.

    Science.gov (United States)

    Wahyuningsih, Hesty; K Cayami, Ferdy; Bahrudin, Udin; A Sobirin, Mochamad; Ep Mundhofir, Farmaditya; Mh Faradz, Sultana; Hisatome, Ichiro

    2017-03-01

    High resolution melting (HRM) is a post-PCR technique for variant screening and genotyping based on the different melting points of DNA fragments. The advantages of this technique are that it is fast, simple, and efficient and has a high output, particularly for screening of a large number of samples. APOA1 encodes apolipoprotein A1 (apoA1) which is a major component of high density lipoprotein cholesterol (HDL-C). This study aimed to obtain an optimal quantitative polymerase chain reaction (qPCR)-HRM condition for screening of APOA1 variance. Genomic DNA was isolated from a peripheral blood sample using the salting out method. APOA1 was amplified using the RotorGeneQ 5Plex HRM. The PCR product was visualized with the HRM amplification curve and confirmed using gel electrophoresis. The melting profile was confirmed by looking at the melting curve. Five sets of primers covering the translated region of APOA1 exons were designed with expected PCR product size of 100-400 bps. The amplified segments of DNA were amplicons 2, 3, 4A, 4B, and 4C. Amplicons 2, 3 and 4B were optimized at an annealing temperature of 60 °C at 40 PCR cycles. Amplicon 4A was optimized at an annealing temperature of 62 °C at 45 PCR cycles. Amplicon 4C was optimized at an annealing temperature of 63 °C at 50 PCR cycles. In addition to the suitable procedures of DNA isolation and quantification, primer design and an estimated PCR product size, the data of this study showed that appropriate annealing temperature and PCR cycles were important factors in optimization of HRM technique for variant screening in APOA1 .

  1. Rapid Detection and Identification of Human Hookworm Infections through High Resolution Melting (HRM) Analysis

    Science.gov (United States)

    Ngui, Romano; Lim, Yvonne A. L.; Chua, Kek Heng

    2012-01-01

    Background Hookworm infections are still endemic in low and middle income tropical countries with greater impact on the socioeconomic and public health of the bottom billion of the world's poorest people. In this study, a real-time polymerase chain reaction (PCR) coupled with high resolution melting-curve (HRM) analysis was evaluated for an accurate, rapid and sensitive tool for species identification focusing on the five human hookworm species. Methods Real-time PCR coupled with HRM analysis targeting the second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA as the genetic marker was used to identify and distinguish hookworm species in human samples. Unique and distinct characteristics of HRM patterns were produced for each of the five hookworm species. The melting curves were characterized by peaks of 79.24±0.05°C and 83.00±0.04°C for Necator americanus, 79.12±0.10°C for Ancylostoma duodenale, 79.40±0.10°C for Ancylostoma ceylanicum, 79.63±0.05°C for Ancylostoma caninum and 79.70±0.14°C for Ancylostoma braziliense. An evaluation of the method's sensitivity and specificity revealed that this assay was able to detect as low as 0.01 ng/µl hookworm DNA and amplification was only recorded for hookworm positive samples. Conclusion The HRM assay developed in this study is a rapid and straightforward method for the diagnosis, identification and discrimination of five human hookworms. This assay is simple compared to other probe-based genotyping methods as it does not require multiplexing, DNA sequencing or post-PCR processing. Therefore, this method offers a new alternative for rapid detection of human hookworm species. PMID:22844538

  2. Rapid screening for targeted genetic variants via high-resolution melting curve analysis.

    Science.gov (United States)

    Chambliss, Allison B; Resnick, Molly; Petrides, Athena K; Clarke, William A; Marzinke, Mark A

    2017-03-01

    Current methods for the detection of single nucleotide polymorphisms (SNPs) associated with aberrant drug-metabolizing enzyme function are hindered by long turnaround times and specialized techniques and instrumentation. In this study, we describe the development and validation of a high-resolution melting (HRM) curve assay for the rapid screening of variant genotypes for targeted genetic polymorphisms in the cytochrome P450 enzymes CYP2C9, CYP2C19, and CYP3A5. Sequence-specific primers were custom-designed to flank nine SNPs within the genetic regions of aforementioned drug metabolizing enzymes. PCR amplification was performed followed by amplicon denaturation by precise temperature ramping in order to distinguish genotypes by melting temperature (Tm). A standardized software algorithm was used to assign amplicons as 'reference' or 'variant' as compared to duplicate reference sequence DNA controls for each SNP. Intra-assay (n=5) precision of Tms for all SNPs was ≤0.19%, while inter-assay (n=20) precision ranged from 0.04% to 0.21%. When compared to a reference method of Sanger sequencing, the HRM assay produced no false negative results, and overcall frequency ranged from 0% to 26%, depending on the SNP. Furthermore, HRM genotyping displayed accuracy over input DNA concentrations ranging from 10 to 200 ng/μL. The presented assay provides a rapid method for the screening for genetic variants in targeted CYP450 regions with a result of 'reference' or 'variant' available within 2 h from receipt of extracted DNA. The method can serve as a screening approach to rapidly identify individuals with variant sequences who should be further investigated by reflexed confirmatory testing for aberrant cytochrome P450 enzymatic activity. Rapid knowledge of variant status may aid in the avoidance of adverse clinical events by allowing for dosing of normal metabolizer patients immediately while identifying the need to wait for confirmatory testing in those patients who are

  3. Rapid detection and identification of human hookworm infections through high resolution melting (HRM analysis.

    Directory of Open Access Journals (Sweden)

    Romano Ngui

    Full Text Available BACKGROUND: Hookworm infections are still endemic in low and middle income tropical countries with greater impact on the socioeconomic and public health of the bottom billion of the world's poorest people. In this study, a real-time polymerase chain reaction (PCR coupled with high resolution melting-curve (HRM analysis was evaluated for an accurate, rapid and sensitive tool for species identification focusing on the five human hookworm species. METHODS: Real-time PCR coupled with HRM analysis targeting the second internal transcribed spacer (ITS-2 of nuclear ribosomal DNA as the genetic marker was used to identify and distinguish hookworm species in human samples. Unique and distinct characteristics of HRM patterns were produced for each of the five hookworm species. The melting curves were characterized by peaks of 79.24±0.05°C and 83.00±0.04°C for Necator americanus, 79.12±0.10°C for Ancylostoma duodenale, 79.40±0.10°C for Ancylostoma ceylanicum, 79.63±0.05°C for Ancylostoma caninum and 79.70±0.14°C for Ancylostoma braziliense. An evaluation of the method's sensitivity and specificity revealed that this assay was able to detect as low as 0.01 ng/µl hookworm DNA and amplification was only recorded for hookworm positive samples. CONCLUSION: The HRM assay developed in this study is a rapid and straightforward method for the diagnosis, identification and discrimination of five human hookworms. This assay is simple compared to other probe-based genotyping methods as it does not require multiplexing, DNA sequencing or post-PCR processing. Therefore, this method offers a new alternative for rapid detection of human hookworm species.

  4. Equation for the melting curve of solids under high pressure

    International Nuclear Information System (INIS)

    Boguslavskii, Yu.Ya.

    1982-01-01

    Simon's equation of the melting curve is obtained using the Clausius-Clapeyron equation in the linear approximation of the pressure dependence of the melting entropy and the volume change at the melting point. The constants in Simon's equation are calculated in this approximation for the alkali metals Li, Na, K, Rb, Cs and also for hydrogen, H 2 , and argon. It is shown that one can obtain the constants of Simon's equation in a pressure range which is wider than the region of the thermodynamical validity of Simon's equation by averaging the values of the constants determined in different points of the melting curves. The constants obtained by this manner agree well with the experimental data. (author)

  5. Use of a New High Resolution Melting Method for Genotyping Pathogenic Leptospira spp.

    Directory of Open Access Journals (Sweden)

    Florence Naze

    Full Text Available Leptospirosis is a worldwide zoonosis that is endemic in tropical areas, such as Reunion Island. The species Leptospira interrogans is the primary agent in human infections, but other pathogenic species, such as L. kirschner and L. borgpetersenii, are also associated with human leptospirosis.In this study, a melting curve analysis of the products that were amplified with the primer pairs lfb1 F/R and G1/G2 facilitated an accurate species classification of Leptospira reference strains. Next, we combined an unsupervised high resolution melting (HRM method with a new statistical approach using primers to amplify a two variable-number tandem-repeat (VNTR for typing at the subspecies level. The HRM analysis, which was performed with ScreenClust Software, enabled the identification of genotypes at the serovar level with high resolution power (Hunter-Gaston index 0.984. This method was also applied to Leptospira DNA from blood samples that were obtained from Reunion Island after 1998. We were able to identify a unique genotype that is identical to that of the L. interrogans serovars Copenhageni and Icterohaemorrhagiae, suggesting that this genotype is the major cause of leptospirosis on Reunion Island.Our simple, rapid, and robust genotyping method enables the identification of Leptospira strains at the species and subspecies levels and supports the direct genotyping of Leptospira in biological samples without requiring cultures.

  6. Applying high-resolution melting (HRM) technology to olive oil and wine authenticity.

    Science.gov (United States)

    Pereira, Leonor; Gomes, Sónia; Barrias, Sara; Fernandes, José Ramiro; Martins-Lopes, Paula

    2018-01-01

    Olive oil and wine production have a worldwide economic impact. Their market reliability is under great concern because of the increasing number of fraud and adulteration attempts. The need for a traceability system in all its extension is crucial particularly for the cases of olive oils and wines with certified labels, in which only a limited number of olives and grapevine varieties, respectively, are allowed in a restricted well-defined geographical area. Molecular markers have been vastly applied to the food sector, and in particular High-Resolution DNA Melting technology has been successfully applied for olive oil and wine authentication, as part of the traceability system. In this review, the applications of HRM and their usefulness for this sector considering, Safety, Security and Authenticity will be reviewed. A broad overview of the HRM technique will be presented, focusing on the aspects that are crucial for its success, in particular the new generation of fluorescent dsDNA dyes used for amplicon detection and quantification, and the data analysis. A brief outlook on the olive oil and wine authenticity procedures, based on new DNA technology advances, and in which way this may influence the future establishment of a traceability system will be discussed. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Comparison of multiple DNA dyes for real-time PCR: effects of dye concentration and sequence composition on DNA amplification and melting temperature

    DEFF Research Database (Denmark)

    Guðnason, Haukur; Dufva, Hans Martin; Bang, Dang Duong

    2007-01-01

    investigate 15 different intercalating DNA dyes for their inhibitory effects on PCR, effects on DNA melting temperature and possible preferential binding to GC-rich sequences. Our results demonstrated that in contrast to the results of SYBR Green I, two intercalating dyes SYTO-13 and SYTO-82 do not inhibit......The importance of real-time polymerase chain reaction (PCR) has increased steadily in clinical applications over the last decade. Many applications utilize SYBR Green I dye to follow the accumulation of amplicons in real time. SYBR Green I has, however, a number of limitations that include...... the inhibition of PCR, preferential binding to GC-rich sequences and effects on melting curve analysis. Although a few alternative dyes without some of these limitations have been recently proposed, no large-scale investigation into the properties of intercalating dyes has been performed. In this study, we...

  8. Evaluation of PCR and high-resolution melt curve analysis for differentiation of Salmonella isolates.

    Science.gov (United States)

    Saeidabadi, Mohammad Sadegh; Nili, Hassan; Dadras, Habibollah; Sharifiyazdi, Hassan; Connolly, Joanne; Valcanis, Mary; Raidal, Shane; Ghorashi, Seyed Ali

    2017-06-01

    Consumption of poultry products contaminated with Salmonella is one of the major causes of foodborne diseases worldwide and therefore detection and differentiation of Salmonella spp. in poultry is important. In this study, oligonucleotide primers were designed from hemD gene and a PCR followed by high-resolution melt (HRM) curve analysis was developed for rapid differentiation of Salmonella isolates. Amplicons of 228 bp were generated from 16 different Salmonella reference strains and from 65 clinical field isolates mainly from poultry farms. HRM curve analysis of the amplicons differentiated Salmonella isolates and analysis of the nucleotide sequence of the amplicons from selected isolates revealed that each melting curve profile was related to a unique DNA sequence. The relationship between reference strains and tested specimens was also evaluated using a mathematical model without visual interpretation of HRM curves. In addition, the potential of the PCR-HRM curve analysis was evaluated for genotyping of additional Salmonella isolates from different avian species. The findings indicate that PCR followed by HRM curve analysis provides a rapid and robust technique for genotyping of Salmonella isolates to determine the serovar/serotype.

  9. Rapid detection of SMARCB1 sequence variation using high resolution melting

    Directory of Open Access Journals (Sweden)

    Ashley David M

    2009-12-01

    Full Text Available Abstract Background Rhabdoid tumors are rare cancers of early childhood arising in the kidney, central nervous system and other organs. The majority are caused by somatic inactivating mutations or deletions affecting the tumor suppressor locus SMARCB1 [OMIM 601607]. Germ-line SMARCB1 inactivation has been reported in association with rhabdoid tumor, epitheloid sarcoma and familial schwannomatosis, underscoring the importance of accurate mutation screening to ascertain recurrence and transmission risks. We describe a rapid and sensitive diagnostic screening method, using high resolution melting (HRM, for detecting sequence variations in SMARCB1. Methods Amplicons, encompassing the nine coding exons of SMARCB1, flanking splice site sequences and the 5' and 3' UTR, were screened by both HRM and direct DNA sequencing to establish the reliability of HRM as a primary mutation screening tool. Reaction conditions were optimized with commercially available HRM mixes. Results The false negative rate for detecting sequence variants by HRM in our sample series was zero. Nine amplicons out of a total of 140 (6.4% showed variant melt profiles that were subsequently shown to be false positive. Overall nine distinct pathogenic SMARCB1 mutations were identified in a total of 19 possible rhabdoid tumors. Two tumors had two distinct mutations and two harbored SMARCB1 deletion. Other mutations were nonsense or frame-shifts. The detection sensitivity of the HRM screening method was influenced by both sequence context and specific nucleotide change and varied from 1: 4 to 1:1000 (variant to wild-type DNA. A novel method involving digital HRM, followed by re-sequencing, was used to confirm mutations in tumor specimens containing associated normal tissue. Conclusions This is the first report describing SMARCB1 mutation screening using HRM. HRM is a rapid, sensitive and inexpensive screening technology that is likely to be widely adopted in diagnostic laboratories to

  10. Rapid detection of SMARCB1 sequence variation using high resolution melting

    International Nuclear Information System (INIS)

    Dagar, Vinod; Chow, Chung-Wo; Ashley, David M; Algar, Elizabeth M

    2009-01-01

    Rhabdoid tumors are rare cancers of early childhood arising in the kidney, central nervous system and other organs. The majority are caused by somatic inactivating mutations or deletions affecting the tumor suppressor locus SMARCB1 [OMIM 601607]. Germ-line SMARCB1 inactivation has been reported in association with rhabdoid tumor, epitheloid sarcoma and familial schwannomatosis, underscoring the importance of accurate mutation screening to ascertain recurrence and transmission risks. We describe a rapid and sensitive diagnostic screening method, using high resolution melting (HRM), for detecting sequence variations in SMARCB1. Amplicons, encompassing the nine coding exons of SMARCB1, flanking splice site sequences and the 5' and 3' UTR, were screened by both HRM and direct DNA sequencing to establish the reliability of HRM as a primary mutation screening tool. Reaction conditions were optimized with commercially available HRM mixes. The false negative rate for detecting sequence variants by HRM in our sample series was zero. Nine amplicons out of a total of 140 (6.4%) showed variant melt profiles that were subsequently shown to be false positive. Overall nine distinct pathogenic SMARCB1 mutations were identified in a total of 19 possible rhabdoid tumors. Two tumors had two distinct mutations and two harbored SMARCB1 deletion. Other mutations were nonsense or frame-shifts. The detection sensitivity of the HRM screening method was influenced by both sequence context and specific nucleotide change and varied from 1: 4 to 1:1000 (variant to wild-type DNA). A novel method involving digital HRM, followed by re-sequencing, was used to confirm mutations in tumor specimens containing associated normal tissue. This is the first report describing SMARCB1 mutation screening using HRM. HRM is a rapid, sensitive and inexpensive screening technology that is likely to be widely adopted in diagnostic laboratories to facilitate whole gene mutation screening

  11. Investigation of melt structure and crystallization processes by high-temperature Raman spectroscopy method

    International Nuclear Information System (INIS)

    Voron'ko, Yu.K.; Kudryavtsev, A.B.; Osiko, V.V.; Sobol', A.A.

    1988-01-01

    A review of studies dealing with the melts of alkali, rare earth and other element phosphates, gallates, germanates, niobates and tungstates, which are carried out by the method of high-temperature Raman spectroscopy, is given. The effect of the melt structure on the mechanism of the substance cystallization is considered. It is shown that vitrification and supercooling of the melt, as well as its crystallization in the from of metastable structures, are related to the effect of nonconformity between the melt and crystal strucure. The effect of nonconformity between anion motives in the melt and crystal creates obstacles for equilibrium structure nucleation, which results in the formation mainly of metastable forms with lattice structure for from the structure of the melt, though cases of equilibrium phase crystallization are also possible. 37 refs.; 13 figs.; 2 tabs

  12. High-resolution melting analysis, a simple and effective method for reliable mutation scanning and frequency studies in the ACADVL gene

    DEFF Research Database (Denmark)

    Olsen, Rikke Katrine Jentoft; Dobrowolski, Steven F; Kjeldsen, Margrethe

    2010-01-01

    -long-chain acyl-CoA dehydrogenase deficiency (VLCADD), the second most common fatty acid oxidation disorder detected by expanded newborn screening, to demonstrate accurate and fast diagnostic evaluation of the ACADVL gene utilizing DNA extracted from the newborn screening dried blood spot and high resolution melt...

  13. Controlled localised melting in silicon by high dose germanium implantation and flash lamp annealing

    International Nuclear Information System (INIS)

    Voelskow, Matthias; Skorupa, Wolfgang; Pezoldt, Joerg; Kups, Thomas

    2009-01-01

    High intensity light pulse irradiation of monocrystalline silicon wafers is usually accompanied by inhomogeneous surface melting. The aim of the present work is to induce homogeneous buried melting in silicon by germanium implantation and subsequent flash lamp annealing. For this purpose high dose, high energy germanium implantation has been employed to lower the melting temperature of silicon in a predetermined depth region. Subsequent flash lamp irradiation at high energy densities leads to local melting of the germanium rich buried layer, whereby the thickness of the molten layer depends on the irradiation energy density. During the cooling down epitaxial crystallization takes place resulting in a largely defect-free layer. The combination of buried melting and dopant segregation has the potential to produce unusually buried doping profiles or to create strained silicon structures.

  14. Genome-wide SNP discovery in tetraploid alfalfa using 454 sequencing and high resolution melting analysis

    Directory of Open Access Journals (Sweden)

    Zhao Patrick X

    2011-07-01

    Full Text Available Abstract Background Single nucleotide polymorphisms (SNPs are the most common type of sequence variation among plants and are often functionally important. We describe the use of 454 technology and high resolution melting analysis (HRM for high throughput SNP discovery in tetraploid alfalfa (Medicago sativa L., a species with high economic value but limited genomic resources. Results The alfalfa genotypes selected from M. sativa subsp. sativa var. 'Chilean' and M. sativa subsp. falcata var. 'Wisfal', which differ in water stress sensitivity, were used to prepare cDNA from tissue of clonally-propagated plants grown under either well-watered or water-stressed conditions, and then pooled for 454 sequencing. Based on 125.2 Mb of raw sequence, a total of 54,216 unique sequences were obtained including 24,144 tentative consensus (TCs sequences and 30,072 singletons, ranging from 100 bp to 6,662 bp in length, with an average length of 541 bp. We identified 40,661 candidate SNPs distributed throughout the genome. A sample of candidate SNPs were evaluated and validated using high resolution melting (HRM analysis. A total of 3,491 TCs harboring 20,270 candidate SNPs were located on the M. truncatula (MT 3.5.1 chromosomes. Gene Ontology assignments indicate that sequences obtained cover a broad range of GO categories. Conclusions We describe an efficient method to identify thousands of SNPs distributed throughout the alfalfa genome covering a broad range of GO categories. Validated SNPs represent valuable molecular marker resources that can be used to enhance marker density in linkage maps, identify potential factors involved in heterosis and genetic variation, and as tools for association mapping and genomic selection in alfalfa.

  15. High-temperature oxidation of tungsten covered by layer of glass-enamel melt

    International Nuclear Information System (INIS)

    Vasnetsova, V.B.; Shardakov, N.T.; Kudyakov, V.Ya.; Deryabin, V.A.

    1997-01-01

    Corrosion losses of tungsten covered by the layer of glass-enamel melt were determined at 800, 850, 900, 950 deg C. It is shown that the rate of high-temperature oxidation of tungsten decreases after application of glass-enamel melt on its surface. This is probably conditioned by reduction of area of metal interaction with oxidizing atmosphere

  16. Microstructure and properties of high chrome steel roller after laser surface melting

    Energy Technology Data Exchange (ETDEWEB)

    Li Meiyan, E-mail: lmy_102411@163.com [College of Electromechanical Engineering, China University of Petroleum, 271 Bei' er Road, Dongying 257061 (China); Wang Yong; Han Bin; Zhao Weimin; Han Tao [College of Electromechanical Engineering, China University of Petroleum, 271 Bei' er Road, Dongying 257061 (China)

    2009-06-15

    Laser surface melting of high chrome steels was achieved by a 5 kW continuous wave CO{sub 2} laser. The microstructure of the laser surface-melted steels was investigated by scanning electron microscopy, transmission electron microscopy and X-ray diffractometry, and the hardness profiles were determined by a Vickers hardness tester. The corrosion behavior in 3.5% NaCl solution was studied by electrochemical corrosion equipment. The large carbides of high chrome steels are completely dissolved and ultrafine dendrites of austenite with submicroscopic M{sub 23}C{sub 6} carbides precipitation are formed in the melted zone. The austenite in the melted zone has a high tempering stability. The corrosion resistance of the laser surface-melted steels is significantly improved due to the dissolution of carbides and the increase of the alloying elements in the solid solution as well as the large amount of austenite.

  17. Microstructure and properties of high chrome steel roller after laser surface melting

    International Nuclear Information System (INIS)

    Li Meiyan; Wang Yong; Han Bin; Zhao Weimin; Han Tao

    2009-01-01

    Laser surface melting of high chrome steels was achieved by a 5 kW continuous wave CO 2 laser. The microstructure of the laser surface-melted steels was investigated by scanning electron microscopy, transmission electron microscopy and X-ray diffractometry, and the hardness profiles were determined by a Vickers hardness tester. The corrosion behavior in 3.5% NaCl solution was studied by electrochemical corrosion equipment. The large carbides of high chrome steels are completely dissolved and ultrafine dendrites of austenite with submicroscopic M 23 C 6 carbides precipitation are formed in the melted zone. The austenite in the melted zone has a high tempering stability. The corrosion resistance of the laser surface-melted steels is significantly improved due to the dissolution of carbides and the increase of the alloying elements in the solid solution as well as the large amount of austenite.

  18. Identification of Brucella spp. isolated from human brucellosis in Malaysia using high-resolution melt (HRM) analysis.

    Science.gov (United States)

    Mohamed Zahidi, Jama'ayah; Bee Yong, Tay; Hashim, Rohaidah; Mohd Noor, Azura; Hamzah, Siti Hawa; Ahmad, Norazah

    2015-04-01

    Molecular approaches have been investigated to overcome difficulties in identification and differentiation of Brucella spp. using conventional phenotypic methods. In this study, high-resolution melt (HRM) analysis was used for rapid identification and differentiation of members of Brucella genus. A total of 41 Brucella spp. isolates from human brucellosis were subjected to HRM analysis using 4 sets of primers, which identified 40 isolates as Brucella melitensis and 1 as Brucella canis. The technique utilized low DNA concentration and was highly reproducible. The assay is shown to be a useful diagnostic tool, which can rapidly differentiate Brucella up to species level. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  19. Polymineralic inclusions in mantle chromitites from the Oman ophiolite indicate a highly magnesian parental melt

    Science.gov (United States)

    Rollinson, Hugh; Mameri, Lucan; Barry, Tiffany

    2018-06-01

    Polymineralic inclusions interpreted as melt inclusions in chromite from the dunitic Moho Transition Zone in the Maqsad area of the Oman ophiolite have been analysed and compositions integrated using a rastering technique on the scanning electron microscope. The inclusions now comprise a range of inter-grown hydrous phases including pargasite, aspidolite, phlogopite and chlorite, indicating that the parental melts were hydrous. Average inclusion compositions for seven samples contain between 23.1 and 26.8 wt% MgO and 1.7-3.6 wt% FeO. Compositions were corrected to allow for the low FeO concentrations using coexisting olivine compositions. These suggest that the primary melt has between 20 and 22 wt% MgO and 7-9.7 wt% FeO and has an affinity with boninitic melts, although the melts have a higher Ti content than most boninites. Average rare earth element concentrations suggest that the melts were derived from a REE depleted mantle source although fluid-mobile trace elements indicate a more enriched source. Given the hydrous nature of the inclusions this enrichment could be fluid driven. An estimate of the melt temperature can be made from the results of homogenisation experiments on these inclusions and suggests 1300 °C, which implies for a harzburgite solidus, relatively shallow melting at depths of <50 km and is consistent with a boninitic origin. The current "basaltic" nature of the chromite host to highly magnesian melt inclusions suggests that the dunitic Moho Transition Zone operated as a reaction filter in which magnesian melts were transformed into basalts by the removal of high magnesian olivines, particularly in areas where the Moho Transition Zone is unusually thick. We propose therefore that podiform mantle chromitites, even those with an apparent MORB-like chemical signature, have crystallised from a highly magnesian parental melt. The data presented here strongly support the view that this took place in a subduction initiation setting.

  20. Identification of Uvaria sp by barcoding coupled with high-resolution melting analysis (Bar-HRM).

    Science.gov (United States)

    Osathanunkul, M; Madesis, P; Ounjai, S; Pumiputavon, K; Somboonchai, R; Lithanatudom, P; Chaowasku, T; Wipasa, J; Suwannapoom, C

    2016-01-13

    DNA barcoding, which was developed about a decade ago, relies on short, standardized regions of the genome to identify plant and animal species. This method can be used to not only identify known species but also to discover novel ones. Numerous sequences are stored in online databases worldwide. One of the ways to save cost and time (by omitting the sequencing step) in species identification is to use available barcode data to design optimized primers for further analysis, such as high-resolution melting analysis (HRM). This study aimed to determine the effectiveness of the hybrid method Bar-HRM (DNA barcoding combined with HRM) to identify species that share similar external morphological features, rather than conduct traditional taxonomic identification that require major parts (leaf, flower, fruit) of the specimens. The specimens used for testing were those, which could not be identified at the species level and could either be Uvaria longipes or Uvaria wrayias, indicated by morphological identification. Primer pairs derived from chloroplast regions (matK, psbA-trnH, rbcL, and trnL) were used in the Bar-HRM. The results obtained from psbA-trnH primers were good enough to help in identifying the specimen while the rest were not. Bar-HRM analysis was proven to be a fast and cost-effective method for plant species identification.

  1. Volatile and light lithophile elements in high-anorthite plagioclase-hosted melt inclusions from Iceland

    Science.gov (United States)

    Neave, David A.; Hartley, Margaret E.; Maclennan, John; Edmonds, Marie; Thordarson, Thorvaldur

    2017-05-01

    Melt inclusions formed during the early stages of magmatic evolution trap primitive melt compositions and enable the volatile contents of primary melts and the mantle to be estimated. However, the syn- and post-entrapment behaviour of volatiles in primitive high-anorthite plagioclase-hosted melt inclusions from oceanic basalts remains poorly constrained. To address this deficit, we present volatile and light lithophile element analyses from a well-characterised suite of nine matrix glasses and 102 melt inclusions from the 10 ka Grímsvötn tephra series (i.e., Saksunarvatn ash) of Iceland's Eastern Volcanic Zone (EVZ). High matrix glass H2O and S contents indicate that eruption-related exsolution was arrested by quenching in a phreatomagmatic setting; Li, B, F and Cl did not exsolve during eruption. The almost uniformly low CO2 content of plagioclase-hosted melt inclusions cannot be explained by either shallow entrapment or the sequestration of CO2 into shrinkage bubbles, suggesting that inclusion CO2 contents were controlled by decrepitation instead. High H2O/Ce values in primitive plagioclase-hosted inclusions (182-823) generally exceed values expected for EVZ primary melts (∼ 180), and can be accounted for by diffusive H2O gain following the entrainment of primitive macrocrysts into evolved and H2O-rich melts a few days before eruption. A strong positive correlation between H2O and Li in plagioclase-hosted inclusions suggests that diffusive Li gain may also have occurred. Extreme F enrichments in primitive plagioclase-hosted inclusions (F/Nd = 51-216 versus ∼15 in matrix glasses) possibly reflect the entrapment of inclusions from high-Al/(Al + Si) melt pools formed by dissolution-crystallisation processes (as indicated by HFSE depletions in some inclusions), and into which F was concentrated by uphill diffusion since F is highly soluble in Al-rich melts. The high S/Dy of primitive inclusions (∼300) indicates that primary melts were S-rich in comparison

  2. Melting point of high-purity germanium stable isotopes

    Science.gov (United States)

    Gavva, V. A.; Bulanov, A. D.; Kut'in, A. M.; Plekhovich, A. D.; Churbanov, M. F.

    2018-05-01

    The melting point (Tm) of germanium stable isotopes 72Ge, 73Ge, 74Ge, 76Ge was determined by differential scanning calorimetry. With the increase in atomic mass of isotope the decrease in Tm is observed. The decrease was equal to 0.15 °C per the unit of atomic mass which qualitatively agrees with the value calculated by Lindemann formula accounting for the effect of "isotopic compression" of elementary cell.

  3. Species identification in meat products: A new screening method based on high resolution melting analysis of cyt b gene.

    Science.gov (United States)

    Lopez-Oceja, A; Nuñez, C; Baeta, M; Gamarra, D; de Pancorbo, M M

    2017-12-15

    Meat adulteration by substitution with lower value products and/or mislabeling involves economic, health, quality and socio-religious issues. Therefore, identification and traceability of meat species has become an important subject to detect possible fraudulent practices. In the present study the development of a high resolution melt (HRM) screening method for the identification of eight common meat species is reported. Samples from Bos taurus, Ovis aries, Sus scrofa domestica, Equus caballus, Oryctolagus cuniculus, Gallus gallus domesticus, Meleagris gallopavo and Coturnix coturnix were analyzed through the amplification of a 148 bp fragment from the cyt b gene with a universal primer pair in HRM analyses. Melting profiles from each species, as well as from several DNA mixtures of these species and blind samples, allowed a successful species differentiation. The results demonstrated that the HRM method here proposed is a fast, reliable, and low-cost screening technique. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Hybrid analysis (barcode-high resolution melting) for authentication of Thai herbal products, Andrographis paniculata (Burm.f.) Wall.ex Nees.

    Science.gov (United States)

    Osathanunkul, Maslin; Suwannapoom, Chatmongkon; Khamyong, Nuttaluck; Pintakum, Danupol; Lamphun, Santisuk Na; Triwitayakorn, Kanokporn; Osathanunkul, Kitisak; Madesis, Panagiotis

    2016-01-01

    Andrographis paniculata Nees is a medicinal plant with multiple pharmacological properties. It has been used over many centuries as a household remedy. A. paniculata products sold on the markets are in processed forms so it is difficult to authenticate. Therefore buying the herbal products poses a high-risk of acquiring counterfeited, substituted and/or adulterated products. Due to these issues, a reliable method to authenticate products is needed. High resolution melting analysis coupled with DNA barcoding (Bar-HRM) was applied to detect adulteration in commercial herbal products. The rbcL barcode was selected to use in primers design for HRM analysis to produce standard melting profile of A. paniculata species. DNA of the tested commercial products was isolated and their melting profiles were then generated and compared with the standard A. paniculata. The melting profiles of the rbcL amplicons of the three closely related herbal species (A. paniculata, Acanthus ebracteatus and Rhinacanthus nasutus) are clearly separated so that they can be distinguished by the developed method. The method was then used to authenticate commercial herbal products. HRM curves of all 10 samples tested are similar to A. paniculata which indicated that all tested products were contained the correct species as labeled. The method described in this study has been proved to be useful in aiding identification and/or authenticating A. paniculata. This Bar-HRM analysis has allowed us easily to determine the A. paniculata species in herbal products on the markets even they are in processed forms. We propose the use of DNA barcoding combined with High Resolution Melting analysis for authenticating of Andrographis paniculata products.The developed method can be used regardless of the type of the DNA template (fresh or dried tissue, leaf, and stem).rbcL region was chosen for the analysis and work well with our samplesWe can easily determine the A. paniculata species in herbal products tested

  5. Characterization of ash melting behaviour at high temperatures under conditions simulating combustible solid waste gasification.

    Science.gov (United States)

    Niu, Miaomiao; Dong, Qing; Huang, Yaji; Jin, Baosheng; Wang, Hongyan; Gu, Haiming

    2018-05-01

    To achieve high-temperature gasification-melting of combustible solid waste, ash melting behaviour under conditions simulating high-temperature gasification were studied. Raw ash (RA) and gasified ash (GA) were prepared respectively by waste ashing and fluidized bed gasification. Results of microstructure and composition of the two-ash indicated that GA showed a more porous structure and higher content of alkali and alkali earth metals among metallic elements. Higher temperature promoted GA melting and could reach a complete flowing state at about 1250°C. The order of melting rate of GA under different atmospheres was reducing condition > inert condition > oxidizing condition, which might be related to different existing forms of iron during melting and different flux content with atmosphere. Compared to RA, GA showed lower melting activity at the same condition due to the existence of an unconverted carbon and hollow structure. The melting temperature for sufficient melting and separation of GA should be at least 1250°C in this work.

  6. A study on structural analysis of highly corrosive melts at high temperature

    CERN Document Server

    Ohtori, N

    2002-01-01

    When sodium is burned at high temperature in the atmosphere, it reacts simultaneously with H sub 2 O in the atmosphere so that it can produce high temperature melt of sodium hydroxide as a solvent. If this melt includes peroxide ion (O sub 2 sup 2 sup -), it will be a considerably active and corrosive for iron so that several sodium iron double oxides will be produced as corrosion products after the reaction with steel structures. The present study was carried out in order to investigate the ability of presence of peroxide ion in sodium hydroxide solvent at high temperature and that of identification of the several corrosion products using laser Raman spectroscopy. The measurement system with ultraviolet laser was developed simultaneously in the present work to improve the ability of the measurement at high temperature. As results from the measurements, the possibility of the presence of peroxide ion was shown up to 823K in sodium peroxide and 823K in the melt of sodium hydroxide mixed with sodium peroxide. A...

  7. High-throughput gender identification of penguin species using melting curve analysis.

    Science.gov (United States)

    Tseng, Chao-Neng; Chang, Yung-Ting; Chiu, Hui-Tzu; Chou, Yii-Cheng; Huang, Hurng-Wern; Cheng, Chien-Chung; Liao, Ming-Hui; Chang, Hsueh-Wei

    2014-04-03

    Most species of penguins are sexual monomorphic and therefore it is difficult to visually identify their genders for monitoring population stability in terms of sex ratio analysis. In this study, we evaluated the suitability using melting curve analysis (MCA) for high-throughput gender identification of penguins. Preliminary test indicated that the Griffiths's P2/P8 primers were not suitable for MCA analysis. Based on sequence alignment of Chromo-Helicase-DNA binding protein (CHD)-W and CHD-Z genes from four species of penguins (Pygoscelis papua, Aptenodytes patagonicus, Spheniscus magellanicus, and Eudyptes chrysocome), we redesigned forward primers for the CHD-W/CHD-Z-common region (PGU-ZW2) and the CHD-W-specific region (PGU-W2) to be used in combination with the reverse Griffiths's P2 primer. When tested with P. papua samples, PCR using P2/PGU-ZW2 and P2/PGU-W2 primer sets generated two amplicons of 148- and 356-bp, respectively, which were easily resolved in 1.5% agarose gels. MCA analysis indicated the melting temperature (Tm) values for P2/PGU-ZW2 and P2/PGU-W2 amplicons of P. papua samples were 79.75°C-80.5°C and 81.0°C-81.5°C, respectively. Females displayed both ZW-common and W-specific Tm peaks, whereas male was positive only for ZW-common peak. Taken together, our redesigned primers coupled with MCA analysis allows precise high throughput gender identification for P. papua, and potentially for other penguin species such as A. patagonicus, S. magellanicus, and E. chrysocome as well.

  8. High resolution laser micro sintering / melting using q-switched and high brilliant laser radiation

    Science.gov (United States)

    Exner, H.; Streek, A.

    2015-03-01

    Since the discovery of selective laser sintering/melting, numerous modifications have been made to upgrade or customize this technology for industrial purposes. Laser micro sintering (LMS) is one of those modifications: Powders with particles in the range of a few micrometers are used to obtain products with highly resolved structures. Pulses of a q-switched laser had been considered necessary in order to generate sinter layers from the micrometer scaled metal powders. LMS has been applied with powders from metals as well as from ceramic and cermet feedstock's to generate micro parts. Recent technological progress and the application of high brilliant continuous laser radiation have now allowed an efficient laser sintering/melting of micrometer scaled metal powders. Thereby it is remarkable that thin sinter layers are generated using high continuous laser power. The principles of the process, the state of the art in LMS concerning its advantages and limitations and furthermore the latest results of the recent development of this technology will be presented. Laser Micro Sintering / Laser Micro Melting (LMM) offer a vision for a new dimension of additive fabrication of miniature and precise parts also with application potential in all engineering fields.

  9. Actinide, lanthanide and fission product speciation and electrochemistry in high and low temperature ionic melts

    Energy Technology Data Exchange (ETDEWEB)

    Bhatt, Anand I.; Kinoshita, Hajime; Koster, Anne L.; May, Iain; Sharrad, Clint A.; Volkovich, Vladimir A.; Fox, O. Danny; Jones, Chris J.; Lewin, Bob G.; Charnock, John M.; Hennig, Christoph

    2004-07-01

    There is currently a great deal of research interest in the development of molten salt technology, both classical high temperature melts and low temperature ionic liquids, for the electrochemical separation of the actinides from spent nuclear fuel. We are interested in gaining a better understanding of actinide and key fission product speciation and electrochemical properties in a range of melts. Our studies in high temperature alkali metal melts (including LiCl and LiCl-KCl and CsCl-NaCl eutectics) have focussed on in-situ species of U, Th, Tc and Ru using X-ray absorption spectroscopy (XAS, both EXAFS and XANES) and electronic absorption spectroscopy (EAS). We report unusual actinide speciation in high temperature melts and an evaluation of the likelihood of Ru or Tc volatilization during plant operation. Our studies in lower temperature melts (ionic liquids) have focussed on salts containing tertiary alkyl group 15 cations and the bis(tri-fluor-methyl)sulfonyl)imide anion, melts which we have shown to have exceptionally wide electrochemical windows. We report Ln, Th, U and Np speciation (XAS, EAS and vibrational spectroscopy) and electrochemistry in these melts and relate the solution studies to crystallographic characterised benchmark species. (authors)

  10. Actinide, lanthanide and fission product speciation and electrochemistry in high and low temperature ionic melts

    International Nuclear Information System (INIS)

    Bhatt, Anand I.; Kinoshita, Hajime; Koster, Anne L.; May, Iain; Sharrad, Clint A.; Volkovich, Vladimir A.; Fox, O. Danny; Jones, Chris J.; Lewin, Bob G.; Charnock, John M.; Hennig, Christoph

    2004-01-01

    There is currently a great deal of research interest in the development of molten salt technology, both classical high temperature melts and low temperature ionic liquids, for the electrochemical separation of the actinides from spent nuclear fuel. We are interested in gaining a better understanding of actinide and key fission product speciation and electrochemical properties in a range of melts. Our studies in high temperature alkali metal melts (including LiCl and LiCl-KCl and CsCl-NaCl eutectics) have focussed on in-situ species of U, Th, Tc and Ru using X-ray absorption spectroscopy (XAS, both EXAFS and XANES) and electronic absorption spectroscopy (EAS). We report unusual actinide speciation in high temperature melts and an evaluation of the likelihood of Ru or Tc volatilization during plant operation. Our studies in lower temperature melts (ionic liquids) have focussed on salts containing tertiary alkyl group 15 cations and the bis(tri-fluor-methyl)sulfonyl)imide anion, melts which we have shown to have exceptionally wide electrochemical windows. We report Ln, Th, U and Np speciation (XAS, EAS and vibrational spectroscopy) and electrochemistry in these melts and relate the solution studies to crystallographic characterised benchmark species. (authors)

  11. Identification of the GST-T1 and GST-M1 null genotypes using high resolution melting analysis.

    Science.gov (United States)

    Drobná, Zuzana; Del Razo, Luz Maria; Garcia-Vargas, Gonzalo; Sánchez-Ramírez, Blanca; González-Horta, Carmen; Ballinas-Casarrubias, Lourdes; Loomis, Dana; Stýblo, Miroslav

    2012-01-13

    Glutathione S-transferases, including GST-T1 and GST-M1, are known to be involved in the phase II detoxification pathways for xenobiotics as well as in the metabolism of endogenous compounds. Polymorphisms in these genes have been linked to an increased susceptibility to carcinogenesis and associated with risk factors that predispose to certain inflammatory diseases. In addition, GST-T1 and GST-M1 null genotypes have been shown to be responsible for interindividual variations in the metabolism of arsenic, a known human carcinogen. To assess the specific GST genotypes in the Mexican population chronically exposed to arsenic, we have developed a multiplex High Resolution Melting PCR (HRM-PCR) analysis using a LightCycler480 instrument. This method is based on analysis of the PCR product melting curve that discriminates PCR products according to their lengths and base sequences. Three pairs of primers that specifically recognize GST-T1, GST-M1, and β-globin, an internal control, to produce amplicons of different length were designed and combined with LightCycler480 High Resolution Melting Master Mix containing ResoLight, a completely saturating DNA dye. Data collected from melting curve analysis were evaluated using LightCycler480 software to determine specific melting temperatures of individual melting curves representing target genes. Using this newly developed multiplex HRM-PCR analysis, we evaluated GST-T1 and GST-M1 genotypes in 504 DNA samples isolated from the blood of individuals residing in Zimapan, Lagunera, and Chihuahua regions in Mexico. We found that the Zimapan and Lagunera populations have similar GST-T1 and GST-M1 genotype frequencies which differ from those of the Chihuahua population. In addition, 14 individuals have been identified as carriers of the double null genotype, i.e., null genotypes in both GST-T1 and GST-M1 genes. Although this procedure does not distinguish between biallelic (+/+) and monoallelic (+/-) genotypes, it can be used in an

  12. Thermodynamic study of rhodamine 123-calf thymus DNA interaction: determination of calorimetric enthalpy by optical melting study.

    Science.gov (United States)

    Masum, Abdulla Al; Chakraborty, Maharudra; Pandya, Prateek; Halder, Umesh Chandra; Islam, Md Maidul; Mukhopadhyay, Subrata

    2014-11-20

    In this paper, the interaction of rhodamine123 (R123) with calf thymus DNA has been studied using molecular modeling and other biophysical methods like UV-vis spectroscopy, fluoremetry, optical melting, isothermal titration calorimetry, and circular dichroic studies. Results showed that the binding energy is about -6 to -8 kcal/mol, and the binding process is favored by both negative enthalpy change and positive entropy change. A new method to determine different thermodynamic properties like calorimetric enthalpy and heat capacity change has been introduced in this paper. The obtained data has been crossed-checked by other methods. After dissecting the free-energy contribution, it was observed that the binding was favored by both negative hydrophobic free energy and negative molecular free energy which compensated for the positive free energies due to the conformational change loss of rotational and transitional freedom of the DNA helix.

  13. Study of the melting of simple substances under high pressures

    International Nuclear Information System (INIS)

    Stishov, S.M.

    Results of studies on the thermodynamic properties of Ar, Na, and Cs are reported. The main laws governing the thermodynamics of melting of these elements were established. Experimental P-V-T data were obtained characterizing the equations of state for the liquid and solid phases of argon, sodium, and cesium at pressures up to 17 x 10 3 kg/cm 2 (Ar), 22 x 10 3 kg/cm 2 (Na,Cs), and temperatures up to 75 0 C (Ar), 220 0 C (Na), and 200 0 C (Cs). Thermodynamic functions and their derivatives were calculated for the liquid and solid phases of these elements using empirical equations of state approximating the experimental P-V-T data with good accuracy

  14. The rolling-circle melting-pot model for porcine circovirus DNA replication

    Science.gov (United States)

    A stem-loop structure, formed by a pair of inverted repeats during DNA replication, is a conserved feature at the origin of DNA replication (Ori) among plant and animal viruses, bacteriophages and plasmids that replicate their genomes via the rolling-circle replication (RCR) mechanism. Porcine circo...

  15. Microfludic device for creating ionic strength gradients over DNA microarrays for efficient DNA melting studies and assay development

    DEFF Research Database (Denmark)

    Petersen, Jesper; Poulsen, Lena; Birgens, Henrik

    2009-01-01

    microfluidic device that creates a gradient comprising zones of defined ionic strength over a glass slide, in which each zone corresponds to a subarray. Using this device, we demonstrated that ionic strength gradients function in a similar fashion as corresponding thermal gradients in assay development. More...... specifically, we noted that (i) the two stringency modulators generated melting curves that could be compared, (ii) both led to increased assay robustness, and (iii) both were associated with difficulties in genotyping the same mutation. These findings demonstrate that ionic strength stringency buffers can...

  16. Research on Melt Degassing Processes of High Conductivity Hard Drawn Aluminum Wire

    Science.gov (United States)

    Xu, Xuexia; Feng, Yanting; Wang, Qing; Li, Wenbin; Fan, Hui; Wang, Yong; Li, Guowei; Zhang, Daoqian

    2018-03-01

    Degassing effects of ultrasonic and vacuum processes on high conductivity hard drawn aluminum melt were studied. Results showed that the degassing efficiency improved with the increase of ultrasonic power within certain range, stabilizing at 70% with 240W. For vacuum degassing process, hydrogen content of aluminum melt decreased with the loading time and was linear with logarithm of vacuum degree. Comparison of degassing effects of ultrasonic, vacuum, vacuum-ultrasonic degassing process showed that vacuum-ultrasonic process presented optimal effect.

  17. Viscosity measurements on metal melts at high pressure and viscosity calculations for the earth's core

    International Nuclear Information System (INIS)

    Mineev, Vladimir N; Funtikov, Aleksandr I

    2004-01-01

    A review is given of experimental and calculated data on the viscosity of iron-based melts on the melting curve. The interest in these data originates in the division of opinion on whether viscosity increases rather moderately or considerably in the high-pressure range. This disagreement is especially pronounced in the interpretation of the values of molten iron and its compounds in the environment of the earth's outer core. The conclusion on a substantial rise in viscosity mostly follows from the universal law, proposed by Brazhkin and Lyapin [1], of viscosity changing along the metal melting curve in the high-pressure range. The review analyzes available experimental and computational data, including the most recent ones. Data on viscosity of metals under shock wave compression in the megabar pressure range are also discussed. It is shown that data on viscosity of metal melts point to a small increase of viscosity on the melting curve. Specifics are discussed of the phase diagram of iron made more complex by the presence of several phase transitions and by the uncertainty in the position of the melting curve in the high-pressure range. Inaccuracies that arise in extrapolating the results of viscosity measurements to the pressure range corresponding to the earth's core environment are pointed out. (reviews of topical problems)

  18. The high resolution melting analysis (HRM) as a molecular tool for monitoring parasites of the wildlife.

    Science.gov (United States)

    Héritier, Laurent; Verneau, Olivier; Breuil, Gregory; Meistertzheim, Anne-Leila

    2017-04-01

    In an interconnected world, the international pet trade on wild animals is becoming increasingly important. As a consequence, non-native parasite species are introduced, which affect the health of wildlife and contribute to the loss of biodiversity. Because the investigation of parasite diversity within vulnerable host species implies the molecular identification of large samples of parasite eggs, the sequencing of DNA barcodes is time-consuming and costly. Thereby, the objectives of our study were to apply the high resolution melting (HRM) approach for species determination from pools of parasite eggs. Molecular assays were validated on flatworm parasites (polystomes) infecting the Mediterranean pond turtle Mauremys leprosa and the invasive red-eared slider Trachemys scripta elegans in French natural environments. HRM analysis results indicated that double or multiple parasitic infections could be detected from wild animal populations. They also showed that the cycle of parasite eggs production was not regular over time and may depend on several factors, among which the ecological niche and the target species. Thereby, monitoring parasites from wild endangered animals implies periodic parasitological surveys to avoid false negative diagnostics, based solely on eggs production.

  19. High-resolution melting analysis for prenatal diagnosis of beta-thalassemia in northern Thailand.

    Science.gov (United States)

    Charoenkwan, Pimlak; Sirichotiyakul, Supatra; Phusua, Arunee; Suanta, Sudjai; Fanhchaksai, Kanda; Sae-Tung, Rattika; Sanguansermsri, Torpong

    2017-12-01

    High-resolution melting (HRM) analysis is a rapid mutation analysis which assesses the pattern of reduction of fluorescence signal after subjecting the amplified PCR product with saturated fluorescence dye to an increasing temperature. We used HRM analysis for prenatal diagnosis of beta-thalassemia disease in northern Thailand. Five PCR-HRM protocols were used to detect point mutations in five different segments of the beta-globin gene, and one protocol to detect the 3.4 kb beta-globin deletion. We sought to characterize the mutations in carriers and to enable prenatal diagnosis in 126 couples at risk of having a fetus with beta-thalassemia disease. The protocols identified 18 common mutations causing beta-thalassemia, including the rare codon 132 (A-T) mutation. Each mutation showed a specific HRM pattern and all results were in concordance with those from direct DNA sequencing or gap-PCR methods. In cases of beta-thalassemia disease resulting from homozygosity for a mutation or compound heterozygosity for two mutations on the same amplified segment, the HRM patterns were different to those of a single mutation and were specific for each combination. HRM analysis is a simple and useful method for mutation identification in beta-thalassemia carriers and prenatal diagnosis of beta-thalassemia in northern Thailand.

  20. Melting point of polymers under high pressure Part I: Influence of the polymer properties

    International Nuclear Information System (INIS)

    Seeger, Andreas; Freitag, Detlef; Freidel, Frank; Luft, Gerhard

    2004-01-01

    The pressure dependence of the melting point of various polymers including homo- and copolymers (HDPE, LDPE, PP and ethylene vinyl acetate copolymers (EVA)) was investigated under nitrogen atmosphere up to 330 MPa within a high pressure differential thermal analysis cell designed by our group. The properties of the polymers (vinylacetate content, melt flow index, molecular weight, isotactic index, crystallinity, density, and frequency of branching) have been correlated with the change of the melting point under pressure (dT m /dp). It could be shown that the melting point always increases linearly with pressure up to 330 MPa. The pressure dependence was found to be in the range of 11-17 K/(100 MPa). From these results it is possible to approximate dT m /dp using the enthalpy of fusion of the polymers at ambient pressure

  1. Influence of gas generation on high-temperature melt/concrete interactions

    International Nuclear Information System (INIS)

    Powers, D.A.

    1979-01-01

    Accidents involving fuel melting and eventual contact between the high temperature melt and structural concrete may be hypothesized for both light water thermal reactors and liquid metal cooled breeder reactors. Though these hypothesized accidents have a quite low probability of occurring, it is necessary to investigate the probable natures of the accidents if an adequate assessment of the risks associated with the use of nuclear reactors is to be made. A brief description is given of a program addressing the nature of melt/concrete interactions which has been underway for three years at Sandia Laboratories. Emphasis in this program has been toward the behavior of prototypic melts of molten core materials with concrete representative of that found in existing or proposed reactors. The goals of the experimentation have been to identify phenomena particularly pertinent to questions of reactor safety, and phenomena particularly pertinent to questions of reactor safety, and provide quantitative data suitable for the purposes of risk assessment

  2. Shear melting and high temperature embrittlement: theory and application to machining titanium.

    Science.gov (United States)

    Healy, Con; Koch, Sascha; Siemers, Carsten; Mukherji, Debashis; Ackland, Graeme J

    2015-04-24

    We describe a dynamical phase transition occurring within a shear band at high temperature and under extremely high shear rates. With increasing temperature, dislocation deformation and grain boundary sliding are supplanted by amorphization in a highly localized nanoscale band, which allows for massive strain and fracture. The mechanism is similar to shear melting and leads to liquid metal embrittlement at high temperature. From simulation, we find that the necessary conditions are lack of dislocation slip systems, low thermal conduction, and temperature near the melting point. The first two are exhibited by bcc titanium alloys, and we show that the final one can be achieved experimentally by adding low-melting-point elements: specifically, we use insoluble rare earth metals (REMs). Under high shear, the REM becomes mixed with the titanium, lowering the melting point within the shear band and triggering the shear-melting transition. This in turn generates heat which remains localized in the shear band due to poor heat conduction. The material fractures along the shear band. We show how to utilize this transition in the creation of new titanium-based alloys with improved machinability.

  3. A novel typing method for Listeria monocytogenes using high-resolution melting analysis (HRMA) of tandem repeat regions.

    Science.gov (United States)

    Ohshima, Chihiro; Takahashi, Hajime; Iwakawa, Ai; Kuda, Takashi; Kimura, Bon

    2017-07-17

    Listeria monocytogenes, which is responsible for causing food poisoning known as listeriosis, infects humans and animals. Widely distributed in the environment, this bacterium is known to contaminate food products after being transmitted to factories via raw materials. To minimize the contamination of products by food pathogens, it is critical to identify and eliminate factory entry routes and pathways for the causative bacteria. High resolution melting analysis (HRMA) is a method that takes advantage of differences in DNA sequences and PCR product lengths that are reflected by the disassociation temperature. Through our research, we have developed a multiple locus variable-number tandem repeat analysis (MLVA) using HRMA as a simple and rapid method to differentiate L. monocytogenes isolates. While evaluating our developed method, the ability of MLVA-HRMA, MLVA using capillary electrophoresis, and multilocus sequence typing (MLST) was compared for their ability to discriminate between strains. The MLVA-HRMA method displayed greater discriminatory ability than MLST and MLVA using capillary electrophoresis, suggesting that the variation in the number of repeat units, along with mutations within the DNA sequence, was accurately reflected by the melting curve of HRMA. Rather than relying on DNA sequence analysis or high-resolution electrophoresis, the MLVA-HRMA method employs the same process as PCR until the analysis step, suggesting a combination of speed and simplicity. The result of MLVA-HRMA method is able to be shared between different laboratories. There are high expectations that this method will be adopted for regular inspections at food processing facilities in the near future. Copyright © 2017. Published by Elsevier B.V.

  4. High porosity harzburgite and dunite channels for the transport of compositionally heterogeneous melts in the mantle: II. Geochemical consequences

    Science.gov (United States)

    Liang, Y.; Schiemenz, A.; Xia, Y.; Parmentier, E.

    2009-12-01

    In a companion numerical study [1], we explored the spatial distribution of high porosity harzburgite and dunite channels produced by reactive dissolution of orthopyroxene (opx) in an upwelling mantle column and identified a number of new features. In this study, we examine the geochemical consequences of channelized melt flow under the settings outlined in [1] with special attention to the transport of compositionally heterogeneous melts and their interactions with the surrounding peridotite matrix during melt migration in the mantle. Time-dependent transport equations for a trace element in the interstitial melt and solids that include advection, dispersion, and melt-rock reaction were solved in a 2-D upwelling column using the high-order numerical methods outlined in [1]. The melt and solid velocities were taken from the steady state or quasi-steady state solutions of [1]. In terms of trace element fractionation, the simulation domain can be divided into 4 distinct regions: (a) high porosity harzburgite channel, overlain by; (b) high porosity dunite channel; (c) low porosity compacting boundary layer surrounding the melt channels; and (d) inter-channel regions outside (c). In the limit of local chemical equilibrium, melting in region (d) is equivalent to batch melting, whereas melting and melt extraction in (c) is more close to fractional melting with the melt suction rate first increase from the bottom of the melting column to a maximum near the bottom of the dunite channel and then decrease upward in the compacting boundary layer. The melt composition in the high porosity harzburgite channel is similar to that produced by high-degree batch melting (up to opx exhaustion), whereas the melt composition in the dunite is a weighted average of the ultra-depleted melt from the harzburgite channel below, the expelled melt from the compacting boundary layer, and melt produced by opx dissolution along the sidewalls of the dunite channel. Compaction within the dunite

  5. Planetesimal core formation with partial silicate melting using in-situ high P, high T, deformation x-ray microtomography

    Science.gov (United States)

    Anzures, B. A.; Watson, H. C.; Yu, T.; Wang, Y.

    2017-12-01

    Differentiation is a defining moment in formation of terrestrial planets and asteroids. Smaller planetesimals likely didn't reach high enough temperatures for widescale melting. However, we infer that core formation must have occurred within a few million years from Hf-W dating. In lieu of a global magma ocean, planetesimals likely formed through inefficient percolation. Here, we used in-situ high temperature, high pressure, x-ray microtomography to track the 3-D evolution of the sample at mantle conditions as it underwent shear deformation. Lattice-Boltzmann simulations for permeability were used to characterize the efficiency of melt percolation. Mixtures of KLB1 peridotite plus 6.0 to 12.0 vol% FeS were pre-sintered to achieve an initial equilibrium microstructure, and then imaged through several consecutive cycles of heating and deformation. The maximum calculated melt segregation velocity was found to be 0.37 cm/yr for 6 vol.% FeS and 0.61 cm/year for 12 vol.% FeS, both below the minimum velocity of 3.3 cm/year required for a 100km planetesimal to fully differentiate within 3 million years. However, permeability is also a function of grain size and thus the samples having smaller grains than predicted for small planetesimals could have contributed to low permeability and also low migration velocity. The two-phase (sulfide melt and silicate melt) flow at higher melt fractions (6 vol.% and 12 vol.% FeS) was an extension of a similar study1 containing only sulfide melt at lower melt fraction (4.5 vol.% FeS). Contrary to the previous study, deformation did result in increased permeability until the sample was sheared by twisting the opposing Drickamer anvils by 360 degrees. Also, the presence of silicate melt caused the FeS melt to coalesce into less connected pathways as the experiment with 6 vol.% FeS was found to be less permeable than the one with 4.5 vol.% FeS but without any partial melt. The preliminary data from this study suggests that impacts as well as

  6. Environmentally friendly and highly productive bi-component melt spinning of thermoregulated smart polymer fibres with high latent heat capacity

    Directory of Open Access Journals (Sweden)

    Ch. Cherif

    2018-03-01

    Full Text Available A stable and reproducible bi-component melt spinning process on an industrial scale incorporating Phase Change Material (PCM into textile fibres has been successfully developed and carried out using a melt spinning machine. The key factor for a successful bi-component melt spinning process is that a deep insight into the thermal and rheological behaviour of PCM using Difference Scanning Calorimetry (DSC, Thermogravimetric Analysis (TGA, and an oscillatory rheological investigation. PCM is very sensitive to the temperature and residence time of the melt spinning process. It is found that the optimal process temperature of PCM is 210 °C. The textile-physical properties and the morphology of the melt spun and further drawn bi-component core and sheath fibres (bico fibres were investigated and interpreted. The heat capacities of PCM incorporated in bico fibres were also determined by means of DSC. The melt spun bico fibres integrating PCM provide a high latent heat of up to 22 J/g, which is three times higher than that of state-of-the-art fibres, which were also obtained using the melt spinning process. Therefore, they have the potential to be used as smart polymer fibres for textile and other technical applications.

  7. Consistent increase in High Asia's runoff due to increasing glacier melt and precipitation

    NARCIS (Netherlands)

    Lutz, A. F.; Immerzeel, W. W.|info:eu-repo/dai/nl/290472113; Shrestha, A. B.; Bierkens, M. F P|info:eu-repo/dai/nl/125022794

    Rivers originating in the high mountains of Asia are among the most meltwater-dependent river systems on Earth, yet large human populations depend on their resources downstream1. Across High Asias river basins, there is large variation in the contribution of glacier and snow melt to total runoff 2,

  8. High-resolution melting curve analysis for rapid detection of mutations in a Medaka TILLING library

    Directory of Open Access Journals (Sweden)

    Deguchi Tomonori

    2010-09-01

    Full Text Available Abstract Background During the last two decades, DNA sequencing has led to the identification of numerous genes in key species; however, in most cases, their functions are still unknown. In this situation, reverse genetics is the most suitable method to assign function to a gene. TILLING (Targeting Induced Local Lesions IN Genomes is a reverse-genetic strategy that combines random chemical mutagenesis with high-throughput discovery of the induced mutations in target genes. The method has been applied to a variety of plant and animal species. Screening of the induced mutations is the most important step in TILLING. Currently, direct sequencing or nuclease-mediated screening of heteroduplexes is widely used for detection of mutations in TILLING. Both methods are useful, but the costs are substantial and turnaround times are relatively long. Thus, there is a need for an alternative method that is of higher throughput and more cost effective. Results In this study, we developed a high resolution melting (HRM assay and evaluated its effectiveness for screening ENU-induced mutations in a medaka TILLING library. We had previously screened mutations in the p53 gene by direct sequencing. Therefore, we first tested the efficiency of the HRM assay by screening mutations in p53, which indicated that the HRM assay is as useful as direct sequencing. Next, we screened mutations in the atr and atm genes with the HRM assay. Nonsense mutations were identified in each gene, and the phenotypes of these nonsense mutants confirmed their loss-of-function nature. Conclusions These results demonstrate that the HRM assay is useful for screening mutations in TILLING. Furthermore, the phenotype of the obtained mutants indicates that medaka is an excellent animal model for investigating genome stability and gene function, especially when combined with TILLING.

  9. A demonstration experiment of steam-driven, high-pressure melt ejection

    International Nuclear Information System (INIS)

    Allen, M.D.; Pitch, M.; Nichols, R.T.

    1990-08-01

    A steam blowdown test was performed at the Surtsey Direct Heating Test Facility to test the steam supply system and burst diaphragm arrangement that will be used in subsequent Surtsey Direct Containment Heating (DCH) experiments. Following successful completion of the steam blowdown test, the HIPS-10S (High-Pressure Melt Streaming) experiment was conducted to demonstrate that the technology to perform steam-driven, high-pressure melt ejection (HPME) experiments has been successfully developed. In addition, the HIPS-10S experiment was used to assess techniques and instrumentation design to create the proper timing of events in HPME experiments. This document discusses the results of this test

  10. A technique of melting temperature measurement and its application for irradiated high-burnup MOX fuels

    International Nuclear Information System (INIS)

    Namekawa, Takashi; Hirosawa, Takashi

    1999-01-01

    A melting temperature measurement technique for irradiated oxide fuels is described. In this technique, the melting temperature was determined from a thermal arrest on a heating curve of the specimen which was enclosed in a tungsten capsule to maintain constant chemical composition of the specimen during measurement. The measurement apparatus was installed in an alpha-tight steel box within a gamma-shielding cell and operated by remote handling. The temperature of the specimen was measured with a two-color pyrometer sighted on a black-body well at the bottom of the tungsten capsule. The diameter of the black-body well was optimized so that the uncertainties of measurement were reduced. To calibrate the measured temperature, two reference melting temperature materials, tantalum and molybdenum, were encapsulated and run before and after every oxide fuel test. The melting temperature data on fast reactor mixed oxide fuels irradiated up to 124 GWd/t were obtained. In addition, simulated high-burnup mixed oxide fuel up to 250 GWd/t by adding non-radioactive soluble fission products was examined. These data shows that the melting temperature decrease with increasing burnup and saturated at high burnup region. (author)

  11. 1D/2D analyses of the lower head vessel in contact with high temperature melt

    International Nuclear Information System (INIS)

    Chang, Jong Eun; Cho, Jae Seon; Suh, Kune Y.; Chung, Chang H.

    1998-01-01

    One- and two-dimensional analyses were performed for the ceramic/metal melt and the vessel to interpret the temperature history of the outer surface of the vessel wall measured from typical Al 2 O 3 /Fe thermite melt tests LAVA (Lower-plenum Arrested Vessel Attack) spanning heatup and cooldown periods. The LAVA tests were conducted at the Korea Atomic Energy Research Institute (KAERI) during the process of high temperature molten material relocation from the delivery duct down into the water in the test vessel pressurized to 2.0 MPa. Both analyses demonstrated reasonable predictions of the temperature history of the LHV (Lower Head Vessel). The comparison sheds light on the thermal hydraulic and material behavior of the high temperature melt within the hemispherical vessel

  12. Systematic prediction of high-pressure melting curves of transition metals

    International Nuclear Information System (INIS)

    Hieu, Ho Khac

    2014-01-01

    The pressure effects on melting temperatures of transition metals have been studied based on the combination of the modified Lindemann criterion with statistical moment method in quantum statistical mechanics. Numerical calculations have been performed for five transition metals including Cu, Pd, Pt, Ni, and Mn up to pressure 100 GPa. Our results are in good and reasonable agreements with available experimental data. This approach gives us a relatively simple method for qualitatively calculating high-pressure melting temperature. Moreover, it can be used to verify future experimental and theoretical works. This research proposes the potential of the combination of statistical moment method and the modified Lindemann criterion on predicting high-pressure melting of materials.

  13. Development of melt compositions for sulphate bearing high level waste

    International Nuclear Information System (INIS)

    Jahagirdar, P.B.; Wattal, P.K.

    1997-09-01

    The report deals with the development and characterization of vitreous matrices for sulphate bearing high level waste. Studies were conducted in sodium borosilicate and lead borosilicate systems with the introduction of CaO, BaO, MgO etc. Lead borosilicate system was found to be compatible with sulphate bearing high level wastes. Detailed product evaluation carried on selected formulations is also described. (author)

  14. Rapid identification and classification of bacteria by 16S rDNA restriction fragment melting curve analyses (RFMCA).

    Science.gov (United States)

    Rudi, Knut; Kleiberg, Gro H; Heiberg, Ragnhild; Rosnes, Jan T

    2007-08-01

    The aim of this work was to evaluate restriction fragment melting curve analyses (RFMCA) as a novel approach for rapid classification of bacteria during food production. RFMCA was evaluated for bacteria isolated from sous vide food products, and raw materials used for sous vide production. We identified four major bacterial groups in the material analysed (cluster I-Streptococcus, cluster II-Carnobacterium/Bacillus, cluster III-Staphylococcus and cluster IV-Actinomycetales). The accuracy of RFMCA was evaluated by comparison with 16S rDNA sequencing. The strains satisfying the RFMCA quality filtering criteria (73%, n=57), with both 16S rDNA sequence information and RFMCA data (n=45) gave identical group assignments with the two methods. RFMCA enabled rapid and accurate classification of bacteria that is database compatible. Potential application of RFMCA in the food or pharmaceutical industry will include development of classification models for the bacteria expected in a given product, and then to build an RFMCA database as a part of the product quality control.

  15. High Resolution Melting Analysis Targeting hsp70 as a Fast and Efficient Method for the Discrimination of Leishmania Species.

    Science.gov (United States)

    Zampieri, Ricardo Andrade; Laranjeira-Silva, Maria Fernanda; Muxel, Sandra Marcia; Stocco de Lima, Ana Carolina; Shaw, Jeffrey Jon; Floeter-Winter, Lucile Maria

    2016-02-01

    Protozoan parasites of the genus Leishmania cause a large spectrum of clinical manifestations known as Leishmaniases. These diseases are increasingly important public health problems in many countries both within and outside endemic regions. Thus, an accurate differential diagnosis is extremely relevant for understanding epidemiological profiles and for the administration of the best therapeutic protocol. Exploring the High Resolution Melting (HRM) dissociation profiles of two amplicons using real time polymerase chain reaction (real-time PCR) targeting heat-shock protein 70 coding gene (hsp70) revealed differences that allowed the discrimination of genomic DNA samples of eight Leishmania species found in the Americas, including Leishmania (Leishmania) infantum chagasi, L. (L.) amazonensis, L. (L.) mexicana, L. (Viannia) lainsoni, L. (V.) braziliensis, L. (V.) guyanensis, L. (V.) naiffi and L. (V.) shawi, and three species found in Eurasia and Africa, including L. (L.) tropica, L. (L.) donovani and L. (L.) major. In addition, we tested DNA samples obtained from standard promastigote culture, naturally infected phlebotomines, experimentally infected mice and clinical human samples to validate the proposed protocol. HRM analysis of hsp70 amplicons is a fast and robust strategy that allowed for the detection and discrimination of all Leishmania species responsible for the Leishmaniases in Brazil and Eurasia/Africa with high sensitivity and accuracy. This method could detect less than one parasite per reaction, even in the presence of host DNA.

  16. High Resolution Melting Analysis Targeting hsp70 as a Fast and Efficient Method for the Discrimination of Leishmania Species.

    Directory of Open Access Journals (Sweden)

    Ricardo Andrade Zampieri

    2016-02-01

    Full Text Available Protozoan parasites of the genus Leishmania cause a large spectrum of clinical manifestations known as Leishmaniases. These diseases are increasingly important public health problems in many countries both within and outside endemic regions. Thus, an accurate differential diagnosis is extremely relevant for understanding epidemiological profiles and for the administration of the best therapeutic protocol.Exploring the High Resolution Melting (HRM dissociation profiles of two amplicons using real time polymerase chain reaction (real-time PCR targeting heat-shock protein 70 coding gene (hsp70 revealed differences that allowed the discrimination of genomic DNA samples of eight Leishmania species found in the Americas, including Leishmania (Leishmania infantum chagasi, L. (L. amazonensis, L. (L. mexicana, L. (Viannia lainsoni, L. (V. braziliensis, L. (V. guyanensis, L. (V. naiffi and L. (V. shawi, and three species found in Eurasia and Africa, including L. (L. tropica, L. (L. donovani and L. (L. major. In addition, we tested DNA samples obtained from standard promastigote culture, naturally infected phlebotomines, experimentally infected mice and clinical human samples to validate the proposed protocol.HRM analysis of hsp70 amplicons is a fast and robust strategy that allowed for the detection and discrimination of all Leishmania species responsible for the Leishmaniases in Brazil and Eurasia/Africa with high sensitivity and accuracy. This method could detect less than one parasite per reaction, even in the presence of host DNA.

  17. Melting temperatures of MgO under high pressure determined by micro-texture observation

    Science.gov (United States)

    Kimura, T.; Ohfuji, H.; Nishi, M.; Irifune, T.

    2016-12-01

    Periclase (MgO) is the second abundant mineral after bridgmanite in the Earth's lower mantle, and its melting temperature (Tm) under pressure is important to constrain the chemical composition of ultra-deep magma formed near the mantle-core boundary. However, the melting behavior is highly controversial among previous studies: a laser-heated diamond anvil cell (LHDAC) study reported a melting curve with a dTm/dP of 30 K/GPa at zero pressure [1], while several theoretical computations gave substantially higher dTm/dP of 90 100 K/GPa [2,3]. We performed a series of LHDAC experiments for measurements of Tm of MgO under high pressure, using single crystal MgO as the starting material. The melting was detected by using micro-texture observations of the quenched samples. We found that the laser-heated area of the sample quenched from the Tm in previous LHDAC experiments [1] showed randomly aggregated granular crystals, which was not caused by melting, but by plastic deformation of the sample. This suggests that the Tms of their study were substantially underestimated. On the other hand, the sample recovered from the temperature higher by 1500-1700 K than the Tms in previous LHDAC experiments showed a characteristic internal texture comparable to the solidification texture typically shown in metal casting. We determined the Tms based on the observation of this texture up to 32 GPa. Fitting our Tms to the Simon equation yields dTm/dP of 82 K/GPa at zero pressure, which is consistent with those of the theoretical predictions (90 100 K/GPa) [2,3]. Extrapolation of the present melting curve of MgO to the pressure of the CMB (135 GPa) gives a melting temperature of 8900 K. The present steep melting slope offers the eutectic composition close to peridotite (in terms of Mg/Si ratio) throughout the lower mantle conditions. According to the model for sink/float relationship between the solid mantle and the magma [4], a considerable amount of iron (Fe/(Mg+Fe) > 0.24) is expected

  18. Process to Continuously Melt, Refine and Cast High Quality Steel

    Energy Technology Data Exchange (ETDEWEB)

    None

    2005-09-01

    The purpose of this project is to conduct research and development targeted at designing a revolutionary steelmaking process. This process will deliver high quality steel from scrap to the casting mold in one continuous process and will be safer, more productive, and less capital intensive to build and operate than conventional steelmaking. The new process will produce higher quality steel faster than traditional batch processes while consuming less energy and other resources.

  19. Behavior of a corium jet in high pressure melt ejection from a reactor pressure vessel

    International Nuclear Information System (INIS)

    Frid, W.

    1988-04-01

    Discharge of the molten core debris from a pressurized reactor vessel has been recognized as an important accident scenario for pressurized water reactors. Recent high-pressure melt streaming experiments conducted at Sandia National Laboratories, designed to study cavity and containment events related to melt ejection, have resulted in two important observations: (1) Expansion and breakup of the ejected molten jet. (2) Significant aerosol generation during the ejection process. The expansion and breakup of the jet in the experiments are attributed to rapid evolution of the pressurizing gas (nitrogen or hydrogen) dissolved in the melt. It has been concluded that aerosol particles may be formed by condensation of melt vapor and mechanical breakup of the melt and generation. It was also shown that the above stated phenomena are likely to occur in reactor accidents. This report provides results from analytical and experimental investigations on the behavior of a gas supersaturated molten jet expelled from a pressurized vessel. Aero-hydrodynamic stability of liquid jets in gas, stream degassing of molten metals, and gas bubble nucleation in molten metals are relevant problems that are addressed in this work

  20. Molecular identification of broomrape species from a single seed by High Resolution Melting analysis

    Directory of Open Access Journals (Sweden)

    Mathieu Rolland

    2016-12-01

    Full Text Available Broomrapes are holoparasitic plants spreading through seeds. Each plant produces hundreds of thousands of seeds which remain viable in the soils for decades. To limit their spread, drastic measures are being taken and the contamination of a commercial seed lot by a single broomrape seed can lead to its rejection. Considering that broomrapes species identification from a single seed is extremely difficult even for trained botanists and that among all the described species, only a few are really noxious for the crops, numerous seed lots are rejected because of the contamination by seeds of non-noxious broomrape species. The aim of this study was to develop and evaluate a High Resolution Melting assay identifying the eight most noxious and common broomrape species (P. aegyptiaca, O. cernua, O. crenata, O. cumana, O. foetida, O. hederae, O. minor, and P. ramosa from a single seed. Based on trnL and rbcL plastidial genes amplification, the designed assay successfully identifies O. cumana, O. cernua, O. crenata, O. minor, O. hederae, and O. foetida; P. ramosa and P. aegyptiaca can be differentiated from other species but not from each other. Tested on 50 seed lots, obtained results perfectly matched identifications performed by sequencing. Through the analysis of common seed lots by different analysts, the reproducibility of the assay was evaluated at 90 %. Despite an original sample preparation process it was not possible to extract enough DNA from some seeds (10% of the samples. The described assay fulfils its objectives and allows an accurate identification of the targeted broomrape species. It can be used to identify contaminants in commercial seed lots or for any other purpose. The assay might be extended to vegetative material.

  1. Microscopic Origins of the Anomalous Melting Behavior of Sodium under High Pressure

    Science.gov (United States)

    Eshet, Hagai; Khaliullin, Rustam Z.; Kühne, Thomas D.; Behler, Jörg; Parrinello, Michele

    2012-03-01

    X-ray diffraction experiments have shown that sodium exhibits a dramatic pressure-induced drop in melting temperature, which extends from 1000 K at ˜30GPa to as low as room temperature at ˜120GPa. Despite significant theoretical effort to understand the anomalous melting, its origins are still debated. In this work, we reconstruct the sodium phase diagram by using an ab initio quality neural-network potential. Furthermore, we demonstrate that the reentrant behavior results from the screening of interionic interactions by conduction electrons, which at high pressure induces a softening in the short-range repulsion.

  2. Method for preparation of melts of alkali metal chlorides with highly volatile polyvalent metal chlorides

    International Nuclear Information System (INIS)

    Salyulev, A.B.; Kudyakov, V.Ya.

    1990-01-01

    A method for production of alkali metal (Cs, Rb, K) chloride melts with highly volatile polyvalent metal chlorides is suggested. The method consists, in saturation of alkali metal chlorides, preheated to the melting point, by volatile component vapours (titanium tetrachloride, molybdenum or tantalum pentachloride) in proportion, corresponding to the composition reguired. The saturation is realized in an evacuated vessel with two heating areas for 1-1.5 h. After gradual levelling of temperature in both areas the product is rapidly cooled. 1 fig.; 1 tab

  3. Melting curves of molecular hydrogen and molecular deuterium under high pressures between 20 and 373 K

    International Nuclear Information System (INIS)

    Diatschenko, V.; Chu, C.W.; Liebenberg, D.H.; Young, D.A.; Ross, M.; Mills, R.L.

    1985-01-01

    We determined the melting curve of molecular hydrogen and molecular deuterium at closely spaced intervals from 20 to 373 K by two different techniques using high-pressure diamond cells. The cells were loaded with liquid at low temperature or with compressed gas at room temperature. Empirical functions for the melting curves were evaluated from least-squares fits of the data. Values of the compressibility and Debye temperature were computed at melting, and the results are compared with those calculated from various theoretical models. The good agreement shows that the models are generally valid, although small systematic deviations may point the way toward refinements in modeling. Our study also demonstrates the need to determine a one-piece intermolecular potential valid over a wide pressure range by refitting all experimental data, including the shock data recently made available

  4. Transmission and selection of macrolide resistant Mycoplasma genitalium infections detected by rapid high resolution melt analysis.

    Directory of Open Access Journals (Sweden)

    Jimmy Twin

    Full Text Available BACKGROUND: Mycoplasma genitalium (MG causes urethritis, cervicitis and pelvic inflammatory disease. The MG treatment failure rate using 1 g azithromycin at an Australian Sexual Health clinic in 2007-9 was 31% (95%CI 23-40%. We developed a rapid high resolution melt analysis (HRMA assay targeting resistance mutations in the MG 23S rRNA gene, and validated it against DNA sequencing by examining pre- and post-treatment archived samples from MG-infected patients. METHODOLOGY/PRINCIPAL FINDINGS: Available MG-positive pre-treatment (n = 82 and post-treatment samples from individuals with clinical treatment failure (n = 20 were screened for 23S rRNA gene mutations. Sixteen (20% pre-treatment samples possessed resistance mutations (A2058G, A2059G, A2059C, which were significantly more common in patients with symptomatic azithromycin-treatment failure (12/26; 44% than in those clinically cured (4/56; 7%, p<0.001. All 20 patients experiencing azithromycin-failure had detectable mutations in their post-treatment samples. In 9 of these cases, the same mutational types were present in both pre- and post-treatment samples indicating transmitted resistance, whilst in 11 of these cases (55%, mutations were absent in pre-treatment samples indicating likely selection of resistant isolates have occurred. HRMA was able to detect all mutational changes determined in this study by DNA sequencing. An additional HRMA assay incorporating an unlabelled probe was also developed to detect type 4 single-nucleotide polymorphisms found in other populations, with a slightly lower sensitivity of 90%. CONCLUSIONS/SIGNIFICANCE: Treatment failure is associated with the detection of macrolide resistance mutations, which appear to be almost equally due to selection of resistant isolates following exposure to 1 g azithromycin and pre-existing transmitted resistance. The application of a rapid molecular assay to detect resistance at the time of initial detection of infection allows

  5. Structure, production and properties of high-melting compounds and systems (hard materials and hard metals)

    International Nuclear Information System (INIS)

    Holleck, H.; Thuemmler, F.

    1979-07-01

    The report contains contributions by various authors to the research project on the production, structure, and physical properties of high-melting compounds and systems (hard metals and hard materials), in particular WC-, TaC-, and MoC-base materials. (GSCH) [de

  6. High-temperature apparatus for chaotic mixing of natural silicate melts

    Energy Technology Data Exchange (ETDEWEB)

    Morgavi, D.; Petrelli, M.; Vetere, F. P.; González-García, D.; Perugini, D., E-mail: diego.perugini@unipg.it [Department of Physics and Geology, Petro-Volcanology Research Group (PVRG), University of Perugia, Piazza Università, Perugia 06100 (Italy)

    2015-10-15

    A unique high-temperature apparatus was developed to trigger chaotic mixing at high-temperature (up to 1800 °C). This new apparatus, which we term Chaotic Magma Mixing Apparatus (COMMA), is designed to carry out experiments with high-temperature and high-viscosity (up to 10{sup 6} Pa s) natural silicate melts. This instrument allows us to follow in time and space the evolution of the mixing process and the associated modulation of chemical composition. This is essential to understand the dynamics of magma mixing and related chemical exchanges. The COMMA device is tested by mixing natural melts from Aeolian Islands (Italy). The experiment was performed at 1180 °C using shoshonite and rhyolite melts, resulting in a viscosity ratio of more than three orders of magnitude. This viscosity ratio is close to the maximum possible ratio of viscosity between high-temperature natural silicate melts. Results indicate that the generated mixing structures are topologically identical to those observed in natural volcanic rocks highlighting the enormous potential of the COMMA to replicate, as a first approximation, the same mixing patterns observed in the natural environment. COMMA can be used to investigate in detail the space and time development of magma mixing providing information about this fundamental petrological and volcanological process that would be impossible to investigate by direct observations. Among the potentials of this new experimental device is the construction of empirical relationships relating the mixing time, obtained through experimental time series, and chemical exchanges between the melts to constrain the mixing-to-eruption time of volcanic systems, a fundamental topic in volcanic hazard assessment.

  7. High-temperature apparatus for chaotic mixing of natural silicate melts

    International Nuclear Information System (INIS)

    Morgavi, D.; Petrelli, M.; Vetere, F. P.; González-García, D.; Perugini, D.

    2015-01-01

    A unique high-temperature apparatus was developed to trigger chaotic mixing at high-temperature (up to 1800 °C). This new apparatus, which we term Chaotic Magma Mixing Apparatus (COMMA), is designed to carry out experiments with high-temperature and high-viscosity (up to 10 6 Pa s) natural silicate melts. This instrument allows us to follow in time and space the evolution of the mixing process and the associated modulation of chemical composition. This is essential to understand the dynamics of magma mixing and related chemical exchanges. The COMMA device is tested by mixing natural melts from Aeolian Islands (Italy). The experiment was performed at 1180 °C using shoshonite and rhyolite melts, resulting in a viscosity ratio of more than three orders of magnitude. This viscosity ratio is close to the maximum possible ratio of viscosity between high-temperature natural silicate melts. Results indicate that the generated mixing structures are topologically identical to those observed in natural volcanic rocks highlighting the enormous potential of the COMMA to replicate, as a first approximation, the same mixing patterns observed in the natural environment. COMMA can be used to investigate in detail the space and time development of magma mixing providing information about this fundamental petrological and volcanological process that would be impossible to investigate by direct observations. Among the potentials of this new experimental device is the construction of empirical relationships relating the mixing time, obtained through experimental time series, and chemical exchanges between the melts to constrain the mixing-to-eruption time of volcanic systems, a fundamental topic in volcanic hazard assessment

  8. Multiscale radar mapping of surface melt over mountain glaciers in High Mountain Asia

    Science.gov (United States)

    Steiner, N.; McDonald, K. C.

    2017-12-01

    Glacier melt dominates input for many hydrologic systems in the Himalayan Hindukush region that feed rivers that are critical for downstream ecosystems and hydropower generation in this highly populated area. Deviation in seasonal surface melt timing and duration with a changing climate has the potential to affect up to a billion people on the Indian Subcontinent. Satellite-borne microwave remote sensing has unique capabilities that allow monitoring of numerous landscape processes associated with snowmelt and freeze/thaw state, without many of the limitations in optical-infrared sensors such as solar illumination or atmospheric conditions. The onset of regional freeze/thaw and surface melting transitions determine important surface hydrologic variables like river discharge. Theses regional events are abrupt therefore difficult to observe with low-frequency observation sensors. Recently launched synthetic aperture radar (SAR) onboard the Sentinel-1 A and B satellites from the European Space Agency (ESA) provide wide-swath and high spatial resolution (50-100 m) C-Band SAR observations with observations frequencies not previously available, on the order of 8 to 16 days. The Sentinel SARs provide unique opportunity to study freeze/thaw and mountain glacier melt dynamics at process level scales, spatial and temporal. The melt process of individual glaciers, being fully resolved by imaging radar, will inform on the radiometric scattering physics associated with surface hydrology during the transition from melted to thawed state and during refreeze. Backscatter observations, along with structural information about the surface will be compared with complimentary coarse spatial resolution C-Band radar scatterometers, Advanced Scatterometer (ASCAT Met Op A+B), to understand the sub-pixel contribution of surface melting and freeze/thaw signals. This information will inform on longer-scale records of backscatter from ASCAT, 2006-2017. We present a comparison of polarimetric C

  9. Development of foams from linear polypropylene (PP) and high melt strength polypropylene (HMSPP) polymeric blends

    International Nuclear Information System (INIS)

    Cardoso, Elisabeth Carvalho Leite

    2009-01-01

    Foamed polymers are future materials, with a comprehensive application field. They can be used in order to improve appearance of insulation structures, for example, or to reduce costs involving materials. This work address to Isotactic Polypropylene / High Melt Strength Polypropylene blends, for foams production. Rheological behavior of polymer melt, especially referring to viscosity in processing temperature, plays a decisive role in applications where dominates extensional flow, as in case of foaming. If the viscosity is very low, it will correspond to a low melt strength, as in case of linear homopolymer (Isotact PP), and the foam will be prejudiced, due to the impossibility of expansion. Otherwise, if the viscosity is very high, with a high melt strength, the foam will collapse immediately after its formation. In order to get foams with an homogeneous and defined cellular structure, there were accomplished blends, 50% in weight, between linear homopolymer (isotactic PP) and HMSPP, from PP modified as per gamma radiation, in acetylene environment and at a 12.5 kGy doses. Extrusion process used a soluble foaming methodology, according to a processing/dissolution principle, which involves the dissolution of a Physical Blowing Agent (PBA), under 30 bar pressure, homogeneously mixed with polymeric melt. Extrusion conditions, that generally involve temperature, pressure and viscoelastic material flow control were experimentally investigated to define prevalent characteristics for producing foams. Nitrogen was the used PBA and process extrusion parameters were adapted to PP, HMSPP and their 50% in weight mixtures thereof. Major PP and HMSPP characteristics were obtained via melt Index and melt strength and thermal analyses (DSC/TGA), in order to make viable and to reproduce foaming as per extrusion process. Foams cellular morphology of PP, HMSPP and their 50% in weight mixtures thereof was investigated, with and without talc addition, as nucleating agent, by using

  10. Performance of High Layer Thickness in Selective Laser Melting of Ti6Al4V

    Directory of Open Access Journals (Sweden)

    Xuezhi Shi

    2016-12-01

    Full Text Available To increase building rate and save cost, the selective laser melting (SLM of Ti6Al4V with a high layer thickness (200 μm and low cost coarse powders (53 μm–106 μm at a laser power of 400 W is investigated in this preliminary study. A relatively large laser beam with a diameter of 200 μm is utilized to produce a stable melt pool at high layer thickness, and the appropriate scanning track, which has a smooth surface with a shallow contact angle, can be obtained at the scanning speeds from 40 mm/s to 80 mm/s. By adjusting the hatch spacings, the density of multi-layer samples can be up to 99.99%, which is much higher than that achieved in previous studies about high layer thickness selective laser melting. Meanwhile, the building rate can be up to 7.2 mm3/s, which is about 2 times–9 times that of the commercial equipment. Besides, two kinds of defects are observed: the large un-melted defects and the small spherical micropores. The formation of the un-melted defects is mainly attributed to the inappropriate overlap rates and the unstable scanning tracks, which can be eliminated by adjusting the processing parameters. Nevertheless, the micropores cannot be completely eliminated. It is worth noting that the high layer thickness plays a key role on surface roughness rather than tensile properties during the SLM process. Although a sample with a relatively coarse surface is generated, the average values of yield strength, ultimate tensile strength, and elongation are 1050 MPa, 1140 MPa, and 7.03%, respectively, which are not obviously different than those with the thin layer thickness used in previous research; this is due to the similar metallurgical bonding and microstructure.

  11. Performance of High Layer Thickness in Selective Laser Melting of Ti6Al4V.

    Science.gov (United States)

    Shi, Xuezhi; Ma, Shuyuan; Liu, Changmeng; Chen, Cheng; Wu, Qianru; Chen, Xianping; Lu, Jiping

    2016-12-01

    To increase building rate and save cost, the selective laser melting (SLM) of Ti6Al4V with a high layer thickness (200 μm) and low cost coarse powders (53 μm-106 μm) at a laser power of 400 W is investigated in this preliminary study. A relatively large laser beam with a diameter of 200 μm is utilized to produce a stable melt pool at high layer thickness, and the appropriate scanning track, which has a smooth surface with a shallow contact angle, can be obtained at the scanning speeds from 40 mm/s to 80 mm/s. By adjusting the hatch spacings, the density of multi-layer samples can be up to 99.99%, which is much higher than that achieved in previous studies about high layer thickness selective laser melting. Meanwhile, the building rate can be up to 7.2 mm³/s, which is about 2 times-9 times that of the commercial equipment. Besides, two kinds of defects are observed: the large un-melted defects and the small spherical micropores. The formation of the un-melted defects is mainly attributed to the inappropriate overlap rates and the unstable scanning tracks, which can be eliminated by adjusting the processing parameters. Nevertheless, the micropores cannot be completely eliminated. It is worth noting that the high layer thickness plays a key role on surface roughness rather than tensile properties during the SLM process. Although a sample with a relatively coarse surface is generated, the average values of yield strength, ultimate tensile strength, and elongation are 1050 MPa, 1140 MPa, and 7.03%, respectively, which are not obviously different than those with the thin layer thickness used in previous research; this is due to the similar metallurgical bonding and microstructure.

  12. Laser melting of groove defect repair on high thermal conductivity steel (HTCS-150)

    Science.gov (United States)

    Norhafzan, B.; Aqida, S. N.; Fazliana, F.; Reza, M. S.; Ismail, I.; Khairil, C. M.

    2018-02-01

    This paper presents laser melting repair of groove defect on HTCS-150 surface using Nd:YAG laser system. Laser melting process was conducted using JK300HPS Nd:YAG twin lamp laser source with 1064 nm wavelength and pulsed mode. The parameters are pulse repetition frequency (PRF) that is set from 70 to 100 Hz, average power ( P A) of 50-70 W, and laser spot size of 0.7 mm. HTCS-150 samples were prepared with groove dimension of 0.3 mm width and depths of 0.5 mm using EDM wire cut. Groove defect repaired using laser melting process on groove surface area with various parameters' process. The melted surface within the groove was characterized for subsurface hardness profile, roughness, phase identification, chemical composition, and metallographic study. The roughness analysis indicates high PRF at large spot size caused high surface roughness and low surface hardness. Grain refinement of repaired layer was analyzed within the groove as a result of rapid heating and cooling. The hardness properties of modified HTCS inside the groove and the bulk surface increased two times from as received HTCS due to grain refinement which is in agreement with Hall-Petch equation. These findings are significant to parameter design of die repair for optimum surface integrity and potential for repairing crack depth and width of less than 0.5 and 0.3 mm, respectively.

  13. Molecular differentiation of Opisthorchis viverrini and Clonorchis sinensis eggs by multiplex real-time PCR with high resolution melting analysis.

    Science.gov (United States)

    Kaewkong, Worasak; Intapan, Pewpan M; Sanpool, Oranuch; Janwan, Penchom; Thanchomnang, Tongjit; Laummaunwai, Porntip; Lulitanond, Viraphong; Doanh, Pham Ngoc; Maleewong, Wanchai

    2013-12-01

    Opisthorchis viverrini and Clonorchis sinensis are parasites known to be carcinogenic and causative agents of cholangiocarcinoma in Asia. The standard method for diagnosis for those parasite infections is stool examination to detect parasite eggs. However, the method has low sensitivity, and eggs of O. viverrini and C. sinensis are difficult to distinguish from each other and from those of some other trematodes. Here, we report a multiplex real-time PCR coupled with high resolution melting (HRM) analysis for the differentiation of O. viverrini and C. sinensis eggs in fecal samples. Using 2 pairs of species-specific primers, DNA sequences from a portion of the mitochondrial NADH dehydrogenase subunit 2 (nad 2) gene, were amplified to generate 209 and 165 bp products for O. viverrini and C. sinensis, respectively. The distinct characteristics of HRM patterns were analyzed, and the melting temperatures peaked at 82.4±0.09℃ and 85.9±0.08℃ for O. viverrini and C. sinensis, respectively. This technique was able to detect as few as 1 egg of O. viverrini and 2 eggs of C. sinensis in a 150 mg fecal sample, which is equivalent to 7 and 14 eggs per gram of feces, respectively. The method is species-specific, rapid, simple, and does not require fluorescent probes or post-PCR processing for discrimination of eggs of the 2 species. It offers a new tool for differentiation and detection of Asian liver fluke infections in stool specimens.

  14. The use of high resolution melting analysis to detect Fabry mutations in heterozygous females via dry bloodspots.

    Science.gov (United States)

    Tai, Chang-Long; Liu, Mei-Ying; Yu, Hsiao-Chi; Chiang, Chiang-Chuan; Chiang, Hung; Suen, Jeng-Hung; Kao, Shu-Min; Huang, Yu-Hsiu; Wu, Tina Jui-Ting; Yang, Chia-Feng; Tsai, Fang-Chih; Lin, Ching-Yuang; Chang, Jan-Gowth; Chen, Hong-Duo; Niu, Dau-Ming

    2012-02-18

    As an X-linked genetic disorder, Fabry disease was first thought to affect males only, and females were generally considered to be asymptomatic carriers. However, recent research suggests that female carriers of Fabry disease may still develop vital organ damage causing severe morbidity and mortality. In the previous newborn screening, from 299,007 newborns, we identified a total of 20 different Fabry mutations and 121 newborns with Fabry mutations. However, we found that most female carriers are not detected by enzyme assays. A streamlined method for high resolution melting (HRM) analysis was designed to screen for GLA gene mutations using a same PCR and melting program. Primer sets were designed to cover the 7 exons and the Chinese common intronic mutation, IVS4+919G>A of GLA gene. The HRM analysis was successful in identifying heterozygous and hemizygous patients with the 20 surveyed mutations. We were also successful in using this method to test dry blood spots of newborns afflicted with Fabry mutations without having to determine DNA concentration before PCR amplification. The results of this study show that HRM could be a reliable and sensitive method for use in the rapid screening of females for GLA mutations. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. High-resolution melt analysis for species identification of coagulase-negative staphylococci derived from bovine milk.

    Science.gov (United States)

    Ajitkumar, Praseeda; Barkema, Herman W; Zadoks, Ruth N; Morck, Douglas W; van der Meer, Frank J U M; De Buck, Jeroen

    2013-03-01

    Coagulase-negative staphylococci (CNS) are the most frequently isolated pathogens isolated from bovine milk. In this study, we report a rapid assay for species identification of CNS using high-resolution melt analysis (HRMA) of 16S rDNA sequences. Real-time polymerase chain reaction amplification of 16S rRNA gene fragment, spanning the variable region V1 and V2, was performed with a resulting amplicon of 215 bp. A library of distinct melt curves of reference strains of 13 common CNS species was created using HRMA. Sequencing of 16S rRNA and rpoB genes, and, when needed, tuf gene, of 100 CNS isolates obtained from Canadian Bovine Mastitis Research Network was done to determine their species identity, allowing for subsequent evaluation of the performance of HRMA for field isolates of bovine CNS. A combination of HRMA and sequencing revealed that Staphylococcus chromogenes, S. xylosus, S. simulans, and S. sciuri had multiple genotypes, complicating their resolution by HRMA. As the 3 genotypes of S. chromogenes had distinct melt curves, the 3 distinct genotypes were employed as reference strains in a blinded trial of 156 CNS isolates to identify S. chromogenes. HRMA correctly identified all S. chromogenes isolates which were later confirmed by sequencing. Staphylococcus chromogenes (68%) was most frequently found among the CNS isolates, followed by S. haemolyticus (10%) and S. xylosus (6%). The present study revealed that HRMA of 16S rRNA gene (V1-V2) could be used as a rapid, efficient, low-cost, and minimally cumbersome technique for S. chromogenes identification, the most common CNS derived from bovine milk. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. High resolution melting curve analysis as a new tool for rapid identification of canine parvovirus type 2 strains.

    Science.gov (United States)

    Bingga, Gali; Liu, Zhicheng; Zhang, Jianfeng; Zhu, Yujun; Lin, Lifeng; Ding, Shuangyang; Guo, Pengju

    2014-01-01

    A high resolution melting (HRM) curve method was developed to identify canine parvovirus type 2 (CPV-2) strains by nested PCR. Two sets of primers, CPV-426F/426R and CPV-87R/87F, were designed that amplified a 52 bp and 53 bp product from the viral VP2 capsid gene. The region amplified by CPV-426F/426R included the A4062G and T4064A mutations in CPV-2a, CPV-2b and CPV-2c. The region amplified by CPV-87F/87R included the A3045T mutation in the vaccine strains of CPV-2 and CPV-2a, CPV-2b and CPV-2c. Faecal samples were obtained from 30 dogs that were CPV antigen-positive. The DNA was isolated from the faecal samples and PCR-amplified using the two sets of primers, and genotyped by HRM curve analysis. The PCR-HRM assay was able to distinguish single nucleotide polymorphisms between CPV-2a, CPV-2b and CPV-2c using CPV-426F/426R. CPV-2a was distinguished from CPV-2b and CPV-2c by differences in the melting temperature. CPV-2b and CPV-2c could be distinguished based on the shape of the melting curve after generating heteroduplexes using a CPV-2b reference sample. The vaccine strains of CPV-2 were identified using CPV-87F/87R. Conventional methods for genotyping CPV strains are labor intensive, expensive or time consuming; the present PCR-based HRM assay might be an attractive alternative. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. Rapid detection and differentiation of Clonorchis sinensis and Opisthorchis viverrini using real-time PCR and high resolution melting analysis.

    Science.gov (United States)

    Cai, Xian-Quan; Yu, Hai-Qiong; Li, Rong; Yue, Qiao-Yun; Liu, Guo-Hua; Bai, Jian-Shan; Deng, Yan; Qiu, De-Yi; Zhu, Xing-Quan

    2014-01-01

    Clonorchis sinensis and Opisthorchis viverrini are both important fish-borne pathogens, causing serious public health problem in Asia. The present study developed an assay integrating real-time PCR and high resolution melting (HRM) analysis for the specific detection and rapid identification of C. sinensis and O. viverrini. Primers targeting COX1 gene were highly specific for these liver flukes, as evidenced by the negative amplification of closely related trematodes. Assays using genomic DNA extracted from the two flukes yielded specific amplification and their identity was confirmed by sequencing, having the accuracy of 100% in reference to conventional methods. The assay was proved to be highly sensitive with a detection limit below 1 pg of purified genomic DNA, 5 EPG, or 1 metacercaria of C. sinensis. Moreover, C. sinensis and O. viverrini were able to be differentiated by their HRM profiles. The method can reduce labor of microscopic examination and the contamination of agarose electrophoresis. Moreover, it can differentiate these two flukes which are difficult to be distinguished using other methods. The established method provides an alternative tool for rapid, simple, and duplex detection of C. sinensis and O. viverrini.

  18. Rapid Detection and Differentiation of Clonorchis sinensis and Opisthorchis viverrini Using Real-Time PCR and High Resolution Melting Analysis

    Directory of Open Access Journals (Sweden)

    Xian-Quan Cai

    2014-01-01

    Full Text Available Clonorchis sinensis and Opisthorchis viverrini are both important fish-borne pathogens, causing serious public health problem in Asia. The present study developed an assay integrating real-time PCR and high resolution melting (HRM analysis for the specific detection and rapid identification of C. sinensis and O. viverrini. Primers targeting COX1 gene were highly specific for these liver flukes, as evidenced by the negative amplification of closely related trematodes. Assays using genomic DNA extracted from the two flukes yielded specific amplification and their identity was confirmed by sequencing, having the accuracy of 100% in reference to conventional methods. The assay was proved to be highly sensitive with a detection limit below 1 pg of purified genomic DNA, 5 EPG, or 1 metacercaria of C. sinensis. Moreover, C. sinensis and O. viverrini were able to be differentiated by their HRM profiles. The method can reduce labor of microscopic examination and the contamination of agarose electrophoresis. Moreover, it can differentiate these two flukes which are difficult to be distinguished using other methods. The established method provides an alternative tool for rapid, simple, and duplex detection of C. sinensis and O. viverrini.

  19. Melt-drop technique for the production of high-purity metal powder

    International Nuclear Information System (INIS)

    Aldinger, F.; Linck, E.; Claussen, N.

    1977-01-01

    The production of high-purity powders of metals and alloys such as beryllium, titanium alloys, or superalloys is a problem. Oxidation of these materials cannot be avoided. Oxidation occurs in inert gases and even in reducing atmospheres when any gas impurities are present. Therefore, the powder production of these materials has to be performed either in high vacuum or at least in a static atmosphere of inert gas purified immediately before coming into contact with the disintegrating material. These requirements are very well met by the melt-drop technique presented in this paper, especially for coarse powders which must not necessarily be cold-workable. This is true, for example, for superalloys where high-temperature applications require large grain sizes; or in titanium alloys because the final microstructure will be achieved by a thermomechanical treatment. In the case of beryllium and beryllium alloys, where grain sizes <5 μm are desired, further milling is necessary. But the melt-drop technique offers a simple and clean method directly from the purifying process of vacuum melting. In melt-drop processes a liquid metal flows through a nozzle at the bottom of a crucible or the melt is just poured through a sieve. The theory of disintegration of a liquid jet into droplets, dates back to the 19th century. More recent investigations attempted to produce uniformly sized droplets by applying a capillary wave of given wave length to the jet. But this has been done only with non-metallic materials. Evidence is presented to prove the theory and show that this concept is applicable to the production of metal powders with controlled particle size

  20. High-density Bi-Pb-Sr-Ca-Cu-O superconductor prepared by rapid thermal melt processing

    Science.gov (United States)

    Moon, B. M.; Lalevic, B.; Kear, B. H.; McCandlish, L. E.; Safari, A.; Meskoob, M.

    1989-10-01

    A high quality, dense Bi-Pb-Sr-Ca-Cu-O superconductor has been successfully synthesized by rapid thermal melt processing. Conventionally sintered pellets were melted at 1200 °C, cooled rapidly, and then annealed. As-melted samples exhibited semiconductor behavior, which upon annealing became superconducting at 115 K [Tc(zero)=105 K]. A detailed study of various processing techniques has been carried out.

  1. Quantifying the surface energy fluxes in South Greenland during the 2012 high melt episodes using in-situ observations

    Directory of Open Access Journals (Sweden)

    Robert S. Fausto

    2016-09-01

    Full Text Available Two high melt episodes occurred on the Greenland ice sheet in July 2012, during which nearly the entire ice sheet surface experienced melting. Observations from an automatic weather station (AWS in the lower ablation area in South Greenland reveal the largest daily melt rates (up to 28 cm d-1 ice equivalent ever recorded on the ice sheet. The two melt episodes lasted 6 days, equivalent to 6% of the June-August melt period, but contributed 14 % to the total annual ablation of 8.5 m ice equivalent. We employ a surface energy balance model driven by AWS data to quantify the relative importance of the energy budget components contributing to melt through the melt season. During the days with largest daily melt rates, surface turbulent heat input peaked at 552 Wm-2, 77 % of the surface melt energy, which is otherwise typically dominated by absorbed solar radiation. We find that rain contributed ca. 7 % to melt during these episodes.

  2. The WAIS Melt Monitor: An automated ice core melting system for meltwater sample handling and the collection of high resolution microparticle size distribution data

    Science.gov (United States)

    Breton, D. J.; Koffman, B. G.; Kreutz, K. J.; Hamilton, G. S.

    2010-12-01

    Paleoclimate data are often extracted from ice cores by careful geochemical analysis of meltwater samples. The analysis of the microparticles found in ice cores can also yield unique clues about atmospheric dust loading and transport, dust provenance and past environmental conditions. Determination of microparticle concentration, size distribution and chemical makeup as a function of depth is especially difficult because the particle size measurement either consumes or contaminates the meltwater, preventing further geochemical analysis. Here we describe a microcontroller-based ice core melting system which allows the collection of separate microparticle and chemistry samples from the same depth intervals in the ice core, while logging and accurately depth-tagging real-time electrical conductivity and particle size distribution data. This system was designed specifically to support microparticle analysis of the WAIS Divide WDC06A deep ice core, but many of the subsystems are applicable to more general ice core melting operations. Major system components include: a rotary encoder to measure ice core melt displacement with 0.1 millimeter accuracy, a meltwater tracking system to assign core depths to conductivity, particle and sample vial data, an optical debubbler level control system to protect the Abakus laser particle counter from damage due to air bubbles, a Rabbit 3700 microcontroller which communicates with a host PC, collects encoder and optical sensor data and autonomously operates Gilson peristaltic pumps and fraction collectors to provide automatic sample handling, melt monitor control software operating on a standard PC allowing the user to control and view the status of the system, data logging software operating on the same PC to collect data from the melting, electrical conductivity and microparticle measurement systems. Because microparticle samples can easily be contaminated, we use optical air bubble sensors and high resolution ice core density

  3. Constraints on the Parental Melts of Enriched Shergottites from Image Analysis and High Pressure Experiments

    Science.gov (United States)

    Collinet, M.; Medard, E.; Devouard, B.; Peslier, A.

    2012-01-01

    Martian basalts can be classified in at least two geochemically different families: enriched and depleted shergottites. Enriched shergottites are characterized by higher incompatible element concentrations and initial Sr-87/Sr-86 and lower initial Nd-143/Nd-144 and Hf-176/Hf-177 than depleted shergottites [e.g. 1, 2]. It is now generally admitted that shergottites result from the melting of at least two distinct mantle reservoirs [e.g. 2, 3]. Some of the olivine-phyric shergottites (either depleted or enriched), the most magnesian Martian basalts, could represent primitive melts, which are of considerable interest to constrain mantle sources. Two depleted olivine-phyric shergottites, Yamato (Y) 980459 and Northwest Africa (NWA) 5789, are in equilibrium with their most magnesian olivine (Fig. 1) and their bulk rock compositions are inferred to represent primitive melts [4, 5]. Larkman Nunatak (LAR) 06319 [3, 6, 7] and NWA 1068 [8], the most magnesian enriched basalts, have bulk Mg# that are too high to be in equilibrium with their olivine megacryst cores. Parental melt compositions have been estimated by subtracting the most magnesian olivine from the bulk rock composition, assuming that olivine megacrysts have partially accumulated [3, 9]. However, because this technique does not account for the actual petrography of these meteorites, we used image analysis to study these rocks history, reconstruct their parent magma and understand the nature of olivine megacrysts.

  4. High-Resolution Melt Analysis for Rapid Comparison of Bacterial Community Compositions

    DEFF Research Database (Denmark)

    Hjelmsø, Mathis Hjort; Hansen, Lars Hestbjerg; Bælum, Jacob

    2014-01-01

    In the study of bacterial community composition, 16S rRNA gene amplicon sequencing is today among the preferred methods of analysis. The cost of nucleotide sequence analysis, including requisite computational and bioinformatic steps, however, takes up a large part of many research budgets. High......-resolution melt (HRM) analysis is the study of the melt behavior of specific PCR products. Here we describe a novel high-throughput approach in which we used HRM analysis targeting the 16S rRNA gene to rapidly screen multiple complex samples for differences in bacterial community composition. We hypothesized...... that HRM analysis of amplified 16S rRNA genes from a soil ecosystem could be used as a screening tool to identify changes in bacterial community structure. This hypothesis was tested using a soil microcosm setup exposed to a total of six treatments representing different combinations of pesticide...

  5. Differentiation of minute virus of mice and mouse parvovirus by high resolution melting curve analysis.

    Science.gov (United States)

    Rao, Dan; Wu, Miaoli; Wang, Jing; Yuan, Wen; Zhu, Yujun; Cong, Feng; Xu, Fengjiao; Lian, Yuexiao; Huang, Bihong; Wu, Qiwen; Chen, Meili; Zhang, Yu; Huang, Ren; Guo, Pengju

    2017-12-01

    Murine parvovirus is one of the most prevalent infectious pathogens in mouse colonies. A specific primer pair targeting the VP2 gene of minute virus of mice (MVM) and mouse parvovirus (MPV) was utilized for high resolution melting (HRM) analysis. The resulting melting curves could distinguish these two virus strains and there was no detectable amplification of the other mouse pathogens which included rat parvovirus (KRV), ectromelia virus (ECT), mouse adenovirus (MAD), mouse cytomegalovirus (MCMV), polyoma virus (Poly), Helicobactor hepaticus (H. hepaticus) and Salmonella typhimurium (S. typhimurium). The detection limit of the standard was 10 copies/μL. This study showed that the PCR-HRM assay could be an alternative useful method with high specificity and sensitivity for differentiating murine parvovirus strains MVM and MPV. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Authenticity analyses of Rhizoma Paridis using barcoding coupled with high resolution melting (Bar-HRM) analysis to control its quality for medicinal plant product.

    Science.gov (United States)

    Duan, Bao-Zhong; Wang, Ya-Ping; Fang, Hai-Lan; Xiong, Chao; Li, Xi-Wen; Wang, Ping; Chen, Shi-Lin

    2018-01-01

    Rhizoma Paridis (Chonglou) is a commonly used and precious traditional Chinese medicine. Paris polyphylla Smith var. yunnanensis (Franch.) Hand. -Mazz. and Paris polyphylla Smith var . chinensis (Franch.) Hara are the two main sources of Chonglou under the monograph of Rhizoma Paridis in Chinese Pharmacopoeia. In the local marketplace, however, this medicine is prone to be accidentally contaminated, deliberately substituted or admixed with other species that are similar to Rhizoma Paridis in shape and color. Consequently, these adulterations might compromise quality control and result in considerable health concerns for consumers. This study aims to develop a rapid and sensitive method for accurate identification of Rhizoma Paridis and its common adulterants. DNA barcoding coupled with high resolution melting analysis was applied in this research to distinguish Rhizoma Paridis from its adulteration. The internal transcribed spacer 2 (ITS2) barcode was selected for HRM analysis to produce standard melting profile of the selected species. DNA of the tested herbal medicines was isolated and their melting profiles were generated and compared with the standard melting profile of P. polyphylla var. chinensis . The results indicate that the ITS2 molecular regions coupled with HRM analysis can effectively differentiate nine herbal species, including two authentic origins of Chonglou and their seven common adulterants. Ten herbal medicines labeled "Chonglou" obtained from a local market were collected and identified with our methods, and their sequence information was analyzed to validate the accuracy of HRM analysis. DNA barcoding coupled with HRM analysis is a accurate, reliable, rapid, cost-effective and robust tool, which could contribute to the quality control of Rhizoma Paridis in the supply chain of the natural health product industry (NHP).

  7. Lambda-Cyhalothrin Nanosuspension Prepared by the Melt Emulsification-High Pressure Homogenization Method

    OpenAIRE

    Pan, Zhenzhong; Cui, Bo; Zeng, Zhanghua; Feng, Lei; Liu, Guoqiang; Cui, Haixin; Pan, Hongyu

    2015-01-01

    The nanosuspension of 5% lambda-cyhalothrin with 0.2% surfactants was prepared by the melt emulsification-high pressure homogenization method. The surfactants composition, content, and homogenization process were optimized. The anionic surfactant (1-dodecanesulfonic acid sodium salt) and polymeric surfactant (maleic rosin-polyoxypropylene-polyoxyethylene ether sulfonate) screened from 12 types of commercially common-used surfactants were used to prepare lambda-cyhalothrin nanosuspension with ...

  8. Atomistic simulation of the premelting of iron and aluminum : Implications for high-pressure melting-curve measurements

    NARCIS (Netherlands)

    Starikov, Sergey V.; Stegailov, Vladimir V.

    2009-01-01

    Using atomistic simulations we show the importance of the surface premelting phenomenon for the melting-curve measurements at high pressures. The model under consideration mimics the experimental conditions deployed for melting studies with diamond-anvil cells. The iron is considered in this work

  9. Thermal depinning and melting of the flux-line lattice in high-Tc superconductors

    International Nuclear Information System (INIS)

    Brandt, E.H.

    1991-01-01

    In high-T c superconductors (HTSC) the thermal fluctuation of the vortex lattice (VL) may become large since the vortex lattice is soft due to the strong overlap of the vortex fields and since the temperature T and b high. It was thus argued that the three-dimensional (3D) vortex lattice is thermally entangled and may melt. This type of transition and the consequences of melting are not clear as yet since the always present pinning of the vortex ores by material inhomogeneities may cause similar disorder. In HTSC the pinning energy may become comparable with k B T because the coherence length ξ(vortex radius) is small and t may be high. Therefore, thermally activated depinning competes with possible effects of flux melting, and the irreversibility line in the B-T-plane (B = magnetic field) should better be called depinning line. In this paper it is argued that theories predicting new phases of vortex matter (flux solid, flux liquid, vortex plasma, vortex glass, and hexatic vortex glass) may be improved by replacing the 2D straight-vortex interaction by the correct 3D interaction between all vortex segments

  10. First report of Tasmanian sheep strain (G2) genotype isolated from Iranian goat using the high resolution melting (HRM) analysis.

    Science.gov (United States)

    Hosseini-Safa, Ahmad; Mohag Hegh, Mohammad Ali; Pestechian, Nader; Ganji, Maryam; Mohammadi, Rasoul; Mahmoudi Lamouki, Reza; Rostami-Nejad, Mohammad

    2016-12-01

    The present study was aimed to evaluate E. granulosus genotypes isolated from goats using HRM analysis in Isfahan province. Cystic echincoccosis, so-called hydatidosis, is widespread infection caused by the larval stage of Echinococcus granulosus . This is an important zoonotic disease worldwide, especially in the developing countries such as Iran. To date, molecular studies mainly based on the mitochondrial DNA sequences have identified distinct genotypes termed G1-G10 which can differ in some characteristics such as the growth and infectivity to different intermediate hosts or the survival rate in the definitive hosts that are important for the development of control strategies. From August to December 2014, 1341 goats were investigated and hydatid cysts were collected from the liver and lungs of 43 infected goats in Isfahan province abattoirs, Isfahan, Iran. Total genomic DNA was extracted from each sample, amplified for the presence of polymorphism of mitochondrial gene coding for cytochrome c oxidase subunit 1 (CO1), using high resolution melting curve (HRM) method. the results of HRM analysis using the sequence of CO1 gene for 43 Echinococcus granulosus isolates from goats showed 31, 2 and 10 isolates were identified as G1, G2, and G3 genotypes, respectively. G1 is the predominant genotype in the isolated goat samples in Isfahan province, and the presence of G2 strain was reported for the first time in goat in Iran.

  11. High-resolution melting analysis (HRM) for differentiation of four major Taeniidae species in dogs Taenia hydatigena, Taenia multiceps, Taenia ovis, and Echinococcus granulosus sensu stricto.

    Science.gov (United States)

    Dehghani, Mansoureh; Mohammadi, Mohammad Ali; Rostami, Sima; Shamsaddini, Saeedeh; Mirbadie, Seyed Reza; Harandi, Majid Fasihi

    2016-07-01

    Tapeworms of the genus Taenia include several species of important parasites with considerable medical and veterinary significance. Accurate identification of these species in dogs is the prerequisite of any prevention and control program. Here, we have applied an efficient method for differentiating four major Taeniid species in dogs, i.e., Taenia hydatigena, T. multiceps, T. ovis, and Echinococcus granulosus sensu stricto. High-resolution melting (HRM) analysis is simpler, less expensive, and faster technique than conventional DNA-based assays and enables us to detect PCR amplicons in a closed system. Metacestode samples were collected from local abattoirs from sheep. All the isolates had already been identified by PCR-sequencing, and their sequence data were deposited in the GenBank. Real-time PCR coupled with HRM analysis targeting mitochondrial cox1 and ITS1 genes was used to differentiate taeniid species. Distinct melting curves were obtained from ITS1 region enabling accurate differentiation of three Taenia species and E. granulosus in dogs. The HRM curves of Taenia species and E .granulosus were clearly separated at Tm of 85 to 87 °C. In addition, double-pick melting curves were produced in mixed infections. Cox1 melting curves were not decisive enough to distinguish four taeniids. In this work, the efficiency of HRM analysis to differentiate four major taeniid species in dogs has been demonstrated using ITS1 gene.

  12. Possible role of rf melted microparticles on the operation of high-gradient accelerating structures

    Directory of Open Access Journals (Sweden)

    G. S. Nusinovich

    2009-10-01

    Full Text Available High-gradient accelerating structures should operate reliably for a long time. Therefore studies of various processes which may lead to disruption of such an operation are so important. In the present paper, the dissipation of rf electromagnetic energy in metallic microparticles is analyzed accounting for the temperature dependence of the skin depth. Such particles may appear in structures, for example, due to mechanical fracture of irises in strong rf electric fields. It is shown that such microparticles with dimensions on the order of the skin depth, being immersed in the region of strong rf magnetic field, can absorb enough energy in long-pulse operation to be melted. Then, the melted clumps can impinge on the surface of a structure and create nonuniformities leading to field enhancement and corresponding emission of dark current. Results are given for several geometries and materials of microparticles.

  13. A refined, rapid and reproducible high resolution melt (HRM-based method suitable for quantification of global LINE-1 repetitive element methylation

    Directory of Open Access Journals (Sweden)

    Tse M Yat

    2011-12-01

    Full Text Available Abstract Background The methylation of DNA is recognized as a key mechanism in the regulation of genomic stability and evidence for its role in the development of cancer is accumulating. LINE-1 methylation status represents a surrogate measure of genome-wide methylation. Findings Using high resolution melt (HRM curve analysis technology, we have established an in-tube assay that is linear (r > 0.9986 with a high amplification efficiency (90-105%, capable of discriminating between partcipant samples with small differences in methylation, and suitable for quantifying a wide range of LINE-1 methylation levels (0-100%--including the biologically relevant range of 50-90% expected in human DNA. We have optimized this procedure to perform using 2 μg of starting DNA and 2 ng of bisulfite-converted DNA for each PCR reaction. Intra- and inter-assay coefficients of variation were 1.44% and 0.49%, respectively, supporting the high reproducibility and precision of this approach. Conclusions In summary, this is a completely linear, quantitative HRM PCR method developed for the measurement of LINE-1 methylation. This cost-efficient, refined and reproducible assay can be performed using minimal amounts of starting DNA. These features make our assay suitable for high throughput analysis of multiple samples from large population-based studies.

  14. The use of radiation for the production of high melt strength polypropylene

    International Nuclear Information System (INIS)

    Lugao, A.B.; Cardoso, E.C.L.; Hustzler, B.; Tokumoto, S.; Mendes, A.N.F.

    2001-01-01

    PP suffers from low melt strength, i.e., the melted PP does not exhibits an increase in resistance to stretching during elongation. It is well known that the melt-strength properties of a polymer increase with molecular weight and with long chain branching due to the increase in the entanglement level (high melt strength PP-HMSPP). In spite of been the most fast growing polymeric commodity nowadays those new grades of PP and its development have been barely studied and its general chemical characterization have been even less studied with few exceptions. HMSPP as proposed by Montell patents are produced by low temperature and low dose irradiation of high molecular weight isotatica PP in N 2 atmosphere. So the well-known reactions would be mostly degradation and crosslinking. Degradation however is supposed to be the first and more intense reaction a the initial steps as already shown. So, according to Montell patents, another reaction is likely to occur, branching or T links competing with crosslinkings or H links. Radical are likely to decay very fast in amorphous phase, but under annealing the radicals entrapped in the crystal phase is likely to move to the boundary and react. The group of professor Silverman has already hypothesized in T links formation and also studied the radical decay of PP. It is easy to understand the difficulties in differentiate Hs from Ts links and even these links if in very small amount from the bulk or from virgin polymer, as the Ts links and even these links if in very small amount from the bulk or from virgin polymer, as the chemical groups and links are chemically speaking essentially the same

  15. Detection of Schistosoma mansoni and Schistosoma haematobium by Real-Time PCR with High Resolution Melting Analysis

    Directory of Open Access Journals (Sweden)

    Hany Sady

    2015-07-01

    Full Text Available The present study describes a real-time PCR approach with high resolution melting-curve (HRM assay developed for the detection and differentiation of Schistosoma mansoni and S. haematobium in fecal and urine samples collected from rural Yemen. The samples were screened by microscopy and PCR for the Schistosoma species infection. A pair of degenerate primers were designed targeting partial regions in the cytochrome oxidase subunit I (cox1 gene of S. mansoni and S. haematobium using real-time PCR-HRM assay. The overall prevalence of schistosomiasis was 31.8%; 23.8% of the participants were infected with S. haematobium and 9.3% were infected with S. mansoni. With regards to the intensity of infections, 22.1% and 77.9% of S. haematobium infections were of heavy and light intensities, respectively. Likewise, 8.1%, 40.5% and 51.4% of S. mansoni infections were of heavy, moderate and light intensities, respectively. The melting points were distinctive for S. mansoni and S. haematobium, categorized by peaks of 76.49 ± 0.25 °C and 75.43 ± 0.26 °C, respectively. HRM analysis showed high detection capability through the amplification of Schistosoma DNA with as low as 0.0001 ng/µL. Significant negative correlations were reported between the real-time PCR-HRM cycle threshold (Ct values and microscopic egg counts for both S. mansoni in stool and S. haematobium in urine (p < 0.01. In conclusion, this closed-tube HRM protocol provides a potentially powerful screening molecular tool for the detection of S. mansoni and S. haematobium. It is a simple, rapid, accurate, and cost-effective method. Hence, this method is a good alternative approach to probe-based PCR assays.

  16. Electron beam melting of high niobium containing TiAl alloy: feasibility investigation

    Energy Technology Data Exchange (ETDEWEB)

    Terner, Mathieu; Biamino, Sara; Epicoco, Paolo; Fino, Paolo; Pavese, Matteo; Badini, Claudio [Dipartimento di Scienza Applicata e Tecnologia, Politecnico di Torino (Italy); Penna, Andrea; Gennaro, Paolo [AvioProp, Novara (Italy); Hedin, Oscar; Ackelid, Ulf [Arcam AB, Molndal (Sweden); Sabbadini, Silvia; Pelissero, Federica [Avio SpA, Torino (Italy)

    2012-08-15

    Third generation {gamma}-TiAl alloys with a high niobium content, Ti-(47-48)Al-2Cr-8Nb, were processed by electron beam melting (EBM). This near-net-shape additive manufacturing process produces complex parts according to a CAD design. The starting powder is deposited layer by layer on the building table and selectively melted to progressively form the massive part. The EBM parameters such as layer thickness, melting temperature, scanning speed, or building strategy were set up to minimize porosity. The chemical composition of the built material is similar to the composition of the base powder despite a slight evaporation of aluminum and reveals a neglectable oxygen pick-up. The very fine equiaxed microstructure resulting after EBM can be then set up by heat treatment (HT). According to the HT temperature in particular, an equiaxed microstructure, a duplex microstructure with different lamellar ratio and a fully lamellar microstructure is obtained. Not only test bars have been produced but also complex parts such as demo low pressure turbine blades. (Copyright copyright 2012 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  17. Synthesis by irradiation and mechanism and structural characterization study of high melt strength polypropylene

    International Nuclear Information System (INIS)

    Lugao, Ademar Benevolo

    2004-01-01

    Polypropylene molecular structure is made only by linear molecules interacting by weak forces. The resulting PP has very low melt strength (MS). MS is important to make feasible to process PP by all the transformation technologies based on the free expansion of the melt. The aim of this work was to develop a new process to synthesize PP with crosslinks and/or long chain branches, known as High Melt Strength Polypropylene (HMSPP) and to characterize its structure and synthesis mechanism. HMSPP was obtained by the irradiation of PP under a crosslinking (acetylene) atmosphere or inert or oxidative one, followed by thermal treatment for radical recombination and thermal treatment for annihilation of the remaining radicals under reactive or inert atmosphere. The results from rheological characterization showed that the highest levels of MS were obtained by conducting irradiation and thermal treatments under crosslinking atmospheres. The results for the elucidation of reaction mechanism by electron spin resonance (ESR) showed that acetylene irradiation is effective in promoting the creation of double bonds, based on the formation of polyenil radicals. The results of structural unraveling showed that radiation promotes predominantly the degradation of atactic molecules or molecules with atactic defects. These results support the hypothesis of formation of branched PP molecules based on the reaction of those fragments with the double bonds created in the PP molecules. (author)

  18. Study of the weathering of high melt strength polypropylene (HMS-PP)

    Energy Technology Data Exchange (ETDEWEB)

    Oliani, Washington L.; Parra, Duclerc F.; Otaguro, Harumi; Lima, Luis F.C.P.; Lugao, Ademar B. [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)], E-mail: dfparra@ipen.br

    2007-07-01

    One of the reasons for the good acceptance of the commercial PP is the fact that market requires products with features of 'engineering plastics' with prices in the range of commodities. High melt strength polypropylene (HMSPP) grades are produced by radiation process and have improved rheology for melt blow processes. The melt strength (MS) properties of a polymer increase with molecular weight and with long chain branching due to the increase in the entanglement level. The main scope of this study was to evaluate the stability of HMS-PP prepared by gamma radiation with doses of 12.5, 20 kGy in comparison with virgin PP. Many variables influence the rate of degradation of polymers by photo-oxidation. The irradiance and permeability to oxygen are the most important factors but other factors such as temperature and moisture have also influenced the degradation rates. Polypropylenes are sensitive to oxidation due to the presence of the tertiary carbon atom. Therefore, effective stabilization against oxidation (thermo and photo oxidation) is required. The samples submitted to the natural aging for a period of six months were characterized by: tensile test, thermogravimetry analysis (TGA), optical microscopy, scanning electronic microscopy (SEM) and infrared spectroscopy (FTIR). SEM analysis showed particular aspects of cracks on the surface. The loss of tensile strength is associated to the presence of fractures. The results showed that pronounced oxidation followed by chain scission occur at the initial periods of weathering exposition of the HMS-PP. (author)

  19. Study of the weathering of high melt strength polypropylene (HMS-PP)

    International Nuclear Information System (INIS)

    Oliani, Washington L.; Parra, Duclerc F.; Otaguro, Harumi; Lima, Luis F.C.P.; Lugao, Ademar B.

    2007-01-01

    One of the reasons for the good acceptance of the commercial PP is the fact that market requires products with features of 'engineering plastics' with prices in the range of commodities. High melt strength polypropylene (HMSPP) grades are produced by radiation process and have improved rheology for melt blow processes. The melt strength (MS) properties of a polymer increase with molecular weight and with long chain branching due to the increase in the entanglement level. The main scope of this study was to evaluate the stability of HMS-PP prepared by gamma radiation with doses of 12.5, 20 kGy in comparison with virgin PP. Many variables influence the rate of degradation of polymers by photo-oxidation. The irradiance and permeability to oxygen are the most important factors but other factors such as temperature and moisture have also influenced the degradation rates. Polypropylenes are sensitive to oxidation due to the presence of the tertiary carbon atom. Therefore, effective stabilization against oxidation (thermo and photo oxidation) is required. The samples submitted to the natural aging for a period of six months were characterized by: tensile test, thermogravimetry analysis (TGA), optical microscopy, scanning electronic microscopy (SEM) and infrared spectroscopy (FTIR). SEM analysis showed particular aspects of cracks on the surface. The loss of tensile strength is associated to the presence of fractures. The results showed that pronounced oxidation followed by chain scission occur at the initial periods of weathering exposition of the HMS-PP. (author)

  20. High resolution melting analysis (HRM) for the assessment of clonality in feline B-cell lymphomas.

    Science.gov (United States)

    Henrich, Manfred; Scheffold, Svenja; Hecht, Werner; Reinacher, Manfred

    2018-06-01

    Analysis of clonality is gaining importance in diagnosing lymphomas in veterinary medicine. Usually, PCR for the analysis of antigen receptor rearrangement (PARR) is followed by electrophoretic separation of the PCR products. Aim of this study was to test the feasibility of HRM for the assessment of clonality in B-cell lymphomas of cats. High resolution melting analysis differentiates PCR products by their different melting point using the decrease in fluorescence of an intercalating dye during melting of the PCR product. Additionally, the method is easy to use with no post-PCR manipulation of the samples. Forty-seven feline B-cell lymphomas and 31 reactive lymphatic proliferations of cats were investigated by PARR followed either by capillary electrophoresis or an HRM assay. To objectify the interpretation of the HRM results a recently published mathematical approach was applied to the melting curve. To overcome discrepancies between the visual interpretation and the mathematical approach, the latter was modified to include testing of reproducibility and recognition of pseudoclonality. In 11 of 47 lymphoma cases clonal populations were detectable by HRM assay compared to 14 of 47 lymphomas in which clonal populations were detected by capillary electrophoresis assay. Neither of the methods showed a clonal pattern in any of the reactive samples. However, the HRM assay showed a unique pattern in cases of follicular lymphatic hyperplasia that had no corresponding pattern in capillary electrophoresis. The capillary electrophoresis assay could identify 3 lymphomas that were not detected by the HRM assay and is therefore regarded superior to the HRM assay. The comparison however, was hampered by the overall bad performance of the PARR, that might be the consequence of insufficient primer binding due to somatic hypermutation of the binding sites during antigen stimulated proliferation of the B lymphocytes. Copyright © 2018 Elsevier B.V. All rights reserved.

  1. Development of high-frequency induction melting system for radioactive solid wastes

    International Nuclear Information System (INIS)

    Kawaguchi, Ichiro; Yamazaki, Seichiro; Takahashi, Noriaki; Kugai, Katsutoshi; Yokozawa, Minoru

    2004-01-01

    Kawasaki Heavy Industries, Ltd. developed an active insulation (AI) method radiofrequency melting system as a new melting treatment system of radioactive solid wastes and proved production of waste satisfied the treatment performances and burying by repeating many practical melting tests. The melting vessel uses a low-priced ceramic canister with nonelectrical conductivity, which is able to treat wastes with large amount of inorganic substances. The wastes melted in the canister is taken out the canister itself from radiofrequency melting reactor and solidified after cooling. The cool canister is stored in 2001 metal drum filling up a gap with mortal for laying underground. New radiofrequency melting reactor, 1/3 scale melting test, estimation of scale effects, melting tests for practical use and the total system are explained. (S.Y.)

  2. High-resolution melting (HRM) analysis as a feasible method for detecting spinal muscular atrophy via dried blood spots.

    Science.gov (United States)

    Er, Tze-Kiong; Kan, Tzu-Min; Su, Yu-Fa; Liu, Ta-Chih; Chang, Jan-Gowth; Hung, Shih-Ya; Jong, Yuh-Jyh

    2012-11-12

    Spinal muscular atrophy (SMA) is a neurodegenerative disease with the leading genetic cause of infant mortality. More than 95% of patients with SMA have a homozygous disruption in the survival motor neuron1 (SMN1) gene, caused by mutation, deletion, or rearrangement. Recently, treatment in humans in the immediate postnatal period, prior to the development of weakness or very early in the course of the disease, may be effective. Therefore, our objective was to establish a feasible method for SMA screening. High-resolution melting (HRM) analysis is rapidly becoming the most important mutation-scanning methodology that allows mutation scanning and genotyping without the need for costly labeled oligonucleotides. In the current study, we aim to develop a method for identifying the substitution of single nucleotide in SMN1 exon 7 (c.840C>T) by HRM analysis. Genomic DNA was extracted from peripheral blood samples and dried blood spots obtained from 30 patients with SMA and 30 normal individuals. All results were previously confirmed by denaturing high-performance liquid chromatography (DHPLC). In order to identify the substitution of single nucleotide in SMN1 exon 7 (c.840C>T) by HRM analysis, a primer set was used in HRM analysis. At first, we failed to identify the substitution of single nucleotide in SMN1 exon 7 (c.840C>T) by HRM analysis because the homozygous CC and homozygous TT cannot be distinguished by HRM analysis. Therefore, all samples were mixed with a known SMN1/SMN2 copy number (SMN1/SMN2=0:3), which we may call driver. This strategy is used to differentiate between homozygous CC and homozygous TT. After mixing with driver, the melting profile of homozygous CC becomes heteroduplex; however, the homozygous TT remains the same in the normalized and temperature-shifted difference plots. HRM analysis can be successfully applied to screen SMA via DNA obtained from whole blood and dried blood spots. We strongly believe that HRM analysis, a high-throughput method

  3. Metallurgy of high-silicon steel parts produced using Selective Laser Melting

    International Nuclear Information System (INIS)

    Garibaldi, Michele; Ashcroft, Ian; Simonelli, Marco; Hague, Richard

    2016-01-01

    The metallurgy of high-silicon steel (6.9%wt.Si) processed using Selective Laser Melting (SLM) is presented for the first time in this study. High-silicon steel has great potential as a soft magnetic alloy, but its employment has been limited due to its poor workability. The effect of SLM-processing on the metallurgy of the alloy is investigated in this work using microscopy, X-Ray Diffraction (XRD) and Electron Backscatter Diffraction (EBSD). XRD analysis suggests that the SLM high-silicon steel is a single ferritic phase (solid solution), with no sign of phase ordering. This is expected to have beneficial effects on the material properties, since ordering has been shown to make silicon steels more brittle and electrically conductive. For near-fully dense samples, columnar grains with a high aspect ratio and oriented along the build direction are found. Most importantly, a <001> fibre-texture along the build direction can be changed into a cube-texture when the qualitative shape of the melt-pool is altered (from shallow to deep) by increasing the energy input of the scanning laser. This feature could potentially open the path to the manufacture of three-dimensional grain-oriented high-silicon steels for electromechanical applications.

  4. extraction of high quality dna from polysaccharides-secreting ...

    African Journals Online (AJOL)

    cistvr

    A DNA extraction method using CTAB was used for the isolation of genomic DNA from ten. Xanthomonas campestris pathovars, ten isolates of Xanthomonas albilineans and one isolate of. Pseudomonas rubrisubalbicans. High quality DNA was obtained that was ideal for molecular analy- ses. Extracellular polysaccharides ...

  5. Meteorological conditions associated to high sublimation amounts in semiarid high-elevation Andes decrease the performance of empirical melt models

    Science.gov (United States)

    Ayala, Alvaro; Pellicciotti, Francesca; MacDonell, Shelley; McPhee, James; Burlando, Paolo

    2015-04-01

    Empirical melt (EM) models are often preferred to surface energy balance (SEB) models to calculate melt amounts of snow and ice in hydrological modelling of high-elevation catchments. The most common reasons to support this decision are that, in comparison to SEB models, EM models require lower levels of meteorological data, complexity and computational costs. However, EM models assume that melt can be characterized by means of a few index variables only, and their results strongly depend on the transferability in space and time of the calibrated empirical parameters. In addition, they are intrinsically limited in accounting for specific process components, the complexity of which cannot be easily reconciled with the empirical nature of the model. As an example of an EM model, in this study we use the Enhanced Temperature Index (ETI) model, which calculates melt amounts using air temperature and the shortwave radiation balance as index variables. We evaluate the performance of the ETI model on dry high-elevation sites where sublimation amounts - that are not explicitly accounted for the EM model - represent a relevant percentage of total ablation (1.1 to 8.7%). We analyse a data set of four Automatic Weather Stations (AWS), which were collected during the ablation season 2013-14, at elevations between 3466 and 4775 m asl, on the glaciers El Tapado, San Francisco, Bello and El Yeso, which are located in the semiarid Andes of central Chile. We complement our analysis using data from past studies in Juncal Norte Glacier (Chile) and Haut Glacier d'Arolla (Switzerland), during the ablation seasons 2008-09 and 2006, respectively. We use the results of a SEB model, applied to each study site, along the entire season, to calibrate the ETI model. The ETI model was not designed to calculate sublimation amounts, however, results show that their ability is low also to simulate melt amounts at sites where sublimation represents larger percentages of total ablation. In fact, we

  6. Experience in melting of high-quality chromium-nickel-molybdenum steel in oxygen converter

    Energy Technology Data Exchange (ETDEWEB)

    Kosoi, L F; Yaburov, S I; Shul' kin, M L; Vedernikov, G G; Bragin, E D; Filork' yan, B K

    1978-10-01

    Technology of melting high-quality medium-carbon constructional chromium-nickel-molybdenum steel has been developed and tested in 130-t converters. The technology envisages metal refinement in a casting laddle using synthetic lime-aluminous slag and argon blowing, as well as liquid ferroallys (master alloys) for steel deoxidation and alloying. Due to a smaller content of sulfur, phosphorus, arsenic and sulphide inclusions, and to a smaller grain size (N 11-12), the steel, produced according to this technology possesses higher plastic properties and impact strength than conventional open-hearth furnace metal after heat treatment for the same strength.

  7. Behavior of a corium jet in high pressure melt ejection from a reactor pressure vessel

    International Nuclear Information System (INIS)

    Frid, W.E.

    1986-01-01

    A model has been developed to calculate the expansion and fragmentation of a corium jet, due to the evolution of dissolved gas, during a postulated core meltdown accident. Parametric calculations have been performed for a PWR high pressure accident scenario. Jet breakup occurs within a few jet diameters from the RPV. The diameter of the fragmented jet at the level of the reactor cavity floor is predicted to be 40-130 times the discharge diameter. Particles generated by fragmentation of corium melt are predicted to be in the 30-150 μm size range

  8. Evaluation of Foaming Behavior of Glass Melts by High-Temperature Microscopy

    DEFF Research Database (Denmark)

    Petersen, Rasmus Rosenlund; König, Jakob; Yue, Yuanzheng

    2016-01-01

    Optical monitoring techniques can record in situ the size of glass samples during a dynamic heating process. This allowed us to study sintering and expansion rate of panel glass from cathode ray tube using MnO2 as foaming agent. We show the maximum expansion rate of glass melt foaming (in situ va...... such as type and concentration of foaming agent, glass composition and particle size to obtain foam glass with high porosity and closed pores. Using this approach we show that the foaming of bottle glass is preferentially conducted at a SiC concentration of 1‒4 wt%....

  9. Recent Advancements in DNA Damage-Transcription Crosstalk and High-Resolution Mapping of DNA Breaks.

    Science.gov (United States)

    Vitelli, Valerio; Galbiati, Alessandro; Iannelli, Fabio; Pessina, Fabio; Sharma, Sheetal; d'Adda di Fagagna, Fabrizio

    2017-08-31

    Until recently, DNA damage arising from physiological DNA metabolism was considered a detrimental by-product for cells. However, an increasing amount of evidence has shown that DNA damage could have a positive role in transcription activation. In particular, DNA damage has been detected in transcriptional elements following different stimuli. These physiological DNA breaks are thought to be instrumental for the correct expression of genomic loci through different mechanisms. In this regard, although a plethora of methods are available to precisely map transcribed regions and transcription start sites, commonly used techniques for mapping DNA breaks lack sufficient resolution and sensitivity to draw a robust correlation between DNA damage generation and transcription. Recently, however, several methods have been developed to map DNA damage at single-nucleotide resolution, thus providing a new set of tools to correlate DNA damage and transcription. Here, we review how DNA damage can positively regulate transcription initiation, the current techniques for mapping DNA breaks at high resolution, and how these techniques can benefit future studies of DNA damage and transcription.

  10. [Rapid detection of hot spot mutations of FGFR3 gene with PCR-high resolution melting assay].

    Science.gov (United States)

    Li, Shan; Wang, Han; Su, Hua; Gao, Jinsong; Zhao, Xiuli

    2017-08-10

    To identify the causative mutations in five individuals affected with dyschondroplasia and develop an efficient procedure for detecting hot spot mutations of the FGFR3 gene. Genomic DNA was extracted from peripheral blood samples with a standard phenol/chloroform method. PCR-Sanger sequencing was used to analyze the causative mutations in the five probands. PCR-high resolution melting (HRM) was developed to detect the identified mutations. A c.1138G>A mutation in exon 8 was found in 4 probands, while a c.1620C>G mutation was found in exon 11 of proband 5 whom had a mild phenotype. All patients were successfully distinguished from healthy controls with the PCR-HRM method. The results of HRM analysis were highly consistent with that of Sanger sequencing. The Gly380Arg and Asn540Lys are hot spot mutations of the FGFR3 gene among patients with ACH/HCH. PCR-HRM analysis is more efficient for detecting hot spot mutations of the FGFR3 gene.

  11. High-resolution melting-curve (HRM) analysis for C. meleagridis identification in stool samples.

    Science.gov (United States)

    Chelbi, Hanen; Essid, Rym; Jelassi, Refka; Bouzekri, Nesrine; Zidi, Ines; Ben Salah, Hamza; Mrad, Ilhem; Ben Sghaier, Ines; Abdelmalek, Rym; Aissa, Sameh; Bouratbine, Aida; Aoun, Karim

    2018-02-01

    Cryptosporidiosis represents a major public health problem. This infection, caused by a protozoan parasite of the genus Cryptosporidium, has been reported worldwide as a frequent cause of diarrhoea. In the immunocompetent host, the typical watery diarrhea can be self-limiting. However, it is severe and chronic, in the immunocompromised host and may cause death. Cryptosporidium spp. are coccidians, which complete their life cycle in both humans and animals. The two species C. hominis and C. parvum are the major cause of human infection. Compared to studies on C. hominis and C. parvum, only a few studies have developed methods to identify C. meleagridis. To develop a new real time PCR-coupled High resolution melting assay allowing the detection for C. meleagridis, in addition of the other dominant species (C. hominis and C. parvum). The polymorphic sequence on the dihydrofolate reductase gene (DHFR) of three species was sequenced to design primers pair and establish a sensitive real-time PCR coupled to a high-resolution melting-curve (HRM) analysis method, allowing the detection of Cryptosporidium sp. and discrimination between three prevalent species in Tunisia. We analyzed a collection of 42 archived human isolates of the three studied species. Real-time PCR coupled to HRM assay allowed detection of Cryptosporidium, using the new designed primers, and basing on melting profile, we can distinguish C. meleagridis species in addition to C. parvum and C. hominis. We developed a qPCR-HRM assay that allows Cryptosporidium genotyping. This method is sensitive and able to distinguish three Cryptosporidium species. Copyright © 2017. Published by Elsevier Ltd.

  12. High-resolution Melting Analysis for Gene Scanning of Adenomatous Polyposis Coli (APC) Gene With Oral Squamous Cell Carcinoma Samples.

    Science.gov (United States)

    Chang, Ya-Sian; Lin, Chien-Yu; Yang, Shu-Fen; Ho, Cheng Mao; Chang, Jan-Gowth

    2016-02-01

    There have been many different mutations reported for the large adenomatous polyposis coli (APC) tumor suppressor gene. APC mutations result in inactivation of APC tumor suppressor action, allowing the progression of tumorigenesis. The present study utilized a highly efficient method to identify APC mutations and investigated the association between the APC genetic variants Y486Y, A545A, T1493T, and D1822V and susceptibility to oral squamous cell carcinoma (OSCC). High-resolution melting (HRM) analysis was used to characterize APC mutations. Genomic DNA was extracted from 83 patient specimens of OSCC and 50 blood samples from healthy control subjects. The 14 exons and mutation cluster region of exon 15 were screened by HRM analysis. All mutations were confirmed by direct DNA sequencing. Three mutations and 4 single nucleotide polymorphisms (SNPs) were found in this study. The mutations were c.573T>C (Y191Y) in exon 5, c.1005A>G (L335L) in exon 9, and c.1488A>T (T496T) in exon 11. Two SNPs, c.4479G>A (T1493T) and c.5465A>T (D1822V), were located in exon 15, whereas c.1458T>C (Y486Y) and c.1635G>A (A545A) were located in exon 11 and 13, respectively. There was no observed association between OSCC risk and genotype for any of the 4 APC SNPs. The mutation of APC is rare in Taiwanese patients with OSCC. HRM analysis is a reliable, accurate, and fast screening method for APC mutations.

  13. Development of high melting point, environmentally friendly solders, using the calphad approach

    DEFF Research Database (Denmark)

    Chidambaram, Vivek; Hald, John; Hattel, Jesper Henri

    2008-01-01

    An attempt has been made using the CALPHAD approach via Thermo-Calc to explore the various possible chemical compositions that adhere to the melting criterion i.e. 270-350 degrees C, required to replace the traditionally used high lead content solders for first level packaging applications. Vario...... tension have also been considered. Special focus has been given to toxicity related issues since the main ideology of looking for an alternative to high lead containing solders is not related to technical issues but due to environmental concerns.......An attempt has been made using the CALPHAD approach via Thermo-Calc to explore the various possible chemical compositions that adhere to the melting criterion i.e. 270-350 degrees C, required to replace the traditionally used high lead content solders for first level packaging applications. Various...... of promising solder alloy candidates. The ternary combinations that satisfied the primary solidification requirement were scrutinized taking into account the commercial interests i.e. availability, cost-effectiveness, recyclability and toxicity issues. Technical issues like manufacturability and surface...

  14. Hydrate-melt electrolytes for high-energy-density aqueous batteries

    Science.gov (United States)

    Yamada, Yuki; Usui, Kenji; Sodeyama, Keitaro; Ko, Seongjae; Tateyama, Yoshitaka; Yamada, Atsuo

    2016-10-01

    Aqueous Li-ion batteries are attracting increasing attention because they are potentially low in cost, safe and environmentally friendly. However, their low energy density (water and the limited selection of suitable negative electrodes, is problematic for their future widespread application. Here, we explore optimized eutectic systems of several organic Li salts and show that a room-temperature hydrate melt of Li salts can be used as a stable aqueous electrolyte in which all water molecules participate in Li+ hydration shells while retaining fluidity. This hydrate-melt electrolyte enables a reversible reaction at a commercial Li4Ti5O12 negative electrode with a low reaction potential (1.55 V versus Li+/Li) and a high capacity (175 mAh g-1). The resultant aqueous Li-ion batteries with high energy density (>130 Wh kg-1) and high voltage (˜2.3-3.1 V) represent significant progress towards performance comparable to that of commercial non-aqueous batteries (with energy densities of ˜150-400 Wh kg-1 and voltages of ˜2.4-3.8 V).

  15. Lambda-Cyhalothrin Nanosuspension Prepared by the Melt Emulsification-High Pressure Homogenization Method

    Directory of Open Access Journals (Sweden)

    Zhenzhong Pan

    2015-01-01

    Full Text Available The nanosuspension of 5% lambda-cyhalothrin with 0.2% surfactants was prepared by the melt emulsification-high pressure homogenization method. The surfactants composition, content, and homogenization process were optimized. The anionic surfactant (1-dodecanesulfonic acid sodium salt and polymeric surfactant (maleic rosin-polyoxypropylene-polyoxyethylene ether sulfonate screened from 12 types of commercially common-used surfactants were used to prepare lambda-cyhalothrin nanosuspension with high dispersity and stability. The mean particle size and polydispersity index of the nanosuspension were 16.01 ± 0.11 nm and 0.266 ± 0.002, respectively. The high zeta potential value of −41.7 ± 1.3 mV and stable crystalline state of the nanoparticles indicated the excellent physical and chemical stability. The method could be widely used for preparing nanosuspension of various pesticides with melting points below boiling point of water. This formulation may avoid the use of organic solvents and reduce surfactants and is perspective for improving bioavailability and reducing residual pollution of pesticide in agricultural products and environment.

  16. High-resolution melting (HRM) re-analysis of a polyposis patients cohort reveals previously undetected heterozygous and mosaic APC gene mutations.

    Science.gov (United States)

    Out, Astrid A; van Minderhout, Ivonne J H M; van der Stoep, Nienke; van Bommel, Lysette S R; Kluijt, Irma; Aalfs, Cora; Voorendt, Marsha; Vossen, Rolf H A M; Nielsen, Maartje; Vasen, Hans F A; Morreau, Hans; Devilee, Peter; Tops, Carli M J; Hes, Frederik J

    2015-06-01

    Familial adenomatous polyposis is most frequently caused by pathogenic variants in either the APC gene or the MUTYH gene. The detection rate of pathogenic variants depends on the severity of the phenotype and sensitivity of the screening method, including sensitivity for mosaic variants. For 171 patients with multiple colorectal polyps without previously detectable pathogenic variant, APC was reanalyzed in leukocyte DNA by one uniform technique: high-resolution melting (HRM) analysis. Serial dilution of heterozygous DNA resulted in a lowest detectable allelic fraction of 6% for the majority of variants. HRM analysis and subsequent sequencing detected pathogenic fully heterozygous APC variants in 10 (6%) of the patients and pathogenic mosaic variants in 2 (1%). All these variants were previously missed by various conventional scanning methods. In parallel, HRM APC scanning was applied to DNA isolated from polyp tissue of two additional patients with apparently sporadic polyposis and without detectable pathogenic APC variant in leukocyte DNA. In both patients a pathogenic mosaic APC variant was present in multiple polyps. The detection of pathogenic APC variants in 7% of the patients, including mosaics, illustrates the usefulness of a complete APC gene reanalysis of previously tested patients, by a supplementary scanning method. HRM is a sensitive and fast pre-screening method for reliable detection of heterozygous and mosaic variants, which can be applied to leukocyte and polyp derived DNA.

  17. Bounding analysis of containment of high pressure melt ejection in advanced light water reactors

    International Nuclear Information System (INIS)

    Additon, S.L.; Fontana, M.H.; Carter, J.C.

    1990-01-01

    This paper reports on the loadings on containment due to direct containment heating (DCH) as a result of high pressure melt ejection (HPME) in advanced light water reactors (ALWR) which were estimated using conservative, bounding analyses. The purpose of the analyses was to scope the magnitude of the possible loadings and to indicate the performance needed from potential mitigation methods, such as a cavity configuration that limits energy transfer to the upper containment volume. Analyses were performed for three cases which examined the effect of availability of high pressure reactor coolant system water at the time of reactor vessel melt through and the effect of preflooding of the reactor cavity. The amount of core ejected from the vessel was varied from 100% to 0% for all cases. Results indicate that all amounts of core debris dispersal could be accommodated by the containment for the case where the reactor cavity was preflooded. For the worst case, all the energy from in-vessel hydrogen generation and combustion plus that from 45% of the entire molten core would be required to equilibrate with the containment upper volume in order to reach containment failure pressure

  18. The Kinetics of Phase Transformations During Tempering in Laser Melted High Chromium Cast Steel

    Science.gov (United States)

    Li, M. Y.; Wang, Y.; Han, B.

    2012-06-01

    The precipitation of secondary carbides in the laser melted high chromium cast steels during tempering at 300-650 °C for 2 h in air furnace was characterized and the present phases was identified, by using transmission electron microscopy. Laser melted high chromium cast steel consists of austenitic dendrites and interdendritic M23C6 carbides. The austenite has such a strong tempering stability that it remains unchanged at temperature below 400 °C and the secondary hardening phenomenon starts from 450 °C to the maximum value of 672 HV at 560 °C. After tempering at 450 °C fine M23C6 carbides precipitate from the supersaturated austenite preferentially. In addition, the dislocation lines and slip bands still exist inside the austenite. While tempering at temperature below 560 °C, the secondary hardening simultaneously results from the martensite phase transformation and the precipitation of carbides as well as dislocation strengthening within a refined microstructure. Moreover, the formation of the ferrite matrix and large quality of coarse lamellar M3C carbides when the samples were tempered at 650 °C contributes to the decrease of hardness.

  19. Differentiation of five enterohepatic Helicobacter species by nested PCR with high-resolution melting curve analysis.

    Science.gov (United States)

    Wu, Miaoli; Rao, Dan; Zhu, Yujun; Wang, Jing; Yuan, Wen; Zhang, Yu; Huang, Ren; Guo, Pengju

    2017-04-01

    Enterohepatic Helicobacter species (EHS) are widespread in rodent species around the world. Several studies have demonstrated that infection with EHS can interfere with the outcomes of animal experiments in cancer research and significantly influence the study results. Therefore, it is essential to establish a rapid detection and identification of EHS for biomedical research using laboratory rodents. Our study aimed to develop a rapid and sensitive method to detect and distinguish five enterohepatic Helicobacter species. Nested PCR followed by high-resolution melting curve analysis (HRM) was developed for identification of H. bilis, H. rodentium, H. muridarum, H. typhlonius, as well as H. hepaticus. To validate the accuracy of nested PCR-HRM analysis, quantitative real-time PCR methods for five different enterohepatic Helicobacter species were developed. A total of 50 cecal samples were tested using both nested PCR-HRM analysis and qPCR method. The nested PCR-HRM method could distinguish five enterohepatic Helicobacter species by different melting temperatures. The melting curve were characterized by peaks of 78.7 ± 0.12°C for H. rodentium, 80.51 ± 0.09°C for H. bilis, 81.6 ± 0.1°C for H. typhlonius, 82.11 ± 0.18°C for H. muridarum, and 82.95 ± 0.09°C for H. hepaticus. The nested PCR-HRM assay is a simple, rapid, and cost-effective assay. This assay could be a useful tool for molecular epidemiology study of enterohepatic Helicobacter infection and an attractive alternative for genotyping of enterohepatic Helicobacter species. © 2016 John Wiley & Sons Ltd.

  20. Theoretical study of melting curves on Ta, Mo, and W at high pressures

    Energy Technology Data Exchange (ETDEWEB)

    Xi Feng [Laboratory for Shock Wave and Detonation Physics Research, Institute of Fluid Physics, P.O. Box 919-102, 621900 Mianyang (China)], E-mail: hawk_0816@yahoo.com.cn; Cai Lingcang [Laboratory for Shock Wave and Detonation Physics Research, Institute of Fluid Physics, P.O. Box 919-102, 621900 Mianyang (China)

    2008-06-01

    The melting curves of tantalum (Ta), molybdenum (Mo), and tungsten (W) are calculated using a dislocation-mediated melting model. The calculated melting curves are in good agreement with shock-wave data, and partially in agreement with wire explosion and piston-cylinder data, but show large discrepancies with diamond-anvil cell (DAC) data. We propose that the melting mechanism caused by shock-wave and laser-heated DAC techniques are probably different, and that a systematic difference exists in the two melting processes.

  1. High Resolution Melting Analysis for Detecting p53 Gene Mutations in Patients with Non-small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Zhihong CHEN

    2011-10-01

    Full Text Available Background and objective It has been proven that p53 gene was related to many human cancers. The mutations in p53 gene play an important role in carcinogensis and mostly happened in exon 5-8. The aim of this study is to establish a high resolution melting (HRM assay to detect p53 mutations from patients with non-small cell lung cancer (NSCLC, to investigate the characteristics of p53 gene mutations, and to analyze the relationship between p53 mutations and evolution regularity of pathogenesis. Methods p53 mutations in exon 5-8 were detected by HRM assay on DNA insolated from 264 NSCLC samples derived from tumor tissues and 54 control samples from pericancerous pulmonary tissues. The mutation samples by the HRM assay were confirmed by sequencing technique. Samples which were positive by HRM but wild type by sequencing were further confirmed by sub-clone and sequencing. Results No mutation was found in 54 pericancerous pulmonary samples by HRM assay. 104 of the 264 tumor tissues demonstrated mutation curves by HRM assay, 102 samples were confirmed by sequencing, including 95 point mutations and 7 frame shift mutations by insertion or deletion. The mutation rate of p53 gene was 39.4%. The mutation rate from exon 5-8 were 11.7%, 8%, 12.5% and 10.6%, respectively and there was no statistically significant difference between them (P=0.35. p53 mutations were significantly more frequent in males than that in females, but not related to the other clinicopathologic characteristics. Conclusion The results indicate that HRM is a sensitive in-tube methodology to detect for mutations in clinical samples. The results suggest that the arising p53 mutations in NSCLC may be due to spontaneous error in DNA synthesis and repair.

  2. Parametric studies on containment thermal hydraulic loads during high pressure melt ejection in a BWR

    Energy Technology Data Exchange (ETDEWEB)

    Silde, A.; Lindholm, I. [VTT Energy, Espoo (Finland)

    1997-12-01

    The containment thermal hydraulic loads during high pressure melt ejection in a Nordic BWR are studied parametrically with the CONTAIN and the MELCOR codes. The work is part of the Nordic RAK-2 project. The containment analyses were divided into two categories according to composition of the discharged debris: metallic and oxidic debris cases. In the base case with highly metallic debris, all sources from the reactor coolant system to the containment were based on the MELCOR/BH calculation. In the base case with the oxidic debris, the source data was specified assuming that {approx} 15% of the whole core material inventory and 34,000 kg of saturated water was discharged from the reactor pressure vessel (RPV) during 30 seconds. In this case, the debris consisted mostly of oxides. The highest predicted containment pressure peaks were about 8.5 bar. In the scenarios with highly metallic debris source, very high gas temperature of about 1900 K was predicted in the pedestal, and about 1400 K in the upper drywell. The calculations with metallic debris were sensititive to model parameters, like the particle size and the parameters, which control the chemical reaction kinetics. In the scenarios with oxidic debris source, the predicted pressure peaks were comparable to the cases with the metallic debris source. The maximum gas temperatures (about 450-500 K) in the containment were, however, significantly lower than in the respective metallic debris case. The temperatures were also insensitive to parametric variations. In addition, one analysis was performed with the MELCOR code for benchmarking of the MELCOR capabilities against the more detailed CONTAIN code. The calculations showed that leak tightness of the containment penetrations could be jeopardized due to high temperature loads, if a high pressure melt ejection occurred during a severe accident. Another consequence would be an early containment venting. (au). 28 refs.

  3. The potential of high resolution melting analysis (hrma) to streamline, facilitate and enrich routine diagnostics in medical microbiology.

    Science.gov (United States)

    Ruskova, Lenka; Raclavsky, Vladislav

    2011-09-01

    Routine medical microbiology diagnostics relies on conventional cultivation followed by phenotypic techniques for identification of pathogenic bacteria and fungi. This is not only due to tradition and economy but also because it provides pure culture needed for antibiotic susceptibility testing. This review focuses on the potential of High Resolution Melting Analysis (HRMA) of double-stranded DNA for future routine medical microbiology. Search of MEDLINE database for publications showing the advantages of HRMA in routine medical microbiology for identification, strain typing and further characterization of pathogenic bacteria and fungi in particular. The results show increasing numbers of newly-developed and more tailor-made assays in this field. For microbiologists unfamiliar with technical aspects of HRMA, we also provide insight into the technique from the perspective of microbial characterization. We can anticipate that the routine availability of HRMA in medical microbiology laboratories will provide a strong stimulus to this field. This is already envisioned by the growing number of medical microbiology applications published recently. The speed, power, convenience and cost effectiveness of this technology virtually predestine that it will advance genetic characterization of microbes and streamline, facilitate and enrich diagnostics in routine medical microbiology without interfering with the proven advantages of conventional cultivation.

  4. Structural Investigation of Fe-Ni-S and Fe-Ni-Si Melts by High-temperature Fluorescence XAFS Measurements

    International Nuclear Information System (INIS)

    Manghnani, Murli H.; Balogh, John; Hong Xinguo; Newville, Matthew; Amulele, G.

    2007-01-01

    Iron-nickel (Fe-Ni) alloy is regarded as the most abundant constituent of Earth's core, with an amount of 5.5 wt% Ni in the core based on geochemical and cosmochemical models. The structural role of nickel in liquid Fe-Ni alloys with light elements such as S or Si is poorly understood, largely because of the experimental difficulties of high-temperature melts. Recently, we have succeeded in acquiring Ni K-edge fluorescence x-ray absorption fine structure (XAFS) spectra of Fe-Ni-S and Fe-Ni-Si melts and alloys. Different structural environment of Ni atoms in Fe-Ni-S and Fe-Ni-Si melts is observed, supporting the effect of light elements in Fe-Ni melts

  5. Applying high-resolution melting (HRM) technology to identify five commonly used Artemisia species.

    Science.gov (United States)

    Song, Ming; Li, Jingjian; Xiong, Chao; Liu, Hexia; Liang, Junsong

    2016-10-04

    Many members of the genus Artemisia are important for medicinal purposes with multiple pharmacological properties. Often, these herbal plants sold on the markets are in processed forms so it is difficult to authenticate. Routine testing and identification of these herbal materials should be performed to ensure that the raw materials used in pharmaceutical products are suitable for their intended use. In this study, five commonly used Artemisia species included Artemisia argyi, Artemisia annua, Artemisia lavandulaefolia, Artemisia indica, and Artemisia atrovirens were analyzed using high resolution melting (HRM) analysis based on the internal transcribed spacer 2 (ITS2) sequences. The melting profiles of the ITS2 amplicons of the five closely related herbal species are clearly separated so that they can be differentiated by HRM method. The method was further applied to authenticate commercial products in powdered. HRM curves of all the commercial samples tested are similar to the botanical species as labeled. These congeneric medicinal products were also clearly separated using the neighbor-joining (NJ) tree. Therefore, HRM method could provide an efficient and reliable authentication system to distinguish these commonly used Artemisia herbal products on the markets and offer a technical reference for medicines quality control in the drug supply chain.

  6. Fluid–fluid–solid triple point on melting curves at high temperatures

    International Nuclear Information System (INIS)

    Norman, G E; Saitov, I M

    2016-01-01

    An analysis is presented of experimental data where fluid-fluid phase transitions are observed for different substances at high temperatures with triple points on melting curves. Viscosity drops point to the structural character of the transition, whereas conductivity jumps remind of both semiconductor-to-metal and plasma nature. The slope of the phase equilibrium dependencies of pressure on temperature and the consequent change of the specific volume, which follows from the Clapeyron-Clausius equation, are discussed. P(V, T) surfaces are presented and discussed for the phase transitions considered in the vicinity of the triple points. The cases of abnormal P(T) dependencies on curves of phase equilibrium are in the focus of discussion. In particular, a P(V, T) surface is presented when both fluid-fluid and melting P(T) curves are abnormal. Particular attention is paid to warm dense hydrogen and deuterium, where remarkable contradictions exist between data of different authors. The possible connection of the P(V, T) surface peculiarities with the experimental data uncertainties is outlined. (paper)

  7. Methods of vitrifying waste with low melting high lithia glass compositions

    Science.gov (United States)

    Jantzen, Carol M.; Pickett, John B.; Cicero-Herman, Connie A.; Marra, James C.

    2001-01-01

    The invention relates to methods of vitrifying waste and for lowering the melting point of glass forming systems by including lithia formers in the glass forming composition in significant amounts, typically from about 0.16 wt % to about 11 wt %, based on the total glass forming oxides. The lithia is typically included as a replacement for alkali oxide glass formers that would normally be present in a particular glass forming system. Replacement can occur on a mole percent or weight percent basis, and typically results in a composition wherein lithia forms about 10 wt % to about 100 wt % of the alkali oxide glass formers present in the composition. The present invention also relates to the high lithia glass compositions formed by these methods. The invention is useful for stabilization of numerous types of waste materials, including aqueous waste streams, sludge solids, mixtures of aqueous supernate and sludge solids, combinations of spent filter aids from waste water treatment and waste sludges, supernate alone, incinerator ash, incinerator offgas blowdown, or combinations thereof, geological mine tailings and sludges, asbestos, inorganic filter media, cement waste forms in need of remediation, spent or partially spent ion exchange resins or zeolites, contaminated soils, lead paint, etc. The decrease in melting point achieved by the present invention desirably prevents volatilization of hazardous or radioactive species during vitrification.

  8. The spa typing of methicillin-resistant Staphylococcus aureus isolates by High Resolution Melting (HRM) analysis.

    Science.gov (United States)

    Fasihi, Yasser; Fooladi, Saba; Mohammadi, Mohammad Ali; Emaneini, Mohammad; Kalantar-Neyestanaki, Davood

    2017-09-06

    Molecular typing is an important tool for control and prevention of infection. A suitable molecular typing method for epidemiological investigation must be easy to perform, highly reproducible, inexpensive, rapid and easy to interpret. In this study, two molecular typing methods including the conventional PCR-sequencing method and high resolution melting (HRM) analysis were used for staphylococcal protein A (spa) typing of 30 Methicillin-resistant Staphylococcus aureus (MRSA) isolates recovered from clinical samples. Based on PCR-sequencing method results, 16 different spa types were identified among the 30 MRSA isolates. Among the 16 different spa types, 14 spa types separated by HRM method. Two spa types including t4718 and t2894 were not separated from each other. According to our results, spa typing based on HRM analysis method is very rapid, easy to perform and cost-effective, but this method must be standardized for different regions, spa types, and real-time machinery.

  9. High thermoelectric performances of Bi–AE–Co–O compounds directionally growth from the melt

    International Nuclear Information System (INIS)

    Diez, J.C.; Rasekh, S.; Madre, M.A.; Torres, M.A.; Sotelo, A.E.

    2018-01-01

    Bi2AE2Co2Ox (AE=Ca, Sr, and Ba) thermoelectric compounds were grown from the melt by the laser floating zone technique. Microstructural analysis of as-grown samples has shown the formation of well-aligned thermoelectric grains together with a relative high amount of secondary phases. On the other hand, a short (24h) thermal treatment (810°C for Sr, 800°C for Ca, and 750°C for Ba) under air, raises of thermoelectric phase content through the recombination of the secondary ones. These microstructural modifications led to a large decrease of electrical resistivity, improving the power factor. These results have been compared with samples prepared by the conventional solid state method and with the best values reported in the literature. From these data, it is possible to deduce that the high thermoelectric characteristics obtained in these samples make them very attractive for practical applications. [es

  10. High-resolution melting analysis for detection of a single-nucleotide polymorphism and the genotype of the myostatin gene in warmblood horses.

    Science.gov (United States)

    Serpa, Priscila B S; Garbade, Petra; Natalini, Cláudio C; Pires, Ananda R; Tisotti, Tainor M

    2017-01-01

    OBJECTIVE To develop a high-resolution melting (HRM) assay to detect the g.66493737C>T polymorphism in the myostatin gene (MSTN) and determine the frequency of 3 previously defined g.66493737 genotypes (T/T, T/C, and C/C) in warmblood horses. SAMPLES Blood samples from 23 horses. PROCEDURES From each blood sample, DNA was extracted and analyzed by standard PCR methods and an HRM assay to determine the MSTN genotype. Three protocols (standard protocol, protocol in which a high-salt solution was added to the reaction mixture before the first melting cycle, and protocol in which an unlabeled probe was added to the reaction mixture before analysis) for the HRM assay were designed and compared. Genotype results determined by the HRM protocol that generated the most consistent melting curves were compared with those determined by sequencing. RESULTS The HRM protocol in which an unlabeled probe was added to the reaction mixture generated the most consistent melting curves. The genotypes of the g.66493737C>T polymorphism were determined for 22 horses (16 by HRM analysis and 20 by sequencing); 14, 7, and 1 had the T/T, T/C, and C/C genotypes, respectively. The genotype determined by HRM analysis agreed with that determined by sequencing for 14 of 16 horses. The frequency of alleles T and C was 79.5% and 20.5%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that HRM analysis may be a faster and more economical alternative than PCR methods for genotyping. Genotyping results might be useful as predictors of athletic performance for horses.

  11. Excised radicle tips as a source of genomic DNA for PCR-based ...

    Indian Academy of Sciences (India)

    2012-12-13

    Dec 13, 2012 ... Cotton; cry1Ac; genomic DNA isolation; high-resolution melting curve analysis; radicle tip; seed purity testing .... cooled to 40°C. Fluorescence data for melting curves were ... greatly increased by introducing automation.

  12. Detecting differential allelic expression using high-resolution melting curve analysis: application to the breast cancer susceptibility gene CHEK2

    Directory of Open Access Journals (Sweden)

    Sinilnikova Olga

    2011-05-01

    Full Text Available Abstract Background The gene CHEK2 encodes a checkpoint kinase playing a key role in the DNA damage pathway. Though CHEK2 has been identified as an intermediate breast cancer susceptibility gene, only a small proportion of high-risk families have been explained by genetic variants located in its coding region. Alteration in gene expression regulation provides a potential mechanism for generating disease susceptibility. The detection of differential allelic expression (DAE represents a sensitive assay to direct the search for a functional sequence variant within the transcriptional regulatory elements of a candidate gene. We aimed to assess whether CHEK2 was subject to DAE in lymphoblastoid cell lines (LCLs from high-risk breast cancer patients for whom no mutation in BRCA1 or BRCA2 had been identified. Methods We implemented an assay based on high-resolution melting (HRM curve analysis and developed an analysis tool for DAE assessment. Results We observed allelic expression imbalance in 4 of the 41 LCLs examined. All four were carriers of the truncating mutation 1100delC. We confirmed previous findings that this mutation induces non-sense mediated mRNA decay. In our series, we ruled out the possibility of a functional sequence variant located in the promoter region or in a regulatory element of CHEK2 that would lead to DAE in the transcriptional regulatory milieu of freely proliferating LCLs. Conclusions Our results support that HRM is a sensitive and accurate method for DAE assessment. This approach would be of great interest for high-throughput mutation screening projects aiming to identify genes carrying functional regulatory polymorphisms.

  13. High-resolution melting genotyping of Enterococcus faecium based on multilocus sequence typing derived single nucleotide polymorphisms.

    Directory of Open Access Journals (Sweden)

    Steven Y C Tong

    Full Text Available We have developed a single nucleotide polymorphism (SNP nucleated high-resolution melting (HRM technique to genotype Enterococcus faecium. Eight SNPs were derived from the E. faecium multilocus sequence typing (MLST database and amplified fragments containing these SNPs were interrogated by HRM. We tested the HRM genotyping scheme on 85 E. faecium bloodstream isolates and compared the results with MLST, pulsed-field gel electrophoresis (PFGE and an allele specific real-time PCR (AS kinetic PCR SNP typing method. In silico analysis based on predicted HRM curves according to the G+C content of each fragment for all 567 sequence types (STs in the MLST database together with empiric data from the 85 isolates demonstrated that HRM analysis resolves E. faecium into 231 "melting types" (MelTs and provides a Simpson's Index of Diversity (D of 0.991 with respect to MLST. This is a significant improvement on the AS kinetic PCR SNP typing scheme that resolves 61 SNP types with D of 0.95. The MelTs were concordant with the known ST of the isolates. For the 85 isolates, there were 13 PFGE patterns, 17 STs, 14 MelTs and eight SNP types. There was excellent concordance between PFGE, MLST and MelTs with Adjusted Rand Indices of PFGE to MelT 0.936 and ST to MelT 0.973. In conclusion, this HRM based method appears rapid and reproducible. The results are concordant with MLST and the MLST based population structure.

  14. High pressure and temperature structure of liquid and solid Cd: implications for the melting curve of Cd

    International Nuclear Information System (INIS)

    Raju, S V; Williams, Q; Geballe, Z M; Godwal, B K; Jeanloz, R; Kalkan, B

    2014-01-01

    The structure of cadmium was characterized in both the solid and liquid forms at pressures to 10 GPa using in situ x-ray diffraction measurements in a resistively heated diamond anvil cell. The distorted hexagonal structure of solid cadmium persists at high pressures and temperatures, with anomalously large c/a ratio of Cd becoming larger as the melting curve is approached. The measured structure factor S(Q) for the melt reveals that the cadmium atoms are spaced about 0.6 Angstroms apart. The melt structure remains notably constant with increasing pressure, with the first peak in the structure factor remaining mildly asymmetric, in accord with the persistence of an anisotropic bonding environment within the liquid. Evolution of powder diffraction patterns up to the temperature of melting revealed the stability of the ambient-pressure hcp structure up to a pressure of 10 GPa. The melting curve has a positive Clausius–Clapeyron slope, and its slope is in good agreement with data from other techniques. We find deviations in the melting curve from Lindemann law type behavior for pressures above 1 GPa. (paper)

  15. Greenland iceberg melt variability from high-resolution satellite observations

    Directory of Open Access Journals (Sweden)

    E. M. Enderlin

    2018-02-01

    Full Text Available Iceberg discharge from the Greenland Ice Sheet accounts for up to half of the freshwater flux to surrounding fjords and ocean basins, yet the spatial distribution of iceberg meltwater fluxes is poorly understood. One of the primary limitations for mapping iceberg meltwater fluxes, and changes over time, is the dearth of iceberg submarine melt rate estimates. Here we use a remote sensing approach to estimate submarine melt rates during 2011–2016 for 637 icebergs discharged from seven marine-terminating glaciers fringing the Greenland Ice Sheet. We find that spatial variations in iceberg melt rates generally follow expected patterns based on hydrographic observations, including a decrease in melt rate with latitude and an increase in melt rate with iceberg draft. However, we find no longitudinal variations in melt rates within individual fjords. We do not resolve coherent seasonal to interannual patterns in melt rates across all study sites, though we attribute a 4-fold melt rate increase from March to April 2011 near Jakobshavn Isbræ to fjord circulation changes induced by the seasonal onset of iceberg calving. Overall, our results suggest that remotely sensed iceberg melt rates can be used to characterize spatial and temporal variations in oceanic forcing near often inaccessible marine-terminating glaciers.

  16. Taxonomic Identification of Mediterranean Pines and Their Hybrids Based on the High Resolution Melting (HRM) and trnL Approaches: From Cytoplasmic Inheritance to Timber Tracing

    Science.gov (United States)

    Ganopoulos, Ioannis; Aravanopoulos, Filippos; Madesis, Panagiotis; Pasentsis, Konstantinos; Bosmali, Irene; Ouzounis, Christos; Tsaftaris, Athanasios

    2013-01-01

    Fast and accurate detection of plant species and their hybrids using molecular tools will facilitate the assessment and monitoring of local biodiversity in an era of climate and environmental change. Herein, we evaluate the utility of the plastid trnL marker for species identification applied to Mediterranean pines (Pinus spp.). Our results indicate that trnL is a very sensitive marker for delimiting species biodiversity. Furthermore, High Resolution Melting (HRM) analysis was exploited as a molecular fingerprint for fast and accurate discrimination of Pinus spp. DNA sequence variants. The trnL approach and the HRM analyses were extended to wood samples of two species (Pinus nigra and Pinus sylvestris) with excellent results, congruent to those obtained using leaf tissue. Both analyses demonstrate that hybrids from the P. brutia (maternal parent) × P. halepensis (paternal parent) cross, exhibit the P. halepensis profile, confirming paternal plastid inheritance in Group Halepensis pines. Our study indicates that a single one-step reaction method and DNA marker are sufficient for the identification of Mediterranean pines, their hybrids and the origin of pine wood. Furthermore, our results underline the potential for certain DNA regions to be used as novel biological information markers combined with existing morphological characters and suggest a relatively reliable and open taxonomic system that can link DNA variation to phenotype-based species or hybrid assignment status and direct taxa identification from recalcitrant tissues such as wood samples. PMID:23577179

  17. Molecular Dynamics Simulation of High Density DNA Arrays

    Directory of Open Access Journals (Sweden)

    Rudolf Podgornik

    2018-01-01

    Full Text Available Densely packed DNA arrays exhibit hexagonal and orthorhombic local packings, as well as a weakly first order transition between them. While we have some understanding of the interactions between DNA molecules in aqueous ionic solutions, the structural details of its ordered phases and the mechanism governing the respective phase transitions between them remains less well understood. Since at high DNA densities, i.e., small interaxial spacings, one can neither neglect the atomic details of the interacting macromolecular surfaces nor the atomic details of the intervening ionic solution, the atomistic resolution is a sine qua non to properly describe and analyze the interactions between DNA molecules. In fact, in order to properly understand the details of the observed osmotic equation of state, one needs to implement multiple levels of organization, spanning the range from the molecular order of DNA itself, the possible ordering of counterions, and then all the way to the induced molecular ordering of the aqueous solvent, all coupled together by electrostatic, steric, thermal and direct hydrogen-bonding interactions. Multiscale simulations therefore appear as singularly suited to connect the microscopic details of this system with its macroscopic thermodynamic behavior. We review the details of the simulation of dense atomistically resolved DNA arrays with different packing symmetries and the ensuing osmotic equation of state obtained by enclosing a DNA array in a monovalent salt and multivalent (spermidine counterions within a solvent permeable membrane, mimicking the behavior of DNA arrays subjected to external osmotic stress. By varying the DNA density, the local packing symmetry, and the counterion type, we are able to analyze the osmotic equation of state together with the full structural characterization of the DNA subphase, the counterion distribution and the solvent structural order in terms of its different order parameters and

  18. Recent progress in the melt-process technique of high-temperature superconductors

    CERN Document Server

    Ikuta, H; Mizutani, U

    1999-01-01

    Recently, the performance of high-temperature super conductors prepared by the melt-process technique has been greatly improved. This progress was accomplished by the addition of Ag into the starting materials of the Sm-Ba-CuO $9 system, which prevents the formation of severe macro-sized cracks in the finished samples. The magnetic flux density trapped by this material has now reached 9 T at 25 K, which is comparable to the magnetic flux density produced by $9 ordinary superconducting magnets. The amount of magnetic flux density that can be trapped by the sample is limited by the mechanical strength rather than superconducting properties of the material. The increase in the mechanical $9 strength of the material is important both for further improvement of the material properties and for ensuring reliability of the material in practical applications. (20 refs).

  19. Genotyping of Listeria monocytogenes isolates from poultry carcasses using high resolution melting (HRM) analysis.

    Science.gov (United States)

    Sakaridis, Ioannis; Ganopoulos, Ioannis; Madesis, Panagiotis; Tsaftaris, Athanasios; Argiriou, Anagnostis

    2014-01-02

    An outbreak situation of human listeriosis requires a fast and accurate protocol for typing Listeria monocytogenes . Existing techniques are either characterized by low discriminatory power or are laborious and require several days to give a final result. Polymerase chain reaction (PCR) coupled with high resolution melting (HRM) analysis was investigated in this study as an alternative tool for a rapid and precise genotyping of L. monocytogenes isolates. Fifty-five isolates of L. monocytogenes isolated from poultry carcasses and the environment of four slaughterhouses were typed by HRM analysis using two specific markers, internalin B and ssrA genes. The analysis of genotype confidence percentage of L. monocytogenes isolates produced by HRM analysis generated dendrograms with two major groups and several subgroups. Furthermore, the analysis of the HRM curves revealed that all L. monocytogenes isolates could easily be distinguished. In conclusion, HRM was proven to be a fast and powerful tool for genotyping isolates of L. monocytogenes .

  20. Forming of High-strength Steels Using a Hot-melt Dry Lubricant

    DEFF Research Database (Denmark)

    Hörnström, Sven-Erik; Karlsson, Erik; Olsson, Mikael

    2008-01-01

    during forming resulting in seizure of the tool/steel sheet contact and extensive scratching of the steel sheet surface. As a result, a number of concepts have been developed in order to reduce the tendency to galling in metal forming, including the development of new dry lubricants, new forming tool...... steel grades and improved surface engineering treatments such as the deposition of low friction CVD and PVD coatings. In the present study the performance of a hot-melt dry lubricant in the forming of hot and cold rolled and hot-dip galvanized high strength steel has been evaluated and compared...... with a conventional rust protection oil using four different tests methods, i.e. a strip reduction test, a bending under tension test, a stretch-forming test and a pin-on disc test. In the tests, two different cold work tool steels, a conventional steel grade and a nitrogen alloyed PM steel grade were evaluated...

  1. High-gravity combustion synthesis and in situ melt infiltration: A new method for preparing cemented carbides

    International Nuclear Information System (INIS)

    Liu, Guanghua; Li, Jiangtao; Yang, Zengchao; Guo, Shibin; Chen, Yixiang

    2013-01-01

    A new method of high-gravity combustion synthesis and in situ melt infiltration is reported for preparing cemented carbides, where hot nickel melt is in situ synthesized from a highly exothermic combustion reaction and then infiltrated into tungsten carbide powder compacts. The as-prepared sample showed a homogeneous microstructure, and its relative density, hardness and flexural strength were 94.4%, 84 HRA and 1.49 GPa, respectively. Compared with conventional powder metallurgy approaches, high-gravity combustion synthesis offers a fast and furnace-free way to produce cemented carbides

  2. Research on High Layer Thickness Fabricated of 316L by Selective Laser Melting

    Directory of Open Access Journals (Sweden)

    Shuo Wang

    2017-09-01

    Full Text Available Selective laser melting (SLM is a potential additive manufacturing (AM technology. However, the application of SLM was confined due to low efficiency. To improve efficiency, SLM fabrication with a high layer thickness and fine powder was systematically researched, and the void areas and hollow powders can be reduced by using fine powder. Single-track experiments were used to narrow down process parameter windows. Multi-layer fabrication relative density can be reached 99.99% at the exposure time-point distance-hatch space of 120 μs-40 μm-240 μm. Also, the building rate can be up to 12 mm3/s, which is about 3–10 times higher than the previous studies. Three typical defects were found by studying deeply, including the un-melted defect between the molten pools, the micro-pore defect within the molten pool, and the irregular distribution of the splashing phenomenon. Moreover, the microstructure is mostly equiaxed crystals and a small amount of columnar crystals. The averages of ultimate tensile strength, yield strength, and elongation are 625 MPa, 525 MPa, and 39.9%, respectively. As exposure time increased from 80 μs to 200 μs, the grain size is gradually grown up from 0.98 μm to 2.23 μm, the grain aspect ratio is close to 1, and the tensile properties are shown as a downward trend. The tensile properties of high layer thickness fabricated are not significantly different than those with a coarse-powder layer thickness of low in previous research.

  3. Research on High Layer Thickness Fabricated of 316L by Selective Laser Melting.

    Science.gov (United States)

    Wang, Shuo; Liu, Yude; Shi, Wentian; Qi, Bin; Yang, Jin; Zhang, Feifei; Han, Dong; Ma, Yingyi

    2017-09-08

    Selective laser melting (SLM) is a potential additive manufacturing (AM) technology. However, the application of SLM was confined due to low efficiency. To improve efficiency, SLM fabrication with a high layer thickness and fine powder was systematically researched, and the void areas and hollow powders can be reduced by using fine powder. Single-track experiments were used to narrow down process parameter windows. Multi-layer fabrication relative density can be reached 99.99% at the exposure time-point distance-hatch space of 120 μs-40 μm-240 μm. Also, the building rate can be up to 12 mm³/s, which is about 3-10 times higher than the previous studies. Three typical defects were found by studying deeply, including the un-melted defect between the molten pools, the micro-pore defect within the molten pool, and the irregular distribution of the splashing phenomenon. Moreover, the microstructure is mostly equiaxed crystals and a small amount of columnar crystals. The averages of ultimate tensile strength, yield strength, and elongation are 625 MPa, 525 MPa, and 39.9%, respectively. As exposure time increased from 80 μs to 200 μs, the grain size is gradually grown up from 0.98 μm to 2.23 μm, the grain aspect ratio is close to 1, and the tensile properties are shown as a downward trend. The tensile properties of high layer thickness fabricated are not significantly different than those with a coarse-powder layer thickness of low in previous research.

  4. Snow cover volumes dynamic monitoring during melting season using high topographic accuracy approach for a Lebanese high plateau witness sinkhole

    Science.gov (United States)

    Abou Chakra, Charbel; Somma, Janine; Elali, Taha; Drapeau, Laurent

    2017-04-01

    Climate change and its negative impact on water resource is well described. For countries like Lebanon, undergoing major population's rise and already decreasing precipitations issues, effective water resources management is crucial. Their continuous and systematic monitoring overs long period of time is therefore an important activity to investigate drought risk scenarios for the Lebanese territory. Snow cover on Lebanese mountains is the most important water resources reserve. Consequently, systematic observation of snow cover dynamic plays a major role in order to support hydrologic research with accurate data on snow cover volumes over the melting season. For the last 20 years few studies have been conducted for Lebanese snow cover. They were focusing on estimating the snow cover surface using remote sensing and terrestrial measurement without obtaining accurate maps for the sampled locations. Indeed, estimations of both snow cover area and volumes are difficult due to snow accumulation very high variability and Lebanese mountains chains slopes topographic heterogeneity. Therefore, the snow cover relief measurement in its three-dimensional aspect and its Digital Elevation Model computation is essential to estimate snow cover volume. Despite the need to cover the all lebanese territory, we favored experimental terrestrial topographic site approaches due to high resolution satellite imagery cost, its limited accessibility and its acquisition restrictions. It is also most challenging to modelise snow cover at national scale. We therefore, selected a representative witness sinkhole located at Ouyoun el Siman to undertake systematic and continuous observations based on topographic approach using a total station. After four years of continuous observations, we acknowledged the relation between snow melt rate, date of total melting and neighboring springs discharges. Consequently, we are able to forecast, early in the season, dates of total snowmelt and springs low

  5. A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis.

    Science.gov (United States)

    Marin, M S; Quintana, S; Leunda, M R; Recavarren, M; Pagnuco, I; Späth, E; Pérez, S; Odeón, A

    2016-01-01

    Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) are antigenically and genetically similar. The aim of this study was to develop a simple and reliable one-step real time PCR assay with high resolution melting (HRM) analysis for the simultaneous detection and differentiation of BoHV-1 and BoHV-5. Optimization of assay conditions was performed with DNA from reference strains. Then, DNA from field isolates, clinical samples and tissue samples of experimentally infected animals were studied by real time PCR-HRM. An efficient amplification of real time PCR products was obtained, and a clear melting curve and appropriate melting peaks for both viruses were achieved in the HRM curve analysis for BoHV type identification. BoHV was identified in all of the isolates and clinical samples, and BoHV types were properly differentiated. Furthermore, viral DNA was detected in 12/18 and 7/18 samples from BoHV-1- and BoHV-5-infected calves, respectively. Real time PCR-HRM achieved a higher sensitivity compared with virus isolation or conventional PCR. In this study, HRM was used as a novel procedure. This method provides rapid, sensitive, specific and simultaneous detection of bovine alpha-herpesviruses DNA. Thus, this technique is an excellent tool for diagnosis, research and epidemiological studies of these viruses in cattle. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Characterisation of Ceramic-Coated 316LN Stainless Steel Exposed to High-Temperature Thermite Melt and Molten Sodium

    Science.gov (United States)

    Ravi Shankar, A.; Vetrivendan, E.; Shukla, Prabhat Kumar; Das, Sanjay Kumar; Hemanth Rao, E.; Murthy, S. S.; Lydia, G.; Nashine, B. K.; Mallika, C.; Selvaraj, P.; Kamachi Mudali, U.

    2017-11-01

    Currently, stainless steel grade 316LN is the material of construction widely used for core catcher of sodium-cooled fast reactors. Design philosophy for core catcher demands its capability to withstand corium loading from whole core melt accidents. Towards this, two ceramic coatings were investigated for its application as a layer of sacrificial material on the top of core catcher to enhance its capability. Plasma-sprayed thermal barrier layer of alumina and partially stabilised zirconia (PSZ) with an intermediate bond coat of NiCrAlY are selected as candidate material and deposited over 316LN SS substrates and were tested for their suitability as thermal barrier layer for core catcher. Coated specimens were exposed to high-temperature thermite melt to simulate impingement of molten corium. Sodium compatibility of alumina and PSZ coatings were also investigated by exposing samples to molten sodium at 400 °C for 500 h. The surface morphology of high-temperature thermite melt-exposed samples and sodium-exposed samples was examined using scanning electron microscope. Phase identification of the exposed samples was carried out by x-ray diffraction technique. Observation from sodium exposure tests indicated that alumina coating offers better protection compared to PSZ coating. However, PSZ coating provided better protection against high-temperature melt exposure, as confirmed during thermite melt exposure test.

  7. Melting of Dense Sodium

    International Nuclear Information System (INIS)

    Gregoryanz, Eugene; Degtyareva, Olga; Hemley, Russell J.; Mao, Ho-kwang; Somayazulu, Maddury

    2005-01-01

    High-pressure high-temperature synchrotron diffraction measurements reveal a maximum on the melting curve of Na in the bcc phase at ∼31 GPa and 1000 K and a steep decrease in melting temperature in its fcc phase. The results extend the melting curve by an order of magnitude up to 130 GPa. Above 103 GPa, Na crystallizes in a sequence of phases with complex structures with unusually low melting temperatures, reaching 300 K at 118 GPa, and an increased melting temperature is observed with further increases in pressure

  8. High efficiency hydrodynamic DNA fragmentation in a bubbling system

    NARCIS (Netherlands)

    Li, Lanhui; Jin, Mingliang; Sun, Chenglong; Wang, Xiaoxue; Xie, Shuting; Zhou, Guofu; Van Den Berg, Albert; Eijkel, Jan C.T.; Shui, Lingling

    2017-01-01

    DNA fragmentation down to a precise fragment size is important for biomedical applications, disease determination, gene therapy and shotgun sequencing. In this work, a cheap, easy to operate and high efficiency DNA fragmentation method is demonstrated based on hydrodynamic shearing in a bubbling

  9. Highly efficient DNA extraction method from skeletal remains

    Directory of Open Access Journals (Sweden)

    Irena Zupanič Pajnič

    2011-03-01

    Full Text Available Background: This paper precisely describes the method of DNA extraction developed to acquire high quality DNA from the Second World War skeletal remains. The same method is also used for molecular genetic identification of unknown decomposed bodies in routine forensic casework where only bones and teeth are suitable for DNA typing. We analysed 109 bones and two teeth from WWII mass graves in Slovenia. Methods: We cleaned the bones and teeth, removed surface contaminants and ground the bones into powder, using liquid nitrogen . Prior to isolating the DNA in parallel using the BioRobot EZ1 (Qiagen, the powder was decalcified for three days. The nuclear DNA of the samples were quantified by real-time PCR method. We acquired autosomal genetic profiles and Y-chromosome haplotypes of the bones and teeth with PCR amplification of microsatellites, and mtDNA haplotypes 99. For the purpose of traceability in the event of contamination, we prepared elimination data bases including genetic profiles of the nuclear and mtDNA of all persons who have been in touch with the skeletal remains in any way. Results: We extracted up to 55 ng DNA/g of the teeth, up to 100 ng DNA/g of the femurs, up to 30 ng DNA/g of the tibias and up to 0.5 ng DNA/g of the humerus. The typing of autosomal and YSTR loci was successful in all of the teeth, in 98 % dekalof the femurs, and in 75 % to 81 % of the tibias and humerus. The typing of mtDNA was successful in all of the teeth, and in 96 % to 98 % of the bones. Conclusions: We managed to obtain nuclear DNA for successful STR typing from skeletal remains that were over 60 years old . The method of DNA extraction described here has proved to be highly efficient. We obtained 0.8 to 100 ng DNA/g of teeth or bones and complete genetic profiles of autosomal DNA, Y-STR haplotypes, and mtDNA haplotypes from only 0.5g bone and teeth samples.

  10. DNA oligonucleotide conformations: high resolution NMR studies

    International Nuclear Information System (INIS)

    Mellema, J.-R.

    1984-01-01

    The present work describes a DNA double-helix model, which is well comparable with the models derived from fibre-diffraction studies. The model has a mononucleotide repeat with torsion angles in accordance with average geometries as derived from 1 H NMR studies. Special attention was paid to reduce the number of short H-H nonbonding contacts, which are abundantly present in the 'classical' fibre-diffraction models. Chapter 3 describes the first complete assignment of a 1 H NMR spectrum of a DNA tetramer, d(TAAT). Preliminary conformational data derived from the spectral parameters recorded at 27 0 C are given. A more detailed analysis employing temperature-dependence studies is given in Chapter 4. (Auth.)

  11. High-speed detection of DNA translocation in nanopipettes

    Science.gov (United States)

    Fraccari, Raquel L.; Ciccarella, Pietro; Bahrami, Azadeh; Carminati, Marco; Ferrari, Giorgio; Albrecht, Tim

    2016-03-01

    We present a high-speed electrical detection scheme based on a custom-designed CMOS amplifier which allows the analysis of DNA translocation in glass nanopipettes on a microsecond timescale. Translocation of different DNA lengths in KCl electrolyte provides a scaling factor of the DNA translocation time equal to p = 1.22, which is different from values observed previously with nanopipettes in LiCl electrolyte or with nanopores. Based on a theoretical model involving electrophoresis, hydrodynamics and surface friction, we show that the experimentally observed range of p-values may be the result of, or at least be affected by DNA adsorption and friction between the DNA and the substrate surface.We present a high-speed electrical detection scheme based on a custom-designed CMOS amplifier which allows the analysis of DNA translocation in glass nanopipettes on a microsecond timescale. Translocation of different DNA lengths in KCl electrolyte provides a scaling factor of the DNA translocation time equal to p = 1.22, which is different from values observed previously with nanopipettes in LiCl electrolyte or with nanopores. Based on a theoretical model involving electrophoresis, hydrodynamics and surface friction, we show that the experimentally observed range of p-values may be the result of, or at least be affected by DNA adsorption and friction between the DNA and the substrate surface. Electronic supplementary information (ESI) available: Gel electrophoresis confirming lengths and purity of DNA samples, comparison between Axopatch 200B and custom-built setup, comprehensive low-noise amplifier characterization, representative I-V curves of nanopipettes used, typical scatter plots of τ vs. peak amplitude for the four LDNA's used, table of most probable τ values, a comparison between different fitting models for the DNA translocation time distribution, further details on the stochastic numerical simulation of the scaling statistics and the derivation of the extended

  12. High-resolution melting PCR assay, applicable for diagnostics and screening studies, allowing detection and differentiation of several Babesia spp. infecting humans and animals.

    Science.gov (United States)

    Rozej-Bielicka, Wioletta; Masny, Aleksander; Golab, Elzbieta

    2017-10-01

    The goal of the study was to design a single tube PCR test for detection and differentiation of Babesia species in DNA samples obtained from diverse biological materials. A multiplex, single tube PCR test was designed for amplification of approximately 400 bp region of the Babesia 18S rRNA gene. Universal primers were designed to match DNA of multiple Babesia spp. and to have low levels of similarity to DNA sequences of other intracellular protozoa and Babesia hosts. The PCR products amplified from Babesia DNA isolated from human, dog, rodent, deer, and tick samples were subjected to high-resolution melting analysis for Babesia species identification. The designed test allowed detection and differentiation of four Babesia species, three zoonotic (B. microti, B. divergens, B. venatorum) and one that is generally not considered zoonotic-Babesia canis. Both detection and identification of all four species were possible based on the HRM curves of the PCR products in samples obtained from the following: humans, dogs, rodents, and ticks. No cross-reactivity with DNA of Babesia hosts or Plasmodium falciparum and Toxoplasma gondii was observed. The lack of cross-reactivity with P. falciparum DNA might allow using the assay in endemic malaria areas. The designed assay is the first PCR-based test for detection and differentiation of several Babesia spp. of medical and veterinary importance, in a single tube reaction. The results of the study show that the designed assay for Babesia detection and identification could be a practical and inexpensive tool for diagnostics and screening studies of diverse biological materials.

  13. SNP discovery and High Resolution Melting Analysis from massive transcriptome sequencing in the California red abalone Haliotis rufescens.

    Science.gov (United States)

    Valenzuela-Muñoz, Valentina; Araya-Garay, José Miguel; Gallardo-Escárate, Cristian

    2013-06-01

    The California red abalone, Haliotis rufescens that belongs to the Haliotidae family, is the largest species of abalone in the world that has sustained the major fishery and aquaculture production in the USA and Mexico. This native mollusk has not been evaluated or assigned a conservation category even though in the last few decades it was heavily exploited until it disappeared in some areas along the California coast. In Chile, the red abalone was introduced in the 1970s from California wild abalone stocks for the purposes of aquaculture. Considering the number of years that the red abalone has been cultivated in Chile crucial genetic information is scarce and critical issues remain unresolved. This study reports and validates novel single nucleotide polymorphisms (SNP) markers for the red abalone H. rufescens using cDNA pyrosequencing. A total of 622 high quality SNPs were identified in 146 sequences with an estimated frequency of 1 SNP each 1000bp. Forty-five SNPs markers with functional information for gene ontology were selected. Of these, 8 were polymorphic among the individuals screened: Heat shock protein 70 (HSP70), vitellogenin (VTG), lysin, alginate lyase enzyme (AL), Glucose-regulated protein 94 (GRP94), fructose-bisphosphate aldolase (FBA), sulfatase 1A precursor (S1AP) and ornithine decarboxylase antizyme (ODC). Two additional sequences were also identified with polymorphisms but no similarities with known proteins were achieved. To validate the putative SNP markers, High Resolution Melting Analysis (HRMA) was conducted in a wild and hatchery-bred population. Additionally, SNP cross-amplifications were tested in two further native abalone species, Haliotis fulgens and Haliotis corrugata. This study provides novel candidate genes that could be used to evaluate loss of genetic diversity due to hatchery selection or inbreeding effects. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Additive Manufacturing of High-Performance 316L Stainless Steel Nanocomposites via Selective Laser Melting

    Science.gov (United States)

    AlMangour, Bandar Abdulaziz

    Austenitic 316L stainless steel alloy is an attractive industrial material combining outstanding corrosion resistance, ductility, and biocompatibility, with promising structural applications and biomedical uses. However, 316L has low strength and wear resistance, limiting its high-performance applicability. Adding secondary hard nanoscale reinforcements to steel matrices, thereby forming steel-matrix nanocomposites (SMCs), can overcome these problems, improving the performance and thereby the applicability of 316L. However, SMC parts with complex-geometry cannot be easily achieved limiting its application. This can be avoided through additive manufacturing (AM) by generating layer-by-layer deposition using computer-aided design data. Expanding the range of AM-applicable materials is necessary to fulfill industrial demand. This dissertation presents the characteristics of new AM-processed high-performance 316L-matrix nanocomposites with nanoscale TiC or TiB2 reinforcements, addressing specific aspects of material design, process control and optimization, and physical metallurgy theory. The nanocomposites were prepared by high-energy ball-milling and consolidated by AM selective laser melting (SLM). Continuous and refined ring-like network structures were obtained with homogenously distributed reinforcements. Additional grain refinement occurred with reinforcement addition, attributed to nanoparticles acting as nuclei for heterogeneous nucleation. The influence of reinforcement content was first investigated; mechanical and tribological behaviors improved with increased reinforcement contents. The compressive yield strengths of composites with TiB2 or TiC reinforcements were approximately five or two times those of 316L respectively. Hot isostatic pressing post-treatment effectively eliminated major cracks and pores in SLM-fabricated components. The effects of the SLM processing parameters on the microstructure and mechanical performance were also investigated. Laser

  15. Mutual interaction between high and low stereo-regularity components for crystallization and melting behaviors of polypropylene blend fibers

    Science.gov (United States)

    Kawai, Kouya; Kohri, Youhei; Takarada, Wataru; Takebe, Tomoaki; Kanai, Toshitaka; Kikutani, Takeshi

    2016-03-01

    Crystallization and melting behaviors of blend fibers of two types of polypropylene (PP), i.e. high stereo-regularity/high molecular weight PP (HPP) and low stereo-regularity/low molecular weight PP (LPP), was investigated. Blend fibers consisting of various HPP/LPP compositions were prepared through the melt spinning process. Differential scanning calorimetry (DSC), temperature modulated DSC (TMDSC) and wide-angle X-ray diffraction (WAXD) analysis were applied for clarifying the crystallization and melting behaviors of individual components. In the DSC measurement of blend fibers with high LPP composition, continuous endothermic heat was detected between the melting peaks of LPP at around 40 °C and that of HPP at around 160 °C. Such endothermic heat was more distinct for the blend fibers with higher LPP composition indicating that the melting of LPP in the heating process was hindered because of the presence of HPP crystals. On the other hand, heat of crystallization was detected at around 90 °C in the case of blend fibers with LPP content of 30 to 70 wt%, indicating that the crystallization of HPP component was taking place during the heating of as-spun blend fibers in the DSC measurement. Through the TMDSC analysis, re-organization of the crystalline structure through the simultaneous melting and re-crystallization was detected in the cases of HPP and blend fibers, whereas re-crystallization was not detected during the melting of LPP fibers. In the WAXD analysis during the heating of fibers, amount of a-form crystal was almost constant up to the melting in the case of single component HPP fibers, whereas there was a distinct increase of the intensity of crystalline reflections from around 100 °C, right after the melting of LPP in the case of blend fibers. These results suggested that the crystallization of HPP in the spinning process as well as during the conditioning process after spinning was hindered by the presence of LPP.

  16. Mutual interaction between high and low stereo-regularity components for crystallization and melting behaviors of polypropylene blend fibers

    International Nuclear Information System (INIS)

    Kawai, Kouya; Takarada, Wataru; Kikutani, Takeshi; Kohri, Youhei; Takebe, Tomoaki; Kanai, Toshitaka

    2016-01-01

    Crystallization and melting behaviors of blend fibers of two types of polypropylene (PP), i.e. high stereo-regularity/high molecular weight PP (HPP) and low stereo-regularity/low molecular weight PP (LPP), was investigated. Blend fibers consisting of various HPP/LPP compositions were prepared through the melt spinning process. Differential scanning calorimetry (DSC), temperature modulated DSC (TMDSC) and wide-angle X-ray diffraction (WAXD) analysis were applied for clarifying the crystallization and melting behaviors of individual components. In the DSC measurement of blend fibers with high LPP composition, continuous endothermic heat was detected between the melting peaks of LPP at around 40 °C and that of HPP at around 160 °C. Such endothermic heat was more distinct for the blend fibers with higher LPP composition indicating that the melting of LPP in the heating process was hindered because of the presence of HPP crystals. On the other hand, heat of crystallization was detected at around 90 °C in the case of blend fibers with LPP content of 30 to 70 wt%, indicating that the crystallization of HPP component was taking place during the heating of as-spun blend fibers in the DSC measurement. Through the TMDSC analysis, re-organization of the crystalline structure through the simultaneous melting and re-crystallization was detected in the cases of HPP and blend fibers, whereas re-crystallization was not detected during the melting of LPP fibers. In the WAXD analysis during the heating of fibers, amount of a-form crystal was almost constant up to the melting in the case of single component HPP fibers, whereas there was a distinct increase of the intensity of crystalline reflections from around 100 °C, right after the melting of LPP in the case of blend fibers. These results suggested that the crystallization of HPP in the spinning process as well as during the conditioning process after spinning was hindered by the presence of LPP.

  17. Mutual interaction between high and low stereo-regularity components for crystallization and melting behaviors of polypropylene blend fibers

    Energy Technology Data Exchange (ETDEWEB)

    Kawai, Kouya; Takarada, Wataru; Kikutani, Takeshi, E-mail: kikutani.t.aa@m.titech.ac.jp [Department of Organic and Polymeric Materials, Graduate School of Science and Engineering, Tokyo Institute of Technology, 2-12-1, O-okayama, Meguro-ku, Tokyo 152-8552 (Japan); Kohri, Youhei; Takebe, Tomoaki [Performance Materials Laboratories, Idemitsu Kosan Co.,Ltd. (Japan); Kanai, Toshitaka [KT Polymer (Japan)

    2016-03-09

    Crystallization and melting behaviors of blend fibers of two types of polypropylene (PP), i.e. high stereo-regularity/high molecular weight PP (HPP) and low stereo-regularity/low molecular weight PP (LPP), was investigated. Blend fibers consisting of various HPP/LPP compositions were prepared through the melt spinning process. Differential scanning calorimetry (DSC), temperature modulated DSC (TMDSC) and wide-angle X-ray diffraction (WAXD) analysis were applied for clarifying the crystallization and melting behaviors of individual components. In the DSC measurement of blend fibers with high LPP composition, continuous endothermic heat was detected between the melting peaks of LPP at around 40 °C and that of HPP at around 160 °C. Such endothermic heat was more distinct for the blend fibers with higher LPP composition indicating that the melting of LPP in the heating process was hindered because of the presence of HPP crystals. On the other hand, heat of crystallization was detected at around 90 °C in the case of blend fibers with LPP content of 30 to 70 wt%, indicating that the crystallization of HPP component was taking place during the heating of as-spun blend fibers in the DSC measurement. Through the TMDSC analysis, re-organization of the crystalline structure through the simultaneous melting and re-crystallization was detected in the cases of HPP and blend fibers, whereas re-crystallization was not detected during the melting of LPP fibers. In the WAXD analysis during the heating of fibers, amount of a-form crystal was almost constant up to the melting in the case of single component HPP fibers, whereas there was a distinct increase of the intensity of crystalline reflections from around 100 °C, right after the melting of LPP in the case of blend fibers. These results suggested that the crystallization of HPP in the spinning process as well as during the conditioning process after spinning was hindered by the presence of LPP.

  18. Quantifying the Evolution of Melt Ponds in the Marginal Ice Zone Using High Resolution Optical Imagery and Neural Networks

    Science.gov (United States)

    Ortiz, M.; Pinales, J. C.; Graber, H. C.; Wilkinson, J.; Lund, B.

    2016-02-01

    Melt ponds on sea ice play a significant and complex role on the thermodynamics in the Marginal Ice Zone (MIZ). Ponding reduces the sea ice's ability to reflect sunlight, and in consequence, exacerbates the albedo positive feedback cycle. In order to understand how melt ponds work and their effect on the heat uptake of sea ice, we must quantify ponds through their seasonal evolution first. A semi-supervised neural network three-class learning scheme using a gradient descent with momentum and adaptive learning rate backpropagation function is applied to classify melt ponds/melt areas in the Beaufort Sea region. The network uses high resolution panchromatic satellite images from the MEDEA program, which are collocated with autonomous platform arrays from the Marginal Ice Zone Program, including ice mass-balance buoys, arctic weather stations and wave buoys. The goal of the study is to capture the spatial variation of melt onset and freeze-up of the ponds within the MIZ, and gather ponding statistics such as size and concentration. The innovation of this work comes from training the neural network as the melt ponds evolve over time; making the machine learning algorithm time-dependent, which has not been previously done. We will achieve this by analyzing the image histograms through quantification of the minima and maxima intensity changes as well as linking textural variation information of the imagery. We will compare the evolution of the melt ponds against several different array sites on the sea ice to explore if there are spatial differences among the separated platforms in the MIZ.

  19. Isolation and analysis of high quality nuclear DNA with reduced organellar DNA for plant genome sequencing and resequencing

    Directory of Open Access Journals (Sweden)

    Zdepski Anna

    2011-05-01

    Full Text Available Abstract Background High throughput sequencing (HTS technologies have revolutionized the field of genomics by drastically reducing the cost of sequencing, making it feasible for individual labs to sequence or resequence plant genomes. Obtaining high quality, high molecular weight DNA from plants poses significant challenges due to the high copy number of chloroplast and mitochondrial DNA, as well as high levels of phenolic compounds and polysaccharides. Multiple methods have been used to isolate DNA from plants; the CTAB method is commonly used to isolate total cellular DNA from plants that contain nuclear DNA, as well as chloroplast and mitochondrial DNA. Alternatively, DNA can be isolated from nuclei to minimize chloroplast and mitochondrial DNA contamination. Results We describe optimized protocols for isolation of nuclear DNA from eight different plant species encompassing both monocot and eudicot species. These protocols use nuclei isolation to minimize chloroplast and mitochondrial DNA contamination. We also developed a protocol to determine the number of chloroplast and mitochondrial DNA copies relative to the nuclear DNA using quantitative real time PCR (qPCR. We compared DNA isolated from nuclei to total cellular DNA isolated with the CTAB method. As expected, DNA isolated from nuclei consistently yielded nuclear DNA with fewer chloroplast and mitochondrial DNA copies, as compared to the total cellular DNA prepared with the CTAB method. This protocol will allow for analysis of the quality and quantity of nuclear DNA before starting a plant whole genome sequencing or resequencing experiment. Conclusions Extracting high quality, high molecular weight nuclear DNA in plants has the potential to be a bottleneck in the era of whole genome sequencing and resequencing. The methods that are described here provide a framework for researchers to extract and quantify nuclear DNA in multiple types of plants.

  20. Refining of high-temperature uranium melt by filtration through foam-ceramic filters

    International Nuclear Information System (INIS)

    Antsiferov, V.N.; Porozova, S.E.; Filippov, V.B.; Shtutsa, M.G.; Il'enko, E.V.; Kolotygina, N.S.

    2004-01-01

    An opportunity of applying foam-ceramic filters of corundum-mullite composition has been studied in refining natural uranium melts. Uranium melting conditions were chosen depending on technical characteristics of the foam ceramic filters. When their using, a portion of nonmetallic inclusions decreases by 20-30% (as little as 2.0-3.5% ingot weight), their size is reduced and their distribution in the ingot volume is equalized, contamination of uranium by the filter material being failed to be noticed. The parameters of foam-ceramic filters are optimized for provision of stable characteristics of uranium melt filtration process [ru

  1. Using an Ablation Gradient Model to Characterize Annual Glacial Melt Contribution to Major Rivers in High Asia

    Science.gov (United States)

    Brodzik, M. J.; Armstrong, R. L.; Khalsa, S. J. S.; Painter, T. H.; Racoviteanu, A.; Rittger, K.

    2014-12-01

    Ice melt from mountain glaciers can represent a significant contribution to freshwater hydrological budgets, along with seasonal snow melt, rainfall and groundwater. In the rivers of High Asia, understanding the proportion of glacier ice melt is critical for water resource management of irrigation and planning for hydropower generation and human consumption. Current climate conditions are producing heterogeneous glacier responses across the Hindu Kush-Karakoram-Himalayan ranges. However, it is not yet clear how contrasting glacier patterns affect regional water resources. For example, in the Upper Indus basin, estimates of glacial contribution to runoff are often not distinguished from seasonal snow contribution, and vary widely, from as little as 15% to as much as 55%. While many studies are based on reasonable concepts, most are based on assumptions uninformed by actual snow or ice cover measurements. While straightforward temperature index models have been used to estimate glacier runoff in some Himalayan basins, application of these models in larger Himalayan basins is limited by difficulties in estimating key model parameters, particularly air temperature. Estimating glacial area from the MODIS Permanent Snow and Ice Extent (MODICE) product for the years 2000-2013, with recently released Shuttle Radar Topography Mission (SRTMGL3) elevation data, we use a simple ablation gradient approach to calculate an upper limit on the contribution of clean glacier ice melt to streamflow data. We present model results for the five major rivers with glaciated headwaters in High Asia: the Bramaputra, Ganges, Indus, Amu Darya and Syr Darya. Using GRDC historical discharge records, we characterize the annual contribution from glacier ice melt. We use MODICE interannual trends in each basin to estimate glacier ice melt uncertainties. Our results are being used in the USAID project, Contribution to High Asia Runoff from Ice and Snow (CHARIS), to inform regional-scale planning for

  2. Evaluation of methods for characterizing the melting curves of a high temperature cobalt-carbon fixed point to define and determine its melting temperature

    Science.gov (United States)

    Lowe, David; Machin, Graham

    2012-06-01

    The future mise en pratique for the realization of the kelvin will be founded on the melting temperatures of particular metal-carbon eutectic alloys as thermodynamic temperature references. However, at the moment there is no consensus on what should be taken as the melting temperature. An ideal melting or freezing curve should be a completely flat plateau at a specific temperature. Any departure from the ideal is due to shortcomings in the realization and should be accommodated within the uncertainty budget. However, for the proposed alloy-based fixed points, melting takes place over typically some hundreds of millikelvins. Including the entire melting range within the uncertainties would lead to an unnecessarily pessimistic view of the utility of these as reference standards. Therefore, detailed analysis of the shape of the melting curve is needed to give a value associated with some identifiable aspect of the phase transition. A range of approaches are or could be used; some purely practical, determining the point of inflection (POI) of the melting curve, some attempting to extrapolate to the liquidus temperature just at the end of melting, and a method that claims to give the liquidus temperature and an impurity correction based on the analytical Scheil model of solidification that has not previously been applied to eutectic melting. The different methods have been applied to cobalt-carbon melting curves that were obtained under conditions for which the Scheil model might be valid. In the light of the findings of this study it is recommended that the POI continue to be used as a pragmatic measure of temperature but where required a specified limits approach should be used to define and determine the melting temperature.

  3. Equations of state and melting curve of boron carbide in the high-pressure range of shock compression

    Energy Technology Data Exchange (ETDEWEB)

    Molodets, A. M., E-mail: molodets@icp.ac.ru; Golyshev, A. A.; Shakhrai, D. V. [Russian Academy of Sciences, Institute for Problems in Chemical Physics (Russian Federation)

    2017-03-15

    We have constructed the equations of state for crystalline boron carbide B{sub 11}C (C–B–C) and its melt under high dynamic and static pressures. A kink on the shock adiabat for boron carbide has been revealed in the pressure range near 100 GPa, and the melting curve with negative curvature in the pressure range 0–120 GPa has been calculated. The results have been used for interpreting the kinks on the shock adiabat for boron carbide in the pressure range of 0–400 GPa.

  4. Kinetics of iron redox reaction in silicate melts: A high temperature Xanes study on an alkali basalt

    Energy Technology Data Exchange (ETDEWEB)

    Cochain, B; Neuville, D R; Roux, J; Strukelj, E; Richet, P [Physique des Mineraux et Magmas, Geochimie-Cosmochimie, CNRS-IPGP, 4 place Jussieu, 75005 Paris (France); Ligny, D de [Universite Claude Bernard Lyon 1, LPCML, F-69622 Villeurbanne (France); Baudelet, F, E-mail: cochain@ipgp.jussieu.f [Synchrotron SOLEIL, L' Orme des Merisiers, Saint Aubin (France)

    2009-11-15

    In Earth and Materials sciences, iron is the most important transition element. Glass and melt properties are strongly affected by iron content and redox state with the consequence that some properties (i.e. viscosity, heat capacity, crystallization...) depend not only on the amounts of Fe{sup 2+} and Fe{sup 3+}, but also on the coordination state of these ions. In this work we investigate iron redox reactions through XANES experiments at the K-edge of iron. Using a high-temperature heating device, pre-edge of XANES spectra exhibits definite advantages to make in-situ measurements and to determine the evolution of redox state with time, temperature and composition of synthetic silicate melts. In this study, new kinetics measurements are presented for a basalt melt from the 31,000-BC eruption of the Puy de Lemptegy Volcano in France. These measurements have been made between 773 K and at superliquidus temperatures up to 1923 K.

  5. Kinetics of iron redox reaction in silicate melts: A high temperature Xanes study on an alkali basalt

    International Nuclear Information System (INIS)

    Cochain, B; Neuville, D R; Roux, J; Strukelj, E; Richet, P; Ligny, D de; Baudelet, F

    2009-01-01

    In Earth and Materials sciences, iron is the most important transition element. Glass and melt properties are strongly affected by iron content and redox state with the consequence that some properties (i.e. viscosity, heat capacity, crystallization...) depend not only on the amounts of Fe 2+ and Fe 3+ , but also on the coordination state of these ions. In this work we investigate iron redox reactions through XANES experiments at the K-edge of iron. Using a high-temperature heating device, pre-edge of XANES spectra exhibits definite advantages to make in-situ measurements and to determine the evolution of redox state with time, temperature and composition of synthetic silicate melts. In this study, new kinetics measurements are presented for a basalt melt from the 31,000-BC eruption of the Puy de Lemptegy Volcano in France. These measurements have been made between 773 K and at superliquidus temperatures up to 1923 K.

  6. High thermoelectric performances of Bi–AE–Co–O compounds directionally growth from the melt

    Directory of Open Access Journals (Sweden)

    Juan-Carlos Diez

    2018-01-01

    Full Text Available Bi2AE2Co2Ox (AE = Ca, Sr, and Ba thermoelectric compounds were grown from the melt by the laser floating zone technique. Microstructural analysis of as-grown samples has shown the formation of well-aligned thermoelectric grains together with a relative high amount of secondary phases. On the other hand, a short (24 h thermal treatment (810 °C for Sr, 800 °C for Ca, and 750 °C for Ba under air, raises of thermoelectric phase content through the recombination of the secondary ones. These microstructural modifications led to a large decrease of electrical resistivity, improving the power factor. These results have been compared with samples prepared by the conventional solid state method and with the best values reported in the literature. From these data, it is possible to deduce that the high thermoelectric characteristics obtained in these samples make them very attractive for practical applications.

  7. High resolution melting: improvements in the genetic diagnosis of hypertrophic cardiomyopathy in a Portuguese cohort

    Directory of Open Access Journals (Sweden)

    Santos Susana

    2012-03-01

    Full Text Available Abstract Background Hypertrophic Cardiomyopathy (HCM is a complex myocardial disorder with a recognized genetic heterogeneity. The elevated number of genes and mutations involved in HCM limits a gene-based diagnosis that should be considered of most importance for basic research and clinical medicine. Methodology In this report, we evaluated High Resolution Melting (HRM robustness, regarding HCM genetic testing, by means of analyzing 28 HCM-associated genes, including the most frequent 4 HCM-associated sarcomere genes, as well as 24 genes with lower reported HCM-phenotype association. We analyzed 80 Portuguese individuals with clinical phenotype of HCM allowing simultaneously a better characterization of this disease in the Portuguese population. Results HRM technology allowed us to identify 60 mutated alleles in 72 HCM patients: 49 missense mutations, 3 nonsense mutations, one 1-bp deletion, one 5-bp deletion, one in frame 3-bp deletion, one insertion/deletion, 3 splice mutations, one 5'UTR mutation in MYH7, MYBPC3, TNNT2, TNNI3, CSRP3, MYH6 and MYL2 genes. Significantly 22 are novel gene mutations. Conclusions HRM was proven to be a technique with high sensitivity and a low false positive ratio allowing a rapid, innovative and low cost genotyping of HCM. In a short return, HRM as a gene scanning technique could be a cost-effective gene-based diagnosis for an accurate HCM genetic diagnosis and hopefully providing new insights into genotype/phenotype correlations.

  8. Additive manufacturing of a high niobium-containing titanium aluminide alloy by selective electron beam melting

    International Nuclear Information System (INIS)

    Tang, H.P.; Yang, G.Y.; Jia, W.P.; He, W.W.; Lu, S.L.; Qian, M.

    2015-01-01

    Additive manufacturing (AM) offers a radical net-shape manufacturing approach for titanium aluminide alloys but significant challenges still remain. A study has been made of the AM of a high niobium-containing titanium aluminide alloy (Ti–45Al–7Nb–0.3W, in at% throughout the paper) using selective electron beam melting (SEBM). The formation of various types of microstructural defects, including banded structures caused by the vaporization of aluminum, was investigated with respect to different processing parameters. To avoid both micro- and macro-cracks, the use of higher preheating temperatures and an intermediate reheating process (to reheat each solidified layer during SEBM) was assessed in detail. These measures enabled effective release of the thermal stress that developed during SEBM and therefore the avoidance of cracks. In addition, the processing conditions for the production of a fine full lamellar microstructure were identified. As a result, the Ti–45Al–7Nb–0.3W alloy fabricated showed outstanding properties (compression strength: 2750 MPa; strain-to-fracture: 37%). SEBM can be used to fabricate high performance titanium aluminide alloys with appropriate processing parameters and pathways

  9. Additive manufacturing of a high niobium-containing titanium aluminide alloy by selective electron beam melting

    Energy Technology Data Exchange (ETDEWEB)

    Tang, H.P., E-mail: thpfys@126.com [State Key Laboratory of Porous Metal Materials, Northwest Institute for Nonferrous Metal Research, Xi' an 710016 (China); Yang, G.Y.; Jia, W.P.; He, W.W.; Lu, S.L. [State Key Laboratory of Porous Metal Materials, Northwest Institute for Nonferrous Metal Research, Xi' an 710016 (China); Qian, M., E-mail: ma.qian@rmit.edu.au [State Key Laboratory of Porous Metal Materials, Northwest Institute for Nonferrous Metal Research, Xi' an 710016 (China); RMIT University, School of Aerospace, Mechanical and Manufacturing Engineering, Centre for Additive Manufacturing, Melbourne, VIC 3001 (Australia)

    2015-06-11

    Additive manufacturing (AM) offers a radical net-shape manufacturing approach for titanium aluminide alloys but significant challenges still remain. A study has been made of the AM of a high niobium-containing titanium aluminide alloy (Ti–45Al–7Nb–0.3W, in at% throughout the paper) using selective electron beam melting (SEBM). The formation of various types of microstructural defects, including banded structures caused by the vaporization of aluminum, was investigated with respect to different processing parameters. To avoid both micro- and macro-cracks, the use of higher preheating temperatures and an intermediate reheating process (to reheat each solidified layer during SEBM) was assessed in detail. These measures enabled effective release of the thermal stress that developed during SEBM and therefore the avoidance of cracks. In addition, the processing conditions for the production of a fine full lamellar microstructure were identified. As a result, the Ti–45Al–7Nb–0.3W alloy fabricated showed outstanding properties (compression strength: 2750 MPa; strain-to-fracture: 37%). SEBM can be used to fabricate high performance titanium aluminide alloys with appropriate processing parameters and pathways.

  10. Highly porous, low elastic modulus 316L stainless steel scaffold prepared by selective laser melting.

    Science.gov (United States)

    Čapek, Jaroslav; Machová, Markéta; Fousová, Michaela; Kubásek, Jiří; Vojtěch, Dalibor; Fojt, Jaroslav; Jablonská, Eva; Lipov, Jan; Ruml, Tomáš

    2016-12-01

    Recently, porous metallic materials have been extensively studied as candidates for use in the fabrication of scaffolds and augmentations to repair trabecular bone defects, e.g. in surroundings of joint replacements. Fabricating these complex structures by using common approaches (e.g., casting and machining) is very challenging. Therefore, rapid prototyping techniques, such as selective laser melting (SLM), have been investigated for these applications. In this study, we characterized a highly porous (87 vol.%) 316L stainless steel scaffold prepared by SLM. 316L steel was chosen because it presents a biomaterial still widely used for fabrication of joint replacements and, from the practical point of view, use of the same material for fabrication of an augmentation and a joint replacement is beneficial for corrosion prevention. The results are compared to the reported properties of two representative nonporous 316L stainless steels prepared either by SLM or casting and subsequent hot forging. The microstructural and mechanical properties and the surface chemical composition and interaction with the cells were investigated. The studied material exhibited mechanical properties that were similar to those of trabecular bone (compressive modulus of elasticity ~0.15GPa, compressive yield strength ~3MPa) and cytocompatibility after one day that was similar to that of wrought 316L stainless steel, which is a commonly used biomaterial. Based on the obtained results, SLM is a suitable method for the fabrication of porous 316L stainless steel scaffolds with highly porous structures. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Melting Penetration Simulation of Fe-U System at High Temperature Using MPS-LER

    International Nuclear Information System (INIS)

    Mustari, A P A; Irwanto, Dwi; Yamaji, A

    2016-01-01

    Melting penetration information of Fe-U system is necessary for simulating the molten core behavior during severe accident in nuclear power plants. For Fe-U system, the information is mainly obtained from experiment, i.e. TREAT experiment. However, there is no reported data on SS304 at temperature above 1350°C. The MPS-LER has been developed and validated to simulate melting penetration on Fe-U system. The MPS-LER modelled the eutectic phenomenon by solving the diffusion process and by applying the binary phase diagram criteria. This study simulates the melting penetration of the system at higher temperature using MPS-LER. Simulations were conducted on SS304 at 1400, 1450 and 1500°C. The simulation results show rapid increase of melting penetration rate. (paper)

  12. Underwound DNA under Tension: Structure, Elasticity, and Sequence-Dependent Behaviors

    Science.gov (United States)

    Sheinin, Maxim Y.; Forth, Scott; Marko, John F.; Wang, Michelle D.

    2011-09-01

    DNA melting under torsion plays an important role in a wide variety of cellular processes. In the present Letter, we have investigated DNA melting at the single-molecule level using an angular optical trap. By directly measuring force, extension, torque, and angle of DNA, we determined the structural and elastic parameters of torsionally melted DNA. Our data reveal that under moderate forces, the melted DNA assumes a left-handed structure as opposed to an open bubble conformation and is highly torsionally compliant. We have also discovered that at low forces melted DNA properties are highly dependent on DNA sequence. These results provide a more comprehensive picture of the global DNA force-torque phase diagram.

  13. Highly temporally resolved response to seasonal surface melt of the Zachariae and 79N outlet glaciers in Northeast Greenland

    DEFF Research Database (Denmark)

    Rathmann, N. M.; Hvidberg, C. S.; Solgaard, A. M.

    2017-01-01

    The seasonal response to surface melting of the Northeast Greenland Ice Stream outlets, Zachariae and 79N, is investigated using new highly temporally resolved surface velocity maps for 2016 combined with numerical modelling. The seasonal speed-up at 79N of 0.15km/yr is suggested to be driven by ...

  14. Effect of Feed Melting, Temperature History, and Minor Component Addition on Spinel Crystallization in High-Level Waste Glass

    Czech Academy of Sciences Publication Activity Database

    Izák, Pavel; Hrma, P.; Arey, B. W.; Plaisted, T. J.

    2001-01-01

    Roč. 289, 1-3 (2001), s. 17-29 ISSN 0022-3093 Grant - others:DOE(US) DE/06/76RL01830 Keywords : feed melting * crystalization * high-level waste glass Subject RIV: CI - Industrial Chemistry, Chemical Engineering Impact factor: 1.363, year: 2001

  15. 46 CFR 153.488 - Design and equipment for tanks carrying high melting point NLSs: Category B.

    Science.gov (United States)

    2010-10-01

    ... (CONTINUED) CERTAIN BULK DANGEROUS CARGOES SHIPS CARRYING BULK LIQUID, LIQUEFIED GAS, OR COMPRESSED GAS... equipment for tanks carrying high melting point NLSs: Category B. Unless waived under § 153.491, for a ship to have its Certificate of Inspection or Certificate of Compliance endorsed allowing a tank to carry...

  16. Rapid Determination of Lymphogranuloma Venereum Serovars of Chlamydia trachomatis by Quantitative High-Resolution Melt Analysis (HRMA)

    Science.gov (United States)

    Stevens, Matthew P.; Garland, Suzanne M.; Zaia, Angelo M.; Tabrizi, Sepehr N.

    2012-01-01

    A quantitative high-resolution melt analysis assay was developed to differentiate lymphogranuloma venereum-causing serovars of Chlamydia trachomatis (L1 to L3) from other C. trachomatis serovars (D to K). The detection limit of this assay is approximately 10 copies per reaction, comparable to the limits of other quantitative-PCR-based methods. PMID:22933594

  17. A comparison of PCR assays for beak and feather disease virus and high resolution melt (HRM) curve analysis of replicase associated protein and capsid genes.

    Science.gov (United States)

    Das, Shubhagata; Sarker, Subir; Ghorashi, Seyed Ali; Forwood, Jade K; Raidal, Shane R

    2016-11-01

    Beak and feather disease virus (BFDV) threatens a wide range of endangered psittacine birds worldwide. In this study, we assessed a novel PCR assay and genetic screening method using high-resolution melt (HRM) curve analysis for BFDV targeting the capsid (Cap) gene (HRM-Cap) alongside conventional PCR detection as well as a PCR method that targets a much smaller fragment of the virus genome in the replicase initiator protein (Rep) gene (HRM-Rep). Limits of detection, sensitivity, specificity and discriminatory power for differentiating BFDV sequences were compared. HRM-Cap had a high positive predictive value and could readily differentiate between a reference genotype and 17 other diverse BFDV genomes with more discriminatory power (genotype confidence percentage) than HRM-Rep. Melt curve profiles generated by HRM-Cap correlated with unique DNA sequence profiles for each individual test genome. The limit of detection of HRM-Cap was lower (2×10 -5 ng/reaction or 48 viral copies) than that for both HRM-Rep and conventional BFDV PCR which had similar sensitivity (2×10 -6 ng or 13 viral copies/reaction). However, when used in a diagnostic setting with 348 clinical samples there was strong agreement between HRM-Cap and conventional PCR (kappa=0.87, PHRM-Cap demonstrated higher specificity (99.9%) than HRM-Rep (80.3%). Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Experimental evidence for flux-lattice melting. [in high-Tc superconductors

    Science.gov (United States)

    Farrell, D. E.; Rice, J. P.; Ginsberg, D. M.

    1991-01-01

    A low-frequency torsional oscillator has been used to search for flux-lattice melting in an untwinned single crystal of YBa2Cu3O(7-delta). The damping of the oscillator was measured as a function of temperature, for applied magnetic fields in the range H = 0.1-2.3 T. A remarkably sharp damping peak has been located. It is suggested that the temperature of the peak corresponds to the melting point of the Abrikosov flux lattice.

  19. Testing Snow Melt Algorithms in High Relief Topography Using Calibrated Enhanced-Resolution Brightness Temperatures, Hunza River Basin, Pakistan

    Science.gov (United States)

    Ramage, J. M.; Brodzik, M. J.; Hardman, M.; Troy, T. J.

    2017-12-01

    Snow is a vital part of the terrestrial hydrological cycle, a crucial resource for people and ecosystems. In mountainous regions snow is extensive, variable, and challenging to document. Snow melt timing and duration are important factors affecting the transfer of snow mass to soil moisture and runoff. Passive microwave brightness temperature (Tb) changes at 36 and 18 GHz are a sensitive way to detect snow melt onset due to their sensitivity to the abrupt change in emissivity. They are widely used on large icefields and high latitude watersheds. The coarse resolution ( 25 km) of historically available data has precluded effective use in high relief, heterogeneous regions, and gaps between swaths also create temporal data gaps at lower latitudes. New enhanced resolution data products generated from a scatterometer image reconstruction for radiometer (rSIR) technique are available at the original frequencies. We use these Calibrated Enhanced-resolution Brightness (CETB) Temperatures Earth System Data Records (ESDR) to evaluate existing snow melt detection algorithms that have been used in other environments, including the cross polarized gradient ratio (XPGR) and the diurnal amplitude variations (DAV) approaches. We use the 36/37 GHz (3.125 km resolution) and 18/19 GHz (6.25 km resolution) vertically and horizontally polarized datasets from the Special Sensor Microwave Imager (SSM/I) and Advanced Microwave Radiometer for EOS (AMSR-E) and evaluate them for use in this high relief environment. The new data are used to assess glacier and snow melt records in the Hunza River Basin [area 13,000 sq. km, located at 36N, 74E], a tributary to the Upper Indus Basin, Pakistan. We compare the melt timing results visually and quantitatively to the corresponding EASE-Grid 2.0 25-km dataset, SRTM topography, and surface temperatures from station and reanalysis data. The new dataset is coarser than the topography, but is able to differentiate signals of melt/refreeze timing for

  20. Simplifying the detection of MUTYH mutations by high resolution melting analysis

    International Nuclear Information System (INIS)

    López-Villar, Isabel; Martínez-López, Joaquín; Ayala, Rosa; Wesselink, Jan; Morillas, Juan Diego; López, Elena; Marín, José Carlos; Díaz-Tasende, José; González, Sara; Robles, Luis

    2010-01-01

    MUTYH-associated polyposis (MAP) is a disorder caused by bi-allelic germline MUTYH mutation, characterized by multiple colorectal adenomas. In order to identify mutations in MUTYH gene we applied High Resolution Melting (HRM) genotyping. HRM analysis is extensively employed as a scanning method for the detection of heterozygous mutations. Therefore, we applied HRM to show effectiveness in detecting homozygous mutations for these clinically important and frequent patients. In this study, we analyzed phenotype and genotype data from 82 patients, with multiple (>= 10) synchronous (19/82) or metachronous (63/82) adenomas and negative APC study (except one case). Analysis was performed by HRM-PCR and direct sequencing, in order to identify mutations in MUTYH exons 7, 12 and 13, where the most prevalent mutations are located. In monoallelic mutation carriers, we evaluated entire MUTYH gene in search of another possible alteration. HRM-PCR was performed with strict conditions in several rounds: the first one to discriminate the heteroduplex patterns and homoduplex patterns and the next ones, in order to refine and confirm parameters. The genotypes obtained were correlated to phenotypic features (number of adenomas (synchronous or metachronous), colorectal cancer (CRC) and family history). MUTYH germline mutations were found in 15.8% (13/82) of patients. The hot spots, Y179C (exon 7) and G396D (exon 13), were readily identified and other mutations were also detected. Each mutation had a reproducible melting profile by HRM, both heterozygous mutations and homozygous mutations. In our study of 82 patients, biallelic mutation is associated with being a carrier of ≥10 synchronous polyps (p = 0.05) and there is no association between biallelic mutation and CRC (p = 0.39) nor family history (p = 0.63). G338H non-pathogenic polymorphism (exon 12) was found in 23.1% (19/82) of patients. In all cases there was concordance between HRM (first and subsequent rounds) and sequencing

  1. Rapid multiplex high resolution melting method to analyze inflammatory related SNPs in preterm birth

    Directory of Open Access Journals (Sweden)

    Pereyra Silvana

    2012-01-01

    Full Text Available Abstract Background Complex traits like cancer, diabetes, obesity or schizophrenia arise from an intricate interaction between genetic and environmental factors. Complex disorders often cluster in families without a clear-cut pattern of inheritance. Genomic wide association studies focus on the detection of tens or hundreds individual markers contributing to complex diseases. In order to test if a subset of single nucleotide polymorphisms (SNPs from candidate genes are associated to a condition of interest in a particular individual or group of people, new techniques are needed. High-resolution melting (HRM analysis is a new method in which polymerase chain reaction (PCR and mutations scanning are carried out simultaneously in a closed tube, making the procedure fast, inexpensive and easy. Preterm birth (PTB is considered a complex disease, where genetic and environmental factors interact to carry out the delivery of a newborn before 37 weeks of gestation. It is accepted that inflammation plays an important role in pregnancy and PTB. Methods Here, we used real time-PCR followed by HRM analysis to simultaneously identify several gene variations involved in inflammatory pathways on preterm labor. SNPs from TLR4, IL6, IL1 beta and IL12RB genes were analyzed in a case-control study. The results were confirmed either by sequencing or by PCR followed by restriction fragment length polymorphism. Results We were able to simultaneously recognize the variations of four genes with similar accuracy than other methods. In order to obtain non-overlapping melting temperatures, the key step in this strategy was primer design. Genotypic frequencies found for each SNP are in concordance with those previously described in similar populations. None of the studied SNPs were associated with PTB. Conclusions Several gene variations related to the same inflammatory pathway were screened through a new flexible, fast and non expensive method with the purpose of analyzing

  2. A simple, high throughput method to locate single copy sequences from Bacterial Artificial Chromosome (BAC libraries using High Resolution Melt analysis

    Directory of Open Access Journals (Sweden)

    Caligari Peter DS

    2010-05-01

    Full Text Available Abstract Background The high-throughput anchoring of genetic markers into contigs is required for many ongoing physical mapping projects. Multidimentional BAC pooling strategies for PCR-based screening of large insert libraries is a widely used alternative to high density filter hybridisation of bacterial colonies. To date, concerns over reliability have led most if not all groups engaged in high throughput physical mapping projects to favour BAC DNA isolation prior to amplification by conventional PCR. Results Here, we report the first combined use of Multiplex Tandem PCR (MT-PCR and High Resolution Melt (HRM analysis on bacterial stocks of BAC library superpools as a means of rapidly anchoring markers to BAC colonies and thereby to integrate genetic and physical maps. We exemplify the approach using a BAC library of the model plant Arabidopsis thaliana. Super pools of twenty five 384-well plates and two-dimension matrix pools of the BAC library were prepared for marker screening. The entire procedure only requires around 3 h to anchor one marker. Conclusions A pre-amplification step during MT-PCR allows high multiplexing and increases the sensitivity and reliability of subsequent HRM discrimination. This simple gel-free protocol is more reliable, faster and far less costly than conventional PCR screening. The option to screen in parallel 3 genetic markers in one MT-PCR-HRM reaction using templates from directly pooled bacterial stocks of BAC-containing bacteria further reduces time for anchoring markers in physical maps of species with large genomes.

  3. Diagnosis of genetic predisposition for lactose intolerance by high resolution melting analysis.

    Science.gov (United States)

    Delacour, Hervé; Leduc, Amandine; Louçano-Perdriat, Andréa; Plantamura, Julie; Ceppa, Franck

    2017-02-01

    Lactose, the principle sugar in milk, is a disaccharide hydrolyzed by intestinal lactase into glucose and galactose, which are absorbed directly by diffusion in the intestine. The decline of lactase expression (or hypolactasia) in intestinal microvilli after weaning is a normal phenomenon in mammals known as lactase deficiency. It is observed in nearly 75% of the world population and is an inherited autosomal recessive trait with incomplete penetrance. It is caused by SNPs in a regulatory element for lactase gene. In Indo-European, lactase deficiency is associated with rs4982235 SNP (or -13910C>T). The aim of this study is to describe a method based on high resolution melting for rapidly detecting genetic predisposition to lactose intolerance. Analytical performance of the assay was assessed by evaluating within and betwwen-run precision and by comparing the results (n = 50 patients) obtained with the HRM assay to those obtained with the gold standard (Sanger sequencing of the region of interest). In silico prediction of HRM curves was performed to evaluate the potential impact of the other SNPs described within the PCR product on the HRM analytical performances. The assay has good performance (CV lactose intolerance.

  4. Kinetic model for quartz and spinel dissolution during melting of high-level-waste glass batch

    International Nuclear Information System (INIS)

    Pokorny, Richard; Rice, Jarrett A.; Crum, Jarrod V.; Schweiger, Michael J.; Hrma, Pavel

    2013-01-01

    The dissolution of quartz particles and the growth and dissolution of crystalline phases during the conversion of batch to glass potentially affects both the glass melting process and product quality. Crystals of spinel exiting the cold cap to molten glass below can be troublesome during the vitrification of iron-containing high-level wastes. To estimate the distribution of quartz and spinel fractions within the cold cap, we used kinetic models that relate fractions of these phases to temperature and heating rate. Fitting the model equations to data showed that the heating rate, apart from affecting quartz and spinel behavior directly, also affects them indirectly via concurrent processes, such as the formation and motion of bubbles. Because of these indirect effects, it was necessary to allow one kinetic parameter (the pre-exponential factor) to vary with the heating rate. The resulting kinetic equations are sufficiently simple for the detailed modeling of batch-to-glass conversion as it occurs in glass melters. The estimated fractions and sizes of quartz and spinel particles as they leave the cold cap, determined in this study, will provide the source terms needed for modeling the behavior of these solid particles within the flow of molten glass in the melter

  5. High-resolution melting (HRM) for genotyping bovine ephemeral fever virus (BEFV).

    Science.gov (United States)

    Erster, Oran; Stram, Rotem; Menasherow, Shopia; Rubistein-Giuni, Marisol; Sharir, Binyamin; Kchinich, Evgeni; Stram, Yehuda

    2017-02-02

    In recent years there have been several major outbreaks of bovine ephemeral disease in the Middle East, including Israel. Such occurrences raise the need for quick identification of the viruses responsible for the outbreaks, in order to rapidly identify the entry of viruses that do not belong to the Middle-East BEFV lineage. This challenge was met by the development of a high-resolution melt (HRM) assay. The assay is based on the viral G gene sequence and generation of an algorithm that calculates and evaluates the GC content of various fragments. The algorithm was designed to scan 50- to 200-base-long segments in a sliding-window manner, compare and rank them using an Order of Technique of Preference by Similarity to Ideal Solution (TOPSIS) the technique for order preference by similarity to ideal solution technique, according to the differences in GC content of homologous fragments. Two fragments were selected, based on a match to the analysis criteria, in terms of size and GC content. These fragments were successfully used in the analysis to differentiate between different virus lineages, thus facilitating assignment of the viruses' geographical origins. Moreover, the assay could be used for differentiating infected from vaccinated animales (DIVA). The new algorithm may therefore be useful for development of improved genotyping studies for other viruses and possibly other microorganisms. Copyright © 2016. Published by Elsevier B.V.

  6. High resolution melting analysis: a rapid and accurate method to detect CALR mutations.

    Directory of Open Access Journals (Sweden)

    Cristina Bilbao-Sieyro

    Full Text Available The recent discovery of CALR mutations in essential thrombocythemia (ET and primary myelofibrosis (PMF patients without JAK2/MPL mutations has emerged as a relevant finding for the molecular diagnosis of these myeloproliferative neoplasms (MPN. We tested the feasibility of high-resolution melting (HRM as a screening method for rapid detection of CALR mutations.CALR was studied in wild-type JAK2/MPL patients including 34 ET, 21 persistent thrombocytosis suggestive of MPN and 98 suspected secondary thrombocytosis. CALR mutation analysis was performed through HRM and Sanger sequencing. We compared clinical features of CALR-mutated versus 45 JAK2/MPL-mutated subjects in ET.Nineteen samples showed distinct HRM patterns from wild-type. Of them, 18 were mutations and one a polymorphism as confirmed by direct sequencing. CALR mutations were present in 44% of ET (15/34, 14% of persistent thrombocytosis suggestive of MPN (3/21 and none of the secondary thrombocytosis (0/98. Of the 18 mutants, 9 were 52 bp deletions, 8 were 5 bp insertions and other was a complex mutation with insertion/deletion. No mutations were found after sequencing analysis of 45 samples displaying wild-type HRM curves. HRM technique was reproducible, no false positive or negative were detected and the limit of detection was of 3%.This study establishes a sensitive, reliable and rapid HRM method to screen for the presence of CALR mutations.

  7. Corium spreading: hydrodynamics, rheology and solidification of a high-temperature oxide melt

    International Nuclear Information System (INIS)

    Journeau, Ch.

    2006-06-01

    In the hypothesis of a nuclear reactor severe accident, the core could melt and form a high- temperature (2000-3000 K) mixture called corium. In the hypothesis of vessel rupture, this corium would spread in the reactor pit and adjacent rooms as occurred in Chernobyl or in a dedicated core-catcher s in the new European Pressurized reactor, EPR. This thesis is dedicated to the experimental study of corium spreading, especially with the prototypic corium material experiments performed in the VULCANO facility at CEA Cadarache. The first step in analyzing these tests consists in interpreting the material analyses, with the help of thermodynamic modelling of corium solidification. Knowing for each temperature the phase repartition and composition, physical properties can be estimated. Spreading termination is controlled by corium rheological properties in the solidification range, which leads to studying them in detail. The hydrodynamical, rheological and solidification aspects of corium spreading are taken into account in models and computer codes which have been validated against these tests and enable the assessment of the EPR spreading core-catcher concept. (author)

  8. Traceability of PDO Olive Oil “Terra di Bari” Using High Resolution Melting

    Directory of Open Access Journals (Sweden)

    Cinzia Montemurro

    2015-01-01

    Full Text Available The aim of the research was to verify the applicability of microsatellite (SSR markers in High Resolution Melting (HRM analysis for the identification of the olive cultivars used in the “Terra di Bari” PDO extra virgin olive oil. A panel of nine cultivars, widespread in Apulia region, was tested with seventeen SSR primer pairs and the PCR products were at first analysed with a Genetic Analyzer automatic sequencer. An identification key was obtained for the nine cultivars, which showed an unambiguous discrimination among the varieties constituting the “Terra di Bari” PDO extra virgin olive oil: Cima di Bitonto, Coratina, and Ogliarola. Subsequently, an SSR based method was set up with the DCA18 marker, coupled with HRM analysis for the distinction of the Terra di Bari olive oil from non-Terra di Bari olive oil using different mixtures. Thus, this analysis enabled the distinction and identification of the PDO mixtures. Hence, this assay provided a flexible, cost-effective, and closed-tube microsatellite genotyping method, well suited to varietal identification and authentication analysis in olive oil.

  9. Application of high-resolution melting analysis for authenticity testing of valuable Dendrobium commercial products.

    Science.gov (United States)

    Dong, Xiaoman; Jiang, Chao; Yuan, Yuan; Peng, Daiyin; Luo, Yuqin; Zhao, Yuyang; Huang, Luqi

    2018-01-01

    The accurate identification of botanical origin in commercial products is important to ensure food authenticity and safety for consumers. The Dendrobium species have long been commercialised as functional food supplements and herbal medicines in Asia. Three valuable Dendrobium species, namely Dendrobium officinale, D. huoshanense and D. moniliforme, are often mutually adulterated in trade products in pursuit of higher profit. In this paper, a rapid and reliable semi-quantitative method for identifying the botanical origin of Dendrobium products in terminal markets was developed using high-resolution melting (HRM) analysis with specific primer pairs to target the trnL-F region. The HRM analysis method detected amounts of D. moniliforme adulterants as low as 1% in D. huoshanense or D. officinale products. The results have demonstrated that HRM analysis is a fast and effective tool for the differentiation of these Dendrobium species both for their authenticity as well as for the semi-quantitative determination of the purity of their processed products. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  10. Theory of high-force DNA stretching and overstretching.

    Science.gov (United States)

    Storm, C; Nelson, P C

    2003-05-01

    Single-molecule experiments on single- and double-stranded DNA have sparked a renewed interest in the force versus extension of polymers. The extensible freely jointed chain (FJC) model is frequently invoked to explain the observed behavior of single-stranded DNA, but this model does not satisfactorily describe recent high-force stretching data. We instead propose a model (the discrete persistent chain) that borrows features from both the FJC and the wormlike chain, and show that it resembles the data more closely. We find that most of the high-force behavior previously attributed to stretch elasticity is really a feature of the corrected entropic elasticity; the true stretch compliance of single-stranded DNA is several times smaller than that found by previous authors. Next we elaborate our model to allow coexistence of two conformational states of DNA, each with its own stretch and bend elastic constants. Our model is computationally simple and gives an excellent fit through the entire overstretching transition of nicked, double-stranded DNA. The fit gives the first value for the bend stiffness of the overstretched state. In particular, we find the effective bend stiffness for DNA in this state to be about 12 nm k(B)T, a value quite different from either the B-form or single-stranded DNA.

  11. Impulsive shock induced single drop steam explosion visualized by high-speed x-ray radiography and photography - metallic melt

    International Nuclear Information System (INIS)

    Park, H. S.; Hansson, R. C.; Sehgal, B. R.

    2003-01-01

    Experimental investigation of fine fragmentation process during vapor explosion was conducted in a small-scale single drop system employing continuous high-speed X-ray radiography and photography. A molten tin drop of about 0.7 g at approximately 1000 .deg. C was dropped into a water pool, at temperatures ranging from 20 to 90 .deg. C, and the explosion was triggered by an external shock pulse of about 1 MPa. X-ray radiographs show that finely fragmented melt particles accelerates to the vapor bubble boundary and forms a particle shell during the period of vapor bubble expansion due to vapor explosions. From the photographs, it was possible to observe a number of counter-jets on the vapor boundary. For tests with highly subcooled coolant, local explosion due to external impulsive shock trigger initiates the stratified mode of explosion along the entire melt surface. For tests with lower subcooled coolant local explosions were initiated by an external impulsive shock trigger and by collapse of vapor/gas pocket attached on the top of the melt drop. Transient spatial distribution map of melt fragments during vapor explosion was obtained by a series of image processing and calibration tests

  12. High-Throughput Block Optical DNA Sequence Identification.

    Science.gov (United States)

    Sagar, Dodderi Manjunatha; Korshoj, Lee Erik; Hanson, Katrina Bethany; Chowdhury, Partha Pratim; Otoupal, Peter Britton; Chatterjee, Anushree; Nagpal, Prashant

    2018-01-01

    Optical techniques for molecular diagnostics or DNA sequencing generally rely on small molecule fluorescent labels, which utilize light with a wavelength of several hundred nanometers for detection. Developing a label-free optical DNA sequencing technique will require nanoscale focusing of light, a high-throughput and multiplexed identification method, and a data compression technique to rapidly identify sequences and analyze genomic heterogeneity for big datasets. Such a method should identify characteristic molecular vibrations using optical spectroscopy, especially in the "fingerprinting region" from ≈400-1400 cm -1 . Here, surface-enhanced Raman spectroscopy is used to demonstrate label-free identification of DNA nucleobases with multiplexed 3D plasmonic nanofocusing. While nanometer-scale mode volumes prevent identification of single nucleobases within a DNA sequence, the block optical technique can identify A, T, G, and C content in DNA k-mers. The content of each nucleotide in a DNA block can be a unique and high-throughput method for identifying sequences, genes, and other biomarkers as an alternative to single-letter sequencing. Additionally, coupling two complementary vibrational spectroscopy techniques (infrared and Raman) can improve block characterization. These results pave the way for developing a novel, high-throughput block optical sequencing method with lossy genomic data compression using k-mer identification from multiplexed optical data acquisition. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. qPCR-High resolution melt analysis for drug susceptibility testing of Mycobacterium leprae directly from clinical specimens of leprosy patients.

    Science.gov (United States)

    Araujo, Sergio; Goulart, Luiz Ricardo; Truman, Richard W; Goulart, Isabela Maria B; Vissa, Varalakshmi; Li, Wei; Matsuoka, Masanori; Suffys, Philip; Fontes, Amanda B; Rosa, Patricia S; Scollard, David M; Williams, Diana L

    2017-06-01

    Real-Time PCR-High Resolution Melting (qPCR-HRM) analysis has been recently described for rapid drug susceptibility testing (DST) of Mycobacterium leprae. The purpose of the current study was to further evaluate the validity, reliability, and accuracy of this assay for M. leprae DST in clinical specimens. The specificity and sensitivity for determining the presence and susceptibility of M. leprae to dapsone based on the folP1 drug resistance determining region (DRDR), rifampin (rpoB DRDR) and ofloxacin (gyrA DRDR) was evaluated using 211 clinical specimens from leprosy patients, including 156 multibacillary (MB) and 55 paucibacillary (PB) cases. When comparing the results of qPCR-HRM DST and PCR/direct DNA sequencing, 100% concordance was obtained. The effects of in-house phenol/chloroform extraction versus column-based DNA purification protocols, and that of storage and fixation protocols of specimens for qPCR-HRM DST, were also evaluated. qPCR-HRM results for all DRDR gene assays (folP1, rpoB, and gyrA) were obtained from both MB (154/156; 98.7%) and PB (35/55; 63.3%) patients. All PCR negative specimens were from patients with low numbers of bacilli enumerated by an M. leprae-specific qPCR. We observed that frozen and formalin-fixed paraffin embedded (FFPE) tissues or archival Fite's stained slides were suitable for HRM analysis. Among 20 mycobacterial and other skin bacterial species tested, only M. lepromatosis, highly related to M. leprae, generated amplicons in the qPCR-HRM DST assay for folP1 and rpoB DRDR targets. Both DNA purification protocols tested were efficient in recovering DNA suitable for HRM analysis. However, 3% of clinical specimens purified using the phenol/chloroform DNA purification protocol gave false drug resistant data. DNA obtained from freshly frozen (n = 172), formalin-fixed paraffin embedded (FFPE) tissues (n = 36) or archival Fite's stained slides (n = 3) were suitable for qPCR-HRM DST analysis. The HRM-based assay was also able to

  14. High-Throughput DNA sequencing of ancient wood.

    Science.gov (United States)

    Wagner, Stefanie; Lagane, Frédéric; Seguin-Orlando, Andaine; Schubert, Mikkel; Leroy, Thibault; Guichoux, Erwan; Chancerel, Emilie; Bech-Hebelstrup, Inger; Bernard, Vincent; Billard, Cyrille; Billaud, Yves; Bolliger, Matthias; Croutsch, Christophe; Čufar, Katarina; Eynaud, Frédérique; Heussner, Karl Uwe; Köninger, Joachim; Langenegger, Fabien; Leroy, Frédéric; Lima, Christine; Martinelli, Nicoletta; Momber, Garry; Billamboz, André; Nelle, Oliver; Palomo, Antoni; Piqué, Raquel; Ramstein, Marianne; Schweichel, Roswitha; Stäuble, Harald; Tegel, Willy; Terradas, Xavier; Verdin, Florence; Plomion, Christophe; Kremer, Antoine; Orlando, Ludovic

    2018-03-01

    Reconstructing the colonization and demographic dynamics that gave rise to extant forests is essential to forecasts of forest responses to environmental changes. Classical approaches to map how population of trees changed through space and time largely rely on pollen distribution patterns, with only a limited number of studies exploiting DNA molecules preserved in wooden tree archaeological and subfossil remains. Here, we advance such analyses by applying high-throughput (HTS) DNA sequencing to wood archaeological and subfossil material for the first time, using a comprehensive sample of 167 European white oak waterlogged remains spanning a large temporal (from 550 to 9,800 years) and geographical range across Europe. The successful characterization of the endogenous DNA and exogenous microbial DNA of 140 (~83%) samples helped the identification of environmental conditions favouring long-term DNA preservation in wood remains, and started to unveil the first trends in the DNA decay process in wood material. Additionally, the maternally inherited chloroplast haplotypes of 21 samples from three periods of forest human-induced use (Neolithic, Bronze Age and Middle Ages) were found to be consistent with those of modern populations growing in the same geographic areas. Our work paves the way for further studies aiming at using ancient DNA preserved in wood to reconstruct the micro-evolutionary response of trees to climate change and human forest management. © 2018 John Wiley & Sons Ltd.

  15. MELT RATE ENHANCEMENT FOR HIGH ALUMINUM HLW (HIGH LEVEL WASTE) GLASS FORMULATION FINAL REPORT 08R1360-1

    Energy Technology Data Exchange (ETDEWEB)

    KRUGER AA; MATLACK KS; KOT W; PEGG IL; JOSEPH I; BARDAKCI T; GAN H; GONG W; CHAUDHURI M

    2010-01-04

    This report describes the development and testing of new glass formulations for high aluminum waste streams that achieve high waste loadings while maintaining high processing rates. The testing was based on the compositions of Hanford High Level Waste (HLW) with limiting concentrations of aluminum specified by the Office of River Protection (ORP). The testing identified glass formulations that optimize waste loading and waste processing rate while meeting all processing and product quality requirements. The work included preparation and characterization of crucible melts and small scale melt rate screening tests. The results were used to select compositions for subsequent testing in a DuraMelter 100 (DM100) system. These tests were used to determine processing rates for the selected formulations as well as to examine the effects of increased glass processing temperature, and the form of aluminum in the waste simulant. Finally, one of the formulations was selected for large-scale confirmatory testing on the HLW Pilot Melter (DM1200), which is a one third scale prototype of the Hanford Tank Waste Treatment and Immobilization Plant (WTP) HLW melter and off-gas treatment system. This work builds on previous work performed at the Vitreous State Laboratory (VSL) for Department of Energy (DOE) to increase waste loading and processing rates for high-iron HLW waste streams as well as previous tests conducted for ORP on the same high-aluminum waste composition used in the present work and other Hanford HLW compositions. The scope of this study was outlined in a Test Plan that was prepared in response to an ORP-supplied statement of work. It is currently estimated that the number of HLW canisters to be produced in the WTP is about 13,500 (equivalent to 40,500 MT glass). This estimate is based upon the inventory of the tank wastes, the anticipated performance of the sludge treatment processes, and current understanding of the capability of the borosilicate glass waste form

  16. MELT RATE ENHANCEMENT FOR HIGH ALUMINUM HLW (HIGH LEVEL WASTE) GLASS FORMULATION. FINAL REPORT 08R1360-1

    International Nuclear Information System (INIS)

    Kruger, A.A.; Matlack, K.S.; Kot, W.; Pegg, I.L.; Joseph, I.; Bardakci, T.; Gan, H.; Gong, W.; Chaudhuri, M.

    2010-01-01

    This report describes the development and testing of new glass formulations for high aluminum waste streams that achieve high waste loadings while maintaining high processing rates. The testing was based on the compositions of Hanford High Level Waste (HLW) with limiting concentrations of aluminum specified by the Office of River Protection (ORP). The testing identified glass formulations that optimize waste loading and waste processing rate while meeting all processing and product quality requirements. The work included preparation and characterization of crucible melts and small scale melt rate screening tests. The results were used to select compositions for subsequent testing in a DuraMelter 100 (DM100) system. These tests were used to determine processing rates for the selected formulations as well as to examine the effects of increased glass processing temperature, and the form of aluminum in the waste simulant. Finally, one of the formulations was selected for large-scale confirmatory testing on the HLW Pilot Melter (DM1200), which is a one third scale prototype of the Hanford Tank Waste Treatment and Immobilization Plant (WTP) HLW melter and off-gas treatment system. This work builds on previous work performed at the Vitreous State Laboratory (VSL) for Department of Energy (DOE) to increase waste loading and processing rates for high-iron HLW waste streams as well as previous tests conducted for ORP on the same high-aluminum waste composition used in the present work and other Hanford HLW compositions. The scope of this study was outlined in a Test Plan that was prepared in response to an ORP-supplied statement of work. It is currently estimated that the number of HLW canisters to be produced in the WTP is about 13,500 (equivalent to 40,500 MT glass). This estimate is based upon the inventory of the tank wastes, the anticipated performance of the sludge treatment processes, and current understanding of the capability of the borosilicate glass waste form

  17. Geotechnical modeling of high-level nuclear waste disposal by rock melting

    International Nuclear Information System (INIS)

    Heuze, F.E.

    1981-12-01

    A new strategy has been developed for the geotechnical modeling of nuclear waste disposal by rock melting (DRM). Three seeparate tasks were performed to reach this objective: a review of the four scenarios which have been proposed for DRM, to date; an evaluation of computer-based numerical models which could be used to analyze the mechanical, thermal, and hydraulic processes involved in DRM; and a critical review of rock mass properties which are relevant to the design and safety of waste disposal by rock melting. It is concluded that several geotechnical aspects of DRM can be studied realistically with current state-of-the-art model capabilities and knowledge of material properties. The next step in the feasibility study of DRM should be a best-estimate calculation of the four cavity-melt and canister-burial concepts. These new analyses will indicate the most critical areas for subsequent research

  18. Genetic high throughput screening in Retinitis Pigmentosa based on high resolution melting (HRM) analysis.

    Science.gov (United States)

    Anasagasti, Ander; Barandika, Olatz; Irigoyen, Cristina; Benitez, Bruno A; Cooper, Breanna; Cruchaga, Carlos; López de Munain, Adolfo; Ruiz-Ederra, Javier

    2013-11-01

    Retinitis Pigmentosa (RP) involves a group of genetically determined retinal diseases caused by a large number of mutations that result in rod photoreceptor cell death followed by gradual death of cone cells. Most cases of RP are monogenic, with more than 80 associated genes identified so far. The high number of genes and variants involved in RP, among other factors, is making the molecular characterization of RP a real challenge for many patients. Although HRM has been used for the analysis of isolated variants or single RP genes, as far as we are concerned, this is the first study that uses HRM analysis for a high-throughput screening of several RP genes. Our main goal was to test the suitability of HRM analysis as a genetic screening technique in RP, and to compare its performance with two of the most widely used NGS platforms, Illumina and PGM-Ion Torrent technologies. RP patients (n = 96) were clinically diagnosed at the Ophthalmology Department of Donostia University Hospital, Spain. We analyzed a total of 16 RP genes that meet the following inclusion criteria: 1) size: genes with transcripts of less than 4 kb; 2) number of exons: genes with up to 22 exons; and 3) prevalence: genes reported to account for, at least, 0.4% of total RP cases worldwide. For comparison purposes, RHO gene was also sequenced with Illumina (GAII; Illumina), Ion semiconductor technologies (PGM; Life Technologies) and Sanger sequencing (ABI 3130xl platform; Applied Biosystems). Detected variants were confirmed in all cases by Sanger sequencing and tested for co-segregation in the family of affected probands. We identified a total of 65 genetic variants, 15 of which (23%) were novel, in 49 out of 96 patients. Among them, 14 (4 novel) are probable disease-causing genetic variants in 7 RP genes, affecting 15 patients. Our HRM analysis-based study, proved to be a cost-effective and rapid method that provides an accurate identification of genetic RP variants. This approach is effective for

  19. Sediment transport during the snow melt period in a Mediterranean high mountain catchment

    Energy Technology Data Exchange (ETDEWEB)

    Alvera, B.; Lana-Renault, N.; Garcia-Ruiz, J. M.

    2009-07-01

    Transport of suspended sediment and solutes during the snow melt period (May-June, 2004) in the Izas catchment (Central Pyrenees) was studied to obtain a sediment balance and to assess the annual importance of sediment transport. The results showed that most sediment was exported in the form of solutes (75,6% of the total); 24.4% was exported as suspended sediment and no bed load was recorded. Sediment transport during the snow melt period represented 42.7% of the annual sediment yield. (Author) 7 refs.

  20. Sediment transport during the snow melt period in a Mediterranean high mountain catchment

    International Nuclear Information System (INIS)

    Alvera, B.; Lana-Renault, N.; Garcia-Ruiz, J. M.

    2009-01-01

    Transport of suspended sediment and solutes during the snow melt period (May-June, 2004) in the Izas catchment (Central Pyrenees) was studied to obtain a sediment balance and to assess the annual importance of sediment transport. The results showed that most sediment was exported in the form of solutes (75,6% of the total); 24.4% was exported as suspended sediment and no bed load was recorded. Sediment transport during the snow melt period represented 42.7% of the annual sediment yield. (Author) 7 refs.

  1. Melting properties of some structured lipids native to high stearic acid soybean oil

    Directory of Open Access Journals (Sweden)

    Dunn, R. O.

    2004-06-01

    Full Text Available A number of structured lipids native to high stearic acid soybean oil were synthesized and their physical properties were determined by pulsed nuclear magnetic resonance (NMR, Mettler dropping point and differential scanning calorimetry (DSC. 1,3 Distearo-2-olein (SOS, 1,3 distearo-2-linolein (SLS and1,3 distearo-2-linolenin (SlnS were synthesized from pure 1,3 diacylglycerols and the appropriate fatty acid. Pulsed NMR determinations over the temperature range 10-50 ºC showed that the symmetrical triacylglycerols (SUS: where S = stearic, U = oleic, linoleic or linolenic are high and sharply melting materials, all showing substantial amounts of solids at temperatures up to 33.3 ºC, yet are completely melted at only a few degrees higher. Mettler dropping points for SOS, SLS and SlnS were 44.1, 37.9 and 36.5 ºC respectively. The heats of fusion for the structured triacylglycerols were determined by DSC and shown to be of the order 29-32 cal/gm compared to 45 cal/gm for SSS. The heats of fusion were also calculated from Mettler dropping point determinations as admixtures with soybean oil and showed consistent agreement with the DSC data.Se sintetizaron algunos lípidos estructurados procedentes del aceite de soja con alto contenido en ácido esteárico y sus propiedades físicas se determinaron por resonancia magnética nuclear pulsada (NMR, punto de goteo Mettler y calorimetría diferencial de barrido (DSC. Se sintetizaron 1,3 diestearo-2-oleina (SOS, 1,3 diestearo-2-linoleina (SLS y 1,3 diestearo-2-linolenina (SlnS a partir de 1,3 diacilgliceroles y de los ácidos grasos adecuados puros. Las determinaciones de NMR pulsada en el rango de temperaturas 10- 50 ºC mostraron que los triacilgliceroles simétricos (SUS: donde S = esteárico, U = oleico, linoleico o linolénico funden a mayor temperatura y más bruscamente, todos presentan altos contenidos en sólidos a todas las temperaturas hasta los 33.3 ºC, estando completamente fundidos

  2. Sequence dependent structure and thermodynamics of DNA oligonucleotides and polynucleotides: uv melting and NMR (nuclear magnetic resonance) studies

    Energy Technology Data Exchange (ETDEWEB)

    Aboul-ela, F.M.

    1987-12-01

    Thermodynamic parameters for double strand formation have been measured for the twenty-five DNA double helices made by mixing deoxyoligonucleotides of the sequence dCA/sub 3/XA/sub 3/G with the complement dCT/sub 3/YT/sub 3/G. Each of the bases A, C, G, T, and I (I = hypoxanthine) have been substituted at the positions labeled X and Y. The results are analyzed in terms of nearest neighbors. At higher temperatures the sequences containing a G)centerreverse arrowdot)C base pair become more stable than those containing only A)centerreverse arrowdot)T. All molecules containing mismatcher are destabilized with respect to those with only Watson-Crick pairing, but there is a wide range of destabilization. Large neighboring base effects upon stability were observed. For example, when (X, Y) = (I, A), the duplex is eightfold more stable than when (X, Y) = (A, I). Independent of sequence effects the order of stabilities is: I)centerreverse arrowdot)C )succ) I)centerreverse arrowdot) A)succ) I)centerreverse arrowdot)T approx. I)centerreverse arrowdot)G. All of these results are discussed within the context of models for sequence dependent DNA secondary structure, replication fidelity and mechanisms of mismatch repair, and implications for probe design. The duplex deoxyoligonucleotide d(GGATGGGAG))centerreverse arrowdot)d(CTCCCATCC) is a portion of the gene recognition sequence of the protein transcription factor IIIA. The crystal structure of this oligonucleotide was shown to be A-form The present study employs Nuclear Magnetic Resonance, optical, chemical and enzymatic techniques to investigate the solution structure of this DNA 9-mer. (157 refs., 19 figs., 10 tabs.

  3. Sequence dependent structure and thermodynamics of DNA oligonucleotides and polynucleotides: uv melting and NMR (nuclear magnetic resonance) studies

    International Nuclear Information System (INIS)

    Aboul-ela, F.M.

    1987-12-01

    Thermodynamic parameters for double strand formation have been measured for the twenty-five DNA double helices made by mixing deoxyoligonucleotides of the sequence dCA 3 XA 3 G with the complement dCT 3 YT 3 G. Each of the bases A, C, G, T, and I (I = hypoxanthine) have been substituted at the positions labeled X and Y. The results are analyzed in terms of nearest neighbors. At higher temperatures the sequences containing a G/center dot/C base pair become more stable than those containing only A/center dot/T. All molecules containing mismatcher are destabilized with respect to those with only Watson-Crick pairing, but there is a wide range of destabilization. Large neighboring base effects upon stability were observed. For example, when (X, Y) = (I, A), the duplex is eightfold more stable than when (X, Y) = (A, I). Independent of sequence effects the order of stabilities is: I/center dot/C /succ/ I/center dot/ A/succ/ I/center dot/T ∼ I/center dot/G. All of these results are discussed within the context of models for sequence dependent DNA secondary structure, replication fidelity and mechanisms of mismatch repair, and implications for probe design. The duplex deoxyoligonucleotide d(GGATGGGAG)/center dot/d(CTCCCATCC) is a portion of the gene recognition sequence of the protein transcription factor IIIA. The crystal structure of this oligonucleotide was shown to be A-form The present study employs Nuclear Magnetic Resonance, optical, chemical and enzymatic techniques to investigate the solution structure of this DNA 9-mer. (157 refs., 19 figs., 10 tabs.)

  4. Evaluation of High Resolution Melting for MTHFR C677T Genotyping in Congenital Heart Disease.

    Directory of Open Access Journals (Sweden)

    Ying Wang

    Full Text Available High resolution melting (HRM is a simple, flexible and low-cost mutation screening technique. The methylenetetrahydrofolate reductase (MTHFR gene encoding a critical enzyme, potentially affects susceptibility to some congenital defects like congenital heart disease (CHD. We evaluate the performance of HRM for genotyping of the MTHFR gene C677T locus in CHD cases and healthy controls of Chinese Han population.A total of 315 blood samples from 147 CHD patients (male72, female 75 and 168 healthy controls (male 92, female 76 were enrolled in the study. HRM was utilized to genotype MTHFR C677T locus of all the samples. The results were compared to that of PCR-RFLP and Sanger sequencing. The association of the MTHFR C677T genotypes and the risk of CHD was analyzed using odds ratio with their 95% confidence interval (CIs from unconditional logistic regression.All the samples were successfully genotyped by HRM within 1 hour and 30 minutes while at least 6 hours were needed for PCR-RFLP and sequencing. The genotypes of MTHFR C677T CC, CT, and TT were 9.52%, 49.66%, and 40.82% in CHD group but 29.17%, 50% and 20.83% in control group, which were identical using both methods of HRM and PCR-RFLP, demonstrating the sensitivity and specificity of HRM were all 100%.MTHFR C677T is a potential risk factor for CHD in our local residents of Shandong province in China. HRM is a fast, sensitive, specific and reliable method for clinical application of genotyping.

  5. High water contents in basaltic melt inclusions from Arenal volcano, Costa Rica

    Science.gov (United States)

    Wade, J. A.; Plank, T.; Hauri, E. H.; Melson, W. G.; Soto, G. J.

    2004-12-01

    Despite the importance of water to arc magma genesis, fractionation and eruption, few quantitative constraints exist on the water content of Arenal magmas. Early estimates, by electron microprobe sum deficit, suggested up to 4 wt% H2O in olivine-hosted basaltic andesite melt inclusions (MI) from pre-historic ET-6 tephra (Melson, 1982), and up to 7 wt% H2O in plagioclase and orthopyroxene-hosted dacitic MI from 1968 lapilli (Anderson, 1979). These high water contents are consistent with abundant hornblende phenocrysts in Arenal volcanics, but inconsistent with geochemical tracers such as 10Be and Ba/La that suggest a low flux of recycled material (and presumably water) from the subduction zone. In order to test these ideas, and provide the first direct measurements of water in mafic Arenal magmas, we have studied olivine-hosted MI from the prehistoric (900 yBP; Soto et al., 1998) ET3 tephra layer. MI range from andesitic (> 58% SiO2) to basaltic compositions ( 4 wt%) found here for Arenal basaltic MI support the semi-quantitative data from earlier studies, but are somewhat unexpected given predictions from slab tracers. Arenal water contents (4%) approach those of the 1995 eruption of Cerro Negro in Nicaragua (4-5 wt% in basaltic MI; Roggensack et al., 1997), despite the fact that the latter has Ba/La of > 100, while Arenal has Ba/La Journal of Geology; Melson, William G. (1982) Boletin de Volcanologia; Roggensack et al. (1997) Science; Soto et al. (1998) OSIVAM; Williams-Jones et al. (2001) Journal of Volc. and Geoth. Res.

  6. Highly CO2-supersaturated melts in the Pannonian lithospheric mantle - A transient carbon reservoir?

    Science.gov (United States)

    Créon, Laura; Rouchon, Virgile; Youssef, Souhail; Rosenberg, Elisabeth; Delpech, Guillaume; Szabó, Csaba; Remusat, Laurent; Mostefaoui, Smail; Asimow, Paul D.; Antoshechkina, Paula M.; Ghiorso, Mark S.; Boller, Elodie; Guyot, François

    2017-08-01

    Subduction of carbonated crust is widely believed to generate a flux of carbon into the base of the continental lithospheric mantle, which in turn is the likely source of widespread volcanic and non-volcanic CO2 degassing in active tectonic intracontinental settings such as rifts, continental margin arcs and back-arc domains. However, the magnitude of the carbon flux through the lithosphere and the budget of stored carbon held within the lithospheric reservoir are both poorly known. We provide new constraints on the CO2 budget of the lithospheric mantle below the Pannonian Basin (Central Europe) through the study of a suite of xenoliths from the Bakony-Balaton Highland Volcanic Field. Trails of secondary fluid inclusions, silicate melt inclusions, networks of melt veins, and melt pockets with large and abundant vesicles provide numerous lines of evidence that mantle metasomatism affected the lithosphere beneath this region. We obtain a quantitative estimate of the CO2 budget of the mantle below the Pannonian Basin using a combination of innovative analytical and modeling approaches: (1) synchrotron X-ray microtomography, (2) NanoSIMS, Raman spectroscopy and microthermometry, and (3) thermodynamic models (Rhyolite-MELTS). The three-dimensional volumes reconstructed from synchrotron X-ray microtomography allow us to quantify the proportions of all petrographic phases in the samples and to visualize their textural relationships. The concentration of CO2 in glass veins and pockets ranges from 0.27 to 0.96 wt.%, higher than in typical arc magmas (0-0.25 wt.% CO2), whereas the H2O concentration ranges from 0.54 to 4.25 wt.%, on the low end for estimated primitive arc magmas (1.9-6.3 wt.% H2O). Trapping pressures for vesicles were determined by comparing CO2 concentrations in glass to CO2 saturation as a function of pressure in silicate melts, suggesting pressures between 0.69 to 1.78 GPa. These values are generally higher than trapping pressures for fluid inclusions

  7. High-resolution melt and morphological analyses of mealybugs (Hemiptera: Pseudococcidae) from cacao: tools for the control of Cacao swollen shoot virus spread.

    Science.gov (United States)

    Wetten, Andy; Campbell, Colin; Allainguillaume, Joël

    2016-03-01

    Mealybugs (Hemiptera: Coccoidea: Pseudococcidae) are key vectors of badnaviruses, including Cacao swollen shoot virus (CSSV), the most damaging virus affecting cacao (Theobroma cacao L.). The effectiveness of mealybugs as virus vectors is species dependent, and it is therefore vital that CSSV resistance breeding programmes in cacao incorporate accurate mealybug identification. In this work, the efficacy of a CO1-based DNA barcoding approach to species identification was evaluated by screening a range of mealybugs collected from cacao in seven countries. Morphologically similar adult females were characterised by scanning electron microscopy, and then, following DNA extraction, were screened with CO1 barcoding markers. A high degree of CO1 sequence homology was observed for all 11 individual haplotypes, including those accessions from distinct geographical regions. This has allowed the design of a high-resolution melt (HRM) assay capable of rapid identification of the commonly encountered mealybug pests of cacao. HRM analysis readily differentiated between mealybug pests of cacao that cannot necessarily be identified by conventional morphological analysis. This new approach, therefore, has potential to facilitate breeding for resistance to CSSV and other mealybug-transmitted diseases. © 2015 Society of Chemical Industry.

  8. Phase relations study on the melting and crystallization regions of the Bi-2223 high temperature superconductor

    Directory of Open Access Journals (Sweden)

    Alexander Polasek

    2004-09-01

    Full Text Available The melting and solidification behavior of Bi2Sr2Ca2Cu3 O10 (Bi-2223 precursors has been studied. Nominal compositions corresponding to excess of liquid, Ca2CuO3 and CuO have been investigated. Each sample was made by packing a precursor powder into a silver crucible, in order to approximately simulate the situation found in 2223 silver-sheathed tapes. The samples were partially melted and then slow-cooled, being quenched from different temperatures and analyzed through X-ray diffraction (XRD and scanning electron microscopy (SEM/EDS. The precursors decomposed peritectically during melting, forming liquid and solid phases. Very long plates with compositions falling in the vicinity of the 2223 primary phase field formed upon slow-cooling. The 2223 phase may have been formed and the results suggest that long grains of this phase might be obtained by melting and crystallization if the exact peritectic region and the optimum processing conditions are found.

  9. Theoretical study of the aluminum melting curve to very high pressure

    International Nuclear Information System (INIS)

    Moriarty, J.A.; Young, D.A.; Ross, M.

    1984-01-01

    A detailed theoretical study of the Al melting curve from normal melting conditions to pressures in the vicinity of 2 Mbar is presented. The analysis is based on two parallel, but distinct, treatments of the metal: the first from rigorous generalized pseudopotential theory involving first-principles nonlocal pseudopotentials and the second from a parametrized local pseudopotential model which has been accurately fit to first-principles band-theory and experimental equation-of-state data. Both treatments utilize full lattice-dynamical calculations of the phonon free energy in the solid, within the harmonic approximation, and fluid variational theory to obtain the free energy of the liquid. Particular attention is focused on the choice of the reference system in implementing the fluid variational theory. It is shown that in Al the soft-sphere model of Ross produces a lower (and hence more accurate) liquid free energy than either the hard-sphere or one-component-plasma reference systems, and is, moreover, necessary to obtain a reasonable quantitative description of the melting properties. With the soft-sphere system, the two theoretical treatments give results in good overall agreement with each other and with experiment. In particular, melting on the shock Hugoniot is predicted to begin at about 1.2 Mbar and to end at about 1.55 Mbar, in excellent agreement with the recent preliminary measurements of McQueen

  10. Highly porous, low elastic modulus 316L stainless steel scaffold prepared by selective laser melting

    Czech Academy of Sciences Publication Activity Database

    Čapek, Jaroslav; Machová, M.; Fousová, M.; Kubásek, J.; Vojtěch, D.; Fojt, J.; Jablonská, E.; Lipov, J.; Ruml, T.

    2016-01-01

    Roč. 69, Dec (2016), 631–639 ISSN 0928-4931 R&D Projects: GA ČR GBP108/12/G043 Institutional support: RVO:68378271 Keywords : selective laser melting * 316L stainless steel * porous implants * scaffolds Subject RIV: BM - Solid Matter Physics ; Magnetism

  11. A high throughput DNA extraction method with high yield and quality

    Directory of Open Access Journals (Sweden)

    Xin Zhanguo

    2012-07-01

    Full Text Available Abstract Background Preparation of large quantity and high quality genomic DNA from a large number of plant samples is a major bottleneck for most genetic and genomic analyses, such as, genetic mapping, TILLING (Targeting Induced Local Lesion IN Genome, and next-generation sequencing directly from sheared genomic DNA. A variety of DNA preparation methods and commercial kits are available. However, they are either low throughput, low yield, or costly. Here, we describe a method for high throughput genomic DNA isolation from sorghum [Sorghum bicolor (L. Moench] leaves and dry seeds with high yield, high quality, and affordable cost. Results We developed a high throughput DNA isolation method by combining a high yield CTAB extraction method with an improved cleanup procedure based on MagAttract kit. The method yielded large quantity and high quality DNA from both lyophilized sorghum leaves and dry seeds. The DNA yield was improved by nearly 30 fold with 4 times less consumption of MagAttract beads. The method can also be used in other plant species, including cotton leaves and pine needles. Conclusion A high throughput system for DNA extraction from sorghum leaves and seeds was developed and validated. The main advantages of the method are low cost, high yield, high quality, and high throughput. One person can process two 96-well plates in a working day at a cost of $0.10 per sample of magnetic beads plus other consumables that other methods will also need.

  12. Establishment of a simple and rapid identification method for Listeria spp. by using high-resolution melting analysis, and its application in food industry.

    Science.gov (United States)

    Ohshima, Chihiro; Takahashi, Hajime; Phraephaisarn, Chirapiphat; Vesaratchavest, Mongkol; Keeratipibul, Suwimon; Kuda, Takashi; Kimura, Bon

    2014-01-01

    Listeria monocytogenes is the causative bacteria of listeriosis, which has a higher mortality rate than that of other causes of food poisoning. Listeria spp., of which L. monocytogenes is a member, have been isolated from food and manufacturing environments. Several methods have been published for identifying Listeria spp.; however, many of the methods cannot identify newly categorized Listeria spp. Additionally, they are often not suitable for the food industry, owing to their complexity, cost, or time consumption. Recently, high-resolution melting analysis (HRMA), which exploits DNA-sequence differences, has received attention as a simple and quick genomic typing method. In the present study, a new method for the simple, rapid, and low-cost identification of Listeria spp. has been presented using the genes rarA and ldh as targets for HRMA. DNA sequences of 9 Listeria species were first compared, and polymorphisms were identified for each species for primer design. Species specificity of each HRM curve pattern was estimated using type strains of all the species. Among the 9 species, 7 were identified by HRMA using rarA gene, including 3 new species. The remaining 2 species were identified by HRMA of ldh gene. The newly developed HRMA method was then used to assess Listeria isolates from the food industry, and the method efficiency was compared to that of identification by 16S rDNA sequence analysis. The 2 methods were in coherence for 92.6% of the samples, demonstrating the high accuracy of HRMA. The time required for identifying Listeria spp. was substantially low, and the process was considerably simplified, providing a useful and precise method for processing multiple samples per day. Our newly developed method for identifying Listeria spp. is highly valuable; its use is not limited to the food industry, and it can be used for the isolates from the natural environment.

  13. High-temperature vacant lattice site formation in solids and free volumes in melts studied by positron lifetime measurements

    Science.gov (United States)

    Schaefer, H.-E.

    1991-05-01

    In the present paper a concise review is given of the application of positron lifetime measurements to the study of high-temperature vacancies in intermetallic compounds (F 76.3Al 23.7), in metal oxides (NiO), in elemental semiconductors (Si, Ge), and of the oxygen loss or uptake in YBa 2Cu 3O 7-δ. Investigations of free volumes in elemental melts (Al, In, Ge) are included.

  14. High-temperature vacant lattice site formation in solids and free volumes in melts studied by positron lifetime measurements

    International Nuclear Information System (INIS)

    Schaefer, H.E.

    1991-01-01

    In the present paper a concise review is given of the application of positron lifetime measurements to the study of high-temperature vacancies in intermetallic compounds (F 76.3 Al 23.7 ), in metal oxides (NiO), in elemntal semiconductors (Si, Ge), and of the oxygen loss or uptake in YBa 2 Cu 3 O 7-δ . Investigations of free volumes in elemental melts (Al, In, Ge) are included. (orig.)

  15. Trace element partitioning between ilmenite, armalcolite and anhydrous silicate melt: Implications for the formation of lunar high-Ti mare basalts

    NARCIS (Netherlands)

    Kan Parker, M. van; Mason, P.R.D.; Westrenen, W. van

    2011-01-01

    We performed a series of experiments at high pressures and temperatures to determine the partitioning of a wide range of trace elements between ilmenite (Ilm), armalcolite (Arm) and anhydrous lunar silicate melt, to constrain geochemical models of the formation of titanium-rich melts in the Moon.

  16. Rapid Identification of Echinococcus granulosus and E. canadensis Using High-Resolution Melting (HRM) Analysis by Focusing on a Single Nucleotide Polymorphism.

    Science.gov (United States)

    Safa, Ahmad Hosseini; Harandi, Majid Fasihi; Tajaddini, Mohammadhasan; Rostami-Nejad, Mohammad; Mohtashami-Pour, Mehdi; Pestehchian, Nader

    2016-07-22

    High-resolution melting (HRM) is a reliable and sensitive scanning method to detect variation in DNA sequences. We used this method to better understand the epidemiology and transmission of Echinococcus granulosus. We tested the use of HRM to discriminate the genotypes of E. granulosus and E. canadensis. One hundred forty-one hydatid cysts were collected from slaughtered animals in different parts of Isfahan-Iran in 2013. After DNA extraction, the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene was amplified using PCR coupled with the HRM curve. The result of HRM analysis using partial the sequences of cox1 gene revealed that 93, 35, and 2 isolates were identified as G1, G3, and G6 genotypes, respectively. A single nucleotide polymorphism (SNP) was found in locus 9867 of the cox1 gene. This is a critical locus for the differentiation between the G6 and G7 genotypes. In the phylogenic tree, the sample with a SNP was located between the G6 and G7 genotypes, which suggest that this isolate has a G6/G7 genotype. The HRM analysis developed in the present study provides a powerful technique for molecular and epidemiological studies on echinococcosis in humans and animals.

  17. Mutation Scanning in a Single and a Stacked Genetically Modified (GM) Event by Real-Time PCR and High Resolution Melting (HRM) Analysis

    Science.gov (United States)

    Ben Ali, Sina-Elisabeth; Madi, Zita Erika; Hochegger, Rupert; Quist, David; Prewein, Bernhard; Haslberger, Alexander G.; Brandes, Christian

    2014-01-01

    Genetic mutations must be avoided during the production and use of seeds. In the European Union (EU), Directive 2001/18/EC requires any DNA construct introduced via transformation to be stable. Establishing genetic stability is critical for the approval of genetically modified organisms (GMOs). In this study, genetic stability of two GMOs was examined using high resolution melting (HRM) analysis and real-time polymerase chain reaction (PCR) employing Scorpion primers for amplification. The genetic variability of the transgenic insert and that of the flanking regions in a single oilseed rape variety (GT73) and a stacked maize (MON88017 × MON810) was studied. The GT73 and the 5' region of MON810 showed no instabilities in the examined regions. However; two out of 100 analyzed samples carried a heterozygous point mutation in the 3' region of MON810 in the stacked variety. These results were verified by direct sequencing of the amplified PCR products as well as by sequencing of cloned PCR fragments. The occurrence of the mutation suggests that the 5' region is more suitable than the 3' region for the quantification of MON810. The identification of the single nucleotide polymorphism (SNP) in a stacked event is in contrast to the results of earlier studies of the same MON810 region in a single event where no DNA polymorphism was found. PMID:25365178

  18. Rapid isolation of high molecular weight DNA from single dried ...

    African Journals Online (AJOL)

    ANAND

    For studying genetic diversity in populations of predatory coccinellid, Cryptolaemus montrouzieri. Mulsant (Coccinellidae: Coleoptera), our attempts to isolate high quality DNA from individual adult beetle using several previously reported protocols and even modifications were quite unsuccessful as the insect size was small ...

  19. High-temperature abnormal behavior of resistivities for Bi-In melts

    International Nuclear Information System (INIS)

    Xi Yun; Zu Fangqiu; Li Xianfen; Yu Jin; Liu Lanjun; Li Qiang; Chen Zhihao

    2004-01-01

    The patterns of electrical resistivities versus temperature in large temperature range have been studied, using the D.C. four-probe method, for liquid Bi-In alloys (Bi-In(33 wt%), Bi-In(38 wt%), Bi-In(50.5 wt%), Bi-In(66 wt%)). The clear turning point of each resistivity-temperature curves of the liquid Bi-In alloys is observed at the temperature much above the melting point, in which temperature range the resistivity-temperature coefficient increases rapidly. Except for the turning temperature range, the resistivities of Bi-In alloys increase linearly with temperature. Because resistivity is sensitive to the structure, this experiment shows the structural transition in Bi-In melts at the temperature much higher than the liquidus. And it is suggested that there are different Bi-In short-range orderings in different Bi-In melts, so the resistivity-temperature curves have the turns at different temperatures and the resistivity-temperature coefficients are also different

  20. High-temperature corrosion of metals in the salt and metallic melts containing rare earths

    Science.gov (United States)

    Karpov, V. V.; Abramov, A. V.; Zhilyakov, A. Yu.; Belikov, S. V.; Volkovich, V. A.; Polovov, I. B.; Rebrin, O. I.

    2016-09-01

    A complex of independent methods was employed to study the corrosion resistance of molybdenum, zirconium, tantalum and tungsten in chloride, chloride-fluoride and fluoride-oxide melts based on LiCl, CaCl2, NaCl- KCl, LiF, and containing rare earths. Tests were conducted for 30 h at 750-1050 °C. The metals showed excellent corrosion resistance in fused chlorides (the corrosion rates were below 0.0005 g/(m2 h). Despite the presence of chemically active fluoride ions in the chloride-fluoride melts, the metals studied also showed very low corrosion rates, except molybdenum, for which the rate of corrosion was 0,8 g/(m2 h). The corrosion resistance of tantalum was considerably reduced in the fluoride-oxide melts; the corrosion rate was over 1 g/(m2 h) corresponding to the 8-th grade of stability and placing tantalum to the group of "low stability" materials.

  1. Growth from the melt of high-quality In2O3 and Ga2O3 single crystals

    Energy Technology Data Exchange (ETDEWEB)

    Fornari, Roberto; Galazka, Zbigniew; Uecker, Reinhard; Irmscher, Klaus [Leibniz Institute for Crystal Growth, Max-Born-Str. 2, 12489 Berlin (Germany)

    2013-07-01

    Because of their interesting properties semiconducting oxides, in particular Ga{sub 2}O{sub 3} In{sub 2}O{sub 3}, have recently received much attention. However, as they were deposited as films on hetero-substrates their quality was quite poor. The growth of high-quality bulk Ga{sub 2}O{sub 3} and In{sub 2}O{sub 3} and manufacture of the corresponding substrates can allow the deposition of high-quality epilayers with lower residual carrier density and fewer extended defects. For this reason IKZ has undertaken an effort to grow large single crystals of these oxide compounds from the melt. Transparent semiconducting Ga{sub 2}O{sub 3} single crystals with diameter of about 20 mm and 50-60 mm long were grown by the Czochralski method along the b-axis, using an Iridium crucible and a dynamic protective atmosphere to minimize the dissociation of Ga{sub 2}O{sub 3} melt and ingot. In the case of In{sub 2}O{sub 3} the Czochralski technique is not applicable and it was necessary to develop a novel melt growth method. This new method indeed supplied crystals from which oriented substrates could be prepared. In this presentation the melt growth of Ga{sub 2}O{sub 3} and In{sub 2}O{sub 3} single crystals will be reviewed. An important feature of both materials is given by their strong sensitivity to thermal processing: the free carrier concentration and the absorption spectra drastically vary as a function of annealing temperature, duration and ambient. The possible causes are discussed.

  2. High Performance Systolic Array Core Architecture Design for DNA Sequencer

    Directory of Open Access Journals (Sweden)

    Saiful Nurdin Dayana

    2018-01-01

    Full Text Available This paper presents a high performance systolic array (SA core architecture design for Deoxyribonucleic Acid (DNA sequencer. The core implements the affine gap penalty score Smith-Waterman (SW algorithm. This time-consuming local alignment algorithm guarantees optimal alignment between DNA sequences, but it requires quadratic computation time when performed on standard desktop computers. The use of linear SA decreases the time complexity from quadratic to linear. In addition, with the exponential growth of DNA databases, the SA architecture is used to overcome the timing issue. In this work, the SW algorithm has been captured using Verilog Hardware Description Language (HDL and simulated using Xilinx ISIM simulator. The proposed design has been implemented in Xilinx Virtex -6 Field Programmable Gate Array (FPGA and improved in the core area by 90% reduction.

  3. Highly multiplexed targeted DNA sequencing from single nuclei.

    Science.gov (United States)

    Leung, Marco L; Wang, Yong; Kim, Charissa; Gao, Ruli; Jiang, Jerry; Sei, Emi; Navin, Nicholas E

    2016-02-01

    Single-cell DNA sequencing methods are challenged by poor physical coverage, high technical error rates and low throughput. To address these issues, we developed a single-cell DNA sequencing protocol that combines flow-sorting of single nuclei, time-limited multiple-displacement amplification (MDA), low-input library preparation, DNA barcoding, targeted capture and next-generation sequencing (NGS). This approach represents a major improvement over our previous single nucleus sequencing (SNS) Nature Protocols paper in terms of generating higher-coverage data (>90%), thereby enabling the detection of genome-wide variants in single mammalian cells at base-pair resolution. Furthermore, by pooling 48-96 single-cell libraries together for targeted capture, this approach can be used to sequence many single-cell libraries in parallel in a single reaction. This protocol greatly reduces the cost of single-cell DNA sequencing, and it can be completed in 5-6 d by advanced users. This single-cell DNA sequencing protocol has broad applications for studying rare cells and complex populations in diverse fields of biological research and medicine.

  4. High Efficiency Hydrodynamic DNA Fragmentation in a Bubbling System.

    Science.gov (United States)

    Li, Lanhui; Jin, Mingliang; Sun, Chenglong; Wang, Xiaoxue; Xie, Shuting; Zhou, Guofu; van den Berg, Albert; Eijkel, Jan C T; Shui, Lingling

    2017-01-18

    DNA fragmentation down to a precise fragment size is important for biomedical applications, disease determination, gene therapy and shotgun sequencing. In this work, a cheap, easy to operate and high efficiency DNA fragmentation method is demonstrated based on hydrodynamic shearing in a bubbling system. We expect that hydrodynamic forces generated during the bubbling process shear the DNA molecules, extending and breaking them at the points where shearing forces are larger than the strength of the phosphate backbone. Factors of applied pressure, bubbling time and temperature have been investigated. Genomic DNA could be fragmented down to controllable 1-10 Kbp fragment lengths with a yield of 75.30-91.60%. We demonstrate that the ends of the genomic DNAs generated from hydrodynamic shearing can be ligated by T4 ligase and the fragmented DNAs can be used as templates for polymerase chain reaction. Therefore, in the bubbling system, DNAs could be hydrodynamically sheared to achieve smaller pieces in dsDNAs available for further processes. It could potentially serve as a DNA sample pretreatment technique in the future.

  5. Progress in atomizing high melting intermetallic titanium based alloys by means of a novel plasma melting induction guiding gas atomization facility (PIGA)

    Energy Technology Data Exchange (ETDEWEB)

    Gerling, R.; Schimansky, F.P.; Wagner, R. [GKSS-Forschungszentrum Geesthacht GmbH (Germany). Inst. fuer Werkstofforschung

    1994-12-31

    For the production of intermetallic titanium based alloy powders a novel gas atomization facility has been put into operation: By means of a plasma torch the alloy is melted in a water cooled copper crucible in skull melting technique. To the tap hole of the crucible, a novel transfer system is mounted which forms a thin melt stream and guides it into the gas nozzle. This transfer system consists of a ceramic free induction heated water cooled copper funnel. Gas atomization of {gamma}-TiAl (melting temperature 1400 C) and Ti{sub 5}Si{sub 3} (2130 C) proved the possibility to produce ceramic free pre-alloyed powders with this novel facility. The TiAl powder particles are spherical; about 20 wt.% are smaller than 45 {mu}m. The oxygen and copper pick up during atomization do not exceed 250 and 35 {mu}g/g respectively. The Ti{sub 5}Si{sub 3} powder particles are almost spherical. Only about 10 wt.% are <45 {mu}m whereas the O{sub 2} and Cu contamination is also kept at a very low level (250 and 20 {mu}g/g respectively). (orig.)

  6. Detection of Sequence Polymorphism in Rubus Occidentalis L. Monomorphic Microsatellite Markers by High Resolution Melting

    Science.gov (United States)

    Microsatellite, or simple sequence repeat (SSR) markers, are valuable as co-dominant genetic markers with a variety of applications such as DNA fingerprinting, linkage mapping, and population structure analysis. Development of microsatellite primers through the identification of appropriate repeate...

  7. Effect of Feed Melting, Temperature History and Minor Component Addition on Spinel Crystallization in High-Level Waste Glass

    International Nuclear Information System (INIS)

    Izak, Pavel; Hrma, Pavel R.; Arey, Bruce W.; Plaisted, Trevor J.

    2001-01-01

    This study was undertaken to help design mathematical models for high-level waste (HLW) glass melter that simulate spinel behavior in molten glass. Spinel, (Fe,Ni,Mn) (Fe,Cr)2O4, is the primary solid phase that precipitates from HLW glasses containing Fe and Ni in sufficient concentrations. Spinel crystallization affects the anticipated cost and risk of HLW vitrification. To study melting reactions, we used simulated HLW feed, prepared with co-precipitated Fe, Ni, Cr, and Mn hydroxides. Feed samples were heated up at a temperature-increase rate (4C/min) close to that which the feed experiences in the HLW glass melter. The decomposition, melting, and dissolution of feed components (such as nitrates, carbonates, and silica) and the formation of intermediate crystalline phases (spinel, sodalite (Na8(AlSiO4)6(NO2)2), and Zr-containing minerals) were characterized using evolved gas analysis, volume-expansion measurement, optical microscope, scanning electron microscope, thermogravimetric analysis, differential scanning calorimetry, and X-ray diffraction. Nitrates and quartz, the major feed components, converted to a glass-forming melt by 880C. A chromium-free spinel formed in the nitrate melt starting from 520C and Sodalite, a transient product of corundum dissolution, appeared above 600C and eventually dissolved in glass. To investigate the effects of temperature history and minor components (Ru,Ag, and Cu) on the dissolution and growth of spinel crystals, samples were heated up to temperatures above liquidus temperature (TL), then subjected to different temperature histories, and analyzed. The results show that spinel mass fraction, crystals composition, and crystal size depend on the chemical and physical makeup of the feed and temperature history

  8. The Silicon Environment in Silica Polymorphs, Aluminosilicate Crystals and Melts: An In Situ High Temperature XAS Study

    International Nuclear Information System (INIS)

    Cormier, L.; Neuville, D. R.; Roux, J.; Ligny, D. de; Henderson, G. S.; Flank, A.-M.; Lagarde, P.

    2007-01-01

    High temperature X-ray absorption spectroscopy at the Si K-edge has been used to obtain in situ information on SiO2 phase transitions upon heating. Important modifications are observed for the XANES spectra of the high temperature polymorphs, in relation to disordering of the SiO4 tetrahedra beyond the short-range correlations. This paper also presents the XANES spectra of anorthite (CaAl2Si2O8) from room temperature up to the melt (1900 K). This study shows the possibilities for determining the Si environment in crystals and glasses up to the liquid state using in situ XANES measurements

  9. Corrosion of inconel in high-temperature borosilicate glass melts containing simulant nuclear waste

    Science.gov (United States)

    Mao, Xianhe; Yuan, Xiaoning; Brigden, Clive T.; Tao, Jun; Hyatt, Neil C.; Miekina, Michal

    2017-10-01

    The corrosion behaviors of Inconel 601 in the borosilicate glass (MW glass) containing 25 wt.% of simulant Magnox waste, and in ZnO, Mn2O3 and Fe2O3 modified Mg/Ca borosilicate glasses (MZMF and CZMF glasses) containing 15 wt.% of simulant POCO waste, were evaluated by dimensional changes, the formation of internal defects and changes in alloy composition near corrosion surfaces. In all three kinds of glass melts, Cr at the inconel surface forms a protective Cr2O3 scale between the metal surface and the glass, and alumina precipitates penetrate from the metal surface or formed in-situ. The corrosion depths of inconel 601 in MW waste glass melt are greater than those in the other two glass melts. In MW glass, the Cr2O3 layer between inconel and glass is fragmented because of the reaction between MgO and Cr2O3, which forms the crystal phase MgCr2O4. In MZMF and CZMF waste glasses the layers are continuous and a thin (Zn, Fe, Ni, B)-containing layer forms on the surface of the chromium oxide layer and prevents Cr2O3 from reacting with MgO or other constituents. MgCr2O4 was observed in the XRD analysis of the bulk MW waste glass after the corrosion test, and ZrSiO4 in the MZMF waste glass, and ZrSiO4 and CaMoO4 in the CZMF waste glass.

  10. GLASS MELTING PHENOMENA, THEIR ORDERING AND MELTING SPACE UTILISATION

    Directory of Open Access Journals (Sweden)

    Němec L.

    2013-12-01

    Full Text Available Four aspects of effective glass melting have been defined – namely the fast kinetics of partial melting phenomena, a consideration of the melting phenomena ordering, high utilisation of the melting space, and effective utilisation of the supplied energy. The relations were defined for the specific melting performance and specific energy consumption of the glass melting process which involve the four mentioned aspects of the process and indicate the potentials of effective melting. The quantity “space utilisation” has been treated in more detail as an aspect not considered in practice till this time. The space utilisation was quantitatively defined and its values have been determined for the industrial melting facility by mathematical modelling. The definitions of the specific melting performance and specific energy consumption have been used for assessment of the potential impact of a controlled melt flow and high space utilisation on the melting process efficiency on the industrial scale. The results have shown that even the partial control of the melt flow, leading to the partial increase of the space utilisation, may considerably increase the melting performance, whereas a decrease of the specific energy consumption was determined to be between 10 - 15 %.

  11. Directed PCR-free engineering of highly repetitive DNA sequences

    Directory of Open Access Journals (Sweden)

    Preissler Steffen

    2011-09-01

    Full Text Available Abstract Background Highly repetitive nucleotide sequences are commonly found in nature e.g. in telomeres, microsatellite DNA, polyadenine (poly(A tails of eukaryotic messenger RNA as well as in several inherited human disorders linked to trinucleotide repeat expansions in the genome. Therefore, studying repetitive sequences is of biological, biotechnological and medical relevance. However, cloning of such repetitive DNA sequences is challenging because specific PCR-based amplification is hampered by the lack of unique primer binding sites resulting in unspecific products. Results For the PCR-free generation of repetitive DNA sequences we used antiparallel oligonucleotides flanked by restriction sites of Type IIS endonucleases. The arrangement of recognition sites allowed for stepwise and seamless elongation of repetitive sequences. This facilitated the assembly of repetitive DNA segments and open reading frames encoding polypeptides with periodic amino acid sequences of any desired length. By this strategy we cloned a series of polyglutamine encoding sequences as well as highly repetitive polyadenine tracts. Such repetitive sequences can be used for diverse biotechnological applications. As an example, the polyglutamine sequences were expressed as His6-SUMO fusion proteins in Escherichia coli cells to study their aggregation behavior in vitro. The His6-SUMO moiety enabled affinity purification of the polyglutamine proteins, increased their solubility, and allowed controlled induction of the aggregation process. We successfully purified the fusions proteins and provide an example for their applicability in filter retardation assays. Conclusion Our seamless cloning strategy is PCR-free and allows the directed and efficient generation of highly repetitive DNA sequences of defined lengths by simple standard cloning procedures.

  12. Device for measuring high temperature heat conductivity of solids and melts

    International Nuclear Information System (INIS)

    Magomedov, Ya.B.; Gadzhiev, G.G.

    1990-01-01

    A modification of a device for measuring heat conductivity by a compensation method when a thermocouple with gadolinium sulfide being used is suggested. Such a device has less error of measurement (8%), wider interval of working temperatures (300-1600K) and it permits to investigate the material in the wide range of heat conductivity values (0.5-30 W/(mxK)). The stainless steel 12Kh18N10T, lanthanum sulfide and melted quartz were used for the device calibration. The results obtained and the literature data on these materials agree well between each other

  13. Properties of structural steels melted out of high-purity charge

    International Nuclear Information System (INIS)

    Marchenko, V.N.; Sergeeva, T.K.; Kondakova, N.K.; Morozov, V.P.; Madorskij, L.L.

    1993-01-01

    A comparative evaluation has been made of impurities, mechanical properties and hydrogen embirittlement parameters for steels type 40Kh and 40KhS produced by electrometallurgical method with the use of direct reduced charge (DR-steels) and melted in an open-hearth furnace. Investigation results have shown that 40Kh and 40KhS Dr-steels have more coarse austenitic grains and experience more complete transformation of martensite into ferritic-pearlitic mixture on tempering. Threshold stresses increase 2.5 times due to purity enhancement at the expense of application of direct reduced charge

  14. Vitrification of radioactive high-level waste by spray calcination and in-can melting

    Science.gov (United States)

    Hanson, M. S.; Bjorklund, W. J.

    1980-07-01

    After several nonradioactive test runs, radioactive waste from the processing of 1.5 t of spent, light water reactor fuel was successfully concentrated, dried and converted to a vitreous product. A total of 97 L of waste glass (in two stainless steel canisters) was produced. The spray calcination process coupled to the in-can melting process, as developed at Pacific Northwest Labortory, was used to vitrify the waste. An effluent system consisting of a variety of condensation of scrubbing steps more than adequately decontaminated the process off gas before it was released to the atmosphere.

  15. Behavior of a corium jet in high pressure melt ejection from a reactor pressure vessel

    International Nuclear Information System (INIS)

    Frid, W.

    1987-01-01

    This report provides results from analytical and experimental investigations on the behavior of a gas supersaturated molten jet expelled from a pressurized vessel. Aero-hydrodynamic stability of liquid jets in gas, stream degassing of molten metals and gas bubble nucleation in molten metals are relevant problems which are addressed in this work. Models are developed for jet expansion, primary breakup of the jet and secondary fragmentation of melt droplets resulting from violent effervescence of dissolved gas. The jet expansion model is based on a general relation for bubble growth which includes both inertia-controlled and diffusion-controlled growth phases. The jet expansion model is able to predict the jet void fraction, jet radius as a function of axial distance from the pressure vessel, bubble size and bubble pressure. The number density of gas bubbles in the melt, which is a basic parameter in the model, was determined experimentally and is about 10 8 per m 3 of liquid. The primary breakup of the jet produces a spray of droplets, about 2-3 mm in diameter. Parametric calculations for a TMLB' reactor accident sequence show that the corium jet is disrupted within a few initial jet diameters from the reactor vessel and that the radius of corium spray at the level of the reactor cavity floor is in the range of 0.8 to 2.6 m. (orig./HP)

  16. Melting curve analysis after T allele enrichment (MelcaTle as a highly sensitive and reliable method for detecting the JAK2V617F mutation.

    Directory of Open Access Journals (Sweden)

    Soji Morishita

    Full Text Available Detection of the JAK2V617F mutation is essential for diagnosing patients with classical myeloproliferative neoplasms (MPNs. However, detection of the low-frequency JAK2V617F mutation is a challenging task due to the necessity of discriminating between true-positive and false-positive results. Here, we have developed a highly sensitive and accurate assay for the detection of JAK2V617F and named it melting curve analysis after T allele enrichment (MelcaTle. MelcaTle comprises three steps: 1 two cycles of JAK2V617F allele enrichment by PCR amplification followed by BsaXI digestion, 2 selective amplification of the JAK2V617F allele in the presence of a bridged nucleic acid (BNA probe, and 3 a melting curve assay using a BODIPY-FL-labeled oligonucleotide. Using this assay, we successfully detected nearly a single copy of the JAK2V617F allele, without false-positive signals, using 10 ng of genomic DNA standard. Furthermore, MelcaTle showed no positive signals in 90 assays screening healthy individuals for JAK2V617F. When applying MelcaTle to 27 patients who were initially classified as JAK2V617F-positive on the basis of allele-specific PCR analysis and were thus suspected as having MPNs, we found that two of the patients were actually JAK2V617F-negative. A more careful clinical data analysis revealed that these two patients had developed transient erythrocytosis of unknown etiology but not polycythemia vera, a subtype of MPNs. These findings indicate that the newly developed MelcaTle assay should markedly improve the diagnosis of JAK2V617F-positive MPNs.

  17. High Efficiency Acetylcholinesterase Immobilization on DNA Aptamer Modified Surfaces

    Directory of Open Access Journals (Sweden)

    Orada Chumphukam

    2014-04-01

    Full Text Available We report here the in vitro selection of DNA aptamers for electric eel acetylcholinesterase (AChE. One selected aptamer sequence (R15/19 has a high affinity towards the enzyme (Kd = 157 ± 42 pM. Characterization of the aptamer showed its binding is not affected by low ionic strength (~20 mM, however significant reduction in affinity occurred at high ionic strength (~1.2 M. In addition, this aptamer does not inhibit the catalytic activity of AChE that we exploit through immobilization of the DNA on a streptavidin-coated surface. Subsequent immobilization of AChE by the aptamer results in a 4-fold higher catalytic activity when compared to adsorption directly on to plastic.

  18. Detection of sdhB Gene Mutations in SDHI-Resistant Isolates of Botrytis cinerea Using High Resolution Melting (HRM) Analysis.

    Science.gov (United States)

    Samaras, Anastasios; Madesis, Panagiotis; Karaoglanidis, George S

    2016-01-01

    Botrytis cinerea , is a high risk pathogen for fungicide resistance development. Pathogen' resistance to SDHIs is associated with several mutations in sdh gene. The diversity of mutations and their differential effect on cross-resistance patterns among SDHIs and the fitness of resistant strains necessitate the availability of a tool for their rapid identification. This study was initiated to develop and validate a high-resolution melting (HRM) analysis for the identification of P225H/F/L//T, N230I, and H272L/R/Y mutations. Based on the sequence of sdh B subunit of resistant and sensitive isolates, a universal primer pair was designed. The specificity of the HRM analysis primers was verified to ensure against the cross-reaction with other fungal species and its sensitivity was evaluated using concentrations of known amounts of mutant's DNA. The melting curve analysis generated nine distinct curve profiles, enabling the discrimination of all the four mutations located at codon 225, the N230I mutation, the three mutations located in codon 272, and the non-mutated isolates (isolates of wild-type sensitivity). Similar results were obtained when DNA was extracted directly from artificially inoculated strawberry fruit. The method was validated by monitoring the presence of sdh B mutations in samples of naturally infected strawberry fruits and stone fruit rootstock seedling plants showing damping-off symptoms. HRM analysis data were compared with a standard PIRA-PCR technique and an absolute agreement was observed suggesting that in both populations the H272R mutation was the predominant one, while H272Y, N230I, and P225H were detected in lower frequencies. The results of the study suggest that HRM analysis can be a useful tool for sensate, accurate, and rapid identification of several sdh B mutations in B. cinerea and it is expected to contribute in routine fungicide resistance monitoring or assessments of the effectiveness of anti-resistance strategies implemented in

  19. Detection of sdhB gene mutations in SDHI-resistant isolates of Botrytis cinerea using high resolution melting (HRM analysis

    Directory of Open Access Journals (Sweden)

    Anastasios Samaras

    2016-11-01

    Full Text Available Botrytis cinerea, is a high-risk pathogen for fungicide resistance development. Pathogen` resistance to SDHIs is associated with several mutations in sdh gene. The diversity of mutations and their differential effect on cross-resistance patterns among SDHIs and the fitness of resistant strains necessitate the availability of a tool for their rapid identification. This study was initiated to develop and validate a high-resolution melting (HRM analysis for the identification of P225H/F/L//T, N230I and H272L/R/Y mutations. Based on the sequence of sdhB subunit of resistant and sensitive isolates, a universal primer pair was designed. The specificity of the HRM analysis primers was verified to ensure against the cross-reaction with other fungal species and its sensitivity was evaluated using concentrations of known amounts of mutant`s DNA. The melting curve analysis generated nine distinct curve profiles, enabling the discrimination of all the 4 mutations located at codon 225, the N230I mutation, the 3 mutations located in codon 272 and the non mutated isolates (isolates of wild type sensitivity. Similar results were obtained when DNA was extracted directly from artificially inoculated strawberry fruit. The method was validated by monitoring the presence of sdhB mutations in samples of naturally infected strawberry fruits and stone fruit rootstock seedling plants showing damping off symptoms. HRM analysis data were compared with a standard PIRA-PCR technique and an absolute agreement was observed suggesting that in both populations the H272R mutation was the predominant one, while H272Y, N230I and P225H were detected in lower frequencies. The results of the study suggest that HRM analysis can be a useful tool for sensate, accurate and rapid identification of several sdhB mutations in B. cinerea and it is expected to contribute in routine fungicide resistance monitoring or assessments of the effectiveness of antiresistance strategies implemented in

  20. The partitioning of barium and lead between silicate melts and aqueous fluids at high pressures and temperatures

    International Nuclear Information System (INIS)

    Bureau, Helene; Menez, Benedicte; Khodja, Hicham; Daudin, Laurent; Gallien, Jean-Paul; Massare, Dominique; Shaw, Cliff; Metrich, Nicole

    2003-01-01

    The origin of subduction-related magmas is still a matter of debate in the Earth Sciences. These magmas are characterised by their distinctive trace element compositions compared to magmas from other tectonic settings, e.g. mid-ocean ridges or rifts. The distinct trace element composition of these magmas is generally attributed to alteration of the source region by a contaminating agent: either a silicate melt or a hydrous fluid, possibly chlorine-enriched. In this study, we have used μPIXE (proton induced X-ray emission) to analyse synthetic samples obtained from a micro-experimental petrology study that aims to determine the partitioning behaviour of two key elements, Ba and Pb, between silicate melt and both pure water and saline fluids. Our experiments were performed at high-pressure (>0.34-1.53 GPa) and high-temperature (697-1082 deg. C) in a hydrothermal diamond anvil cell, that was used as a transparent rapid quench autoclave. We observed that at high pressure and temperature, in the presence of pure water, Ba and Pb are not strongly fractionated into one phase or the other. The partition coefficient of Pb is ranging from 0.46 to 1.28. Results from one experiment performed at 0.83 GPa and 847 deg. C, in the presence of a saline fluid indicate that the presence of Cl induces strong fractionation of Pb and moderate fractionation of Ba both into the silicate melt. In addition, our data indicate that Cl is strongly partitioned into the fluid phase

  1. In situ study at high pressure and temperature of the environment of water in hydrous Na and Ca aluminosilicate melts and coexisting aqueous fluids

    Science.gov (United States)

    Le Losq, Charles; Dalou, Célia; Mysen, Bjorn O.

    2017-07-01

    The bonding and speciation of water dissolved in Na silicate and Na and Ca aluminosilicate melts were inferred from in situ Raman spectroscopy of the samples, in hydrothermal diamond anvil cells, while at crustal temperature and pressure conditions. Raman data were also acquired on Na silicate and Na and Ca aluminosilicate glasses, quenched from hydrous melts equilibrated at high temperature and pressure in a piston cylinder apparatus. In the hydrous melts, temperature strongly influences O-H stretching ν(O-H) signals, reflecting its control on the bonding of protons between different molecular complexes. Pressure and melt composition effects are much smaller and difficult to discriminate with the present data. However, the chemical composition of the melt + fluid system influences the differences between the ν(O-H) signals from the melts and the fluids and, hence, between their hydrogen partition functions. Quenching modifies the O-H stretching signals: strong hydrogen bonds form in the glasses below the glass transition temperature Tg, and this phenomenon depends on glass composition. Therefore, glasses do not necessarily record the O-H stretching signal shape in melts near Tg. The melt hydrogen partition function thus cannot be assessed with certainty using O-H stretching vibration data from glasses. From the present results, the ratio of the hydrogen partition functions of hydrous silicate melts and aqueous fluids mostly depends on temperature and the bulk melt + fluid system chemical composition. This implies that the fractionation of hydrogen isotopes between magmas and aqueous fluids in water-saturated magmatic systems with differences in temperature and bulk chemical composition will be different.

  2. Characterization of high-purity niobium structures fabricated using the electron beam melting process

    Science.gov (United States)

    Terrazas Najera, Cesar Adrian

    Additive Manufacturing (AM) refers to the varied set of technologies utilized for the fabrication of complex 3D components from digital data in a layer-by-layer fashion. The use of these technologies promises to revolutionize the manufacturing industry. The electron beam melting (EBM) process has been utilized for the fabrication of fully dense near-net-shape components from various metallic materials. This process, catalogued as a powder bed fusion technology, consists of the deposition of thin layers (50 - 120microm) of metallic powder particles which are fused by the use of a high energy electron beam and has been commercialized by Swedish company Arcam AB. Superconducting radio frequency (SRF) cavities are key components that are used in linear accelerators and other light sources for studies of elemental physics. Currently, cavity fabrication is done by employing different forming processes including deep-drawing and spinning. In both of the latter techniques, a feedstock high-purity niobium sheet with a thickness ranging from 3-4 mm is mechanically deformed and shaped into the desired geometry. In this manner, half cavities are formed that are later joined by electron beam welding (EBW). The welding step causes variability in the shape of the cavity and can also introduce impurities at the surface of the weld interface. The processing route and the purity of niobium are also of utmost importance since the presence of impurities such as inclusions or defects can be detrimental for the SRF properties of cavities. The focus of this research was the use of the EBM process in the manufacture of high purity niobium parts with potential SRF applications. Reactor grade niobium was plasma atomized and used as the precursor material for fabrication using EBM. An Arcam A2 system was utilized for the fabrication. The system had all internal components of the fabrication chamber replaced and was cleaned to prevent contamination of niobium powder. A mini-vat, developed at

  3. High resolution melting detects sequence polymorphism in rubus occidentalis L. monomorphic microsatellite markers

    Science.gov (United States)

    Microsatellite, or simple sequence repeat (SSR) markers, are valuable as co-dominant genetic markers with a variety of applications such as DNA fingerprinting, linkage mapping, and population structure analysis. However, primer pairs designed from the regions that flank SSRs often generate fragment...

  4. Rheological Link Between Polymer Melts with a High Molecular Weight Tail and Enhanced Formation of Shish-Kebabs

    DEFF Research Database (Denmark)

    Wingstrand, Sara Lindeblad; Shen, Bo; Kornfield, Julie A.

    2017-01-01

    Presence of an ultra high molecular weight (UHMw) fraction in flowingpolymer melts is known to facilitate formation of oriented crystalline structures significantly. The UHMw fraction manifests itself as a minor tail in the molar mass distribution and is hardly detectable in the canonical...... a clear increase in extensional stress that is directly correlated with the crystalline orientation of the quenched samples. Extensional rheology, particularly, in combination with linear creep measurements, thus, enables the conformational evolution of the UHMw-tail to be studied and linked...

  5. High Resolution Melting (HRM) analysis for mutation screening of RGSL1,RGS16 and RGS8 in breast cancer

    DEFF Research Database (Denmark)

    Wiechec, Emilia; Wiuf, Carsten Henrik; Overgaard, Jens

    2011-01-01

    coding exons of RGSL1, RGS16, and RGS8 in tumors from 200 breast cancer patients. All sequence variants detected by HRM resulted in abnormal shape of the melting curves. The identified mutations and known single nucleotide polymorphisms (SNP) were subsequently confirmed by sequencing, and distribution...... cancer patients. In addition, a total of seven known SNPs were identified in this study. Genotype distributions were not significantly different between breast cancer patients and controls. CONCLUSIONS AND IMPACT: Identification of novel mutations within RGSL1 provides a new insight...... into the pathophysiology of breast cancer. Moreover, the HRM analysis represents a reliable and highly sensitive method for mutation scanning of multiple exons....

  6. Lattice stability and high-pressure melting mechanism of dense hydrogen up to 1.5 TPa

    KAUST Repository

    Geng, Hua Y.; Hoffmann, R.; Wu, Q.

    2015-01-01

    to estimate the well depth and the potential barriers that must be overcome when the crystal melts. Inclusion of nuclear quantum effects (NQE) using path integral molecular dynamics (PIMD) predicts that both superheating limit and melting temperature

  7. Microstructural Characterization of Melt Extracted High-Nb-Containing TiAl-Based Fiber

    Directory of Open Access Journals (Sweden)

    Shuzhi Zhang

    2017-02-01

    Full Text Available The microstructure of melt extracted Ti-44Al-8Nb-0.2W-0.2B-1.5Si fiber were investigated. When the rotation speed increased from 2000 to 2600 r/min, the appearance of the wire was uniform with no Rayleigh-wave default. The structure was mainly composed of fine α2 (α phase dendritic crystal and a second phase between dendrite arms and grain boundaries. The precipitated second phases were confirmed to be Ti5Si3 from the eutectic reaction L→Ti5Si3 + α and TiB. As the lower content of Si and higher cooling rate, a divorced eutectic microstructure was obtained. Segregation of Ti, Nb, B, Si, and Al occurred during rapid solidification.

  8. Study of the Melting Latent Heat of Semicrystalline PVDF applied to High Gamma Dose Dosimetry

    Energy Technology Data Exchange (ETDEWEB)

    Batista, Adriana S.M. [Departamento de Anatomia e Imagem - IMA, Universidade Federal de Minas Gerais, Av. Prof. Alfredo Balena, 190, 30130-100, Belo Horizonte, MG (Brazil); Gual, Maritza R.; Faria, Luiz O. [Centro de Desenvolvimento da Tecnologia Nuclear - CDTN, Av. Antonio Carlos 6627, C.P. 941, 31270-901, Belo Horizonte, MG (Brazil); Lima, Claubia P.B. [Departamento de Engenharia Nuclear - DEN, Universidade Federal de Minas Gerais - UFMG, Av. Antonio Carlos 6627, 31270-970 Belo Horizonte, MG (Brazil)

    2015-07-01

    Poly(vinylidene fluoride) homopolymers [PVDF] homopolymers were irradiated with gamma doses ranging from 0.5 to 2.75 MGy. Differential scanning calorimetry (DSC) and FTIR spectrometry were used in order to study the effects of gamma radiation in the amorphous and crystalline polymer structures. The FTIR data revealed absorption bands at 1730 and 1853 cm{sup -1} which were attributed to the stretch of C=O bonds, at 1715 and 1754 cm{sup -1} which were attributed to the C=C stretching and at 3518, 3585 and 3673 cm{sup -1} which were associated with NH stretch of NH{sub 2} and OH. The melting latent heat (LM) measured by DSC was used to construct an unambiguous relationship with the delivered dose. Regression analyses revealed that the best mathematical function that fits the experimental calibration curve is a 4-degree polynomial function, with an adjusted Rsquare of 0.99817. (authors)

  9. Reactive nanophase oxide additions to melt-processed high-{Tc} superconductors

    Energy Technology Data Exchange (ETDEWEB)

    Goretta, K.C.; Brandel, B.P.; Lanagan, M.T.; Hu, J.; Miller, D.J.; Sengupta, S. [Argonne National Lab., IL (United States); Parker, J.C.; Ali, M.N. [Nanophase Technologies Corp., Darien, IL (United States); Chen, Nan [Illinois Superconductor Corp., Evanston, IL (United States)

    1994-10-01

    Nanophase TiO{sub 2} and Al{sub 2}O{sub 3} powders were synthesized by a vapor-phase process and mechanically mixed with stoichiometric YBa{sub 2}Cu{sub 3}O{sub x} and TlBa{sub 2}Ca{sub 2}Cu{sub 3}O{sub x} powders in 20 mole % concentrations. Pellets produced from powders with and without nanophase oxides were heated in air or O{sub 2} above the peritectic melt temperature and slow-cooled. At 4.2 K, the intragranular critical current density (J{sub c}) increased dramatically with the oxide additions. At 35--50 K, effects of the oxide additions were positive, but less pronounced. At 77 K, the additions decreased J{sub c}, probably because of inducing a depresion of the transition temperature.

  10. Design and analysis of high current DC power supply for vacuum arc melting furnace

    International Nuclear Information System (INIS)

    Adhikary, Santu; Sharma, Vishnu Kumar; Sharma, Archana

    2015-01-01

    Vacuum Arc furnace (VAR), is used for melting of ingot in many industrial units. Till now in many industries the existing power supply for VAR is based on magnetic amplifier, which is a lossy component. Thus an efficient topology is needed to develop as a suitable alternative for the existing power supply. Basically Arc in electrical furnace is an unstable phenomena, it has drooping characteristic in nature so to stabilize the arc we need a power supply across the load (arc), which is more drooping in nature than arc characteristics. So this paper highlights the stability and response analysis of several alternative topologies and Stabilization of arc using the feedback and firing angle control in MATLAB. The work also covers comparison among those topologies to choose the optimized topology as a suitable alternative of the existing magnetic amplifier based power supply and the detail design of the proposed topology with a tested trail circuit in PROTEUS. (author)

  11. Switchable DNA interfaces for the highly sensitive detection of label-free DNA targets.

    Science.gov (United States)

    Rant, Ulrich; Arinaga, Kenji; Scherer, Simon; Pringsheim, Erika; Fujita, Shozo; Yokoyama, Naoki; Tornow, Marc; Abstreiter, Gerhard

    2007-10-30

    We report a method to detect label-free oligonucleotide targets. The conformation of surface-tethered probe nucleic acids is modulated by alternating electric fields, which cause the molecules to extend away from or fold onto the biased surface. Binding (hybridization) of targets to the single-stranded probes results in a pronounced enhancement of the layer-height modulation amplitude, monitored optically in real time. The method features an exceptional detection limit of <3 x 10(8) bound targets per cm(2) sensor area. Single base-pair mismatches in the sequences of DNA complements may readily be identified; moreover, binding kinetics and binding affinities can be determined with high accuracy. When driving the DNA to oscillate at frequencies in the kHz regime, distinct switching kinetics are revealed for single- and double-stranded DNA. Molecular dynamics are used to identify the binding state of molecules according to their characteristic kinetic fingerprints by using a chip-compatible detection format.

  12. High temperature (salt melt) corrosion tests with ceramic-coated steel

    Energy Technology Data Exchange (ETDEWEB)

    Schütz, Adelheid [University Bayreuth, Metals and Alloys, Ludwig-Thoma-Str. 36b, D-95447 Bayreuth (Germany); Günthner, Martin; Motz, Günter [University Bayreuth, Ceramic Materials Engineering, L.-Thoma-Str. 36b, D-95447 Bayreuth (Germany); Greißl, Oliver [EnBW Kraftwerke AG, Schelmenwasenstraße 13-15, D-70567 Stuttgart (Germany); Glatzel, Uwe, E-mail: uwe.glatzel@uni-bayreuth.de [University Bayreuth, Metals and Alloys, Ludwig-Thoma-Str. 36b, D-95447 Bayreuth (Germany)

    2015-06-01

    Thermal recycling of refuse in waste-to-energy plants reduces the problems connected to waste disposal, and is an alternative source of electric energy. However, the combustion process in waste incinerators results in a fast degradation of the steam-carrying superheater steel tubes by corrosive attack and abrasive wear. Higher firing temperatures are used to increase their efficiency but lead to higher corrosion rates. It is more economical to apply protective coatings on the superheater steel tubes than to replace the base material. In-situ tests were conducted in a waste-to-energy plant first in order to identify and quantify all involved corrosive elements. Laboratory scale experiments with salt melts were developed accordingly. The unprotected low-alloyed steel displayed substantial local corrosion. Corrosion was predominant along the grain boundaries of α-ferrite. The corrosion rate was further increased by FeCl{sub 3} and a mixture of HCL and FeCl{sub 3}. Coatings based on pre-ceramic polymers with specific filler particles were engineered to protect superheater tubes. Tests proved their suitability to protect low-alloYed steel tubes from corrosive attack under conditions typical for superheaterS in waste incinerators, rendering higher firing temperatures in waste-to-energy plants possible. - Highlights: • Corrosion wall thickness losses of 400 μm/2 weeks occurred in a waste incinerator. • Abrasion is a major problem on superheater tubes in waste incinerators. • Laboratory salt melt tests can simulate metal corrosion in waste incinerators. • Corrosion protection coatings for steel (temperature: max. 530 °C) were developed. • Higher steam temperatures are possible in WIs with the developed coatings.

  13. Fabrication and heat treatment of high strength Al-Cu-Mg alloy processed using selective laser melting

    Science.gov (United States)

    Zhang, Hu; Zhu, Haihong; Nie, Xiaojia; Qi, Ting; Hu, Zhiheng; Zeng, Xiaoyan

    2016-04-01

    The proposed paper illustrates the fabrication and heat treatment of high strength Al-Cu-Mg alloy produced by selective laser melting (SLM) process. Al-Cu-Mg alloy is one of the heat treatable aluminum alloys regarded as difficult to fusion weld. SLM is an additive manufacturing technique through which components are built by selectively melting powder layers with a focused laser beam. The process is characterized by short laser-powder interaction times and localized high heat input, which leads to steep thermal gradients, rapid solidification and fast cooling. In this research, 3D Al-Cu-Mg parts with relative high density of 99.8% are produced by SLM from gas atomized powders. Room temperature tensile tests reveal a remarkable mechanical behavior: the samples show yield and tensile strengths of about 276 MPa and 402 MPa, respectively, along with fracture strain of 6%. The effect of solution treatment on microstructure and related tensile properties is examined and the results demonstrate that the mechanical behavior of the SLMed Al-Cu-Mg samples can be greatly enhanced through proper heat treatment. After T4 solution treatment at 540°C, under the effect of precipitation strengthening, the tensile strength and the yield strength increase to 532 MPa and 338 MPa, respectively, and the elongation increases to 13%.

  14. Quantification of damage in DNA recovered from highly degraded samples – a case study on DNA in faeces

    Directory of Open Access Journals (Sweden)

    Eveson J Paige

    2006-08-01

    Full Text Available Abstract Background Poorly preserved biological tissues have become an important source of DNA for a wide range of zoological studies. Measuring the quality of DNA obtained from these samples is often desired; however, there are no widely used techniques available for quantifying damage in highly degraded DNA samples. We present a general method that can be used to determine the frequency of polymerase blocking DNA damage in specific gene-regions in such samples. The approach uses quantitative PCR to measure the amount of DNA present at several fragment sizes within a sample. According to a model of random degradation the amount of available template will decline exponentially with increasing fragment size in damaged samples, and the frequency of DNA damage (λ can be estimated by determining the rate of decline. Results The method is illustrated through the analysis of DNA extracted from sea lion faecal samples. Faeces contain a complex mixture of DNA from several sources and different components are expected to be differentially degraded. We estimated the frequency of DNA damage in both predator and prey DNA within individual faecal samples. The distribution of fragment lengths for each target fit well with the assumption of a random degradation process and, in keeping with our expectations, the estimated frequency of damage was always less in predator DNA than in prey DNA within the same sample (mean λpredator = 0.0106 per nucleotide; mean λprey = 0.0176 per nucleotide. This study is the first to explicitly define the amount of template damage in any DNA extracted from faeces and the first to quantify the amount of predator and prey DNA present within individual faecal samples. Conclusion We present an approach for characterizing mixed, highly degraded PCR templates such as those often encountered in ecological studies using non-invasive samples as a source of DNA, wildlife forensics investigations and ancient DNA research. This method will

  15. Coordination Environments of Highly Charged Cations (Ti, Cr, and Light REE's) in Borosilicate Glass/Melts to 1120C

    Energy Technology Data Exchange (ETDEWEB)

    Farges, Francois; /Museum Natl. Hist. Natur. /Stanford U., Geo. Environ. Sci.; Brown, Gordon E., Jr.; /Stanford U., Geo. Environ Sci. /SLAC, SSRL

    2007-01-02

    The local environments around Ti, Cr, and several light rare-earth elements (La, Ce, and Nd) were investigated by in-situ XANES spectroscopy in a number of complex borosilicate glasses and melts (to 1120 C) that are used for nuclear waste storage. Examination of the high-resolution XANES spectra at the Ti K-edge shows that the average coordination of Ti changes from {approx}5 to {approx}4.5. Cr is dominantly trivalent in the melts studied. However, its average coordination is probably lower in the melt (tetrahedral ?) as revealed by the more intense Cr-K pre-edge feature. Ce also changes its average valence from dominantly +4 to +3.5 upon glass melting. These changes are reversible at T{sub g}, the glass transition temperature ({approx}500-550 C for these glasses). In contrast, the local environments of Nd, Pr, and La are unaffected by melting. Therefore, structural reorganization of these borosilicate glass/melts above T{sub g} is variable, not only in terms of valence (as for Ce) but also speciation (Ti and Cr). Both the ability of B to adopt various coordination geometries (triangular and tetrahedral) and the chemical complexity of the glass/melts explain these changes.

  16. Continuum model of tensile fracture of metal melts and its application to a problem of high-current electron irradiation of metals

    International Nuclear Information System (INIS)

    Mayer, Alexander E.; Mayer, Polina N.

    2015-01-01

    A continuum model of the metal melt fracture is formulated on the basis of the continuum mechanics and theory of metastable liquid. A character of temperature and strain rate dependences of the tensile strength that is predicted by the continuum model is verified, and parameters of the model are fitted with the use of the results of the molecular dynamics simulations for ultra-high strain rates (≥1–10/ns). A comparison with experimental data from literature is also presented for Al and Ni melts. Using the continuum model, the dynamic tensile strength of initially uniform melts of Al, Cu, Ni, Fe, Ti, and Pb within a wide range of strain rates (from 1–10/ms to 100/ns) and temperatures (from melting temperature up to 70–80% of critical temperature) is calculated. The model is applied to numerical investigation of a problem of the high-current electron irradiation of Al, Cu, and Fe targets

  17. Solid Lipid Nanoparticle Formulations of Docetaxel Prepared with High Melting Point Triglycerides: In Vitro and in Vivo Evaluation

    Science.gov (United States)

    2015-01-01

    Docetaxel (DCX) is a second generation taxane. It is approved by the U.S. Food and Drug Administration for the treatment of various types of cancer, including breast, non-small cell lung, and head and neck cancers. However, side effects, including those related to Tween 80, an excipient in current DCX formulations, can be severe. In the present study, we developed a novel solid lipid nanoparticle (SLN) composition of DCX. Trimyristin was selected from a list of high melting point triglycerides as the core lipid component of the SLNs, based on the rate at which the DCX was released from the SLNs and the stability of the SLNs. The trimyristin-based, PEGylated DCX-incorporated SLNs (DCX-SLNs) showed significantly higher cytotoxicity against various human and murine cancer cells in culture, as compared to DCX solubilized in a Tween 80/ethanol solution. Moreover, in a mouse model with pre-established tumors, the new DCX-SLNs were significantly more effective than DCX solubilized in a Tween 80/ethanol solution in inhibiting tumor growth without toxicity, likely because the DCX-SLNs increased the concentration of DCX in tumor tissues, but decreased the levels of DCX in major organs such as liver, spleen, heart, lung, and kidney. DCX-incorporated SLNs prepared with one or more high-melting point triglycerides may represent an improved DCX formulation. PMID:24621456

  18. Identification case of evidence in timber tracing of Pinus radiate, using high-resolution melting (HRM) analysis.

    Science.gov (United States)

    Solano, Jaime; Anabalón, Leonardo; Encina, Francisco

    2016-03-01

    Fast, accurate detection of plant species and their hybrids using molecular tools will facilitate assessment and monitoring of timber tracing evidence. In this study the origin of unknown pine samples is determined for a case of timber theft in the region of Araucania southern Chile. We evaluate the utility of the trnL marker region for species identification applied to pine wood based on High Resolution Melting. This efficient tracing methods can be incorporated into forestry applications such as certification of origin. The object of this work was genotype identification using high-resolution melting (HRM) and trnL approaches for Pinus radiata (Don) in timber tracing evidence. Our results indicate that trnL is a very sensitive marker for delimiting species and HRM analysis was used successfully for genotyping Pinus samples for timber tracing purposes. Genotyping samples by HRM analysis with the trnL1 approach allowed us to differentiate two wood samples from the Pinaceae family: Pinus radiata (Don) and Pseudotsuga menziesii (Mirb.) Franco. The same approach with Pinus trnL wood was not able to discriminate between samples of Pinus radiata, indicating that the samples were genetically indistinguishable, possibly because they have the same genotype at this locus. Timber tracing with HRM analysis is expected to contribute to future forest certification schemes, control of illegal trading, and molecular traceability of Pinus spp. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  19. Identification of TNIP1 Polymorphisms by High Resolution Melting Analysis with Unlabelled Probe: Association with Systemic Lupus Erythematosus

    Directory of Open Access Journals (Sweden)

    Jie Zhang

    2012-01-01

    Full Text Available Background. TNFα-induced protein 3 (TNFAIP3 interacting with protein 1 (TNIP1 acts as a negative regulator of NF-κB and plays an important role in maintaining the homeostasis of immune system. A recent genome-wide association study (GWAS showed that the polymorphism of TNIP1 was associated with the disease risk of SLE in Caucasian. In this study, we investigated whether the association of TNIP1 with SLE was replicated in Chinese population. Methods. The association of TNIP1 SNP rs7708392 (G/C was determined by high resolution melting (HRM analysis with unlabeled probe in 285 SLE patients and 336 healthy controls. Results. A new SNP rs79937737 located on 5 bp upstream of rs7708392 was discovered during the HRM analysis. No association of rs7708392 or rs79937737 with the disease risk of SLE was found. Furthermore, rs7708392 and rs79937737 were in weak linkage disequilibrium (LD. Hypotypes analysis of the two SNPs also showed no association with SLE in Chinese population. Conclusions. High resolution melting analysis with unlabeled probes proves to be a powerful and efficient genotyping method for identifying and screening SNPs. No association of rs7708392 or rs79937737 with the disease risk of SLE was observed in Chinese population.

  20. Comparison of high-intensity sound and mechanical vibration for cleaning porous titanium cylinders fabricated using selective laser melting.

    Science.gov (United States)

    Seiffert, Gary; Hopkins, Carl; Sutcliffe, Chris

    2017-01-01

    Orthopedic components, such as the acetabular cup in total hip joint replacement, can be fabricated using porous metals, such as titanium, and a number of processes, such as selective laser melting. The issue of how to effectively remove loose powder from the pores (residual powder) of such components has not been addressed in the literature. In this work, we investigated the feasibility of two processes, acoustic cleaning using high-intensity sound inside acoustic horns and mechanical vibration, to remove residual titanium powder from selective laser melting-fabricated cylinders. With acoustic cleaning, the amount of residual powder removed was not influenced by either the fundamental frequency of the horn used (75 vs. 230 Hz) or, for a given horn, the number of soundings (between 1 and 20). With mechanical vibration, the amount of residual powder removed was not influenced by the application time (10 vs. 20 s). Acoustic cleaning was found to be more reliable and effective in removal of residual powder than cleaning with mechanical vibration. It is concluded that acoustic cleaning using high-intensity sound has significant potential for use in the final preparation stages of porous metal orthopedic components. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 117-123, 2017. © 2015 The Authors Journal of Biomedical Materials Research Part B: Applied Biomaterials Published by Wiley Periodicals, Inc.

  1. Development of a high speed extrusion concept using a floating screw sleeve for solid-melt-separation

    Science.gov (United States)

    Karrenberg, Gregor; Wortberg, Johannes

    2015-05-01

    The High-Speed-S-Truder with floating screw sleeve is an alternative extrusion concept with solid-melt-separation. A fairly conventional 35 mm screw with a length of 21 D, which is accelerated by a 75 kW gearless, water cooled synchronous drive, conveys the resin into a 60 mm screw sleeve with a length of 10 D. Inside the sleeve the material is plasticizied and discharged into the outer screw channel of the sleeve through radial bores. Only the solid bed remains inside. The development of a melt pool - and thus a decrease of the plasticizing capacity - is avoided. The sleeve is rotated by drag forces only (approximately 10 - 15 % of the screw speed). Due to the low speed of the screw sleeve molten material is conveyed to a 4 D Dynamic Mixing Ring in a gentle manner. The DMRs floating ring and the screw sleeve are directly coupled. The granules in the screw channel are stopped by a barrier on the screw in front of the mixing device. So nearly no unmelted material can pass the system. For temperature management in the plastification and mixing zone a 3-zone heating/air-cooling system is used. Various kinds of experiments with the High-Speed S-Truder were conducted. Reachable throughputs with different types of material (LDPE, LLDPE, PP, PS) have been tested. Also three screw geometries, which are mainly varying in the channel depth, were compared. Experimental results and theoretical background will be described in this paper.

  2. Defect, Microstructure, and Mechanical Property of Ti-6Al-4V Alloy Fabricated by High-Power Selective Laser Melting

    Science.gov (United States)

    Cao, Sheng; Chen, Zhuoer; Lim, Chao Voon Samuel; Yang, Kun; Jia, Qingbo; Jarvis, Tom; Tomus, Dacian; Wu, Xinhua

    2017-12-01

    To improve the selective laser melting (SLM) productivity, a high laser power and accordingly adjusted parameters are employed to facilitate a high build rate. Three distinct processing strategies with incremental build rate are developed for SLM Ti-6Al-4V. Various types of defects are investigated. Further studies were carried out by heat-treatment and hot isostatic pressing to evaluate the influence of microstructure and porosity on mechanical properties. The anisotropic mechanical property in horizontally and vertically build samples were observed, which was attributable to the columnar grains and spatial arrangement of defects. Regardless of anisotropy, a post-SLM heat-treatment at 800°C for 2 h produces a combined high strength and ductility.

  3. Colloidal silica films for high-capacity DNA arrays

    Science.gov (United States)

    Glazer, Marc Irving

    The human genome project has greatly expanded the amount of genetic information available to researchers, but before this vast new source of data can be fully utilized, techniques for rapid, large-scale analysis of DNA and RNA must continue to develop. DNA arrays have emerged as a powerful new technology for analyzing genomic samples in a highly parallel format. The detection sensitivity of these arrays is dependent on the quantity and density of immobilized probe molecules. We have investigated substrates with a porous, "three-dimensional" surface layer as a means of increasing the surface area available for the synthesis of oligonucleotide probes, thereby increasing the number of available probes and the amount of detectable bound target. Porous colloidal silica films were created by two techniques. In the first approach, films were deposited by spin-coating silica colloid suspensions onto flat glass substrates, with the pores being formed by the natural voids between the solid particles (typically 23nm pores, 35% porosity). In the second approach, latex particles were co-deposited with the silica and then pyrolyzed, creating films with larger pores (36 nm), higher porosity (65%), and higher surface area. For 0.3 mum films, enhancements of eight to ten-fold and 12- to 14-fold were achieved with the pure silica films and the films "templated" with polymer latex, respectively. In gene expression assays for up to 7,000 genes using complex biological samples, the high-capacity films provided enhanced signals and performed equivalently or better than planar glass on all other functional measures, confirming that colloidal silica films are a promising platform for high-capacity DNA arrays. We have also investigated the kinetics of hybridization on planar glass and high-capacity substrates. Adsorption on planar arrays is similar to ideal Langmuir-type adsorption, although with an "overshoot" at high solution concentration. Hybridization on high-capacity films is

  4. Polypyrrole–gold nanoparticle composites for highly sensitive DNA detection

    International Nuclear Information System (INIS)

    Spain, Elaine; Keyes, Tia E.; Forster, Robert J.

    2013-01-01

    DNA capture surfaces represent a powerful approach to developing highly sensitive sensors for identifying the cause of infection. Electrochemically deposited polypyrrole, PPy, films have been functionalized with electrodeposited gold nanoparticles to give a nanocomposite material, PPy–AuNP. Thiolated capture strand DNA, that is complementary to the sequence from the pathogen Staphylococcus aureus that causes mammary gland inflammation, was then immobilized onto the gold nanoparticles and any of the underlying gold electrode that is exposed. A probe strand, labelled with horse radish peroxidase, HRP, was then hybridized to the target. The concentration of the target was determined by measuring the current generated by reducing benzoquinone produced by the HRP label. Semi-log plots of the pathogen DNA concentration vs. faradaic current are linear from 150 pM to 1 μM and pM concentrations can be detected without the need for molecular, e.g., PCR or NASBA, amplification. The nanocomposite also exhibits excellent selectivity and single base mismatches in a 30 mer sequence can be detected

  5. Critical factors for assembling a high volume of DNA barcodes

    Science.gov (United States)

    Hajibabaei, Mehrdad; deWaard, Jeremy R; Ivanova, Natalia V; Ratnasingham, Sujeevan; Dooh, Robert T; Kirk, Stephanie L; Mackie, Paula M; Hebert, Paul D.N

    2005-01-01

    Large-scale DNA barcoding projects are now moving toward activation while the creation of a comprehensive barcode library for eukaryotes will ultimately require the acquisition of some 100 million barcodes. To satisfy this need, analytical facilities must adopt protocols that can support the rapid, cost-effective assembly of barcodes. In this paper we discuss the prospects for establishing high volume DNA barcoding facilities by evaluating key steps in the analytical chain from specimens to barcodes. Alliances with members of the taxonomic community represent the most effective strategy for provisioning the analytical chain with specimens. The optimal protocols for DNA extraction and subsequent PCR amplification of the barcode region depend strongly on their condition, but production targets of 100K barcode records per year are now feasible for facilities working with compliant specimens. The analysis of museum collections is currently challenging, but PCR cocktails that combine polymerases with repair enzyme(s) promise future success. Barcode analysis is already a cost-effective option for species identification in some situations and this will increasingly be the case as reference libraries are assembled and analytical protocols are simplified. PMID:16214753

  6. Highly parallel translation of DNA sequences into small molecules.

    Directory of Open Access Journals (Sweden)

    Rebecca M Weisinger

    Full Text Available A large body of in vitro evolution work establishes the utility of biopolymer libraries comprising 10(10 to 10(15 distinct molecules for the discovery of nanomolar-affinity ligands to proteins. Small-molecule libraries of comparable complexity will likely provide nanomolar-affinity small-molecule ligands. Unlike biopolymers, small molecules can offer the advantages of cell permeability, low immunogenicity, metabolic stability, rapid diffusion and inexpensive mass production. It is thought that such desirable in vivo behavior is correlated with the physical properties of small molecules, specifically a limited number of hydrogen bond donors and acceptors, a defined range of hydrophobicity, and most importantly, molecular weights less than 500 Daltons. Creating a collection of 10(10 to 10(15 small molecules that meet these criteria requires the use of hundreds to thousands of diversity elements per step in a combinatorial synthesis of three to five steps. With this goal in mind, we have reported a set of mesofluidic devices that enable DNA-programmed combinatorial chemistry in a highly parallel 384-well plate format. Here, we demonstrate that these devices can translate DNA genes encoding 384 diversity elements per coding position into corresponding small-molecule gene products. This robust and efficient procedure yields small molecule-DNA conjugates suitable for in vitro evolution experiments.

  7. Development and assessment of multiplex high resolution melting assay as a tool for rapid single-tube identification of five Brucella species.

    Science.gov (United States)

    Gopaul, Krishna K; Sells, Jessica; Lee, Robin; Beckstrom-Sternberg, Stephen M; Foster, Jeffrey T; Whatmore, Adrian M

    2014-12-11

    The zoonosis brucellosis causes economically significant reproductive problems in livestock and potentially debilitating disease of humans. Although the causative agent, organisms from the genus Brucella, can be differentiated into a number of species based on phenotypic characteristics, there are also significant differences in genotype that are concordant with individual species. This paper describes the development of a five target multiplex assay to identify five terrestrial Brucella species using real-time polymerase chain reaction (PCR) and subsequent high resolution melt curve analysis. This technology offers a robust and cost effective alternative to previously described hydrolysis-probe Single Nucleotide Polymorphism (SNP)-based species defining assays. Through the use of Brucella whole genome sequencing five species defining SNPs were identified. Individual HRM assays were developed to these target these changes and, following optimisation of primer concentrations, it was possible to multiplex all five assays in a single tube. In a validation exercise using a panel of 135 Brucella strains of terrestrial and marine origin, it was possible to distinguish the five target species from the other species within this panel. The HRM multiplex offers a number of diagnostic advantages over previously described SNP-based typing approaches. Further, and uniquely for HRM, the successful multiplexing of five assays in a single tube allowing differentiation of five Brucella species in the diagnostic laboratory in a cost-effective and timely manner is described. However there are possible limitations to using this platform on DNA extractions direct from clinical material.

  8. Immiscible silicate liquids at high pressure: the influence of melt structure on elemental partitioning

    Energy Technology Data Exchange (ETDEWEB)

    Vicenzi, E [Princeton Materials Laboratory, Princeton, NJ (United States); Green, T H [Macquarie Univ., North Ryde, NSW (Australia); Sie, S H [Commonwealth Scientific and Industrial Research Organisation (CSIRO), North Ryde, NSW (Australia). Div. of Exploration Geoscience

    1994-12-31

    Micro-PIXE analyses have been applied to study partitioning of trace elements between immiscible silicate melts stabilised at 0.5 and 1.0 GPa over a temperature range of 1160-1240 deg C in the system SiO{sub 2}-FeO-Al{sub 2}0{sub 3}-K{sub 2}0 (+P{sub 2}0{sub 5}). The system was doped with a suite of trace elements of geochemical interest: Rb, Ba, Pb, Sr, La, Ce, Sm, Ho, Y, Lu, Th, U, Zr, Hf, Nb and Ta at approximately 200 ppm level for all elements except for the REE`s, Ba and Ta (600-1200 ppm). Trace element partitioning was found to be a complex function of cation field strength (charge/radius{sup 2}). Although field strength is important in determining the nature and degree of partitioning, the authors emphasised that it is only one component of the underlying mechanism for the way in which elements distribute themselves between two silicate liquids. 8 refs., 2 figs.

  9. Immiscible silicate liquids at high pressure: the influence of melt structure on elemental partitioning

    Energy Technology Data Exchange (ETDEWEB)

    Vicenzi, E. [Princeton Materials Laboratory, Princeton, NJ (United States); Green, T.H. [Macquarie Univ., North Ryde, NSW (Australia); Sie, S.H. [Commonwealth Scientific and Industrial Research Organisation (CSIRO), North Ryde, NSW (Australia). Div. of Exploration Geoscience

    1993-12-31

    Micro-PIXE analyses have been applied to study partitioning of trace elements between immiscible silicate melts stabilised at 0.5 and 1.0 GPa over a temperature range of 1160-1240 deg C in the system SiO{sub 2}-FeO-Al{sub 2}0{sub 3}-K{sub 2}0 (+P{sub 2}0{sub 5}). The system was doped with a suite of trace elements of geochemical interest: Rb, Ba, Pb, Sr, La, Ce, Sm, Ho, Y, Lu, Th, U, Zr, Hf, Nb and Ta at approximately 200 ppm level for all elements except for the REE`s, Ba and Ta (600-1200 ppm). Trace element partitioning was found to be a complex function of cation field strength (charge/radius{sup 2}). Although field strength is important in determining the nature and degree of partitioning, the authors emphasised that it is only one component of the underlying mechanism for the way in which elements distribute themselves between two silicate liquids. 8 refs., 2 figs.

  10. Structure and dynamics of confined flexible and unentangled polymer melts in highly adsorbing cylindrical pores

    International Nuclear Information System (INIS)

    Carrillo, Jan-Michael Y.; Sumpter, Bobby G.

    2014-01-01

    Coarse-grained molecular dynamics simulations are used to probe the dynamic phenomena of polymer melts confined in nanopores. The simulation results show excellent agreement in the values obtained for the normalized coherent single chain dynamic structure factor, (S(Q,Δt))/(S(Q,0)) . In the bulk configuration, both simulations and experiments confirm that the polymer chains follow Rouse dynamics. However, under confinement, the Rouse modes are suppressed. The mean-square radius of gyration 〈R g 2 〉 and the average relative shape anisotropy 〈κ 2 〉 of the conformation of the polymer chains indicate a pancake-like conformation near the surface and a bulk-like conformation near the center of the confining cylinder. This was confirmed by direct visualization of the polymer chains. Despite the presence of these different conformations, the average form factor of the confined chains still follows the Debye function which describes linear ideal chains, which is in agreement with small angle neutron scattering experiments (SANS). The experimentally inaccessible mean-square displacement (MSD) of the confined monomers, calculated as a function of radial distance from the pore surface, was obtained in the simulations. The simulations show a gradual increase of the MSD from the adsorbed, but mobile layer, to that similar to the bulk far away from the surface

  11. Ab Initio Investigations of High-Pressure Melting of Dense Lithium

    Science.gov (United States)

    Clay, Raymond; Morales, Miguel; Bonev, Stanimir

    Lithium at ambient conditions is the simplest alkali metal and exhibits textbook nearly-free electron behavior. As the density is increased, however, significant core/valence overlap leads to surprisingly complex chemistry. We have systematically investigated the phase diagram of lithium at pressures ranging between two and six million atmospheres. Through a combination of density functional theory based path-integral and classical molecular dynamics simulations, we have investigated the impact of both nuclear quantum effects and anharmonicity on the melting line and solid phase boundaries. We also investigate how the inclusion of nuclear quantum effects and approximations in the treatment of electronic exchange-correlation impact the robustness of previous predictions of tetrahedral clustering in dense liquid Li. Sandia National Laboratories is a multi-mission laboratory managed and operated by Sandia Corporation, a wholly owned subsidiary of Lockheed Martin Corporation, for the U.S. Department of Energy's National Nuclear Security Administration under contract DE-AC04-94AL85000.

  12. High melt strength, tear resistant blown film based on poly(lactic acid)

    Science.gov (United States)

    Edmonds, Neil R.; Plimmer, Peter N.; Tanner, Chris

    2015-05-01

    A major problem associated with the commercial manufacture of thin films from PLA is inferior processing characteristics on blown film lines compared to low density polyethylene. PLA has poor melt strength (leading to bubble instability) and develops a permanent crease in the flattened film as it exits the tower of the film line. In addition, the thin film product has poor tear strength and an unacceptable `noise' level when converted into flexible packaging. Furthermore, fabricated articles based on PLA are known to show an unattractive tendency toward dimensional instability. This behaviour is associated with `cold crystallization', a phenomenon which also causes exudation of any plasticizer added for improving flexibility. Blow moulded articles based on PLA also exhibit dimensional sensitivity above 60°C. All of these issues have been overcome by the technology described in this paper. This has been accomplished without loss of the valuable compostability characteristic of PLA; this was confirmed by evaluation of film in a commercial composting operation. These results have been achieved through novel reactive compounding technology which: (a) Creates a PLA-rich structure containing long chain crosslinks, (b) generates a low glass transition temperature phase covalently bonded to the PLA structure, and (c) provides a material which performs like LDPE in a blown film manufacturing operation. The technology developed is covered by NZ Patent 580231 (3). The patent is held by UniServices Ltd, The University of Auckland, New Zealand.

  13. Generation and emplacement of shear-related highly mobile crustal melts: the synkinematic leucogranites from the Variscan Tormes Dome, Western Spain

    Science.gov (United States)

    López-Moro, Francisco Javier; López-Plaza, Miguel; Romer, Rolf L.

    2012-07-01

    The Tormes dome consists of S-type granites that intruded into Ordovician augen gneisses and Neoproterozoic-Lower Cambrian metapelites/metagreywackes at different extents of migmatization. S-type granites are mainly equigranular two-mica granites, occurring as: (1) enclave-laden subvertical feeder dykes, (2) small external sill-like bodies with size and shape relations indicative for self-similar pluton growth, and (3) as large pluton bodies, emplaced at higher levels than the external ones. These magmas were highly mobile as it is inferred from the high contents of fluxing components, the disintegration and alignment of pelitic xenoliths in feeder dykes and at the bottom of some sill-like bodies. Field relations relate this 311 Ma magmatism (U-Pb monazite) to the regional shearing of the D3 Variscan event. Partial melting modeling and the relatively high estimated liquidus temperatures indicate biotite-dehydration partial melting (800-840°C and 400-650 MPa) rather than water-fluxed melting, implying that there was no partial melting triggered by externally derived fluids in the shear zones. Instead, the subvertical shear zones favored extraction of melts that formed during the regional migmatization event around 320 Ma. Nd isotope variation among the granites might reflect disequilibrium partial melting or different protoliths. Mass-balance and trace element partial melting modeling strongly suggest two kinds of fertile crustal protoliths: augen gneisses and metapelites. Slight compositional variation among the leucogranites does not reflect different extent of protolith melting but is related to a small amount of fractional crystallization (bodies. The lower extent of fractional crystallization and the higher-pressure emplacement conditions of the sill-like bodies support a more restricted movement through the crust than for batholitic leucogranites.

  14. Study of mechanical, rheological and thermal properties of nanocomposite HMSPP (high melt strength polypropylene) with Brazilian bentonite

    International Nuclear Information System (INIS)

    Fermino, Danilo Marin

    2011-01-01

    This work concerns to the study of the mechanical, thermal and rheological behavior of the nano composite HMSPP - Polypropylene High Melt Strength (obtained at a dose of 12.5 kGy) and a bentonite clay Brazilian Paraiba, known as 'Chocolate' in concentrations of 5 and 10% by weight, comparison of to one American Clay, Cloisite 20A nanocomposite was done. Agent compatibilizer polypropylene-graft, known as maleic anhydride (PP-g-AM) was addict 3% concentration thought technique melt intercalation using a twin-screw extruder and the specimens were prepared by injection process. The mechanical behavior was evaluated by strength, flexural strength and impact tests. The thermal behavior was evaluated by the techniques of differential scanning calorimetry (DSC) and thermogravimetry (TGM). The rheological behavior was evaluated in rheometer. The morphology of the nanocomposites was studied by the technique of scanning electron microscopy (SEM). The organophilic bentonite and the nanocomposites were characterized by X-ray diffraction (XRD) and infrared (FTIR). (author)

  15. A high-resolution melting (HRM) assay for the differentiation between Israeli field and Neethling vaccine lumpy skin disease viruses.

    Science.gov (United States)

    Menasherow, Sophia; Erster, Oran; Rubinstein-Giuni, Marisol; Kovtunenko, Anita; Eyngor, Evgeny; Gelman, Boris; Khinich, Evgeny; Stram, Yehuda

    2016-06-01

    Lumpy skin disease (LSD) is a constant threat to the Middle East including the State of Israel. During vaccination programs it is essential for veterinary services and farmers to be able to distinguish between animals affected by the cattle-borne virulent viruses and vaccinated animals, subsequently affected by the vaccine strain. This study describes an improved high resolution-melting (HRM) test that exploits a 27 base pair (bp) fragment of the LSDV126 extracellular enveloped virion (EEV) gene that is present in field viruses but is absent from the Neethling vaccine strain. This difference leads to ∼0.5 °C melting point change in the HRM assay, when testing the quantitative PCR (qPCR) products generated from the virulent field viruses compared to the attenuated vaccine. By exploiting this difference, it could be shown using the newly developed HRM assay that virus isolated from vaccinated cattle that developed disease symptoms behave similarly to vaccine virus control, indicating that the vaccine virus can induce disease symptoms. This assay is not only in full agreement with the previously published PCR gradient and restriction fragment length polymorphism (RFLP) tests but it is faster with, fewer steps, cheaper and dependable. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Test results on direct containment heating by high-pressure melt ejection into the Surtsey vessel: The TDS test series

    International Nuclear Information System (INIS)

    Allen, M.D.; Blanchat, T.K.; Pilch, M.M.

    1994-08-01

    The Technology Development and Scoping (TDS) test series was conducted to test and develop instrumentation and procedures for performing steam-driven, high-pressure melt ejection (HPME) experiments at the Surtsey Test Facility to investigate direct containment heating (DCH). Seven experiments, designated TDS-1 through TDS-7, were performed in this test series. These experiments were conducted using similar initial conditions; the primary variable was the initial pressure in the Surtsey vessel. All experiments in this test series were performed with a steam driving gas pressure of ≅ 4 MPa, 80 kg of lumina/iron/chromium thermite melt simulant, an initial hole diameter of 4.8 cm (which ablated to a final hole diameter of ≅ 6 cm), and a 1/10th linear scale model of the Surry reactor cavity. The Surtsey vessel was purged with argon ( 2 ) to limit the recombination of hydrogen and oxygen, and gas grab samples were taken to measure the amount of hydrogen produced

  17. Development of a two-step high-resolution melting (HRM) analysis for screening sequence variants associated with resistance to the QoIs, benzimidazoles and dicarboximides in airborne inoculum of Botrytis cinerea.

    Science.gov (United States)

    Chatzidimopoulos, Michael; Ganopoulos, Ioannis; Vellios, Evangelos; Madesis, Panagiotis; Tsaftaris, Athanasios; Pappas, Athanassios C

    2014-11-01

    A rapid, high-resolution melting (HRM) analysis protocol was developed to detect sequence variations associated with resistance to the QoIs, benzimidazoles and dicarboximides in Botrytis cinerea airborne inoculum. HRM analysis was applied directly in fungal DNA collected from air samplers with selective medium. Three and five different genotypes were detected and classified according to their melting profiles in BenA and bos1 genes associated with resistance to benzimidazoles and dicarboximides, respectively. The sensitivity of the methodology was evident in the case of the QoIs, where genotypes varying either by a single nucleotide polymorphism or an additional 1205-bp intron were separated accurately with a single pair of primers. The developed two-step protocol was completed in 82 min and showed reduced variation in the melting curves' formation. HRM analysis rapidly detected the major mutations found in greenhouse strains providing accurate data for successfully controlling grey mould. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  18. High-throughput screening of suppression subtractive hybridization cDNA libraries using DNA microarray analysis

    CSIR Research Space (South Africa)

    Van den Berg, N

    2004-11-01

    Full Text Available Efficient construction of cDNA libraries enriched for differentially expressed transcripts is an important first step in many biological investigations. We present a quantitative procedure for screening cDNA libraries constructed by suppression...

  19. Fundamentals of and experiences with forming by rolling of sintered rods of the high-melting metals molybdenum and tungsten

    International Nuclear Information System (INIS)

    Kohlstrung, G.; Marx, H.; Bresch, J.; Leich, M.; Kalning, I.

    1986-01-01

    An efficient and economical technique for rolling sintered rods of the high-melting metals molybdenum and tungsten which comprises only a minimum of processing steps and provides crucial advantages in comparison with the conventional hammering procedure has been developed for application in medium-sized powder-metallurgical plants. The advantages are, in particular, given with the favourable structure development and the elimination of pores from the sintered bar as a result of the higher degree of deformation, increased labour productivity, savings of electrical energy and consumables, as well as a reduction of the working inconveniencies. Experiences gained in test series as well as in industrial practice indicate that final wire diameters can be obtained with a high material economy, provided that the preceding and subsequent process stages are adapted in the optimum manner dependent on the physical and chemical parameters of the starting material and the strain-hardening behaviour in the process of forming by rolling. (orig.) [de

  20. Formation of recent martian debris flows by melting of near-surface ground ice at high obliquity.

    Science.gov (United States)

    Costard, F; Forget, F; Mangold, N; Peulvast, J P

    2002-01-04

    The observation of small gullies associated with recent surface runoff on Mars has renewed the question of liquid water stability at the surface of Mars. The gullies could be formed by groundwater seepage from underground aquifers; however, observations of gullies originating from isolated peaks and dune crests question this scenario. We show that these landforms may result from the melting of water ice in the top few meters of the martian subsurface at high obliquity. Our conclusions are based on the analogy between the martian gullies and terrestrial debris flows observed in Greenland and numerical simulations that show that above-freezing temperatures can occur at high obliquities in the near surface of Mars, and that such temperatures are only predicted at latitudes and for slope orientations corresponding to where the gullies have been observed on Mars.

  1. Precipitation strengthened high strength, high conductivity Cu-Cr-Nb alloys produced by chill block melt spinning. Final Report Ph.D. Thesis

    Science.gov (United States)

    Ellis, David L.; Michal, Gary M.

    1989-01-01

    A series of Cu-based alloys containing 2 to 10 a/o Cr and 1 to 5 a/o Nb were produced by chill block melt spinning (CBMS). The melt spun ribbons were consolidated and hot rolled to sheet to produce a supersaturated Cu-Cr-Nb solid solution from which the high melting point intermetallic compound Cr2Nb could be precipitated to strengthen the Cu matrix. The results show that the materials possess electrical conductivities in excess of 90 percent that of pure Cu at 200 C and above. The strengths of the Cu-Cr-Nb alloys were much greater than Cu, Cu-0.6 Cr, NARloy-A, and NARloy-Z in the as-melt spun condition. The strengths of the consolidated materials were less than Cu-Cr and Cu-Cr-Zr below 500 C and 600 C respectively, but were significantly better above these temperatures. The strengths of the consolidated materials were greater than NARloy-Z, at all temperatures. The GLIDCOP possessed similar strength levels up to 750 C when the strength of the Cu-Cr-Nb alloys begins to degrade. The long term stability of the Cu-Cr-Nb alloys was measured by the microhardness of aged samples and the growth of precipitates. The microhardness measurements indicate that the alloys overage rapidly, but do not suffer much loss in strength between 10 and 100 hours which confirms the results of the electrical resistivity measurements taken during the aging of the alloys at 500 C. The loss in strength from peak strength levels is significant, but the strength remains exceptionally good. Transmission electron microscopy (TEM) of the as-melt spun samples revealed that Cr2Nb precipitates formed in the liquid Cu during the chill block melt spinning, indicating a very strong driving force for the formation of the precipitates. The TEM of the aged and consolidated materials indicates that the precipitates coarsen considerably, but remain in the submicron range.

  2. Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure.

    OpenAIRE

    Felgner, P L; Gadek, T R; Holm, M; Roman, R; Chan, H W; Wenz, M; Northrop, J P; Ringold, G M; Danielsen, M

    1987-01-01

    A DNA-transfection protocol has been developed that makes use of a synthetic cationic lipid, N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA). Small unilamellar liposomes containing DOTMA interact spontaneously with DNA to form lipid-DNA complexes with 100% entrapment of the DNA, DOTMA facilitates fusion of the complex with the plasma membrane of tissue culture cells, resulting in both uptake and expression of the DNA. The technique is simple, highly reproducible, and eff...

  3. In-situ, high pressure and temperature experimental determination of hydrogen isotope fractionation between coexisting hydrous melt and silicate-saturated aqueous fluid

    Science.gov (United States)

    Mysen, B. O.

    2012-12-01

    Hydrogen isotope fractionation between water-saturated silicate melt and silicate-saturated aqueous fluid has been determined experimentally, in-situ with the samples in the 450-800C and 101-1567 MPa temperature and pressure range, respectively. The temperatures are, therefore higher than those where hydrogen bonding in fluids and melts is important [1]. The experiments were conducted with a hydrothermal diamond anvil cell (HDAC) as the high-temperature/-pressure tool and vibrational spectroscopy to determine D/H fractionation. Compositions were along the haploandesite join, Na2Si4O9 - Na2(NaAl)4O9 [Al/(Al+Si)=0-0.1], and a 50:50 (by volume) H2O:D2O fluid mixture as starting material. Platinum metal was used to enhance equilibration rate. Isotopic equilibrium was ascertained by using variable experimental duration at given temperature and pressure. In the Al-free Na-silicate system, the enthalpy change of the (D/H) equilibrium of fluid is 3.1±0.7 kJ/mol, whereas for coexisting melt, ΔH=0 kJ/mol within error. With Al/(Al+Si)=0.1, ΔH=5.2±0.9 kJ/mol for fluid and near 0 within error for coexisting melt melt. For the exchange equilibrium between melt and fluid, H2O(melt)+D2O(fluid)=H2O(fluid)+D2O(melt), the ΔH=4.6±0.7 and 6.5±0.7 kJ/mol for the two Al-free and Al-bearing compositions, respectively, respectively. The D/H equilibration within fluids and melts and, therefore, D/H partitioning between coexisting fluid and melt reflect the influence of dissolved H2O(D2O) in melts and dissolved silicate components in H2O(D2O) fluid on their structure. The positive temperature- and pressure-dependence of silicate solubility and on silicate structure in silicate-saturated aqueous fluid governs the D/H fractionation in the fluid because increasing silicate solute concentration in fluid results in silicate polymerization [2]. These structural effects may be analogous to observed solute-dependent oxygen isotope fractionation between brine and CO2 [3]. In the temperature

  4. Mixing-to-eruption timescales: an integrated model combining numerical simulations and high-temperature experiments with natural melts

    Science.gov (United States)

    Montagna, Chiara; Perugini, Diego; De Campos, Christina; Longo, Antonella; Dingwell, Donald Bruce; Papale, Paolo

    2015-04-01

    Arrival of magma from depth into shallow reservoirs and associated mixing processes have been documented as possible triggers of explosive eruptions. Quantifying the timing from beginning of mixing to eruption is of fundamental importance in volcanology in order to put constraints about the possible onset of a new eruption. Here we integrate numerical simulations and high-temperature experiment performed with natural melts with the aim to attempt identifying the mixing-to-eruption timescales. We performed two-dimensional numerical simulations of the arrival of gas-rich magmas into shallow reservoirs. We solve the fluid dynamics for the two interacting magmas evaluating the space-time evolution of the physical properties of the mixture. Convection and mingling develop quickly into the chamber and feeding conduit/dyke. Over time scales of hours, the magmas in the reservoir appear to have mingled throughout, and convective patterns become harder to identify. High-temperature magma mixing experiments have been performed using a centrifuge and using basaltic and phonolitic melts from Campi Flegrei (Italy) as initial end-members. Concentration Variance Decay (CVD), an inevitable consequence of magma mixing, is exponential with time. The rate of CVD is a powerful new geochronometer for the time from mixing to eruption/quenching. The mingling-to-eruption time of three explosive volcanic eruptions from Campi Flegrei (Italy) yield durations on the order of tens of minutes. These results are in perfect agreement with the numerical simulations that suggest a maximum mixing time of a few hours to obtain a hybrid mixture. We show that integration of numerical simulation and high-temperature experiments can provide unprecedented results about mixing processes in volcanic systems. The combined application of numerical simulations and CVD geochronometer to the eruptive products of active volcanoes could be decisive for the preparation of hazard mitigation during volcanic unrest.

  5. Management of High-Throughput DNA Sequencing Projects: Alpheus.

    Science.gov (United States)

    Miller, Neil A; Kingsmore, Stephen F; Farmer, Andrew; Langley, Raymond J; Mudge, Joann; Crow, John A; Gonzalez, Alvaro J; Schilkey, Faye D; Kim, Ryan J; van Velkinburgh, Jennifer; May, Gregory D; Black, C Forrest; Myers, M Kathy; Utsey, John P; Frost, Nicholas S; Sugarbaker, David J; Bueno, Raphael; Gullans, Stephen R; Baxter, Susan M; Day, Steve W; Retzel, Ernest F

    2008-12-26

    High-throughput DNA sequencing has enabled systems biology to begin to address areas in health, agricultural and basic biological research. Concomitant with the opportunities is an absolute necessity to manage significant volumes of high-dimensional and inter-related data and analysis. Alpheus is an analysis pipeline, database and visualization software for use with massively parallel DNA sequencing technologies that feature multi-gigabase throughput characterized by relatively short reads, such as Illumina-Solexa (sequencing-by-synthesis), Roche-454 (pyrosequencing) and Applied Biosystem's SOLiD (sequencing-by-ligation). Alpheus enables alignment to reference sequence(s), detection of variants and enumeration of sequence abundance, including expression levels in transcriptome sequence. Alpheus is able to detect several types of variants, including non-synonymous and synonymous single nucleotide polymorphisms (SNPs), insertions/deletions (indels), premature stop codons, and splice isoforms. Variant detection is aided by the ability to filter variant calls based on consistency, expected allele frequency, sequence quality, coverage, and variant type in order to minimize false positives while maximizing the identification of true positives. Alpheus also enables comparisons of genes with variants between cases and controls or bulk segregant pools. Sequence-based differential expression comparisons can be developed, with data export to SAS JMP Genomics for statistical analysis.

  6. High-resolution melting analysis using unlabeled probe and amplicon scanning simultaneously detects several lactase persistence variants

    DEFF Research Database (Denmark)

    Janukonyté, Jurgita; Vestergaard, Else M; Ladefoged, Søren A

    2010-01-01

    Lactase persistence and thereby tolerance to lactose is a common trait in people of Northern European descent. It is linked to the LCT -13910C>T variant located in intron 13 of the MCM6 gene 13.9 kb upstream of the lactase (LCT) gene. In people of African and Middle Eastern descent, lactase...... persistence can be associated with other variants nearby the -13910C>T variant, limiting the use of the -13910C>T-based SNP analysis, e.g. TaqMan assays for the diagnosis of lactose intolerance. Using high-resolution melting analysis, we identified five samples that were heterozygous for the -13915T>G variant...... the -13910C>T and -13915T>G variants in addition to rarer variants surrounding the -13910 site. This new method may contribute to improve the diagnostic performance of the genetic analysis for lactose intolerance....

  7. Improving Understanding of Glacier Melt Contribution to High Asian River Discharge through Collaboration and Capacity Building with High Asian CHARIS Partner Institutions

    Science.gov (United States)

    Armstrong, Richard; Brodzik, Mary Jo; Armstrong, Betsy; Barrett, Andrew; Fetterer, Florence; Hill, Alice; Jodha Khalsa, Siri; Racoviteanu, Adina; Raup, Bruce; Rittger, Karl; Williams, Mark; Wilson, Alana; Ye, Qinghua

    2017-04-01

    The Contribution to High Asia Runoff from Ice & Snow (CHARIS) project uses remote sensing data combined with modeling from 2000 to the present to improve proportional estimates of melt from glaciers and seasonal snow surfaces. Based at the National Snow and Ice Data Center (NSIDC), University of Colorado, Boulder, USA, the CHARIS project objectives are twofold: 1) capacity-building efforts with CHARIS partners from eight High Asian countries to better forecast future availability and vulnerability of water resources in the region, and 2) improving our ability to systematically assess the role of glaciers and seasonal snow in the freshwater resources of High Asia. Capacity-building efforts include working with CHARIS partners from Bhutan, Nepal, India, Pakistan, Afghanistan, Kazakhstan, Kyrgyzstan and Tajikistan. Our capacity-building activities include training, data sharing, supporting fieldwork, graduate student education and infrastructure development. Because of the scarcity of in situ data in this High Asian region, we are using the wealth of available remote sensing data to characterize digital elevation, daily maps of fractional snow-cover, annual maps of glacier and permanent snow cover area and downscaled reanalysis temperature data in snow melt models to estimate the relative proportions of river runoff from glacierized and seasonally snow-covered surfaces. Current collaboration with Qinghua Ye, visiting scientist at NSIDC from the Institute of Tibetan Plateau Research, CAS, focuses on remote sensing methods to detect changes in the mountain cryosphere. Collaboration with our Asian partners supports the systematic analysis of the annual cycle of seasonal snow and glacier ice melt across the High Mountain Asia region. With our Asian partners, we have derived reciprocal benefits, learning from their specialized local knowledge and obtaining access to their in situ data. We expect that the improved understanding of runoff from snow and glacier surfaces will

  8. Bacterial natural transformation by highly fragmented and damaged DNA

    DEFF Research Database (Denmark)

    Overballe-Petersen, Søren; Harms, Klaus; Orlando, Ludovic Antoine Alexandre

    2013-01-01

    for microbes, but not as potential substrate for bacterial evolution. Here, we show that fragmented DNA molecules (≥20 bp) that additionally may contain abasic sites, cross-links, or miscoding lesions are acquired by the environmental bacterium Acinetobacter baylyi through natural transformation. With uptake......DNA molecules are continuously released through decomposition of organic matter and are ubiquitous in most environments. Such DNA becomes fragmented and damaged (often DNA is recognized as nutrient source...... of DNA from a 43,000-y-old woolly mammoth bone, we further demonstrate that such natural transformation events include ancient DNA molecules. We find that the DNA recombination is RecA recombinase independent and is directly linked to DNA replication. We show that the adjacent nucleotide variations...

  9. High LET radiation and mechanism of DNA damage repair

    International Nuclear Information System (INIS)

    Furusawa, Yoshiya

    2004-01-01

    Clarifying the mechanism of repair from radiation damage gives most important information on radiation effects on cells. Approximately 10% of biological experiments groups in Heavy Ion Medical Accelerator in Chiba (HIMAC) cooperative research group has performed the subject. They gave a lot of new findings on the mechanism, and solved some open questions. The reason to show the peak of relative biological effectiveness RBE at around 100-200 keV/μm causes miss-repair of DNA damage. Sub-lethal damage generated by high linear energy transfer (LET) radiation can be repaired fully. Potentially lethal damages by high-LET radiation also repaired, but the efficiency decreased with the LET, and so on. (author)

  10. Rapid, dynamic segregation of core forming melts: Results from in-situ High Pressure- High Temperature X-ray Tomography

    Science.gov (United States)

    Watson, H. C.; Yu, T.; Wang, Y.

    2011-12-01

    The timing and mechanisms of core formation in the Earth, as well as in Earth-forming planetesimals is a problem of significant importance in our understanding of the early evolution of terrestrial planets . W-Hf isotopic signatures in meteorites indicate that core formation in small pre-differentiated planetesimals was relatively rapid, and occurred over the span of a few million years. This time scale is difficult to achieve by percolative flow of the metallic phase through a silicate matrix in textural equilibrium. It has been suggested that during this active time in the early solar system, dynamic processes such as impacts may have caused significant deformation in the differentiating planetesimals, which could lead to much higher permeability of the core forming melts. Here, we have measured the change in permeability of core forming melts in a silicate matrix due to deformation. Mixtures of San Carlos olivine and FeS close to the equilibrium percolation threshold (~5 vol%FeS) were pre-synthesized to achieve an equilibrium microstructure, and then loaded into the rotational Drickamer apparatus at GSE-CARS, sector 13-BMD, at the Advanced Photon Source (Argonne National Laboratory). The samples were subsequently pressed to ~2GPa, and heated to 1100°C. Alternating cycles of rotation to collect X-ray tomography images, and twisting to deform the sample were conducted until the sample had been twisted by 1080°. Qualitative and quantitative analyses were performed on the resulting 3-dimensional x-ray tomographic images to evaluate the effect of shear deformation on permeability and migration velocity. Lattice-Boltzmann simulations were conducted, and show a marked increase in the permeability with increasing deformation, which would allow for much more rapid core formation in planetesimals.

  11. Damage of plasmid DNA by high energy ions

    International Nuclear Information System (INIS)

    Michaelidesova, A.; Pachnerova Brabcova, K.; Davidkova, M.

    2018-01-01

    The aim of the study was to determine the degree of direct DNA damage by high-energy ions, which are one of the components of cosmic rays, and therefore the knowledge of the biological effects of these ions is key to long-term space missions with human crew. The pBR322 plasmid containing 4361 base pairs was used in this study. The aqueous solution of plasmid pBR322 was transferred on ice to Japan to the Heavy Ion Medical Accelerator in Chiba, the Research Center for Charged Particle Therapy. Just before the experiment, the droplets of solution of known concentration were applied to the slides and the water was allowed to evaporate to produce dry DNA samples. Half of the slides were irradiated with 290 MeV/u of carbon ions and a dose rate of 20 Gy/min. The other half of the slides were irradiated with helium nuclei of 150 MeV/hr and a dose rate of 12.6 Gy/min. Both sets of slides were irradiated with doses of 0-1,400 Gy with a 200 Gy step. After irradiation, the samples were re-dissolved in distilled water, frozen and transported on ice to the Czech Republic for processing. Samples were analyzed by agarose gel electrophoresis. The plasmid was evaluated separately to determine the degree of radiation induced lesions and further to incubation with enzymes recognizing basal damage. (authors)

  12. Simultaneous effect of mechanical alloying and arc-melting processes in the microstructure and hardness of an AlCoFeMoNiTi high-entropy alloy

    International Nuclear Information System (INIS)

    Baldenebro-Lopez, F.J.; Herrera-Ramírez, J.M.; Arredondo-Rea, S.P.; Gómez-Esparza, C.D.; Martínez-Sánchez, R.

    2015-01-01

    Highlights: • Multi-component systems of AlCoFeMoNiTi were produced by mechanical alloying. • Consolidated samples were fabricated by two different processing routes, sintering and arc melting. • Effect of routes of consolidation on microstructural evolution and microhardness is reported. • High hardness values are found in consolidated samples. • Alloying elements, grain size, and precipitates have a high effect on microhardness. - Abstract: A nanostructured AlCoFeMoNiTi high entropy alloy was synthesized through the mechanical alloying process. Bulk samples were obtained by two different routes to compare the microstructural evolution and hardness behavior: sintering and arc melting. Through electron microscopy analyses the formation of Mo-rich and Ti-rich phases were identified in the melted sample, while Ti-rich nano-precipitates were observed in the sintered sample. A higher microhardness value was achieved on the sintered sample than for the melted sample. The disadvantage of porosity in the sintered sample in comparison to the melted one was overcome by the hardening effect produced by the mechanical alloying

  13. Simultaneous effect of mechanical alloying and arc-melting processes in the microstructure and hardness of an AlCoFeMoNiTi high-entropy alloy

    Energy Technology Data Exchange (ETDEWEB)

    Baldenebro-Lopez, F.J. [Centro de Investigación en Materiales Avanzados (CIMAV), Laboratorio Nacional de Nanotecnología, Miguel de Cervantes 120, 31109 Chihuahua, Chih. (Mexico); Facultad de Ingeniería Mochis, Universidad Autónoma de Sinaloa, Prol. Ángel Flores y Fuente de Poseidón, S.N., 81223 Los Mochis, Sinaloa (Mexico); Herrera-Ramírez, J.M. [Centro de Investigación en Materiales Avanzados (CIMAV), Laboratorio Nacional de Nanotecnología, Miguel de Cervantes 120, 31109 Chihuahua, Chih. (Mexico); Arredondo-Rea, S.P. [Facultad de Ingeniería Mochis, Universidad Autónoma de Sinaloa, Prol. Ángel Flores y Fuente de Poseidón, S.N., 81223 Los Mochis, Sinaloa (Mexico); Gómez-Esparza, C.D. [Centro de Investigación en Materiales Avanzados (CIMAV), Laboratorio Nacional de Nanotecnología, Miguel de Cervantes 120, 31109 Chihuahua, Chih. (Mexico); Martínez-Sánchez, R., E-mail: roberto.martinez@cimav.edu.mx [Centro de Investigación en Materiales Avanzados (CIMAV), Laboratorio Nacional de Nanotecnología, Miguel de Cervantes 120, 31109 Chihuahua, Chih. (Mexico)

    2015-09-15

    Highlights: • Multi-component systems of AlCoFeMoNiTi were produced by mechanical alloying. • Consolidated samples were fabricated by two different processing routes, sintering and arc melting. • Effect of routes of consolidation on microstructural evolution and microhardness is reported. • High hardness values are found in consolidated samples. • Alloying elements, grain size, and precipitates have a high effect on microhardness. - Abstract: A nanostructured AlCoFeMoNiTi high entropy alloy was synthesized through the mechanical alloying process. Bulk samples were obtained by two different routes to compare the microstructural evolution and hardness behavior: sintering and arc melting. Through electron microscopy analyses the formation of Mo-rich and Ti-rich phases were identified in the melted sample, while Ti-rich nano-precipitates were observed in the sintered sample. A higher microhardness value was achieved on the sintered sample than for the melted sample. The disadvantage of porosity in the sintered sample in comparison to the melted one was overcome by the hardening effect produced by the mechanical alloying.

  14. Chemical synthesis of perfectly isotactic and high melting bacterial poly(3-hydroxybutyrate) from bio-sourced racemic cyclic diolide.

    Science.gov (United States)

    Tang, Xiaoyan; Chen, Eugene Y-X

    2018-06-11

    Bacterial poly(3-hydroxybutyrate) (P3HB) is a perfectly isotactic, crystalline material possessing properties suitable for substituting petroleum plastics, but high costs and low volumes of its production are impractical for commodity applications. The chemical synthesis of P3HB via ring-opening polymerization (ROP) of racemic β-butyrolactone has attracted intensive efforts since the 1960s, but not yet produced P3HB with high isotacticity and molecular weight. Here, we report a route utilizing racemic cyclic diolide (rac-DL) derived from bio-sourced succinate. With stereoselective racemic catalysts, the ROP of rac-DL under ambient conditions produces rapidly P3HB with perfect isotacticity ([mm] > 99%), high melting temperature (T m  = 171 °C), and high molecular weight (M n  = 1.54 × 10 5  g mol -1 , Đ = 1.01). With enantiomeric catalysts, kinetic resolution polymerizations of rac-DL automatically stops at 50% conversion and yields enantiopure (R,R)-DL and (S,S)-DL with >99% e.e. and the corresponding poly[(S)-3HB] and poly[(R)-3HB] with high T m  = 175 °C.

  15. DNA Origami Reorganizes upon Interaction with Graphite: Implications for High-Resolution DNA Directed Protein Patterning

    Directory of Open Access Journals (Sweden)

    Masudur Rahman

    2016-10-01

    Full Text Available Although there is a long history of the study of the interaction of DNA with carbon surfaces, limited information exists regarding the interaction of complex DNA-based nanostructures with the important material graphite, which is closely related to graphene. In view of the capacity of DNA to direct the assembly of proteins and optical and electronic nanoparticles, the potential for combining DNA-based materials with graphite, which is an ultra-flat, conductive carbon substrate, requires evaluation. A series of imaging studies utilizing Atomic Force Microscopy has been applied in order to provide a unified picture of this important interaction of structured DNA and graphite. For the test structure examined, we observe a rapid destabilization of the complex DNA origami structure, consistent with a strong interaction of single-stranded DNA with the carbon surface. This destabilizing interaction can be obscured by an intentional or unintentional primary intervening layer of single-stranded DNA. Because the interaction of origami with graphite is not completely dissociative, and because the frustrated, expanded structure is relatively stable over time in solution, it is demonstrated that organized structures of pairs of the model protein streptavidin can be produced on carbon surfaces using DNA origami as the directing material.

  16. DNA Origami Reorganizes upon Interaction with Graphite: Implications for High-Resolution DNA Directed Protein Patterning

    Science.gov (United States)

    Rahman, Masudur; Neff, David; Green, Nathaniel; Norton, Michael L.

    2016-01-01

    Although there is a long history of the study of the interaction of DNA with carbon surfaces, limited information exists regarding the interaction of complex DNA-based nanostructures with the important material graphite, which is closely related to graphene. In view of the capacity of DNA to direct the assembly of proteins and optical and electronic nanoparticles, the potential for combining DNA-based materials with graphite, which is an ultra-flat, conductive carbon substrate, requires evaluation. A series of imaging studies utilizing Atomic Force Microscopy has been applied in order to provide a unified picture of this important interaction of structured DNA and graphite. For the test structure examined, we observe a rapid destabilization of the complex DNA origami structure, consistent with a strong interaction of single-stranded DNA with the carbon surface. This destabilizing interaction can be obscured by an intentional or unintentional primary intervening layer of single-stranded DNA. Because the interaction of origami with graphite is not completely dissociative, and because the frustrated, expanded structure is relatively stable over time in solution, it is demonstrated that organized structures of pairs of the model protein streptavidin can be produced on carbon surfaces using DNA origami as the directing material. PMID:28335324

  17. Melting temperature of H2, D2, N2 and СH4 under high pressure

    Indian Academy of Sciences (India)

    the analysis indicates the presence of the melting maximum in these solids. ... values of the melting temperature in case of hydrogen up to a pressure of 4800 ... temperature, Tm, will rise with the increase in pressure, reach to a maximum and.

  18. Tungsten disulfide nanosheet and exonuclease III co-assisted amplification strategy for highly sensitive fluorescence polarization detection of DNA glycosylase activity

    International Nuclear Information System (INIS)

    Zhao, Jingjin; Ma, Yefei; Kong, Rongmei; Zhang, Liangliang; Yang, Wen; Zhao, Shulin

    2015-01-01

    Herein, we introduced a tungsten disulfide (WS 2 ) nanosheet and exonuclease III (Exo III) co-assisted signal amplification strategy for highly sensitive fluorescent polarization (FP) assay of DNA glycosylase activity. Two DNA glycosylases, uracil-DNA glycosylase (UDG) and human 8-oxoG DNA glycosylase 1 (hOGG1), were tested. A hairpin-structured probe (HP) which contained damaged bases in the stem was used as the substrate. The removal of damaged bases from substrate by DNA glycosylase would lower the melting temperature of HP. The HP was then opened and hybridized with a FAM dye-labeled single strand DNA (DP), generating a duplex with a recessed 3′-terminal of DP. This design facilitated the Exo III-assisted amplification by repeating the hybridization and digestion of DP, liberating numerous FAM fluorophores which could not be adsorbed on WS 2 nanosheet. Thus, the final system exhibited a small FP signal. However, in the absence of DNA glycosylases, no hybridization between DP and HP was occurred, hampering the hydrolysis of DP by Exo III. The intact DP was then adsorbed on the surface of WS 2 nanosheet that greatly amplified the mass of the labeled-FAM fluorophore, resulting in a large FP value. With the co-assisted amplification strategy, the sensitivity was substantially improved. In addition, this method was applied to detect UDG activity in cell extracts. The study of the inhibition of UDG was also performed. Furthermore, this method is simple in design, easy in implementation, and selective, which holds potential applications in the DNA glycosylase related mechanism research and molecular diagnostics. - Highlights: • A fluorescence polarization strategy for DNA glycosylase activity detection was developed. • The present method was based on WS 2 nanosheet and exonuclease III co-assisted signal amplification. • A high sensitivity and desirable selectivity were achieved. • This method provides a promising universal platform for DNA glycosylase

  19. Rapid and inexpensive body fluid identification by RNA profiling-based multiplex High Resolution Melt (HRM) analysis.

    Science.gov (United States)

    Hanson, Erin K; Ballantyne, Jack

    2013-01-01

    Positive identification of the nature of biological material present on evidentiary items can be crucial for understanding the circumstances surrounding a crime. However, traditional protein-based methods do not permit the identification of all body fluids and tissues, and thus molecular based strategies for the conclusive identification of all forensically relevant biological fluids and tissues need to be developed. Messenger RNA (mRNA) profiling is an example of such a molecular-based approach. Current mRNA body fluid identification assays involve capillary electrophoresis (CE) or quantitative RT-PCR (qRT-PCR) platforms, each with its own limitations. Both platforms require the use of expensive fluorescently labeled primers or probes. CE-based assays require separate amplification and detection steps thus increasing the analysis time. For qRT-PCR assays, only 3-4 markers can be included in a single reaction since each requires a different fluorescent dye. To simplify mRNA profiling assays, and reduce the time and cost of analysis, we have developed single- and multiplex body fluid High Resolution Melt (HRM) assays for the identification of common forensically relevant biological fluids and tissues. The incorporated biomarkers include IL19 (vaginal secretions), IL1F7 (skin), ALAS2 (blood), MMP10 (menstrual blood), HTN3 (saliva) and TGM4 (semen).  The HRM assays require only unlabeled PCR primers and a single saturating intercalating fluorescent dye (Eva Green). Each body-fluid-specific marker can easily be identified by the presence of a distinct melt peak. Usually, HRM assays are used to detect variants or isoforms for a single gene target. However, we have uniquely developed duplex and triplex HRM assays to permit the simultaneous detection of multiple targets per reaction. Here we describe the development and initial performance evaluation of the developed HRM assays. The results demonstrate the potential use of HRM assays for rapid, and relatively inexpensive

  20. Lab-on-a-chip platform for high throughput drug discovery with DNA-encoded chemical libraries

    Science.gov (United States)

    Grünzner, S.; Reddavide, F. V.; Steinfelder, C.; Cui, M.; Busek, M.; Klotzbach, U.; Zhang, Y.; Sonntag, F.

    2017-02-01

    The fast development of DNA-encoded chemical libraries (DECL) in the past 10 years has received great attention from pharmaceutical industries. It applies the selection approach for small molecular drug discovery. Because of the limited choices of DNA-compatible chemical reactions, most DNA-encoded chemical libraries have a narrow structural diversity and low synthetic yield. There is also a poor correlation between the ranking of compounds resulted from analyzing the sequencing data and the affinity measured through biochemical assays. By combining DECL with dynamical chemical library, the resulting DNA-encoded dynamic library (EDCCL) explores the thermodynamic equilibrium of reversible reactions as well as the advantages of DNA encoded compounds for manipulation/detection, thus leads to enhanced signal-to-noise ratio of the selection process and higher library quality. However, the library dynamics are caused by the weak interactions between the DNA strands, which also result in relatively low affinity of the bidentate interaction, as compared to a stable DNA duplex. To take advantage of both stably assembled dual-pharmacophore libraries and EDCCLs, we extended the concept of EDCCLs to heat-induced EDCCLs (hi-EDCCLs), in which the heat-induced recombination process of stable DNA duplexes and affinity capture are carried out separately. To replace the extremely laborious and repetitive manual process, a fully automated device will facilitate the use of DECL in drug discovery. Herein we describe a novel lab-on-a-chip platform for high throughput drug discovery with hi-EDCCL. A microfluidic system with integrated actuation was designed which is able to provide a continuous sample circulation by reducing the volume to a minimum. It consists of a cooled and a heated chamber for constant circulation. The system is capable to generate stable temperatures above 75 °C in the heated chamber to melt the double strands of the DNA and less than 15 °C in the cooled chamber

  1. Disentangled solid state and metastable polymer melt; a solvent free route to high-modulus high-strength tapes and films of UHMWPE

    Science.gov (United States)

    Rastogi, Sanjay

    2013-03-01

    Ultra High Molecular Weight Polyethylene (UHMWPE) having average molar mass greater than a million g/mol is an engineering polymer. Due to its light-weight, high abrasion resistance and biocompatibility it is used for demanding applications such as body armour, prostheses etc. At present, because of its high melt viscosity to achieve the uniaxial/biaxial properties in the form of fibers/films the polymer is processed via solution route where nearly 95wt% of the solvent is used to process 5wt% of the polymer. In past several attempts have been made to process the polymer without using any solvent. However, compared to the solvent processing route the achieved mechanical properties were rather poor. Here we show that by controlled synthesis it is feasible to obtain UHMWPE that could be processed free of solvent to make uniaxial tapes and biaxial films, having unprecedented mechanical properties, exceeding that of the solution spun fibers. We address some of the fundamental aspects of chemistry, physics, rheology and processing for the development of desired morphological features to achieve the ultimate mechanical properties in tapes and films. The paper will also address the metastable melt state obtained on melting of the disentangled crystals and its implication on rheology in linear and nonlinear viscoelastic region. Solid state NMR studies will be applied to establish disentangled state in solid state to the polymerisation conditions. References: Macromolecules 2011, 44(14), 5558-5568; Nature Materials 2005, 4, 635-641; Phys Rev Lett 2006, 96(21), 218303-218205. The authors acknowledge financial support by the Dutch Polymer Institute.

  2. Identification and Evaluation of Single-Nucleotide Polymorphisms in Allotetraploid Peanut (Arachis hypogaea L.) Based on Amplicon Sequencing Combined with High Resolution Melting (HRM) Analysis.

    Science.gov (United States)

    Hong, Yanbin; Pandey, Manish K; Liu, Ying; Chen, Xiaoping; Liu, Hong; Varshney, Rajeev K; Liang, Xuanqiang; Huang, Shangzhi

    2015-01-01

    The cultivated peanut (Arachis hypogaea L.) is an allotetraploid (AABB) species derived from the A-genome (Arachis duranensis) and B-genome (Arachis ipaensis) progenitors. Presence of two versions of a DNA sequence based on the two progenitor genomes poses a serious technical and analytical problem during single nucleotide polymorphism (SNP) marker identification and analysis. In this context, we have analyzed 200 amplicons derived from expressed sequence tags (ESTs) and genome survey sequences (GSS) to identify SNPs in a panel of genotypes consisting of 12 cultivated peanut varieties and two diploid progenitors representing the ancestral genomes. A total of 18 EST-SNPs and 44 genomic-SNPs were identified in 12 peanut varieties by aligning the sequence of A. hypogaea with diploid progenitors. The average frequency of sequence polymorphism was higher for genomic-SNPs than the EST-SNPs with one genomic-SNP every 1011 bp as compared to one EST-SNP every 2557 bp. In order to estimate the potential and further applicability of these identified SNPs, 96 peanut varieties were genotyped using high resolution melting (HRM) method. Polymorphism information content (PIC) values for EST-SNPs ranged between 0.021 and 0.413 with a mean of 0.172 in the set of peanut varieties, while genomic-SNPs ranged between 0.080 and 0.478 with a mean of 0.249. Total 33 SNPs were used for polymorphism detection among the parents and 10 selected lines from mapping population Y13Zh (Zhenzhuhei × Yueyou13). Of the total 33 SNPs, nine SNPs showed polymorphism in the mapping population Y13Zh, and seven SNPs were successfully mapped into five linkage groups. Our results showed that SNPs can be identified in allotetraploid peanut with high accuracy through amplicon sequencing and HRM assay. The identified SNPs were very informative and can be used for different genetic and breeding applications in peanut.

  3. High-resolution melting analysis of the single nucleotide polymorphism hot-spot region in the rpoB gene as an indicator of reduced susceptibility to rifaximin in Clostridium difficile.

    Science.gov (United States)

    Pecavar, Verena; Blaschitz, Marion; Hufnagl, Peter; Zeinzinger, Josef; Fiedler, Anita; Allerberger, Franz; Maass, Matthias; Indra, Alexander

    2012-06-01

    Clostridium difficile, a Gram-positive, spore-forming, anaerobic bacterium, is the main causative agent of hospital-acquired diarrhoea worldwide. In addition to metronidazole and vancomycin, rifaximin, a rifamycin derivative, is a promising antibiotic for the treatment of recurring C. difficile infections (CDI). However, exposure of C. difficile to this antibiotic has led to the development of rifaximin-resistance due to point mutations in the β-subunit of the RNA polymerase (rpoB) gene. In the present study, 348 C. difficile strains with known PCR-ribotypes were investigated for respective single nucleotide polymorphisms (SNPs) within the proposed rpoB hot-spot region by using high-resolution melting (HRM) analysis. This method allows the detection of SNPs by comparing the altered melting behaviour of dsDNA with that of wild-type DNA. Discrimination between wild-type and mutant strains was enhanced by creating heteroduplexes by mixing sample DNA with wild-type DNA, leading to characteristic melting curve shapes from samples containing SNPs in the respective rpoB section. In the present study, we were able to identify 16 different rpoB sequence-types (ST) by sequencing analysis of a 325 bp fragment. The 16 PCR STs displayed a total of 24 different SNPs. Fifteen of these 24 SNPs were located within the proposed 151 bp SNP hot-spot region, resulting in 11 different HRM curve profiles (CP). Eleven SNPs (seven of which were within the proposed hot-spot region) led to amino acid substitutions associated with reduced susceptibility to rifaximin and 13 SNPs (eight of which were within the hot-spot region) were synonymous. This investigation clearly demonstrates that HRM analysis of the proposed SNP hot-spot region in the rpoB gene of C. difficile is a fast and cost-effective method for the identification of C. difficile samples with reduced susceptibility to rifaximin and even allows simultaneous SNP subtyping of the respective C. difficile isolates.

  4. Melting of tantalum at high pressure determined by angle dispersive x-ray diffraction in a double-sided laser-heated diamond-anvil cell

    International Nuclear Information System (INIS)

    Errandonea, D; Somayazulu, M; Haeusermann, D; Mao, H K

    2003-01-01

    The high-pressure and high-temperature phase diagram of Ta has been studied in a laser-heated diamond-anvil cell (DAC) using x-ray diffraction measurements up to 52 deg. GPa and 3800 deg. K. The melting was observed at nine different pressures, the melting temperature being in good agreement with previous laser-heated DAC experiments, but in contradiction with several theoretical calculations and previous piston-cylinder apparatus experiments. A small slope for the melting curve of Ta is estimated (dT m /dP ≅ 24 GPa -1 at 1 deg. bar) and a possible explanation for this behaviour is given. Finally, a P-V -T equation of states is obtained, the temperature dependence of the thermal expansion coefficient and the bulk modulus being estimated

  5. Two high-mobility group box domains act together to underwind and kink DNA

    Energy Technology Data Exchange (ETDEWEB)

    Sánchez-Giraldo, R.; Acosta-Reyes, F. J. [Universitat Politecnica de Catalunya, 08028 Barcelona (Spain); Malarkey, C. S. [University of Colorado School of Medicine, Aurora, CO 80045 (United States); Saperas, N. [Universitat Politecnica de Catalunya, 08028 Barcelona (Spain); Churchill, M. E. A., E-mail: mair.churchill@ucdenver.edu [University of Colorado School of Medicine, Aurora, CO 80045 (United States); Campos, J. L., E-mail: mair.churchill@ucdenver.edu [Universitat Politecnica de Catalunya, 08028 Barcelona (Spain)

    2015-06-30

    The crystal structure of HMGB1 box A bound to an unmodified AT-rich DNA fragment is reported at a resolution of 2 Å. A new mode of DNA recognition for HMG box proteins is found in which two box A domains bind in an unusual configuration generating a highly kinked DNA structure. High-mobility group protein 1 (HMGB1) is an essential and ubiquitous DNA architectural factor that influences a myriad of cellular processes. HMGB1 contains two DNA-binding domains, box A and box B, which have little sequence specificity but have remarkable abilities to underwind and bend DNA. Although HMGB1 box A is thought to be responsible for the majority of HMGB1–DNA interactions with pre-bent or kinked DNA, little is known about how it recognizes unmodified DNA. Here, the crystal structure of HMGB1 box A bound to an AT-rich DNA fragment is reported at a resolution of 2 Å. Two box A domains of HMGB1 collaborate in an unusual configuration in which the Phe37 residues of both domains stack together and intercalate the same CG base pair, generating highly kinked DNA. This represents a novel mode of DNA recognition for HMGB proteins and reveals a mechanism by which structure-specific HMG boxes kink linear DNA.

  6. Two high-mobility group box domains act together to underwind and kink DNA

    International Nuclear Information System (INIS)

    Sánchez-Giraldo, R.; Acosta-Reyes, F. J.; Malarkey, C. S.; Saperas, N.; Churchill, M. E. A.; Campos, J. L.

    2015-01-01

    The crystal structure of HMGB1 box A bound to an unmodified AT-rich DNA fragment is reported at a resolution of 2 Å. A new mode of DNA recognition for HMG box proteins is found in which two box A domains bind in an unusual configuration generating a highly kinked DNA structure. High-mobility group protein 1 (HMGB1) is an essential and ubiquitous DNA architectural factor that influences a myriad of cellular processes. HMGB1 contains two DNA-binding domains, box A and box B, which have little sequence specificity but have remarkable abilities to underwind and bend DNA. Although HMGB1 box A is thought to be responsible for the majority of HMGB1–DNA interactions with pre-bent or kinked DNA, little is known about how it recognizes unmodified DNA. Here, the crystal structure of HMGB1 box A bound to an AT-rich DNA fragment is reported at a resolution of 2 Å. Two box A domains of HMGB1 collaborate in an unusual configuration in which the Phe37 residues of both domains stack together and intercalate the same CG base pair, generating highly kinked DNA. This represents a novel mode of DNA recognition for HMGB proteins and reveals a mechanism by which structure-specific HMG boxes kink linear DNA

  7. An equation of state for high pressure-temperature liquids (RTpress) with application to MgSiO3 melt

    Science.gov (United States)

    Wolf, Aaron S.; Bower, Dan J.

    2018-05-01

    The thermophysical properties of molten silicates at extreme conditions are crucial for understanding the early evolution of Earth and other massive rocky planets, which is marked by giant impacts capable of producing deep magma oceans. Cooling and crystallization of molten mantles are sensitive to the densities and adiabatic profiles of high-pressure molten silicates, demanding accurate Equation of State (EOS) models to predict the early evolution of planetary interiors. Unfortunately, EOS modeling for liquids at high P-T conditions is difficult due to constantly evolving liquid structure. The Rosenfeld-Tarazona (RT) model provides a physically sensible and accurate description of liquids but is limited to constant volume heating paths (Rosenfeld and Tarazona, 1998). We develop a high P-T EOS for liquids, called RTpress, which uses a generalized Rosenfeld-Tarazona model as a thermal perturbation to isothermal and adiabatic reference compression curves. This approach provides a thermodynamically consistent EOS which remains accurate over a large P-T range and depends on a limited number of physically meaningful parameters that can be determined empirically from either simulated or experimental datasets. As a first application, we model MgSiO3 melt representing a simplified rocky mantle chemistry. The model parameters are fitted to the MD simulations of both Spera et al. (2011) and de Koker and Stixrude (2009), recovering pressures, volumes, and internal energies to within 0.6 GPa, 0.1 Å3 , and 6 meV per atom on average (for the higher resolution data set), as well as accurately predicting liquid densities and temperatures from shock-wave experiments on MgSiO3 glass. The fitted EOS is used to determine adiabatic thermal profiles, revealing the approximate thermal structure of a fully molten magma ocean like that of the early Earth. These adiabats, which are in strong agreement for both fitted models, are shown to be sufficiently steep to produce either a center

  8. High level active n+ doping of strained germanium through co-implantation and nanosecond pulsed laser melting

    Science.gov (United States)

    Pastor, David; Gandhi, Hemi H.; Monmeyran, Corentin P.; Akey, Austin J.; Milazzo, Ruggero; Cai, Yan; Napolitani, Enrico; Gwilliam, Russell M.; Crowe, Iain F.; Michel, Jurgen; Kimerling, L. C.; Agarwal, Anuradha; Mazur, Eric; Aziz, Michael J.

    2018-04-01

    Obtaining high level active n+ carrier concentrations in germanium (Ge) has been a significant challenge for further development of Ge devices. By ion implanting phosphorus (P) and fluorine (F) into Ge and restoring crystallinity using Nd:YAG nanosecond pulsed laser melting (PLM), we demonstrate 1020 cm-3 n+ carrier concentration in tensile-strained epitaxial germanium-on-silicon. Scanning electron microscopy shows that after laser treatment, samples implanted with P have an ablated surface, whereas P + F co-implanted samples have good crystallinity and a smooth surface topography. We characterize P and F concentration depth profiles using secondary ion mass spectrometry and spreading resistance profiling. The peak carrier concentration, 1020 cm-3 at 80 nm below the surface, coincides with the peak F concentration, illustrating the key role of F in increasing donor activation. Cross-sectional transmission electron microscopy of the co-implanted sample shows that the Ge epilayer region damaged during implantation is a single crystal after PLM. High-resolution X-ray diffraction and Raman spectroscopy measurements both indicate that the as-grown epitaxial layer strain is preserved after PLM. These results demonstrate that co-implantation and PLM can achieve the combination of n+ carrier concentration and strain in Ge epilayers necessary for next-generation, high-performance Ge-on-Si devices.

  9. Exploiting Process-Related Advantages of Selective Laser Melting for the Production of High-Manganese Steel.

    Science.gov (United States)

    Haase, Christian; Bültmann, Jan; Hof, Jan; Ziegler, Stephan; Bremen, Sebastian; Hinke, Christian; Schwedt, Alexander; Prahl, Ulrich; Bleck, Wolfgang

    2017-01-11

    Metal additive manufacturing has strongly gained scientific and industrial importance during the last decades due to the geometrical flexibility and increased reliability of parts, as well as reduced equipment costs. Within the field of metal additive manufacturing methods, selective laser melting (SLM) is an eligible technique for the production of fully dense bulk material with complex geometry. In the current study, we addressed the application of SLM for processing a high-manganese TRansformation-/TWinning-Induced Plasticity (TRIP/TWIP) steel. The solidification behavior was analyzed by careful characterization of the as-built microstructure and element distribution using optical and scanning electron microscopy (SEM). In addition, the deformation behavior was studied using uniaxial tensile testing and SEM. Comparison with conventionally produced TRIP/TWIP steel revealed that elemental segregation, which is normally very pronounced in high-manganese steels and requires energy-intensive post processing, is reduced due to the high cooling rates during SLM. Also, the very fast cooling promoted ε- and α'-martensite formation prior to deformation. The superior strength and pronounced anisotropy of the SLM-produced material was correlated with the microstructure based on the process-specific characteristics.

  10. Exploiting Process-Related Advantages of Selective Laser Melting for the Production of High-Manganese Steel

    Directory of Open Access Journals (Sweden)

    Christian Haase

    2017-01-01

    Full Text Available Metal additive manufacturing has strongly gained scientific and industrial importance during the last decades due to the geometrical flexibility and increased reliability of parts, as well as reduced equipment costs. Within the field of metal additive manufacturing methods, selective laser melting (SLM is an eligible technique for the production of fully dense bulk material with complex geometry. In the current study, we addressed the application of SLM for processing a high-manganese TRansformation-/TWinning-Induced Plasticity (TRIP/TWIP steel. The solidification behavior was analyzed by careful characterization of the as-built microstructure and element distribution using optical and scanning electron microscopy (SEM. In addition, the deformation behavior was studied using uniaxial tensile testing and SEM. Comparison with conventionally produced TRIP/TWIP steel revealed that elemental segregation, which is normally very pronounced in high-manganese steels and requires energy-intensive post processing, is reduced due to the high cooling rates during SLM. Also, the very fast cooling promoted ε- and α’-martensite formation prior to deformation. The superior strength and pronounced anisotropy of the SLM-produced material was correlated with the microstructure based on the process-specific characteristics.

  11. High quality vacuum induction melting of small quantities of NiTi shape memory alloys in graphite crucibles

    International Nuclear Information System (INIS)

    Frenzel, J.; Zhang, Z.; Neuking, K.; Eggeler, G.

    2004-01-01

    Binary NiTi based shape memory alloys can be produced starting from the pure elements (Ni-pellets, Ti-rods) by using vacuum induction melting (VIM). VIM ingot metallurgy is known to produce materials with a good chemical homogeneity; it, moreover, is cheaper than vacuum arc melting (VAM) when small quantities of laboratory materials are needed. In a VIM procedure, graphite crucibles are attractive because they have appropriate electrical properties. For NiTi melting, graphite crucibles are interesting because they are reasonably priced and they show a good resistance against thermal cracking. On the other hand, it is well known that melting of Ti alloys in graphite crucibles is associated with a vigorous interface reaction. And the carbon concentration of NiTi alloys needs to be kept below a certain minimum in order to assure that the functional properties of the alloys meet the required targets. Therefore, it is important to minimize the carbon pick up of the melt. The present work presents experimental results and discusses thermodynamic and kinetic aspects of the reaction of NiTi melts with graphite crucibles; a method is suggested to keep the carbon dissolution into the melt at a minimum

  12. Using high resolution tritium profiles to quantify the effects of melt on two Spitsbergen ice cores

    NARCIS (Netherlands)

    van der Wel, L.G.; Streurman, H.J.; Isaksson, E.; Helsen, M.M.; van de Wal, R.S.W.; Martma, T.; Pohjola, V.A.; Moore, J.C.; Meijer, H.A.J.

    2011-01-01

    Ice cores from small ice caps provide valuable climatic information, additional to that of Greenland and Antarctica. However, their integrity is usually compromised by summer meltwater percolation. To determine to what extent this can affect such ice cores, we performed high-resolution tritium

  13. Highly sensitive polymerase chain reaction-free quantum dot-based quantification of forensic genomic DNA

    International Nuclear Information System (INIS)

    Tak, Yu Kyung; Kim, Won Young; Kim, Min Jung; Han, Eunyoung; Han, Myun Soo; Kim, Jong Jin; Kim, Wook; Lee, Jong Eun; Song, Joon Myong

    2012-01-01

    Highlights: ► Genomic DNA quantification were performed using a quantum dot-labeled Alu sequence. ► This probe provided PCR-free determination of human genomic DNA. ► Qdot-labeled Alu probe-hybridized genomic DNAs had a 2.5-femtogram detection limit. ► Qdot-labeled Alu sequence was used to assess DNA samples for human identification. - Abstract: Forensic DNA samples can degrade easily due to exposure to light and moisture at the crime scene. In addition, the amount of DNA acquired at a criminal site is inherently limited. This limited amount of human DNA has to be quantified accurately after the process of DNA extraction. The accurately quantified extracted genomic DNA is then used as a DNA template in polymerase chain reaction (PCR) amplification for short tandem repeat (STR) human identification. Accordingly, highly sensitive and human-specific quantification of forensic DNA samples is an essential issue in forensic study. In this work, a quantum dot (Qdot)-labeled Alu sequence was developed as a probe to simultaneously satisfy both the high sensitivity and human genome selectivity for quantification of forensic DNA samples. This probe provided PCR-free determination of human genomic DNA and had a 2.5-femtogram detection limit due to the strong emission and photostability of the Qdot. The Qdot-labeled Alu sequence has been used successfully to assess 18 different forensic DNA samples for STR human identification.

  14. Visualization of DNA in highly processed botanical materials.

    Science.gov (United States)

    Lu, Zhengfei; Rubinsky, Maria; Babajanian, Silva; Zhang, Yanjun; Chang, Peter; Swanson, Gary

    2018-04-15

    DNA-based methods have been gaining recognition as a tool for botanical authentication in herbal medicine; however, their application in processed botanical materials is challenging due to the low quality and quantity of DNA left after extensive manufacturing processes. The low amount of DNA recovered from processed materials, especially extracts, is "invisible" by current technology, which has casted doubt on the presence of amplifiable botanical DNA. A method using adapter-ligation and PCR amplification was successfully applied to visualize the "invisible" DNA in botanical extracts. The size of the "invisible" DNA fragments in botanical extracts was around 20-220 bp compared to fragments of around 600 bp for the more easily visualized DNA in botanical powders. This technique is the first to allow characterization and visualization of small fragments of DNA in processed botanical materials and will provide key information to guide the development of appropriate DNA-based botanical authentication methods in the future. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Highly sensitive DNA sensors based on cerium oxide nanorods

    Science.gov (United States)

    Nguyet, Nguyen Thi; Hai Yen, Le Thi; Van Thu, Vu; lan, Hoang; Trung, Tran; Vuong, Pham Hung; Tam, Phuong Dinh

    2018-04-01

    In this work, a CeO2 nanorod (NR)-based electrochemical DNA sensor was developed to identify Salmonella that causes food-borne infections. CeO2 NRs were synthesized without templates via a simple and unexpensive hydrothermal approach at 170 °C for 12 h by using CeO(NO3)3·6H2O as a Ce source. The DNA probe was immobilized onto the CeO2 NR-modified electrode through covalent attachment. The characteristics of the hybridized DNA were analyzed through electrochemical impedance spectroscopy (EIS) with [Fe(CN)6]3-/4- as a redox probe. Experimental results showed that electron transfer resistance (Ret) increased after the DNA probe was attached to the electrode surface and increased further after the DNA probe hybridized with its complementary sequence. A linear response of Ret to the target DNA concentration was found from 0.01 μM to 2 μM. The detection limit and sensitivity of the DNA sensor were 0.01 μM and 3362.1 Ω μM-1 cm-2, respectively. Various parameters, such as pH value, ionic strength, DNA probe concentration, and hybridization time, influencing DNA sensor responses were also investigated.

  16. Fusion surface material melting, ablation, and ejection under high heat loading

    International Nuclear Information System (INIS)

    Holliday, M.R.; Doster, J.M.; Gilligan, J.G.

    1986-01-01

    Limiters, divertor plates, and sections of the first wall are exposed to intense heat loads during normal operation and plasma disruptions. This results in severe thermal stresses as well as erosion of the surface material. Large surface areas of compact high-field tokamaks are expected to be exposed to these high heat loads. The need for a fast and accurate computational model describing the heat transfer and phase change process has arisen as a part of the larger model of the plasma-edge region. The authors report on a solution scheme that has been developed that minimizes computational time for this time-dependent, one-dimensional, moving boundary problem. This research makes use of the heat balance integral technique, which is at least an order of magnitude faster than previous finite difference techniques. In addition, we report on the effect of molten material ejection (by external forces) on the total surface erosion rate

  17. High Temperature Corrosion of Nickel in NaVO3-V2O5 Melts

    Directory of Open Access Journals (Sweden)

    J. Porcayo-Calderon

    2017-01-01

    Full Text Available Many alloys used at high temperature in industrial processes are Ni-based and many others contain it in appreciable quantities, so it is of interest to evaluate the performance of pure nickel in order to determine the behavior of its alloys once the elements responsible for their protection have been depleted due to accelerated corrosion processes in the presence of vanadium-rich molten salts. Due to this, this work presents the study of Ni behavior in NaVO3-V2O5 mixtures at different temperatures. The behavior of pure nickel was determined by both electrochemical and mass loss measurements. The results show that the aggressiveness of the vanadium salts is increased by increasing both the V2O5 content and temperature. V2O5 addition considerably increases the current densities of the anodic and cathodic reactions. The corrosion process of Ni is modified due to the presence of its corrosion products, and its presence increases the activation energy by at least one order of magnitude. Although nickel shows a high reactivity in vanadium-rich salts, its reaction products are highly stable and protect it from the corrosive medium because the corrosion reactions trap the vanadium and block the migration of nickel ions.

  18. Scanning fluorescence detector for high-throughput DNA genotyping

    Science.gov (United States)

    Rusch, Terry L.; Petsinger, Jeremy; Christensen, Carl; Vaske, David A.; Brumley, Robert L., Jr.; Luckey, John A.; Weber, James L.

    1996-04-01

    A new scanning fluorescence detector (SCAFUD) was developed for high-throughput genotyping of short tandem repeat polymorphisms (STRPs). Fluorescent dyes are incorporated into relatively short DNA fragments via polymerase chain reaction (PCR) and are separated by electrophoresis in short, wide polyacrylamide gels (144 lanes with well to read distances of 14 cm). Excitation light from an argon laser with primary lines at 488 and 514 nm is introduced into the gel through a fiber optic cable, dichroic mirror, and 40X microscope objective. Emitted fluorescent light is collected confocally through a second fiber. The confocal head is translated across the bottom of the gel at 0.5 Hz. The detection unit utilizes dichroic mirrors and band pass filters to direct light with 10 - 20 nm bandwidths to four photomultiplier tubes (PMTs). PMT signals are independently amplified with variable gain and then sampled at a rate of 2500 points per scan using a computer based A/D board. LabView software (National Instruments) is used for instrument operation. Currently, three fluorescent dyes (Fam, Hex and Rox) are simultaneously detected with peak detection wavelengths of 543, 567, and 613 nm, respectively. The detection limit for fluorescein-labeled primers is about 100 attomoles. Planned SCAFUD upgrades include rearrangement of laser head geometry, use of additional excitation lasers for simultaneous detection of more dyes, and the use of detector arrays instead of individual PMTs. Extensive software has been written for automatic analysis of SCAFUD images. The software enables background subtraction, band identification, multiple- dye signal resolution, lane finding, band sizing and allele calling. Whole genome screens are currently underway to search for loci influencing such complex diseases as diabetes, asthma, and hypertension. Seven production SCAFUDs are currently in operation. Genotyping output for the coming year is projected to be about one million total genotypes (DNA

  19. Characterisation of 100 kW electron beam melting gun and its adaptation as electron gun for high power DC electron accelerators

    International Nuclear Information System (INIS)

    Banerjee, Srutarshi; Bhattacharjee, Dhruva; Waghmare, Abhay; Tiwari, Rajnish; Bakhtsingh, R.I.; Dasgupta, K.; Gupta, Sachin; Prakash, Baibhaw; Jha, M.N.

    2015-01-01

    The paper deals with the characterization of the 100 kW electron beam melting gun for its adaptation in high power DC Electron Accelerators. The indigenously designed electron beam melting system at BARC is chosen for characterization. It comprises of electron gun as source of electrons, two electromagnetic focusing lenses viz. upper focusing lens and lower focusing lens for beam focusing, intermediate beam aperture for vacuum decoupling between gun region and melt zone, deflection and oscillation lens for maneuvering the beam on the melt charge and water cooled crucible that acts as a beam dump. In this system, the electron gun is designed for 40 kV and 100 kW corresponding to a maximum beam current of 2.5 A. The electron gun uses directly heated spiral tungsten filament. The operating temperature of the filament is 2800 °K. The focusing electrode and the anode profile are designed based on Pierce geometry. High Power DC Electron Accelerators require high currents of 1 A. The beam must comply with the requirement of 40 mm beam diameter and 10 mrad divergence at the exit of the electron gun. The characterization of the existing electron gun was done to find out all the beam parameters, for e.g. beam size, beam divergence, perveance etc. to be adapted or to be modified for the design of electron gun for high power DC accelerators. This paper shows limitations and the possible solutions for design of high power DC accelerators. (author)

  20. Progade PT path, prograde fluid flow, metasomatism and hydrous melting in the Osor high-grade HT-LP complex (Catalan Coastal Ranges-CCR, NE Iberia).

    Science.gov (United States)

    Reche, Joan; Martínez, Francisco; Leoz, Gisela

    2015-04-01

    Fast thermal pulses related to HT-LP metamorphism may imply dehydration reaction overstepping, higher than normal fluid production rates, quick local increases in Pfluid and common situations of Pfluid >> Plitostatic and surpassing locally the tensile stresses. This ambient would be favorable to transient hydrofracturing and fluid flow even if the ongoing HT-LP event develops on dominantly ductile crustal levels. In inner zones where temperatures are high enough, hydrous melting and melt migration would be favored as well. Such movement of fluids and melts would tend to be sustained if non-hydrostatic stresses are active during heating, and would be favored in high strain domains such high-T shear zones or along foliation planes. In such scenario, local metasomatic processes and mass-transfer phenomena are expected to occur along these high strain zones and so distributed along tectonic anisotropies. A variety of features found in high T Garnet - biotite-sillimanite±cordierite±plagioclase±K-feldspar±quartz metapelitic gneisses from the Osor Complex (Guilleries massif, CCR), testify from this kind of processes operating in the lower crustal section, at the amphibolite to granulite transition zone during a prograde Variscan HT-LP thermal pulse. Such features include: syn-D2 quartz veining, leucogranitoid (leucotonalite, trondhjemitic) lenses sub parallel to S2 dominant foliation, fibrolite-rich foliation planes and prograde sub-idiomorphic garnet developing preferentially near fluid migration channels (quartz veins) or near melt lenses.

  1. Evaluation of the Capacity of PCR and High-Resolution Melt Curve Analysis for Identification of Mixed Infection with Mycoplasma gallisepticum Strains.

    Directory of Open Access Journals (Sweden)

    Seyed A Ghorashi

    Full Text Available Pathogenicity and presentation of Mycoplasma gallisepticum (MG infection may differ from one strain to another and this may have implications on control measures. Infection of individual birds with more than one MG strain has been reported. A PCR followed by high resolution melt (HRM curve analysis has been developed in our laboratory and routinely used for detection and differentiation of MG strains. However the potential of this test for identification of MG strains in a mixed specimen has not been evaluated. In the present study, the capability of PCR-HRM curve analysis technique, targeting vlhA and pvpA genes was assessed for identification of individual MG strains in a mixed population. Different DNA ratios of two MG strains from 1 to 10(-4 ng were tested with some generated conventional and normalized curves distinct from those of individual strains alone. Using genotype confidence percentages (GCP generated from HRM curve analysis, it was found that vlhA PCR-HRM was more consistent than pvpA PCR-HRM for the detection of MG ts-11 vaccine strain mixed with any of the MG strains 6/85, F, S6 or a field isolate. The potential of vlhA PCR-HRM to detect mixed MG strains in a specimen was found to be primarily dependent on quantity and proportion of the target DNAs in the mixture. This is the first study examining the capacity of PCR-HRM technique for identification of individual MG strains in a mixed strain population.

  2. Application of support vector regression for optimization of vibration flow field of high-density polyethylene melts characterized by small angle light scattering

    Science.gov (United States)

    Xian, Guangming

    2018-03-01

    In this paper, the vibration flow field parameters of polymer melts in a visual slit die are optimized by using intelligent algorithm. Experimental small angle light scattering (SALS) patterns are shown to characterize the processing process. In order to capture the scattered light, a polarizer and an analyzer are placed before and after the polymer melts. The results reported in this study are obtained using high-density polyethylene (HDPE) with rotation speed at 28 rpm. In addition, support vector regression (SVR) analytical method is introduced for optimization the parameters of vibration flow field. This work establishes the general applicability of SVR for predicting the optimal parameters of vibration flow field.

  3. High-pressure anatectic paragneisses from the Namche Barwa, Eastern Himalayan Syntaxis: Textural evidence for partial melting, phase equilibria modeling and tectonic implications

    Science.gov (United States)

    Guilmette, C.; Indares, A.; Hébert, R.

    2011-05-01

    Rare kyanite-bearing anatectic paragneisses are found as boudins within sillimanite-bearing paragneisses of the core of the Namche Barwa Antiform, Tibet. In the present study, we document an occurrence from the NW side of the Yarlung Zangbo River. These rocks mainly consist of the assemblage garnet + K-feldspar + kyanite ± biotite + quartz + rutile ± plagioclase with kyanite locally pseudomorphed by sillimanite. The documented textures are consistent with the rocks having undergone biotite-dehydration melting in the kyanite stability field, under high-P granulite facies conditions, and having experienced melt extraction. However textures related to melt crystallization are ubiquitous both in polymineralic inclusions in garnet and in the matrix, suggesting that a melt fraction had remained in these rocks. Phase equilibria modelling was undertaken in the NCKFMASTHO system with THERMOCALC. P-T pseudosections built with the bulk compositions of one aluminous and one sub-aluminous paragneiss samples predict a biotite-kyanite-garnet-quartz-plagioclase-K-feldspar-liquid-rutile ± ilmenite field, in which biotite-dehydration melting occurs, located in the P-T range of ~ 800-875 °C and ~ 10-17 kbar. In addition, the topologies of these pseudosections are consistent with substantial melt loss during prograde metamorphism. A second set of P-T pseudosections with melt-reintegrated model bulk compositions were thus constructed to evaluate the effect of melt loss. The integration of textural information, precise mineral modes, mineral chemistry, and phase equilibria modelling allowed to constrain a P-T path where the rocks are buried to lower crustal depths at peak P-T conditions higher than 14 kbar and 825 °C, possibly in the order of 15-16 kbar and 850 °C, followed by decompression and cooling to P-T conditions of around 9 kbar and 810 °C, under which the remaining melt was solidified. The implications for granite production at the NBA and for Himalayan tectonic models

  4. Method of melting solid waste

    International Nuclear Information System (INIS)

    Ootsuka, Katsuyuki; Mizuno, Ryokichi; Kuwana, Katsumi; Sawada, Yoshihisa; Komatsu, Fumiaki.

    1982-01-01

    Purpose: To enable the volume reduction treatment of a HEPA filter containing various solid wastes, particularly acid digestion residue, or an asbestos separator at a relatively low temperature range. Method: Solid waste to be heated and molten is high melting point material treated by ''acid digestion treatment'' for treating solid waste, e.g. a HEPA filter or polyvinyl chloride, etc. of an atomic power facility treated with nitric acid or the like. When this material is heated and molten by an electric furnace, microwave melting furnace, etc., boron oxide, sodium boride, sodium carbonate, etc. is added as a melting point lowering agent. When it is molten in this state, its melting point is lowered, and it becomes remarkably fluid, and the melting treatment is facilitated. Solidified material thus obtained through the melting step has excellent denseness and further large volume reduction rate of the solidified material. (Yoshihara, H.)

  5. Clarithromycin highly-loaded gastro-floating fine granules prepared by high-shear melt granulation can enhance the efficacy of Helicobacter pylori eradication.

    Science.gov (United States)

    Aoki, Hajime; Iwao, Yasunori; Mizoguchi, Midori; Noguchi, Shuji; Itai, Shigeru

    2015-05-01

    In an effort to develop a new gastro-retentive drug delivery system (GRDDS) without a large amount of additives, 75% clarithromycin (CAM) loaded fine granules were prepared with three different hydrophobic binders by high-shear melt granulation and their properties were evaluated. Granules containing the higher hydrophobic binder showed sustained drug release and were able to float over 24h. The synchrotron X-ray CT measurement indicated that both the high hydrophobicity of the binder and the void space inside the granules might be involved in their buoyancy. In an in vivo experiment, the floating granules more effectively eradicated Helicobacter pylori than a CAM suspension by remaining in the stomach for a longer period. In short, CAM highly-loaded gastro-floating fine granules can enhance the eradication efficiency of H. pylori compared with CAM alone. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Theory of high-force DNA stretching and overstretching

    NARCIS (Netherlands)

    Storm, C.; Nelson, P.

    2003-01-01

    Single-molecule experiments on single- and double-stranded DNA have sparked a renewed interest in the force versus extension of polymers. The extensible freely jointed chain (FJC) model is frequently invoked to explain the observed behavior of single-stranded DNA, but this model does not

  7. Determination of a Screening Metric for High Diversity DNA Libraries.

    Science.gov (United States)

    Guido, Nicholas J; Handerson, Steven; Joseph, Elaine M; Leake, Devin; Kung, Li A

    2016-01-01

    The fields of antibody engineering, enzyme optimization and pathway construction rely increasingly on screening complex variant DNA libraries. These highly diverse libraries allow researchers to sample a maximized sequence space; and therefore, more rapidly identify proteins with significantly improved activity. The current state of the art in synthetic biology allows for libraries with billions of variants, pushing the limits of researchers' ability to qualify libraries for screening by measuring the traditional quality metrics of fidelity and diversity of variants. Instead, when screening variant libraries, researchers typically use a generic, and often insufficient, oversampling rate based on a common rule-of-thumb. We have developed methods to calculate a library-specific oversampling metric, based on fidelity, diversity, and representation of variants, which informs researchers, prior to screening the library, of the amount of oversampling required to ensure that the desired fraction of variant molecules will be sampled. To derive this oversampling metric, we developed a novel alignment tool to efficiently measure frequency counts of individual nucleotide variant positions using next-generation sequencing data. Next, we apply a method based on the "coupon collector" probability theory to construct a curve of upper bound estimates of the sampling size required for any desired variant coverage. The calculated oversampling metric will guide researchers to maximize their efficiency in using highly variant libraries.

  8. Determination of a Screening Metric for High Diversity DNA Libraries.

    Directory of Open Access Journals (Sweden)

    Nicholas J Guido

    Full Text Available The fields of antibody engineering, enzyme optimization and pathway construction rely increasingly on screening complex variant DNA libraries. These highly diverse libraries allow researchers to sample a maximized sequence space; and therefore, more rapidly identify proteins with significantly improved activity. The current state of the art in synthetic biology allows for libraries with billions of variants, pushing the limits of researchers' ability to qualify libraries for screening by measuring the traditional quality metrics of fidelity and diversity of variants. Instead, when screening variant libraries, researchers typically use a generic, and often insufficient, oversampling rate based on a common rule-of-thumb. We have developed methods to calculate a library-specific oversampling metric, based on fidelity, diversity, and representation of variants, which informs researchers, prior to screening the library, of the amount of oversampling required to ensure that the desired fraction of variant molecules will be sampled. To derive this oversampling metric, we developed a novel alignment tool to efficiently measure frequency counts of individual nucleotide variant positions using next-generation sequencing data. Next, we apply a method based on the "coupon collector" probability theory to construct a curve of upper bound estimates of the sampling size required for any desired variant coverage. The calculated oversampling metric will guide researchers to maximize their efficiency in using highly variant libraries.

  9. Methods for High-throughput Characterisation of Environmental DNA

    DEFF Research Database (Denmark)

    Andersen, Kenneth

    This PhD thesis examines the potential of describing biodiversity of green plants (Viridiplantae), birds (Aves) and mammals (Mammalia), in the context of next-generation sequencing, from the DNA that all organisms segregate into the environment (eDNA). The research is based on case studies...... of species assemblages described by eDNA recovered from contemporary surface soil and Holocene sediment sequences, to assess the accuracy and limitations of the approach. Biodiversity incorporates two aspects of ecological communities, including both the taxonomic richness and abundance of individual taxa...... inhibition. In chapter four, alternative DNA extraction protocols and pipelines for characterising plant eDNA are tested on samples from contrasting environments including modern, Holocene and Pleistocene sediment samples. These results are compared to pollen and macrofossil records described from earlier...

  10. Lattice stability and high-pressure melting mechanism of dense hydrogen up to 1.5 TPa

    KAUST Repository

    Geng, Hua Y.

    2015-09-01

    © 2015 American Physical Society. Lattice stability and metastability, as well as melting, are important features of the physics and chemistry of dense hydrogen. Using ab initio molecular dynamics (AIMD), the classical superheating limit and melting line of metallic hydrogen are investigated up to 1.5 TPa. The computations show that the classical superheating degree is about 100 K, and the classical melting curve becomes flat at a level of 350 K when beyond 500 GPa. This information allows us to estimate the well depth and the potential barriers that must be overcome when the crystal melts. Inclusion of nuclear quantum effects (NQE) using path integral molecular dynamics (PIMD) predicts that both superheating limit and melting temperature are lowered to below room temperature, but the latter never reaches absolute zero. Detailed analysis indicates that the melting is thermally activated, rather than driven by pure zero-point motion (ZPM). This argument was further supported by extensive PIMD simulations, demonstrating the stability of Fddd structure against liquefaction at low temperatures.

  11. Melting temperature of graphite

    International Nuclear Information System (INIS)

    Korobenko, V.N.; Savvatimskiy, A.I.

    2001-01-01

    Full Text: Pulse of electrical current is used for fast heating (∼ 1 μs) of metal and graphite specimens placed in dielectric solid media. Specimen consists of two strips (90 μm in thick) placed together with small gap so they form a black body model. Quasy-monocrystal graphite specimens were used for uniform heating of graphite. Temperature measurements were fulfilled with fast pyrometer and with composite 2-strip black body model up to melting temperature. There were fulfilled experiments with zirconium and tungsten of the same black body construction. Additional temperature measurements of liquid zirconium and liquid tungsten are made. Specific heat capacity (c P ) of liquid zirconium and of liquid tungsten has a common feature in c P diminishing just after melting. It reveals c P diminishing after melting in both cases over the narrow temperature range up to usual values known from steady state measurements. Over the next wide temperature range heat capacity for W (up to 5000 K) and Zr (up to 4100 K) show different dependencies of heat capacity on temperature in liquid state. The experiments confirmed a high quality of 2-strip black body model used for graphite temperature measurements. Melting temperature plateau of tungsten (3690 K) was used for pyrometer calibration area for graphite temperature measurement. As a result, a preliminary value of graphite melting temperature of 4800 K was obtained. (author)

  12. Ankaramite: A New Type of High-Magnesium and High-Calcium Primitive Melt in the Magnitogorsk Island-Arc Zone (Southern Urals)

    Science.gov (United States)

    Pushkarev, E. V.; Ryazancev, A. V.; Gottman, I. A.; Degtyarev, K. E.; Kamenetsky, V. S.

    2018-04-01

    This work describes the geological position, mineral and chemical composition of high-Mg effusive ankaramites occurring as dykes and lava flows. They were found in the mélange zone of the western margin of the Magnitogorsk island arc zone in the Southern Urals. Data on the liquidus association of phenocrysts and on the composition of the matrix of effusives are given. According to the data obtained, the conclusion was drawn that the ankaramites studied can be attributed to the primary island arc melts, which were not subject to essential differentiation. This type of effusives has not been distinguished previously among island arc volcanogenic formations of the Urals. It is shown that ankaramites can be considered to be primary melts parental for dunite-clinopyroxenites-gabbro complexes of Ural-Alaskan type. The occurrence of ankaramites in the Paleozoic island arc formations of the Urals indicates the wehrlite composition of the mantle as the reason for the extremely wide development of wehrlites and clinopyroxenites in different mafic-ultramafic complexes of the Urals.

  13. Assessment of the potential for high-pressure melt ejection resulting from a Surry station blackout transient

    International Nuclear Information System (INIS)

    Knudson, D.L.; Dobbe, C.A.

    1993-11-01

    Containment integrity could be challenged by direct heating associated with a high pressure melt ejection (HPME) of core materials following reactor vessel breach during certain severe accidents. Intentional reactor coolant system (RCS) depressurization, where operators latch pressurizer relief valves open, has been proposed as an accident management strategy to reduce risks by mitigating the severity of HPME. However, decay heat levels, valve capacities, and other plant-specific characteristics determine whether the required operator action will be effective. Without operator action, natural circulation flows could heat ex-vessel RCS pressure boundaries (surge line and hot leg piping, steam generator tubes, etc.) to the point of failure before vessel breach, providing an alternate mechanism for RCS depressurization and HPME mitigation. This report contains an assessment of the potential for HPME during a Surry station blackout transient without operator action and without recovery. The assessment included a detailed transient analysis using the SCDAP/RELAP5/MOD3 computer code to calculate the plant response with and without hot leg countercurrent natural circulation, with and without reactor coolant pump seal leakage, and with variations on selected core damage progression parameters. RCS depressurization-related probabilities were also evaluated, primarily based on the code results

  14. Identification of six New World Leishmania species through the implementation of a High-Resolution Melting (HRM) genotyping assay.

    Science.gov (United States)

    Hernández, Carolina; Alvarez, Catalina; González, Camila; Ayala, Martha Stella; León, Cielo Maritza; Ramírez, Juan David

    2014-11-14

    Leishmaniases are tropical zoonotic diseases, caused by parasites from the genus Leishmania. New World (NW) species are related to sylvatic cycles although urbanization processes have been reported in some South American Countries such as Colombia. This eco-epidemiological complexity imposes a challenge to the detection of circulating parasite species, not only related to human cases but also infecting vectors and reservoirs. Currently, no harmonized methods have been deployed to discriminate the NW Leishmania species. Herein, we conducted a systematic and mechanistic High-Resolution Melting (HRM) assay targeted to HSP70 and ITS1. Specific primers were designed that coupled with a HRM analyses permitted to discriminate six NW Leishmania species. In order to validate the herein described algorithm, we included 35 natural isolates obtained from human cases, insect vectors and mammals. Our genotyping assay allowed the correct assignment of the six NW Leishmania species (L. mexicana, L. infantum (chagasi), L. amazonensis, L. panamensis, L. guyanensis and L. braziliensis) based on reference strains. When the algorithm was applied to a set of well-characterized strains by means of PCR-RFLP, MLEE and monoclonal antibodies (MA) we observed a tailored concordance between the HRM and PCR-RFLP/MLEE/MA (KI = 1.0). Additionally, we tested the limit of detection for the HRM method showing that this is able to detect at least 10 equivalent-parasites per mL. This is a rapid and reliable method to conduct molecular epidemiology and host-parasite association studies in endemic areas.

  15. Identification of molecular markers associated with Verticillium wilt resistance in alfalfa (Medicago sativa L.) using high-resolution melting.

    Science.gov (United States)

    Zhang, Tiejun; Yu, Long-Xi; McCord, Per; Miller, David; Bhamidimarri, Suresh; Johnson, David; Monteros, Maria J; Ho, Julie; Reisen, Peter; Samac, Deborah A

    2014-01-01

    Verticillium wilt, caused by the soilborne fungus, Verticillium alfalfae, is one of the most serious diseases of alfalfa (Medicago sativa L.) worldwide. To identify loci associated with resistance to Verticillium wilt, a bulk segregant analysis was conducted in susceptible or resistant pools constructed from 13 synthetic alfalfa populations, followed by association mapping in two F1 populations consisted of 352 individuals. Simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers were used for genotyping. Phenotyping was done by manual inoculation of the pathogen to replicated cloned plants of each individual and disease severity was scored using a standard scale. Marker-trait association was analyzed by TASSEL. Seventeen SNP markers significantly associated with Verticillium wilt resistance were identified and they were located on chromosomes 1, 2, 4, 7 and 8. SNP markers identified on chromosomes 2, 4 and 7 co-locate with regions of Verticillium wilt resistance loci reported in M. truncatula. Additional markers identified on chromosomes 1 and 8 located the regions where no Verticillium resistance locus has been reported. This study highlights the value of SNP genotyping by high resolution melting to identify the disease resistance loci in tetraploid alfalfa. With further validation, the markers identified in this study could be used for improving resistance to Verticillium wilt in alfalfa breeding programs.

  16. Identification of molecular markers associated with Verticillium wilt resistance in alfalfa (Medicago sativa L. using high-resolution melting.

    Directory of Open Access Journals (Sweden)

    Tiejun Zhang

    Full Text Available Verticillium wilt, caused by the soilborne fungus, Verticillium alfalfae, is one of the most serious diseases of alfalfa (Medicago sativa L. worldwide. To identify loci associated with resistance to Verticillium wilt, a bulk segregant analysis was conducted in susceptible or resistant pools constructed from 13 synthetic alfalfa populations, followed by association mapping in two F1 populations consisted of 352 individuals. Simple sequence repeat (SSR and single nucleotide polymorphism (SNP markers were used for genotyping. Phenotyping was done by manual inoculation of the pathogen to replicated cloned plants of each individual and disease severity was scored using a standard scale. Marker-trait association was analyzed by TASSEL. Seventeen SNP markers significantly associated with Verticillium wilt resistance were identified and they were located on chromosomes 1, 2, 4, 7 and 8. SNP markers identified on chromosomes 2, 4 and 7 co-locate with regions of Verticillium wilt resistance loci reported in M. truncatula. Additional markers identified on chromosomes 1 and 8 located the regions where no Verticillium resistance locus has been reported. This study highlights the value of SNP genotyping by high resolution melting to identify the disease resistance loci in tetraploid alfalfa. With further validation, the markers identified in this study could be used for improving resistance to Verticillium wilt in alfalfa breeding programs.

  17. High resolution melting curve analysis, a rapid and affordable method for mutation analysis in childhood acute myeloid leukemia

    Directory of Open Access Journals (Sweden)

    Yin eLiu

    2014-09-01

    Full Text Available Background: Molecular genetic alterations with prognostic significance have been described in childhood acute myeloid leukemia (AML. The aim of this study was to establish cost-effective techniques to detect mutations of FMS-like tyrosine kinase 3 (FLT3, Nucleophosmin 1 (NPM1, and a partial tandem duplication within the mixed lineage leukemia (MLL-PTD genes in childhood AML. Procedure: Ninety-nine children with newly diagnosed AML were included in this study. We developed a fluoresent dye SYTO-82 based high resolution melting curve (HRM anaylsis to detect FLT3 internal tandem duplication (FLT3-ITD, FLT3 tyrosine kinase domain (FLT3-TKD and NPM1 mutations. MLL-PTD was screened by real-time quantitative PCR. Results: The HRM methodology correlated well with gold standard Sanger sequencing with less cost. Among the 99 patients studied, the FLT3-ITD mutation was associated with significantly worse event free survival (EFS. Patients with the NPM1 mutation had significantly better EFS and overall survival. However, HRM was not sensitive enough for minimal residual disease monitoring. Conclusions: HRM was a rapid and efficient method for screening of FLT3 and NPM1 gene mutations. It was both affordable and accurate, especially in resource underprivileged regions. Our results indicated that HRM could be a useful clinical tool for rapid and cost effective screening of the FLT3 and NPM1 mutations in AML patients.

  18. Stripped-down DNA repair in a highly reduced parasite

    Directory of Open Access Journals (Sweden)

    Fast Naomi M

    2007-03-01

    Full Text Available Abstract Background Encephalitozoon cuniculi is a member of a distinctive group of single-celled parasitic eukaryotes called microsporidia, which are closely related to fungi. Some of these organisms, including E. cuniculi, also have uniquely small genomes that are within the prokaryotic range. Thus, E. cuniculi has undergone a massive genome reduction which has resulted in a loss of genes from diverse biological pathways, including those that act in DNA repair. DNA repair is essential to any living cell. A loss of these mechanisms invariably results in accumulation of mutations and/or cell death. Six major pathways of DNA repair in eukaryotes include: non-homologous end joining (NHEJ, homologous recombination repair (HRR, mismatch repair (MMR, nucleotide excision repair (NER, base excision repair (BER and methyltransferase repair. DNA polymerases are also critical players in DNA repair processes. Given the close relationship between microsporidia and fungi, the repair mechanisms present in E. cuniculi were compared to those of the yeast Saccharomyces cerevisiae to ascertain how the process of genome reduction has affected the DNA repair pathways. Results E. cuniculi lacks 16 (plus another 6 potential absences of the 56 DNA repair genes sought via BLASTP and PSI-BLAST searches. Six of 14 DNA polymerases or polymerase subunits are also absent in E. cuniculi. All of these genes are relatively well conserved within eukaryotes. The absence of genes is not distributed equally among the different repair pathways; some pathways lack only one protein, while there is a striking absence of many proteins that are components of both double strand break repair pathways. All specialized repair polymerases are also absent. Conclusion Given the large number of DNA repair genes that are absent from the double strand break repair pathways, E. cuniculi is a prime candidate for the study of double strand break repair with minimal machinery. Strikingly, all of the

  19. High Resolution Melt (HRM) analysis is an efficient tool to genotype EMS mutants in complex crop genomes.

    Science.gov (United States)

    Lochlainn, Seosamh Ó; Amoah, Stephen; Graham, Neil S; Alamer, Khalid; Rios, Juan J; Kurup, Smita; Stoute, Andrew; Hammond, John P; Østergaard, Lars; King, Graham J; White, Phillip J; Broadley, Martin R

    2011-12-08

    Targeted Induced Loci Lesions IN Genomes (TILLING) is increasingly being used to generate and identify mutations in target genes of crop genomes. TILLING populations of several thousand lines have been generated in a number of crop species including Brassica rapa. Genetic analysis of mutants identified by TILLING requires an efficient, high-throughput and cost effective genotyping method to track the mutations through numerous generations. High resolution melt (HRM) analysis has been used in a number of systems to identify single nucleotide polymorphisms (SNPs) and insertion/deletions (IN/DELs) enabling the genotyping of different types of samples. HRM is ideally suited to high-throughput genotyping of multiple TILLING mutants in complex crop genomes. To date it has been used to identify mutants and genotype single mutations. The aim of this study was to determine if HRM can facilitate downstream analysis of multiple mutant lines identified by TILLING in order to characterise allelic series of EMS induced mutations in target genes across a number of generations in complex crop genomes. We demonstrate that HRM can be used to genotype allelic series of mutations in two genes, BraA.CAX1a and BraA.MET1.a in Brassica rapa. We analysed 12 mutations in BraA.CAX1.a and five in BraA.MET1.a over two generations including a back-cross to the wild-type. Using a commercially available HRM kit and the Lightscanner™ system we were able to detect mutations in heterozygous and homozygous states for both genes. Using HRM genotyping on TILLING derived mutants, it is possible to generate an allelic series of mutations within multiple target genes rapidly. Lines suitable for phenotypic analysis can be isolated approximately 8-9 months (3 generations) from receiving M3 seed of Brassica rapa from the RevGenUK TILLING service.

  20. High Resolution Melt (HRM analysis is an efficient tool to genotype EMS mutants in complex crop genomes

    Directory of Open Access Journals (Sweden)

    Lochlainn Seosamh Ó

    2011-12-01

    Full Text Available Abstract Background Targeted Induced Loci Lesions IN Genomes (TILLING is increasingly being used to generate and identify mutations in target genes of crop genomes. TILLING populations of several thousand lines have been generated in a number of crop species including Brassica rapa. Genetic analysis of mutants identified by TILLING requires an efficient, high-throughput and cost effective genotyping method to track the mutations through numerous generations. High resolution melt (HRM analysis has been used in a number of systems to identify single nucleotide polymorphisms (SNPs and insertion/deletions (IN/DELs enabling the genotyping of different types of samples. HRM is ideally suited to high-throughput genotyping of multiple TILLING mutants in complex crop genomes. To date it has been used to identify mutants and genotype single mutations. The aim of this study was to determine if HRM can facilitate downstream analysis of multiple mutant lines identified by TILLING in order to characterise allelic series of EMS induced mutations in target genes across a number of generations in complex crop genomes. Results We demonstrate that HRM can be used to genotype allelic series of mutations in two genes, BraA.CAX1a and BraA.MET1.a in Brassica rapa. We analysed 12 mutations in BraA.CAX1.a and five in BraA.MET1.a over two generations including a back-cross to the wild-type. Using a commercially available HRM kit and the Lightscanner™ system we were able to detect mutations in heterozygous and homozygous states for both genes. Conclusions Using HRM genotyping on TILLING derived mutants, it is possible to generate an allelic series of mutations within multiple target genes rapidly. Lines suitable for phenotypic analysis can be isolated approximately 8-9 months (3 generations from receiving M3 seed of Brassica rapa from the RevGenUK TILLING service.

  1. Melting of contaminated metallic waste

    International Nuclear Information System (INIS)

    Lee, Y.-S.; Cheng, S.-Y.; Kung, H.-T.; Lin, L.-F.

    2004-01-01

    Approximately 100 tons of contaminated metallic wastes were produced each year due to maintenance for each TPC's nuclear power reactor and it was roughly estimated that there will be 10,000 tons of metallic scraps resulted from decommissioning of each reactor in the future. One means of handling the contaminated metal is to melt it. Melting process owns not only volume reduction which saves the high cost of final disposal but also resource conservation and recycling benefits. Melting contaminated copper and aluminum scraps in the laboratory scale have been conducted at INER. A total of 546 kg copper condenser tubes with a specific activity of about 2.7 Bq/g was melted in a vacuum induction melting facility. Three types of products, ingot, slag and dust were derived from the melting process, with average activities of 0.10 Bq/g, 2.33 Bq/g and 84.3 Bq/g respectively. After the laboratory melting stage, a pilot plant with a 500 kg induction furnace is being designed to melt the increasingly produced contaminated metallic scraps from nuclear facilities and to investigate the behavior of different radionuclides during melting. (author)

  2. Comparative study of the microstructures and mechanical properties of direct laser fabricated and arc-melted Al{sub x}CoCrFeNi high entropy alloys

    Energy Technology Data Exchange (ETDEWEB)

    Joseph, Jithin, E-mail: jithin@deakin.edu.au [Institute for Frontier Materials, Deakin University, Waurn Ponds 3216 (Australia); Jarvis, Tom; Wu, Xinhua [Monash Centre for Additive Manufacturing, Monash University, Clayton 3168 (Australia); Stanford, Nicole; Hodgson, Peter; Fabijanic, Daniel Mark [Institute for Frontier Materials, Deakin University, Waurn Ponds 3216 (Australia)

    2015-05-01

    High entropy alloys (HEA) are a relatively new metal alloy system that have promising potential in high temperature applications. These multi-component alloys are typically produced by arc-melting, requiring several remelts to achieve chemical homogeneity. Direct laser fabrication (DLF) is a rapid prototyping technique, which produces complex components from alloy powder by selectively melting micron-sized powder in successive layers. However, studies of the fabrication of complex alloys from simple elemental powder blends are sparse. In this study, DLF was employed to fabricate bulk samples of three alloys based on the Al{sub x}CoCrFeNi HEA system, where x was 0.3, 0.6 and 0.85 M fraction of Al. This produced FCC, FCC/BCC and BCC crystal structures, respectively. Corresponding alloys were also produced by arc-melting, and all microstructures were characterised and compared longitudinal and transverse to the build/solidification direction by x-ray diffraction, glow discharge optical emission spectroscopy and scanning electron microscopy (EDX and EBSD). Strong similarities were observed between the single phase FCC and BCC alloys produced by both techniques, however the FCC/BCC structures differed significantly. This has been attributed to a difference in the solidification rate and thermal gradient in the melt pool between the two different techniques. Room temperature compression testing showed very similar mechanical behaviour and properties for the two different processing routes. DLF was concluded to be a successful technique to manufacture bulk HEA's.

  3. Capillary gel electrophoresis for rapid, high resolution DNA sequencing.

    OpenAIRE

    Swerdlow, H; Gesteland, R

    1990-01-01

    Capillary gel electrophoresis has been demonstrated for the separation and detection of DNA sequencing samples. Enzymatic dideoxy nucleotide chain termination was employed, using fluorescently tagged oligonucleotide primers and laser based on-column detection (limit of detection is 6,000 molecules per peak). Capillary gel separations were shown to be three times faster, with better resolution (2.4 x), and higher separation efficiency (5.4 x) than a conventional automated slab gel DNA sequenci...

  4. High-resolution melting-curve analysis of ligation-mediated real-time PCR for rapid evaluation of an epidemiological outbreak of extended-spectrum-beta-lactamase-producing Escherichia coli.

    Science.gov (United States)

    Woksepp, Hanna; Jernberg, Cecilia; Tärnberg, Maria; Ryberg, Anna; Brolund, Alma; Nordvall, Michaela; Olsson-Liljequist, Barbro; Wisell, Karin Tegmark; Monstein, Hans-Jürg; Nilsson, Lennart E; Schön, Thomas

    2011-12-01

    Methods for the confirmation of nosocomial outbreaks of bacterial pathogens are complex, expensive, and time-consuming. Recently, a method based on ligation-mediated PCR (LM/PCR) using a low denaturation temperature which produces specific melting-profile patterns of DNA products has been described. Our objective was to further develop this method for real-time PCR and high-resolution melting analysis (HRM) in a single-tube system optimized in order to achieve results within 1 day. Following the optimization of LM/PCR for real-time PCR and HRM (LM/HRM), the method was applied for a nosocomial outbreak of extended-spectrum-beta-lactamase (ESBL)-producing and ST131-associated Escherichia coli isolates (n = 15) and control isolates (n = 29), including four previous clusters. The results from LM/HRM were compared to results from pulsed-field gel electrophoresis (PFGE), which served as the gold standard. All isolates from the nosocomial outbreak clustered by LM/HRM, which was confirmed by gel electrophoresis of the LM/PCR products and PFGE. Control isolates that clustered by LM/PCR (n = 4) but not by PFGE were resolved by confirmatory gel electrophoresis. We conclude that LM/HRM is a rapid method for the detection of nosocomial outbreaks of bacterial infections caused by ESBL-producing E. coli strains. It allows the analysis of isolates in a single-tube system within a day, and the discriminatory power is comparable to that of PFGE.

  5. High-Resolution Melting-Curve Analysis of Ligation-Mediated Real-Time PCR for Rapid Evaluation of an Epidemiological Outbreak of Extended-Spectrum-Beta-Lactamase-Producing Escherichia coli ▿

    Science.gov (United States)

    Woksepp, Hanna; Jernberg, Cecilia; Tärnberg, Maria; Ryberg, Anna; Brolund, Alma; Nordvall, Michaela; Olsson-Liljequist, Barbro; Wisell, Karin Tegmark; Monstein, Hans-Jürg; Nilsson, Lennart E.; Schön, Thomas

    2011-01-01

    Methods for the confirmation of nosocomial outbreaks of bacterial pathogens are complex, expensive, and time-consuming. Recently, a method based on ligation-mediated PCR (LM/PCR) using a low denaturation temperature which produces specific melting-profile patterns of DNA products has been described. Our objective was to further develop this method for real-time PCR and high-resolution melting analysis (HRM) in a single-tube system optimized in order to achieve results within 1 day. Following the optimization of LM/PCR for real-time PCR and HRM (LM/HRM), the method was applied for a nosocomial outbreak of extended-spectrum-beta-lactamase (ESBL)-producing and ST131-associated Escherichia coli isolates (n = 15) and control isolates (n = 29), including four previous clusters. The results from LM/HRM were compared to results from pulsed-field gel electrophoresis (PFGE), which served as the gold standard. All isolates from the nosocomial outbreak clustered by LM/HRM, which was confirmed by gel electrophoresis of the LM/PCR products and PFGE. Control isolates that clustered by LM/PCR (n = 4) but not by PFGE were resolved by confirmatory gel electrophoresis. We conclude that LM/HRM is a rapid method for the detection of nosocomial outbreaks of bacterial infections caused by ESBL-producing E. coli strains. It allows the analysis of isolates in a single-tube system within a day, and the discriminatory power is comparable to that of PFGE. PMID:21956981

  6. Melt layer erosion of pure and lanthanum doped tungsten under VDE-like high heat flux loads

    Science.gov (United States)

    Yuan, Y.; Greuner, H.; Böswirth, B.; Luo, G.-N.; Fu, B. Q.; Xu, H. Y.; Liu, W.

    2013-07-01

    Heat loads expected for VDEs in ITER were applied in the neutral beam facility GLADIS at IPP Garching. Several ˜3 mm thick rolled pure W and W-1 wt% La2O3 plates were exposed to pulsed hydrogen beams with a central heat flux of 23 MW/m2 for 1.5-1.8 s. The melting thresholds are determined, and melt layer motion as well as material structure evolutions are shown. The melting thresholds of the two W grades are very close in this experimental setup. Lots of big bubbles with diameters from several μm to several 10 μm in the re-solidified layer of W were observed and they spread deeper with increasing heat flux. However, for W-1 wt% La2O3, no big bubbles were found in the corrugated melt layer. The underlying mechanisms referred to the melt layer motion and bubble issues are tentatively discussed based on comparison of the erosion characteristics between the two W grades.

  7. Melt layer erosion of pure and lanthanum doped tungsten under VDE-like high heat flux loads

    International Nuclear Information System (INIS)

    Yuan, Y.; Greuner, H.; Böswirth, B.; Luo, G.-N.; Fu, B.Q.; Xu, H.Y.; Liu, W.

    2013-01-01

    Heat loads expected for VDEs in ITER were applied in the neutral beam facility GLADIS at IPP Garching. Several ∼3 mm thick rolled pure W and W–1 wt% La 2 O 3 plates were exposed to pulsed hydrogen beams with a central heat flux of 23 MW/m 2 for 1.5–1.8 s. The melting thresholds are determined, and melt layer motion as well as material structure evolutions are shown. The melting thresholds of the two W grades are very close in this experimental setup. Lots of big bubbles with diameters from several μm to several 10 μm in the re-solidified layer of W were observed and they spread deeper with increasing heat flux. However, for W–1 wt% La 2 O 3 , no big bubbles were found in the corrugated melt layer. The underlying mechanisms referred to the melt layer motion and bubble issues are tentatively discussed based on comparison of the erosion characteristics between the two W grades

  8. An optimized rapid bisulfite conversion method with high recovery of cell-free DNA.

    Science.gov (United States)

    Yi, Shaohua; Long, Fei; Cheng, Juanbo; Huang, Daixin

    2017-12-19

    Methylation analysis of cell-free DNA is a encouraging tool for tumor diagnosis, monitoring and prognosis. Sensitivity of methylation analysis is a very important matter due to the tiny amounts of cell-free DNA available in plasma. Most current methods of DNA methylation analysis are based on the difference of bisulfite-mediated deamination of cytosine between cytosine and 5-methylcytosine. However, the recovery of bisulfite-converted DNA based on current methods is very poor for the methylation analysis of cell-free DNA. We optimized a rapid method for the crucial steps of bisulfite conversion with high recovery of cell-free DNA. A rapid deamination step and alkaline desulfonation was combined with the purification of DNA on a silica column. The conversion efficiency and recovery of bisulfite-treated DNA was investigated by the droplet digital PCR. The optimization of the reaction results in complete cytosine conversion in 30 min at 70 °C and about 65% of recovery of bisulfite-treated cell-free DNA, which is higher than current methods. The method allows high recovery from low levels of bisulfite-treated cell-free DNA, enhancing the analysis sensitivity of methylation detection from cell-free DNA.

  9. mtDNA point and length heteroplasmy in high- and low radiation areas of Kerala

    International Nuclear Information System (INIS)

    Forster, L.; Forster, P.; Gurney, S.M.; Spencer, M.; Huang, C.; Röhl, A.; Brinkmann, B.

    2010-01-01

    A coastal peninsula in Kerala (India) contains the world's highest level of natural radioactivity in a densely populated area, offering an opportunity to characterize radiation-associated DNA mutations. Here, we focus on mitochondrial DNA (mtDNA) mutations, which are passed exclusively from the mother to her children. To analyse point mutations, we sampled 248 pedigrees (988 individuals) in the high-radiation peninsula and in nearby low-radiation islands as a control population. Then, in an extended sample of 1,172 mtDNA sequences (containing some non-Indians for comparison), we also analysed length mutations, which in mtDNA can lead to the phenomenon of length heteroplasmy, i.e. the existence of different DNA types in the same cell. We wished to find out how fast mtDNA mutates between generations, and whether the mutation rate is increased in radioactive conditions compared to the low-irradiation sample

  10. Galvanic high temperature cell with solid negative electrode and an electrolyte melt. Galvanische Hochtemperaturzelle mit fester negativer Elektrode und einem Schmelzelektrolyten

    Energy Technology Data Exchange (ETDEWEB)

    Kappus, W; Borger, W

    1987-01-08

    The purpose of the invention is to make an electrolyte melt available for high temperature cells (e.g. LiFeS cells), which guarantees ion transport and also acts as a separator. The invention starts from the fact that binary melts of the LiCl/KCl type are only liquid (i.e. without solid components) at a certain temperature at certain concentrations. With suitable mixing conditions, which apart from a eutectic composition, are mainly on the side of one of the two components, one can ensure that this component is present in the solid phase. In this way, a solid framework of LiCl, for example, is formed between the electrode plates in situ as a separator, in the pores of which the excess melt (e.g. LiCl/KCl) can carry out ion conduction. The volumetric ratio of the electrolyte melt in which liquid and solid phases are present at the working temperature of the cell should preferably be in the range of 2:1 to 1:2.

  11. DNA barcode detects high genetic structure within neotropical bird species.

    Directory of Open Access Journals (Sweden)

    Erika Sendra Tavares

    Full Text Available BACKGROUND: Towards lower latitudes the number of recognized species is not only higher, but also phylogeographic subdivision within species is more pronounced. Moreover, new genetically isolated populations are often described in recent phylogenies of Neotropical birds suggesting that the number of species in the region is underestimated. Previous COI barcoding of Argentinean bird species showed more complex patterns of regional divergence in the Neotropical than in the North American avifauna. METHODS AND FINDINGS: Here we analyzed 1,431 samples from 561 different species to extend the Neotropical bird barcode survey to lower latitudes, and detected even higher geographic structure within species than reported previously. About 93% (520 of the species were identified correctly from their DNA barcodes. The remaining 41 species were not monophyletic in their COI sequences because they shared barcode sequences with closely related species (N = 21 or contained very divergent clusters suggestive of putative new species embedded within the gene tree (N = 20. Deep intraspecific divergences overlapping with among-species differences were detected in 48 species, often with samples from large geographic areas and several including multiple subspecies. This strong population genetic structure often coincided with breaks between different ecoregions or areas of endemism. CONCLUSIONS: The taxonomic uncertainty associated with the high incidence of non-monophyletic species and discovery of putative species obscures studies of historical patterns of species diversification in the Neotropical region. We showed that COI barcodes are a valuable tool to indicate which taxa would benefit from more extensive taxonomic revisions with multilocus approaches. Moreover, our results support hypotheses that the megadiversity of birds in the region is associated with multiple geographic processes starting well before the Quaternary and extending to more recent

  12. High-resolution analysis of cytosine methylation in ancient DNA.

    Directory of Open Access Journals (Sweden)

    Bastien Llamas

    Full Text Available Epigenetic changes to gene expression can result in heritable phenotypic characteristics that are not encoded in the DNA itself, but rather by biochemical modifications to the DNA or associated chromatin proteins. Interposed between genes and environment, these epigenetic modifications can be influenced by environmental factors to affect phenotype for multiple generations. This raises the possibility that epigenetic states provide a substrate for natural selection, with the potential to participate in the rapid adaptation of species to changes in environment. Any direct test of this hypothesis would require the ability to measure epigenetic states over evolutionary timescales. Here we describe the first single-base resolution of cytosine methylation patterns in an ancient mammalian genome, by bisulphite allelic sequencing of loci from late Pleistocene Bison priscus remains. Retrotransposons and the differentially methylated regions of imprinted loci displayed methylation patterns identical to those derived from fresh bovine tissue, indicating that methylation patterns are preserved in the ancient DNA. Our findings establish the biochemical stability of methylated cytosines over extensive time frames, and provide the first direct evidence that cytosine methylation patterns are retained in DNA from ancient specimens. The ability to resolve cytosine methylation in ancient DNA provides a powerful means to study the role of epigenetics in evolution.

  13. Complete mtDNA genomes of Filipino ethnolinguistic groups: a melting pot of recent and ancient lineages in the Asia-Pacific region

    Science.gov (United States)

    Delfin, Frederick; Min-Shan Ko, Albert; Li, Mingkun; Gunnarsdóttir, Ellen D; Tabbada, Kristina A; Salvador, Jazelyn M; Calacal, Gayvelline C; Sagum, Minerva S; Datar, Francisco A; Padilla, Sabino G; De Ungria, Maria Corazon A; Stoneking, Mark

    2014-01-01

    The Philippines is a strategic point in the Asia-Pacific region for the study of human diversity, history and origins, as it is a cross-road for human migrations and consequently exhibits enormous ethnolinguistic diversity. Following on a previous in-depth study of Y-chromosome variation, here we provide new insights into the maternal genetic history of Filipino ethnolinguistic groups by surveying complete mitochondrial DNA (mtDNA) genomes from a total of 14 groups (11 groups in this study and 3 groups previously published) including previously published mtDNA hypervariable segment (HVS) data from Filipino regional center groups. Comparison of HVS data indicate genetic differences between ethnolinguistic and regional center groups. The complete mtDNA genomes of 14 ethnolinguistic groups reveal genetic aspects consistent with the Y-chromosome, namely: diversity and heterogeneity of groups, no support for a simple dichotomy between Negrito and non-Negrito groups, and different genetic affinities with Asia-Pacific groups that are both ancient and recent. Although some mtDNA haplogroups can be associated with the Austronesian expansion, there are others that associate with South Asia, Near Oceania and Australia that are consistent with a southern migration route for ethnolinguistic group ancestors into the Asia-Pacific, with a timeline that overlaps with the initial colonization of the Asia-Pacific region, the initial colonization of the Philippines and a possible separate post-colonization migration into the Philippine archipelago. PMID:23756438

  14. High resolution melting analysis for the detection of EMS induced mutations in wheat SbeIIa genes

    Directory of Open Access Journals (Sweden)

    Botticella Ermelinda

    2011-11-01

    Full Text Available Abstract Background Manipulation of the amylose-amylopectin ratio in cereal starch has been identified as a major target for the production of starches with novel functional properties. In wheat, silencing of starch branching enzyme genes by a transgenic approach reportedly caused an increase of amylose content up to 70% of total starch, exhibiting novel and interesting nutritional characteristics. In this work, the functionality of starch branching enzyme IIa (SBEIIa has been targeted in bread wheat by TILLING. An EMS-mutagenised wheat population has been screened using High Resolution Melting of PCR products to identify functional SNPs in the three homoeologous genes encoding the target enzyme in the hexaploid genome. Results This analysis resulted in the identification of 56, 14 and 53 new allelic variants respectively for SBEIIa-A, SBEIIa-B and SBEIIa-D. The effects of the mutations on protein structure and functionality were evaluated by a bioinformatic approach. Two putative null alleles containing non-sense or splice site mutations were identified for each of the three homoeologous SBEIIa genes; qRT-PCR analysis showed a significant decrease of their gene expression and resulted in increased amylose content. Pyramiding of different single null homoeologous allowed to isolate double null mutants showing an increase of amylose content up to 21% compared to the control. Conclusion TILLING has successfully been used to generate novel alleles for SBEIIa genes known to control amylose content in wheat. Single and double null SBEIIa genotypes have been found to show a significant increase in amylose content.

  15. Melting method for miscellaneous radioactive solid waste and melting furnace

    International Nuclear Information System (INIS)

    Osaki, Toru; Furukawa, Hirofumi; Uda, Nobuyoshi; Katsurai, Kiyomichi

    1998-01-01

    A vessel containing miscellaneous solid wastes is inserted in a crucible having a releasable material on the inner surface, they are induction-heated from the outside of the crucible by way of low temperature heating coils to melt low melting point materials in the miscellaneous wastes within a temperature range at which the vessel does not melt. Then, they are induction-heated by way of high temperature heating coils to melt the vessel and not yet melted materials, those molten materials are cooled, solidified molten material and the releasable material are taken out, and then the crucible is used again. Then, the crucible can be used again, so that it can be applied to a large scaled melting furnace which treats wastes by a unit of drum. In addition, since the cleaning of the used crucible and the application of the releasable material can be conducted without interrupting the operation of the melting furnace, the operation cycle of the melting furnace can be shortened. (N.H.)

  16. High-resolution DNA content analysis of microbiopsy samples in oral lichen planus.

    Science.gov (United States)

    Pentenero, M; Monticone, M; Marino, R; Aiello, C; Marchitto, G; Malacarne, D; Giaretti, W; Gandolfo, S; Castagnola, P

    2017-04-01

    DNA aneuploidy has been reported to be a predictor of poor prognosis in both premalignant and malignant lesions. In oral lichen planus (OLP), this hypothesis remains to be proved. This study aimed to determine the rate of occurrence of DNA aneuploidy in patients with OLP by high-resolution DNA flow cytometry. Patients with OLP were consecutively enrolled. Tissue samples were subdivided for formalin fixation and routine histological assessment and for immediate storage at -20°C for later DNA ploidy analysis, which was performed by DAPI staining of the extracted nuclei and excitation with a UV lamp. The DNA aneuploid sublines were characterized by the DNA Index. A DNA aneuploid status was observed in two of 77 patients with OLP (2.6%). When considering the clinical aspect of the OLP lesions, both DNA aneuploid cases had a reticular clinical aspect. DNA aneuploidy is an uncommon event in OLP and less frequent compared to other non-dysplastic and non-OLP oral potentially malignant disorders. The extremely low rate of DNA aneuploidy could represent an occasional finding or reflect the low rate of malignant transformation observed in patients with OLP even if the real prognostic value of DNA ploidy analysis in patients with OLP remains to be confirmed. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  17. High-Throughput Analysis of T-DNA Location and Structure Using Sequence Capture.

    Directory of Open Access Journals (Sweden)

    Soichi Inagaki

    Full Text Available Agrobacterium-mediated transformation of plants with T-DNA is used both to introduce transgenes and for mutagenesis. Conventional approaches used to identify the genomic location and the structure of the inserted T-DNA are laborious and high-throughput methods using next-generation sequencing are being developed to address these problems. Here, we present a cost-effective approach that uses sequence capture targeted to the T-DNA borders to select genomic DNA fragments containing T-DNA-genome junctions, followed by Illumina sequencing to determine the location and junction structure of T-DNA insertions. Multiple probes can be mixed so that transgenic lines transformed with different T-DNA types can be processed simultaneously, using a simple, index-based pooling approach. We also developed a simple bioinformatic tool to find sequence read pairs that span the junction between the genome and T-DNA or any foreign DNA. We analyzed 29 transgenic lines of Arabidopsis thaliana, each containing inserts from 4 different T-DNA vectors. We determined the location of T-DNA insertions in 22 lines, 4 of which carried multiple insertion sites. Additionally, our analysis uncovered a high frequency of unconventional and complex T-DNA insertions, highlighting the needs for high-throughput methods for T-DNA localization and structural characterization. Transgene insertion events have to be fully characterized prior to use as commercial products. Our method greatly facilitates the first step of this characterization of transgenic plants by providing an efficient screen for the selection of promising lines.

  18. High-Throughput Analysis of T-DNA Location and Structure Using Sequence Capture.

    Science.gov (United States)

    Inagaki, Soichi; Henry, Isabelle M; Lieberman, Meric C; Comai, Luca

    2015-01-01

    Agrobacterium-mediated transformation of plants with T-DNA is used both to introduce transgenes and for mutagenesis. Conventional approaches used to identify the genomic location and the structure of the inserted T-DNA are laborious and high-throughput methods using next-generation sequencing are being developed to address these problems. Here, we present a cost-effective approach that uses sequence capture targeted to the T-DNA borders to select genomic DNA fragments containing T-DNA-genome junctions, followed by Illumina sequencing to determine the location and junction structure of T-DNA insertions. Multiple probes can be mixed so that transgenic lines transformed with different T-DNA types can be processed simultaneously, using a simple, index-based pooling approach. We also developed a simple bioinformatic tool to find sequence read pairs that span the junction between the genome and T-DNA or any foreign DNA. We analyzed 29 transgenic lines of Arabidopsis thaliana, each containing inserts from 4 different T-DNA vectors. We determined the location of T-DNA insertions in 22 lines, 4 of which carried multiple insertion sites. Additionally, our analysis uncovered a high frequency of unconventional and complex T-DNA insertions, highlighting the needs for high-throughput methods for T-DNA localization and structural characterization. Transgene insertion events have to be fully characterized prior to use as commercial products. Our method greatly facilitates the first step of this characterization of transgenic plants by providing an efficient screen for the selection of promising lines.

  19. High molecular weight DNA assembly in vivo for synthetic biology applications.

    Science.gov (United States)

    Juhas, Mario; Ajioka, James W

    2017-05-01

    DNA assembly is the key technology of the emerging interdisciplinary field of synthetic biology. While the assembly of smaller DNA fragments is usually performed in vitro, high molecular weight DNA molecules are assembled in vivo via homologous recombination in the host cell. Escherichia coli, Bacillus subtilis and Saccharomyces cerevisiae are the main hosts used for DNA assembly in vivo. Progress in DNA assembly over the last few years has paved the way for the construction of whole genomes. This review provides an update on recent synthetic biology advances with particular emphasis on high molecular weight DNA assembly in vivo in E. coli, B. subtilis and S. cerevisiae. Special attention is paid to the assembly of whole genomes, such as those of the first synthetic cell, synthetic yeast and minimal genomes.

  20. Revisiting the melting temperature of NpO2 and the challenges associated with high temperature actinide compound measurements

    NARCIS (Netherlands)

    Böhler, R.; Welland, M.J.; De Bruycker, F.; Boboridis, K.; Janssen, A.; Eloirdi, R.; Konings, R.J.M.; Manara, D.

    2012-01-01

    This work revisits the melting behaviour of neptunium dioxide, an actinide compound which can be produced in the nuclear fuel during operation, and which has an important impact on the nuclear fuel and waste radioactivity especially on the very long term. The present experimental approach employs

  1. A Microneedle Functionalized with Polyethyleneimine and Nanotubes for Highly Sensitive, Label-Free Quantification of DNA

    OpenAIRE

    Saadat-Moghaddam, Darius; Kim, Jong-Hoon

    2017-01-01

    The accurate measure of DNA concentration is necessary for many DNA-based biological applications. However, the current methods are limited in terms of sensitivity, reproducibility, human error, and contamination. Here, we present a microneedle functionalized with polyethyleneimine (PEI) and single-walled carbon nanotubes (SWCNTs) for the highly sensitive quantification of DNA. The microneedle was fabricated using ultraviolet (UV) lithography and anisotropic etching, and then functionalized w...

  2. Targeted DNA Methylation Analysis by High Throughput Sequencing in Porcine Peri-attachment Embryos

    OpenAIRE

    MORRILL, Benson H.; COX, Lindsay; WARD, Anika; HEYWOOD, Sierra; PRATHER, Randall S.; ISOM, S. Clay

    2013-01-01

    Abstract The purpose of this experiment was to implement and evaluate the effectiveness of a next-generation sequencing-based method for DNA methylation analysis in porcine embryonic samples. Fourteen discrete genomic regions were amplified by PCR using bisulfite-converted genomic DNA derived from day 14 in vivo-derived (IVV) and parthenogenetic (PA) porcine embryos as template DNA. Resulting PCR products were subjected to high-throughput sequencing using the Illumina Genome Analyzer IIx plat...

  3. A novel FRET pair for detection of parallel DNA triplexes by the LightCycler

    DEFF Research Database (Denmark)

    Schneider, Uffe V; Severinsen, Jette K; Géci, Imrich

    2010-01-01

    BACKGROUND: Melting temperature of DNA structures can be determined on the LightCycler using quenching of FAM. This method is very suitable for pH independent melting point (Tm) determination performed at basic or neutral pH, as a high throughput alternative to UV absorbance measurements. At acid...

  4. Assessment for Melting Temperature Measurement of Nucleic Acid by HRM.

    Science.gov (United States)

    Wang, Jing; Pan, Xiaoming; Liang, Xingguo

    2016-01-01

    High resolution melting (HRM), with a high sensitivity to distinguish the nucleic acid species with small variations, has been widely applied in the mutation scanning, methylation analysis, and genotyping. For the aim of extending HRM for the evaluation of thermal stability of nucleic acid secondary structures on sequence dependence, we investigated effects of the dye of EvaGreen, metal ions, and impurities (such as dNTPs) on melting temperature ( T m ) measurement by HRM. The accuracy of HRM was assessed as compared with UV melting method, and little difference between the two methods was found when the DNA T m was higher than 40°C. Both insufficiency and excessiveness of EvaGreen were found to give rise to a little bit higher T m , showing that the proportion of dye should be considered for precise T m measurement of nucleic acids. Finally, HRM method was also successfully used to measure T m s of DNA triplex, hairpin, and RNA duplex. In conclusion, HRM can be applied in the evaluation of thermal stability of nucleic acid (DNA or RNA) or secondary structural elements (even when dNTPs are present).

  5. Stability of Human Telomere Quadruplexes at High DNA Concentrations

    Czech Academy of Sciences Publication Activity Database

    Kejnovská, Iva; Vorlíčková, Michaela; Brázdová, Marie; Sagi, J.

    2014-01-01

    Roč. 101, č. 4 (2014), s. 428-438 ISSN 0006-3525 R&D Projects: GA ČR(CZ) GAP205/12/0466 Institutional support: RVO:68081707 Keywords : quadruplex * DNA concentration * folding topology Subject RIV: BO - Biophysics Impact factor: 2.385, year: 2014

  6. No variation and low synonymous substitution rates in coral mtDNA despite high nuclear variation

    Directory of Open Access Journals (Sweden)

    Hellberg Michael E

    2006-03-01

    Full Text Available Abstract Background The mitochondrial DNA (mtDNA of most animals evolves more rapidly than nuclear DNA, and often shows higher levels of intraspecific polymorphism and population subdivision. The mtDNA of anthozoans (corals, sea fans, and their kin, by contrast, appears to evolve slowly. Slow mtDNA evolution has been reported for several anthozoans, however this slow pace has been difficult to put in phylogenetic context without parallel surveys of nuclear variation or calibrated rates of synonymous substitution that could permit quantitative rate comparisons across taxa. Here, I survey variation in the coding region of a mitochondrial gene from a coral species (Balanophyllia elegans known to possess high levels of nuclear gene variation, and estimate synonymous rates of mtDNA substitution by comparison to another coral (Tubastrea coccinea. Results The mtDNA surveyed (630 bp of cytochrome oxidase subunit I was invariant among individuals sampled from 18 populations spanning 3000 km of the range of B. elegans, despite high levels of variation and population subdivision for allozymes over these same populations. The synonymous substitution rate between B. elegans and T. coccinea (0.05%/site/106 years is similar to that in most plants, but 50–100 times lower than rates typical for most animals. In addition, while substitutions to mtDNA in most animals exhibit a strong bias toward transitions, mtDNA from these corals does not. Conclusion Slow rates of mitochondrial nucleotide substitution result in low levels of intraspecific mtDNA variation in corals, even when nuclear loci vary. Slow mtDNA evolution appears to be the basal condition among eukaryotes. mtDNA substitution rates switch from slow to fast abruptly and unidirectionally. This switch may stem from the loss of just one or a few mitochondrion-specific DNA repair or replication genes.

  7. Effect of the distribution of saturated fatty acids in the melting and crystallization profiles of high-oleic high-stearic oils

    International Nuclear Information System (INIS)

    Bootello, M.A.; Garces, R.; Martinez-Force, E.; Salas, J.J.

    2016-01-01

    The composition and distribution of fatty acids in triacylglycerol (TAG) molecules are commonly considered as factors that determine the physical properties of a given oil or fat. The distribution of any fatty acid in fats and oils can be described through the α coefficient of asymmetry, which can be calculated from the TAG composition and fatty acid composition of the Sn-2 position of the TAGs determined through lipase hydrolysis. High-oleic high-stearic oils and fats are considered stable and healthy, and they are good substitutes for hydrogenated vegetable oils and palm fractions in many food products, such as spreads and confectionery. Here, different high-oleic high-stearic acid oils were formulated which contained different distributions of saturated fatty acids in their TAGs, while maintaining a similar fatty acid composition. The aim of this work was to discuss the possibility of using the α coefficient to predict the physical properties of fats in function of their chemical composition and their melting and crystallization behavior as examined by differential scanning calorimetry.

  8. Dynamic DNA binding, junction recognition and G4 melting activity underlie the telomeric and genome-wide roles of human CST.

    Science.gov (United States)

    Bhattacharjee, Anukana; Wang, Yongyao; Diao, Jiajie; Price, Carolyn M

    2017-12-01

    Human CST (CTC1-STN1-TEN1) is a ssDNA-binding complex that helps resolve replication problems both at telomeres and genome-wide. CST resembles Replication Protein A (RPA) in that the two complexes harbor comparable arrays of OB-folds and have structurally similar small subunits. However, the overall architecture and functions of CST and RPA are distinct. Currently, the mechanism underlying CST action at diverse replication issues remains unclear. To clarify CST mechanism, we examined the capacity of CST to bind and resolve DNA structures found at sites of CST activity. We show that CST binds preferentially to ss-dsDNA junctions, an activity that can explain the incremental nature of telomeric C-strand synthesis following telomerase action. We also show that CST unfolds G-quadruplex structures, thus providing a mechanism for CST to facilitate replication through telomeres and other GC-rich regions. Finally, smFRET analysis indicates that CST binding to ssDNA is dynamic with CST complexes undergoing concentration-dependent self-displacement. These findings support an RPA-based model where dissociation and re-association of individual OB-folds allow CST to mediate loading and unloading of partner proteins to facilitate various aspects of telomere replication and genome-wide resolution of replication stress. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. Large-scale DNA Barcode Library Generation for Biomolecule Identification in High-throughput Screens.

    Science.gov (United States)

    Lyons, Eli; Sheridan, Paul; Tremmel, Georg; Miyano, Satoru; Sugano, Sumio

    2017-10-24

    High-throughput screens allow for the identification of specific biomolecules with characteristics of interest. In barcoded screens, DNA barcodes are linked to target biomolecules in a manner allowing for the target molecules making up a library to be identified by sequencing the DNA barcodes using Next Generation Sequencing. To be useful in experimental settings, the DNA barcodes in a library must satisfy certain constraints related to GC content, homopolymer length, Hamming distance, and blacklisted subsequences. Here we report a novel framework to quickly generate large-scale libraries of DNA barcodes for use in high-throughput screens. We show that our framework dramatically reduces the computation time required to generate large-scale DNA barcode libraries, compared with a naїve approach to DNA barcode library generation. As a proof of concept, we demonstrate that our framework is able to generate a library consisting of one million DNA barcodes for use in a fragment antibody phage display screening experiment. We also report generating a general purpose one billion DNA barcode library, the largest such library yet reported in literature. Our results demonstrate the value of our novel large-scale DNA barcode library generation framework for use in high-throughput screening applications.

  10. Association between high risk papillomavirus DNA and nitric oxide release in the human uterine cervix.

    Science.gov (United States)

    Rahkola, Paivi; Mikkola, Tomi S; Ylikorkala, Olavi; Vaisanen-Tommiska, Mervi

    2009-08-01

    Local cervical factors may determine the outcome of human papillomavirus (HPV) infection. Nitric oxide (NO) may be one such factor, since it is produced by uterine cervical cells and it takes part in both immunological and carcinogenic reactions. We studied the association between the presence of cervical high risk (hr) HPV DNA and NO in the cervical canal in women. High risk HPV DNA status was assessed from 328 women by using a specific DNA test and the release of cervical NO was assessed as nitrate/nitrite in cervical fluid. Cervical NO was then compared between women showing different status of hr HPV DNA and different cytological and histological findings. High risk HPV DNA was present in 175/328 (53%) women. The cervical NO release in women with hr HPV DNA was 90% higher compared to hr HPV DNA negative women (poral contraception, intrauterine devices, or signs of bacterial vaginosis or candida infection. Cytologically healthy epithelium and epithelium with mild cytological or histological changes showed elevated NO release if hr HPV DNA was present. The presence of hr HPV DNA is associated with an increased release of NO in the human uterine cervix. The clinical significance of this phenomenon remains open.

  11. High-resolution melt PCR analysis for rapid identification of Chlamydia abortus live vaccine strain 1B among C. abortus strains and field isolates.

    Science.gov (United States)

    Vorimore, Fabien; Cavanna, Noémie; Vicari, Nadia; Magnino, Simone; Willems, Hermann; Rodolakis, Annie; Siarkou, Victoria I; Laroucau, Karine

    2012-09-01

    We describe a novel high-resolution melt assay that clearly differentiates Chlamydia abortus live vaccine strain 1B from field C. abortus strains and field wild-type isolates based on previously described single nucleotide polymorphisms. This modern genotyping technique is inexpensive, easy to use, and less time-consuming than PCR-RFLP. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. High resolution melt-curve analysis to fine map a locus controlling the paternal sorting of mitochondria

    Science.gov (United States)

    Mitochondria are required for normal growth and development and play an important role in programmed cell death and aging. The mitochondrial DNA is maternally transmitted in the vast majority of eukaryotes. One exception is cucumber (Cucumis sativus), whose mitochondrial DNA is paternally transmit...

  13. Identification and discrimination of Toxoplasma gondii, Sarcocystis spp., Neospora spp., and Cryptosporidium spp. by righ-resolution melting analysis.

    Directory of Open Access Journals (Sweden)

    Hllytchaikra Ferraz Fehlberg

    Full Text Available The objective of this study was to standardize the high-resolution melting method for identification and discrimination of Toxoplasma gondii, Sarcocystis spp., Neospora spp., and Cryptosporidium spp. by amplification of 18S ribosomal DNA (rDNA using a single primer pair. The analyses were performed on individual reactions (containing DNA from a single species of a protozoan, on duplex reactions (containing DNA from two species of protozoa in each reaction, and on a multiplex reaction (containing DNA of four parasites in a single reaction. The proposed method allowed us to identify and discriminate the four species by analyzing the derivative, normalized, and difference melting curves, with high reproducibility among and within the experiments, as demonstrated by low coefficients of variation (less than 2.2% and 2.0%, respectively. This is the first study where this method is used for discrimination of these four species of protozoa in a single reaction.

  14. Lipofection: A Highly Efficient, Lipid-Mediated DNA-Transfection Procedure

    Science.gov (United States)

    Felgner, Philip L.; Gadek, Thomas R.; Holm, Marilyn; Roman, Richard; Chan, Hardy W.; Wenz, Michael; Northrop, Jeffrey P.; Ringold, Gordon M.; Danielsen, Mark

    1987-11-01

    A DNA-transfection protocol has been developed that makes use of a synthetic cationic lipid, N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA). Small unilamellar liposomes containing DOTMA interact spontaneously with DNA to form lipid-DNA complexes with 100% entrapment of the DNA. DOTMA facilitates fusion of the complex with the plasma membrane of tissue culture cells, resulting in both uptake and expression of the DNA. The technique is simple, highly reproducible, and effective for both transient and stable expression of transfected DNA. Depending upon the cell line, lipofection is from 5- to >100-fold more effective than either the calcium phosphate or the DEAE-dextran transfection technique.

  15. Serum induced degradation of 3D DNA box origami observed by high speed atomic force microscope

    DEFF Research Database (Denmark)

    Jiang, Zaixing; Zhang, Shuai; Yang, Chuanxu

    2015-01-01

    3D DNA origami holds tremendous potential to encapsulate and selectively release therapeutic drugs. Observations of real-time performance of 3D DNA origami structures in physiological environment will contribute much to its further applications. Here, we investigate the degradation kinetics of 3D...... DNA box origami in serum using high-speed atomic force microscope optimized for imaging 3D DNA origami in real time. The time resolution allows characterizing the stages of serum effects on individual 3D DNA box origami with nanometer resolution. Our results indicate that the whole digest process...... is a combination of a rapid collapse phase and a slow degradation phase. The damages of box origami mainly happen in the collapse phase. Thus, the structure stability of 3D DNA box origami should be further improved, especially in the collapse phase, before clinical applications...

  16. High resolution, high sensitivity imaging and analysis of minerals and inclusions (fluid and melt) using the new CSIRO-GEMOC nuclear microprobe

    International Nuclear Information System (INIS)

    Ryan, C.G.; McInnes, B.M.; Van Achterbergh, E.; Williams, P.J.; Dong, G.; Zaw, K.

    1999-01-01

    .g. Yankee Lode, Mole Granite, NSW [Heinrich el al., 1993] and Batu Hijau, Indonesia [McInnes et al., 1999]), and the high concentrations of some elements in many ore-related fluid inclusions [e.g. Pb ∼4 wt% at Hellyer, Tasmania (Khin Zaw et al., 1996) and Ba ∼9 wt% at Starra, Cloncurry district, Queensland (Williams et al., 2000)]. Now, using the NMP, the internal contents of individual fluid inclusions can be imaged to show clearly that these elements reside within the fluid inclusions, and to discrimination against solid phases outside the inclusion volume. Melt Inclusion Analysis and Imaging Samples of melts and fluids, responsible for metasomatic change and evolution of the earth's upper mantle are often preserved as inclusions in xenoliths. However, their quench textures can often conceal rare minor phases that concentrate important trace elements (e.g. HFSE and REE). The penetration of MeV protons enables the detection of these contributions to ∼40 μm depth, thus providing a tool to determine reliable melt composition, with detection sensitivities down to 0.2 ppm, and to image spatial variation in component elements at 1-2 μm resolution. Copyright (1999) Geological Society of Australia

  17. High-fidelity DNA replication in Mycobacterium tuberculosis relies on a trinuclear zinc center.

    Science.gov (United States)

    Baños-Mateos, Soledad; van Roon, Anne-Marie M; Lang, Ulla F; Maslen, Sarah L; Skehel, J Mark; Lamers, Meindert H

    2017-10-11

    High-fidelity DNA replication depends on a proofreading 3'-5' exonuclease that is associated with the replicative DNA polymerase. The replicative DNA polymerase DnaE1 from the major pathogen Mycobacterium tuberculosis (Mtb) uses its intrinsic PHP-exonuclease that is distinct from the canonical DEDD exonucleases found in the Escherichia coli and eukaryotic replisomes. The mechanism of the PHP-exonuclease is not known. Here, we present the crystal structure of the Mtb DnaE1 polymerase. The PHP-exonuclease has a trinuclear zinc center, coordinated by nine conserved residues. Cryo-EM analysis reveals the entry path of the primer strand in the PHP-exonuclease active site. Furthermore, the PHP-exonuclease shows a striking similarity to E. coli endonuclease IV, which provides clues regarding the mechanism of action. Altogether, this work provides important insights into the PHP-exonuclease and reveals unique properties that make it an attractive target for novel anti-mycobacterial drugs.The polymerase and histidinol phosphatase (PHP) domain in the DNA polymerase DnaE1 is essential for mycobacterial high-fidelity DNA replication. Here, the authors determine the DnaE1 crystal structure, which reveals the PHP-exonuclease mechanism that can be exploited for antibiotic development.

  18. Tin in granitic melts: The role of melting temperature and protolith composition

    Science.gov (United States)

    Wolf, Mathias; Romer, Rolf L.; Franz, Leander; López-Moro, Francisco Javier

    2018-06-01

    Granite bound tin mineralization typically is seen as the result of extreme magmatic fractionation and late exsolution of magmatic fluids. Mineralization, however, also could be obtained at considerably less fractionation if initial melts already had enhanced Sn contents. We present chemical data and results from phase diagram modeling that illustrate the dominant roles of protolith composition, melting conditions, and melt extraction/evolution for the distribution of Sn between melt and restite and, thus, the Sn content of melts. We compare the element partitioning between leucosome and restite of low-temperature and high-temperature migmatites. During low-temperature melting, trace elements partition preferentially into the restite with the possible exception of Sr, Cd, Bi, and Pb, that may be enriched in the melt. In high-temperature melts, Ga, Y, Cd, Sn, REE, Pb, Bi, and U partition preferentially into the melt whereas Sc, V, Cr, Co, Ni, Mo, and Ba stay in the restite. This contrasting behavior is attributed to the stability of trace element sequestering minerals during melt generation. In particular muscovite, biotite, titanite, and rutile act as host phases for Sn and, therefore prevent Sn enrichment in the melt as long as they are stable phases in the restite. As protolith composition controls both the mineral assemblage and modal contents of the various minerals, protolith composition eventually also controls the fertility of a rock during anatexis, restite mineralogy, and partitioning behavior of trace metals. If a particular trace element is sequestered in a phase that is stable during partial melting, the resulting melt is depleted in this element whereas the restite becomes enriched. Melt generation at high temperature may release Sn when Sn-hosts become unstable. If melt has not been lost before the breakdown of Sn-hosts, Sn contents in the melt will increase but never will be high. In contrast, if melt has been lost before the decomposition of Sn

  19. On the phase evolution of AlCoCrCuFeMnSix high entropy alloys prepared by mechanical alloying and arc melting route

    Science.gov (United States)

    Kumar, Anil; Chopkar, Manoj

    2018-05-01

    Effect of Si addition on phase formation of AlCoCrCuFeMnSix (x=0, 0.3, 0.6 and 0.9) high entropy alloy have been investigated in this work. The alloys are prepared by mechanical alloying and vacuum arc melting technique. The X-ray diffraction results reveals the formation of mixture of face centered and body centered cubic solid solution phases in milled powders. The addition of Si favours body centered cubic structure formation during milling process. Whereas, after melting the milled powders, body centered phases formed during milling is partial transformed into sigma phases. XRD results were also correlated with the SEM elemental mapping of as casted samples. Addition of Si favours σ phase formation in the as cast samples.

  20. Anomalous density and elastic properties of basalt at high pressure: Reevaluating of the effect of melt fraction on seismic velocity in the Earth's crust and upper mantle

    Science.gov (United States)

    Clark, Alisha N.; Lesher, Charles E.; Jacobsen, Steven D.; Wang, Yanbin

    2016-06-01

    Independent measurements of the volumetric and elastic properties of Columbia River basalt glass were made up to 5.5 GPa by high-pressure X-ray microtomography and GHz-ultrasonic interferometry, respectively. The Columbia River basalt displays P and S wave velocity minima at 4.5 and 5 GPa, respectively, violating Birch's law. These data constrain the pressure dependence of the density and elastic moduli at high pressure, which cannot be modeled through usual equations of state nor determined by stepwise integrating the bulk sound velocity as is common practice. We propose a systematic variation in compression behavior of silicate glasses that is dependent on the degree of polymerization and arises from the flexibility of the aluminosilicate network. This behavior likely persists into the liquid state for basaltic melts resulting in weak pressure dependence for P wave velocities perhaps to depths of the transition zone. Modeling the effect of partial melt on P wave velocity reductions suggests that melt fraction determined by seismic velocity variations may be significantly overestimated in the crust and upper mantle.

  1. CO partial pressure dependence of the kinetics of melting of HbS aggregates studied in high concentration phosphate buffer

    Science.gov (United States)

    Aroutiounian, Svetlana

    2002-10-01

    Deoxygenated sickle cell hemoglobin (HbS) monomers enter the polymer phase either by incorporation into a critical nucleus, through heterogeneous nucleation and or through linear growth of the polymers when the concentration of monomers exceeds the solubility. CO-bound, R-state HbS monomers do not polymerize. Thus, polymer melting is enhanced by binding of carbon monoxide (CO) to HbS polymerized monomers. In our study, the melting of HbS aggregates mediated by dilution and CO binding to polymerized monomers is observed with time-resolved extinction spectroscopy. The CO partial pressure (pCO) dependence of the kinetics of melting is studied for pCO = 0, 0.25, 0.5, 0.75, 1 atm with difference progress curves. A phenomenological description with slow and fast relaxation modes reveals a variable relaxation time near the pCO=0.5 due to competition of kinetic mechanisms. The slow component increases with increasing pCO. It has a positive intercept due to the combined action of dilution of the sample and CO-ligation. The pCO dependence is near linear due to non-cooperative CO binding. Significant slowing down of aged samples, most likely due to gelation, is observed. As possible mechanism for variable relaxation time near pCO=0.5atm the fractional percolation threshold is discussed. This work was supported by NIH grant HL58091 (awarded to Daniel. B. Kim-Shapiro).

  2. Fabrication of the tetrathiafulvalene–2,3,5,6-tetrafluoro-7,7,8,8-tetracyanoquinodimethane charge transfer complex with high crystallinity by eutectic melting method

    Science.gov (United States)

    Kim, Jueun; Kang, Youngjong; Lee, Jaejong

    2018-06-01

    We show that high crystallinity and charge transporting gain can be obtained in a noble donor–acceptor system (CT complex) composed of organic complex: tetrathiafulvalene–2,3,5,6-tetrafluoro-7,7,8,8-tetracyanoquinodimethane (TTF–F4TCNQ). The complex is small-gap organic metallic or semiconductor (less than 1 eV), and we predict having a high conductivity. We perform an approach to fabricate organic CT complex with high crystallinity by eutectic melting method. Our process is simple and shows crystal growth with improved crystallinity when combined with soft-lithography.

  3. Production of highly knotted DNA by means of cosmid circularization inside phage capsids

    Directory of Open Access Journals (Sweden)

    Trigueros Sonia

    2007-12-01

    Full Text Available Abstract Background The formation of DNA knots is common during biological transactions. Yet, functional implications of knotted DNA are not fully understood. Moreover, potential applications of DNA molecules condensed by means of knotting remain to be explored. A convenient method to produce abundant highly knotted DNA would be highly valuable for these studies. Results We had previously shown that circularization of the 11.2 kb linear DNA of phage P4 inside its viral capsid generates complex knots by the effect of confinement. We demonstrate here that this mechanism is not restricted to the viral genome. We constructed DNA cosmids as small as 5 kb and introduced them inside P4 capsids. Such cosmids were then recovered as a complex mixture of highly knotted DNA circles. Over 250 μg of knotted cosmid were typically obtained from 1 liter of bacterial culture. Conclusion With this biological system, DNA molecules of varying length and sequence can be shaped into very complex and heterogeneous knotted forms. These molecules can be produced in preparative amounts suitable for systematic studies and applications.

  4. Analysis of JC virus DNA replication using a quantitative and high-throughput assay

    International Nuclear Information System (INIS)

    Shin, Jong; Phelan, Paul J.; Chhum, Panharith; Bashkenova, Nazym; Yim, Sung; Parker, Robert; Gagnon, David; Gjoerup, Ole; Archambault, Jacques; Bullock, Peter A.

    2014-01-01

    Progressive Multifocal Leukoencephalopathy (PML) is caused by lytic replication of JC virus (JCV) in specific cells of the central nervous system. Like other polyomaviruses, JCV encodes a large T-antigen helicase needed for replication of the viral DNA. Here, we report the development of a luciferase-based, quantitative and high-throughput assay of JCV DNA replication in C33A cells, which, unlike the glial cell lines Hs 683 and U87, accumulate high levels of nuclear T-ag needed for robust replication. Using this assay, we investigated the requirement for different domains of T-ag, and for specific sequences within and flanking the viral origin, in JCV DNA replication. Beyond providing validation of the assay, these studies revealed an important stimulatory role of the transcription factor NF1 in JCV DNA replication. Finally, we show that the assay can be used for inhibitor testing, highlighting its value for the identification of antiviral drugs targeting JCV DNA replication. - Highlights: • Development of a high-throughput screening assay for JCV DNA replication using C33A cells. • Evidence that T-ag fails to accumulate in the nuclei of established glioma cell lines. • Evidence that NF-1 directly promotes JCV DNA replication in C33A cells. • Proof-of-concept that the HTS assay can be used to identify pharmacological inhibitor of JCV DNA replication

  5. Analysis of JC virus DNA replication using a quantitative and high-throughput assay

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Jong; Phelan, Paul J.; Chhum, Panharith; Bashkenova, Nazym; Yim, Sung; Parker, Robert [Department of Developmental, Molecular and Chemical Biology, Tufts University School of Medicine, Boston, MA 02111 (United States); Gagnon, David [Institut de Recherches Cliniques de Montreal (IRCM), 110 Pine Avenue West, Montreal, Quebec, Canada H2W 1R7 (Canada); Department of Biochemistry and Molecular Medicine, Université de Montréal, Montréal, Quebec (Canada); Gjoerup, Ole [Molecular Oncology Research Institute, Tufts Medical Center, Boston, MA 02111 (United States); Archambault, Jacques [Institut de Recherches Cliniques de Montreal (IRCM), 110 Pine Avenue West, Montreal, Quebec, Canada H2W 1R7 (Canada); Department of Biochemistry and Molecular Medicine, Université de Montréal, Montréal, Quebec (Canada); Bullock, Peter A., E-mail: Peter.Bullock@tufts.edu [Department of Developmental, Molecular and Chemical Biology, Tufts University School of Medicine, Boston, MA 02111 (United States)

    2014-11-15

    Progressive Multifocal Leukoencephalopathy (PML) is caused by lytic replication of JC virus (JCV) in specific cells of the central nervous system. Like other polyomaviruses, JCV encodes a large T-antigen helicase needed for replication of the viral DNA. Here, we report the development of a luciferase-based, quantitative and high-throughput assay of JCV DNA replication in C33A cells, which, unlike the glial cell lines Hs 683 and U87, accumulate high levels of nuclear T-ag needed for robust replication. Using this assay, we investigated the requirement for different domains of T-ag, and for specific sequences within and flanking the viral origin, in JCV DNA replication. Beyond providing validation of the assay, these studies revealed an important stimulatory role of the transcription factor NF1 in JCV DNA replication. Finally, we show that the assay can be used for inhibitor testing, highlighting its value for the identification of antiviral drugs targeting JCV DNA replication. - Highlights: • Development of a high-throughput screening assay for JCV DNA replication using C33A cells. • Evidence that T-ag fails to accumulate in the nuclei of established glioma cell lines. • Evidence that NF-1 directly promotes JCV DNA replication in C33A cells. • Proof-of-concept that the HTS assay can be used to identify pharmacological inhibitor of JCV DNA replication.

  6. Scalable whole-exome sequencing of cell-free DNA reveals high concordance with metastatic tumors.

    Science.gov (United States)

    Adalsteinsson, Viktor A; Ha, Gavin; Freeman, Samuel S; Choudhury, Atish D; Stover, Daniel G; Parsons, Heather A; Gydush, Gregory; Reed, Sarah C; Rotem, Denisse; Rhoades, Justin; Loginov, Denis; Livitz, Dimitri; Rosebrock, Daniel; Leshchiner, Ignaty; Kim, Jaegil; Stewart, Chip; Rosenberg, Mara; Francis, Joshua M; Zhang, Cheng-Zhong; Cohen, Ofir; Oh, Coyin; Ding, Huiming; Polak, Paz; Lloyd, Max; Mahmud, Sairah; Helvie, Karla; Merrill, Margaret S; Santiago, Rebecca A; O'Connor, Edward P; Jeong, Seong H; Leeson, Rachel; Barry, Rachel M; Kramkowski, Joseph F; Zhang, Zhenwei; Polacek, Laura; Lohr, Jens G; Schleicher, Molly; Lipscomb, Emily; Saltzman, Andrea; Oliver, Nelly M; Marini, Lori; Waks, Adrienne G; Harshman, Lauren C; Tolaney, Sara M; Van Allen, Eliezer M; Winer, Eric P; Lin, Nancy U; Nakabayashi, Mari; Taplin, Mary-Ellen; Johannessen, Cory M; Garraway, Levi A; Golub, Todd R; Boehm, Jesse S; Wagle, Nikhil; Getz, Gad; Love, J Christopher; Meyerson, Matthew

    2017-11-06

    Whole-exome sequencing of cell-free DNA (cfDNA) could enable comprehensive profiling of tumors from blood but the genome-wide concordance between cfDNA and tumor biopsies is uncertain. Here we report ichorCNA, software that quantifies tumor content in cfDNA from 0.1× coverage whole-genome sequencing data without prior knowledge of tumor mutations. We apply ichorCNA to 1439 blood samples from 520 patients with metastatic prostate or breast cancers. In the earliest tested sample for each patient, 34% of patients have ≥10% tumor-derived cfDNA, sufficient for standard coverage whole-exome sequencing. Using whole-exome sequencing, we validate the concordance of clonal somatic mutations (88%), copy number alterations (80%), mutational signatures, and neoantigens between cfDNA and matched tumor biopsies from 41 patients with ≥10% cfDNA tumor content. In summary, we provide methods to identify patients eligible for comprehensive cfDNA profiling, revealing its applicability to many patients, and demonstrate high concordance of cfDNA and metastatic tumor whole-exome sequencing.

  7. Functional role of a highly repetitive DNA sequence in anchorage of the mouse genome.

    Science.gov (United States)

    Neuer-Nitsche, B; Lu, X N; Werner, D

    1988-09-12

    The major portion of the eukaryotic genome consists of various categories of repetitive DNA sequences which have been studied with respect to their base compositions, organizations, copy numbers, transcription and species specificities; their biological roles, however, are still unclear. A novel quality of a highly repetitive mouse DNA sequence is described which points to a functional role: All copies (approximately 50,000 per haploid genome) of this DNA sequence reside on genomic Alu I DNA fragments each associated with nuclear polypeptides that are not released from DNA by proteinase K, SDS and phenol extraction. By this quality the repetitive DNA sequence is classified as a member of the sub-set of DNA sequences involved in tight DNA-polypeptide complexes which have been previously shown to be components of the subnuclear structure termed 'nuclear matrix'. From these results it has to be concluded that the repetitive DNA sequence characterized in this report represents or comprises a signal for a large number of site specific attachment points of the mouse genome in the nuclear matrix.

  8. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

    International Nuclear Information System (INIS)

    Wang, Deng-Liang; Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan; Yang, Hai-Tao; Wang, Jiang-Jie; Yao, Pei-Sen; Pan, Ru-Jun; Yang, Chaoyong James; Kang, De-Zhi

    2014-01-01

    Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K d 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K d 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy

  9. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Deng-Liang [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Yang, Hai-Tao; Wang, Jiang-Jie [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yao, Pei-Sen [Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Pan, Ru-Jun [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yang, Chaoyong James, E-mail: cyyang@xmu.edu.cn [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Kang, De-Zhi, E-mail: kdzy99988@163.com [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China)

    2014-10-31

    Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K{sub d} 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K{sub d} 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy.

  10. A high-throughput and quantitative method to assess the mutagenic potential of translesion DNA synthesis

    Science.gov (United States)

    Taggart, David J.; Camerlengo, Terry L.; Harrison, Jason K.; Sherrer, Shanen M.; Kshetry, Ajay K.; Taylor, John-Stephen; Huang, Kun; Suo, Zucai

    2013-01-01

    Cellular genomes are constantly damaged by endogenous and exogenous agents that covalently and structurally modify DNA to produce DNA lesions. Although most lesions are mended by various DNA repair pathways in vivo, a significant number of damage sites persist during genomic replication. Our understanding of the mutagenic outcomes derived from these unrepaired DNA lesions has been hindered by the low throughput of existing sequencing methods. Therefore, we have developed a cost-effective high-throughput short oligonucleotide sequencing assay that uses next-generation DNA sequencing technology for the assessment of the mutagenic profiles of translesion DNA synthesis catalyzed by any error-prone DNA polymerase. The vast amount of sequencing data produced were aligned and quantified by using our novel software. As an example, the high-throughput short oligonucleotide sequencing assay was used to analyze the types and frequencies of mutations upstream, downstream and at a site-specifically placed cis–syn thymidine–thymidine dimer generated individually by three lesion-bypass human Y-family DNA polymerases. PMID:23470999

  11. Pressure melting and ice skating

    Science.gov (United States)

    Colbeck, S. C.

    1995-10-01

    Pressure melting cannot be responsible for the low friction of ice. The pressure needed to reach the melting temperature is above the compressive failure stress and, if it did occur, high squeeze losses would result in very thin films. Pure liquid water cannot coexist with ice much below -20 °C at any pressure and friction does not increase suddenly in that range. If frictional heating and pressure melting contribute equally, the length of the wetted contact could not exceed 15 μm at a speed of 5 m/s, which seems much too short. If pressure melting is the dominant process, the water films are less than 0.08 μm thick because of the high pressures.

  12. Poly(1,4-cyclohexanedimethylene 2,6-naphthalate polyester with high melting point: Effect of different synthesis methods on molecular weight and properties

    Directory of Open Access Journals (Sweden)

    N. Kasmi

    2018-03-01

    Full Text Available In the current manuscript, a new approach for the synthesis of poly(1,4- cyclohexanedimethylene 2,6-naphthalate (PCHDMN derived from dimethyl 2,6-naphthalenedicarboxylate (2,6-DMN and 1,4-Cyclohexanedimethanol (CHDM via melt polycondensation method is introduced. The effect of three different synthesis pathways, polycondensation time and temperature on polyesters molecular weight increase has been investigated. All of the prepared samples were characterized measuring their intrinsic viscosity (IV, thermal properties and morphology with differential scanning calorimetry (DSC and wide-angle X-ray diffraction (WAXD, respectively. The results demonstrated the effectiveness of the synthesis pathway proposed for the preparation of PCHDMN, resulting in high molecular weight (IV value around 0.5 dL/g and much shorter reaction time. Melt polycondensation temperatures above melting point of polyester should be avoided to be used due to the decomposition of polyester. This was proved by thermogravimetric analysis (TGA and Pyrolysis-gas chromatography–mass spectroscopy analysis (Py-GC/MS.

  13. Assessment of genetic mutations in the XRCC2 coding region by high resolution melting curve analysis and the risk of differentiated thyroid carcinoma in Iran

    Directory of Open Access Journals (Sweden)

    Shima Fayaz

    2012-01-01

    Full Text Available Homologous recombination (HR is the major pathway for repairing double strand breaks (DSBs in eukaryotes and XRCC2 is an essential component of the HR repair machinery. To evaluate the potential role of mutations in gene repair by HR in individuals susceptible to differentiated thyroid carcinoma (DTC we used high resolution melting (HRM analysis, a recently introduced method for detecting mutations, to examine the entire XRCC2 coding region in an Iranian population. HRM analysis was used to screen for mutations in three XRCC2 coding regions in 50 patients and 50 controls. There was no variation in the HRM curves obtained from the analysis of exons 1 and 2 in the case and control groups. In exon 3, an Arg188His polymorphism (rs3218536 was detected as a new melting curve group (OR: 1.46; 95%CI: 0.432-4.969; p = 0.38 compared with the normal melting curve. We also found a new Ser150Arg polymorphism in exon 3 of the control group. These findings suggest that genetic variations in the XRCC2 coding region have no potential effects on susceptibility to DTC. However, further studies with larger populations are required to confirm this conclusion.

  14. A Novel Low Temperature PCR Assured High-Fidelity DNA Amplification

    Directory of Open Access Journals (Sweden)

    Shaoxia Zhou

    2013-06-01

    Full Text Available As previously reported, a novel low temperature (LoTemp polymerase chain reaction (PCR catalyzed by a moderately heat-resistant (MHR DNA polymerase with a chemical-assisted denaturation temperature set at 85 °C instead of the conventional 94–96 °C can achieve high-fidelity DNA amplification of a target DNA, even after up to 120 PCR thermal cycles. Furthermore, such accurate amplification is not achievable with conventional PCR. Now, using a well-recognized L1 gene segment of the human papillomavirus (HPV type 52 (HPV-52 as the template for experiments, we demonstrate that the LoTemp high-fidelity DNA amplification is attributed to an unusually high processivity and stability of the MHR DNA polymerase whose high fidelity in template-directed DNA synthesis is independent of non-existent 3'–5' exonuclease activity. Further studies and understanding of the characteristics of the LoTemp PCR technology may facilitate implementation of DNA sequencing-based diagnostics at the point of care in community hospital laboratories.

  15. Tungsten disulfide nanosheet and exonuclease III co-assisted amplification strategy for highly sensitive fluorescence polarization detection of DNA glycosylase activity

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Jingjin; Ma, Yefei [Key Laboratory for the Chemistry and Molecular Engineering of Medicinal Resources of Education Ministry, Guangxi Normal University, Guilin, 541004 (China); Kong, Rongmei [The Key Laboratory of Life-Organic Analysis, College of Chemistry and Chemical Engineering, Qufu Normal University, Qufu, Shandong 273165 (China); Zhang, Liangliang, E-mail: liangzhang319@163.com [Key Laboratory for the Chemistry and Molecular Engineering of Medicinal Resources of Education Ministry, Guangxi Normal University, Guilin, 541004 (China); Yang, Wen; Zhao, Shulin [Key Laboratory for the Chemistry and Molecular Engineering of Medicinal Resources of Education Ministry, Guangxi Normal University, Guilin, 541004 (China)

    2015-08-05

    Herein, we introduced a tungsten disulfide (WS{sub 2}) nanosheet and exonuclease III (Exo III) co-assisted signal amplification strategy for highly sensitive fluorescent polarization (FP) assay of DNA glycosylase activity. Two DNA glycosylases, uracil-DNA glycosylase (UDG) and human 8-oxoG DNA glycosylase 1 (hOGG1), were tested. A hairpin-structured probe (HP) which contained damaged bases in the stem was used as the substrate. The removal of damaged bases from substrate by DNA glycosylase would lower the melting temperature of HP. The HP was then opened and hybridized with a FAM dye-labeled single strand DNA (DP), generating a duplex with a recessed 3′-terminal of DP. This design facilitated the Exo III-assisted amplification by repeating the hybridization and digestion of DP, liberating numerous FAM fluorophores which could not be adsorbed on WS{sub 2} nanosheet. Thus, the final system exhibited a small FP signal. However, in the absence of DNA glycosylases, no hybridization between DP and HP was occurred, hampering the hydrolysis of DP by Exo III. The intact DP was then adsorbed on the surface of WS{sub 2} nanosheet that greatly amplified the mass of the labeled-FAM fluorophore, resulting in a large FP value. With the co-assisted amplification strategy, the sensitivity was substantially improved. In addition, this method was applied to detect UDG activity in cell extracts. The study of the inhibition of UDG was also performed. Furthermore, this method is simple in design, easy in implementation, and selective, which holds potential applications in the DNA glycosylase related mechanism research and molecular diagnostics. - Highlights: • A fluorescence polarization strategy for DNA glycosylase activity detection was developed. • The present method was based on WS{sub 2} nanosheet and exonuclease III co-assisted signal amplification. • A high sensitivity and desirable selectivity were achieved. • This method provides a promising universal platform for