WorldWideScience

Sample records for hybridization fish denaturing

  1. Marine Fish Hybridization

    KAUST Repository

    He, Song

    2017-04-01

    Natural hybridization is reproduction (without artificial influence) between two or more species/populations which are distinguishable from each other by heritable characters. Natural hybridizations among marine fishes were highly underappreciated due to limited research effort; it seems that this phenomenon occurs more often than is commonly recognized. As hybridization plays an important role in biodiversity processes in the marine environment, detecting hybridization events and investigating hybridization is important to understand and protect biodiversity. The first chapter sets the framework for this disseration study. The Cohesion Species Concept was selected as the working definition of a species for this study as it can handle marine fish hybridization events. The concept does not require restrictive species boundaries. A general history and background of natural hybridization in marine fishes is reviewed during in chapter as well. Four marine fish hybridization cases were examed and documented in Chapters 2 to 5. In each case study, at least one diagnostic nuclear marker, screened from among ~14 candidate markers, was found to discriminate the putative hybridizing parent species. To further investigate genetic evidence to support the hybrid status for each hybrid offspring in each case, haploweb analysis on diagnostic markers (nuclear and/or mitochondrial) and the DAPC/PCA analysis on microsatellite data were used. By combining the genetic evidences, morphological traits, and ecological observations together, the potential reasons that triggered each hybridization events and the potential genetic/ecology effects could be discussed. In the last chapter, sequences from 82 pairs of hybridizing parents species (for which COI barcoding sequences were available either on GenBank or in our lab) were collected. By comparing the COI fragment p-distance between each hybridizing parent species, some general questions about marine fish hybridization were discussed: Is

  2. Detection of the End Point Temperature of Thermal Denatured Protein in Fish and Chicken Meat Through SDS-PAGE Electrophoresis

    Institute of Scientific and Technical Information of China (English)

    GAO Hongwei; MAO Mao; LIANG Chengzhu; LIN Chao; XIANG Jianhai

    2009-01-01

    Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied in the detection of the end point temperature (EPT) of thermal denatured protein in fish and meat in this study. It was also used in studying the thermal denatured temperature range of proteins in salmon and chicken meat. The results show that the temperature ranges of denatured proteins were from 65℃ to 75℃, and these temperature ranges were influenced by the processing methods. Through SDS-PAGE, the features of repeated heating thermal denatured proteins under the same temperature and processing time were studied. The electrophoresis pat-terns of thermal denatured proteins determined through repeated heating at the same temperature did not exhibit any change. For the detection of cooked fish and meat samples, they were subjected to applying the SDS-PAGE method, which revealed an EPT ranging from 60℃ to 80℃.

  3. Molecular assessment of collagen denaturation in decellularized tissues using a collagen hybridizing peptide.

    Science.gov (United States)

    Hwang, Jeongmin; San, Boi Hoa; Turner, Neill J; White, Lisa J; Faulk, Denver M; Badylak, Stephen F; Li, Yang; Yu, S Michael

    2017-04-15

    Decellularized extracellular matrix (ECM) derived from tissues and organs are emerging as important scaffold materials for regenerative medicine. Many believe that preservation of the native ECM structure during decellularization is highly desirable. However, because effective techniques to assess the structural damage in ECM are lacking, the disruptive effects of a decellularization method and the impact of the associated structural damage upon the scaffold's regenerative capacity are often debated. Using a novel collagen hybridizing peptide (CHP) that specifically binds to unfolded collagen chains, we investigated the molecular denaturation of collagen in the ECM decellularized by four commonly used cell-removing detergents: sodium dodecyl sulfate (SDS), 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), sodium deoxycholate (SD), and Triton X-100. Staining of the detergent-treated porcine ligament and urinary bladder matrix with carboxyfluorescein-labeled CHP demonstrated that SDS and Triton X-100 denature the triple helical collagen molecule while CHAPS and SD do not, although second harmonic generation imaging and transmission electron microscopy (TEM) revealed that all four detergents disrupt collagen fibrils. Our findings from the CHP staining were further confirmed by the circular dichroism spectra of intact triple helical collagen molecules in CHAPS and SD solutions, and the TEM images of CHP-conjugated gold nanoparticles binding only to the SDS and Triton X-100 treated collagen fibrils. CHP is a powerful new tool for direct and reliable measurement of denatured collagen molecules in decellularized tissues. It is expected to have wide applications in the development and standardization of the tissue/organ decellularization technology. Preservation of the native ECM structure in decellularized tissues is highly desirable, since denaturation of ECM molecules (e.g., collagen) during decellularization can strongly influence the cellular response

  4. Distant hybridization leads to different ploidy fishes

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Distant hybridization makes it possible to transfer the genome of one species to another, which results in changes in phenotypes and genotypes of the progenies. This study shows that distant hybridization or the combination of this method with gynogenesis or androgenesis lead to different ploidy fishes with genetic variation, including fertile tetraploid hybrids, sterile triploid hybrids, fertile diploid hybrids, fertile diploid gynogenetic fish, and their derived progenies. The formations of the different ploidy fishes depend on the genetic relationship between the parents. In this study, several types of distant hybridization, including red crucian carp (Carassius auratus red var.) (2n=100, abbreviated as RCC) (♀)×common carp (Cyprinus carpio L.) (2n=100, abbreviated as CC) (♂), and RCC (2n=100) (♀)×blunt snout bream (Megalobrama amblycephala) (2n=48, abbreviated as BSB) (♂) are described. In the distant hybridization of RCC (♀)×CC (♂), bisexual fertile F3–F18 allotetraploid hybrids (4n=200, abbreviated as 4nAT) were formed. The diploid hybrid eggs and diploid sperm generated by the females and males of 4nAT developed into diploid gynogenetic hybrids and diploid androgenetic hybrids, respectively, by gynogenesis and androgenesis, without treatment for doubling the chromosome. Improved tetraploid hybrids and improved diploid fishes with genetic variation were derived from the gynogenetic hybrid line. The improved diploid fishes included the high-body RCC and high-body goldfish. The formation of the tetraploid hybrids was related to the occurrence of unreduced gametes generated from the diploid hybrids, which involved in premeiotic endoreduplication, endomitosis, or fusion of germ cells. The sterile triploid hybrids (3n=150) were produced on a large scale by crossing the males of tetraploid hybrids with females of diploid fish (2n=100). In another distant hybridization of RCC (♀)×BSB (♂), different ploidy fishes were obtained, including

  5. Distant hybridization leads to different ploidy fishes.

    Science.gov (United States)

    Liu, ShaoJun

    2010-04-01

    Distant hybridization makes it possible to transfer the genome of one species to another, which results in changes in phenotypes and genotypes of the progenies. This study shows that distant hybridization or the combination of this method with gynogenesis or androgenesis lead to different ploidy fishes with genetic variation, including fertile tetraploid hybrids, sterile triploid hybrids, fertile diploid hybrids, fertile diploid gynogenetic fish, and their derived progenies. The formations of the different ploidy fishes depend on the genetic relationship between the parents. In this study, several types of distant hybridization, including red crucian carp (Carassius auratus red var.) (2n=100, abbreviated as RCC) (female) x common carp (Cyprinus carpio L.) (2n=100, abbreviated as CC) (male), and RCC (2n=100) (female) x blunt snout bream (Megalobrama amblycephala) (2n=48, abbreviated as BSB) (male) are described. In the distant hybridization of RCC (female) x CC (male), bisexual fertile F(3)-F(18) allotetraploid hybrids (4n=200, abbreviated as 4nAT) were formed. The diploid hybrid eggs and diploid sperm generated by the females and males of 4nAT developed into diploid gynogenetic hybrids and diploid androgenetic hybrids, respectively, by gynogenesis and androgenesis, without treatment for doubling the chromosome. Improved tetraploid hybrids and improved diploid fishes with genetic variation were derived from the gynogenetic hybrid line. The improved diploid fishes included the high-body RCC and high-body goldfish. The formation of the tetraploid hybrids was related to the occurrence of unreduced gametes generated from the diploid hybrids, which involved in premeiotic endoreduplication, endomitosis, or fusion of germ cells. The sterile triploid hybrids (3n=150) were produced on a large scale by crossing the males of tetraploid hybrids with females of diploid fish (2n=100). In another distant hybridization of RCC (female) x BSB (male), different ploidy fishes were

  6. Third-strand in situ hybridization (TISH) to non-denatured metaphase spreads and interphase nuclei.

    Science.gov (United States)

    Johnson, M D; Fresco, J R

    1999-07-01

    A methodology has been developed for binding oligodeoxyribonucleotide 'third strands' to duplex DNA targets in fixed but not additionally denatured metaphase spreads and interphase nuclei under conditions found to be optimal in solution. Third-strand in situ hybridization (TISH) at pH 6.0 of a psoralen- and biotin-modified 16-nucleotide homopyrimidine third strand to a unique multicopy target sequence in human chromosome 17 alpha-satellite (D17Z1 locus) is described. UVA-photofixed third strands, rendered fluorescent by fluorescein isothiocyanate-labeled avidin, are reproducibly centromere-specific for chromosome 17, and visible without amplification of the signal in lymphocyte and somatic cell hybrid spreads and interphase nuclei. Two third-strand-specific D17Z1 haplotypes were identified. TISH has potential diagnostic, biochemical, and flow cytometric applicability to native metaphase and interphase chromatin.

  7. Immobilization of denatured DNA to macroporous supports: II. Steric and kinetic parameters of heterogeneous hybridization reactions.

    Science.gov (United States)

    Bünemann, H

    1982-11-25

    The accessibility of immobilized DNA has been shown to depend more crucially on the method of immobilization than on the type of support used for fixation. When sonicated denatured DNA is coupled via diazotization or via cyanogen bromide reaction to solid Sephadex G-25 and Cellex 410 or to macroporous Sephacryl S-500 and Sepharose C1-6B its accessibility varies from 100 to 24 percent. Generally the loss of accessibility is linked to a depression of the melting temperature of DNA helices formed on the support. This correlation shows a characteristic course for a particular coupling method. DNA coupled under denaturing conditions may become totally inaccessible when only 3 percent of its bases are involved in the covalent linkage. Kinetic experiments with sonicated E.coli DNA have shown that the rate constants for renaturation or hybridization reactions are very similar for DNA immobilized by different methods to solid or macroporous supports. Generally the second order rate constant for a heterogeneous reaction (between mobile and immobilized DNA) is about one order of magnitude smaller than that of the analogous homogeneous reaction (in solution).

  8. Hybrid seine for full fish community collections

    Science.gov (United States)

    McKenna, James E.; Waldt, Emily M.; Abbett, Ross; David, Anthony; Snyder, James

    2013-01-01

    Seines are simple and effective fish collection gears, but the net mesh size influences how well the catch represents the fish communities. We designed and tested a hybrid seine with a dual-mesh bag (1/4″ and 1/8″) and compared the fish assemblage collected by each mesh. The fine-mesh net retained three times as many fish and collected more species (as many as eight), including representatives of several rare species, than did the coarser mesh. The dual-mesh bag permitted us to compare both sizes and species retained by each layer and to develop species-specific abundance correction factors, which allowed comparison of catches with the coarse-mesh seine used for earlier collections. The results indicate that a hybrid seine with coarse-mesh wings and a fine-mesh bag would enhance future studies of fish communities, especially when small-bodied fishes or early life stages are the research focus.

  9. Application of fluorescence in situ hybridization (FISH) to the ...

    African Journals Online (AJOL)

    Application of fluorescence in situ hybridization (FISH) to the analysis of ... In this study, fluorescent in situ hybridization (FISH) as a culture-independent molecular ... a high percentage and took place in an oily biological system under aerobic ...

  10. Effect of enzymatic fish frame protein hydrolysate on freeze-induced denaturation of myofibrillar protein from bighead carp fish (Aristichthys nobilis) during frozen storage

    Institute of Scientific and Technical Information of China (English)

    Xue Yong; Xue Changhu; Zhang Yongqin; Li Zhaojie; Gao Ruichang; Gao Xin

    2006-01-01

    Three kinds of fish frame protein hydrolysates (PPH, APH and FPH) were prepared from fish frame of red drum (Sciaenops ocellatus) by papain, alkaline proteinase and flavorzyme treatment. The hydrolysates were mainly composed of peptide (83.5%-84.6%) and displayed different molecular weight distribution pattern. The protective effects of hydrolysates on the freeze-induced denaturation of myofibrillar protein (Mf) from bighead carp (Aristichthys nobilis) mince during storage at -20℃ for 12 weeks were investigated. The hydrolysate (5% dried weight/wet weight) reduced the freeze-induced denaturation of Mf as evidenced by the lowered decrease in Ca-ATPase activity and reactive sulfhydryl contents as well as the impeded increase in surface hydrophobicity. Microscopic photographs indicated that the hydrolysates inhibited the growth of ice crystal in fish mince, and then prevented the aggregation of Mf during frozen storage. The protective effects of hydrolysates on freeze-induced denaturation of Mf were influenced by the molecular weight distribution. PPH had strongest cryoprotective ability among three hydrolysates.

  11. Isothermal DNA origami folding: avoiding denaturing conditions for one-pot, hybrid-component annealing

    Science.gov (United States)

    Kopielski, Andreas; Schneider, Anne; Csáki, Andrea; Fritzsche, Wolfgang

    2015-01-01

    The DNA origami technique offers great potential for nanotechnology. Using biomolecular self-assembly, defined 2D and 3D nanoscale DNA structures can be realized. DNA origami allows the positioning of proteins, fluorophores or nanoparticles with an accuracy of a few nanometers and enables thereby novel nanoscale devices. Origami assembly usually includes a thermal denaturation step at 90 °C. Additional components used for nanoscale assembly (such as proteins) are often thermosensitive, and possibly damaged by such harsh conditions. They have therefore to be attached in an extra second step to avoid defects. To enable a streamlined one-step nanoscale synthesis - a so called one-pot folding - an adaptation of the folding procedures is required. Here we present a thermal optimization of this process for a 2D DNA rectangle-shaped origami resulting in an isothermal assembly protocol below 60 °C without thermal denaturation. Moreover, a room temperature protocol is presented using the chemical additive betaine, which is biocompatible in contrast to chemical denaturing approaches reported previously.The DNA origami technique offers great potential for nanotechnology. Using biomolecular self-assembly, defined 2D and 3D nanoscale DNA structures can be realized. DNA origami allows the positioning of proteins, fluorophores or nanoparticles with an accuracy of a few nanometers and enables thereby novel nanoscale devices. Origami assembly usually includes a thermal denaturation step at 90 °C. Additional components used for nanoscale assembly (such as proteins) are often thermosensitive, and possibly damaged by such harsh conditions. They have therefore to be attached in an extra second step to avoid defects. To enable a streamlined one-step nanoscale synthesis - a so called one-pot folding - an adaptation of the folding procedures is required. Here we present a thermal optimization of this process for a 2D DNA rectangle-shaped origami resulting in an isothermal assembly

  12. Immobilization of denatured DNA to macroporous supports: II. Steric and kinetic parameters of heterogeneous hybridization reactions.

    OpenAIRE

    Bünemann, H

    1982-01-01

    The accessibility of immobilized DNA has been shown to depend more crucially on the method of immobilization than on the type of support used for fixation. When sonicated denatured DNA is coupled via diazotization or via cyanogen bromide reaction to solid Sephadex G-25 and Cellex 410 or to macroporous Sephacryl S-500 and Sepharose C1-6B its accessibility varies from 100 to 24 percent. Generally the loss of accessibility is linked to a depression of the melting temperature of DNA helices forme...

  13. Marine genetic swamping: hybrids replace an obligately estuarine fish.

    Science.gov (United States)

    Roberts, David G; Gray, Charles A; West, Ronald J; Ayre, David J

    2010-02-01

    Populations of obligately estuarine taxa are potentially small and isolated and may lack genetic variation and display regional differentiation as a result of drift and inbreeding. Hybridization with a wide-ranging marine congener should introduce genetic variation and reduce the effects of inbreeding depression and genetic drift. However, high levels of hybridization can cause demographic and genetic swamping. In southeastern Australia hybridization occurs between obligately estuarine Black bream (Acanthopagrus butcheri) and migratory marine Yellowfin bream (Acanthopagrus australis). Here, we surveyed genetic variation at eight microsatellite loci and the mitochondrial control region of juvenile fish from five coastal lagoons (including temporal replication in two lagoons) (total n = 970) to determine the frequency and persistence of hybridization, and its likely consequence for the estuarine restricted A. butcheri. Of 688 juvenile fish genotyped 95% were either A. australis (347) or hybrids (309); only 5% (32) were A. butcheri. Most hybrids were later generation hybrids or A. butcheri backcrosses, which are likely multi-generational residents within lagoons. Far greater proportions of hybrid juveniles were found within two lagoons that are generally closed to the ocean (>90% hybrid fish within generally closed lagoons vs. 12-27% in permanently or intermittently open lagoons). In both lagoons, this was consistent across multiple cohorts of fish [79-97% hybrid fish (n = 282)]. Hybridization and introgression represent a major threat to the persistence of A. butcheri and have yet to be investigated for large numbers of estuarine taxa.

  14. Human Papillomavirus Genotyping After Denaturation of Specimens for Hybrid Capture 2 Testing: Feasibility Study for the HPV Persistence and Progression Cohort†

    OpenAIRE

    2007-01-01

    Human papillomavirus (HPV) genotyping could be clinically useful, depending on the results of large, prospective studies like the HPV Persistence and Progression cohort. The cohort is based on genotyping and follow-up of Hybrid Capture-positive women at Kaiser Permanente, Northern California. HPV DNA testing by Hybrid Capture 2 requires denaturation with alkali, possibly damaging the DNA for optimal PCR-based genotyping. A feasibility study was conducted on paired aliquots of anonymized speci...

  15. Human papillomavirus genotyping after denaturation of specimens for Hybrid Capture 2 testing: feasibility study for the HPV persistence and progression cohort.

    Science.gov (United States)

    LaMere, Brandon J; Kornegay, Janet; Fetterman, Barbara; Sadorra, Mark; Shieh, Jen; Castle, Philip E

    2007-12-01

    Human papillomavirus (HPV) genotyping could be clinically useful, depending on the results of large, prospective studies like the HPV persistence and progression (PaP) cohort. The cohort is based on genotyping and follow-up of Hybrid Capture-positive women at Kaiser Permanente, Northern California. HPV DNA testing by Hybrid Capture 2 requires denaturation with alkali, possibly damaging the DNA for optimal PCR-based genotyping. A feasibility study was conducted on paired aliquots of anonymized specimens from 100 women with low-grade intraepithelial lesion cytology. Test aliquots were left in denaturant for 10 or 18h at 4 degrees C and then neutralized; comparison aliquots were not denatured but diluted to match the timing, temperature, concentration and salt conditions of the treated specimens. The masked aliquots were tested using a commercialized PCR-based assay that detects of 37 HPV genotypes. There was no overall effect of treatment on test positivity or number of types. HPV16 was marginally more likely to be detected in untreated versus treated aliquots (P=0.09) but HPV45 was marginally more likely to be detected in treated than untreated aliquots (P=0.07), suggesting that these differences represented chance (intra-test variability). It can be concluded that residual Hybrid Capture-positive specimens can be genotyped by PCR after Hybrid Capture 2 processing.

  16. Ancient hybridization and genomic stabilization in a swordtail fish.

    Science.gov (United States)

    Schumer, Molly; Cui, Rongfeng; Powell, Daniel L; Rosenthal, Gil G; Andolfatto, Peter

    2016-06-01

    A rapidly increasing body of work is revealing that the genomes of distinct species often exhibit hybrid ancestry, presumably due to postspeciation hybridization between closely related species. Despite the growing number of documented cases, we still know relatively little about how genomes evolve and stabilize following hybridization, and to what extent hybridization is functionally relevant. Here, we examine the case of Xiphophorus nezahualcoyotl, a teleost fish whose genome exhibits significant hybrid ancestry. We show that hybridization was relatively ancient and is unlikely to be ongoing. Strikingly, the genome of X. nezahualcoyotl has largely stabilized following hybridization, distinguishing it from examples such as human-Neanderthal hybridization. Hybridization-derived regions are remarkably distinct from other regions of the genome, tending to be enriched in genomic regions with reduced constraint. These results suggest that selection has played a role in removing hybrid ancestry from certain functionally important regions. Combined with findings in other systems, our results raise many questions about the process of genomic stabilization and the role of selection in shaping patterns of hybrid ancestry in the genome. © 2016 John Wiley & Sons Ltd.

  17. Parasitic fauna in hybrid tambacu from fish farms

    Directory of Open Access Journals (Sweden)

    Ronilson Macedo Silva

    2013-08-01

    Full Text Available The objective of this work was to evaluate the parasitic fauna of hybrid tambacu (Colossoma macropomum x Piaractus mesopotamicus from fish farms and the host-parasite relationship. A hundred and fourteen fish were collected from four fish farms in Macapá, in the state of Amapá, Brazil, 80.7% of which were infected by: Ichthyophthirius multifiliis (Ciliophora; Piscinoodinium pillulare (Dinoflagellida; Anacanthorus spatulatus, Notozothecium janauachensis, and Mymarothecium viatorum (Monogenoidea; Neoechinorhynchus buttnerae (Acanthocephala; Cucullanus colossomi (Nematoda; Perulernaea gamitanae (Lernaeidae; and Proteocephalidae larvae (Cestoda. A total of 8,136,252 parasites were collected from the examined fish. This is the first record of N. buttnerae, C. colossomi, N. janauachensis, M. viatorum, and Proteocephalidae for hybrid tambacu in Brazil. Ichthyophthirius multifiliis was the most prevalent parasite, whereas endohelminths were the less. A positive correlation was observed between number of I. multifiliis and total length and weight of fish, as well as between number of P. gamitanae and total length. The infection by I. multifiliis had association with the parasitism by Monogenoidea. Low water quality contributes to high parasitism of hybrid tambacu by ectoparasites, which, however, does not influence the relative condition factor of fish.

  18. The chromosomal constitution of fish hybrid lineage revealed by 5S rDNA FISH.

    Science.gov (United States)

    Zhang, Chun; Ye, Lihai; Chen, Yiyi; Xiao, Jun; Wu, Yanhong; Tao, Min; Xiao, Yamei; Liu, Shaojun

    2015-12-03

    The establishment of the bisexual fertile fish hybrid lineage including the allodiploid and allotetraploid hybrids, from interspecific hybridization of red crucian carp (Carassius auratus red var. 2n = 100, 2n = AA) (♀) × common carp (Cyprinus carpio L. 2n = 100, 2n = BB) (♂), provided a good platform to investigate genetic relationship between the parents and their hybrid progenies. The chromosomal inheritance of diploid and allotetraploid hybrid progenies in successive generations, was studied by applying 5S rDNA fluorescence in situ hybridization. Signals of 5S rDNA distinguished the chromosomal constitution of common carp (B-genome) from red crucian carp (A-genome), in which two strong signals were observed on the first submetacentric chromosome, while no major signal was found in common carp. After fish hybridization, one strong signal of 5S rDNA was detected in the same locus on the chromosome of diploid hybrids. As expected, two strong signals were observed in 4nF3 tetraploid hybrids offspring and it is worth mentioning that two strong signals were detected in a separating bivalent of a primary spermatocyte in 4nF3. Furthermore, the mitosis of heterozygous chromosomes was shown normal and stable with blastular tissue histological studies. We revealed that 5S rDNA signal can be applied to discern A-genome from B-genome, and that 5S rDNA bearing chromosomes can be stably passed down in successive generations. Our work provided a significant method in fish breeding and this is important for studies in fish evolutionary biology.

  19. [Denaturalized psychoanalysts].

    Science.gov (United States)

    Peglau, Andreas

    2011-01-01

    This paper presents hitherto unknown material from the German Foreign Office referring to the denaturalization of Therese Benedek, Bruno Bettelheim, Adolf Storfer and Wilhelm Reich by Nazi Germany. It corroborates the finding that nobody was persecuted by the Nazis solely on the basis of psychoanalytic activities or membership in a psychoanalytic organization.

  20. Study of the Artificial Fish Swarm Algorithm for Hybrid Clustering

    Directory of Open Access Journals (Sweden)

    Hongwei Zhao

    2015-06-01

    Full Text Available The basic Artificial Fish Swarm (AFS Algorithm is a new type of an heuristic swarm intelligence algorithm, but it is difficult to optimize to get high precision due to the randomness of the artificial fish behavior, which belongs to the intelligence algorithm. This paper presents an extended AFS algorithm, namely the Cooperative Artificial Fish Swarm (CAFS, which significantly improves the original AFS in solving complex optimization problems. K-medoids clustering algorithm is being used to classify data, but the approach is sensitive to the initial selection of the centers with low quality of the divided cluster. A novel hybrid clustering method based on the CAFS and K-medoids could be used for solving clustering problems. In this work, first, CAFS algorithm is used for optimizing six widely-used benchmark functions, coming up with comparative results produced by AFS and CAFS, then Particle Swarm Optimization (PSO is studied. Second, the hybrid algorithm with K-medoids and CAFS algorithms is used for data clustering on several benchmark data sets. The performance of the hybrid algorithm based on K-medoids and CAFS is compared with AFS and CAFS algorithms on a clustering problem. The simulation results show that the proposed CAFS outperforms the other two algorithms in terms of accuracy and robustness.

  1. Ancient hybridization fuels rapid cichlid fish adaptive radiations

    Science.gov (United States)

    Meier, Joana I.; Marques, David A.; Mwaiko, Salome; Wagner, Catherine E.; Excoffier, Laurent; Seehausen, Ole

    2017-01-01

    Understanding why some evolutionary lineages generate exceptionally high species diversity is an important goal in evolutionary biology. Haplochromine cichlid fishes of Africa's Lake Victoria region encompass >700 diverse species that all evolved in the last 150,000 years. How this ‘Lake Victoria Region Superflock' could evolve on such rapid timescales is an enduring question. Here, we demonstrate that hybridization between two divergent lineages facilitated this process by providing genetic variation that subsequently became recombined and sorted into many new species. Notably, the hybridization event generated exceptional allelic variation at an opsin gene known to be involved in adaptation and speciation. More generally, differentiation between new species is accentuated around variants that were fixed differences between the parental lineages, and that now appear in many new combinations in the radiation species. We conclude that hybridization between divergent lineages, when coincident with ecological opportunity, may facilitate rapid and extensive adaptive radiation. PMID:28186104

  2. Chromogenic in situ hybridization: a multicenter study comparing silver in situ hybridization with FISH.

    Science.gov (United States)

    Bartlett, J M S; Campbell, Fiona M; Ibrahim, Merdol; Wencyk, Peter; Ellis, Ian; Kay, Elaine; Connolly, Yvonne; O'Grady, Anthony; Di Palma, Silvana; Starczynski, Jane; Morgan, John M; Jasani, Bharat; Miller, Keith

    2009-10-01

    Our purposes were to perform a robust assessment of a new HER2 chromogenic in situ hybridization test and report on concordance of silver in situ hybridization (SISH) data with fluorescence in situ hybridization (FISH) data and on intraobserver and interlaboratory scoring consistency. HER2 results were scored from 45 breast cancers in 7 laboratories using the Ventana (Tucson, AZ) INFORM HER-2 SISH assay and in 1 central laboratory using a standard FISH assay. Overall, 94.8% of cases were successfully analyzed by SISH across the 6 participating laboratories that reported data. Concordance for diagnosis of HER2 amplification by SISH compared with FISH was high (96.0% overall). Intraobserver variability (8.0%) and intersite variability (12.66%) of absolute HER2/chromosome 17 ratios appear to be tightly controlled across all 6 participating laboratories. The Ventana INFORM HER-2 SISH assay is robust and reproducible, shows good concordance with a standard FISH assay, and complies with requirements in national guidelines for performance of diagnostic tests.

  3. Fluorescent in situ hybridization (FISH on the veterinary diagnostic field

    Directory of Open Access Journals (Sweden)

    Ján Dianovský

    2016-09-01

    Full Text Available Proceeding deals with of genomic changes detectable by FISH . The DSD syndrom in Yorkshire terrier 78,XY t (Y-;6p+ was observed by the use of X and Y FISH WCP probes. Following results indicated numerous genomic changes in cancers. Using comparative genomic hybridization numerous chromosomal rearrangements were detected, which indicated the heterogeneity in tumour growth. In Bernese Mountain Dog bitch,8 loses on chromosomes and gains on 18 different of chromosomes were detected. The last study was focused on chromosomal position and nucleotide sequencing of the LCA5L exons. Those were analysed in cattle of BTA1q44, sheep OAR1q43 and of CHI1q44 in the goats.

  4. Temperature controlling system based on FISH probe hybridization facility%基于FISH探针杂交设备的温度控制系统

    Institute of Scientific and Technical Information of China (English)

    李天庆; 张化; 张婉尧

    2015-01-01

    以玻片为基础的FISH,它的变性杂交过程完全实现自动化.FISH探针杂交仪内置一个ARM7微处理器,以控制不同温度变化,满足变性、杂交和固定温度等不同模式,可预置用按键编程进去的程序(显示屏上能指导编程),方便客户调用,提高工作效率.温度控制系统是FISH探针杂交设备的核心部分,决定了杂交仪的可靠性、稳定性及精准性.%By using FISH based on slide,automation in the degeneration process of hybrid can be achieved.Installed an internal ARM7 microprocessor,the FISH probe hybridization facility can control temperature,meet different modes such as denaturation,hybridization,fixed temperature,and so on.Procedure input by means of push-button can be preset(there are guidance on display screen)to make things easy for user,as a result,work efficiency can be improved.The temperature controlling system,which is the core of FISH probe hybridization facility and ensure its reliability,stability and accuracy.

  5. A New Type of Hybrid Fish-like Microrobot

    Institute of Scientific and Technical Information of China (English)

    Wei Zhang; Shu-Xiang Guo; Kinji Asaka

    2006-01-01

    In order to develop a new type of fish-like microrobot with swimming, walking, and floating motions, in our past research, we developed a hybrid microrobot actuated by ionic conducting polymer film (ICPF) actuators. But the microrobot had some problems in walking and floating motions. In this paper, we propose a concept of hybrid microrobot (see Fig. 1). The microrobot is actuated by a pair of caudal fins, a base with legs and an array of artificial swim bladders. We have developed a prototype of the base with legs and one artificial swim bladder, respectively, and carried out experiments for evaluating their characteristics. Experimental results show the base with legs can realize walking speed of 6 mm/s and rotating speed of 7.1 degrees/s respectively, and the prototype of the artificial swim bladder has a maximum floatage of 2.6 mN. The experimental results also indicate that the microrobot has some advantages, such as walking motion with 2 degrees of freedom, the walking ability on rough surface (sand paper), the controllable floatage, etc. This kind of fish-like microrobot is expected for industrial and medical applications.

  6. Parallel characterization of anaerobic toluene- and ethylbenzene-degrading microbial consortia by PCR-denaturing gradient gel electrophoresis, RNA-DNA membrane hybridization, and DNA microarray technology

    Science.gov (United States)

    Koizumi, Yoshikazu; Kelly, John J.; Nakagawa, Tatsunori; Urakawa, Hidetoshi; El-Fantroussi, Said; Al-Muzaini, Saleh; Fukui, Manabu; Urushigawa, Yoshikuni; Stahl, David A.

    2002-01-01

    A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis.

  7. Introgressive hybridization as a promoter of genome reshuffling in natural homoploid fish hybrids (Cyprinidae, Leuciscinae).

    Science.gov (United States)

    Pereira, C S A; Aboim, M A; Ráb, P; Collares-Pereira, M J

    2014-03-01

    Understanding the mechanisms underlying diversification and speciation by introgressive hybridization is currently one of the major challenges in evolutionary biology. Here, the analysis of hybridization between two pairs of Iberian Leuciscinae provided new data on independent hybrid zones involving Achondrostoma oligolepis (AOL) and Pseudochondrostoma duriense (PDU), and confirmed the occurrence of hybrids between AOL and Pseudochondrostoma polylepis (PPO). A multilevel survey combining morphological, genetic and cytogenomic markers on a vast population screening successfully sorted the selected fishes as admixed. Results were similar in both AOL × PDU and AOL × PPO systems. Overall, hybrid morphotypes, cytogenomic data and genetic profiling indicated preferential backcrossing and suggested AOL as a major genomic contributor. Moreover, results implied AOL as more permissive to introgression than PDU or PPO. Although PDU- and PPO-like individuals appeared more resilient to genome modifications, AOL appeared to be more involved and affected by the ongoing hybridization events, as chromosomal translocations were only found in AOL-like individuals. All hybrids analysed evidenced extensive ribosomal DNA (rDNA) polymorphism that was not found in parental species, but usually seen falling within the range of possible parental combinations. Yet, transgressive phenotypes that cannot be explained by normal recombination, including more rDNA clusters than expected or the occurrence of syntenic rDNAs, were also detected. Present results proved rapid genomic evolution providing the genetic novelty for species to persist. In addition, although the ultimate consequences of such apparently extensive and recurrent events remain unknown, modern genome-wide methodologies are of great promise towards answering questions concerning the causes, dynamics and impacts of hybridization.

  8. Metaphase FISH on a Chip: Miniaturized Microfluidic Device for Fluorescence in situ Hybridization

    DEFF Research Database (Denmark)

    Vedarethinam, Indumathi; Shah, Pranjul Jaykumar; Dimaki, Maria;

    2010-01-01

    Fluorescence in situ Hybridization (FISH) is a major cytogenetic technique for clinical genetic diagnosis of both inherited and acquired chromosomal abnormalities. Although FISH techniques have evolved and are often used together with other cytogenetic methods like CGH, PRINS and PNA-FISH, the pr......Fluorescence in situ Hybridization (FISH) is a major cytogenetic technique for clinical genetic diagnosis of both inherited and acquired chromosomal abnormalities. Although FISH techniques have evolved and are often used together with other cytogenetic methods like CGH, PRINS and PNA...

  9. Characterization of B chromosomes in Lilium hybrids through GISH and FISH

    NARCIS (Netherlands)

    Xie, S.L.; Marasek-Ciolakowska, A.; Ramanna, M.S.; Arens, P.F.P.; Visser, R.G.F.; Tuyl, van J.M.

    2014-01-01

    Supernumerary (B) chromosomes and small aberrant chromosomes were detected in Lilium hybrids and characterized through genomic in situ hybridization (GISH) and florescence in situ hybridization (FISH). Two small, supernumerary or B chromosomes were detected as extra chromosomes in a tetraploid plant

  10. Mitochondrial Genome Variation after Hybridization and Differences in the First and Second Generation Hybrids of Bream Fishes.

    Directory of Open Access Journals (Sweden)

    Wei-Zhuo Zhang

    Full Text Available Hybridization plays an important role in fish breeding. Bream fishes contribute a lot to aquaculture in China due to their economically valuable characteristics and the present study included five bream species, Megalobrama amblycephala, Megalobrama skolkovii, Megalobrama pellegrini, Megalobrama terminalis and Parabramis pekinensis. As maternal inheritance of mitochondrial genome (mitogenome involves species specific regulation, we aimed to investigate in which way the inheritance of mitogenome is affected by hybridization in these fish species. With complete mitogenomes of 7 hybrid groups of bream species being firstly reported in the present study, a comparative analysis of 17 mitogenomes was conducted, including representatives of these 5 bream species, 6 first generation hybrids and 6 second generation hybrids. The results showed that these 17 mitogenomes shared the same gene arrangement, and had similar gene size and base composition. According to the phylogenetic analyses, all mitogenomes of the hybrids were consistent with a maternal inheritance. However, a certain number of variable sites were detected in all F1 hybrid groups compared to their female parents, especially in the group of M. terminalis (♀ × M. amblycephala (♂ (MT×MA, with a total of 86 variable sites between MT×MA and its female parent. Among the mitogenomes genes, the protein-coding gene nd5 displayed the highest variability. The number of variation sites was found to be related to phylogenetic relationship of the parents: the closer they are, the lower amount of variation sites their hybrids have. The second generation hybrids showed less mitogenome variation than that of first generation hybrids. The non-synonymous and synonymous substitution rates (dN/dS were calculated between all the hybrids with their own female parents and the results indicated that most PCGs were under negative selection.

  11. Effect of graded fingerlings on hybrid catfish food fish size distribution

    Science.gov (United States)

    It is not unusual to have both 0.5 lb and 5 lb fish harvested from a single-batch hybrid catfish production pond at the end of the growing season. When that happens, farmers may be docked for fish that are either larger or smaller than the processor’s preferred size range. This study was conducted t...

  12. Role of fluorescence in situ hybridization (FISH) in sequencing the tomato genome

    Science.gov (United States)

    Chromosomes at various stages of the cell cycle can be used for localization of DNA probes via Fluorescence in situ hybridization (FISH). While mitotic metaphase chromosomes are demonstrably too short and compact for this purpose, long pachytene chromosomes are ideal. BACs that hybridize to euchrom...

  13. The Formation of the Polyploid Hybrids From Different Subfamily Fish Crossings and Its Evolutionary Significance

    OpenAIRE

    Liu, Shaojun; Qin, Qinbo; Xiao, Jun; Lu, Wenting; Shen, Jiamin; Li, Wei; Liu, Jifang; Duan, Wei; Zhang, Chun; Tao, De Min; Zhao, Rurong; Yan, Jinpeng; Liu, Yun

    2007-01-01

    This study provides genetic evidences at the chromosome, DNA content, DNA fragment and sequence, and morphological levels to support the successful establishment of the polyploid hybrids of red crucian carp × blunt snout bream, which belonged to a different subfamily of fish (Cyprininae subfamily and Cultrinae subfamily) in the catalog. We successfully obtained the sterile triploid hybrids and bisexual fertile tetraploid hybrids of red crucian carp (RCC) (♀) × blunt snout bream (BSB) (♂) as w...

  14. The formation of the polyploid hybrids from different subfamily fish crossings and its evolutionary significance.

    Science.gov (United States)

    Liu, Shaojun; Qin, Qinbo; Xiao, Jun; Lu, Wenting; Shen, Jiamin; Li, Wei; Liu, Jifang; Duan, Wei; Zhang, Chun; Tao, Min; Zhao, Rurong; Yan, Jinpeng; Liu, Yun

    2007-06-01

    This study provides genetic evidences at the chromosome, DNA content, DNA fragment and sequence, and morphological levels to support the successful establishment of the polyploid hybrids of red crucian carp x blunt snout bream, which belonged to a different subfamily of fish (Cyprininae subfamily and Cultrinae subfamily) in the catalog. We successfully obtained the sterile triploid hybrids and bisexual fertile tetraploid hybrids of red crucian carp (RCC) (female symbol) x blunt snout bream (BSB) (male symbol) as well as their pentaploid hybrids. The triploid hybrids possessed 124 chromosomes with two sets from RCC and one set from BSB; the tetraploid hybrids had 148 chromosomes with two sets from RCC and two sets from BSB. The females of tetraploid hybrids produced unreduced tetraploid eggs that were fertilized with the haploid sperm of BSB to generate pentaploid hybrids with 172 chromosomes with three sets from BSB and two sets from RCC. The ploidy levels of triploid, tetraploid, and pentaploid hybrids were confirmed by counting chromosomal number, forming chromosomal karyotype, and measuring DNA content and erythrocyte nuclear volume. The similar and different DNA fragments were PCR amplified and sequenced in triploid, tetraploid hybrids, and their parents, indicating their molecular genetic relationship and genetic markers. In addition, this study also presents results about the phenotypes and feeding habits of polyploid hybrids and discusses the formation mechanism of the polyploid hybrids. It is the first report on the formation of the triploid, tetraploid, and pentaploid hybrids by crossing parents with a different chromosome number in vertebrates. The formation of the polyploid hybrids is potentially interesting in both evolution and fish genetic breeding.

  15. Current and historical hybridization with differential introgression among three species of cyprinid fishes (genus Cyprinella).

    Science.gov (United States)

    Broughton, Richard E; Vedala, Krishna C; Crowl, Tessa M; Ritterhouse, Lauren L

    2011-05-01

    Hybridization is common among freshwater fishes, particular among the Cyprinidae. We used two mitochondrial genes and one nuclear gene to characterize hybridization among two species pairs of Cyprinella in southwestern North America. Genalogical patterns revealed that C. lutrensis and C. venusta are currently hybridizing in several localities producing apparent F(1), F(2) and backcross generations, yet there was no evidence for introgression outside of local hybrid zones. Alternatively, mitochondrial haplotypes from C. lutrensis appear to have introgressed into a C. lepida population in the Nueces River completely replacing the native C. lepida haplotype. There was no evidence of introgression of nuclear DNA and there does not appear to be ongoing hybridization. The population of C. lepida from the nearby Frio River exhibits no evidence of hybridization with C. lutrensis. Thus, contact between C. lutrensis and C. venusta results in the formation of localized hybrid swarms, while contact between C. lutrensis and C. lepida has resulted in complete mitochondrial introgression in the Nueces River or no apparent hybridization in the Frio River. The three different outcomes of contact between these species illustrate the variable nature of interspecific reproductive interactions and provide an excellent system in which to better understand the factors influencing hybridization among freshwater fishes.

  16. Comparison Between Fluorescent In Situ Hybridization (FISH) and ...

    African Journals Online (AJOL)

    ... were collected, for culture method and bacteria characterized by biochemical ... for 24 hours and later preserved in 70% alcohol before in situ hybridization test. ... were observed in 47 to 75% while the bacterium was isolated on culture in 7 ...

  17. DNA breakage detection-fluorescence in situ hybridization (DBD-FISH in buccal cells

    Directory of Open Access Journals (Sweden)

    E. I. Cortés-Gutiérrez

    2012-12-01

    Full Text Available DNA breakage detection-fluorescence in situ hybridization (DBD-FISH is a recently developed technique that allows cell-by-cell detection and quantification of DNA breakage in the whole genome or within specific DNA sequences. The present investigation was conducted to adapt the methodology of DBD-FISH to the visualization and evaluation of DNA damage in buccal epithelial cells. DBD-FISH revealed that DNA damage increased significantly according to H2O2 concentration (r2=0.91. In conclusion, the DBD-FISH technique is easy to apply in buccal cells and provides prompt results that are easy to interpret. Future studies are needed to investigate the potential applicability of a buccal cell DBD-FISH model to human biomonitoring and nutritional work.

  18. DNA breakage detection-fluorescence in situ hybridization (DBD-FISH) in buccal cells.

    Science.gov (United States)

    Cortés-Gutiérrez, E I; Dávila-Rodríguez, M I; Fernández, J L; López-Fernández, C; Gosálvez, J

    2012-12-28

    DNA breakage detection-fluorescence in situ hybridization (DBD-FISH) is a recently developed technique that allows cell-by-cell detection and quantification of DNA breakage in the whole genome or within specific DNA sequences. The present investigation was conducted to adapt the methodology of DBD-FISH to the visualization and evaluation of DNA damage in buccal epithelial cells. DBD-FISH revealed that DNA damage increased significantly according to H2O2 concentration (r2=0.91). In conclusion, the DBD-FISH technique is easy to apply in buccal cells and provides prompt results that are easy to interpret. Future studies are needed to investigate the potential applicability of a buccal cell DBD-FISH model to human biomonitoring and nutritional work.

  19. Interphase cytogenetics in pathology: principles, methods, and applications of fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Werner, M; Wilkens, L; Aubele, M; Nolte, M; Zitzelsberger, H; Komminoth, P

    1997-01-01

    Characteristic chromosome aberrations have been identified in various tumors. Fluorescence in situ hybridization (FISH) using specific probes that are generated by vector cloning or in vitro amplification and labeled with fluorescent dyes allow for the detection of these genetic changes in interphase cells. This technique, that is also referred to as "interphase cytogenetics", can be performed in cytological preparations as well as in sections of routinely formaldehyde-fixed and paraffin-embedded tissue. In cancer research and diagnostics, interphase cytogenetics by FISH is used to detect numerical chromosome changes and structural aberrations, e.g., translocations, deletions, or amplifications. In this technical overview, we explain the principles of the FISH method and provide protocols for FISH in cytological preparations and paraffin sections. Moreover, possible applications of FISH are discussed.

  20. Characterization of the Arachis (Leguminosae) D genome using fluorescence in situ hybridization (FISH) chromosome markers and total genome DNA hybridization

    OpenAIRE

    Germán Robledo; Guillermo Seijo

    2008-01-01

    Chromosome markers were developed for Arachis glandulifera using fluorescence in situ hybridization (FISH) of the 5S and 45S rRNA genes and heterochromatic 4'-6-diamidino-2-phenylindole (DAPI) positive bands. We used chromosome landmarks identified by these markers to construct the first Arachis species ideogram in which all the homologous chromosomes were precisely identified. The comparison of this ideogram with those published for other Arachis species revealed very poor homeologies with a...

  1. Metaphase FISH on a Chip: Miniaturized Microfluidic Device for Fluorescence in situ Hybridization

    Directory of Open Access Journals (Sweden)

    Niels Tommerup

    2010-11-01

    Full Text Available Fluorescence in situ Hybridization (FISH is a major cytogenetic technique for clinical genetic diagnosis of both inherited and acquired chromosomal abnormalities. Although FISH techniques have evolved and are often used together with other cytogenetic methods like CGH, PRINS and PNA-FISH, the process continues to be a manual, labour intensive, expensive and time consuming technique, often taking over 3–5 days, even in dedicated labs. We have developed a novel microFISH device to perform metaphase FISH on a chip which overcomes many shortcomings of the current laboratory protocols. This work also introduces a novel splashing device for preparing metaphase spreads on a microscope glass slide, followed by a rapid adhesive tape-based bonding protocol leading to rapid fabrication of the microFISH device. The microFISH device allows for an optimized metaphase FISH protocol on a chip with over a 20-fold reduction in the reagent volume. This is the first demonstration of metaphase FISH on a microfluidic device and offers a possibility of automation and significant cost reduction of many routine diagnostic tests of genetic anomalies.

  2. Inconsistent phylogeographic pattern between a sperm dependent fish and its host: in situ hybridization vs dispersal.

    Science.gov (United States)

    Vergilino, Roland; Leung, Christelle; Angers, Bernard

    2016-09-06

    Co-dispersal of sperm-dependent hybrids and their sexual relatives is expected to result in consistent spatial patterns between assemblages of hybrids and genetic structure of parental species. However, local hybridization events may blur this signal as assemblages could be organized under different connectivity constraints. This study aims at testing the hypothesis of local hybridization events by comparing the assemblage of hybrid fish Chrosomus eos-neogaeus to the genetic diversity of one of its parental species, Chrosomus eos. An extensive survey performed on a total of 132 sites located in two regions of Southern Quebec (West-Qc and East-Qc) revealed a distinct organization of hybrid lineages. One of the six hybrid lineages detected in West-Qc is widespread throughout this region resulting in a low α-diversity (1.38) and β-diversity (4.35). On the other hand, 36 hybrid lineages were detected in East-Qc and displayed narrow geographic distributions leading to a high α-diversity (2.30) and β-diversity (15.68). In addition, the C. eos multilocus haplotype of several of these hybrids is assigned to their respective sympatric C. eos population. Finally, contrasting with hybrids, the paternal species C. eos displayed a higher ρST in West-Qc (0.2300) than in East-Qc (0.0734). The unusually high diversity of hybrid lineages in East-Qc as well as the spatial organization and the close genetic relationship with C. eos sympatric populations support the hypothesis that multiple hybridization events occurred in situ. These findings coupled to the near absence of the maternal species Chrosomus neogeaus suggest that the decline of this species could be the trigger event at the origin of the high rates of spontaneous hybridization in this region.

  3. Behavioural isolation may facilitate homoploid hybrid speciation in cichlid fish

    NARCIS (Netherlands)

    Selz, O. M.; Thommen, R.; Maan, M. E.; Seehausen, O.

    Hybrid speciation is constrained by the homogenizing effects of gene flow from the parental species. In the absence of post-mating isolation due to structural changes in the genome, or temporal or spatial premating isolation, another form of reproductive isolation would be needed for homoploid

  4. Water quality in hybrid catfish ponds after partial fish harvest

    Science.gov (United States)

    Intensification of United States catfish aquaculture involves hybrid catfish ('channel catfish Ictalurus punctatus x ' blue catfish I. furcatus) grown in ponds with abundant aeration and high feeding rates. High feeding rates cause water quality deterioration because most of the nitrogen, phosphorus...

  5. Behavioural isolation may facilitate homoploid hybrid speciation in cichlid fish

    NARCIS (Netherlands)

    Selz, O. M.; Thommen, R.; Maan, M. E.; Seehausen, O.

    2014-01-01

    Hybrid speciation is constrained by the homogenizing effects of gene flow from the parental species. In the absence of post-mating isolation due to structural changes in the genome, or temporal or spatial premating isolation, another form of reproductive isolation would be needed for homoploid hybri

  6. Telomere analysis of platyhelminths and acanthocephalans by FISH and Southern hybridization.

    Science.gov (United States)

    Bombarová, Marta; Vítková, Magda; Spakulová, Marta; Koubková, Bozena

    2009-11-01

    We examined the composition of telomeres in chromosomes of parasitic worms, representatives of the flatworm groups Monogenea and Cestoda (Platyhelminthes), and thorny-headed worms (Syndermata: Acanthocephala) by fluorescence in situ hybridization (FISH) with different telomeric repeat probes. Our results show that the (TTAGGG)n sequence, supposed to be the ancestral telomeric repeat motif of Metazoa, is conserved in Monogenea (Paradiplozoon homoion) and Cestoda (Caryophyllaeus laticeps, Caryophyllaeides fennica, and Nippotaenia mogurndae) but not in Acanthocephala (Pomphorhynchus laevis and Pomphorhynchus tereticollis). In the Pomphorhynchus species, no hybridization signals were obtained with the "nematode" (TTAGGC)n, "arthropod" (TTAGG)n, and bdelloid (TGTGGG)n telomeric probes using FISH with their chromosomes and Southern hybridization with P. laevis DNA. Therefore, we suggest that parasitic Acanthocephala have evolved yet unknown telomeric repeat motifs or different mechanisms of telomere maintenance.

  7. Clinical Application of Fluorescence in Situ Hybridization (FISH) to Detect HER-2 Gene in Breast Cancer

    Institute of Scientific and Technical Information of China (English)

    Maurie Buehler; Ellie Guardino; Jung Sik Park; Eun Jeong Jang

    2014-01-01

    Objective: To investigate the clinical application of the detection of HER-2 gene by lfuorescence in situ hybridization (FISH) in breast cancer and the correlation between HER-2 gene ampliifcation and clinicopathology of breast cancer. Methods:Parafifn-embedded breast inifltrating ductal carcinoma from 48 patients were detected by FISH and immunohistochemistry (IHC) respectively for comparing the results of two methods. Results: HER-2 protein expressions were classified into three groups (3+/2+/1+ or 0) and the positive rates of HER-2 gene ampliifcation by FISH were 77.8%, 57.1% and 10.5%, respectively. Of the 29 cases with positive axillary lymph node, 12 were with HER-2 gene ampliifcation (P0.05). Conclusion:The false positive and negative rates are higher in HER-2 protein expression by IHC. Compared with IHC, FISH, being more effective and precise, can be applied extensively in clinic. HER-2 gene ampliifcation is concerned with axillary nodes metastases.

  8. Ocean warming, a rapid distributional shift, and the hybridization of a coastal fish species.

    Science.gov (United States)

    Potts, Warren M; Henriques, Romina; Santos, Carmen V; Munnik, Kate; Ansorge, Isabelle; Dufois, Francois; Booth, Anthony J; Kirchner, Carola; Sauer, Warwick H H; Shaw, Paul W

    2014-09-01

    Despite increasing awareness of large-scale climate-driven distribution shifts in the marine environment, no study has linked rapid ocean warming to a shift in distribution and consequent hybridization of a marine fish species. This study describes rapid warming (0.8 °C per decade) in the coastal waters of the Angola-Benguela Frontal Zone over the last three decades and a concomitant shift by a temperature sensitive coastal fish species (Argyrosomus coronus) southward from Angola into Namibia. In this context, rapid shifts in distribution across Economic Exclusive Zones will complicate the management of fishes, particularly when there is a lack of congruence in the fisheries policy between nations. Evidence for recent hybridization between A. coronus and a congener, A. inodorus, indicate that the rapid shift in distribution of A. coronus has placed adults of the two species in contact during their spawning events. Ocean warming may therefore revert established species isolation mechanisms and alter the evolutionary history of fishes. While the consequences of the hybridization on the production of the resource remain unclear, this will most likely introduce additional layers of complexity to their management.

  9. Prediction of melting temperatures in fluorescence in situ hybridization (FISH) procedures using thermodynamic models.

    Science.gov (United States)

    Fontenete, Sílvia; Guimarães, Nuno; Wengel, Jesper; Azevedo, Nuno Filipe

    2016-01-01

    The thermodynamics and kinetics of DNA hybridization, i.e. the process of self-assembly of one, two or more complementary nucleic acid strands, has been studied for many years. The appearance of the nearest-neighbor model led to several theoretical and experimental papers on DNA thermodynamics that provide reasonably accurate thermodynamic information on nucleic acid duplexes and allow estimation of the melting temperature. Because there are no thermodynamic models specifically developed to predict the hybridization temperature of a probe used in a fluorescence in situ hybridization (FISH) procedure, the melting temperature is used as a reference, together with corrections for certain compounds that are used during FISH. However, the quantitative relation between melting and experimental FISH temperatures is poorly described. In this review, various models used to predict the melting temperature for rRNA targets, for DNA oligonucleotides and for nucleic acid mimics (chemically modified oligonucleotides), will be addressed in detail, together with a critical assessment of how this information should be used in FISH.

  10. itFISH: Enhanced Staining by Iterative Fluorescent In Situ Hybridization.

    Science.gov (United States)

    Row, Richard H; Martin, Benjamin L

    2017-03-20

    Fluorescent in situ hybridization (FISH) is an important tool for zebrafish research, particularly when observing the expression of two different genes in the same embryo. Peroxidase-catalyzed deposition of tyramide-conjugated dyes is a widely used and cost-effective approach to performing FISH. A major limitation of the technique is that it does not work well for weakly expressed genes. Here we present a method adapted from planarian research for use in zebrafish that provides a dramatic enhancement of weak staining. By iterating the antibody staining and development steps, a strong signal can be obtained from probes that were previously too weak to detect.

  11. Characterization of the Arachis (Leguminosae D genome using fluorescence in situ hybridization (FISH chromosome markers and total genome DNA hybridization

    Directory of Open Access Journals (Sweden)

    Germán Robledo

    2008-01-01

    Full Text Available Chromosome markers were developed for Arachis glandulifera using fluorescence in situ hybridization (FISH of the 5S and 45S rRNA genes and heterochromatic 4'-6-diamidino-2-phenylindole (DAPI positive bands. We used chromosome landmarks identified by these markers to construct the first Arachis species ideogram in which all the homologous chromosomes were precisely identified. The comparison of this ideogram with those published for other Arachis species revealed very poor homeologies with all A and B genome taxa, supporting the special genome constitution (D genome of A. glandulifera. Genomic affinities were further investigated by dot blot hybridization of biotinylated A. glandulifera total DNA to DNA from several Arachis species, the results indicating that the D genome is positioned between the A and B genomes.

  12. Analysis of Single-cell Gene Transcription by RNA Fluorescent In Situ Hybridization (FISH)

    DEFF Research Database (Denmark)

    Ronander, Elena; Bengtsson, Dominique C; Joergensen, Louise;

    2012-01-01

    and the consequence of differential binding on the clinical outcome of P. falciparum infections. Recently, the mutually exclusive transcription paradigm has been called into doubt by transcription assays based on individual P. falciparum transcript identification in single infected erythrocytic cells using RNA...... fluorescent in situ hybridization (FISH) analysis of var gene transcription by the parasite in individual nuclei of P. falciparum IE(1). Here, we present a detailed protocol for carrying out the RNA-FISH methodology for analysis of var gene transcription in single-nuclei of P. falciparum infected human...... erythrocytes. The method is based on the use of digoxigenin- and biotin- labeled antisense RNA probes using the TSA Plus Fluorescence Palette System(2) (Perkin Elmer), microscopic analyses and freshly selected P. falciparum IE. The in situ hybridization method can be used to monitor transcription...

  13. Design of Hybrid Solar and Wind Energy Harvester for Fishing Boat

    Science.gov (United States)

    Banjarnahor, D. A.; Hanifan, M.; Budi, E. M.

    2017-07-01

    In southern beach of West Java, Indonesia, there are many villagers live as fishermen. They use small boats for fishing, in one to three days. Therefore, they need a fish preservation system. Fortunately, the area has high potential of solar and wind energy. This paper presents the design of a hybrid solar and wind energy harvester to power a refrigerator in the fishing boat. The refrigerator should keep the fish in 2 - 4 °C. The energy needed is 720 Wh daily. In the area, the daily average wind velocity is 4.27 m/s and the sun irradiation is 672 W/m2. The design combined two 100W solar panels and a 300W wind turbine. The testing showed that the solar panels can harvest 815 - 817 Wh of energy, while the wind turbine can harvest 43 - 62 Wh of energy daily. Therefore, the system can fulfil the energy requirement in fishing boat, although the solar panels were more dominant. To install the wind turbine on the fishing-boat, a computational design had been conducted. The boat hydrostatic dimension was measured to determine its stability condition. To reach a stable equilibrium condition, the wind turbine should be installed no more than 1.7 m of height.

  14. Detection of VHSV IVb within the gonads of Great Lakes fish using in situ hybridization.

    Science.gov (United States)

    Al-Hussinee, L; Lumsden, J S

    2011-05-24

    Viral haemorrhagic septicaemia virus (VHSV) genotype IVb was recently detected as the cause of numerous mortality events in Great Lakes fish. In situ hybridization was used to examine the gonads from 13 fish, including freshwater drum Aplodinotus grunniens and muskellunge Esox masquinongy that were infected naturally, as well as rainbow trout Oncorhynchus mykiss and fathead minnows Pimphales promelas, which were experimentally infected. Although the ovaries and testes of fish infected by VHSV IVb had few lesions, viral RNA was present in the ovaries of the rainbow trout and fathead minnow and was abundant in the gonads of muskellunge and in the ovaries of freshwater drum. Viral RNA was present mainly surrounding yolk vacuoles/granules or adjacent to the germinal vesicle, with lesser amounts found within the germinal vesicle, in the mesovarium and/or tunica albuginea and blood vessels of the ovary. Viral RNA was also found in and surrounding primary and secondary spermatocytes of the muskellunge.

  15. Detection of integrated herpesvirus genomes by fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Kaufer, Benedikt B

    2013-01-01

    Fluorescence in situ hybridization (FISH) is widely used to visualize nucleotide sequences in interphase cells or on metaphase chromosomes using specific probes that are complementary to the respective targets. Besides its broad application in cytogenetics and cancer research, FISH facilitates the localization of virus genomes in infected cells. Some herpesviruses, including human herpesvirus 6 (HHV-6) and Marek's disease virus (MDV), have been shown to integrate their genetic material into host chromosomes, which allows transmission of HHV-6 via the germ line and is required for efficient MDV-induced tumor formation. We describe here the detection by FISH of integrated herpesvirus genomes in metaphase chromosomes and interphase nuclei of herpesvirus-infected cells.

  16. Risk stratification of plasma cell neoplasm: insights from plasma cell-specific cytoplasmic immunoglobulin fluorescence in situ hybridization (cIg FISH) vs. conventional FISH.

    Science.gov (United States)

    Dong, Henry; Yang, Hai-Su; Jagannath, Sundar; Stephenson, Christine F; Brenholz, Pauline; Mazumder, Amitabha; Chari, Ajai

    2012-10-01

    We directly compared the results of routine fluorescence in situ hybridization (FISH) and plasma cell-specific cytoplasmic immunoglobulin (cIg) FISH from 75 paired samples for myeloma risk stratification. CIg FISH improves test specificity and sensitivity and tends to eliminate borderline results. It proves that most plasma cells (PCs) consistently carry the abnormality in myelomas with an IGH translocation, whereas routine FISH detects these cells only at variably low levels. Routine cytogenetic analysis of plasma cell neoplasms (PCNs) has a low sensitivity. Conventional fluorescence in situ hybridization (FISH) is not plasma cell (PC) specific and results are diluted by other cells in the sample. Although PC-specific FISH testing has been recommended for multiple myeloma (MM) risk stratification, eg, by combining cytoplasmic immunoglobulin (cIg) staining with FISH, the benefits of cIg FISH have never been directly demonstrated in a controlled study. Seventy-five samples from patients with PCNs were analyzed by concomitant conventional FISH and cIg FISH with probes for t(4;14), t(11;14), t(14;16), -13, 17p-, and +3. The results were compared for their reliability, specificity, and consistency. Apart from marginally improving detection threshold in samples with low PC burden, cIg FISH identified more abnormal cases (50 vs. 47 cases) and more chromosome abnormalities (113 vs. 103 events) than did conventional FISH. It differentiated del(13q) in myelodysplasia from MM. Remarkably, cIg FISH consistently identified a high percentage of abnormal PCs in all cases. It detected IGH translocation in 78% to 100% of PCs in all but 2 positive cases, whereas conventional FISH detected 0% to 46% in these cases (median, 91% vs. 9%). The abnormal cells found in patients with 17p- were 19% to 96% by cIg FISH vs. 0% to 13% by conventional FISH (median, 54% vs. 9%). Cases with insufficient PCs for cIg FISH had only normal conventional FISH results. CIg FISH improves reliability of

  17. A simple protocol for attenuating the auto-fluorescence of cyanobacteria for optimized fluorescence in situ hybridization (FISH) imaging.

    Science.gov (United States)

    Zeller, Perrine; Ploux, Olivier; Méjean, Annick

    2016-03-01

    Cyanobacteria contain pigments, which generate auto-fluorescence that interferes with fluorescence in situ hybridization (FISH) imaging of cyanobacteria. We describe simple chemical treatments using CuSO4 or H2O2 that significantly reduce the auto-fluorescence of Microcystis strains. These protocols were successfully applied in FISH experiments using 16S rRNA specific probes and filamentous cyanobacteria.

  18. The complete mitochondrial genome of Channa argus, Channa maculata and hybrid snakehead fish [Channa maculata (♀) × Channa argus (♂)].

    Science.gov (United States)

    Zhu, Shu-Ren; Ma, Ke-Yi; Xing, Zhi-Jun; Xie, Nan; Wang, Yu-Xi; Wang, Qun; Li, Jia-Le

    2013-06-01

    We sequenced and characterized the complete mitochondrial genome of Channa argus, Channa maculata and their hybrid [C. maculata (♀) and C. argus (♂)]. All the three mitochondrial genomes contained the typical complement of 13 protein-coding genes, 22 transfer RNAs (tRNAs), 2 ribosomal RNAs (rRNAs) and 1 control region. The entire mitochondrial DNA (mtDNA) molecule of C. maculata was 16,559 bp long while the complete mtDNA molecule of C. argus and hybrid snakehead fish was 16,558 bp long. This is the first report on the complete mitogenome sequence of C. maculata and hybrid snakehead fish.

  19. Detection of a complex translocation using fluorescent in situ hybridization (FISH)

    Energy Technology Data Exchange (ETDEWEB)

    Rosen, B.A. [Brandeis Univ., Waltham, MA (United States); Abuelo, D.N. [Rhode Island Hospital, Providence, RI (United States); Mark, H.F. [Brown Univ. School of Medicine, Providence, RI (United States)

    1994-09-01

    The use of fluorescent in situ hybridization (FISH) allowed the detection of a complex 3-way translocation in a patient with multiple congenital malformations and mental retardation. The patient was a 10-year-old girl with mental retardation, seizures, repaired cleft palate, esotropia, epicanthal folds, broad nasal bridge, upward slanting palpebral fissures, single transverse palmar crease, brachydactyly, hypoplastic nails, ectrodactyly between the third and fourth right toes, and hypoplasia of the left third toe. Chromosome analysis performed at birth was reported as normal. We performed high resolution banding analysis which revealed an apparently balanced translocation between chromosomes 2 and 9. However, because of her multiple abnormalities, further studies were ordered. Fluorescent in situ hybridization (FISH) using chromosome painting probes revealed a karyotype of 46,XX,t(2;8;9) (2pter{yields}q31::8q21.2{yields}8qter; 8pter{yields}q21.2::2q31{yields}q34::9q34{yields}qter; 9pter{yields}q34::2q34{yields}qter). The 3-way translocation appears to be de novo, as neither parent is a translocation carrier. This case illustrates the importance of using FISH to further investigate cases of apparently balanced translocations in the presence of phenotypic abnormalities and/or mental retardation.

  20. Detection of airborne Legionella while showering using liquid impingement and fluorescent in situ hybridization (FISH).

    Science.gov (United States)

    Deloge-Abarkan, Magali; Ha, Thi-Lan; Robine, Enric; Zmirou-Navier, Denis; Mathieu, Laurence

    2007-01-01

    Aerosols of water contaminated with Legionella bacteria constitute the only mode of exposure for humans. However, the prevention strategy against this pathogenic bacteria risk is managed through the survey of water contamination. No relationship linked the Legionella bacteria water concentration and their airborne abundance. Therefore, new approaches in the field of the metrological aspects of Legionella bioaerosols are required. This study was aimed at testing the main principles for bioaerosol collection (solid impaction, liquid impingement and filtration) and the in situ hybridization (FISH) method, both in laboratory and field assays, with the intention of applying such methodologies for airborne Legionella bacteria detection while showering. An aerosolization chamber was developed to generate controlled and reproducible L. pneumophila aerosols. This tool allowed the identification of the liquid impingement method as the most appropriate one for collecting airborne Legionella bacteria. The culturable fraction of airborne L. pneumophila recovered with the liquid impingement principle was 4 and 700 times higher compared to the impaction and filtration techniques, respectively. Moreover, the concentrations of airborne L. pneumophila in the impinger fluid were on average 7.0 x 10(5) FISH-cells m(-3) air with the fluorescent in situ hybridization (FISH) method versus 9.0 x 10(4) CFU m(-3) air with the culture method. These results, recorded under well-controlled conditions, were confirmed during the field experiments performed on aerosols generated by hot water showers in health institutions. This new approach may provide a more accurate characterization of aerobiocontamination by Legionella bacteria.

  1. Detection of Helicobacter pylori in raw bovine milk by fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Angelidis, Apostolos S; Tirodimos, Ilias; Bobos, Mattheos; Kalamaki, Mary S; Papageorgiou, Demetrios K; Arvanitidou, Malamatenia

    2011-12-02

    The transmission pathways of Helicobacter pylori in humans have not been fully elucidated. Research in the last decade has proposed that foodborne transmission, among others, may be a plausible route of human infection. Owing to the organism's fastidious growth characteristics and its ability to convert to viable, yet unculturable states upon exposure to stress conditions, the detection of H. pylori in foods via culture-dependent methods has been proven to be laborious, difficult and in most cases unsuccessful. Hence, nucleic acid-based methods have been proposed as alternative methods but, to date, only PCR-based methods have been reported in the literature. In the current study, fluorescence in situ hybridization (FISH) was used for the detection of H. pylori in raw, bulk-tank bovine milk. After repeated milk centrifugation and washing steps, the bacterial flora of raw milk was subjected to fixation and permeabilization and H. pylori detection was conducted by FISH after hybridization with an H. pylori-specific 16S rRNA-directed fluorescent oligonucleotide probe. Using this protocol, H. pylori was detected in four out of the twenty (20%) raw milk samples examined. The data presented in this manuscript indicate that FISH can serve as an alternative molecular method for screening raw bovine milk for the presence of H. pylori.

  2. Segregation of Species-Specific Male Attractiveness in F2 Hybrid Lake Malawi Cichlid Fish

    Directory of Open Access Journals (Sweden)

    Ola Svensson

    2011-01-01

    Full Text Available Among the huge radiations of haplochromine cichlid fish in Lakes Malawi and Victoria, closely related species are often reproductively isolated via female mate choice although viable fertile hybrids can be produced when females are confined only with heterospecific males. We generated F2 hybrid males from a cross between a pair of closely related sympatric cichlid fish from Lake Malawi. Laboratory mate choice experiments using microsatellite paternity analysis demonstrated that F2 hybrid males differed significantly in their attractiveness to females of the two parental species, indicating heritable variation in traits involved in mate choice that may contribute to reproductive isolation between these species. We found no significant correlation between male mating success and any measurement of male colour pattern. A simple quantitative genetic model of reproductive isolation suggests that there may be as few as two chromosomal regions controlling species-specific attractiveness. We propose that adaptive radiation of Lake Malawi cichlids could be facilitated by the presence of genes with major effects on mate choice and reproductive isolation.

  3. Water quality and plankton communities in hybrid catfish (female channel catfish, Ictalurus punctatus x male blue catfish, I. furcatus) ponds after partial fish harvest

    Science.gov (United States)

    Twelve, 0.4-ha ponds were stocked with 10,000 hybrid catfish fingerlings in March 2015. Six ponds were partially harvested in August to remove fish larger than ~ 0.57 kg. All remaining fish were removed in October and November. Partial harvest of faster-growing fish removed ~26% of fish initially st...

  4. Discrimination of bacteriophage infected cells using locked nucleic acid fluorescent in situ hybridization (LNA-FISH).

    Science.gov (United States)

    Vilas Boas, Diana; Almeida, Carina; Sillankorva, Sanna; Nicolau, Ana; Azeredo, Joana; Azevedo, Nuno F

    2016-01-01

    Bacteriophage-host interaction studies in biofilm structures are still challenging due to the technical limitations of traditional methods. The aim of this study was to provide a direct fluorescence in situ hybridization (FISH) method based on locked nucleic acid (LNA) probes, which targets the phage replication phase, allowing the study of population dynamics during infection. Bacteriophages specific for two biofilm-forming bacteria, Pseudomonas aeruginosa and Acinetobacter, were selected. Four LNA probes were designed and optimized for phage-specific detection and for bacterial counterstaining. To validate the method, LNA-FISH counts were compared with the traditional plaque forming unit (PFU) technique. To visualize the progression of phage infection within a biofilm, colony-biofilms were formed and infected with bacteriophages. A good correlation (r = 0.707) was observed between LNA-FISH and PFU techniques. In biofilm structures, LNA-FISH provided a good discrimination of the infected cells and also allowed the assessment of the spatial distribution of infected and non-infected populations.

  5. HER2 in situ hybridization in gastric and gastroesophageal adenocarcinoma: comparison of automated dual ISH to FISH.

    Science.gov (United States)

    Grin, Andrea; Brezden-Masley, Christine; Bauer, Sharon; Streutker, Catherine J

    2013-12-01

    Patients with gastric and gastroesophageal junction (GEJ) adenocarcinomas that are HER2 positive by immunohistochemistry (IHC) or in situ hybridization show a significant survival benefit with trastuzumab therapy. In situ hybridization is traditionally done by fluorescence in situ hybridization (FISH), despite some limitations. An alternative is the dual in situ hybridization (Dual ISH) technique that is fully automated and uses differentially labeled CEP17 and HER2 probes that can be read by light microscopy on 1 slide. The aim of this study was to assess the utility of Dual ISH in gastric/GEJ cancer and to compare the results with those obtained by IHC and FISH. Cases of gastric/GEJ adenocarcinoma were analyzed by IHC, FISH, and Dual ISH and the correlation between methods calculated. Results for 50 patients were available. There was a 98% (49/50) concordance rate between Dual ISH and FISH. One discrepant case was nonamplified by FISH but showed focal amplification by Dual ISH. Discrepancy was attributed to tumor heterogeneity, which was a frequent finding (78% of HER2-positive cases). There was excellent correlation between Dual ISH and FISH for assessment of HER2 amplification. Dual ISH was rapid, easy to interpret, and maintained cell morphology, which was valuable in identifying tumor heterogeneity.

  6. Insertional translocation detected using FISH confirmation of array-comparative genomic hybridization (aCGH) results.

    Science.gov (United States)

    Kang, Sung-Hae L; Shaw, Chad; Ou, Zhishuo; Eng, Patricia A; Cooper, M Lance; Pursley, Amber N; Sahoo, Trilochan; Bacino, Carlos A; Chinault, A Craig; Stankiewicz, Pawel; Patel, Ankita; Lupski, James R; Cheung, Sau Wai

    2010-05-01

    Insertional translocations (ITs) are rare events that require at least three breaks in the chromosomes involved and thus qualify as complex chromosomal rearrangements (CCR). In the current study, we identified 40 ITs from approximately 18,000 clinical cases (1:500) using array-comparative genomic hybridization (aCGH) in conjunction with fluorescence in situ hybridization (FISH) confirmation of the aCGH findings, and parental follow-up studies. Both submicroscopic and microscopically visible IT events were detected. They were divided into three major categories: (1) simple intrachromosomal and interchromosomal IT resulting in pure segmental trisomy, (2) complex IT involving more than one abnormality, (3) deletion inherited from a parent with a balanced IT resulting in pure segmental monosomy. Of the cases in which follow-up parental studies were available, over half showed inheritance from an apparently unaffected parent carrying the same unbalanced rearrangement detected in the propositi, thus decreasing the likelihood that these IT events are clinically relevant. Nevertheless, we identified six cases in which small submicroscopic events were detected involving known disease-associated genes/genomic segments and are likely to be pathogenic. We recommend that copy number gains detected by clinical aCGH analysis should be confirmed using FISH analysis whenever possible in order to determine the physical location of the duplicated segment. We hypothesize that the increased use of aCGH in the clinic will demonstrate that IT occurs more frequently than previously considered but can identify genomic rearrangements with unclear clinical significance.

  7. Stability of collagen during denaturation.

    Science.gov (United States)

    Penkova, R; Goshev, I; Gorinstein, S; Nedkov, P

    1999-05-01

    The stability of calf skin collagen (CSC) type I during thermal and chemical denaturation in the presence of glycerol was investigated. Thermal denaturation of type I collagen was performed in the presence of glycerol or in combination with urea and sodium chloride. The denaturation curves obtained in the presence of urea or sodium chloride retained their original shape without glycerol. These curves were shifted upward proportionally to the glycerol concentration in the reaction medium. This means that glycerol and the denaturants act independently. The explanation is based on the difference in the mechanism of their action on the collagen molecule.

  8. Optimized Fast-FISH with a-satellite probes: acceleration by microwave activation

    Directory of Open Access Journals (Sweden)

    Durm M.

    1997-01-01

    Full Text Available It has been shown for several DNA probes that the recently introduced Fast-FISH (fluorescence in situ hybridization technique is well suited for quantitative microscopy. For highly repetitive DNA probes the hybridization (renaturation time and the number of subsequent washing steps were reduced considerably by omitting denaturing chemical agents (e.g., formamide. The appropriate hybridization temperature and time allow a clear discrimination between major and minor binding sites by quantitative fluorescence microscopy. The well-defined physical conditions for hybridization permit automatization of the procedure, e.g., by a programmable thermal cycler. Here, we present optimized conditions for a commercially available X-specific a-satellite probe. Highly fluorescent major binding sites were obtained for 74oC hybridization temperature and 60 min hybridization time. They were clearly discriminated from some low fluorescent minor binding sites on metaphase chromosomes as well as in interphase cell nuclei. On average, a total of 3.43 ± 1.59 binding sites were measured in metaphase spreads, and 2.69 ± 1.00 in interphase nuclei. Microwave activation for denaturation and hybridization was tested to accelerate the procedure. The slides with the target material and the hybridization buffer were placed in a standard microwave oven. After denaturation for 20 s at 900 W, hybridization was performed for 4 min at 90 W. The suitability of a microwave oven for Fast-FISH was confirmed by the application to a chromosome 1-specific a-satellite probe. In this case, denaturation was performed at 630 W for 60 s and hybridization at 90 W for 5 min. In all cases, the results were analyzed quantitatively and compared to the results obtained by Fast-FISH. The major binding sites were clearly discriminated by their brightness

  9. Fluorescent in situ hybridization (FISH) assessment of chromosome copy number in sperm

    Energy Technology Data Exchange (ETDEWEB)

    Sheu, M.; Sigman, M.; Mark, H.F.L. [Brown Univ. School of Medicine, Providence, RI (United States)

    1994-09-01

    Approximately 15% of all recognized pregnancies end in spontaneous abortions. The overall frequency of chromosome abnormalities in spontaneous abortions is approximately 50%. Thus aneuploidy is a significant cause of fetal wastage. In addition, structural and numerical abnormalities of chromosomes can also lead to birth defects, developmental delay, mental retardation and infertility. Conventional cytogenetic analysis via GTG- and other banding techniques is a powerful tool in the elucidation of the nature of chromosomal abnormalities. Fluorescent in situ hybridization (FISH) enables detection of numerical chromosomal abnormalities, especially trisomies, in intact cells. Using FISH and commercially available biotin-labeled probes, we have initiated a prospective study to assess specific chromosome copy number of preparations of unstained smears from men referred for a male infertility evaluation as well as smears from normal control males chosen randomly from the sample of sperm donors. A total of approximately 19,000 sperm nuclei have been examined thus far. Of those suitable for analysis, 7382 (38.75%) were normal possessing one copy of chromosome 8, 155 (0.81%) were disomic, and 15 (0.079%) had more than two copies of chromosome 8. Comparisons with data available in the literature will be discussed. Work is ongoing to increase the efficiency of hybridization using both reported and previously untried pretreatment and fixation protocols. We have also initiated studies using multicolor FISH with various chromosome enumeration probes. The assay described here is a potentially powerful tool for detecting rare events such as spontaneous germ cell aneuploidy, aneuploidy detected in semen from men with carcinoma in situ of the testis and aneuploidy induced by potential environmental genotoxicants. It can also be utilized for segregation analysis and for correlating chromosome copy number with germ cell morphology.

  10. Fluorescence in situ hybridization (FISH analysis of primary ocular adnexal MALT lymphoma

    Directory of Open Access Journals (Sweden)

    Harada Mine

    2006-10-01

    Full Text Available Abstract Background It remains unknown whether primary ocular adnexal extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma is a homogeneous entity, as there are few reports of the results of cytogenetic or molecular analyses of these tumors. Methods We performed interphase fluorescence in situ hybridization (FISH analysis to detect translocations and aneuploidy in 34 cases of primary ocular adnexal MALT lymphoma, and reviewed the histopathological findings. Correlations between the results of FISH analysis, the histopathological features and the clinical data were also analyzed. Results Among the 34 cases, FISH analysis revealed t(14;18(q32;q21 in one case, trisomy 3 in 21 cases (62%, and trisomy 18 in 16 cases (47%. The cases with trisomy 18 had significantly more prominent lymphoepithelial lesions (LELs and less nodularity in the tumors. In regard to the clinical correlations, tumors with trisomy 18 were observed predominantly in females and younger patients; also, in the majority of the cases, the tumor was of conjunctival origin. All the cases with recurrence showed trisomy 18 in the tumor. Conclusion Primary ocular adnexal MALT lymphoma is a significantly heterogeneous entity. Cases with trisomy 18 may have unique clinicopathological features.

  11. Trade-off between morphological convergence and opportunistic diet behavior in fish hybrid zone

    Directory of Open Access Journals (Sweden)

    Grey Jonathan

    2009-10-01

    Full Text Available Abstract Background The invasive Chondrostoma nasus nasus has colonized part of the distribution area of the protected endemic species Chondrostoma toxostoma toxostoma. This hybrid zone is a complex system where multiple effects such as inter-species competition, bi-directional introgression, strong environmental pressure and so on are combined. Why do sympatric Chondrostoma fish present a unidirectional change in body shape? Is this the result of inter-species interactions and/or a response to environmental effects or the result of trade-offs? Studies focusing on the understanding of a trade-off between multiple parameters are still rare. Although this has previously been done for Cichlid species flock and for Darwin finches, where mouth or beak morphology were coupled to diet and genetic identification, no similar studies have been done for a fish hybrid zone in a river. We tested the correlation between morphology (body and mouth morphology, diet (stable carbon and nitrogen isotopes and genomic combinations in different allopatric and sympatric populations for a global data set of 1330 specimens. To separate the species interaction effect from the environmental effect in sympatry, we distinguished two data sets: the first one was obtained from a highly regulated part of the river and the second was obtained from specimens coming from the less regulated part. Results The distribution of the hybrid combinations was different in the two part of the sympatric zone, whereas all the specimens presented similar overall changes in body shape and in mouth morphology. Sympatric specimens were also characterized by a larger diet behavior variance than reference populations, characteristic of an opportunistic diet. No correlation was established between the body shape (or mouth deformation and the stable isotope signature. Conclusion The Durance River is an untamed Mediterranean river despite the presence of numerous dams that split the river from

  12. Trade-off between morphological convergence and opportunistic diet behavior in fish hybrid zone.

    Science.gov (United States)

    Corse, Emmanuel; Costedoat, Caroline; Pech, Nicolas; Chappaz, Rémi; Grey, Jonathan; Gilles, André

    2009-10-27

    The invasive Chondrostoma nasus nasus has colonized part of the distribution area of the protected endemic species Chondrostoma toxostoma toxostoma. This hybrid zone is a complex system where multiple effects such as inter-species competition, bi-directional introgression, strong environmental pressure and so on are combined. Why do sympatric Chondrostoma fish present a unidirectional change in body shape? Is this the result of inter-species interactions and/or a response to environmental effects or the result of trade-offs? Studies focusing on the understanding of a trade-off between multiple parameters are still rare. Although this has previously been done for Cichlid species flock and for Darwin finches, where mouth or beak morphology were coupled to diet and genetic identification, no similar studies have been done for a fish hybrid zone in a river. We tested the correlation between morphology (body and mouth morphology), diet (stable carbon and nitrogen isotopes) and genomic combinations in different allopatric and sympatric populations for a global data set of 1330 specimens. To separate the species interaction effect from the environmental effect in sympatry, we distinguished two data sets: the first one was obtained from a highly regulated part of the river and the second was obtained from specimens coming from the less regulated part. The distribution of the hybrid combinations was different in the two part of the sympatric zone, whereas all the specimens presented similar overall changes in body shape and in mouth morphology. Sympatric specimens were also characterized by a larger diet behavior variance than reference populations, characteristic of an opportunistic diet. No correlation was established between the body shape (or mouth deformation) and the stable isotope signature. The Durance River is an untamed Mediterranean river despite the presence of numerous dams that split the river from upstream to downstream. The sympatric effect on morphology and

  13. Salmonellae in fish feces analyzed by in situ hybridization and quantitative polymerase chain reaction.

    Science.gov (United States)

    Sha, Qiong; Forstner, Michael R J; Bonner, Timothy H; Hahn, Dittmar

    2013-09-01

    The potential of fish to transfer salmonellae from heterogeneous aquatic biofilms into feces was assessed in controlled aquarium studies with Suckermouth Catfish Hypostomus plecostomus and with biofilms inoculated with salmonellae. Neither the presence of catfish nor inoculation with salmonellae had detectable effects on the abundance of the microbial community. Densities of the microbial community were about 10(5) cells/mL in the water during a 1-week period, whereas densities of the microbial community increased 10-fold (10(6) to 10(7) cells/mg) in catfish feces during the same period. Salmonellae were detected by both quantitative polymerase chain reaction (qPCR) and situ hybridization in water samples immediately after inoculation, in numbers of about 10(4) cells/mL, representing up to 20% of the cells of the microbial community. Numbers decreased by three orders of magnitude within the first 3 d of the study, which represented only 0.01% of the community, and became undetectable after day 5. In catfish feces, numbers of Salmonella initially increased to up to 6% of the cells of the community but then declined. These results suggest that Salmonella are not biomagnified during gut passage, and thus, fish only provide a means for the translocation of this pathogen.

  14. CONGESTION MANAGEMENT BY OPTIMAL ALLOCATION OF FACTS CONTROLLERS USING HYBRID FISH BEE OPTIMIZATION

    Directory of Open Access Journals (Sweden)

    S. Thangalakshmi

    2014-01-01

    Full Text Available The role of Independent System Operator (ISO in the restructured power industry includes system control, capacity planning, transmission tariff and congestion management; the challenging task being minimizing the congestion. One of the popular techniques used to alleviate congestion is using Flexible AC Transmission Systems (FACTS devices. The power system generally operates near its rated capacity in deregulated market because of intensive usage of transmission grids. So, the major issues that need to be addressed are improving the voltage profile and reducing the power loss in the electrical network. Motivation: The location of FACTS devices can improve the power flow in the line, maintain the bus profile and reduce the losses. However locating the ideal location is a NP problem. This study presents a novel heuristic method to determine the types of FACTS devices and its optimal location in a power system without violating the thermal and voltage limits. Power flow sensitivity index to find the optimal location of UPFC is suggested in this study. A hybrid fish bee swarm optimization is proposed which is based on Artificial Bee Colony (ABC and Fish School Search (FSS methods. This proposed algorithm is tested based on IEEE 30 bus system and line performances are studied.

  15. Trypanosomiasis in hybrid catfish (Clarias macrocephalus x Clarias gariepinus and other freshwater fishes

    Directory of Open Access Journals (Sweden)

    Promkhunthong, W.

    2005-02-01

    Full Text Available The infestation of Trypanosoma sp. (Kinetoplastida in the hybrid catfish and some fresh water fish was studied showing such parasitic infestation was specific only in catfish. The virulence as determined by LD50 for 5 days was 2.28x1010 cell in a fish sample. The parasitic infestation caused hematological changes by the reduction of red as well as white blood cells. The reductions were highly significant as compared to the healthy sample (p<0.05 as noted by the red and white blood cell count which dropped from 2.14±0.48x106 to 1.62±0.27x106 cells/ml and from 1.45±3.76x105 to 2.42±0.78x104 cells/ml blood in the infested samples, respectively. Similar trend was noted for hemoglobin and hematocrit which dropped significantly (p<0.05. The hemoglobin in healthy fish is 7.075±0.929g/100g, which dropped to 6.268±0.697g/100g in the samples with infestation. The percentage of hematocrit in healthy sample is 25.275±3.318%, which dropped to 21.722±3.068% in the samples with infestation. The reverse trend was recognized for serum protein andleukocrit which increased in the samples with Trypanosoma sp. infestation. The density gradient centrifugation technique was employed in the isolation of parasites in which 50% Percoll solution in 0.85% final preparation of saline solution was capable of removing Trypanosoma sp. from the blood. The study of antibody levels in serum showed that the infested hybrid catfish could develop the antibody which reached a peak 14 days after the infestation. Trypanosoma sp. was unable to cause histological changes in the tissues of gill, liver, kidney, spleen, stomach and intestine. Minor inflammations were observed, even the cases that large number of parasites were found in the tissues, blood streams and sinuses. Marked reductions were recorded for mature red blood cells while there were the formation of immature red blood and phagocytotic cells at higher rates as compared to the healthy individual.

  16. Karyotype of Zea luxurians and Z. mays subsp. mays using FISH/DAPI, and analysis of meiotic behavior of hybrids.

    Science.gov (United States)

    González, Graciela E; Poggio, Lidia

    2011-01-01

    The karyotypes of Zea luxurians and a race of maize from northwestern Argentina are described and compared using 4′,6-diamidino-2-phenylindole (DAPI) banding and fluorescent in situ hybridization (FISH) to localize the 180 bp knobs. The meiotic behavior of the F₁ artificial hybrids Z. luxurians × maize is also analyzed to determine the genomic relationships between both species. Neocentromere activity at knobs in the meiosis of the hybrids is particularly discussed. The meiotic behavior and the high pollen sterility of the hybrid revealed genetical and (or) chromosomal divergences, leading to postzygotic reproductive isolation among their parents. Here, we propose that maize shows lower genomic affinity to Z. luxurians than to other species of the genus with 2n = 20.

  17. Multiplex fluorescence in situ hybridization (M-FISH) and confocal laser scanning microscopy (CLSM) to analyze multispecies oral biofilms.

    Science.gov (United States)

    Karygianni, Lamprini; Hellwig, Elmar; Al-Ahmad, Ali

    2014-01-01

    Multiplex fluorescence in situ hybridization (M-FISH) constitutes a favorable microbiological method for the analysis of spatial distribution of highly variable phenotypes found in multispecies oral biofilms. The combined use of confocal laser scanning microscopy (CLSM) produces high-resolution three-dimensional (3D) images of individual bacteria in their natural environment. Here, we describe the application of M-FISH on early (Streptococcus spp., Actinomyces naeslundii) and late colonizers (Fusobacterium nucleatum, Veillonella spp.) of in situ-formed oral biofilms, the acquisition of CLSM images, as well as the qualitative and quantitative analysis of these digitally obtained and processed images.

  18. Studies of the Ecophysiology of Single Cells in Microbial Communities by (Quantitative) Microautoradiography and Fluorescence In Situ Hybridization (MAR-FISH)

    DEFF Research Database (Denmark)

    Nierychlo, Marta; Nielsen, Jeppe Lund; Nielsen, Per Halkjær

    2015-01-01

    Microautoradiography (MAR) in combination with fluorescence in situ hybridization (FISH) is a powerful method of obtaining information about the ecophysiology of probe-defined single cells in mixed microbial communities. The incorporation of radiolabelled substrates can be quantified by automated...

  19. Development of a fluorescent in situ hybridization (FISH) technique for visualizing CGMMV in plant tissues.

    Science.gov (United States)

    Shargil, D; Zemach, H; Belausov, E; Lachman, O; Kamenetsky, R; Dombrovsky, A

    2015-10-01

    Cucumber green mottle mosaic virus (CGMMV), which belongs to the genus Tobamovirus, is a major pathogen of cucurbit crops grown indoors and in open fields. Currently, immunology (e.g., ELISA) and molecular amplification techniques (e.g., RT-PCR) are employed extensively for virus detection in plant tissues and commercial seed lots diagnostics. In this study, a fluorescent in situ hybridization (FISH) technique, using oligonucleotides whose 5'-terminals were labeled with red cyanine 3 (Cy3) or green fluorescein isothiocyanate (FITC), was developed for the visualization of the pathogen in situ. This simple and reliable method allows detection and localization of CGMMV in the vegetative and reproductive tissues of cucumber and melon. When this technique was applied in male flowers, anther tissues were found to be infected; whereas the pollen grains were found to be virus-free. These results have meaningful epidemiological implications for the management of CGMMV, particularly with regard to virus transfer via seed and the role of insects as CGMMV vectors.

  20. When biogeographical provinces collide: Hybridization of reef fishes at the crossroads of marine biogeographical provinces in the Arabian Sea

    KAUST Repository

    DiBattista, Joseph

    2015-04-01

    Aim: Suture zones are areas where closely related species from different biogeographical regions come into contact and interbreed. This concept originated from the study of terrestrial ecosystems but it remains unclear whether a similar phenomenon occurs in the marine environment. Here we investigate a potential suture zone from a previously unknown hybrid hotspot at the Socotra Archipelago (Yemen), located in the Arabian Sea, where fauna from the Red Sea, Gulf of Aden, Arabian Sea, western Indian Ocean and greater Indo-Polynesian Province intersect. Location: Red Sea, Gulf of Aden, Arabian Sea and Indian Ocean. Methods: Putative hybrid reef fish were identified based on intermediate coloration and morphology. Underwater observations and collections were conducted to determine: (1) whether parent species form heterospecific social groups or breeding pairs; (2) the sex and reproductive status of morphologically intermediate individuals; and (3) whether parent species were forming mixed species associations owing to a dearth of conspecific partners. To support hybrid status, morphologically intermediate and parental individuals were genotyped using mitochondrial DNA cytochrome c oxidase subunit I (COI), nuclear recombination-activating gene 2 (RAG2) and the nuclear TMO-4C4 (TMO) gene. Results: We observed putative hybrids involving 14 species from four reef fish families at Socotra. Most cases involved a parental species with a restricted distribution (e.g. Red Sea or Arabian Sea) and a broadly distributed Indo-Pacific species. In most cases, at least one of the parent species was rare at Socotra. Hybrid gene flow was largely unidirectional, and although introgression was rare, we found evidence that some butterflyfish and surgeonfish hybrids were fertile and formed breeding groups with parental species. Main conclusions: The rate of hybrid discovery at Socotra is much greater than that recorded elsewhere in the marine environment and involved both allopatric and

  1. Effect of thermal denaturation on the properties of collagen sponges from fish%热变性对鱼胶原海绵材料性能的影响∗

    Institute of Scientific and Technical Information of China (English)

    江颖; 邓明霞; 汪海婴; 张含俊; 汪海波

    2015-01-01

    In this paper,native collagen (NC)was extracted from skin of grass carp,and it was proved typeⅠcollagen with intact triple helix structure via the DSC and circular dichroism and SDS-PAGE analysis.Low ther-mal denaturation (LD),high thermal denaturation (HD)and complete thermal denaturation(CD)collagen sam-ples were obtained by heat treatment of NC under different conditions,and the degrees of thermal denaturation of them were 16.9%,57.9 and 100% respectively,according to the changes of thermal denaturation enthalpy from DSC determination.On this basis,the research focused on the influences of heat treatment on the collagen structure and the corresponding properties of collagen sponges.The results show that the onset endothermic temperature (To )is the threshold temperature for the molecule structure and sponges properties stability. When subj ected to temperatures above this threshold temperature even in a short time,the triple helix structure of collagen would completely or partially unfold.At the same time,the ordering of material structure,mechani-cal properties and enzymatic sensitivities of collagen sponges reduced,the cell proliferation ability enhanced a-long with the rise of degrees of thermal denaturation of collagen.The findings demonstrate To is the crucial temperature and this temperature should not be exceeded during the production and utilization of native colla-gens.%以草鱼皮为原料提取天然胶原蛋白,经DSC、圆二色谱和 SDS-PAGE 分析证实为具有完整三螺旋分子结构的Ⅰ型胶原.在不同条件下对该胶原进行热处理后分别获得了低变性、高变性和完全变性的胶原样品,热变性焓测定结果表明热变性程度分别为16.9%,57.9%和100%.在此基础上,重点研究了热变性处理后胶原结构和胶原海绵材料性能的变化.结果表明,草鱼皮胶原变性的起始温度(To,33℃)是其分子结构和胶原海绵材料性能改变的阀值温度.高于该温度条件下

  2. Variability of ribosomal DNA sites in Festuca pratensis, Lolium perenne, and their intergeneric hybrids, revealed by FISH and GISH.

    Science.gov (United States)

    Ksiazczyk, T; Taciak, M; Zwierzykowski, Z

    2010-01-01

    This study focuses on the variability of chromosomal location and number of ribosomal DNA (rDNA) sites in some diploid and autotetraploid Festuca pratensis and Lolium perenne cultivars, as well as on identification of rDNA-bearing chromosomes in their triploid and tetraploid F. pratensis × L. perenne hybrids. The rDNA loci were mapped using fluorescence in situ hybridization (FISH) with 5S and 25S rDNA probes, and the origin of parental genomes was verified by genomic in situ hybridization (GISH) with L. perenne genomic DNA as a probe, and F. pratensis genomic DNA as a block. FISH detected variation in the number and chromosomal location of both 5S and 45S rDNA sites. In F. pratensis mostly additional signals of 5S rDNA loci occurred, as compared with standard F. pratensis karyotypes. Losses of 45S rDNA loci were more frequent in L. perenne cultivars and intergeneric hybrids. Comparison of the F. pratensis and L. perenne genomes approved a higher number of rDNA sites as well as variation in chromosomal rDNA location in L. perenne. A greater instability of F. pratensis-genome-like and L. perenne-genome-like chromosomes in tetraploid hybrids was revealed, indicating gains and losses of rDNA loci, respectively. Our data indicate that the rDNA loci physically mapped on chromosomes 2 and 3 in F. pratensis and on chromosome 3 in L. perenne are useful markers for these chromosomes in intergeneric Festuca × Lolium hybrids.

  3. 21 CFR 866.4700 - Automated fluorescence in situ hybridization (FISH) enumeration systems.

    Science.gov (United States)

    2010-04-01

    ... detection and enumeration of FISH signals in interphase nuclei of formalin-fixed, paraffin-embedded human.... This device is intended for in vitro diagnostic use with FISH assays as an aid in the...

  4. Fluorescent in-situ hybridization (FISH for BCR/ABL in chronic myeloid leukemia after bone marrow transplantation

    Directory of Open Access Journals (Sweden)

    Maria de Lourdes Lopes Ferrari Chauffaille

    2001-01-01

    Full Text Available CONTEXT: Identification of Philadelphia chromosome or BCR/ABL gene rearrangement in chronic myeloid leukemia is important at diagnosis as well as after treatment. OBJECTIVE: To compare the results of karyotyping using fluorescent in-situ hybridization (FISH upon diagnosis and 1 year after bone marrow transplantation in 12 patients. TYPE OF STUDY: Diagnostic test and residual disease detection. SETTING: Hematology and Hemotherapy Department, Federal University of São Paulo/Escola Paulista de Medicina, São Paulo, Brazil. SAMPLE: 12 patients with chronic myeloid leukemia at diagnosis and 1 year after bone marrow transplantation. DIAGNOSTIC TEST: Karyotyping was done in the usual way and the BCR/ABL gene-specific probe was used for FISH. MAIN MEASUREMENTS: Disease at diagnosis and residual. RESULTS: At diagnosis, 10 patients presented t(9;22(q34.1;q11 as well as positive FISH. Two cases did not have metaphases but FISH was positive. After bone marrow transplantation, 8 patients presented normal karyotype, 1 had persistence of identifiable Philadelphia chromosome and 3 had no metaphases. Two cases showed complete chimera and 2 had donor and host cells simultaneously. FISH was possible in all cases after bone marrow transplantation and confirmed the persistence of identifiable Philadelphia chromosome clone in one patient, and identified another that did not present metaphases for analysis. Cases that showed mixed chimera in karyotype were negative for BCR/ABL by FISH. CONCLUSION: The applicability of FISH is clear, particularly for residual disease detection. Classical and molecular cytogenetics are complementary methods.

  5. Identification and characterization of marker chromosomes, de novo rearrangements and microdeletions in 100 cases with fluorescence in situ hybridization (FISH)

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, S.M.; Liu, Y.; Papenhausen, P.R. [Roche Biomedical Labs., Research Triangle Park, NC (United States)

    1994-09-01

    Results of molecular cytogenetic analysis are presented for 100 cases in which fluorescence in situ hybridization (FISH) was used as an adjunct to standard cytogenetics. Commercially available centromeric, telomeric, chromosome painting and unique sequence probes were used. Cases were from a 12-month period (June 1993-May 1994) and included examples of sex chromosome abnormalities (8), duplications (5), de novo translocations (6), satellited (12) and non-satellited (7) markers, and microdeletion syndromes (62). Satellited marker chromosomes were evaluated using a combination of DAPI/Distamycin A staining, hybridization with a classical satellite probe for chromosome 15 and hybridization with alpha-satellite probes for chromosomes 13, 14, 21 and 22. Markers positive for the chromosome 15 probe were further evaluated using unique sequence probes for the Prader-Willi/Angelman region. Microdeletion analysis was performed for Prader-Willi/Angelman (49) and DiGeorge/VCF (13) syndromes. Seven cases evaluated for Prader-Willi/Angelman syndrome demonstrated evidence of a deletion within this region. Uniparental disomy analysis was available in cases where a deletion was not detected by FISH, yet follow-up was clinically indicated. Two cases evaluated for DiGeorge/VCF syndrome demonstrated molecular evidence of a deletion. Included in our analysis is an example of familial DiGeorge syndrome.

  6. Fishing

    Institute of Scientific and Technical Information of China (English)

    姜群山

    2002-01-01

    @@ Last Saturday my cousin (表兄) came to my home. We were very happy to see each other. We decided that the next day we went to fish. We got up very early that day. When we left home,the moon could still be seen in the sky.

  7. Human disturbance causes the formation of a hybrid swarm between two naturally sympatric fish species.

    Science.gov (United States)

    Hasselman, Daniel J; Argo, Emily E; McBride, Meghan C; Bentzen, Paul; Schultz, Thomas F; Perez-Umphrey, Anna A; Palkovacs, Eric P

    2014-03-01

    Most evidence for hybrid swarm formation stemming from anthropogenic habitat disturbance comes from the breakdown of reproductive isolation between incipient species, or introgression between allopatric species following secondary contact. Human impacts on hybridization between divergent species that naturally occur in sympatry have received considerably less attention. Theory predicts that reinforcement should act to preserve reproductive isolation under such circumstances, potentially making reproductive barriers resistant to human habitat alteration. Using 15 microsatellites, we examined hybridization between sympatric populations of alewife (Alosa pseudoharengus) and blueback herring (A. aestivalis) to test whether the frequency of hybridization and pattern of introgression have been impacted by the construction of a dam that isolated formerly anadromous populations of both species in a landlocked freshwater reservoir. The frequency of hybridization and pattern of introgression differed markedly between anadromous and landlocked populations. The rangewide frequency of hybridization among anadromous populations was generally 0-8%, whereas all landlocked individuals were hybrids. Although neutral introgression was observed among anadromous hybrids, directional introgression leading to increased prevalence of alewife genotypes was detected among landlocked hybrids. We demonstrate that habitat alteration can lead to hybrid swarm formation between divergent species that naturally occur sympatrically, and provide empirical evidence that reinforcement does not always sustain reproductive isolation under such circumstances.

  8. Fluorescence in situ hybridizations (FISH) for the localization of viruses and endosymbiotic bacteria in plant and insect tissues.

    Science.gov (United States)

    Kliot, Adi; Kontsedalov, Svetlana; Lebedev, Galina; Brumin, Marina; Cathrin, Pakkianathan Britto; Marubayashi, Julio Massaharu; Skaljac, Marisa; Belausov, Eduard; Czosnek, Henryk; Ghanim, Murad

    2014-02-24

    Fluorescence in situ hybridization (FISH) is a name given to a variety of techniques commonly used for visualizing gene transcripts in eukaryotic cells and can be further modified to visualize other components in the cell such as infection with viruses and bacteria. Spatial localization and visualization of viruses and bacteria during the infection process is an essential step that complements expression profiling experiments such as microarrays and RNAseq in response to different stimuli. Understanding the spatiotemporal infections with these agents complements biological experiments aimed at understanding their interaction with cellular components. Several techniques for visualizing viruses and bacteria such as reporter gene systems or immunohistochemical methods are time-consuming, and some are limited to work with model organisms and involve complex methodologies. FISH that targets RNA or DNA species in the cell is a relatively easy and fast method for studying spatiotemporal localization of genes and for diagnostic purposes. This method can be robust and relatively easy to implement when the protocols employ short hybridizing, commercially-purchased probes, which are not expensive. This is particularly robust when sample preparation, fixation, hybridization, and microscopic visualization do not involve complex steps. Here we describe a protocol for localization of bacteria and viruses in insect and plant tissues. The method is based on simple preparation, fixation, and hybridization of insect whole mounts and dissected organs or hand-made plant sections, with 20 base pairs short DNA probes conjugated to fluorescent dyes on their 5' or 3' ends. This protocol has been successfully applied to a number of insect and plant tissues, and can be used to analyze expression of mRNAs or other RNA or DNA species in the cell.

  9. On the Translation of Hybridity in the Chinese Version of Saving Fish from Drowning in the Light of Bakhtin’s Theory of Dialogue

    Institute of Scientific and Technical Information of China (English)

    HU Zheng-yan; WANG Ji-ping

    2015-01-01

    In translation, hybridity often poses a big challenge to the translator, especially when the hybrid text is a novel. The au⁃thor of this paper hopes to contribute to the study of the translation of hybridity in English novels by drawing on Bakhtin ’s theory of dialogue. By analyzing the translation of linguistic hybridity in Saving Fish from Drowning, the author of this paper finds that the translation by Caiju is not good enough and tries to provide a new direction by virtue of which translators can choose right transla⁃tion strategies.

  10. Fish predation on a Daphnia hybrid species complex: A factor explaining species coexistence?

    NARCIS (Netherlands)

    Spaak, P.; Hoekstra, J.F.

    1997-01-01

    Recent studies on the life histories of Daphnia hybrids and their parental species have revealed that hybrids can combine an intermediate size with a relatively high reproductive rate, which might explain their success in many European lakes. Based on this information, we formulated the temporal

  11. Fully automated fluorescent in situ hybridization (FISH staining and digital analysis of HER2 in breast cancer: a validation study.

    Directory of Open Access Journals (Sweden)

    Elise M J van der Logt

    Full Text Available HER2 assessment is routinely used to select patients with invasive breast cancer that might benefit from HER2-targeted therapy. The aim of this study was to validate a fully automated in situ hybridization (ISH procedure that combines the automated Leica HER2 fluorescent ISH system for Bond with supervised automated analysis with the Visia imaging D-Sight digital imaging platform. HER2 assessment was performed on 328 formalin-fixed/paraffin-embedded invasive breast cancer tumors on tissue microarrays (TMA and 100 (50 selected IHC 2+ and 50 random IHC scores full-sized slides of resections/biopsies obtained for diagnostic purposes previously. For digital analysis slides were pre-screened at 20x and 100x magnification for all fluorescent signals and supervised-automated scoring was performed on at least two pictures (in total at least 20 nuclei were counted with the D-Sight HER2 FISH analysis module by two observers independently. Results were compared to data obtained previously with the manual Abbott FISH test. The overall agreement with Abbott FISH data among TMA samples and 50 selected IHC 2+ cases was 98.8% (κ = 0.94 and 93.8% (κ = 0.88, respectively. The results of 50 additionally tested unselected IHC cases were concordant with previously obtained IHC and/or FISH data. The combination of the Leica FISH system with the D-Sight digital imaging platform is a feasible method for HER2 assessment in routine clinical practice for patients with invasive breast cancer.

  12. Automation of ALK gene rearrangement testing with fluorescence in situ hybridization (FISH): a feasibility study.

    Science.gov (United States)

    Zwaenepoel, Karen; Merkle, Dennis; Cabillic, Florian; Berg, Erica; Belaud-Rotureau, Marc-Antoine; Grazioli, Vittorio; Herelle, Olga; Hummel, Michael; Le Calve, Michele; Lenze, Dido; Mende, Stefanie; Pauwels, Patrick; Quilichini, Benoit; Repetti, Elena

    2015-02-01

    In the past several years we have observed a significant increase in our understanding of molecular mechanisms that drive lung cancer. Specifically in the non-small cell lung cancer sub-types, ALK gene rearrangements represent a sub-group of tumors that are targetable by the tyrosine kinase inhibitor Crizotinib, resulting in significant reductions in tumor burden. Phase II and III clinical trials were performed using an ALK break-apart FISH probe kit, making FISH the gold standard for identifying ALK rearrangements in patients. FISH is often considered a labor and cost intensive molecular technique, and in this study we aimed to demonstrate feasibility for automation of ALK FISH testing, to improve laboratory workflow and ease of testing. This involved automation of the pre-treatment steps of the ALK assay using various protocols on the VP 2000 instrument, and facilitating automated scanning of the fluorescent FISH specimens for simplified enumeration on various backend scanning and analysis systems. The results indicated that ALK FISH can be automated. Significantly, both the Ikoniscope and BioView system of automated FISH scanning and analysis systems provided a robust analysis algorithm to define ALK rearrangements. In addition, the BioView system facilitated consultation of difficult cases via the internet.

  13. Action spectrum for melanoma induction in hybrid fish of the genus Xiphophorus

    Energy Technology Data Exchange (ETDEWEB)

    Setlow, R.B.

    1997-03-01

    Cutaneous malignant melanoma (CMM) is a complicated disease that is dependent on a number of factors that influence its incidence in ways that are quantitatively uncertain. The incidence of CMM increases with proximity to the Equator -- an observation in line with the conclusion that sun exposure is the most important etiologic agent. However, the latitude effect does not implicate UVB because the intensities of all spectral regions increase toward the Equator. An understanding of the useful public health measures to lower the incidence of CMM would benefit greatly if the spectral region of sunlight implicated in melanoma incidence were known. Such knowledge requires animal models to evaluate the incidence as a function of wavelength. There are marsupial models, a transgenic mouse model, and a fish model. To date, only the fish model has been employed to obtain an action spectrum. The paper describes a fish model, implications of the fish spectrum, and epidemiological data.

  14. In vitro formation and thermal transition of novel hybrid fibrils from type I fish scale collagen and type I porcine collagen

    OpenAIRE

    Song Chen, Toshiyuki Ikoma, Nobuhiro Ogawa, Satoshi Migita, Hisatoshi Kobayashi and Nobutaka Hanagata

    2010-01-01

    Novel type I collagen hybrid fibrils were fabricated by neutralizing a mixture of type I fish scale collagen solution and type I porcine collagen solution with a phosphate buffer saline at 28 °C. Their structure was discussed in terms of the volume ratio of fish/porcine collagen solution. Scanning electron and atomic force micrographs showed that the diameter of collagen fibrils derived from the collagen mixture was larger than those derived from each collagen, and all resultant fibrils exhib...

  15. Prediction of melting temperatures in fluorescence in situ hybridization (FISH) procedures using thermodynamic models

    DEFF Research Database (Denmark)

    Fontenete, Sílvia; Guimarães, Nuno; Wengel, Jesper

    2016-01-01

    Abstract The thermodynamics and kinetics of DNA hybridization, i.e. the process of self-assembly of one, two or more complementary nucleic acid strands, has been studied for many years. The appearance of the nearest-neighbor model led to several theoretical and experimental papers on DNA thermody......Abstract The thermodynamics and kinetics of DNA hybridization, i.e. the process of self-assembly of one, two or more complementary nucleic acid strands, has been studied for many years. The appearance of the nearest-neighbor model led to several theoretical and experimental papers on DNA...... thermodynamics that provide reasonably accurate thermodynamic information on nucleic acid duplexes and allow estimation of the melting temperature. Because there are no thermodynamic models specifically developed to predict the hybridization temperature of a probe used in a fluorescence in situ hybridization...

  16. Single DNA denaturation and bubble dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Metzler, Ralf [Physics Department, Technical University of Munich, James Franck Strasse, 85747 Garching (Germany); Ambjoernsson, Tobias [Chemistry Department, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139 (United States); Hanke, Andreas [Department of Physics and Astronomy, University of Texas, 80 Fort Brown, Brownsville (United States); Fogedby, Hans C [Department of Physics and Astronomy, University of Arhus, Ny Munkegade, 8000 Arhus C (Denmark)], E-mail: metz@ph.tum.de

    2009-01-21

    While the Watson-Crick double-strand is the thermodynamically stable state of DNA in a wide range of temperature and salt conditions, even at physiological conditions local denaturation bubbles may open up spontaneously due to thermal activation. By raising the ambient temperature, titration, or by external forces in single molecule setups bubbles proliferate until full denaturation of the DNA occurs. Based on the Poland-Scheraga model we investigate both the equilibrium transition of DNA denaturation and the dynamics of the denaturation bubbles with respect to recent single DNA chain experiments for situations below, at, and above the denaturation transition. We also propose a new single molecule setup based on DNA constructs with two bubble zones to measure the bubble coalescence and extract the physical parameters relevant to DNA breathing. Finally we consider the interplay between denaturation bubbles and selectively single-stranded DNA binding proteins.

  17. Single DNA denaturation and bubble dynamics

    DEFF Research Database (Denmark)

    Metzler, Ralf; Ambjörnsson, Tobias; Hanke, Andreas

    2009-01-01

    for situations below, at, and above the denaturation transition. We also propose a new single molecule setup based on DNA constructs with two bubble zones to measure the bubble coalescence and extract the physical parameters relevant to DNA breathing. Finally we consider the interplay between denaturation......While the Watson-Crick double-strand is the thermodynamically stable state of DNA in a wide range of temperature and salt conditions, even at physiological conditions local denaturation bubbles may open up spontaneously due to thermal activation. By raising the ambient temperature, titration......, or by external forces in single molecule setups bubbles proliferate until full denaturation of the DNA occurs. Based on the Poland-Scheraga model we investigate both the equilibrium transition of DNA denaturation and the dynamics of the denaturation bubbles with respect to recent single DNA chain experiments...

  18. The dynamics of the DNA denaturation transition

    CERN Document Server

    van Erp, Titus S

    2012-01-01

    The dynamics of the DNA denaturation is studied using the Peyrard-Bishop-Dauxois model. The denaturation rate of double stranded polymers decreases exponentially as function of length below the denaturation temperature. Above Tc, the rate shows a minimum, but then increases as function of length. We also examine the influence of sequence and solvent friction. Molecules having the same number of weak and strong base-pairs can have significantly different opening rates depending on the order of base-pairs.

  19. Assessment of chromosomal abnormalities in sperm of infertile men using sperm karyotyping and multicolour fluorescence in situ hybridization (FISH)

    Energy Technology Data Exchange (ETDEWEB)

    Moosani, N.; Martin, R.H. [Alberta Children`s Hospital and Univ. of Calgary (Canada)

    1994-09-01

    Individuals with male factor infertility resulting from idiopathic oligo-, astheno- or teratozoospermia are frequently offered IVF in an attempt to increase their chances of having a child. A concern remains whether these infertile males have an elevated risk of transmitting chromosomal abnormalities to their offspring. Sperm chromosomal complements from these men were assayed using the human sperm/hamster oocyte fusion system and fluorescence in situ hybridization (FISH) on sperm nuclei. For each of 5 infertile patients, 100 sperm karyotypes were analyzed and multicolour FISH analysis was performed on a minimum of 10,000 sperm nuclei for each chromosome-specific DNA probe for chromosomes 1 (pUC1.77), 12 (D12Z3), X (XC) and Y (DYZ3). As a group, the infertile patients showed increased frequencies of both numerical ({chi}{sup 2}=17.26, {proportional_to} <0.001) and total abnormalities ({chi}{sup 2}=7.78, {proportional_to} <0.01) relative to control donors when assessed by sperm karyotypes. Analysis of sperm nuclei by FISH indicated a significant increase in the frequency of disomy for chromosome 1 in three of the five patients as compared to control donors ({chi}{sup 2}>8.35, {proportional_to} <0.005). In addition, the frequency of XY disomy was significantly higher in four of the five patients studied by FISH ({chi}{sup 2}>10.58, {proportional_to}<0.005), suggesting that mis-segregation caused by the failure of the XY bivalent to pair may play a role in idiopathic male infertility.

  20. Bubbles and denaturation in DNA

    CERN Document Server

    Van Erp, T S; Peyrard, M; Erp, Titus S. van; Cuesta-Lopez, Santiago; Peyrard, Michel

    2006-01-01

    The local opening of DNA is an intriguing phenomenon from a statistical physics point of view, but is also essential for its biological function. For instance, the transcription and replication of our genetic code can not take place without the unwinding of the DNA double helix. Although these biological processes are driven by proteins, there might well be a relation between these biological openings and the spontaneous bubble formation due to thermal fluctuations. Mesoscopic models, like the Peyrard-Bishop-Dauxois model, have fairly accurately reproduced some experimental denaturation curves and the sharp phase transition in the thermodynamic limit. It is, hence, tempting to see whether these models could be used to predict the biological activity of DNA. In a previous study, we introduced a method that allows to obtain very accurate results on this subject, which showed that some previous claims in this direction, based on molecular dynamics studies, were premature. This could either imply that the present...

  1. DNA denaturation in ionic solution

    Science.gov (United States)

    Maity, Arghya; Singh, Amar; Singh, Navin

    2016-05-01

    Salt or cations, present in solution play an important role in DNA denaturation and folding kinetics of DNA helix. In this work we study the thermal melting of double stranded DNA (dsDNA) molecule using Peyrard Bishop Dauxois (PBD) model. We modify the potential of H-bonding between the bases of the complimentary strands to introduce the salt and solvent effect. We choose different DNA sequences having different contents of GC pairs and calculate the melting temperatures. The melting temperature increases logarithmically with the salt concentration of the solution. The more GC base pairs in the chain enhance the stability of DNA chain at a fix salt concentration. The obtained results are in good accordance with experimental findings.

  2. Gill monogeneans of Nile tilapia (Oreochromis niloticus) and red hybrid tilapia (Oreochromis spp.) from the wild and fish farms in Perak, Malaysia: infection dynamics and spatial distribution.

    Science.gov (United States)

    Lim, Shen-Yin; Ooi, Ai-Lin; Wong, Wey-Lim

    2016-01-01

    Tilapia is one of the commercially important fish in Malaysia as well as in other parts of the world. An understanding of monogenean infection dynamics in tilapia fish may assist us in searching for some intervention measures in reducing the loss of fish caused by parasitic diseases. The present study aimed (1) to compare infection level of monogeneans between the wild and cultured Oreochromis niloticus, and between the cultured O. niloticus and cultured red hybrid tilapia, and (2) to examine the spatial distribution of monogenean species over the gills of the different host species. From a total of 75 fish specimens, six species of monogeneans from two genera: Cichlidogyrus (C. halli, C. mbirizei, C. sclerosus, C. thurstonae, C. tilapiae) and Scutogyrus (S. longicornis) were identified. Data showed that the infection level of cultured O. niloticus was higher than that of the wild O. niloticus, however, the former was lower than that of the cultured red hybrid tilapia. Higher species richness of monogeneans was observed in the cultured red hybrid tilapia as compared to the others. Results for spatial distribution showed that the monogeneans have no preference on the left or right sides of the gills. However, C. halli, C. mbirizei, and C. tilapiae showed preferences on specific gill arches in the cultured O. niloticus and red hybrid tilapia. In general, the gill arch IV harboured the least number of monogeneans. The susceptibility of monogenean infection between the different types of tilapia is discussed.

  3. Phylogeny-function analysis of (meta)genomic libraries: screening for expression of ribosomal RNA genes by large-insert library fluorescent in situ hybridization (LIL-FISH)

    NARCIS (Netherlands)

    Leveau, J.H.J.; Gerards, S.; De Boer, W.; Van Veen, J.A.

    2004-01-01

    We assessed the utility of fluorescent in situ hybridization (FISH) in the screening of clone libraries of (meta)genomic or environmental DNA for the presence and expression of bacterial ribosomal RNA (rRNA) genes. To establish proof-of-principle, we constructed a fosmid-based library in Escherichia

  4. Distribution characteristics of ammonia-oxidizing bacteria in the Typha latifolia constructed wetlands using fluorescent in situ hybridization (FISH).

    Science.gov (United States)

    Yan, Li; Inamori, Ryuhei; Gui, Ping; Xu, Kai-qin; Kong, Hai-nan; Matsumura, Masatoshi; Inamori, Yuhei

    2005-01-01

    A molecular biology method, fluorescent in situ hybridization (FISH), in which the pre-treatment was improved in allusion to the media of the constructed wetlands (CW), e.g. the soil and the grit, was used to investigate the vertical distribution characteristics of ammonia-oxidizing bacteria (AOB) quantity and the relation with oxidation-reduction potential (ORP) in the Typha latifolia constructed wetlands under three different loadings in summer from May to September. Results showed that the quantity of the AOB decreased in the Typha latifolia CW with the increase of vertical depth. However, the AOB quantity was 2-4 times the quantity of the control in the root area. Additionally, ORP in the rhizosphere was found to be higher than other areas, which showed that Typha latifolia CW was in an aerobic state in summer when using simulated non-point sewage at the rural area of Taihu Lake in China and small town combined sewage.

  5. Validation of interphase fluorescence in situ hybridization (iFISH for multiple myeloma using CD138 positive cells

    Directory of Open Access Journals (Sweden)

    Renata Kiyomi Kishimoto

    2016-06-01

    Full Text Available ABSTRACT BACKGROUND: Multiple myeloma is a plasma cell neoplasm with acquired genetic abnormalities of clinical and prognostic importance. Multiple myeloma differs from other hematologic malignancies due to a high fraction of low proliferating malignant plasma cells and the paucity of plasma cells in bone marrow aspiration samples, making cytogenetic analysis a challenge. An abnormal karyotype is found in only one-third of patients with multiple myeloma and interphase fluorescence in situ hybridization is the most useful test for studying the chromosomal abnormalities present in almost 90% of cases. However, it is necessary to study the genetic abnormalities in plasma cells after their identification or selection by morphology, immunophenotyping or sorting. Other challenges are the selection of the most informative FISH panel and determining cut-off levels for FISH probes. This study reports the validation of interphase fluorescence in situ hybridization using CD138 positive cells, according to proposed guidelines published by the European Myeloma Network (EMN in 2012. METHOD: Bone marrow samples from patients with multiple myeloma were used to standardize a panel of five probes [1q amplification, 13q14 deletion, 17p deletion, t(4;14, and t(14;16] in CD138+ cells purified by magnetic cell sorting. RESULTS: This test was validated with a low turnaround time and good reproducibility. Five of six samples showed genetic abnormalities. Monosomy/deletion 13 plus t(4;14 were found in two cases. CONCLUSION: This technique together with magnetic cell sorting is effective and can be used in the routine laboratory practice. In addition, magnetic cell sorting provides a pure plasma cell population that allows other molecular and genomic studies.

  6. Hybrid striped bass feeds based on fish oil, beef tallow, and eicosapentaenoic acid/docosahexaenoic acid supplements: Insight regarding fish oil sparing and demand for -3 long-chain polyunsaturated fatty acids.

    Science.gov (United States)

    Bowzer, J; Jackson, C; Trushenski, J

    2016-03-01

    Previous research suggests that saturated (SFA) and monounsaturated fatty acid (MUFA) rich lipids, including beef tallow, can make utilization or diet-to-tissue transfer of long-chain polyunsaturated fatty acids (LC-PUFA) more efficient. We hypothesized that using beef tallow as an alternative to fish oil may effectively reduce the LC-PUFA demand of hybrid striped bass × and allow for greater fish oil sparing. Accordingly, we evaluated growth performance and tissue fatty acid profiles of juvenile fish (23.7 ± 0.3 g) fed diets containing menhaden fish oil (considered an ideal source of LC-PUFA for this taxon), beef tallow (BEEF ONLY), or beef tallow amended with purified sources of eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA) to achieve levels corresponding to 50 or 100% of those observed in the FISH ONLY feed. Diets were randomly assigned to quadruplicate tanks of fish ( = 4; 10 fish/tank), and fish were fed assigned diets to apparent satiation once daily for 10 wk. Survival (98-100%) was equivalent among treatments, but weight gain (117-180%), specific growth rate (1.1-1.5% BW/d), feed intake (1.4-1.8% BW/d), thermal growth coefficient (0.50-0.70), and feed conversion ratio (FCR; 1.1-1.4, DM basis) varied. Except for FCR, no differences were observed between the FISH ONLY and BEEF ONLY treatments, but performance was generally numerically superior among fish fed the diets containing beef tallow supplemented with DHA at the 100% or both EPA and DHA at the 50% or 100% level. Tissue fatty acid composition was significantly distorted in favor among fish fed the beef tallow-based feeds; however, profile distortion was most overt in peripheral tissues. Results suggest that beef tallow may be used as a primary lipid source in practical diets for hybrid striped bass, but performance may be improved by supplementation with LC-PUFA, particularly DHA. Furthermore, our results suggest that -3 LC-PUFA requirements reported for hybrid striped bass may not be

  7. In vitro formation and thermal transition of novel hybrid fibrils from type I fish scale collagen and type I porcine collagen

    Energy Technology Data Exchange (ETDEWEB)

    Chen Song; Ogawa, Nobuhiro; Migita, Satoshi; Kobayashi, Hisatoshi [Biomaterials Center, National Institute for Materials Science, 1-2-1 Sengen, Tsukuba, Ibaraki 305-0047 (Japan); Ikoma, Toshiyuki [Department of Metallurgy and Ceramics Science, Tokyo Institute of Technology, O-okayama 2-12-1, Meguro-ku, Tokyo 152-8550 (Japan); Hanagata, Nobutaka, E-mail: HANAGATA.Nobutaka@nims.go.j [Nanotechnology Innovation Center, National Institute for Materials Science, 1-2-1 Sengen, Tsukuba, Ibaraki 305-0047 (Japan)

    2010-06-15

    Novel type I collagen hybrid fibrils were fabricated by neutralizing a mixture of type I fish scale collagen solution and type I porcine collagen solution with a phosphate buffer saline at 28 {sup 0}C. Their structure was discussed in terms of the volume ratio of fish/porcine collagen solution. Scanning electron and atomic force micrographs showed that the diameter of collagen fibrils derived from the collagen mixture was larger than those derived from each collagen, and all resultant fibrils exhibited a typical D-periodic unit of {approx}67 nm, irrespective of volume ratio of both collagens. Differential scanning calorimetry revealed only one endothermic peak for the fibrils derived from collagen mixture or from each collagen solution, indicating that the resultant collagen fibrils were hybrids of type I fish scale collagen and type I porcine collagen.

  8. In vitro formation and thermal transition of novel hybrid fibrils from type I fish scale collagen and type I porcine collagen

    Science.gov (United States)

    Chen, Song; Ikoma, Toshiyuki; Ogawa, Nobuhiro; Migita, Satoshi; Kobayashi, Hisatoshi; Hanagata, Nobutaka

    2010-06-01

    Novel type I collagen hybrid fibrils were fabricated by neutralizing a mixture of type I fish scale collagen solution and type I porcine collagen solution with a phosphate buffer saline at 28 °C. Their structure was discussed in terms of the volume ratio of fish/porcine collagen solution. Scanning electron and atomic force micrographs showed that the diameter of collagen fibrils derived from the collagen mixture was larger than those derived from each collagen, and all resultant fibrils exhibited a typical D-periodic unit of ~67 nm, irrespective of volume ratio of both collagens. Differential scanning calorimetry revealed only one endothermic peak for the fibrils derived from collagen mixture or from each collagen solution, indicating that the resultant collagen fibrils were hybrids of type I fish scale collagen and type I porcine collagen.

  9. In vitro formation and thermal transition of novel hybrid fibrils from type I fish scale collagen and type I porcine collagen

    Directory of Open Access Journals (Sweden)

    Song Chen, Toshiyuki Ikoma, Nobuhiro Ogawa, Satoshi Migita, Hisatoshi Kobayashi and Nobutaka Hanagata

    2010-01-01

    Full Text Available Novel type I collagen hybrid fibrils were fabricated by neutralizing a mixture of type I fish scale collagen solution and type I porcine collagen solution with a phosphate buffer saline at 28 °C. Their structure was discussed in terms of the volume ratio of fish/porcine collagen solution. Scanning electron and atomic force micrographs showed that the diameter of collagen fibrils derived from the collagen mixture was larger than those derived from each collagen, and all resultant fibrils exhibited a typical D-periodic unit of ~67 nm, irrespective of volume ratio of both collagens. Differential scanning calorimetry revealed only one endothermic peak for the fibrils derived from collagen mixture or from each collagen solution, indicating that the resultant collagen fibrils were hybrids of type I fish scale collagen and type I porcine collagen.

  10. Feasibility of using fluorescence in situ hybridization (FISH) to detect early gene changes in sputum cells from uranium miners

    Energy Technology Data Exchange (ETDEWEB)

    Neft, R.E.; Rogers, J.L.; Belinsky, S.A. [and others

    1995-12-01

    Epidemiological studies have shown that combined exposure to radon progeny and tobacco smoke produce a greater than additive or synergistic increase in lung cancer risk. Lung cancer results from multiple genetic changes over a long period of time. An early change that occurs in lung cancer is trisomy 7 which is found in 50% of non-small cell lung cancer and in the far margins of resected lung tumors. The 80% mortality associated with lung cancer is in part related to the high proportion of patients who present with an advanced, unresectable tumor. Therefore, early detection of patients at risk for tumor development is critical to improve treatment of this disease. Currently, it is difficult to detect lung cancer early while it is still amendable by surgery. Saccomanno, G. has shown that premalignant cytologic changes in sputum cells collected from uranium miners can be detected by a skilled, highly trained cytopathologist. A more objective alternative for identifying premalignant cells in sputum may be to determine whether an early genetic change such as trisomy 7 is present in these cells. Fluorescence in situ hybridization (FISH) can be used to identify cells with trisomy 7. The results of this investigation indicate that FISH may prove to be an accurate, efficient method to test at-risk individuals for genetic alterations in bronchial epithelial cells from sputum.

  11. New DAPI and FISH findings on egg maturation processes in related hybridogenetic and parthenogenetic Bacillus hybrids (Insecta, Phasmatodea).

    Science.gov (United States)

    Marescalchi, O; Scali, V

    2001-10-01

    Bacillus stick insects have proved adequate for studying a wide array of reproductive modes: sexual, parthenogenetic, hybridogenetic, androgenetic. Hybridogenetic strains (B. rossius-grandii) were thought to discard the paternal "grandii" haploset during first meiotic division and keep the "rossius" hemiclone, whereas the clonal B. whitei (=rossius/grandii) would maintain its hybrid structure by fusing back two nonsister nuclei-each derived from previously segregated heterospecific complements-by the end of the 2(nd) meiotic division. New investigations on laid eggs and ovariole squashes, either DAPI stained or FISH labeled, revealed that in hybridogens the "grandii" set is excluded from the germ line prior to meiosis and that a DNA extra-synthesis should occur to produce hemiclonal eggs after two cytologically normal meiotic divisions. On the other hand, in B. whitei eggs no genome segregation appears to occur and an intrameiotic DNA extra-synthesis must take place to produce 2n tetrachromatidic oocytes I; these divide twice and give unreduced clonal eggs. The new findings bring hybridogenetic oogenesis of Bacillus to be coincident with that of the known hemiclonal organisms and point to an independent onset of B. whitei from hemiclonal strains. In addition, B. whitei gains a closer resemblance to B. lynceorum owing to the sharing of a cytologically identical egg maturation mechanism, of the same maternal ancestor and of peculiar chromosomal features. It is here suggested that B. lynceorum originated from the incorporation of an "atticus" genome into a B. whitei egg, according to a pathway of repeated hybridization often occurred with other polyploid hybrids.

  12. Diagnosis of a constitutional five-chromosome rearrangement by fluorescent in situ hybridization (FISH)

    Energy Technology Data Exchange (ETDEWEB)

    Tsien, F.; Shapira, E. [Tulane Univ. School of Medicine, New Orleans, LA (United States); Carvalho, T. [Hospital Sarah Kubitschek, Brasilia (Brazil)] [and others

    1994-09-01

    Complex chromosomal rearrangements are structural rearrangements involving at least three chromosomes and three or more chromosome breakpoints. Such karyotypes are often acquired during cancer multi-step development and in chromosome instability syndromes. However, extremely rare constitutional forms have been reported, most of which are incompatible with life. We present a 2-year-old female with de novo complex rearrangement consisting of five chromosomes and nine breakpoints. Clinical evaluation at two years of age revealed a weight of 5 kg, length of 66 cm, and had circumference of 38 cm, all below the 5th percentile, microcephaly, trigonocephaly, epicanthal folds, inguinal hernia, left clubfoot, hypertonicity, and developmental delay. The neurological examination revealed chorea-acanthocytosis and psychomotor delay. Cultured lymphocytes and fibroblasts revealed a karyotype consisting of five derivative chromosomes. The metaphases were further analyzed by FISH using chromosome-specific libraries and telomeric probes in order to delineate the composition of the rearranged chromosomes; FISH results demonstrated a karyotype of: 46,XX,1pter{r_arrow}1q25::1q42.1{r_arrow}1qter, 2pter{r_arrow}q32.3::1q32.3{r_arrow}2q41::2q37.3{r_arrow}2qter, 7qter{r_arrow}7q21.2::6q22.3{r_arrow}6qter::1q31{r_arrow}1q32.3::6p23{r_arrow}6q22.3, 7pter{r_arrow}7q21.1::6p23{r_arrow}6pter, 2q33{r_arrow}2q37, 1::9p21{r_arrow}9qter. This analysis demonstrates the usefulness of FISH in characterizing complex chromosome rearrangements otherwise difficult to correctly interpret using classical cytogenetics alone.

  13. Calcium Binding and Disulfide Bonds Regulate the Stability of Secretagogin towards Thermal and Urea Denaturation

    Science.gov (United States)

    Weiffert, Tanja; Ní Mhurchú, Niamh; O’Connell, David; Linse, Sara

    2016-01-01

    Secretagogin is a calcium-sensor protein with six EF-hands. It is widely expressed in neurons and neuro-endocrine cells of a broad range of vertebrates including mammals, fishes and amphibia. The protein plays a role in secretion and interacts with several vesicle-associated proteins. In this work, we have studied the contribution of calcium binding and disulfide-bond formation to the stability of the secretagogin structure towards thermal and urea denaturation. SDS-PAGE analysis of secretagogin in reducing and non-reducing conditions identified a tendency of the protein to form dimers in a redox-dependent manner. The denaturation of apo and Calcium-loaded secretagogin was studied by circular dichroism and fluorescence spectroscopy under conditions favoring monomer or dimer or a 1:1 monomer: dimer ratio. This analysis reveals significantly higher stability towards urea denaturation of Calcium-loaded secretagogin compared to the apo protein. The secondary and tertiary structure of the Calcium-loaded form is not completely denatured in the presence of 10 M urea. Reduced and Calcium-loaded secretagogin is found to refold reversibly after heating to 95°C, while both oxidized and reduced apo secretagogin is irreversibly denatured at this temperature. Thus, calcium binding greatly stabilizes the structure of secretagogin towards chemical and heat denaturation. PMID:27812162

  14. Identification of intracellular bacteria in adenoid and tonsil tissue specimens: the efficiency of culture versus fluorescent in situ hybridization (FISH).

    Science.gov (United States)

    Stępińska, M; Olszewska-Sosińska, O; Lau-Dworak, M; Zielnik-Jurkiewicz, B; Trafny, E A

    2014-01-01

    Monocyte/macrophage cells from human nasopharyngeal lymphoid tissue can be a source of bacteria responsible for human chronic and recurrent upper respiratory tract infection. Detection and characterization of pathogens surviving intracellularly could be a key element in bacteriological diagnosis of the infections as well as in the study on interactions between bacteria and their host. The present study was undertaken to assess the possibility of isolation of viable bacteria from the cells expressing monocyte/macrophage marker CD14 in nasopharyngeal lymphoid tissue. Overall, 74 adenotonsillectomy specimens (adenoids and tonsils) from 37 children with adenoid hypertrophy and recurrent infections as well as 15 specimens from nine children with adenoid hypertrophy, which do not suffer from upper respiratory tract infections (the control group), were studied. The suitability of immunomagnetic separation for extraction of CD14(+) cells from lymphoid tissue and for further isolation of the intracellular pathogens has been shown. The coexistence of living pathogens including Haemophilus influenzae, Staphylococcus aureus, and Streptococcus pyogenes with the bacteria representing normal nasopharyngeal microbiota inside CD14(+) cells was demonstrated. Twenty-four strains of these pathogens from 32.4 % of the lysates of CD14(+) cells were isolated. Concurrently, the fluorescent in situ hybridization (FISH) with a universal EUB388, and the species-specific probes demonstrated twice more often the persistence of these bacterial species in the lysates of CD14(+) cells than conventional culture. Although the FISH technique appears to be more sensitive than traditional culture in the intracellular bacteria identification, the doubts on whether the bacteria are alive, and therefore, pathogenic would still exist without the strain cultivation.

  15. Locked Nucleic Acid Flow Cytometry-fluorescence in situ Hybridization (LNA flow-FISH): A Method for Bacterial Small RNA Detection

    Science.gov (United States)

    2012-01-10

    Friedrich, U. & Lenke, J. Improved Enumeration of Lactic Acid Bacteria in Mesophilic Dairy Starter Cultures by Using Multiplex Quantitative Real...messenger RNA using locked nucleic acid probes. Anal. Biochem. 390, 109-114 (2009). 13. Waters, L. & Storz, G. Regulatory RNAs in bacteria . Cell. 136, 615...Video Article Locked Nucleic Acid Flow Cytometry-fluorescence in situ Hybridization (LNA flow-FISH): a Method for Bacterial Small RNA Detection Kelly

  16. Phylogeny-function analysis of (meta)genomic libraries: screening for expression of ribosomal RNA genes by large-insert library fluorescent in situ hybridization (LIL-FISH).

    Science.gov (United States)

    Leveau, Johan H J; Gerards, Saskia; de Boer, Wietse; van Veen, Johannes A

    2004-09-01

    We assessed the utility of fluorescent in situ hybridization (FISH) in the screening of clone libraries of (meta)genomic or environmental DNA for the presence and expression of bacterial ribosomal RNA (rRNA) genes. To establish proof-of-principle, we constructed a fosmid-based library in Escherichia coli of large-sized genomic DNA fragments of the mycophagous soil bacterium Collimonas fungivorans, and hybridized 768 library clones with the Collimonas-specific fluorescent probe CTE998-1015. Critical to the success of this approach (which we refer to as large-insert library FISH or LIL-FISH) was the ability to induce fosmid copy number, the exponential growth status of library clones in the FISH assay and the use of a simple pooling strategy to reduce the number of hybridizations. Twelve out of 768 E. coli clones were suspected to harbour and express Collimonas 16S rRNA genes based on their hybridization to CTE998-1015. This was confirmed by the finding that all 12 clones were also identified in an independent polymerase chain reaction-based screening of the same 768 clones using a primer set for the specific detection of Collimonas 16S ribosomal DNA (rDNA). Fosmids isolated from these clones were grouped by restriction analysis into two distinct contigs, confirming that C. fungivorans harbours at least two 16S rRNA genes. For one contig, representing 1-2% of the genome, the nucleotide sequence was determined, providing us with a narrow but informative view of Collimonas genome structure and content.

  17. Three sympatric karyomorphs in the fish Astyanax fasciatus (Teleostei, Characidae) do not seem to hybridize in natural populations

    Science.gov (United States)

    Ferreira-Neto, Maressa; Artoni, Roberto Ferreira; Vicari, Marcelo Ricardo; Moreira-Filho, Orlando; Camacho, Juan Pedro Martínez; Bakkali, Mohammed; de Oliveira, Claudio; Foresti, Fausto

    2012-01-01

    Abstract Ninety individuals of the characid fish Astyanax fasciatus (Cuvier, 1819) were collected at Água da Madalena stream (Botucatu, São Paulo, Brazil) and analyzed for diploid chromosome number 2n and karyotype composition as well as for the chromosomal location of the 5S and 18S ribosomal DNA (rDNA). Whereas no chromosome differences were associated with sex, three different karyomorphs with diploid chromosome numbers 2n=46, 2n=48 and 2n=50 were found. No intermediate 2n numbers were discovered. The 2n=50 karyomorph showed some differences in 18S rDNA location compared to the two other karyomorphs. Finally, all specimens with the 2n=46 karyomorph showed the presence of a partly heterochromatic macro supernumerary chromosome, which was absent in all individuals with the two other karyomorphs. All these results suggest that indviduals of the three different karyomorphs are not likely to hybridize in the examined populations. Our findings strongly suggest the presence of three separate species (sensu biological species concept) easily diagnosed on the basis of differences in the diploid chromosome numbers and other chromosomal markers. PMID:24260650

  18. Three sympatric karyomorphs in the fish Astyanax fasciatus (Teleostei, Characidae do not seem to hybridize in natural populations

    Directory of Open Access Journals (Sweden)

    Maressa Ferreira-Neto

    2012-01-01

    Full Text Available Ninety individuals of the characid fish Astyanax fasciatus (Cuvier, 1819 were collected at Água da Madalena stream (Botucatu, São Paulo, Brazil and analyzed for diploid chromosome number 2n and karyotype composition as well as for the chromosomal location of the 5S and 18S ribosomal DNA (rDNA. Whereas no chromosome differences were associated with sex, three different karyomorphs with diploid chromosome numbers 2n=46, 2n=48 and 2n=50 were found. No intermediate 2n numbers were discovered. The 2n=50 karyomorph showed some differences in 18S rDNA location compared to the two other karyomorphs. Finally, all specimens with the 2n=46 karyomorph showed the presence of a partly heterochromatic macro supernumerary chromosome, which was absent in all individuals with the two other karyomorphs. All these results suggest that indviduals of the three different karyomorphs are not likely to hybridize in the examined populations. Our findings strongly suggest the presence of three separate species (sensu biological species concept easily diagnosed on the basis of differences in the diploid chromosome numbers and other chromosomal markers.

  19. Detection of Inter-chromosomal Stable Aberrations by Multiple Fluorescence In Situ Hybridization (mFISH) and Spectral Karyotyping (SKY) in Irradiated Mice.

    Science.gov (United States)

    Pathak, Rupak; Koturbash, Igor; Hauer-Jensen, Martin

    2017-01-11

    Ionizing radiation (IR) induces numerous stable and unstable chromosomal aberrations. Unstable aberrations, where chromosome morphology is substantially compromised, can easily be identified by conventional chromosome staining techniques. However, detection of stable aberrations, which involve exchange or translocation of genetic materials without considerable modification in the chromosome morphology, requires sophisticated chromosome painting techniques that rely on in situ hybridization of fluorescently labeled DNA probes, a chromosome painting technique popularly known as fluorescence in situ hybridization (FISH). FISH probes can be specific for whole chromosome/s or precise sub-region on chromosome/s. The method not only allows visualization of stable aberrations, but it can also allow detection of the chromosome/s or specific DNA sequence/s involved in a particular aberration formation. A variety of chromosome painting techniques are available in cytogenetics; here two highly sensitive methods, multiple fluorescence in situ hybridization (mFISH) and spectral karyotyping (SKY), are discussed to identify inter-chromosomal stable aberrations that form in the bone marrow cells of mice after exposure to total body irradiation. Although both techniques rely on fluorescent labeled DNA probes, the method of detection and the process of image acquisition of the fluorescent signals are different. These two techniques have been used in various research areas, such as radiation biology, cancer cytogenetics, retrospective radiation biodosimetry, clinical cytogenetics, evolutionary cytogenetics, and comparative cytogenetics.

  20. [Interphase cytogenetics of the breast tumors with fluorescence in situ hybridization (FISH) on cytologic preparation--its practice and clinical applications].

    Science.gov (United States)

    Shibuya, M; Osamura, R Y

    1996-05-01

    Fluorescence in situ hybridization (FISH) study with the chromosome specific probes is performed in the interphase nuclei of the routinely processed cytologic preparation of the breast tumors. Numerical aberrations on the chromosomes 1, 3, 11 or 17 were detected in more than 80% of the malignant tumors, but not in the benign tumors. Marked heterogeneity of the polysomies is noted in the malignant tumor cells. A few malignant cases revealed monosomy of chromosome 17. No apparent correlation between the numerical abnormalities and the histological features in malignant tumors is identified. These results suggest that the interphase cytogenetics with FISH for the breast tumors may be useful for differential diagnosis of malignancy. The practice and the clinical applications of the FISH study are discussed.

  1. Simultaneous RNA-DNA FISH.

    Science.gov (United States)

    Lai, Lan-Tian; Meng, Zhenyu; Shao, Fangwei; Zhang, Li-Feng

    2016-01-01

    A highly useful tool for studying lncRNAs is simultaneous RNA-DNA FISH, which reveals the localization and quantitative information of RNA and DNA in cellular contexts. However, a simple combination of RNA FISH and DNA FISH often generates disappointing results because the fragile RNA signals are often damaged by the harsh conditions used in DNA FISH for denaturing the DNA. Here, we describe a robust and simple RNA-DNA FISH protocol, in which amino-labeled nucleic acid probes are used for RNA FISH. The method is suitable to detect single-RNA molecules simultaneously with DNA.

  2. Comparison of high resolution chromosome banding and fluorescence in situ hybridization (FISH) for the laboratory evaluation of Prader-Willi syndrome and Angelman syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Delach, J.A.; Rosengren, S.S.; Kaplan, L.; Greenstein, R.M.; Cassidy, S.B.; Benn, P.A.

    1994-08-01

    The development of probes containing segments of DNA from chromosome region 15q11-q13 provides the opportunity to confirm the diagnosis of Prader-Willi syndrome (PWS) and Angelman syndrome (AS) by fluorescence in situ hybridization (FISH). We have evaluated FISH studies and high resolution chromosome banding studies in 14 patients referred to confirm or rule out AS. In four patients (three from the PWS category and 1 from the AS group) chromosome analysis suggested that a deletion was present but FISH failed to confirm the finding. In one AS group patient, FISH identified a deletion not detectable by high resolution banding. Review of the clinical findings in the discrepant cases suggested that FISH results were correct and high resolution findings were erroneous. Studies with a chromosome 15 alpha satellite probe (D15Z) on both normal and abnormal individuals suggested that incorrect interpretation of chromosome banding may occasionally be attributable to alpha satellite polymorphism but other variation of 15q11-q13 chromosome bands also contributes to misinterpretation. We conclude that patients who have been reported to have a cytogenetic deletion of 15q11-q13 and who have clinical findings inconsistent with PWS and AS should be re-evaluated by molecular genetic techniques. 31 refs., 3 figs., 2 tabs.

  3. mRNA-targeted Fluorescent in Situ Hybridization (FISH) of Gram-negative Bacteria Without Template Amplification or Tyramide Signal Amplification

    Energy Technology Data Exchange (ETDEWEB)

    Coleman, James R.; Culley, David E.; Chrisler, William B.; Brockman, Fred J.

    2007-12-01

    Technologies are needed to study gene expression at the level of individual cells within a population or microbial community. Fluorescent in situ hybridization (FISH) supplies high-resolution spatial information and has been widely applied to study microbial communities at the rRNA level. While mRNA-targeted FISH has been popular for studying gene expression in eukaryotic cells, very little success has been achieved with prokaryotes. At present, detection of specific mRNAs in individual prokaryotic cells requires the use of in situ-RT-PCR or tyramide signal amplification (TSA). In this study we used DNA oligonucleotide probes labeled with a single near-infrared dye in FISH assays to detect multicopy plasmid-based and endogenous mRNA molecules in Escherichia coli and Shewanella oneidensis MR-1. We took advantage of the fact there is much less background signal produced by biological materials and support matrices in the near-infrared spectrum and thus long camera exposure times could be used. In addition, we demonstrate that a combination of probes targeting both rRNA and mRNA could be successfully employed within the same FISH assay. These results, as well as ongoing R&D improvements in NIR and infrared dyes, indicate the FISH approach we demonstrated could be applied in certain environmental settings to monitor gene expression in mixed populations.

  4. Numbers and locations of native bacteria on field-grown wheat roots quantified by fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Watt, Michelle; Hugenholtz, Philip; White, Rosemary; Vinall, Kerry

    2006-05-01

    Native bacteria, Pseudomonas and filamentous bacteria were quantified and localized on wheat roots grown in the field using fluorescence in situ hybridization (FISH). Seminal roots were sampled through the season from unploughed soil in a conservation farming system. Such soils are spatially heterogeneous, and many roots grow slowly through hard soil with cracks and pores containing dead roots remnant from previous crops. Root and rhizosphere morphology, and contact with soil particles were preserved, and autofluorescence was avoided by observing sections in the far-red with Cy5 and Cy5.5 fluorochromes. Spatial analyses showed that bacteria were embedded in a stable matrix (biofilm) within 11 microm of the root surface (range 2-30 microm) and were clustered on 40% of roots. Half the clusters co-located with axial grooves between epidermal cells, soil particles, cap cells or root hairs; the other half were not associated with visible features. Across all wheat roots, although variable, bacteria averaged 15.4 x 10(5) cells per mm(3) rhizosphere, and of these, Pseudomonas and filaments comprised 10% and 4%, respectively, with minor effects of sample time, and no effect of plant age. Root caps were most heavily colonized by bacteria along roots, and elongation zones least heavily colonized. Pseudomonas varied little with root development and were 17% of bacteria on the elongation zone. Filamentous bacteria were not found on the elongation zone. The most significant factor to rhizosphere populations along a wheat root, however, was contact with dead root remnants, where Pseudomonas were reduced but filaments increased to 57% of bacteria (P < 0.001). This corresponded with analyses of root remnants showing they were heavily colonized by bacteria, with 48% filaments (P < 0.001) and 1.4%Pseudomonas (P = 0.014). Efforts to manage rhizosphere bacteria for sustainable agricultural systems should continue to focus on root cap and mucilage chemistry, and remnant roots as

  5. 5-bp Classical Satellite DNA Loci from Chromosome-1 Instability in Cervical Neoplasia Detected by DNA Breakage Detection/Fluorescence in Situ Hybridization (DBD-FISH).

    Science.gov (United States)

    Cortés-Gutiérrez, Elva I; Ortíz-Hernández, Brenda L; Dávila-Rodríguez, Martha I; Cerda-Flores, Ricardo M; Fernández, José Luis; López-Fernández, Carmen; Gosálvez, Jaime

    2013-02-19

    We aimed to evaluate the association between the progressive stages of cervical neoplasia and DNA damage in 5-bp classical satellite DNA sequences from chromosome-1 in cervical epithelium and in peripheral blood lymphocytes using DNA breakage detection/fluorescence in situ hybridization (DBD-FISH). A hospital-based unmatched case-control study was conducted in 2011 with a sample of 30 women grouped according to disease stage and selected according to histological diagnosis; 10 with low-grade squamous intraepithelial lesions (LG-SIL), 10 with high-grade SIL (HG-SIL), and 10 with no cervical lesions, from the Unidad Medica de Alta Especialidad of The Mexican Social Security Institute, IMSS, Mexico. Specific chromosome damage levels in 5-bp classical satellite DNA sequences from chromosome-1 were evaluated in cervical epithelium and peripheral blood lymphocytes using the DBD-FISH technique. Whole-genome DNA hybridization was used as a reference for the level of damage. Results of Kruskal-Wallis test showed a significant increase according to neoplastic development in both tissues. The instability of 5-bp classical satellite DNA sequences from chromosome-1 was evidenced using chromosome-orientation FISH. In conclusion, we suggest that the progression to malignant transformation involves an increase in the instability of 5-bp classical satellite DNA sequences from chromosome-1.

  6. Fluorescence in situ hybridization and sequential catalysed reporter deposition (2C-FISH for the flow cytometric sorting of freshwater ultramicrobacteria

    Directory of Open Access Journals (Sweden)

    Stefan M Neuenschwander

    2015-03-01

    Full Text Available Flow cytometric sorting is a powerful tool to physically separate cells within mixed microbial communities. If combined with phylogenetic staining (fluorescence in situ hybridization, FISH it allows to specifically sort defined genotypic microbial populations from complex natural samples. However, the targeted enrichment of freshwater ultramicrobacteria, such as members of the LD12 clade of Alphaproteobacteria (SAR11-IIIb, is still challenging. Current FISH protocols, even in combination with signal amplification by catalysed reporter deposition (CARD, are not sufficiently sensitive for the distinction of these bacteria from background noise by flow cytometry, presumably due to their low ribosome content and small cell sizes. We, therefore, modified a CARD based flow sorting protocol with the aim of increasing its sensitivity to a level sufficient for ultramicrobacteria. This was achieved by a second signal amplification step mediated by horseradish peroxidase labelled antibodies targeted to the fluorophores that were previously deposited by CARD-FISH staining. The protocol was tested on samples from an oligo-mesotrophic lake. Ultramicrobacteria affiliated with LD12 Alphaproteobacteria could be successfully sorted to high purity by flow cytometry. The ratios of median fluorescence signal to background ranged around 20, and hybridization rates determined by flow cytometry were comparable to those obtained by fluorescence microscopy. Potential downstream applications of our modified cell staining approach range from the analysis of microdiversity within 16S rRNA-defined populations to that of functional properties, such as the taxon-specific incorporation rates of organic substrates.

  7. 5-bp Classical Satellite DNA Loci from Chromosome-1 Instability in Cervical Neoplasia Detected by DNA Breakage Detection/Fluorescence in Situ Hybridization (DBD-FISH)

    Science.gov (United States)

    Cortés-Gutiérrez, Elva I.; Ortíz-Hernández, Brenda L.; Dávila-Rodríguez, Martha I.; Cerda-Flores, Ricardo M; Fernández, José Luis; López-Fernández, Carmen; Gosálvez, Jaime

    2013-01-01

    We aimed to evaluate the association between the progressive stages of cervical neoplasia and DNA damage in 5-bp classical satellite DNA sequences from chromosome-1 in cervical epithelium and in peripheral blood lymphocytes using DNA breakage detection/fluorescence in situ hybridization (DBD-FISH). A hospital-based unmatched case-control study was conducted in 2011 with a sample of 30 women grouped according to disease stage and selected according to histological diagnosis; 10 with low-grade squamous intraepithelial lesions (LG-SIL), 10 with high-grade SIL (HG-SIL), and 10 with no cervical lesions, from the Unidad Medica de Alta Especialidad of The Mexican Social Security Institute, IMSS, Mexico. Specific chromosome damage levels in 5-bp classical satellite DNA sequences from chromosome-1 were evaluated in cervical epithelium and peripheral blood lymphocytes using the DBD-FISH technique. Whole-genome DNA hybridization was used as a reference for the level of damage. Results of Kruskal-Wallis test showed a significant increase according to neoplastic development in both tissues. The instability of 5-bp classical satellite DNA sequences from chromosome-1 was evidenced using chromosome-orientation FISH. In conclusion, we suggest that the progression to malignant transformation involves an increase in the instability of 5-bp classical satellite DNA sequences from chromosome-1. PMID:23429197

  8. 5-bp Classical Satellite DNA Loci from Chromosome-1 Instability in Cervical Neoplasia Detected by DNA Breakage Detection/Fluorescence in Situ Hybridization (DBD-FISH

    Directory of Open Access Journals (Sweden)

    Jaime Gosálvez

    2013-02-01

    Full Text Available We aimed to evaluate the association between the progressive stages of cervical neoplasia and DNA damage in 5-bp classical satellite DNA sequences from chromosome-1 in cervical epithelium and in peripheral blood lymphocytes using DNA breakage detection/fluorescence in situ hybridization (DBD-FISH. A hospital-based unmatched case-control study was conducted in 2011 with a sample of 30 women grouped according to disease stage and selected according to histological diagnosis; 10 with low-grade squamous intraepithelial lesions (LG-SIL, 10 with high-grade SIL (HG-SIL, and 10 with no cervical lesions, from the Unidad Medica de Alta Especialidad of The Mexican Social Security Institute, IMSS, Mexico. Specific chromosome damage levels in 5-bp classical satellite DNA sequences from chromosome-1 were evaluated in cervical epithelium and peripheral blood lymphocytes using the DBD-FISH technique. Whole-genome DNA hybridization was used as a reference for the level of damage. Results of Kruskal-Wallis test showed a significant increase according to neoplastic development in both tissues. The instability of 5-bp classical satellite DNA sequences from chromosome-1 was evidenced using chromosome-orientation FISH. In conclusion, we suggest that the progression to malignant transformation involves an increase in the instability of 5-bp classical satellite DNA sequences from chromosome-1.

  9. Nitrifying bacterial communities in an aquaculture wastewater treatment system using fluorescence in situ hybridization (FISH), 16S rRNA gene cloning, and phylogenetic analysis.

    Science.gov (United States)

    Paungfoo, Chanyarat; Prasertsan, Poonsuk; Burrell, Paul C; Intrasungkha, Nugul; Blackall, Linda L

    2007-07-01

    Aquaculture, especially shrimp farming, has played a major role in the growth of Thailand's economy in recent years, as well as in many South East Asian countries. However, the nutrient discharges from these activities have caused adverse impacts on the quality of the receiving waterways. In particular nitrogenous compounds, which may accumulate in aquaculture ponds, can be toxic to aquatic animals and cause environmental problems such as eutrophication. The mineralization process is well known, but certain aspects of the microbial ecology of nitrifiers, the microorganisms that convert ammonia to nitrate, are poorly understood. A previously reported enrichment of nitrifying bacteria (ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB)) from a shrimp farm inoculated in a sequencing batch reactor (SBR) was studied by molecular methods. The initial identification and partial quantification of the nitrifying bacteria (AOB and NOB) were carried out by fluorescence in situ hybridization (FISH) using previously published 16S rRNA-targeting oligonucleotide probes. The two dominant bacterial groups detected by FISH were from the Cytophaga-Flavobacterium-Bacteroides and Proteobacteria (beta subdivision) phyla. Published FISH probes for Nitrobacter and Nitrospira did not hybridize to any of the bacterial cells. Therefore it is likely that new communities of NOBs, differing from previously reported ones, exist in the enrichments. Molecular genetic techniques (cloning, sequencing, and phylogenetic analysis) targeting the 16S rRNA genes from the nitrifying enrichments were performed to identify putative AOBs and NOBs.

  10. Asexual Reproduction Does Not Apparently Increase the Rate of Chromosomal Evolution: Karyotype Stability in Diploid and Triploid Clonal Hybrid Fish (Cobitis, Cypriniformes, Teleostei).

    Science.gov (United States)

    Majtánová, Zuzana; Choleva, Lukáš; Symonová, Radka; Ráb, Petr; Kotusz, Jan; Pekárik, Ladislav; Janko, Karel

    2016-01-01

    Interspecific hybridization, polyploidization and transitions from sexuality to asexuality considerably affect organismal genomes. Especially the last mentioned process has been assumed to play a significant role in the initiation of chromosomal rearrangements, causing increased rates of karyotype evolution. We used cytogenetic analysis and molecular dating of cladogenetic events to compare the rate of changes of chromosome morphology and karyotype in asexually and sexually reproducing counterparts in European spined loach fish (Cobitis). We studied metaphases of three sexually reproducing species and their diploid and polyploid hybrid clones of different age of origin. The material includes artificial F1 hybrid strains, representatives of lineage originated in Holocene epoch, and also individuals of an oldest known age to date (roughly 0.37 MYA). Thereafter we applied GISH technique as a marker to differentiate parental chromosomal sets in hybrids. Although the sexual species accumulated remarkable chromosomal rearrangements after their speciation, we observed no differences in chromosome numbers and/or morphology among karyotypes of asexual hybrids. These hybrids possess chromosome sets originating from respective parental species with no cytogenetically detectable recombinations, suggesting their integrity even in a long term. The switch to asexual reproduction thus did not provoke any significant acceleration of the rate of chromosomal evolution in Cobitis. Asexual animals described in other case studies reproduce ameiotically, while Cobitis hybrids described here produce eggs likely through modified meiosis. Therefore, our findings indicate that the effect of asexuality on the rate of chromosomal change may be context-dependent rather than universal and related to particular type of asexual reproduction.

  11. Magnetic Fe3O4/Ag Hybrid Nanoparticles as Surface-Enhanced Raman Scattering Substrate for Trace Analysis of Furazolidone in Fish Feeds

    Directory of Open Access Journals (Sweden)

    Wansong Yu

    2014-01-01

    Full Text Available Nanoparticles (NPs composed of ferromagnetic and noble metal materials show dual functions of magnetic activity and local surface plasmon response and have great potential as substrates for surface-enhanced Raman scattering (SERS in trace analysis. Easy-to-prepare superparamagnetic Fe3O4/Ag hybrid NPs were synthesized and optimized by adjusting the ratio of silver particles aggregated with APTMS-modified Fe3O4 NPs. The hybrid NPs were assembled under an external magnetic field before being used as substrate for SERS analysis. The SERS spectral features of furazolidone standard solution were clearly identified at concentrations as low as 40 ng mL−1, and furazolidone in fish feeds could be detected at 500 ng g−1. The results indicated that the Fe3O4/Ag hybrid NPs as SERS substrates had a great potential for detection of trace amount of furazolidone and other prohibited or restricted antibiotics in the animal and fish feeds.

  12. Combining M-FISH and Quantum Dot technology for fast chromosomal assignment of transgenic insertions

    Directory of Open Access Journals (Sweden)

    Yusuf Mohammed

    2011-12-01

    Full Text Available Abstract Background Physical mapping of transgenic insertions by Fluorescence in situ Hybridization (FISH is a reliable and cost-effective technique. Chromosomal assignment is commonly achieved either by concurrent G-banding or by a multi-color FISH approach consisting of iteratively co-hybridizing the transgenic sequence of interest with one or more chromosome-specific probes at a time, until the location of the transgenic insertion is identified. Results Here we report a technical development for fast chromosomal assignment of transgenic insertions at the single cell level in mouse and rat models. This comprises a simplified 'single denaturation mixed hybridization' procedure that combines multi-color karyotyping by Multiplex FISH (M-FISH, for simultaneous and unambiguous identification of all chromosomes at once, and the use of a Quantum Dot (QD conjugate for the transgene detection. Conclusions Although the exploitation of the unique optical properties of QD nanocrystals, such as photo-stability and brightness, to improve FISH performance generally has been previously investigated, to our knowledge this is the first report of a purpose-designed molecular cytogenetic protocol in which the combined use of QDs and standard organic fluorophores is specifically tailored to assist gene transfer technology.

  13. Chemical shift prediction for denatured proteins

    Energy Technology Data Exchange (ETDEWEB)

    Prestegard, James H., E-mail: jpresteg@ccrc.uga.edu; Sahu, Sarata C.; Nkari, Wendy K.; Morris, Laura C.; Live, David; Gruta, Christian

    2013-02-15

    While chemical shift prediction has played an important role in aspects of protein NMR that include identification of secondary structure, generation of torsion angle constraints for structure determination, and assignment of resonances in spectra of intrinsically disordered proteins, interest has arisen more recently in using it in alternate assignment strategies for crosspeaks in {sup 1}H-{sup 15}N HSQC spectra of sparsely labeled proteins. One such approach involves correlation of crosspeaks in the spectrum of the native protein with those observed in the spectrum of the denatured protein, followed by assignment of the peaks in the latter spectrum. As in the case of disordered proteins, predicted chemical shifts can aid in these assignments. Some previously developed empirical formulas for chemical shift prediction have depended on basis data sets of 20 pentapeptides. In each case the central residue was varied among the 20 amino common acids, with the flanking residues held constant throughout the given series. However, previous choices of solvent conditions and flanking residues make the parameters in these formulas less than ideal for general application to denatured proteins. Here, we report {sup 1}H and {sup 15}N shifts for a set of alanine based pentapeptides under the low pH urea denaturing conditions that are more appropriate for sparse label assignments. New parameters have been derived and a Perl script was created to facilitate comparison with other parameter sets. A small, but significant, improvement in shift predictions for denatured ubiquitin is demonstrated.

  14. Rousseau's Philosophy of Transformative, "Denaturing" Education

    Science.gov (United States)

    Riley, Patrick

    2011-01-01

    Rousseau's political philosophy presents the great legislator as a civic educator who must over time transform naturally self-loving egoists into citizens animated by a general will without destroying freedom. This is an educational process which is "denaturing" but which aims to produce autonomous adults who can ultimately say to their teacher…

  15. speciation through asexuality in fish: postzygotic reproductive isolation may be completed in spite of fertility of hybrids

    Directory of Open Access Journals (Sweden)

    Karel Janko

    2015-12-01

    Altogether, it appears that initiation of hybrid asexuality and the completion of speciation process through formation of postRIMs are interconnected phenomena. Both processes are linked to the genetic divergence of hybridizing taxa: initially, hybridization between little diverged species leads to recombinant and fertile hybrids allowing intensive gene flow. As the hybridizing taxa continue to diverge, clonally reproducing hybrid females and sterile males become dominant and the gene flow ceases. The speciation may therefore be completed through asexuality of hybrids The work was supported by grant no. 13-12580S provided by the Czech Science Foundation (www.gacr.cz. Further support was provided by the Academy of Sciences of the Czech Republic (www.cas.cz by the grant no. RVO 67985904

  16. Comparison of immunohistochemistry (IHC and fluorescence in situ hybridization (FISH assessment for Her-2 status in breast cancer

    Directory of Open Access Journals (Sweden)

    Qi Minfang

    2009-11-01

    Full Text Available Abstract Background The concordance rate between IHC and FISH according to clinical performance is still controversial. We report a prospective study to reflect the concordance between IHC and FISH in Guilin city, People's Republic of China. Methods Fifty cases of invasive ductal carcinoma of breast tested by IHC and scored as 0, 1+, 2+ and 3+ by pathologists were further analyzed by FISH using a commercially available double-color probe, and the FISH findings were compared with IHC test results. Results A total concordance of 82.0% was observed with a Kappa coefficient of 0.640 (P Conclusion The IHC can be used firstly to screen the HER-2 status, and FISH can be used as a supplementary role to IHC and 2+ and some negative cases. And only those cases with Her-2 status of IHC 3+ or FISH positive should be treated with Herceptin.

  17. Fluorescence in situ hybridization (FISH using non-commercial probes in the diagnosis of clinically suspected microdeletion syndromes

    Directory of Open Access Journals (Sweden)

    Ashutosh Halder

    2013-01-01

    Interpretation & conclusion: We conclude that FISH should not be the method of choice for clinically suspected microdeletion syndromes. We propose to follow strict clinical criteria for FISH testing or preferably to follow better methods (genotype first approach. Whole genome screening may be used as first line of test and FISH may be used for confirmation of screening result, screening of family members and prenatal diagnosis.

  18. A populational survey of 45S rDNA polymorphism in the Jefferson salamander Ambystoma jeffersonianum revealed by fluorescence in situ hybridization (FISH)

    Institute of Scientific and Technical Information of China (English)

    Ke BI; James P.BOGART; Jinzhong FU

    2009-01-01

    The chromosomal localization of 45S ribosomal RNA genes in Ambystoma jeffersonianum was determined by fluorescence in situ hybridization with 18S rDNA fragment as a probe (FISH-rDNA). Our results revealed the presence of rDNA polymorphism among A.jeffersonianum populations in terms of number, location and FISH signal intensity on the chromosomes. Nine rDNA cytotypes were found in ten geographically isolated populations and most of them contained derivative rDNA sites. Our preliminary study provides strong indication of karyotypic diversification of A.jeffersonianum that is demonstrated by intraspecific variation of 45S rDNA cytotypes. rDNA cytotype polymorphism has been described in many other caudate amphibians. We predict that habitat isolation, low dispersal ability and decline of effective population size could facilitate the fixation and accumulation of variable rDNA cytotypes during their chromosome evolution [Current Zoology 55(2):145-149,2009].

  19. A populational survey of 45S rDNA polymorphism in the Jefferson salamander Ambystoma jeffersonianum revealed by fluorescence in situ hybridization (FISH

    Directory of Open Access Journals (Sweden)

    Jinzhong FU

    2009-04-01

    Full Text Available The chromosomal localization of 45S ribosomal RNA genes in Ambystoma jeffersonianum was determined by fluorescence in situ hybridization with 18S rDNA fragment as a probe (FISH-rDNA. Our results revealed the presence of rDNA polymorphism among A.jeffersonianum populations in terms of number, location and FISH signal intensity on the chromosomes. Nine rDNA cytotypes were found in ten geographically isolated populations and most of them contained derivative rDNA sites. Our preliminary study provides strong indication of karyotypic diversification of A.jeffersonianum that is demonstrated by intraspecific variation of 45S rDNA cytotypes. rDNA cytotype polymorphism has been described in many other caudate amphibians. We predict that habitat isolation, low dispersal ability and decline of effective population size could facilitate the fixation and accumulation of variable rDNA cytotypes during their chromosome evolution.

  20. Effects of the acute exposition to glyphosate-based herbicide on oxidative stress parameters and antioxidant responses in a hybrid Amazon fish surubim (Pseudoplatystoma sp).

    Science.gov (United States)

    Sinhorin, Valéria Dornelles Gindri; Sinhorin, Adilson Paulo; Teixeira, Jhonnes Marcos dos Santos; Miléski, Kelly Márcia Lazarotto; Hansen, Paula Carine; Moreira, Paula Sueli Andrade; Kawashita, Nair Honda; Baviera, Amanda Martins; Loro, Vania Lúcia

    2014-08-01

    The aim of this study was to investigate the effects of acute glyphosate (active ingredient) exposure on the oxidative stress biomarkers and antioxidant defenses of a hybrid surubim (Pseudoplatystoma sp). The fish were exposed to different herbicide concentrations for 96 h. The thiobarbituric acid-reactive substances (TBARS), protein carbonyls and antioxidant responses were verified. The 15 mg a.pL(-1) of herbicide resulted in the death of 50% of the fish after 96 h. An increase in liver and muscle TBARS levels was observed when fish were exposed to the herbicide. The protein carbonyl content was also increased in the liver (4.5mg a.pL(-1) concentration) and brain (2.25 mg a.pL(-1) concentration). The antioxidant activities decreased in the liver and brain after exposure to herbicide. Levels of ascorbic acid in the liver (2.25 mg a.pL(-1) and 4.5 mg a.pL(-1) concentrations) and brain (2.25 mg a.pL(-1) concentration) were increased post-treatment. Levels of total thiols were increased in the liver and brain (2.25 mg L(-1) and 7.5mg a.pL(-1), respectively). Glyphosate exposure, at the tested concentrations affects surubim health by promoting changes that can affect their survival in natural environment. Some parameters as TBARS and protein carbonyl could be early biomarkers for Roundup exposure in this fish species.

  1. Fluorescence in situ hybridization (FISH screening for the 22q11.2 deletion in patients with clinical features of velocardiofacial syndrome but without cardiac anomalies

    Directory of Open Access Journals (Sweden)

    Paula Sandrin-Garcia

    2007-01-01

    Full Text Available The velocardiofacial syndrome (VCFS, a condition associated with 22q11.2 deletions, is characterized by a typical facies, palatal anomalies, learning disabilities, behavioral disturbances and cardiac defects. We investigated the frequency of these chromosomal deletions in 16 individuals with VCFS features who presented no cardiac anomalies, one of the main characteristics of VCFS. Fluorescent in situ hybridization (FISH with the N25 (D22S75; 22q11.2 probe revealed deletions in ten individuals (62%. Therefore, even in the absence of cardiac anomalies testing for the 22q11.2 microdeletions in individuals showing other clinical features of this syndrome is recommended.

  2. Guanidinium-induced denaturation by breaking of salt bridges

    NARCIS (Netherlands)

    Meuzelaar, H.; Panman, M.R.; Woutersen, S.

    2015-01-01

    Despite its wide use as a denaturant, the mechanism by which guanidinium (Gdm+) induces protein unfolding remains largely unclear. Herein, we show evidence that Gdm+ can induce denaturation by disrupting salt bridges that stabilize the folded conformation. We study the Gdm+-​induced denaturation of

  3. Hysteresis in pressure-driven DNA denaturation.

    Directory of Open Access Journals (Sweden)

    Enrique Hernández-Lemus

    Full Text Available In the past, a great deal of attention has been drawn to thermal driven denaturation processes. In recent years, however, the discovery of stress-induced denaturation, observed at the one-molecule level, has revealed new insights into the complex phenomena involved in the thermo-mechanics of DNA function. Understanding the effect of local pressure variations in DNA stability is thus an appealing topic. Such processes as cellular stress, dehydration, and changes in the ionic strength of the medium could explain local pressure changes that will affect the molecular mechanics of DNA and hence its stability. In this work, a theory that accounts for hysteresis in pressure-driven DNA denaturation is proposed. We here combine an irreversible thermodynamic approach with an equation of state based on the Poisson-Boltzmann cell model. The latter one provides a good description of the osmotic pressure over a wide range of DNA concentrations. The resulting theoretical framework predicts, in general, the process of denaturation and, in particular, hysteresis curves for a DNA sequence in terms of system parameters such as salt concentration, density of DNA molecules and temperature in addition to structural and configurational states of DNA. Furthermore, this formalism can be naturally extended to more complex situations, for example, in cases where the host medium is made up of asymmetric salts or in the description of the (helical-like charge distribution along the DNA molecule. Moreover, since this study incorporates the effect of pressure through a thermodynamic analysis, much of what is known from temperature-driven experiments will shed light on the pressure-induced melting issue.

  4. Hybridization leads to sensory repertoire expansion in a gynogenetic fish, the Amazon molly (poecilia formosa): a test of the hybrid-sensory expansion hypothesis.

    Science.gov (United States)

    Sandkam, Benjamin A; Joy, Jeffrey B; Watson, Corey T; Gonzalez-Bendiksen, Pablo; Gabor, Caitlin R; Breden, Felix

    2013-01-01

    Expansions in sensory systems usually require processes such as gene duplication and divergence, and thus evolve slowly. We evaluate a novel mechanism leading to rapid sensory repertoire expansion: hybrid-sensory expansion (HSE). HSE occurs when two species with differently tuned sensory systems form a hybrid, bringing together alleles from each of the parental species. In one generation, a sensory repertoire is created that is the sum of the variance between parental species. The Amazon molly presents a unique opportunity to test the HSE hypothesis in a "frozen" hybrid. We compared opsin sequences of the Amazon molly, Poecilia formosa, to those of the parental species. Both parental species are homozygous at the RH2-1 locus and each of the four long wavelength sensitive loci, while P. formosa possess two different alleles at these loci; one matching each parental allele. Gene expression analysis showed P. formosa use the expanded opsin repertoire that was the result of HSE. Additionally, behavioral tests revealed P. formosa respond to colored stimuli in a manner similar or intermediate to the parental species P. mexicana and P. latipinna. Together these results strongly support the HSE hypothesis. Hybrid-sensory repertoire expansion is likely important in other hybrid species and in other sensory systems.

  5. Simultaneous 16S and 18S rRNA fluorescence in situ hybridization (FISH) on LR White sections demonstrated in Vestimentifera (Siboglinidae) tubeworms.

    Science.gov (United States)

    Schimak, Mario P; Toenshoff, Elena R; Bright, Monika

    2012-02-01

    Traditional morphological identification of invertebrate marine species is limited in early life history stages for many taxa. In this study, we demonstrate, by example of Vestimentiferan tubeworms (Siboglinidae, Polychaeta), that the simultaneous fluorescence in situ hybridization (FISH) of both eukaryotic host and bacterial symbiont cells is possible on a single semi-thin (1 μm) section. This allows the identification of host specimens to species level as well as offering visualization of bacteria distributed within the host tissue. Previously published 18S rRNA host-specific oligonucleotide probes for Riftia pachyptila, Tevnia jerichonana and a newly designed Oasisia alvinae probe, as well as a 16S rRNA probe targeting symbionts found in all host species, were applied. A number of standard fixation and hybridization parameters were tested and optimized for the best possible signal intensity and cellular resolution. Ethanol conserved samples embedded in LR White low viscosity resin yielded the best results with regard to both signal intensity and resolution. We show that extended storage times of specimens does not affect the quality of signals attained by FISH and use our protocol to identify morphologically unidentifiable tubeworm individuals from a small data set, conforming to previous findings in succession studies of the Siboglinidae family.

  6. Validation of microsatellite multiplexes for parentage analysis and species discrimination in two hybridizing species of coral reef fish (Plectropomus spp., Serranidae).

    Science.gov (United States)

    Harrison, Hugo B; Feldheim, Kevin A; Jones, Geoffrey P; Ma, Kayan; Mansour, Hicham; Perumal, Sadhasivam; Williamson, David H; Berumen, Michael L

    2014-06-01

    Microsatellites are often considered ideal markers to investigate ecological processes in animal populations. They are regularly used as genetic barcodes to identify species, individuals, and infer familial relationships. However, such applications are highly sensitive the number and diversity of microsatellite markers, which are also prone to error. Here, we propose a novel framework to assess the suitability of microsatellite datasets for parentage analysis and species discrimination in two closely related species of coral reef fish, Plectropomus leopardus and P. maculatus (Serranidae). Coral trout are important fisheries species throughout the Indo-Pacific region and have been shown to hybridize in parts of the Great Barrier Reef, Australia. We first describe the development of 25 microsatellite loci and their integration to three multiplex PCRs that co-amplify in both species. Using simulations, we demonstrate that the complete suite of markers provides appropriate power to discriminate between species, detect hybrid individuals, and resolve parent-offspring relationships in natural populations, with over 99.6% accuracy in parent-offspring assignments. The markers were also tested on seven additional species within the Plectropomus genus with polymorphism in 28-96% of loci. The multiplex PCRs developed here provide a reliable and cost-effective strategy to investigate evolutionary and ecological dynamics and will be broadly applicable in studies of wild populations and aquaculture brood stocks for these closely related fish species.

  7. Validation of microsatellite multiplexes for parentage analysis and species discrimination in two hybridizing species of coral reef fish (Plectropomus spp., Serranidae)

    KAUST Repository

    Harrison, H.B.

    2014-04-24

    Microsatellites are often considered ideal markers to investigate ecological processes in animal populations. They are regularly used as genetic barcodes to identify species, individuals, and infer familial relationships. However, such applications are highly sensitive the number and diversity of microsatellite markers, which are also prone to error. Here, we propose a novel framework to assess the suitability of microsatellite datasets for parentage analysis and species discrimination in two closely related species of coral reef fish, Plectropomus leopardus and P. maculatus (Serranidae). Coral trout are important fisheries species throughout the Indo-Pacific region and have been shown to hybridize in parts of the Great Barrier Reef, Australia. We first describe the development of 25 microsatellite loci and their integration to three multiplex PCRs that co-amplify in both species. Using simulations, we demonstrate that the complete suite of markers provides appropriate power to discriminate between species, detect hybrid individuals, and resolve parent-offspring relationships in natural populations, with over 99.6% accuracy in parent-offspring assignments. The markers were also tested on seven additional species within the Plectropomus genus with polymorphism in 28-96% of loci. The multiplex PCRs developed here provide a reliable and cost-effective strategy to investigate evolutionary and ecological dynamics and will be broadly applicable in studies of wild populations and aquaculture brood stocks for these closely related fish species. 2014 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.

  8. Application and analysis of LNG cold energy for hybrid fishing vessel%混合动力渔船LNG冷量的应用分析

    Institute of Scientific and Technical Information of China (English)

    魏鹏威; 张建一; 詹鋆

    2012-01-01

    液化天然气(LNC)是清洁能源,同时又蕴含大量冷量,我国已经提出渔船推广使用LNG作为动力燃料.目前,渔船制冷能耗占全船能耗30%-40%.为了确定混合动力渔船中LNG的应用优势,本文结合实际生产中的50m型围网渔船进行了计算分析.结果显示,在该型渔船上采用柴油/天然气混合动力,替油率80%时每年可节省燃料费用61万元.将LNG冷量用于冷海水系统,每天可制取0℃冷海水10.4t,减少冷海水制冷系统工作时长1.6h,部分冷量用于系统的过冷循环,可使系统COP提高5.2%.分析表明,利用混合动力渔船的LNG冷能可显著节能,降低营运成本.%Liquified Natural Gas (LNG) is a clean energy with a lot of cold energy.Its use as engine fuel for fishing vessel is promoted currently in China.The energy consumption of refrigeration system in fishing vessels has accounted for 30% to 40% in total energy consumption.In order to determine the advantage of LNG applications in hybrid fishing vessel,some calculation and analysis are conducted based on 50 m model purse seine vessel.The results show that 610 thousands yuan can be saved annually if 80% of diesel oil is replaced by LNG on this type of fishing vessels.Meanwhile,10.4 t cold seawater in 0℃ can be produced and 1.6 hour of the system's working time can be reduced each day by use of the LNG cold energy.The COP can be improved by 5.2% when the surplus LNG cold energy is used for subcooled cycle of the refrigeration system.Energy cost can be significantly reduced by use of LNG cold energy in the hybrid fishing vessels based on the analysis.

  9. Fluorescence In Situ Hybridization (FISH-Based Karyotyping Reveals Rapid Evolution of Centromeric and Subtelomeric Repeats in Common Bean (Phaseolus vulgaris and Relatives

    Directory of Open Access Journals (Sweden)

    Aiko Iwata-Otsubo

    2016-04-01

    Full Text Available Fluorescence in situ hybridization (FISH-based karyotyping is a powerful cytogenetics tool to study chromosome organization, behavior, and chromosome evolution. Here, we developed a FISH-based karyotyping system using a probe mixture comprised of centromeric and subtelomeric satellite repeats, 5S rDNA, and chromosome-specific BAC clones in common bean, which enables one to unambiguously distinguish all 11 chromosome pairs. Furthermore, we applied the karyotyping system to several wild relatives and landraces of common bean from two distinct gene pools, as well as other related Phaseolus species, to investigate repeat evolution in the genus Phaseolus. Comparison of karyotype maps within common bean indicates that chromosomal distribution of the centromeric and subtelomeric satellite repeats is stable, whereas the copy number of the repeats was variable, indicating rapid amplification/reduction of the repeats in specific genomic regions. In Phaseolus species that diverged approximately 2–4 million yr ago, copy numbers of centromeric repeats were largely reduced or diverged, and chromosomal distributions have changed, suggesting rapid evolution of centromeric repeats. We also detected variation in the distribution pattern of subtelomeric repeats in Phaseolus species. The FISH-based karyotyping system revealed that satellite repeats are actively and rapidly evolving, forming genomic features unique to individual common bean accessions and Phaseolus species.

  10. DSC study of denaturation of β-lactoglobulin B

    Institute of Scientific and Technical Information of China (English)

    王邦宁; 谈夫

    1995-01-01

    The denaturation of bovine β-lactoglobulin B (β-Lg B) has been studied in phosphate solutions with various concentrations of GuHCl with differential scanning calorimetry The experiments demonstrated that the presence of GuHCl made the β-Lg B undergo both cold denaturation and heat denaturation under the condition of a high concentration of the protein. The enthalpy changes of both kinds of denaturation exhibit opposite signs. Both the cold denaturation and the renaturation of the protein are reproducible, but its heat denaturation is irreversible. The cooperation among monomer molecules of the protein is involved in its heat denaturation The heat denaturation of the protein can be represented by the thermodynamic model Nc D→F. The activation energy of heat denaturation is 285 kJ/mol, which imples that the depression of temperature and enthalpy of heat denaturation of the P-Lg B does not result from decreasing considerably the activation energy by GuHCl As for the cold denaturation of the protein, es

  11. Behavior of meiotic chromosomes in Pinus wallichiana, P. strobus and their hybrid and nrDNA localization in pollen mother cells of the hybrid by using FISH.

    Science.gov (United States)

    Deng, Hui-Sheng; Zhang, Da-Ming; Fu, Cheng-Xin; Hong, De-Yuan

    2008-03-01

    The complete process of meiosis was investigated in Pinus wallichiana, P. strobus and their artificial hybrid (F1) using microsporocytes. It is revealed that there were slightly lower chiasma frequency, lower ring bivalent frequency, lower meiotic index and distinctly higher frequency of aberrance (chromosomal bridges, fragments or micronuclei) in pollen mother cells (PMCs) of the hybrid (F1) than those of the parental species, which showed a certain degree of differentiation between homologous chromosomes of the two parents. However, relatively higher frequency of ring bivalents and higher meiotic index in all the three entities indicate the great stability of genomes of parental species, and the differentiation of genomes between the two parents must have been slight. Total nineteen signal loci of 18S rDNA were observed in nine bivalents of the hybrid (F1), among which one bivalent bears two loci, while the others have only one. It is suggested that distinct differentiation at genetic level existed in homologous chromosomes of the two parental species, whereas only slight differentiation at karyotypic and genomic levels take place between the parent species.

  12. Behavior of Meiotic Chromosomes in Pinus wallichiana, P. strobus and Their Hybrid and nrDNA Localization in Pollen Mother Cells of the Hybrid by Using FISH

    Institute of Scientific and Technical Information of China (English)

    Hui-Sheng Deng; Da-Ming Zhang; Cheng-Xin Fu; De-Yuan Hong

    2008-01-01

    The complete process of meiosis was investigated in Pinus wallichiana, P. strobus and their artificial hybrid (F1) using microsporocytes. It is revealed that there were slightly lower chiasma frequency, lower ring bivalent frequency, lower meiotic Index and distinctly higher frequency of aberrance (chromosomal bridges, fragments or micronuclei) in pollen mother cells (PMCs) of the hybrid (F1) than those of the parental species, which showed a certain degree of differentiation between homologous chromosomes of the two parents. However, relatively higher frequency of ring bivalents and higher meiotic index in all the three entities indicate the great stability of genomes of parental species, and the differentiation of genomes between the two parents must have been slight. Total nineteen signal loci of 18S rDNA were observed in nine bivalents of the hybrid (F1), among which one bivalent bears two loci, while the others have only one. It is suggested that distinct differentiation at genetic level existed in homologous chromosomes of the two parental species, whereas only slight differentiation at karyotypic and genomic levels take place between the parent species.

  13. Supercoiling induces denaturation bubbles in circular DNA.

    Science.gov (United States)

    Jeon, Jae-Hyung; Adamcik, Jozef; Dietler, Giovanni; Metzler, Ralf

    2010-11-12

    We present a theoretical framework for the thermodynamic properties of supercoiling-induced denaturation bubbles in circular double-stranded DNA molecules. We explore how DNA supercoiling, ambient salt concentration, and sequence heterogeneity impact on the bubble occurrence. An analytical derivation of the probability distribution to find multiple bubbles is derived and the relevance for supercoiled DNA discussed. We show that in vivo sustained DNA bubbles are likely to occur due to partial twist release in regions rich in weaker AT base pairs. Single DNA plasmid imaging experiments clearly demonstrate the existence of bubbles in free solution.

  14. 27 CFR 21.151 - List of denaturants authorized for denatured spirits.

    Science.gov (United States)

    2010-04-01

    ... TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS FORMULAS FOR DENATURED ALCOHOL AND RUM... (Glycerol), U.S.P S.D.A. 31-A. Green soap, U.S.P S.D.A. 27-B. Guaiacol, N.F.X S.D.A. 38-B. Heptane C.D.A....

  15. 78 FR 38628 - Reclassification of Specially Denatured Spirits and Completely Denatured Alcohol Formulas and...

    Science.gov (United States)

    2013-06-27

    ... alcohol. 13-A 10 gallons of ethyl ether. 19 100 gallons of ethyl ether. 23-A 8 gallons of acetone, U.S.P... alcohol. 32 5 gallons of ethyl ether. 35-A 4.25 gallons of ethyl acetate having an ester content of 100... regulations regarding the production, warehousing, denaturing, distribution, sale, export, and use...

  16. Partially folded intermediates during trypsinogen denaturation

    Directory of Open Access Journals (Sweden)

    Martins N.F.

    1999-01-01

    Full Text Available The equilibrium unfolding of bovine trypsinogen was studied by circular dichroism, differential spectra and size exclusion HPLC. The change in free energy of denaturation was = 6.99 ± 1.40 kcal/mol for guanidine hydrochloride and = 6.37 ± 0.57 kcal/mol for urea. Satisfactory fits of equilibrium unfolding transitions required a three-state model involving an intermediate in addition to the native and unfolded forms. Size exclusion HPLC allowed the detection of an intermediate population of trypsinogen whose Stokes radii varied from 24.1 ± 0.4 Å to 26.0 ± 0.3 Å for 1.5 M and 2.5 M guanidine hydrochloride, respectively. During urea denaturation, the range of Stokes radii varied from 23.9 ± 0.3 Å to 25.7 ± 0.6 Å for 4.0 M and 6.0 M urea, respectively. Maximal intrinsic fluorescence was observed at about 3.8 M urea with 8-aniline-1-naphthalene sulfonate (ANS binding. These experimental data indicate that the unfolding of bovine trypsinogen is not a simple transition and suggest that the equilibrium intermediate population comprises one intermediate that may be characterized as a molten globule. To obtain further insight by studying intermediates representing different stages of unfolding, we hope to gain a better understanding of the complex interrelations between protein conformation and energetics.

  17. Apparent digestibility of Asian carp and common carp-derived fish meals in feeds for hybrid striped bass Morone saxatilis female x M. chrysops male and rainbow trout Oncorhynchus mykiss

    Science.gov (United States)

    Apparent digestibility coefficients (ADCs) of nutrients (crude protein, amino acids, crude lipid, fatty acids, and minerals) were determined for fish meals derived from menhaden, Asian carp (combination of silver and bighead carps), and common carp in feeds for hybrid striped bass and rainbow trout....

  18. Fully Automated Fluorescent in situ Hybridization (FISH) Staining and Digital Analysis of HER2 in Breast Cancer : A Validation Study

    NARCIS (Netherlands)

    van der Logt, Elise M. J.; Kuperus, Deborah A. J.; van Setten, Jan W.; van den Heuvel, Marius C.; Boers, James. E.; Schuuring, Ed; Kibbelaar, Robby E.

    2015-01-01

    HER2 assessment is routinely used to select patients with invasive breast cancer that might benefit from HER2-targeted therapy. The aim of this study was to validate a fully automated in situ hybridization (ISH) procedure that combines the automated Leica HER2 fluorescent ISH system for Bond with su

  19. Comprehensive meiotic segregation analysis of a 4-breakpoint t(1;3;6) complex chromosome rearrangement using single sperm array comparative genomic hybridization and FISH.

    Science.gov (United States)

    Hornak, Miroslav; Vozdova, Miluse; Musilova, Petra; Prinosilova, Petra; Oracova, Eva; Linkova, Vlasta; Vesela, Katerina; Rubes, Jiri

    2014-10-01

    Complex chromosomal rearrangements (CCR) represent rare structural chromosome abnormalities frequently associated with infertility. In this study, meiotic segregation in spermatozoa of an infertile normospermic carrier of a 4-breakpoint t(1;3;6) CCR was analysed. A newly developed array comparative genomic hybridization protocol was used, and all chromosomes in 50 single sperm cells were simultaneously examined. Three-colour FISH was used to analyse chromosome segregation in 1557 other single sperm cells. It was also used to measure an interchromosomal effect; sperm chromatin structure assay was used to measure chromatin integrity. A high-frequency of unbalanced spermatozoa (84%) was observed, mostly arising from the 3:3 symmetrical segregation mode. Array comparative genomic hybridization was used to detect additional aneuploidies in two out of 50 spermatozoa (4%) in chromosomes not involved in the complex chromosome rearrangement. Significantly increased rates of diploidy and XY disomy were found in the CCR carrier compared with the control group (P < 0.001). Defective condensation of sperm chromatin was also found in 22.7% of spermatozoa by sperm chromatin structure assay. The results indicate that the infertility in the man with CCR and normal spermatozoa was caused by a production of chromosomally unbalanced, XY disomic and diploid spermatozoa and spermatozoa with defective chromatin condensation.

  20. Can A Denaturant Stabilize DNA? Pyridine Reverses DNA Denaturation in Acidic pH.

    Science.gov (United States)

    Portella, Guillem; Terrazas, Montserrat; Villegas, Núria; González, Carlos; Orozco, Modesto

    2015-09-01

    The stability of DNA is highly dependent on the properties of the surrounding solvent, such as ionic strength, pH, and the presence of denaturants and osmolytes. Addition of pyridine is known to unfold DNA by replacing π-π stacking interactions between bases, stabilizing conformations in which the nucleotides are solvent exposed. We show here experimental and theoretical evidences that pyridine can change its role and in fact stabilize the DNA under acidic conditions. NMR spectroscopy and MD simulations demonstrate that the reversal in the denaturing role of pyridine is specific, and is related to its character as pseudo groove binder. The present study sheds light on the nature of DNA stability and on the relationship between DNA and solvent, with clear biotechnological implications.

  1. Phenotypic consequences of a mosaic marker chromosome identified by fluorescence in situ hybridization (FISH) as being derived from chromosome 16

    Energy Technology Data Exchange (ETDEWEB)

    Ray, J.H.; Zhou, X.; Pletcher, B.A. [Cornell Univ. Medical College, Manhasset, NY (United States)] [and others

    1994-09-01

    De novo marker chromosomes are detected in 1 in 2500 amniotic fluid samples and are associated with a 10-15% risk for phenotypic abnormality. FISH can be utilized as a research tool to identify the origins of marker chromosomes. The phenotypic consequences of a marker chromosome derived from the short arm of chromosome 16 are described. A 26-year-old woman underwent amniocentesis at 28 weeks gestation because of a prenatally diagnosed tetralogy of Fallot. Follow-up ultrasounds also showed ventriculomegaly and cleft lip and palate. 32 of 45 cells had the karyotype 47,XY,+mar; the remaining cells were 46,XY. The de novo marker chromosome was C-band positive and non-satellited and failed to stain with distamycin A/DAPI. At birth the ultrasound findings were confirmed and dysmorphic features and cryptorchidism were noted. Although a newborn blood sample contained only normal cells, mosaicism was confirmed in 2 skin biopsies. FISH using whole-chromosome painting and alpha-satellite DNA probes showed that the marker chromosome had originated from chromosome 16. As proximal 16q is distamycin A/DAPI positive, the marker is apparently derived from proximal 16p. At 15 months of age, this child is hypotonic, globally delayed and is gavage-fed. His physical examination is significant for microbrachycephaly, a round face, sparse scalp hair, ocular hypertelorism, exotropia, a flat, wide nasal bridge and tip, mild micrognathia, and tapered fingers with lymphedema of hands and feet. Inguinal hernias have been repaired. His features are consistent with those described for patients trisomic for most or all of the short arm of chromosome 16. Marker chromosomes derived from the short arm of chromosome 16 appear to have phenotypic consequences. As the origin of more marker chromosomes are identified using FISH, their karyotype/phenotype correlations will become more apparent, which will permit more accurate genetic counseling.

  2. Kinetically controlled refolding of a heat-denatured hyperthermostable protein

    NARCIS (Netherlands)

    Koutsopoulos, Sotirios; van der Oost, John; Norde, Willem

    2007-01-01

    The thermal denaturation of endo-beta-1,3-glucanase from the hyperthermophilic microorganism Pyrococcus furiosus was studied by calorimetry. The calorimetric profile revealed two transitions at 109 and 144 degrees C, corresponding to protein denaturation and complete unfolding, respectively, as

  3. Kinetically controlled refolding of a heat denatured hyperthermostable protein

    NARCIS (Netherlands)

    Koutsopoulos, S.; Oost, van der J.; Norde, W.

    2007-01-01

    The thermal denaturation of endo-ß-1,3-glucanase from the hyperthermophilic microorganism Pyrococcus furiosus was studied by calorimetry. The calorimetric profile revealed two transitions at 109 and 144¿°C, corresponding to protein denaturation and complete unfolding, respectively, as shown by

  4. 19 CFR 10.56 - Vegetable oils, denaturing; release.

    Science.gov (United States)

    2010-04-01

    ... 19 Customs Duties 1 2010-04-01 2010-04-01 false Vegetable oils, denaturing; release. 10.56 Section... Vegetable Oils § 10.56 Vegetable oils, denaturing; release. (a) Olive, palm-kernel, rapeseed, sunflower, and sesame oil shall be classifiable under subheadings 1509.10.20, 1509.10.40, 1509.90.20, 1509.90.40,...

  5. 27 CFR 19.464 - Denatured spirits inventories.

    Science.gov (United States)

    2010-04-01

    ... inventories. 19.464 Section 19.464 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE... of Articles Inventories § 19.464 Denatured spirits inventories. Each proprietor shall take a physical inventory of all denatured spirits in the processing account at the close of each calendar quarter and...

  6. 27 CFR 20.144 - Packages of completely denatured alcohol.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Packages of completely denatured alcohol. 20.144 Section 20.144 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS DISTRIBUTION AND USE OF DENATURED ALCOHOL AND RUM...

  7. 27 CFR 20.261 - Records of completely denatured alcohol.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Records of completely denatured alcohol. 20.261 Section 20.261 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS DISTRIBUTION AND USE OF DENATURED ALCOHOL AND...

  8. [Evaluation of peptide nucleic acid fluorescent in situ hybridization (PNA FISH) method in the identifi cation of Candida species isolated from blood cultures].

    Science.gov (United States)

    Aydemir, Gonca; Koç, Ayşe Nedret; Atalay, Mustafa Altay

    2016-04-01

    In recent years, increased number of patients who are hospitalized in intensive care units, received immunosuppressive therapy and treated with broad-spectrum antibiotics that can lead an increase in the incidence of systemic candidiasis. In these patients, the most common clinical manifestation is candidemia. Since the identification of Candida species isolated from blood cultures is time consuming by conventional (morphological and biochemical) methods, rapid, reliable and accurate methods are needed. For this purpose novel systems have been developed to identify the agent directly. The aim of this study was to evaluate the peptide nucleic acid fluorescent in situ hybridization (PNA FISH) method for the identification of Candida species by comparing with the conventional methods. A total of 50 patients who were admitted to Erciyes University Medical Faculty Hospital clinics and followed with prediagnosis of systemic fungal infections whose blood cultures were positive for the yeasts between July 2011 and July 2012 were included in the study. The conventional identification of Candida isolates was performed by considering macroscopic and microscopic morphology, germ tube test, cycloheximide sensitivity, urease activity and carbohydrate assimilation patterns with API 20C AUX (bioMerieux, France) test. PNA FISH method was conducted by the use of a commercial kit namely Yeast Traffic Light(®) PNA FISH (AdvanDx, USA). According to morphological and biochemical characteristics (conventional methods), 19 (38%) out of 50 Candida isolates were identified as C.albicans, 12 (24%) as C.glabrata, five (10%) as C.parapsilosis, five (10%) as C.kefyr, four (8%) as C.krusei, two (4%) as C.guilliermondii, two (4%) as C.tropicalis and one (2%) as C.lusitaniae. On the other hand, 24 (48%) of the isolates were identified as C.albicans/C.parapsilosis (with green fluorescence), 16 (32%) as C.glabrata/C.krusei (with red fluorescence) and one (%2) as C.tropicalis (with yellow

  9. Simultaneous labeling of single- and double-strand DNA breaks by DNA breakage detection-FISH (DBD-FISH).

    Science.gov (United States)

    Fernández, José Luis; Cajigal, Dioleyda; Gosálvez, Jaime

    2011-01-01

    DNA Breakage Detection-Fluorescence In Situ Hybridization (DBD-FISH) permits simultaneous and selective labeling of single- and double-strand DNA breaks in individual cells, either in the whole genome or within specific DNA sequences. In this technique, cells are embedded into agarose microgels, lysed and subjected to electrophoresis under nondenaturing conditions. Subsequently, the produced "comets" are exposed to a controlled denaturation step which transforms DNA breaks into single-stranded DNA regions, detected by hybridization with whole genome fluorescent probes or the probes to specific DNA sequences. This makes possible a targeted analysis of various chromatin areas for the presence of DNA breaks. The migration length of the DBD-FISH signal is proportional to the number of double strand breaks, whereas its fluorescence intensity depends on numbers of single-strand breaks.The detailed protocol for detection of two types of DNA breaks produced by ionizing radiation is presented. The technique can be used to determine intragenomic and intercellular heterogeneity in the induction and repair of DNA damage.

  10. Fuel consumption analysis of diesel-electric hybrid system of fishing boat%渔船柴电混合动力系统油耗分析

    Institute of Scientific and Technical Information of China (English)

    曹建军; 李胜勇

    2016-01-01

    为了探索海洋渔船推进系统的节油减排,以一艘安装有柴电混合动力系统,成功实现节能减排的金枪鱼延绳钓船为例,通过剖析柴电混合动力系统的结构,柴油机理论负荷特性,结合渔船实际使用情况,分析其节能原因。结果显示,渔船柴电混合动力系统在使用过程中,可根据工况需要,随意切换常规动力与电力推进系统;通过选择合理的主机工况,提高柴油机输出效率,降低柴油机油耗率;综合各种工况,能节油15%以上。研究表明,现阶段在金枪鱼延绳钓渔船上使用混合动力技术是可行的,能起到节能减排的作用。在核心技术不变的情况下,运用一些创新技术手段,如采用多机多桨拆分系统,赋予系统更高的智能调配作用,可以把混合动力系统推广到拖网渔船和围网渔船上。%In order to explore the energy conservation performance of ocean fishing boat propulsion systems,in this paper,a tuna longline vessel that has realized energy-saving and emission-reducing was taken as an example to analyze the reasons behind,through probing into the structure of its diesel-electric hybrid system, engine load characteristics and the actual using conditions of the vessel. The results showed that the diesel-electric hybrid system of the vessel in use can easily switch between the conventional power and the electricity propulsion systems in accordance with the actual working conditions; by selecting the reasonable working conditions of the main engine,the output efficiency of the diesel engine could thus be improved and the diesel fuel consumption rate decreased,and considering the various working conditions,the system could save energy by more than 15%.The research showed that,at present,using hybrid power technology in tuna longline vessels is feasible and can play a role in energy conservation. With the core technology remaining the same and by adopting some

  11. Thermal denaturation of A-DNA

    Science.gov (United States)

    Valle-Orero, J.; Wildes, A. R.; Theodorakopoulos, N.; Cuesta-López, S.; Garden, J.-L.; Danilkin, S.; Peyrard, M.

    2014-11-01

    The DNA molecule can take various conformational forms. Investigations focus mainly on the so-called ‘B-form’, schematically drawn in the famous paper by Watson and Crick [1]. This is the usual form of DNA in a biological environment and is the only form that is stable in an aqueous environment. Other forms, however, can teach us much about DNA. They have the same nucleotide base pairs for ‘building blocks’ as B-DNA, but with different relative positions, and studying these forms gives insight into the interactions between elements under conditions far from equilibrium in the B-form. Studying the thermal denaturation is particularly interesting because it provides a direct probe of those interactions which control the growth of the fluctuations when the ‘melting’ temperature is approached. Here we report such a study on the ‘A-form’ using calorimetry and neutron scattering. We show that it can be carried further than a similar study on B-DNA, requiring the improvement of thermodynamic models for DNA.

  12. Microwave-enhanced folding and denaturation of globular proteins

    DEFF Research Database (Denmark)

    Bohr, Henrik; Bohr, Jakob

    2000-01-01

    It is shown that microwave irradiation can affect the kinetics of the folding process of some globular proteins, especially beta-lactoglobulin. At low temperature the folding from the cold denatured phase of the protein is enhanced, while at a higher temperature the denaturation of the protein from...... its folded state is enhanced. In the latter case, a negative temperature gradient is needed for the denaturation process, suggesting that the effects of the microwaves are nonthermal. This supports the notion that coherent topological excitations can exist in proteins. The application of microwaves...

  13. Effect of tannic acid-fish scale gelatin hydrolysate hybrid nanoparticles on intestinal barrier function and α-amylase activity.

    Science.gov (United States)

    Wu, Shao-Jung; Ho, Yi-Cheng; Jiang, Shun-Zhou; Mi, Fwu-Long

    2015-07-01

    Practical application of tannic acid is limited because it readily binds proteins to form insoluble aggregates. In this study, tannic acid was self-assembled with fish scale gelatin hydrolysates (FSGH) to form stable colloidal complex nanoparticles. The nanoparticles prepared from 4 mg ml(-1) tannic acid and 4 mg ml(-1) FSGH had a mean particle size of 260.8 ± 3.6 nm, and showed a positive zeta potential (20.4 ± 0.4 mV). The nanoparticles acted as effective nano-biochelators and free radical scavengers because they provided a large number of adsorption sites for interaction with heavy metal ions and scavenging free radicals. The maximum adsorption capacity for Cu(2+) ions was 123.5 mg g(-1) and EC50 of DPPH radical scavenging activity was 21.6 ± 1.2 μg ml(-1). Hydroxyl radical scavenging effects of the nanoparticles were investigated by electron spin resonance spectroscopy. The copper-chelating capacity and free radical scavenging activity of the nanoparticles were associated with their capacity to inhibit Cu(2+) ion-induced barrier impairment and hyperpermeability of Caco-2 intestinal epithelial tight junction (TJ). However, α-amylase inhibitory activity of the nanoparticles was significantly lower than that of free tannic acid. The results suggest that the nanoparticles can ameliorate Cu(2+) ion induced intestinal epithelial TJ dysfunction without severely inhibiting the activity of the digestive enzymes.

  14. A radiation hybrid map of the European sea bass (Dicentrarchus labrax) based on 1581 markers: Synteny analysis with model fish genomes.

    Science.gov (United States)

    Guyon, Richard; Senger, Fabrice; Rakotomanga, Michaelle; Sadequi, Naoual; Volckaert, Filip A M; Hitte, Christophe; Galibert, Francis

    2010-10-01

    The selective breeding of fish for aquaculture purposes requires the understanding of the genetic basis of traits such as growth, behaviour, resistance to pathogens and sex determinism. Access to well-developed genomic resources is a prerequisite to improve the knowledge of these traits. Having this aim in mind, a radiation hybrid (RH) panel of European sea bass (Dicentrarchus labrax) was constructed from splenocytes irradiated at 3000 rad, allowing the construction of a 1581 marker RH map. A total of 1440 gene markers providing ~4400 anchors with the genomes of three-spined stickleback, medaka, pufferfish and zebrafish, helped establish synteny relationships with these model species. The identification of Conserved Segments Ordered (CSO) between sea bass and model species allows the anticipation of the position of any sea bass gene from its location in model genomes. Synteny relationships between sea bass and gilthead seabream were addressed by mapping 37 orthologous markers. The sea bass genetic linkage map was integrated in the RH map through the mapping of 141 microsatellites. We are thus able to present the first complete gene map of sea bass. It will facilitate linkage studies and the identification of candidate genes and Quantitative Trait Loci (QTL). The RH map further positions sea bass as a genetic and evolutionary model of Perciformes and supports their ongoing aquaculture expansion.

  15. Statistical mechanics of thermal denaturation of DNA oligomers

    Indian Academy of Sciences (India)

    Navin Singh; Yashwant Singh

    2003-08-01

    Double stranded DNA chain is known to have non-trivial elasticity. We study the effect of this elasticity on the denaturation profile of DNA oligomer by constraining one base pair at one end of the oligomer to remain in unstretched (or intact) state. The effect of this constraint on the denaturation profile of the oligomer has been calculated using the Peyrard–Bishop Hamiltonian. The denaturation profile is found to be very different from the free (i.e. without the constraint) oligomer. We have also examined how this constraint affects the denaturation profile of the oligomer having a segment of defect sites located at different parts of the chain.

  16. Molecular cytogenetics using fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Gray, J.W.; Kuo, Wen-Lin; Lucas, J.; Pinkel, D.; Weier, H-U.; Yu, Loh-Chung.

    1990-12-07

    Fluorescence in situ hybridization (FISH) with chromosome-specific probes enables several new areas of cytogenetic investigation by allowing visual determination of the presence and normality of specific genetic sequences in single metaphase or interphase cells. in this approach, termed molecular cytogenetics, the genetic loci to be analyzed are made microscopically visible in single cells using in situ hybridization with nucleic acid probes specific to these loci. To accomplish this, the DNA in the target cells is made single stranded by thermal denaturation and incubated with single-stranded, chemically modified probe under conditions where the probe will anneal only with DNA sequences to which it has high DNA sequence homology. The bound probe is then made visible by treatment with a fluorescent reagent such as fluorescein that binds to the chemical modification carried by the probe. The DNA to which the probe does not bind is made visible by staining with a dye such as propidium iodide that fluoresces at a wavelength different from that of the reagent used for probe visualization. We show in this report that probes are now available that make this technique useful for biological dosimetry, prenatal diagnosis and cancer biology. 31 refs., 3 figs.

  17. Thermal denaturation of type I collagen vitrified gels

    Energy Technology Data Exchange (ETDEWEB)

    Xia, Zhiyong, E-mail: zhiyong.xia@jhuapl.edu [The Johns Hopkins University, Applied Physics Laboratory, Laurel, MD 20723 (United States); Calderon-Colon, Xiomara [The Johns Hopkins University, Applied Physics Laboratory, Laurel, MD 20723 (United States); Trexler, Morgana, E-mail: morgana.trexler@jhuapl.edu [The Johns Hopkins University, Applied Physics Laboratory, Laurel, MD 20723 (United States); Elisseeff, Jennifer; Guo, Qiongyu [The Johns Hopkins University, Department of Biomedical Engineering, Baltimore, MD 21231 (United States)

    2012-01-10

    Highlights: Black-Right-Pointing-Pointer We analyzed the denaturation of vitrigels synthesized under different conditions. Black-Right-Pointing-Pointer Overall denaturation kinetics consisted of both reversible and irreversible steps. Black-Right-Pointing-Pointer More stable vitrigels were formed under high level of vitrification. - Abstract: The denaturation kinetics of type I collagen vitrigels synthesized under different vitrification time and temperature were analyzed by the classical Kissinger approach and the advanced model free kinetics (AMFK) using the Vyazovkin algorithm. The AMFK successfully elucidated the overall denaturation into reversible and irreversible processes. Depending on vitrification conditions, the activation energy for the irreversible process ranged from 100 to 200 kJ/mol, and the reversible enthalpy ranged from 250 to 300 kJ/mol. All of these values increased with the vitrification time and temperature, indicating that a more stable and complex structure formed with increased vitrification. The classical Kissinger method predicted the presence of a critical temperate of approximately 60 Degree-Sign C for the transition between reversible and irreversible processes. Scanning electron microscopy revealed the presence of fibril structures in vitrigels both before and after full denaturation; however the fibrils had became thicker and rougher after denaturation.

  18. Chromosome painting by GISH and multi-color FISH

    Science.gov (United States)

    Fluorescent in situ hybridization (FISH) is a powerful cytogenetic technique for identifying chromosomes and mapping specific genes and DNA sequences on individual chromosomes. Genomic in situ hybridization (GISH) and multi-color FISH (mc-FISH) represent two special types of FISH techniques. Both ...

  19. 27 CFR 19.57 - Recovery and reuse of denatured spirits in manufacturing processes.

    Science.gov (United States)

    2010-04-01

    ... denatured spirits in manufacturing processes. 19.57 Section 19.57 Alcohol, Tobacco Products and Firearms... denatured spirits in manufacturing processes. The following persons are not, by reason of the activities...) Manufacturers who use denatured spirits, or articles or substances containing denatured spirits, in a process...

  20. Dodine as a transparent protein denaturant for circular dichroism and infrared studies.

    Science.gov (United States)

    Guin, Drishti; Sye, Kori; Dave, Kapil; Gruebele, Martin

    2016-05-01

    The fungicide dodine combines the cooperative denaturation properties of guanidine with the mM denaturation activity of SDS. It was previously tested only on two small model proteins. Here we show that it can be used as a chemical denaturant for phosphoglycerate kinase (PGK), a much larger two-domain enzyme. In addition to its properties as a chemical denaturant, dodine facilitates thermal denaturation of PGK, and we show for the first time that it also facilitates pressure denaturation of a protein. Much higher quality circular dichroism and amide I' infrared spectra of PGK can be obtained in dodine than in guanidine, opening the possibility for use of dodine as a denaturant when UV or IR detection is desirable. One caution is that dodine denaturation, like other detergent-based denaturants, is less reversible than guanidine denaturation. © 2016 The Protein Society.

  1. Distinguishing between incomplete lineage sorting and genomic introgressions: complete fixation of allospecific mitochondrial DNA in a sexually reproducing fish (Cobitis; Teleostei, despite clonal reproduction of hybrids.

    Directory of Open Access Journals (Sweden)

    Lukas Choleva

    Full Text Available Distinguishing between hybrid introgression and incomplete lineage sorting causing incongruence among gene trees in that they exhibit topological differences requires application of statistical approaches that are based on biologically relevant models. Such study is especially challenging in hybrid systems, where usual vectors mediating interspecific gene transfers--hybrids with Mendelian heredity--are absent or unknown. Here we study a complex of hybridizing species, which are known to produce clonal hybrids, to discover how one of the species, Cobitis tanaitica, has achieved a pattern of mito-nuclear mosaic genome over the whole geographic range. We appplied three distinct methods, including the method using solely the information on gene tree topologies, and found that the contrasting mito-nuclear signal might not have resulted from the retention of ancestral polymorphism. Instead, we found two signs of hybridization events related to C. tanaitica; one concerning nuclear gene flow and the other suggested mitochondrial capture. Interestingly, clonal inheritance (gynogenesis of contemporary hybrids prevents genomic introgressions and non-clonal hybrids are either absent or too rare to be detected among European Cobitis. Our analyses therefore suggest that introgressive hybridizations are rather old episodes, mediated by previously existing hybrids whose inheritance was not entirely clonal. Cobitis complex thus supports the view that the type of resulting hybrids depends on a level of genomic divergence between sexual species.

  2. Virus inactivation by protein denaturants used in affinity chromatography.

    Science.gov (United States)

    Roberts, Peter L; Lloyd, David

    2007-10-01

    Virus inactivation by a number of protein denaturants commonly used in gel affinity chromatography for protein elution and gel recycling has been investigated. The enveloped viruses Sindbis, herpes simplex-1 and vaccinia, and the non-enveloped virus polio-1 were effectively inactivated by 0.5 M sodium hydroxide, 6 M guanidinium thiocyanate, 8 M urea and 70% ethanol. However, pH 2.6, 3 M sodium thiocyanate, 6 M guanidinium chloride and 20% ethanol, while effectively inactivating the enveloped viruses, did not inactivate polio-1. These studies demonstrate that protein denaturants are generally effective for virus inactivation but with the limitation that only some may inactivate non-enveloped viruses. The use of protein denaturants, together with virus reduction steps in the manufacturing process should ensure that viral cross contamination between manufacturing batches of therapeutic biological products is prevented and the safety of the product ensured.

  3. Two conformational states in D-shaped DNA: Effects of local denaturation

    Science.gov (United States)

    Lee, O.-Chul; Kim, Cheolhee; Kim, Jae-Yeol; Lee, Nam Ki; Sung, Wokyung

    2016-06-01

    The bending of double-stranded(ds) DNA on the nano-meter scale plays a key role in many cellular processes such as nucleosome packing, transcription-control, and viral-genome packing. In our recent study, a nanometer-sized dsDNA bent into a D shape was formed by hybridizing a circular single-stranded(ss) DNA and a complementary linear ssDNA. Our fluorescence resonance energy transfer (FRET) measurement of D-DNA revealed two types of conformational states: a less-bent state and a kinked state, which can transform into each other. To understand the origin of the two deformed states of D-DNA, here we study the presence of open base-pairs in the ds portion by using the breathing-DNA model to simulate the system. We provide strong evidence that the two states are due to the emergence of local denaturation, i.e., a bubble in the middle and two forks at ends of the dsDNA portion. We also study the system analytically and find that the free-energy landscape is bistable with two minima representative of the two states. The kink and fork sizes estimated by the analytical calculation are also in excellent agreement with the results of the simulation. Thus, this combined experimental-simulation-analytical study corroborates that highly bent D-DNA reduces bending stress via local denaturation.

  4. Unusual cold denaturation of a small protein domain.

    Science.gov (United States)

    Buchner, Ginka S; Shih, Natalie; Reece, Amy E; Niebling, Stephan; Kubelka, Jan

    2012-08-21

    A thermal unfolding study of the 45-residue α-helical domain UBA(2) using circular dichroism is presented. The protein is highly thermostable and exhibits a clear cold unfolding transition with the onset near 290 K without denaturant. Cold denaturation in proteins is rarely observed in general and is quite unique among small helical protein domains. The cold unfolding was further investigated in urea solutions, and a simple thermodynamic model was used to fit all thermal and urea unfolding data. The resulting thermodynamic parameters are compared to those of other small protein domains. Possible origins of the unusual cold unfolding of UBA(2) are discussed.

  5. OBSERVATION OF DNA PARTIAL DENATURATION BY ATOMIC FORCE MICROSCOPY

    Institute of Scientific and Technical Information of China (English)

    Xin-hua Dai; Zhi-gang Wang; Bo Xiao; Yong-jun Zhang; Chen Wang; Chun-li Bai; Xiao-li Zhang; Jian Xu

    2004-01-01

    Atomic force microscopy was used to investigate the DNA-cetyltrimethylammonium bromide (CTAB) complexes adsorbed on highly ordered pyrolytic graphite (HOPG). These complexes, at low concentrations, can automatically spread out on the surface of HOPG. The DNA-CTAB complexes display a typically extended structure rather than a globular structure. Partially denaturated DNA produced by binding CTAB to DNA is directly observed by AFM with high resolution.The three-dimensional resolution of partially denaturated DNA obtained by AFM is not available by any other technique at present.

  6. Single-molecule denaturation mapping of DNA in nanofluidic channels

    DEFF Research Database (Denmark)

    Reisner, Walter; Larsen, Niels Bent; Silahtaroglu, Asli

    2010-01-01

    Here we explore the potential power of denaturation mapping as a single-molecule technique. By partially denaturing YOYO (R)-1-labeled DNA in nanofluidic channels with a combination of formamide and local heating, we obtain a sequence-dependent "barcode" corresponding to a series of local dips....... Consequently, the technique is sensitive to sequence variation without requiring enzymatic labeling or a restriction step. This technique may serve as the basis for a new mapping technology ideally suited for investigating the long-range structure of entire genomes extracted from single cells....

  7. Leptin stimulates hepatic growth hormone receptor and insulin-like growth factor gene expression in a teleost fish, the hybrid striped bass.

    Science.gov (United States)

    Won, Eugene T; Douros, Jonathan D; Hurt, David A; Borski, Russell J

    2016-04-01

    Leptin is an anorexigenic peptide hormone that circulates as an indicator of adiposity in mammals, and functions to maintain energy homeostasis by balancing feeding and energy expenditure. In fish, leptin tends to be predominantly expressed in the liver, another important energy storing tissue, rather than in fat depots as it is in mammals. The liver also produces the majority of circulating insulin-like growth factors (IGFs), which comprise the mitogenic component of the growth hormone (GH)-IGF endocrine growth axis. Based on similar regulatory patterns of leptin and IGFs that we have documented in previous studies on hybrid striped bass (HSB: Morone saxatilis×Morone chrysops), and considering the co-localization of these peptides in the liver, we hypothesized that leptin might regulate the endocrine growth axis in a manner that helps coordinate somatic growth with energy availability. Using a HSB hepatocyte culture system to simulate autocrine or paracrine exposure that might occur within the liver, this study examines the potential for leptin to modulate metabolism and growth through regulation of IGF gene expression directly, or indirectly through the regulation of GH receptors (GHR), which mediate GH-induced IGF expression. First, we verified that GH (50nM) has a classical stimulatory effect on IGF-1 and additionally show it stimulates IGF-2 transcription in hepatocytes. Leptin (5 and/or 50nM) directly stimulated in vitro GHR2 gene expression within 8h of exposure, and both GHR1 and GHR2 as well as IGF-1 and IGF-2 gene expression after 24h. Cells were then co-incubated with submaximal concentrations of leptin and GH (25nM each) to test if they had a synergistic effect on IGF gene expression, possibly through increased GH sensitivity following GHR upregulation by leptin. In combination, however, the treatments only had an additive effect on stimulating IGF-1 mRNA despite their capacity to increase GHR mRNA abundance. This suggests that leptin's stimulatory

  8. BIOTECHNOLOGY OF THE FISH AQUACULTURE

    Directory of Open Access Journals (Sweden)

    L. P. Buchatsky

    2013-12-01

    Full Text Available The latest progress in biotechnology on fish aquaculture and different modern methods of investigations for increasing of fish productivity in aquaculture are analyzed. Except for the applied aspect, the use of modern biotechnological methods of investigations opens new possibilities for fundamental researches of sex-determining mechanisms, polyploidy, distant hybridization, and developmental biology of bony fishes. Review contains examples of utilizing modern biotechnology methods to obtain transgenic fishes with accelerated growth and for designing surrogate fishes. Methods for receiving unisexual shoals of salmon and sturgeon female fishes with the view of obtaining a large quantity of caviar, as well as receiving sterile (triploid fishes are analyzed. Great attention is given to androgenesis, particularly to disperm one, in connection with the problem of conserving rare and vanishing fish species using only sperm genetic material. Examples how distant hybrids may be obtained with the use of disperm androgenesis and alkylated DNA are given. Methods of obtaining fish primordium germ cells, recent developments in cultivation of fish stem cells and their use in biotechnology, as well as ones of transplantation of oogonium and spermatogonium to obtain surrogate fishes. The examples of successful experiments on spermatogonial xenotransplantation and characteristic of antifreezing fish proteins and also the prospect of their practical usage are given.

  9. Denaturation of membrane proteins and hyperthermic cell killing

    NARCIS (Netherlands)

    Burgman, Paulus Wilhelmus Johannes Jozef

    1993-01-01

    Summarizing: heat induced denaturation of membrane proteins is probably related to hyperthermic cell killing. Induced resistance of heat sensitive proteins seems to be involved in the development of thermotolerance. Although many questions remain still to be answered, it appears that HSP72, when

  10. Fish Allergy

    Science.gov (United States)

    ... Old Feeding Your 1- to 2-Year-Old Fish Allergy KidsHealth > For Parents > Fish Allergy Print A ... From Home en español Alergia al pescado About Fish Allergy A fish allergy is not exactly the ...

  11. Urea Induced Denaturation of Pre-Q1 Riboswitch

    Science.gov (United States)

    Yoon, Jeseong; Thirumalai, Devarajan; Hyeon, Changbong

    2013-01-01

    Urea, a polar molecule with a large dipole moment, not only destabilizes the folded RNA structures, but can also enhance the folding rates of large ribozymes. Unlike the mechanism of urea-induced unfolding of proteins, which is well understood, the action of urea on RNA has barely been explored. We performed extensive all atom molecular dynamics (MD) simulations to determine the molecular underpinnings of urea-induced RNA denaturation. Urea displays its denaturing power in both secondary and tertiary motifs of the riboswitch (RS) structure. Our simulations reveal that the denaturation of RNA structures is mainly driven by the hydrogen bonds and stacking interactions of urea with the bases. Through detailed studies of the simulation trajectories, we found that geminate pairs between urea and bases due to hydrogen bonds and stacks persist only ~ (0.1-1) ns, which suggests that urea-base interaction is highly dynamic. Most importantly, the early stage of base pair disruption is triggered by penetration of water molecules into the hydrophobic domain between the RNA bases. The infiltration of water into the narrow space between base pairs is critical in increasing the accessibility of urea to transiently disrupted bases, thus allowing urea to displace inter base hydrogen bonds. This mechanism, water-induced disruption of base-pairs resulting in the formation of a "wet" destabilized RNA followed by solvation by urea, is the exact opposite of the two-stage denaturation of proteins by urea. In the latter case, initial urea penetration creates a dry-globule, which is subsequently solvated by water penetration leading to global protein unfolding. Our work shows that the ability to interact with both water and polar, non-polar components of nucleotides makes urea a powerful chemical denaturant for nucleic acids.

  12. Insights into food preference in hybrid F1 of Siniperca chuatsi (♀) × Siniperca scherzeri (♂) mandarin fish through transcriptome analysis.

    Science.gov (United States)

    He, Shan; Liang, Xu-Fang; Sun, Jian; Li, Ling; Yu, Ying; Huang, Wei; Qu, Chun-Mei; Cao, Liang; Bai, Xiao-Li; Tao, Ya-Xiong

    2013-09-05

    As economically relevant traits, feeding behavior and food preference domestication determine production cost and profitability. Although there are intensive research efforts on feeding behavior and food intake, little is known about food preference. Mandarin fish accept only live prey fish and refuse dead prey fish or artificial diets. Very little is currently known about the genes regulating this unique food preference. Using transcriptome sequencing and digital gene expression profiling, we identified 1,986 and 4,526 differentially expressed genes in feeders and nonfeeders of dead prey fish, respectively. Up-regulation of Crbp, Rgr and Rdh8, and down-regulation of Gc expression, consistent with greater visual ability in feeders, could promote positive phototaxis. Altered expressions of period, casein kinase and Rev-erbα might reset circadian phase. Down-regulation of orexigenic and up-regulation of anorexigenic genes in feeders were associated with lower appetite. The mRNA levels of Creb, c-fos, C/EBP, zif268, Bdnf and Syt were dramatically decreased in feeders, which might result in significant deficiency in memory retention of its natural food preference (live prey fish). There were roughly 100 times more potential SNPs in feeders than in nonfeeders. In summary, differential expression in the genes identified shed new light on why mandarin fish only feed on live prey fish, with pathways regulating retinal photosensitivity, circadian rhythm, appetite control, learning and memory involved. We also found dramatic difference in SNP abundance in feeders vs nonfeeders. These differences together might account for the different food preferences. Elucidating the genes regulating the unique food preference (live prey fish) in mandarin fish could lead to a better understanding of mechanisms controlling food preference in animals, including mammals.

  13. Denaturing and non-denaturing gel electrophoresis as methods for the detection ofjunctional diversity in rearranged T cell receptor sequences

    NARCIS (Netherlands)

    Offermans, M.T.C.; Sonneveld, R.D.; Bakker, E.; Deutz-Terlouw, P.P.; Geus, B. de; Rozing, J.

    1995-01-01

    Two nucleic acid gel electrophoresis techniques were tested as a possible tool for analyzing junctional diversity in rearranged T cell receptor (TcR) sequences in order to define the extent of T cell heterogeneity. For this purpose denaturing gradient gel electrophoresis (DGGE) as well as

  14. BIOTECHNOLOGY OF THE FISH AQUACULTURE

    OpenAIRE

    L. P. Buchatsky

    2013-01-01

    The latest progress in biotechnology on fish aquaculture and different modern methods of investigations for increasing of fish productivity in aquaculture are analyzed. Except for the applied aspect, the use of modern biotechnological methods of investigations opens new possibilities for fundamental researches of sex-determining mechanisms, polyploidy, distant hybridization, and developmental biology of bony fishes. Review contains examples of utilizing modern biotechnology methods to obtain ...

  15. Progress in molecular diagnosis of Charcot-Marie-Tooth-disease type 1 (CMT 1, HMSN I) and hereditary neuropathy with liability to pressure palsies (HNPP) by fluorescence in situ hybridization (FISH)-detection of a potential genetic mosaicism

    Energy Technology Data Exchange (ETDEWEB)

    Bathke, K.; Liehr. T.; Ekici, A. [Institute for Human Genetics, Erlange (Germany)] [and others

    1994-09-01

    We tested 20 CMT 1 patients characterized according to the criteria of the European CMT consortium by Southern hybridization of MspI restricted genomic DNA with probes pVAW409R1, pVAW412Hec and pEW401HE. In 11 of the 20 CMT 1 cases (55%), we observed a duplication in 17q11.2; one patient had a dinucleotide insertion in exon 6 of the PO-gene (5%). One HNPP case had a typical 17p11.2 deletion. Analysis of CA-repeats was performed with primers RM11GT and Mfd41; SSCP-analysis of the PO, PMP22 and Cx32-genes is in progress. FISH was carried out with probe pVAW409R1. 125 interphase nuclei were analyzed for each proband by counting the signals per nucleus. Normal cells show a characteristic distribution of signals: 1 signal in 5.9% of nuclei, 2 in 86.3% and 3 in 7.8%. A duplication is indicated by a shift to 3 signals in more than approximately 60% and 2 in less than 25% of the nuclei. In contrast, the 17p11.2 deletion of the HNPP patient shifts to 82.4% of nuclei with a single hybridization signal versus 14.4% with 2 signals. We detected one case with significantly abnormal distribution of interphase nuclei hybridization signals compared to cultures of normal cells and to those with 17p11.2 duplication or deletion: 3.2% nuclei revealed 1 signal, 48.0% two signals and 48.8% 3 signals, indicating a pathogenic but moderate dosis increase compared to the throughout duplicated cases. FISH with probe pVAW409R1 is a versatile tool to detect the HNPP deletion both in interphase nuclei and in metaphase chromosomes. In CMT 1 disease interphase nuclei are required for FISH analysis due to the small duplication of 1.5 Mbp. In contrast to Southern techniques, FISH is able to detect genetic mosaicism.

  16. Theoretical Study on Effects of Salt and Temperature on Denaturation Transition of Double-stranded DNA

    Institute of Scientific and Technical Information of China (English)

    DONG Rui-Xin; YAN Xun-Ling; PANG Xiao-Feng; JIANG Shan; LIU Sheng-Gang

    2004-01-01

    We investigate the statistical mechanics properties of a nonlinear dynamics model of the denaturation of the DNA double-helix and study the effects of salt concentration and temperature on denaturation transition of DNA. The specific heat, entropy, and denaturation temperature of the system versus salt concentration are obtained. These results show that the denaturation of DNA not only depends on the temperature but also is influenced by the salt concentration in the solution of DNA, which are in agreement with experimental measurement.

  17. Rheological properties of sweet potato starch before and after denaturalization

    Institute of Scientific and Technical Information of China (English)

    肖华西; 林亲录; 夏新剑; 李丽辉; 林利忠; 吴卫国

    2008-01-01

    Based on the sweet potato starch,cationic starch,acetic starch and cationic-acetic compoundedly modified starch were made through chemical denaturalization.The above three kinds of static rheological parameter and dynamic rheological parameter were measured,respectively.The experimental result reveals that the thermal stability of starchy viscosity increases after chemical denaturalization.Under the condition of identical shearing rate,the shear stress of cationic-acetic ester compoundedly modified sweet potato starch paste is the largest among these kinds of sweet potato starch.This attributes to a phenomenon of shearing thinning.Furthermore,raw sweet potato starch has a larger gel intensity than that of modified starch.

  18. How water contributes to pressure and cold denaturation of proteins

    CERN Document Server

    Bianco, Valentino

    2015-01-01

    The mechanisms of cold- and pressure-denaturation of proteins are matter of debate and are commonly understood as due to water-mediated interactions. Here we study several cases of proteins, with or without a unique native state, with or without hydrophilic residues, by means of a coarse-grain protein model in explicit solvent. We show, using Monte Carlo simulations, that taking into account how water at the protein interface changes its hydrogen bond properties and its density fluctuations is enough to predict protein stability regions with elliptic shapes in the temperature-pressure plane, consistent with previous theories. Our results clearly identify the different mechanisms with which water participates to denaturation and open the perspective to develop advanced computational design tools for protein engineering.

  19. Influence of Ficoll on urea induced denaturation of fibrinogen

    Science.gov (United States)

    Sankaranarayanan, Kamatchi; Meenakshisundaram, N.

    2016-03-01

    Ficoll is a neutral, highly branched polymer used as a molecular crowder in the study of proteins. Ficoll is also part of Ficoll-Paque used in biology laboratories to separate blood to its components (erythrocytes, leukocytes etc.,). Role of Ficoll in the urea induced denaturation of protein Fibrinogen (Fg) has been analyzed using fluorescence, circular dichroism, molecular docking and interfacial studies. Fluorescence studies show that Ficoll prevents quenching of Fg in the presence of urea. From the circular dichroism spectra, Fg shows conformational transition to random coil with urea of 6 M concentration. Ficoll helps to shift this denaturation concentration to 8 M and thus constraints by shielding Fg during the process. Molecular docking studies indicate that Ficoll interacts favorably with the protein than urea. The surface tension and shear viscosity analysis shows clearly that the protein is shielded by Ficoll.

  20. Temperature induced denaturation of collagen in acidic solution.

    Science.gov (United States)

    Mu, Changdao; Li, Defu; Lin, Wei; Ding, Yanwei; Zhang, Guangzhao

    2007-07-01

    The denaturation of collagen solution in acetic acid has been investigated by using ultra-sensitive differential scanning calorimetry (US-DSC), circular dichroism (CD), and laser light scattering (LLS). US-DSC measurements reveal that the collagen exhibits a bimodal transition, i.e., there exists a shoulder transition before the major transition. Such a shoulder transition can recover from a cooling when the collagen is heated to a temperature below 35 degrees C. However, when the heating temperature is above 37 degrees C, both the shoulder and major transitions are irreversible. CD measurements demonstrate the content of triple helix slowly decreases with temperature at a temperature below 35 degrees C, but it drastically decreases at a higher temperature. Our experiments suggest that the shoulder transition and major transition arise from the defibrillation and denaturation of collagen, respectively. LLS measurements show the average hydrodynamic radius R(h), radius of gyration R(g)of the collagen gradually decrease before a sharp decrease at a higher temperature. Meanwhile, the ratio R(g)/R(h) gradually increases at a temperature below approximately 34 degrees C and drastically increases in the range 34-40 degrees C, further indicating the defibrillation of collagen before the denaturation.

  1. [DNA degradation during standard alkaline of thermal denaturation].

    Science.gov (United States)

    Drozhdeniuk, A P; Sulimova, G E; Vaniushin, B F

    1976-01-01

    Essential degradation 8 DNA (up to 10 per cent) with liberation of acid-soluble fragments takes place on the standard alkaline (0,01 M sodium phosphate, pH 12, 60 degrees, 15 min) or thermal (0.06 M sodium phosphate buffer, pH 6.8, 102 degrees C, 15 min) denaturation. This degradation is more or less selective: fraction of low molecular weight fragments, isolated by hydroxyapatite cromatography and eluted by 0.06 M sodium phosphate buffer, pH 6.8 is rich in adenine and thymine and contains about 2 times less 5-methylcytosine than the total wheat germ DNA. The degree of degradation of DNA on thermal denaturation is higher than on alkaline degradation. Therefore while studying reassociation of various DNA, one and the same standard method of DNA denaturation should be used. Besides, both the level of DNA degradation and the nature of the resulting products (fragments) should be taken into account.

  2. 荧光原位杂交法(FISH)快速检测尿液中金黄色葡萄球菌%Fluorescence in Situ Hybridization Rapidly Detects Staphylococcus Aureus in Urinary Tract Infection Samples

    Institute of Scientific and Technical Information of China (English)

    吴青; 李艳; 汪明; 顾剑; 胡慧霞; 徐万洲; 韦传东; 吴泽刚

    2011-01-01

    目的 利用荧光原位杂交法快速检测尿液中的金黄色葡萄球菌,筛查金黄色葡萄球菌所致的尿路感染.方法 针对金黄色葡萄球菌16sRNA设计荧光标记的核苷酸探针,利用荧光原位杂交技术(FISH)对132例疑似尿路感染患者中段尿标本进行检测;同时进行中段尿培养.结果 荧光原位杂交法检测阳性的有9例,与中段尿培养比较,其敏感度为100.0%,特异度为99.2%,阳性预期值为90.0%,阴性预期值为100.0%.结论 荧光原位杂交能快速检测尿液中的金黄色葡萄球菌,对金黄色葡萄球菌所致的尿路感染进行快速诊断.%Objective The rapid detection of Staphylococcus aureus in urine by fluorescence in situ hybridization (FISH)method. Screening urinary tract infection (UTIs) caused by Staphylococcus aureus. Methods Probes that were specific for Staphylococcus aureus were designed based on the conserved 16 s RNA sequences. Detected a total of 132 urine samples using FISH method. And all of tbese samples tested via traditional culturing techniques. Results 9 of these samples tested positive for a UTI via FISH analysis. Compared to conventional methods of bacterial identification,the FISH method had a sensitivity of 100.0% ,specificity of 99. 2%,positive expected value of 90. 0% and negative expected value of 100. 0%. Conclusion FISH had the potential to become an extremely useful diagnostic tool for UTIs because it has a quick turnaround time and high accuracy.

  3. Protein's native state stability in a chemically induced denaturation mechanism.

    Science.gov (United States)

    Olivares-Quiroz, L; Garcia-Colin, L S

    2007-05-21

    In this work, we present a generalization of Zwanzig's protein unfolding analysis [Zwanzig, R., 1997. Two-state models of protein folding kinetics. Proc. Natl Acad. Sci. USA 94, 148-150; Zwanzig, R., 1995. Simple model of protein folding kinetics. Proc. Natl Acad. Sci. USA 92, 9801], in order to calculate the free energy change Delta(N)(D)F between the protein's native state N and its unfolded state D in a chemically induced denaturation. This Extended Zwanzig Model (EZM) is both based on an equilibrium statistical mechanics approach and the inclusion of experimental denaturation curves. It enables us to construct a suitable partition function Z and to derive an analytical formula for Delta(N)(D)F in terms of the number K of residues of the macromolecule, the average number nu of accessible states for each single amino acid and the concentration C(1/2) where the midpoint of the ND transition occurs. The results of the EZM for proteins where chemical denaturation follows a sigmoidal-type profile, as it occurs for the case of the T70N human variant of lysozyme (PDB code: T70N) [Esposito, G., et al., 2003. J. Biol. Chem. 278, 25910-25918], can be splitted into two lines. First, EZM shows that for sigmoidal denaturation profiles, the internal degrees of freedom of the chain play an outstanding role in the stability of the native state. On the other hand, that under certain conditions DeltaF can be written as a quadratic polynomial on concentration C(1/2), i.e., DeltaF approximately aC(1/2)(2)+bC(1/2)+c, where a,b,c are constant coefficients directly linked to protein's size K and the averaged number of non-native conformations nu. Such functional form for DeltaF has been widely known to fit experimental measures in chemically induced protein denaturation [Yagi, M., et al., 2003. J. Biol. Chem. 278, 47009-47015; Asgeirsson, B., Guojonsdottir, K., 2006. Biochim. Biophys. Acta 1764, 190-198; Sharma, S., et al., 2006. Protein Pept. Lett. 13(4), 323-329; Salem, M., et al

  4. International, collaborative assessment of limitations of chromosome-specific probes (CSP) and fluorescent in situ hybridization (FISH): Analysis of expected detections in 73,000 prenatal cases

    Energy Technology Data Exchange (ETDEWEB)

    Evans, M.I.; Henry, G.P.; Miller, W.A. [Wayne State Univ., Detroit, MI (United States)] [and others

    1994-09-01

    FISH and CSP have been proposed to reduce karyotyping need. The purpose of this study was to assess the potential efficacy of CSP-FISH using currently available probes (13, 18, 21, X, & Y) in large, prenatal diagnostic centers. Results (1990-1993) from 7 centers in 4 countries were divided by those expected to be detectable by currently available probes, and those which would be missed assuming 10% probe efficacy. 72,994 karyotypes included 699 trisomy 21`s, 352 trisomy 18`s, 136 trisomy 13`s, 358 sex chromosome aneuploidies, 70 triploidies, and 855 others (translocations, inversions, deletions, markers). Of 2,613 abnormalities, 1,745 would be detectable (66.8%). [Detroit 55.7%, Stockholm 68.3%, Boston 52.6%, Denver 61.3%, Muenster 77.0%, London 84.5%, Philadelphia 69.4%]. Centers with high proportions of referrals for ultrasound anomalies had the highest CSP-FISH positives secondary to increased T 18 & 13. We conclude: (1) 73,000 karyotypes show relatively consistent incidences of the common trisomies, sex chromosome abnormalities, and other chromosome abnormalities among the centers. (2) The proportion expected detectable by FISH-CSP technology varies from 52.6% to 84.5%, averaging 66.8%. (3) 1/3 of the karyotypic abnormalities would be missed, and therefore, replacement of complete karyotyping with FISH would have unacceptably high false-negative rates for routine evaluation. (4) FISH-CSP, while useful when positive for anomalies, is not sufficient when negative to obviate the need for a complete karyotype.

  5. Fish Hearing.

    Science.gov (United States)

    Blaxter, J. H. S.

    1980-01-01

    Provides related information about hearing in fish, including the sensory stimulus of sound in the underwater environment, mechanoreceptors in fish, pressure perception and the swimbladder, specializations in sound conduction peculiar to certain fish families. Includes numerous figures. (CS)

  6. Fish Allergy

    Science.gov (United States)

    ... in a clear and consistent manner, so that consumers with food allergies and their caregivers can be informed as ... the menu, cross-contact with fish is possible. Ethnic ... fish. Avoid foods like fish sticks and anchovies. Some individuals with ...

  7. Fluorescence in situ hybridization(FISH) technique combined with karyotyping analysis in prenatal diagnosis%FISH技术在产前诊断中的应用价值研究

    Institute of Scientific and Technical Information of China (English)

    沈国松; 张甦; 何平亚; 方嵘

    2011-01-01

    目的 使用荧光原位杂交(FISH)技术对常见胎儿染色体数目异常进行快速诊断,结合羊水细胞培养染色体核型分析技术形成产前诊断体系,并对其临床应用价值进行评价.方法 应用诊断最常见染色体病的5种染色体(13、18、21、X和Y)特异性FISH探针对480例未经培养羊水细胞进行产前诊断,并和同时进行的羊水培养染色体核型分析相比较.结果 480例未经培养羊水细胞FISH实验全部获得检测结果,并发现21-三体综合征2例,18-三体综合征1例,克氏综合征1例,特纳综合征 1例,与羊水培养染色体分析结果一致,但报告时间(2至3d)与羊水染色体分析报告时间(2~3周)相比大为缩短.受固有技术限制有6例结构异常未能检出,但应用FISH技术对其中1例与性染色体有关的结构异常进行辅助诊断,获得了成功.结论 FISH技术在常见染色体数目异常产前诊断中应用行之有效,与羊水染色体核型分析技术相结合,将使产前诊断更加高效和安全.%To evaluate the clinical application of the combination of fluorescence in situ hybridization (FISH)technique and karyotyping analysis in prenatal diagnosis.MethodsFISH technology with the centromeric probes of chromo-some 13, 18, 21 ,X and Y was applied in 480 amniotic fluid specimens, the results were compared with those of the G banding karyotypes from standard cytogenetic analysis in cultured amniotic fluid cells.ResultsTotal 480 uncultured amniotic fluid speci-mens were successfully tested by FISH and two Down' s syndrome, one Edwards syndrome, one Klinefelter syndrome and one Turner syndrome were detected. The results obtained by FISH were consistent with those from karyotyping analysis. FISH (2-3 days) was faster than karyotyping(2-3 weeks) to get results. Karyotypes analysis detected 6 cases of chromosome structural abnormalities which were not detected by FISH, however, one of them with a sex chromosome-related structural

  8. Role of immunohistochemistry and fluorescence in-situ hybridization (FISH in the diagnosis of spindle and round cell tumors of the kidney

    Directory of Open Access Journals (Sweden)

    M. Abbas

    2015-09-01

    Conclusion: In summary we advise an immunohistochemical panel for round/spindle cell tumors of the kidney and for unclear cases we advise to add (FISH to get the correct diagnosis, as they are completely different regarding surgical approach and post-operative adjuvant therapy.

  9. Collagen thermal denaturation study for thermal angioplasty based on modified kinetic model: relation between the artery mechanical properties and collagen denaturation rate

    Science.gov (United States)

    Shimazaki, N.; Hayashi, T.; Kunio, M.; Arai, T.

    2010-02-01

    We have been developing the novel short-term heating angioplasty in which sufficient artery lumen-dilatation was attained with thermal softening of collagen fiber in artery wall. In the present study, we investigated on the relation between the mechanical properties of heated artery and thermal denaturation fractures of arterial collagen in ex vivo. We employed Lumry-Eyring model to estimate temperature- and time-dependent thermal denaturation fractures of arterial collagen fiber during heating. We made a kinetic model of arterial collagen thermal denaturation by adjustment of K and k in this model, those were the equilibrium constant of reversible denaturation and the rate constant of irreversible denaturation. Meanwhile we demonstrated that the change of reduced scattering coefficient of whole artery wall during heating reflected the reversible denaturation of the collagen in artery wall. Based on this phenomenon, the K was determined experimentally by backscattered light intensity measurement (at 633nm) of extracted porcine carotid artery during temperature elevation and descending (25°C-->80°C-->25°C). We employed the value of according to our earlier report in which the time-and temperature- dependent irreversible denaturation amount of the artery collagen fiber that was assessed by the artery birefringence. Then, the time- and temperature- dependent reversible (irreversible) denaturation fraction defined as the reversible ((irreversible) denatured collagen amount) / (total collagen amount) was calculated by the model. Thermo-mechanical analysis of artery wall was performed to compare the arterial mechanical behaviors (softening, shrinkage) during heating with the calculated denaturation fraction with the model. In any artery temperature condition in 70-80°, the irreversible denaturation fraction at which the artery thermal shrinkage started was estimated to be around 20%. On the other hand, the calculated irreversible denaturation fraction remained below

  10. Detection of MDM2/CDK4 amplification in lipomatous soft tissue tumors from formalin-fixed, paraffin-embedded tissue: comparison of multiplex ligation-dependent probe amplification (MLPA) and fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Creytens, David; van Gorp, Joost; Ferdinande, Liesbeth; Speel, Ernst-Jan; Libbrecht, Louis

    2015-02-01

    In this study, the detection of MDM2 and CDK4 amplification was evaluated in lipomatous soft tissue tumors using multiplex ligation-dependent probe amplification (MLPA), a PCR-based technique, in comparison with fluorescence in situ hybridization (FISH). These 2 techniques were evaluated in a series of 77 formalin-fixed, paraffin-embedded lipomatous tumors (27 benign adipose tumors, 28 atypical lipomatous tumors/well-differentiated liposarcomas, 18 dedifferentiated liposarcomas, and 4 pleomorphic liposarcomas). Using MLPA, with a cut-off ratio of >2, 36/71 samples (22 atypical lipomatous tumors/well-differentiated liposarcomas, and 14 dedifferentiated liposarcomas) showed MDM2 and CDK4 amplification. Using FISH as gold standard, MLPA showed a sensitivity of 90% (36/40) and a specificity of 100% (31/31) in detecting amplification of MDM2 and CDK4 in lipomatous soft tissue tumors. In case of high-level amplification (MDM2-CDK4/CEP12 ratio >5), concordance was 100%. Four cases of atypical lipomatous tumor/well-differentiated liposarcoma (4/26, 15%) with a low MDM2 and CDK4 amplification level (MDM2-CDK4/CEP12 ratio ranging between 2 and 2.5) detected by FISH showed no amplification by MLPA, although gain of MDM2 and CDK4 (ratios ranging between 1.6 and 1.9) was seen with MLPA. No amplification was detected in benign lipomatous tumors and pleomorphic liposarcomas. Furthermore, there was a very high concordance between the ratios obtained by FISH and MLPA. In conclusion, MLPA proves to be an appropriate and straightforward technique for screening MDM2/CDK4 amplification in lipomatous tumors, especially when a correct cut-off value and reference samples are chosen, and could be considered a good alternative to FISH to determine MDM2 and CDK4 amplification in liposarcomas. Moreover, because MLPA, as a multiplex technique, allows simultaneous detection of multiple chromosomal changes of interest, it could be in the future a very reliable and fast molecular analysis on

  11. Denatured ethanol release into gasoline residuals, Part 1: Source behaviour

    Science.gov (United States)

    Freitas, Juliana G.; Barker, James F.

    2013-05-01

    With the increasing use of ethanol in fuels, it is important to evaluate its fate when released into the environment. While ethanol is less toxic than other organic compounds present in fuels, one of the concerns is the impact ethanol might have on the fate of gasoline hydrocarbons in groundwater. One possible concern is the spill of denatured ethanol (E95: ethanol containing 5% denaturants, usually hydrocarbons) in sites with pre-existing gasoline contamination. In that scenario, ethanol is expected to increase the mobility of the NAPL phase by acting as a cosolvent and decreasing interfacial tension. To evaluate the E95 behaviour and its impacts on pre-existing gasoline, a field test was performed at the CFB-Borden aquifer. Initially gasoline contamination was created releasing 200 L of E10 (gasoline with 10% ethanol) into the unsaturated zone. One year later, 184 L of E95 was released on top of the gasoline contamination. The site was monitored using soil cores, multilevel wells and one glass access tube. At the end of the test, the source zone was excavated and the compounds remaining were quantified. E95 ethanol accumulated and remained within the capillary fringe and unsaturated zone for more than 200 days, despite ~ 1 m oscillations in the water table. The gasoline mobility increased and it was redistributed in the source zone. Gasoline NAPL saturations in the soil increased two fold in the source zone. However, water table oscillations caused a separation between the NAPL and ethanol: NAPL was smeared and remained in deeper positions while ethanol moved upwards following the water table rise. Similarly, the E95 denaturants that initially were within the ethanol-rich phase became separated from ethanol after the water table oscillation, remaining below the ethanol rich zone. The separation between ethanol and hydrocarbons in the source after water table oscillation indicates that ethanol's impact on hydrocarbon residuals is likely limited to early times.

  12. Annual trends in catchability and fish stock assessments

    OpenAIRE

    2003-01-01

    A key assumption of many fish stock assessment models is that catchability is constant over time. We assume here that trends in catchability may occur through fishing power creeping. The tuning fleets, which are prone to fishing power development, may be identified using the "Hybrid" method. A range of catchability trends, including values derived from the "Hybrid" method, is then implemented to standardise the fishing effort of some tuning fleets used in the stock assessments performed by XS...

  13. Strategies for denaturing the weapons-grade plutonium stockpile

    Energy Technology Data Exchange (ETDEWEB)

    Buckner, M.R.; Parks, P.B.

    1992-10-01

    In the next few years, approximately 50 metric tons of weapons-grade plutonium and 150 metric tons of highly-enriched uranium (HEU) may be removed from nuclear weapons in the US and declared excess. These materials represent a significant energy resource that could substantially contribute to our national energy requirements. HEU can be used as fuel in naval reactors, or diluted with depleted uranium for use as fuel in commercial reactors. This paper proposes to use the weapons-grade plutonium as fuel in light water reactors. The first such reactor would demonstrate the dual objectives of producing electrical power and denaturing the plutonium to prevent use in nuclear weapons.

  14. RNA Denaturation: Excluded Volume, Pseudoknots, and Transition Scenarios

    Science.gov (United States)

    Baiesi, M.; Orlandini, E.; Stella, A. L.

    2003-11-01

    A lattice model of RNA denaturation which fully accounts for the excluded volume effects among nucleotides is proposed. A numerical study shows that interactions forming pseudoknots must be included in order to get a sharp continuous transition. Otherwise a smooth crossover occurs from the swollen linear polymer behavior to highly ramified, almost compact conformations with secondary structures. In the latter scenario, which is appropriate when these structures are much more stable than pseudoknot links, probability distributions for the lengths of both loops and main branches obey scaling with nonclassical exponents.

  15. DSC study of cold and heat denaturation processes of β-lactoglobulin A with guanidine hydrochloride

    Institute of Scientific and Technical Information of China (English)

    王邦宁; 谈夫

    1997-01-01

    The cold and heat denaturations of bovine β-lactoglobuhn A (β-lg A) has been studied in solutions of guanidine hydrochloride (GuHCl) by differential scanning calorimelry (DSC) The experimental results are presented and discussed.It is shown that the number of protons bound by the monomeric molecules of β-lg A was unchanged before and after its heat denaturation below pH 3,and that the activation energy of the heat denaturation was depressed owing to the presence of GuHCl.In the solutions with 2.50 and 3.06 mol/L of GuHCl,both the cold and heat denat-urations of β-lg A were observed.In comparison with the heat denaturation,the activation energy of cold denaturation was far lower and the number of GuHCl molecules bound by the unfolded polypeptide chains after cold denaturation increased a lot.The absolute value of the enthalpy of cold denaturation was larger than that of heat denaturation It was found by the analysis that the contribution to the total denaturational enthalpy of conformational change i

  16. Rapid discrimination of Haemophilus influenzae, H. parainfluenzae, and H. haemolyticus by fluorescence in situ hybridization (FISH and two matrix-assisted laser-desorption-ionization time-of-flight mass spectrometry (MALDI-TOF-MS platforms.

    Directory of Open Access Journals (Sweden)

    Hagen Frickmann

    Full Text Available BACKGROUND: Due to considerable differences in pathogenicity, Haemophilus influenzae, H. parainfluenzae and H. haemolyticus have to be reliably discriminated in routine diagnostics. Retrospective analyses suggest frequent misidentifications of commensal H. haemolyticus as H. influenzae. In a multi-center approach, we assessed the suitability of fluorescence in situ hybridization (FISH and matrix-assisted laser-desorption-ionization time-of-flight mass-spectrometry (MALDI-TOF-MS for the identification of H. influenzae, H. parainfluenzae and H. haemolyticus to species level. METHODOLOGY: A strain collection of 84 Haemophilus spp. comprising 50 H. influenzae, 25 H. parainfluenzae, 7 H. haemolyticus, and 2 H. parahaemolyticus including 77 clinical isolates was analyzed by FISH with newly designed DNA probes, and two different MALDI-TOF-MS systems (Bruker, Shimadzu with and without prior formic acid extraction. PRINCIPAL FINDINGS: Among the 84 Haemophilus strains analyzed, FISH led to 71 correct results (85%, 13 uninterpretable results (15%, and no misidentifications. Shimadzu MALDI-TOF-MS resulted in 59 correct identifications (70%, 19 uninterpretable results (23%, and 6 misidentifications (7%, using colony material applied directly. Bruker MALDI-TOF-MS with prior formic acid extraction led to 74 correct results (88%, 4 uninterpretable results (5% and 6 misidentifications (7%. The Bruker MALDI-TOF-MS misidentifications could be resolved by the addition of a suitable H. haemolyticus reference spectrum to the system's database. In conclusion, no analyzed diagnostic procedure was free of errors. Diagnostic results have to be interpreted carefully and alternative tests should be applied in case of ambiguous test results on isolates from seriously ill patients.

  17. Resolving colocalization of bacteria and metal(loid)s on plant root surfaces by combining fluorescence in situ hybridization (FISH) with multiple-energy micro-focused X-ray fluorescence (ME μXRF).

    Science.gov (United States)

    Honeker, Linnea K; Root, Robert A; Chorover, Jon; Maier, Raina M

    2016-12-01

    Metal(loid)-contamination of the environment due to anthropogenic activities is a global problem. Understanding the fate of contaminants requires elucidation of biotic and abiotic factors that influence metal(loid) speciation from molecular to field scales. Improved methods are needed to assess micro-scale processes, such as those occurring at biogeochemical interfaces between plant tissues, microbial cells, and metal(loid)s. Here we present an advanced method that combines fluorescence in situ hybridization (FISH) with synchrotron-based multiple-energy micro-focused X-ray fluorescence microprobe imaging (ME μXRF) to examine colocalization of bacteria and metal(loid)s on root surfaces of plants used to phytostabilize metalliferous mine tailings. Bacteria were visualized on a small root section using SytoBC nucleic acid stain and FISH probes targeting the domain Bacteria and a specific group (Alphaproteobacteria, Gammaproteobacteria, or Actinobacteria). The same root region was then analyzed for elemental distribution and metal(loid) speciation of As and Fe using ME μXRF. The FISH and ME μXRF images were aligned using ImageJ software to correlate microbiological and geochemical results. Results from quantitative analysis of colocalization show a significantly higher fraction of As colocalized with Fe-oxide plaques on the root surfaces (fraction of overlap 0.49±0.19) than to bacteria (0.072±0.052) (pbacteria that colocalized with metal(loid)s, Actinobacteria, known for their metal tolerance, had a higher correlation with both As and Fe than Alphaproteobacteria or Gammaproteobacteria. This method demonstrates how coupling these micro-techniques can expand our understanding of micro-scale interactions between roots, metal(loid)s and microbes, information that should lead to improved mechanistic models of metal(loid) speciation and fate. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Subtelomeric rearrangements in Indian children with idiopathic intellectual disability/developmental delay: Frequency estimation & clinical correlation using fluorescence in situ hybridization (FISH)

    Science.gov (United States)

    Mohan, Shruthi; Koshy, Teena; Vekatachalam, Perumal; Nampoothiri, Sheela; Yesodharan, Dhanya; Gowrishankar, Kalpana; Kumar, Jeevan; Ravichandran, Latha; Joseph, Santhosh; Chandrasekaran, Anupama; Paul, Solomon F. D.

    2016-01-01

    Background & objectives: Subtelomeres are prone to deleterious rearrangements owing to their proximity to unique sequences on the one end and telomeric repetitive sequences, which increase their tendency to recombine, on the other end. These subtelomeric rearrangements resulting in segmental aneusomy are reported to contribute to the aetiology of idiopathic intellectual disability/developmental delay (ID/DD). We undertook this study to estimate the frequency of subtelomeric rearrangements in children with ID/DD. Methods: One hundred and twenty seven children with idiopathic ID/DD were tested for subtelomeric rearrangements using karyotyping and FISH. Blood samples were cultured, harvested, fixed and GTG-banded using the standard protocols. Results: Rearrangements involving the subtelomeres were observed in 7.8 per cent of the tested samples. Detection of rearrangements visible at the resolution of the karyotype constituted 2.3 per cent, while those rearrangements detected only with FISH constituted 5.5 per cent. Five deletions and five unbalanced translocations were detected. Analysis of parental samples wherever possible was informative regarding the inheritance of the rearrangement. Interpretation & conclusions: The frequency of subtelomeric rearrangements observed in this study was within the reported range of 0-35 per cent. All abnormal genotypes were clinically correlated. Further analysis with array technologies presents a future prospect. Our results suggest the need to test individuals with ID/DD for subtelomeric rearrangements using sensitive methods such as FISH. PMID:27934799

  19. Subtelomeric rearrangements in Indian children with idiopathic intellectual disability/developmental delay: Frequency estimation & clinical correlation using fluorescence in situ hybridization (FISH

    Directory of Open Access Journals (Sweden)

    Shruthi Mohan

    2016-01-01

    Full Text Available Background & objectives: Subtelomeres are prone to deleterious rearrangements owing to their proximity to unique sequences on the one end and telomeric repetitive sequences, which increase their tendency to recombine, on the other end. These subtelomeric rearrangements resulting in segmental aneusomy are reported to contribute to the aetiology of idiopathic intellectual disability/developmental delay (ID/DD. We undertook this study to estimate the frequency of subtelomeric rearrangements in children with ID/DD. Methods: One hundred and twenty seven children with idiopathic ID/DD were tested for subtelomeric rearrangements using karyotyping and FISH. Blood samples were cultured, harvested, fixed and GTG-banded using the standard protocols. Results: Rearrangements involving the subtelomeres were observed in 7.8 per cent of the tested samples. Detection of rearrangements visible at the resolution of the karyotype constituted 2.3 per cent, while those rearrangements detected only with FISH constituted 5.5 per cent. Five deletions and five unbalanced translocations were detected. Analysis of parental samples wherever possible was informative regarding the inheritance of the rearrangement. Interpretation & conclusions: The frequency of subtelomeric rearrangements observed in this study was within the reported range of 0-35 per cent. All abnormal genotypes were clinically correlated. Further analysis with array technologies presents a future prospect. Our results suggest the need to test individuals with ID/DD for subtelomeric rearrangements using sensitive methods such as FISH.

  20. The application of fluorescence in situ hybridization (FISH technique for studying the microbial communities in intestinal tissues of white shrimp (Penaeus vannamei

    Directory of Open Access Journals (Sweden)

    Supamattaya, K.

    2005-02-01

    Full Text Available Fluorescence in situ hybridization technique is very useful for the evaluation of microbial communities in various environments. It is possible to apply this technique to study the intestinal microflora in white shrimp (Penaeus vannamei. Different fixatives and storage temperature were tested in this technique. It was found that fixation with 10% buffered formalin for 12 hours and changed to 70% ethanol shown positive results when compared to the fixation with Davidson's fixative or RF fixative. The best signaling was obtainedfrom the samples which were stored in -20ºC. By using the DNA probe targeted to the Eubacteria domain (EUB338 probe, 5′-GCT GCC TCC CGT AGG AGT-3′ labeled with fluorescein as a hybridizing probe, it was found that most intestinal microflora were aggregated with the intestinal contents, or dispersed in the lumen. There was not evidence of the attachment of the microflora with the intestinal epithelium in this study.

  1. Calorimetric Study of Thermal Denaturation of Superoxide Dismutase

    Institute of Scientific and Technical Information of China (English)

    王邦宁; 谈夫

    1994-01-01

    The thermal denaturation of superoxide dismutase (SOD) from bovine erythrocytes was studied at various pH values of different buffers and at various concentrations of solutions of two neutral salts by differential scanning calorimetry. The experiments performed indicate that the PIPES is a buffer non-coordinating with the SOD, and that the binding of the anions studied influences more or less the thermal denaturation of SOD, but the effect on the oxidation form of SOD is more apparent. A new conformer of SOD with lower thermostability was discovered by the experiments performed in different buffers at certain pH values higher than the isoelectric point of SOD, or at higher concentrations of neutral salt solutions. The new conformer may be converted irreversibly into the usual conformer with high thermostability during heating. Based on the thermodynamic parameters obtained in distilled water and by thermodynamic analysis using the Ooi’s model, it is revealed that the large enthalpy △Hdc contributed by

  2. [Characterization of thermal denaturation process of proteinase K by spectrometry].

    Science.gov (United States)

    Zhang, Qi-Bing; Na, Xin-Zhu; Yin, Zong-Ning

    2013-07-01

    The effect of different temperatures on the activity and conformational changes of proteinase K was studied. Methods Proteinase K was treated with different temperatures, then denatured natural substrate casein was used to assay enzyme activity, steady-state and time-resolved fluorescence spectroscopy was used to study tertiary structure, and circular dichroism was used to study secondary structure. Results show with the temperature rising from 25 to 65 degrees C, the enzyme activity and half-life of proteinase K dropped, maximum emission wavelength red shifted from 335 to 354 nm with fluorescence intensity decreasing. Synchronous fluorescence intensity of tryptophan residues decreased and that of tyrosine residues increased. Fluorescence lifetime of tryptophan residues reduced from 4. 427 1 to 4. 032 4 ns and the fraction of alpha-helix dropped. It was concluded that it is simple and accurate to use steady-state/time-resolved fluorescence spectroscopy and circular dichroism to investigate thermal stability of proteinase K. Thermal denaturation of proteinase K followed a three-state process. Fluorescence intensity of proteinase K was affected by fluorescence resonance energy transfer from tyrosine to tryptophan residues. The alpha-helix was the main structure to maintain conformational stability of enzyme active site of proteinase K.

  3. Denatured Thermodynamics of Proteins in Weak Cation-exchange Chromatography

    Institute of Scientific and Technical Information of China (English)

    LI Rong; CHEN Guo-Liang

    2003-01-01

    The thermostability of some proteins in weak cation-exchange chromatography was investigated at 20-80 ℃. The results show that there is a fixed thermal denaturation transition temperature for each protein. The appearance of the thermal transition temperature indicates that the conformations of the proteins are destroyed seriously. The thermal behavior of the proteins in weak cation-exchange and hydrophobic interaction chromatographies were compared in a wide temperature range. It was found that the proteins have a higher thermostability in a weak cation-exchange chromatography system. The thermodynamic parameters(ΔH0, ΔS0) of those proteins were determined by means of Vant Hoff relationship(lnk-1/T). According to standard entropy change(ΔS0), the conformational change of the proteins was judged in the chromatographic process. The linear relationships between ΔH0 and ΔS0 can be used to evaluate "compensation temperature"(β) at the protein denaturation and identify the identity of the protein retention mechanism in weak cation-exchange chromatography.

  4. Optical real-time measurement of collagen denaturation

    Science.gov (United States)

    Sankaran, Vanitha; Walsh, Joseph T., Jr.

    1997-06-01

    Linear birefringence is a property of collagenous tissue that results from both its composition and structure. Previous investigations have shown that birefringence provides an indication of structural changes in collagen during slow heating. We now report the birefringent response of both mature and young rat tail tendon to laser-heating. The results indicate that denaturation of collagen from mature rats induced by a 200-microsecond(s) -long Ho:YAG laser pulse may not be described accurately by kinetic parameters. Several second-long pulses of CO2 laser pulse may not be described from young rats fit an Arrhenius model with Ea equals 12.1 kcal/mol and A equals e18.03 s-1. Typically, for slow-heating of collagen, Ea equals kcal/mol and A equals e120 s-1. Thus, it seems likely that the temperature and energy needed to initiate collagen denaturation is lower in young collagen, possibly due to its decreased hydroxyproline content and consequent decreased thermal stability.

  5. Second-harmonic generation investigation of collagen thermal denaturation

    Science.gov (United States)

    Chen, Wei-Liang; Sun, Yen; Lin, Sung-Jan; Jee, Shiou-Hwa; Chen, Yang-Fang; Lin, Ling-Chih; So, Peter T. C.; Dong, Chen-Yuan

    2007-02-01

    Using the technique of second-harmonic generation (SHG) microscopy we obtained large area image of type I collagen from rat tail tendon as it is heated from 40°C to 70°C for 0 to 180 minutes. The high resolution images allowed us to investigate the collagen structural change. We observed that heating the tendon below the temperature of 54°C does not produce any change in the averaged SHG intensity. At the heating temperature of 54°C and above, we find that increasing the heating temperature and time leads to decreasing SHG intensity. As the tendon is heated above 54°C, a decrease in the SHG signal occurs uniformly throughout the tendon, but the regions where the SHG signal vanishes form a tiger-tail like pattern. By comparing the relative SHG intensities in small and large areas, we found that the denaturation process responsible for forming the tiger-tail like pattern occurs at a higher rate than the global denaturation process occurring throughout the tendon. Our results show that second-harmonic generation microscopy is effective in monitoring the thermal damage to collagen and has potential applications in biomedicine.

  6. Fish allergy and fish allergens

    DEFF Research Database (Denmark)

    Kuehn, A; Hilger, Christiane; Ollert, Markus

    2016-01-01

    but patients with this phenotype constitute an important sub-group among fish-allergic individuals. 2. Newly identified fish allergens, enolases, aldolases, and fish gelatin, are of high relevance as the majority of the fish-allergic individuals seem to develop specific IgE against these proteins. The present...

  7. Heterogeneity of mitochondrial DNA from Saccharomyces carlsbergensis. Denaturation mapping by electron microscopy.

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C; Bak, AL

    1975-01-01

    Electronmicroscopic observation of the denaturation pattern of 130 partially denaturated linear mitochondrial DNA molecules from Saccharomyces carlsbergensis was used to investigate the distribution of AT-rich sequences within the mitochondrial genome. The molecules were observed after heating...... denaturated sequences in the mitochondrial DNA. These sequences which presumably correspond to the very AT-rich regions, known to exist in the yeast mitochondrial DNA, were found at intervals of about 0.5 - 3 mum on the map....

  8. 27 CFR 21.92 - Denaturants listed as U.S.P. or N.F.

    Science.gov (United States)

    2010-04-01

    ....P. or N.F. 21.92 Section 21.92 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND... for Denaturants § 21.92 Denaturants listed as U.S.P. or N.F. Denaturing materials and products listed in this part as “U.S.P.” or “N.F.” shall meet the specifications set forth in the current...

  9. Detection of denitrification genes by in situ rolling circle amplification - fluorescence in situ hybridization (in situ RCA-FISH) to link metabolic potential with identity inside bacterial cells

    DEFF Research Database (Denmark)

    Hoshino, Tatsuhiko; Schramm, Andreas

    2010-01-01

    A target-primed in situ rolling circle amplification (in situ RCA) protocol was developed for detection of single-copy genes inside bacterial cells and optimized with Pseudomonas stutzeri, targeting nitrite and nitrous oxide reductase genes (nirS and nosZ). Two padlock probes were designed per gene......). Nevertheless, multiple genes (nirS and nosZ; nirS and the 16S rRNA gene) could be detected simultaneously in P. stutzeri. Environmental application of in situ RCA-FISH was demonstrated on activated sludge by the differential detection of two types of nirS-defined denitrifiers; one of them was identified...

  10. Electrochemical behaviour of denatured ethanol for use in direct ethanol fuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Bayer, Domnik; Kintzel, Birgit; Joos, Martin; Cremers, Carsten; Tuebke, Jens [Fraunhofer-Institut fuer Chemische Technologie (ICT), Pfinztal (Germany)

    2010-07-01

    In the present study, two denaturing agents, an adjuvant to a denaturing agent and a mixture of a denaturing agend and the adjuvant were tested with regard to their fuel cell compatibility. Therefore, various electrochemical tests including cyclic voltammetry, chronoamperometry and differential electrochemical mass spectroscopy have been conducted at platinum as model catalyst in acidic and alkaline medium. In addition, the most promising denaturing agent, a mixture of tert-butyl ethyl ether (ETBE) with the adjuvant Bitrex {sup registered}, has also been tested at commercial fuel cell catalysts in both acidic and alkaline media. (orig.)

  11. Visualization of early events in acetic acid denaturation of HIV-1 protease: a molecular dynamics study.

    Directory of Open Access Journals (Sweden)

    Aditi Narendra Borkar

    Full Text Available Protein denaturation plays a crucial role in cellular processes. In this study, denaturation of HIV-1 Protease (PR was investigated by all-atom MD simulations in explicit solvent. The PR dimer and monomer were simulated separately in 9 M acetic acid (9 M AcOH solution and water to study the denaturation process of PR in acetic acid environment. Direct visualization of the denaturation dynamics that is readily available from such simulations has been presented. Our simulations in 9 M AcOH reveal that the PR denaturation begins by separation of dimer into intact monomers and it is only after this separation that the monomer units start denaturing. The denaturation of the monomers is flagged off by the loss of crucial interactions between the α-helix at C-terminal and surrounding β-strands. This causes the structure to transit from the equilibrium dynamics to random non-equilibrating dynamics. Residence time calculations indicate that denaturation occurs via direct interaction of the acetic acid molecules with certain regions of the protein in 9 M AcOH. All these observations have helped to decipher a picture of the early events in acetic acid denaturation of PR and have illustrated that the α-helix and the β-sheet at the C-terminus of a native and functional PR dimer should maintain both the stability and the function of the enzyme and thus present newer targets for blocking PR function.

  12. Cold denaturation induces inversion of dipole and spin transfer in chiral peptide monolayers

    Science.gov (United States)

    Eckshtain-Levi, Meital; Capua, Eyal; Refaely-Abramson, Sivan; Sarkar, Soumyajit; Gavrilov, Yulian; Mathew, Shinto P.; Paltiel, Yossi; Levy, Yaakov; Kronik, Leeor; Naaman, Ron

    2016-02-01

    Chirality-induced spin selectivity is a recently-discovered effect, which results in spin selectivity for electrons transmitted through chiral peptide monolayers. Here, we use this spin selectivity to probe the organization of self-assembled α-helix peptide monolayers and examine the relation between structural and spin transfer phenomena. We show that the α-helix structure of oligopeptides based on alanine and aminoisobutyric acid is transformed to a more linear one upon cooling. This process is similar to the known cold denaturation in peptides, but here the self-assembled monolayer plays the role of the solvent. The structural change results in a flip in the direction of the electrical dipole moment of the adsorbed molecules. The dipole flip is accompanied by a concomitant change in the spin that is preferred in electron transfer through the molecules, observed via a new solid-state hybrid organic-inorganic device that is based on the Hall effect, but operates with no external magnetic field or magnetic material.

  13. PMC- FISHs in Hybrids of Tetraploid Cultivated and Diploid Wild Gossypium Species%四倍体栽培棉与二倍体野生棉杂种PMC-FISH研究

    Institute of Scientific and Technical Information of China (English)

    凌键; 王坤波; 邹美娟; 宋国立; 王春英; 彭仁海; 刘方; 黎绍惠; 张香娣; 王玉红

    2009-01-01

    首次将PMC-FISH(花粉母细胞荧光原位杂交)技术应用于四倍体栽培棉与二倍体野生棉杂交所形成的三倍体杂种棉F1中,成功的鉴定了目标染色体中的单价体、双价体、多价体,还发现在非目标染色体上有星状和片段状的杂交信号.在以A组棉基因组DNA作为探针的PMC-FISH中,分别对三倍体杂种棉F1及其母本四倍体栽培棉的减数分裂中期I的染色体构型进行了鉴定.结果显示,四个三倍体杂种棉F1的减数分裂时期染色体的构型分别是:陆地棉(AD)1×雷蒙德氏棉(D5)为1IIIaad + 1IIaa + 4IIad + 7IIdd + 5Ⅰa + 7Ⅰd;海岛棉(AD)2×旱地棉(D4)为1Ⅴaaaad +1IIIadd + 2IIad + 8IIdd + 6Ⅰa + 5Ⅰd;陆地棉(AD)1×澳洲棉(C3)为2IIIadd (adc or acc) + 1IIaa + 9Ⅰa + 4IIcc (dc or dd) + 14Ⅰd (c);陆地棉(AD)1×南岱华棉(C1-n)为:1IIaa + 1IIad (ac)+ 10Ⅰa + 6IIcc (dc or dd) + 13Ⅰd (c).然而,两个四倍体棉染色体的构型都是13IIaa + 13IIdd.上述结果还显示, AD×D的两个杂种组合中,二价体多,单价体少,AD×C的两个杂种组合中,二价体少,单价体多;并且AD×D型杂种中A亚组染色体单体的数目比其在AD×C型杂种中的更少.这说明,与C染色体组相比较,D染色体组与四倍体棉种的亲缘关系更近.同时,我们的结果也证明PMC-FISH技术在分析三倍体杂种棉F1的亲缘关系中起着不可取代的作用.%PMC-FISH (pollen mother cell-fluorescence in situ hybridization) technology was firstly applied to identify probed chromosomes in univalent, bivalent and multivalent, as well as to distinguish star- or fragment-like hybridization signals from non-probed chromosomes in four triploid F1 hybrids produced by cultivated allotetraploid cotton crossed with different diploid cotton species in the genus Gossypium. Using genomic DNA from A genome species as probe, the meiotic chromosome configurations in metaphase Ⅰof four triploid F1 hybrids were identified as 1Ⅲaad + 1Ⅱaa + 4

  14. Antarctic Fishes.

    Science.gov (United States)

    Eastman, Joseph T.; DeVries, Arthur L.

    1986-01-01

    Explains the adaptations to Antarctic waters that Notothenioidei, a group of advanced bony fishes, have exhibited. Discusses the fishes' mechanisms of production of antifreeze properties and their capacities for neutral buoyancy in water. (ML)

  15. Evaluating seasonal dynamics of bacterial communities in marine fish aquaculture: a preliminary study before applying phage therapy.

    Science.gov (United States)

    Pereira, Carla; Salvador, Sara; Arrojado, Cátia; Silva, Yolanda; Santos, Ana L; Cunha, Angela; Gomes, Newton C M; Gomes, Newton; Almeida, Adelaide

    2011-04-01

    The increasing problem of antibiotic resistance in common pathogenic bacteria and the concern about the spreading of antibiotics in the environment bring the need to find new methods to control fish pathogens. Phage therapy represents a potential alternative to antibiotics, but its use in aquaculture requires a detailed understanding of bacterial communities, namely of fish pathogenic bacteria. Therefore, in this study the seasonal dynamics of the overall bacterial communities, microbiological water quality and disease-causing bacteria were followed in a marine aquaculture system of Ria de Aveiro (Portugal). Analysis of the bacterial diversity of the water samples by denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments indicates that the bacterial community structure varied seasonally, showing a higher complexity during the warm season. The diversity of the main fish pathogenic bacteria, assessed by DGGE targeting the Vibrio genus, showed lower seasonal variation, with new dominating populations appearing mainly in the spring. Bacterial indicators, faecal coliforms and enterococci, enumerated by the filter-membrane method, also varied seasonally. The fluorescent in situ hybridization (FISH) results showed that the specific groups of bacteria varied during the study period and that the non-indigenous Enterobactereaceae family was the most abundant group followed by Vibrio and Aeromonas. The seasonal variation detected in terms of density and structure of total and pathogenic bacterial communities demonstrates the need for a careful monitoring of water through the year in order to select the suitable phages to inactivate fish pathogenic bacteria. The spring season seems to be the critical time period when phage therapy should be applied.

  16. Consistent View of Polypeptide Chain Expansion in Chemical Denaturants from Multiple Experimental Methods.

    Science.gov (United States)

    Borgia, Alessandro; Zheng, Wenwei; Buholzer, Karin; Borgia, Madeleine B; Schüler, Anja; Hofmann, Hagen; Soranno, Andrea; Nettels, Daniel; Gast, Klaus; Grishaev, Alexander; Best, Robert B; Schuler, Benjamin

    2016-09-14

    There has been a long-standing controversy regarding the effect of chemical denaturants on the dimensions of unfolded and intrinsically disordered proteins: A wide range of experimental techniques suggest that polypeptide chains expand with increasing denaturant concentration, but several studies using small-angle X-ray scattering (SAXS) have reported no such increase of the radius of gyration (Rg). This inconsistency challenges our current understanding of the mechanism of chemical denaturants, which are widely employed to investigate protein folding and stability. Here, we use a combination of single-molecule Förster resonance energy transfer (FRET), SAXS, dynamic light scattering (DLS), and two-focus fluorescence correlation spectroscopy (2f-FCS) to characterize the denaturant dependence of the unfolded state of the spectrin domain R17 and the intrinsically disordered protein ACTR in two different denaturants. Standard analysis of the primary data clearly indicates an expansion of the unfolded state with increasing denaturant concentration irrespective of the protein, denaturant, or experimental method used. This is the first case in which SAXS and FRET have yielded even qualitatively consistent results regarding expansion in denaturant when applied to the same proteins. To more directly illustrate this self-consistency, we used both SAXS and FRET data in a Bayesian procedure to refine structural ensembles representative of the observed unfolded state. This analysis demonstrates that both of these experimental probes are compatible with a common ensemble of protein configurations for each denaturant concentration. Furthermore, the resulting ensembles reproduce the trend of increasing hydrodynamic radius with denaturant concentration obtained by 2f-FCS and DLS. We were thus able to reconcile the results from all four experimental techniques quantitatively, to obtain a comprehensive structural picture of denaturant-induced unfolded state expansion, and to

  17. Irreversible denaturation of maltodextrin glucosidase studied by differential scanning calorimetry, circular dichroism, and turbidity measurements.

    Science.gov (United States)

    Goyal, Megha; Chaudhuri, Tapan K; Kuwajima, Kunihiro

    2014-01-01

    Thermal denaturation of Escherichia coli maltodextrin glucosidase was studied by differential scanning calorimetry, circular dichroism (230 nm), and UV-absorption measurements (340 nm), which were respectively used to monitor heat absorption, conformational unfolding, and the production of solution turbidity. The denaturation was irreversible, and the thermal transition recorded at scan rates of 0.5-1.5 K/min was significantly scan-rate dependent, indicating that the thermal denaturation was kinetically controlled. The absence of a protein-concentration effect on the thermal transition indicated that the denaturation was rate-limited by a mono-molecular process. From the analysis of the calorimetric thermograms, a one-step irreversible model well represented the thermal denaturation of the protein. The calorimetrically observed thermal transitions showed excellent coincidence with the turbidity transitions monitored by UV-absorption as well as with the unfolding transitions monitored by circular dichroism. The thermal denaturation of the protein was thus rate-limited by conformational unfolding, which was followed by a rapid irreversible formation of aggregates that produced the solution turbidity. It is thus important to note that the absence of the protein-concentration effect on the irreversible thermal denaturation does not necessarily means the absence of protein aggregation itself. The turbidity measurements together with differential scanning calorimetry in the irreversible thermal denaturation of the protein provided a very effective approach for understanding the mechanisms of the irreversible denaturation. The Arrhenius-equation parameters obtained from analysis of the thermal denaturation were compared with those of other proteins that have been reported to show the one-step irreversible thermal denaturation. Maltodextrin glucosidase had sufficiently high kinetic stability with a half-life of 68 days at a physiological temperature (37°C).

  18. Fish Dishes.

    Science.gov (United States)

    Derby, Marie

    2003-01-01

    Describes an art project that was inspired by Greek pottery, specifically dishes shaped as fish. Explains that fourth-grade students drew a fish shape that was later used to create their clay version of the fish. Discusses how the students examined the pottery to make decisions about color and design. (CMK)

  19. Evaluation of the damage in fish spermatozoa cryopreservation

    Institute of Scientific and Technical Information of China (English)

    LI Jun; LIU Qinghua; ZHANG Shicui

    2006-01-01

    Cryodamages occur during sperm cryopreservation. Cryopreservation of fish sperm usually results in marked decrease in sperm quality, such as swelling or disruption of the plasma membrane, mitochondrial dysfunction, diminished sperm motility, impaired velocity, shorter motility period, denaturation, and release of some enzymes from spermatozoa. In this paper, damages in morphology, physiology,biochemistry and metabolism, and genetic integrity of fish semen after cryopreservation are discussed.New approaches in assessment of fish thawed sperm quality such as computer assisted sperm analysis,flow cytometic analysis combined with fluorescent probes and single cell gel electrophoresis are also briefly reviewed.

  20. Residual ordered structure in denatured proteins and the problem of protein folding.

    Science.gov (United States)

    Basharov, Mahmud A

    2012-02-01

    Structural characteristics of numerous globular proteins in the denatured state have been reviewed using literature data. Recent more precise experiments show that in contrast to the conventional standpoint, proteins under strongly denaturing conditions do not unfold completely and adopt a random coil state, but contain significant residual ordered structure. These results cast doubt on the basis of the conventional approach representing the process of protein folding as a spontaneous transition of a polypeptide chain from the random coil state to the unique globular structure. The denaturation of proteins is explained in terms of the physical properties of proteins such as stability, conformational change, elasticity, irreversible denaturation, etc. The spontaneous renaturation of some denatured proteins most probably is merely the manifestation of the physical properties (e.g., the elasticity) of the proteins per se, caused by the residual structure present in the denatured state. The pieces of the ordered structure might be the centers of the initiation of renaturation, where the restoration of the initial native conformation of denatured proteins begins. Studies on the denaturation of proteins hardly clarify how the proteins fold into the native conformation during the successive residue-by-residue elongation of the polypeptide chain on the ribosome.

  1. The generation of denatured reactor plutonium by different options of the fuel cycle

    Energy Technology Data Exchange (ETDEWEB)

    Broeders, C.H.M.; Kessler, G. [Inst. for Neutron Physics and Reactor Technology, Research Center Karlsruhe (Germany)

    2006-11-15

    Denatured (proliferation resistant) reactor plutonium can be generated in a number of different fuel cycle options. First denatured reactor plutonium can be obtained if, instead of low enriched U-235 PWR fuel, re-enriched U-235/U-236 from reprocessed uranium is used (fuel type A). Also the envisaged existing 2,500 t of reactor plutonium (being generated world wide up to the year 2010), mostly stored in intermediate fuel storage facilities at present, could be converted during a transition phase into denatured reactor plutonium by the options fuel type B and D. Denatured reactor plutonium could have the same safeguards standard as present low enriched (<20% U-235) LWR fuel. It could be incinerated by recycling once or twice in PWRs and subsequently by multi-recycling in FRs (CAPRA type or IFRs). Once denatured, such reactor plutonium could remain denatured during multiple recycling. In a PWR, e.g., denatured reactor plutonium could be destroyed at a rate of about 250 kg/GWey. While denatured reactor plutonium could be recycled and incinerated under relieved IAEA safeguards, neptunium would still have to be monitored by the IAEA in future for all cases in which considerable amounts of neptunium are produced. (orig.)

  2. In Vitro Reassembly of Tobacco Ribulose-1,5-bisphosphate Carboxylase/ Oxygenase from Fully Denatured Subunits

    Institute of Scientific and Technical Information of China (English)

    Zhen-Hua YONG; Gen-Yun CHEN; Jiao-Nai SHI; Da-Quan XU

    2006-01-01

    It has been generally proved impossible to reassemble ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) from fully denatured subunits in vitro in higher plant, because large subunit of fully denatured Rubisco is liable to precipitate when the denaturant is removed by common methods of direct dilution and one-step dialysis. In our experiment, the problem of precipitation was resolved by an improved gradual dialysis method, which gradually decreased the concentration of denaturant. However, fully denatured Rubisco subunits still could not be reassembled into holoenzyme using gradual dialysis unless chaperonin 60was added. The restored activity of reassembled Rubisco was approximately 8% of natural enzyme. The quantity of reassembled Rubisco increased greatly when heat shock protein 70 was present in the reassembly process. ATP and Mg2+ were unnecessary for in vitro reassembly of Rubisco, and Mg2+ inhibited the reassembly process. The reassembly was weakened when ATP, Mg2+ and K+ existed together in the reassembly process.

  3. Dynamic light scattering study of peanut agglutinin: Size, shape and urea denaturation

    Indian Academy of Sciences (India)

    Sagarika Dev; Avadhesha Surolia

    2006-12-01

    Peanut agglutinin (PNA) is a homotetrameric protein with a unique open quaternary structure. PNA shows non-two state profile in chaotrope induced denaturation. It passes through a monomeric molten globule like state before complete denaturation (Reddy et al 1999). This denaturation profile is associated with the change in hydrodynamic radius of the native protein. Though the molten globule-like state is monomeric in nature it expands in size due to partial denaturation. The size and shape of the native PNA as well as the change in hydrodynamic radius of the protein during denaturation has been studied by dynamic light scattering (DLS). The generation of two species is evident from the profile of hydrodynamic radii. This study also reveals the extent of compactness of the intermediate state.

  4. In situ observation of collagen thermal denaturation by second harmonic generation microscopy

    Science.gov (United States)

    Liao, C.-S.; Zhuo, Z.-Y.; Yu, J.-Y.; Chao, P.-H. G.; Chu, S.-W.

    2010-02-01

    Collagen denaturation is of fundamental importance for clinical treatment. Conventionally, the denaturation process is quantified by the shrinkage of collagen fibers, but the underlying molecular origin has not been fully understood. Since second harmonic generation (SHG) is related to the molecular packing of the triple helix in collagen fibers, this nonlinear signal provides an insight of molecular dynamics during thermal denaturation. With the aid of SHG microscopy, we found a new step in collagen thermal denaturation process, de-crimp. During the de-crimp step, the characteristic crimp pattern of collagen fascicles disappeared due to the breakage of interconnecting bonds between collagen fibrils, while SHG intensity remained unchanged, suggesting the intactness of the triple helical molecules. At higher temperature, shrinkage is observed with strongly reduced SHG intensity, indicating denaturation at the molecular level.

  5. Refolding of detergent-denatured lysozyme using β-cyclodextrin-assisted ion exchange chromatography.

    Science.gov (United States)

    Zhang, Li; Zhang, Qinming; Wang, Chaozhan

    2013-03-01

    Chromatography-based protein refolding is widely used. Detergent is increasingly used for protein solubilization from inclusion bodies. Therefore, it is necessary to develop a refolding method for detergent-denatured/solubilized proteins based on liquid chromatography. In the present work, sarkosyl-denatured/dithiothreitol-reduced lysozyme was used as a model, and a refolding method based on ion exchange chromatography, assisted by β-cyclodextrin, was developed for refolding detergent-denatured proteins. Many factors affecting the refolding, such as concentration of urea, concentration of β-cyclodextrin, pH and flow rate of mobile phases, were investigated to optimize the refolding conditions for sarkosyl-denatured lysozymes. The results showed that the sarkosyl-denatured lysozyme could be successfully refolded using β-cyclodextrin-assisted ion exchange chromatography.

  6. 应用FISH-AFLP技术分析平欧杂种榛主栽品种的遗传关系%Analysis of the Genetic Relationship of the Main Cultivars of Ping’ou Hybrid Hazelnut (C. heterophylla×C. avellana) by FISH-AFLP Markers

    Institute of Scientific and Technical Information of China (English)

    马庆华; 陈新; 赵天田; 刘庆忠; 王贵禧

    2013-01-01

    [目的]建立平欧杂种榛FISH-AFLP技术体系,并应用该技术分析平欧杂种榛主栽品种的遗传关系。[方法]以达维等10个主栽品种为试材,经 DNA 提取、PstⅠ/MseⅠ双酶切,进行 FISH-AFLP 反应,数据转换为“0-1”矩阵后,使用NTSYS pc 2.11F和Popgene 1.32软件进行数据分析并作图。[结果]从64对引物中,筛选出15对多态性强的PstⅠ/MseⅠ引物,共获得1739条谱带,引物平均多态带比率97.94%;平欧杂种榛主栽品种的相似性系数为0.7556-0.8543,当阈值为0.8398时,可分成4个AFLP群,其中,玉坠(84-310)、辽榛4号(85-41)和平欧69号(84-69)单独为1群,其他品种为1群;平欧杂种榛主栽品种的有效等位基因数、基因多样度、Shannon信息指数分别为1.3921、0.2482和0.3957,具有较高的遗传多样性;研究获得的特征条带可用于平欧杂种榛主栽品种的快速鉴别。[结论]由于平欧杂种榛的实生选优群体是多亲本组合的混合后代,因此,该群体是一个具有高度遗传多样性的群体,主栽品种间存在复杂的亲缘关系;研究构建的FISH-AFLP技术流程、筛选得到的引物以及分析得到的各项遗传参数,可为其他榛属植物的相关研究提供参考。%[Objective]The objective of this study is to establish FISH-AFLP analysis system for Ping’ou hybrid hazelnut and analyze the genetic relationship of the main cultivars.[Method]Ten main cultivars, such as Dawei, were selected as materials. High-quality genomic DNA was extracted with improved CTAB method, and the purified DNA samples were digested with PstⅠ/MseⅠ. After ligation reaction, the samples were used to perform FISH-AFLP detection. The information of all bands was converted into“0-1”matrix. NTSYS pc 2.11F and Popgene 1.32 software were used to conduct the data analysis and plotting.[Result]The results showed that:15 PstⅠ/MseⅠ primer pairs were selected from 64

  7. Breaking the host range: mandarin fish is susceptible to a vesiculovirus derived from snakehead fish.

    Science.gov (United States)

    Liu, Xiaodan; Wen, Yi; Hu, Xianqin; Wang, Wenwen; Liang, Xufang; Li, Jun; Vakharia, Vikram; Lin, Li

    2015-04-01

    Members of the genus Vesiculovirus, which belongs to the family Rhabdoviridae, can cause great economic loss in fish culture. In the present report, a vesiculovirus [named snakehead fish vesiculovirus (SHVV)] was isolated from diseased hybrid snakehead fish. SHVV shared 94 % nucleotide sequence identity at the genomic level with Siniperca chuatsi rhabdovirus (SCRV), which infects mandarin fish (S. chuatsi). We showed that SHVV was able to replicate and proliferate well in SSN-1 cells, which originate from striped snakehead fish (Channa striatus). Furthermore, mandarin fish was susceptible to SHVV by bath exposure, as well as by intraperitoneal injection. The infected fish showed typical clinical signs of rhabdovirus infection, including haemorrhage and oedema. Histopathological analysis revealed that extensive inflammation and necrosis were observed in the spleen, kidney, liver, heart and brain of the moribund mandarin fish. These results will shed new light on the epidemic of vesiculovirus infections among fish.

  8. Community analysis of ammonia oxidizer in the oxygen-limited nitritation stage of OLAND system by DGGE of PCR amplified 16S rDNA Fragments and FISH

    Institute of Scientific and Technical Information of China (English)

    ZHANG Dan; ZHANG De-min; LIU Yao-ping; CAO Wen-wei; CHEN Guan-xiong

    2004-01-01

    OLAND(oxygen limited autotrophic nitrification and denitrification) nitrogen removal system was constructed by coupling with oxygen limited nitritation stage and anaerobic ammonium oxidation stage. Ammonia oxidizer, as a kind of key bacteria in N cycle, plays an important role at the oxygen limited nitritation stage of OLAND nitrogen removal system. In this study, specific amplification of 16S rDNA fragment of ammonia oxidizer by nested PCR, separation of mixed PCR samples by denaturing gradient gel electrophoresis(DGGE), and the quantification of ammonia oxidizer by Fluorescence in situ hybridization(FISH) were combined to investigate the shifts of community composition and quantity of ammonia oxidizer of the oxygen limited nitritation stage in OLAND system. It showed that the community composition of ammonia oxidizer changed drastically when dissolved oxygen was decreased gradually, and the dominant ammonia oxidizer of the steady nitrite accumulation stage were completely different from that of the early stage of oxygen limited nitritation identified by DGGE . It was concluded that the Nitrosomonas may be the dominant genus of ammonia oxidizer at the oxygen limited nitritation stage of OLAND system characterized by nested PCR-DGGE and FISH, and the percentage of Nitrosomonas was 72.5% ( 0.8% of ammonia oxidizer at the steady nitrite accumulation stage detected by FISH.

  9. Annual trends in catchability and fish stock assessments

    DEFF Research Database (Denmark)

    Marchal, P.; Ulrich, Clara; Korsbrekke, K.;

    2003-01-01

    A key assumption of many fish stock assessment models is that catchability is constant over time. We assume here that trends in catchability may occur through fishing power creeping. The tuning fleets, which are prone to fishing power development, may be identified using the "Hybrid" method...

  10. Dissociative mechanism of F-actin thermal denaturation.

    Science.gov (United States)

    Mikhailova, V V; Kurganov, B I; Pivovarova, A V; Levitsky, D I

    2006-11-01

    We have applied differential scanning calorimetry to investigate thermal unfolding of F-actin. It has been shown that the thermal stability of F-actin strongly depends on ADP concentration. The transition temperature, T(m), increases with increasing ADP concentration up to 1 mM. The T(m) value also depends on the concentration of F-actin: it increases by almost 3 degrees C as the F-actin concentration is increased from 0.5 to 2.0 mg/ml. Similar dependence of the T(m) value on protein concentration was demonstrated for F-actin stabilized by phalloidin, whereas it was much less pronounced in the presence of AlF4(-). However, T(m) was independent of protein concentration in the case of monomeric G-actin. The results suggest that at least two reversible stages precede irreversible thermal denaturation of F-actin; one of them is dissociation of ADP from actin subunits, and another is dissociation of subunits from the ends of actin filaments. The model explains why unfolding of F-actin depends on both ADP and protein concentration.

  11. Nonisothermal denaturation kinetics of human hair and the effects of oxidation.

    Science.gov (United States)

    Wortmann, F-J; Popescu, C; Sendelbach, G

    2006-12-15

    Human hair as alpha-keratin fiber exhibits a complex morphology, which for the context of this investigation is considered as a filament/matrix-composite, comprising the intermediate filaments (IF) and a variety of amorphous protein components as matrix. Differential scanning calorimetry (DSC) under aqueous conditions was used to analyze the denaturation of the alpha-helical material in the IFs and to assess the changes imparted by repeated, oxidative bleaching processes. The DSC curves were submitted to kinetic analysis by applying the Friedman method and assuming first order kinetics. It was found that the course of the denaturation process remains largely unchanged through oxidation, despite the fact that pronounced decreases of denaturation temperature as well as of enthalpy occur. In parallel, the reaction rate constant at the denaturation temperature, k(TD), increases with repeated treatments, that is with cumulative chemical modification. However, this effect is in fact small compared to the overall change of k(T) through the denaturation process. This leads to conclude that once the temperature rise in combination with the chemical change has induced a suitable drop of the viscosity of the matrix around the IFs, denaturation of the remaining helical material occurs along a pathway that is largely independent of temperature and of the pretreatment history. This emphasizes the kinetic control of the matrix over the denaturation process of the helical segments in the filament/matrix composite. Copyright 2006 Wiley Periodicals, Inc.

  12. Studies on the refolding of the reduced-denatured insulin with size exclusion chromatography

    Institute of Scientific and Technical Information of China (English)

    BAI Quan; KONG Yu; DONG Cuihua; GENG Xindu

    2005-01-01

    The refolding of the reduced-denatured insulin from bovine pancreas was investigated with the size exclusion chromatography (SEC). It was shown that the reduced-denatured insulin originally denatured with 7.0 mol·L-1 guanidine hydrochloride (GuHCl) or 8.0 mol·L-1 urea could not be refolded with a non-oxidized mobile phase. Although the oxidized and reduced glutathione (GSSG and GSH) were employed in the oxidized mobile phase, the reduced-denatured insulin still could not be renatured. However, in the presence of 2.0 mol·L-1 urea in the oxidized mobile phase employed, the reduced-denatured insulin can be refolded with SEC, and the aggregation of denatured insulin can be diminished by urea. In addition, the disulfide exchange of reduced-denatured insulin also can be accelerated with GSSG/GSH in the oxidized mobile phase. The three disulfide bridges of insulin were formed correctly and the reduced-unfolded insulin can be renatured completely. The results were further tested with reversed-phase liquid chromatography (RPLC) and hydrophobic interaction chromatography (HIC).

  13. Le diagnostic anténatal de la trisomie 21 par l'hybridation in situ en fluorescence (FISH): à propos des premiers tests réalisés au Maroc

    Science.gov (United States)

    Lamzouri, Afaf; Natiq, Abdelhafid; Tajir, Mariam; Sendid, Mohamed; Sefiani, Abdelaziz

    2012-01-01

    Introduction Le but de cette étude était de présenter les premiers résultats de diagnostic anténatal de la trisomie 21 par la technique d'hybridation in situ en fluorescence (FISH) au Maroc et discuter son intérêt dans le diagnostic rapide de cette aneuploïdie. Méthodes Ce travail a été réalisé chez 23 femmes avec des grossesses à haut risque de trisomie 21. La moyenne d’âge des gestantes étaient de 37,43 ans avec des extrêmes de 21 et 43 ans. Toutes étaient musulmanes mariées, mariage légitimé par la Charia, dont trois mariages consanguins, sauf une originaire de la République Démocratique du Congo qui était chrétienne et concubine. La majorité des femmes étaient fonctionnaires et avaient un niveau de scolarisation moyen à élevé. Toutes les patientes ont bénéficié d'une consultation de génétique médicale au cours de laquelle il leur a été donné des informations sur la technique, son intérêt et ses limites. Il s'agit de femmes enceintes qui avaient soit un âge maternel élevé ou des signes d'appel échographiques et/ ou biochimiques. Une des patientes était porteuse d'une translocation robertsonienne t(14;21) équilibrée. Une amniocentèse a été réalisée chez toutes les gestantes et aucun avortement n'a était induit par ce geste invasif. L’âge gestationnel moyen à la première consultation était de 14 semaines d'aménorrhée (SA) et à l'amniocentèse était de 16 SA et 5 jours. L'analyse FISH a été réalisée, après consentement des couples, sur des cellules non cultivées à partir des échantillons de liquides amniotiques, en utilisant des sondes spécifiques du chromosome 21. Résultats Parmi les 23 patientes qui ont bénéficiées d'un diagnostic anténatal de la trisomie 21 par la technique FISH, nous avons pu rassurer 21 d'entre elles, et nous avons détecté deux cas de trisomie 21 fœtal. Conclusion La technique FISH permet un diagnostic anténatal rapide, en moins de 48h, de la trisomie 21 sur

  14. Assessment of collagen crosslinking and denaturation for the design of regenerative scaffolds.

    Science.gov (United States)

    Madaghiele, Marta; Calò, Emanuela; Salvatore, Luca; Bonfrate, Valentina; Pedone, Deborah; Frigione, Mariaenrica; Sannino, Alessandro

    2016-01-01

    Crosslinking and denaturation were two variables that deeply affected the performance of collagen-based scaffolds designed for tissue regeneration. If crosslinking enhances the mechanical properties and the enzymatic resistance of collagen, while masking or reducing the available cell binding sites, denaturation has very opposite effects, as it impairs the mechanical and the enzymatic stability of collagen, but increases the number of exposed cell adhesive domains. The quantification of both crosslinking and denaturation was thus fundamental to the design of collagen-based scaffolds for selected applications. The aim of this work was to investigate the extents of crosslinking and denaturation of collagen-based films upon dehydrothermal (DHT) treatment, that is, one of the most commonly employed methods for zero-length crosslinking that shows the unique ability to induce partial denaturation. Swelling measurements, differential scanning calorimetry, Fourier transform infrared spectroscopy, colorimetric assays for the quantification of primary amines, and mechanical tests were performed to analyze the effect of the DHT temperature on crosslinking and denaturation. In particular, chemically effective and elastically effective crosslink densities were evaluated. Both crosslinking and denaturation were found to increase with the DHT temperature, although according to different trends. The results also showed that DHT treatments performed at temperatures up to 120°C maintained the extent of denaturation under 25%. Coupling a mild DHT treatment with further crosslinking may thus be very useful not only to modulate the crosslink density, but also to induce a limited amount of denaturation, which shows potential to partially compensate the loss of cell binding sites caused by crosslinking.

  15. Urea-temperature phase diagrams capture the thermodynamics of denatured state expansion that accompany protein unfolding.

    Science.gov (United States)

    Tischer, Alexander; Auton, Matthew

    2013-09-01

    We have analyzed the thermodynamic properties of the von Willebrand factor (VWF) A3 domain using urea-induced unfolding at variable temperature and thermal unfolding at variable urea concentrations to generate a phase diagram that quantitatively describes the equilibrium between native and denatured states. From this analysis, we were able to determine consistent thermodynamic parameters with various spectroscopic and calorimetric methods that define the urea-temperature parameter plane from cold denaturation to heat denaturation. Urea and thermal denaturation are experimentally reversible and independent of the thermal scan rate indicating that all transitions are at equilibrium and the van't Hoff and calorimetric enthalpies obtained from analysis of individual thermal transitions are equivalent demonstrating two-state character. Global analysis of the urea-temperature phase diagram results in a significantly higher enthalpy of unfolding than obtained from analysis of individual thermal transitions and significant cross correlations describing the urea dependence of ΔH0 and ΔCP0 that define a complex temperature dependence of the m-value. Circular dichroism (CD) spectroscopy illustrates a large increase in secondary structure content of the urea-denatured state as temperature increases and a loss of secondary structure in the thermally denatured state upon addition of urea. These structural changes in the denatured ensemble make up ∼40% of the total ellipticity change indicating a highly compact thermally denatured state. The difference between the thermodynamic parameters obtained from phase diagram analysis and those obtained from analysis of individual thermal transitions illustrates that phase diagrams capture both contributions to unfolding and denatured state expansion and by comparison are able to decipher these contributions.

  16. Fish health and fish quality

    DEFF Research Database (Denmark)

    Ingerslev, Hans-Christian

    Aquaculture is an expanding worldwide industry producing an increasing amount of fish every year. The quality of the fish meat is dependent upon many biological and non-biological factors. Infectious diseases are known to cause bleedings and damage of the muscle tissue that may lead to scarring...... are poorly described in fish. The present work in this thesis focused on: 1) examination of potential changes in the quality regarding texture of the muscle tissue in rainbow trout (Oncorhynchus mykiss) after previous infection with the bacterial pathogens Yersinia ruckeri and Vibrio anguillarum; 2...... of these studies showed that previous infections by Yersinia ruckeri and Vibrio anguillarum gave rise to subsequent changes regarding textural quality parameters in fresh fish meat, while no differences were seen for cold-smoked meat from the same fish. The texture in previous infected fish was less flaky and less...

  17. Relaxation rate for an ultrafast folding protein is independent of chemical denaturant concentration.

    Science.gov (United States)

    Cellmer, Troy; Henry, Eric R; Kubelka, Jan; Hofrichter, James; Eaton, William A

    2007-11-28

    The connection between free-energy surfaces and chevron plots has been investigated in a laser temperature jump kinetic study of a small ultrafast folding protein, the 35-residue subdomain from the villin headpiece. Unlike all other proteins that have been studied so far, no measurable dependence of the unfolding/refolding relaxation rate on denaturant concentration was observed over a wide range of guanidinium chloride concentration. Analysis with a simple Ising-like theoretical model shows that this denaturant-invariant relaxation rate can be explained by a large movement of the major free energy barrier, together with a denaturant- and reaction coordinate-dependent diffusion coefficient.

  18. Heat Denaturation of Protein Structures and Chlorophyll States in PSII Membranes

    Institute of Scientific and Technical Information of China (English)

    李冬海; 阮翔; 许强; 王可玢; 公衍道; 匡廷云; 赵南明

    2002-01-01

    Heat denaturation is an important technique in the study of the structure and function of photosynthetic proteins. Heat denaturation of photosystem II (PSII) membrane was studied using circular dichroism (CD) spectroscopy, differential scanning calorimetry (DSC) and oxygen electrode. Complete loss of oxygen-evolving activity of the PSII membrane was observed at temperatures below 45℃. The decrease of excitonic interaction between chlorophyll molecules occurred more rapidly than the change of the protein secondary structure of the PSII membrane at temperatures above 45℃. The results indicate that the protein secondary structure of the membrane proteins in PSII membranes is more stable than the excitonic interaction between chlorophyll molecules during heat denaturation.

  19. Protein denaturation and functional properties of Lenient Steam Injection heat treated whey protein concentrate

    DEFF Research Database (Denmark)

    Dickow, Jonatan Ahrens; Kaufmann, Niels; Wiking, Lars

    2012-01-01

    Whey protein concentrate (WPC) was heat treated by use of the novel heat treatment method of Lenient Steam Injection (LSI) to elucidate new functional properties in relation to heat-induced gelation of heat treated WPC. Denaturation was measured by both DSC and FPLC, and the results of the two...... methods were highly correlated. Temperatures of up to 90 °C were applicable using LSI, whereas only 68 °C could be reached by plate heat exchange before coagulation/fouling. Denaturation of whey proteins increased with increasing heat treatment temperature up to a degree of 30–35% denaturation at 90 °C...

  20. On the Effect of Sodium Chloride and Sodium Sulfate on Cold Denaturation.

    Directory of Open Access Journals (Sweden)

    Andrea Pica

    Full Text Available Both sodium chloride and sodium sulfate are able to stabilize yeast frataxin, causing an overall increase of its thermodynamic stability curve, with a decrease in the cold denaturation temperature and an increase in the hot denaturation one. The influence of low concentrations of these two salts on yeast frataxin stability can be assessed by the application of a theoretical model based on scaled particle theory. First developed to figure out the mechanism underlying cold denaturation in water, this model is able to predict the stabilization of globular proteins provided by these two salts. The densities of the salt solutions and their temperature dependence play a fundamental role.

  1. Fish parasites

    DEFF Research Database (Denmark)

    This book contains 22 chapters on some of the most important parasitic diseases in wild and farmed fish. International experts give updated reviews and provide solutions to the problems......This book contains 22 chapters on some of the most important parasitic diseases in wild and farmed fish. International experts give updated reviews and provide solutions to the problems...

  2. The influence of applied heat treatments on whey protein denaturation

    Directory of Open Access Journals (Sweden)

    Fetahagić Safet

    2002-01-01

    . Distribution of nitrogen matter from milk 8%+3%DWP heat treated at 85ºC/10 min, 90ºC/10 min and 95ºC/10 min to sera samples were 9.64%, 8.66% and 8.67%, respectively. Whey protein denaturation increased with increasing of the temperature of the applied heat treatment. Denaturation was the most significant for milk sample 11%.

  3. Fishing TaSC Interacting Protein in Wheat Using Split Ubiquitin Yeast Two Hybrid System%利用分裂泛素酵母双杂交技术钓取小麦TaSC互作蛋白质

    Institute of Scientific and Technical Information of China (English)

    蔺芳芳; 杨旭; 武小翠; 刘晓梅; 葛荣朝; 赵宝存

    2013-01-01

      TaSC (Triticum asetivum L. salt-tolerance related gene, GenBank 登录号为 AY956330)是从小麦耐盐突变体RH8706-49中克隆的高盐诱导表达的耐盐相关基因.以该基因编码的蛋白质TaSC为诱饵,运用分裂泛素酵母双杂交技术从小麦 cDNA 表达文库中钓取互作蛋白质,筛选到一个编码小麦未知功能蛋白质的基因(GenBank 登录号为AK336035),命名为TaSCIP1(TaSC interaction protein 1).双分子荧光互补(BiFC)实验证实TaSCIP1与TaSC存在互作.该互作蛋白的分离有利于进一步研究TaSC基因的耐盐机制.%The TaSC (Triticum asetivum L. salt-tolerance related gene, GenBank accession number AY956330) has been cloned from wheat salt tolerant mutant RH 8706-49, which is induced by high salinity. To fish interacting proteins of this gene, we used TaSC protein as bait to hybrid with the wheat cDNA expression library using the split ubiquitin yeast two hybrid system in this experiment. A novel wheat gene (GenBank accession number AK336035) encoding a protein with unknown function was isolated, which was designated TaSCIP1 (TaSC interaction protein 1). Bimolecular fluorescence complementation (BiFC) experiment con-firmed the interaction between proteins TaSCIP1 and TaSC. The separation of TaSC-interacting protein is beneficial to disclose the mechanism of salt tolerance gene TaSC.

  4. Suppression subtractive hybridization and its application to fish gene cloning%抑制消减杂交(SSH)及其在鱼类基因克隆中的应用

    Institute of Scientific and Technical Information of China (English)

    程起群

    2004-01-01

    There are 10 percent to 15 percent genes expression in certain cells during the life time of fishes like other vertebrates. The genes were different at different development stage, under different physiological conditions, and in different kinds of cells. So comparing the differences of gene expression in different cells can help us understand the genetic nature of phenotypic differences, and understand the basic information of life period, and find the genes in relation to development and diseases, and finally benefit mankind. Several methods were developed to clone differential expression gene in recent years. They are subtractive hybridization (SH), differential display (DD),representional difference analysis (RDA), and so on. These methods all have postive influences on cloning special genes, but they all have some defects, such as higher false-positive, lower replication, lower sensitive and difficulty to manipulate. Suppression subtractive hybridization (SSH) was developed by Diatchenko et al in 1996. SSH was based on suppression PCR and combines normalization and subtraction in a single procedure. It is a more effective and convenient method than all others mentioned above. The principle and the rules of manipulation of SSH in detail was illuminated and the novel genes cloned by SSH was listed. They are immune related genes and reproduction and development related genes. The reproduction and development related genes are as follows: ZP3, Cyclin A2,CBI02, YA2, FSTRAP. The immune related genes are as follows: NKEF(natural killer enhancing factor), CC chemokine, CXCR1, CXCR2, CXCR4, AIF-1(allograft inflammatory factor-1), IL-1β(inteleukin-1), FcεRIγ(γ submit of high affinity Fc receptor for IgE), SSA(serum amyloid A), LECT2 (leucocyte cell-derived hemotaxin 2), GMFβ(glia maturation factorβ), CD45, Lysozyme C, PBEF (Pre-B cell enhancing factor), C-type lectin,PTX(Pentraxin), IL-1RⅡ, IL-8-1ike CXC chemokine, TF(tissue factor), trout chemokine 2, TNF decoy

  5. Protein folding by distributed computing and the denatured state ensemble.

    Science.gov (United States)

    Marianayagam, Neelan J; Fawzi, Nicolas L; Head-Gordon, Teresa

    2005-11-15

    The distributed computing (DC) paradigm in conjunction with the folding@home (FH) client server has been used to study the folding kinetics of small peptides and proteins, giving excellent agreement with experimentally measured folding rates, although pathways sampled in these simulations are not always consistent with the folding mechanism. In this study, we use a coarse-grain model of protein L, whose two-state kinetics have been characterized in detail by using long-time equilibrium simulations, to rigorously test a FH protocol using approximately 10,000 short-time, uncoupled folding simulations starting from an extended state of the protein. We show that the FH results give non-Poisson distributions and early folding events that are unphysical, whereas longer folding events experience a correct barrier to folding but are not representative of the equilibrium folding ensemble. Using short-time, uncoupled folding simulations started from an equilibrated denatured state ensemble (DSE), we also do not get agreement with the equilibrium two-state kinetics because of overrepresented folding events arising from higher energy subpopulations in the DSE. The DC approach using uncoupled short trajectories can make contact with traditionally measured experimental rates and folding mechanism when starting from an equilibrated DSE, when the simulation time is long enough to sample the lowest energy states of the unfolded basin and the simulated free-energy surface is correct. However, the DC paradigm, together with faster time-resolved and single-molecule experiments, can also reveal the breakdown in the two-state approximation due to observation of folding events from higher energy subpopulations in the DSE.

  6. Denaturing gradient gel electrophoresis profiling of bacterial communities composition in Arabian Sea

    Digital Repository Service at National Institute of Oceanography (India)

    Singh, S.K.; Ramaiah, N.

    Denaturing gradient gel electrophoresis (DGGE) was used to elucidate spatial and temporal variations in bacterial community composition (BCC) from four locations along the central west coast of India. DNA extracts from 36 water samples collected...

  7. Streptokinase Recovery by Cross-Flow Microfiltration: Study of Enzyme Denaturation

    National Research Council Canada - National Science Library

    HERNANDEZ-PINZON, Inmaculada; MILLAN, Francisco; BAUTISTA, Juan

    1997-01-01

    ...% remained in the retentate. Immunological experiments using polyclonal antibodies against SK have demonstrated that SK activity loss during CFMF processes could be related to denaturation of SK, forming molecules of lower or no activity...

  8. Single molecule study of the DNA denaturation phase transition in the force-torsion space

    CERN Document Server

    Salerno, D; Mai, I; Brogioli, D; Ziano, R; Cassina, V; Mantegazza, F

    2012-01-01

    We use the "magnetic tweezers" technique to reveal the structural transitions that DNA undergoes in the force-torsion space. In particular, we focus on regions corresponding to negative supercoiling. These regions are characterized by the formation of so-called denaturation bubbles, which have an essential role in the replication and transcription of DNA. We experimentally map the region of the force-torsion space where the denaturation takes place. We observe that large fluctuations in DNA extension occur at one of the boundaries of this region, i.e., when the formation of denaturation bubbles and of plectonemes are competing. To describe the experiments, we introduce a suitable extension of the classical model. The model correctly describes the position of the denaturation regions, the transition boundaries, and the measured values of the DNA extension fluctuations.

  9. How Chain Length and Charge Affect Surfactant Denaturation of Acyl Coenzyme A Binding Protein (ACBP)

    DEFF Research Database (Denmark)

    Andersen, Kell Kleiner; Otzen, Daniel

    2009-01-01

    Using intrinsic tryptophan fluorescence, equilibria and kinetics of unfolding of acyl coenzyme A binding protein (ACBP) have been investigated in sodium alkyl sulfate surfactants of different chain length (8-16 carbon atoms) and with different proportions of the nonionic surfactant dodecyl...... maltoside (DDM). The aim has been to determine how surfactant chain length and micellar charge affect the denaturation mechanism. ACBP denatures in two steps irrespective of surfactant chain length, but with increasing chain length, the potency of the denaturant rises more rapidly than the critical micelle...... constants increases linearly with denaturant concentration below the cmc but declines at higher concentrations. Both shortening chain length and decreasing micellar charge reduce the overall kinetics of unfolding and makes the dependence of unfolding rate constants on surfactant concentration more complex...

  10. Protocols for Pachytene Chromosome and DNA Fiber FISH in Cotton

    Institute of Scientific and Technical Information of China (English)

    PENG Ren-hai; LING Jian; WANG Kun-bo; WANG Chun-ying; SONG Guo-li; LIU Fang

    2008-01-01

    @@ Fluorescence in situ hybridization (FISH) has become the most important technique in plant molecular cytogenetics research.FISH-based physical mapping provides a valuable complementary approach in genome sequeneing to measure the physical distances between adjacent BAC contigs and delineating the structure and DNA composition of genomic regions of centromere and telomere.The accuracy and precision of the FISH-base physical maps depend on the resolution power of FISH.

  11. Effect of mechanical denaturation on surface free energy of protein powders

    OpenAIRE

    Mohammad, Mohammad Amin; Grimsey, Ian M.; Forbes, Robert T.; Blagbrough, Ian S; Barbara R. Conway

    2016-01-01

    Globular proteins are important both as therapeutic agents and excipients. However, their fragile native\\ud conformations can be denatured during pharmaceutical processing, which leads to modification of\\ud the surface energy of their powders and hence their performance. Lyophilized powders of hen eggwhite\\ud lysozyme and �-galactosidase from Aspergillus oryzae were used as models to study the effects\\ud of mechanical denaturation on the surface energies of basic and acidic protein powders, r...

  12. Avoiding adsorption of DNA to polypropylene tubes and denaturation of short DNA fragments

    OpenAIRE

    Gaillard, Claire; Strauss, Francois

    1998-01-01

    Two problems can arise when working with small quantities of DNA in polypropylene tubes: first, significant amounts of DNA can become lost by sticking to the tube walls; second, short DNA fragments tend to denature when binding to polypropylene. In addition, DNA also tends to denature upon dehydration. We have found that a simple way to solve these problems is by using polyallomer tubes instead of polypropylene and by avoiding certain salts, such as sodium acetate, when drying DNA.

  13. One Fish, Two Fish, Redfish, You Fish!

    Science.gov (United States)

    White, Katherine; Timmons, Maryellen; Medders, Paul

    2011-01-01

    The recreational fishing activity presented in this article provides a hands-on, problem-based experience for students; it unites biology, math, economics, environmental policy, and population dynamics concepts. In addition, the activity allows students to shape environmental policy in a realistic setting and evaluate their peers' work. By…

  14. Thermal denaturation behavior of collagen fibrils in wet and dry environment.

    Science.gov (United States)

    Suwa, Yosuke; Nam, Kwangwoo; Ozeki, Kazuhide; Kimura, Tsuyoshi; Kishida, Akio; Masuzawa, Toru

    2016-04-01

    We have developed a new minimally invasive technique--integrated low-level energy adhesion technique (ILEAT)--which uses heat, pressure, and low-frequency vibrations for binding living tissues. Because the adhesion mechanism of the living tissues is not fully understood, we investigated the effect of thermal energy on the collagen structure in living tissues using ILEAT. To study the effect of thermal energy and heating time on the structure of the collagen fibril, samples were divided in two categories-wet and dry. Further, atomic force microscopy was used to analyze the collagen fibril structure before and after heating. Results showed that collagen fibrils in water denatured after 1 minute at temperatures higher than 80 °C, while partial denaturation was observed at temperatures of 80 °C and a heating time of 1 min. Furthermore, complete denaturation was achieved after 90 min, suggesting that the denaturation rate is temperature and time dependent. Moreover, the collagen fibrils in dry condition maintained their native structure even after being heated to 120 °C for 90 min in the absence of water, which specifically suppressed denaturation. However, partial denaturation of collagen fibrils could not be prevented, because this determines the adhesion between the collagen molecules, and stabilizes tissue bonding.

  15. Interim assessment of the denatured /sup 233/U fuel cycle: feasibility and nonproliferation characteristics

    Energy Technology Data Exchange (ETDEWEB)

    Abbott, L.S.; Bartine, D.E.; Burns, T.J. (eds.)

    1978-12-01

    A fuel cycle that employs /sup 233/U denatured with /sup 238/U and mixed with thorium fertile material is examined with respect to its proliferation-resistance characteristics and its technical and economic feasibility. The rationale for considering the denatured /sup 233/U fuel cycle is presented, and the impact of the denatured fuel on the performance of Light-Water Reactors, Spectral-Shift-Controlled Reactors, Gas-Cooled Reactors, Heavy-Water Reactors, and Fast Breeder Reactors is discussed. The scope of the R, D and D programs to commercialize these reactors and their associated fuel cycles is also summarized and the resource requirements and economics of denatured /sup 233/U cycles are compared to those of the conventional Pu/U cycle. In addition, several nuclear power systems that employ denatured /sup 233/U fuel and are based on the energy center concept are evaluated. Under this concept, dispersed power reactors fueled with denatured or low-enriched uranium fuel are supported by secure energy centers in which sensitive activities of the nuclear cycle are performed. These activities include /sup 233/U production by Pu-fueled transmuters (thermal or fast reactors) and reprocessing. A summary chapter presents the most significant conclusions from the study and recommends areas for future work.

  16. Effects of protein and phosphate buffer concentrations on thermal denaturation of lysozyme analyzed by isoconversional method.

    Science.gov (United States)

    Cao, X M; Tian, Y; Wang, Z Y; Liu, Y W; Wang, C X

    2016-07-03

    Thermal denaturation of lysozymes was studied as a function of protein concentration, phosphate buffer concentration, and scan rate using differential scanning calorimetry (DSC), which was then analyzed by the isoconversional method. The results showed that lysozyme thermal denaturation was only slightly affected by the protein concentration and scan rate. When the protein concentration and scan rate increased, the denaturation temperature (Tm) also increased accordingly. On the contrary, the Tm decreased with the increase of phosphate buffer concentration. The denaturation process of lysozymes was accelatated and the thermal stability was reduced with the increase of phosphate concentration. One part of degeneration process was not reversible where the aggregation occurred. The other part was reversible. The apparent activation energy (Ea) was computed by the isoconversional method. It decreased with the increase of the conversion ratio (α). The observed denaturation process could not be described by a simple reaction mechanism. It was not a process involving 2 standard reversible states, but a multi-step process. The new opportunities for investigating the kinetics process of protein denaturation can be supplied by this novel isoconversional method.

  17. Effect of mechanical denaturation on surface free energy of protein powders.

    Science.gov (United States)

    Mohammad, Mohammad Amin; Grimsey, Ian M; Forbes, Robert T; Blagbrough, Ian S; Conway, Barbara R

    2016-10-01

    Globular proteins are important both as therapeutic agents and excipients. However, their fragile native conformations can be denatured during pharmaceutical processing, which leads to modification of the surface energy of their powders and hence their performance. Lyophilized powders of hen egg-white lysozyme and β-galactosidase from Aspergillus oryzae were used as models to study the effects of mechanical denaturation on the surface energies of basic and acidic protein powders, respectively. Their mechanical denaturation upon milling was confirmed by the absence of their thermal unfolding transition phases and by the changes in their secondary and tertiary structures. Inverse gas chromatography detected differences between both unprocessed protein powders and the changes induced by their mechanical denaturation. The surfaces of the acidic and basic protein powders were relatively basic, however the surface acidity of β-galactosidase was higher than that of lysozyme. Also, the surface of β-galactosidase powder had a higher dispersive energy compared to lysozyme. The mechanical denaturation decreased the dispersive energy and the basicity of the surfaces of both protein powders. The amino acid composition and molecular conformation of the proteins explained the surface energy data measured by inverse gas chromatography. The biological activity of mechanically denatured protein powders can either be reversible (lysozyme) or irreversible (β-galactosidase) upon hydration. Our surface data can be exploited to understand and predict the performance of protein powders within pharmaceutical dosage forms. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Biophysical analysis of phaseolin denaturation induced by urea, guanidinium chloride, pH, and temperature.

    Science.gov (United States)

    Dyer, J M; Nelson, J W; Murai, N

    1992-06-01

    The structural stability of phaseolin was determined by using absorbance, circular dichroism (CD), fluorescence emission, and fluorescence polarization anisotropy to monitor denaturation induced by urea, guanidinium chloride (GdmCl), pH changes, increasing temperature, or a combination thereof. Initial results indicated that phaseolin remained folded to a similar extent in the presence or absence of 6.0 M urea or GdmCl at room temperature. In 6.0 M GdmCl, phaseolin denatures at approximately 65 degrees C when probed with absorbance, CD, and fluorescence polarization anisotropy. The transition occurs at lower temperatures by decreasing pH. Kinetic measurements of denaturation using CD indicated that the denaturation is slow below 55 degrees C and is associated with an activation energy of 52 kcal/mol in 6.0 M GdmCl. In addition, kinetic measurement using fluorescence emission indicated that the single tryptophan residue was sensitive to at least two steps of the denaturation process. The fluorescence emission appeared to reflect some other structural perturbation than protein denaturation, as fluorescence inflection occurred approximately 5 degrees C prior to the changes observed in absorbance, CD, and fluorescence polarization anisotropy.

  19. Tissue sealing device associated thermal spread: a comparison of histologic methods for detecting adventitial collagen denaturation

    Science.gov (United States)

    Jones, Ryan M.; Grisez, Brian T.; Thomas, Aaron C.; Livengood, Ryan H.; Coad, James E.

    2013-02-01

    Thermal spread (thermal tissue damage) results from heat conduction through the tissues immediately adjacent to a hyperthermic tissue sealing device. The extent of such heat conduction can be assessed by the detection of adventitial collagen denaturation. Several histologic methods have been reported to measure adventitial collagen denaturation as a marker of thermal spread. This study compared hematoxylin and eosin staining, Gomori trichrome staining and loss of collagen birefringence for the detection of collagen denaturation. Twenty-eight ex vivo porcine carotid arteries were sealed with a commercially available, FDA-approved tissue sealing device. Following formalin fixation and paraffin embedding, two 5-micron tissue sections were hematoxylin and eosin and Gomori trichrome stained. The hematoxylin and eosin-stained section was evaluated by routine bright field microscopy and under polarized light. The trichromestained section was evaluated by routine bright field microscopy. Radial and midline adventitial collagen denaturation measurements were made for both the top and bottom jaw sides of each seal. The adventitial collagen denaturation lengths were determined using these three methods and statistically compared. The results showed that thermal spread, as represented by histologically detected collagen denaturation, is technique dependent. In this study, the trichrome staining method detected significantly less thermal spread than the hematoxylin and eosin staining and birefringence methods. Of the three methods, hematoxylin and eosin staining provided the most representative results for true thermal spread along the adjacent artery.

  20. Fighting fish

    Science.gov (United States)

    Duchi, E.; Guerrini, V.; Rinaldi, S.; Schaeffer, G.

    2017-01-01

    We introduce new combinatorial structures, called fighting fish, that generalize directed convex polyominoes by allowing them to branch out of the plane into independent substructures. On the one hand the combinatorial structure of fighting fish appears to be particularly rich: we show that their generating function with respect to the perimeter and number of tails is algebraic, and we conjecture a mysterious multivariate equidistribution property with the left ternary trees introduced by Del Lungo et al On the other hand, fighting fish provide a simple and natural model of random branching surfaces which displays original features: in particular, we show that the average area of a uniform random fighting fish with perimeter 2n is of order n 5/4: to the best of our knowledge this behaviour is non-standard and suggests that we have identified a new universality class of random structures. Dedicated to Tony Guttmann on the occasion of his 70th birthday.

  1. Recent Advances in Fluorescence in situ Hybridization

    OpenAIRE

    吉田, 廸弘; Michihiro C., Yoshida; 北海道大学理学部附属動物染色体研究施設; Chromosome Research Unit, Faculty of Science, Hokkaido University

    1992-01-01

    Fluorescence in situ hybridization (FISH) procedures that directly couple molecular and cytological information allow precise visualization of DNA sequences on metaphase chromosomes and interphase nuclei. These techniques can be used to identify chromosomes, detect chromosomal aberrations, and analyze linear and spetial genome organization. FISH procedures are also used to clinical fields for diagnosis of disease-related chromosome changes and tumor biology.

  2. Denaturant-Dependent Conformational Changes in a [beta]-Trefoil Protein: Global and Residue-Specific Aspects of an Equilibrium Denaturation Process

    Energy Technology Data Exchange (ETDEWEB)

    Latypov, Ramil F.; Liu, Dingjiang; Jacob, Jaby; Harvey, Timothy S.; Bondarenko, Pavel V.; Kleemann, Gerd R.; Brems, David N.; Raibekas, Andrei A.; (Amgen)

    2010-01-12

    Conformational properties of the folded and unfolded ensembles of human interleukin-1 receptor antagonist (IL-1ra) are strongly denaturant-dependent as evidenced by high-resolution two-dimensional nuclear magnetic resonance (NMR), limited proteolysis, and small-angle X-ray scattering (SAXS). The folded ensemble was characterized in detail in the presence of different urea concentrations by 1H-15N HSQC NMR. The {beta}-trefoil fold characteristic of native IL-1ra was preserved until the unfolding transition region beginning at 4 M urea. At the same time, a subset of native resonances disappeared gradually starting at low denaturant concentrations, indicating noncooperative changes in the folded state. Additional evidence of structural perturbations came from the chemical shift analysis, nonuniform and bell-shaped peak intensity profiles, and limited proteolysis. In particular, the following nearby regions of the tertiary structure became progressively destabilized with increasing urea concentrations: the {beta}-hairpin interface of trefoils 1 and 2 and the H2a-H2 helical region. These regions underwent small-scale perturbations within the native baseline region in the absence of populated molten globule-like states. Similar regions were affected by elevated temperatures known to induce irreversible aggregation of IL-1ra. Further evidence of structural transitions invoking near-native conformations came from an optical spectroscopy analysis of its single-tryptophan variant W17A. The increase in the radius of gyration was associated with a single equilibrium unfolding transition in the case of two different denaturants, urea and guanidine hydrochloride (GuHCl). However, the compactness of urea- and GuHCl-unfolded molecules was comparable only at high denaturant concentrations and deviated under less denaturing conditions. Our results identified the role of conformational flexibility in IL-1ra aggregation and shed light on the nature of structural transitions within the

  3. Virginia ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains sensitive biological resource data for marine, estuarine, anadromous, and brackishwater fish species in Virginia. Vector polygons in this data...

  4. Hawaii ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains sensitive biological resource data for reef, marine, estuarine, and native stream fish species in coastal Hawaii. Vector polygons in this data...

  5. Alabama ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains sensitive biological resource data for marine, estuarine, and freshwater fish species in Alabama. Vector polygons in this data set represent...

  6. Louisiana ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains sensitive biological resource data for freshwater (inland) fish species in coastal Louisiana. Vector polygons represent water-bodies and other...

  7. Correlated parameter fit of arrhenius model for thermal denaturation of proteins and cells.

    Science.gov (United States)

    Qin, Zhenpeng; Balasubramanian, Saravana Kumar; Wolkers, Willem F; Pearce, John A; Bischof, John C

    2014-12-01

    Thermal denaturation of proteins is critical to cell injury, food science and other biomaterial processing. For example protein denaturation correlates strongly with cell death by heating, and is increasingly of interest in focal thermal therapies of cancer and other diseases at temperatures which often exceed 50 °C. The Arrhenius model is a simple yet widely used model for both protein denaturation and cell injury. To establish the utility of the Arrhenius model for protein denaturation at 50 °C and above its sensitivities to the kinetic parameters (activation energy E a and frequency factor A) were carefully examined. We propose a simplified correlated parameter fit to the Arrhenius model by treating E a, as an independent fitting parameter and allowing A to follow dependently. The utility of the correlated parameter fit is demonstrated on thermal denaturation of proteins and cells from the literature as a validation, and new experimental measurements in our lab using FTIR spectroscopy to demonstrate broad applicability of this method. Finally, we demonstrate that the end-temperature within which the denaturation is measured is important and changes the kinetics. Specifically, higher E a and A parameters were found at low end-temperature (50 °C) and reduce as end-temperatures increase to 70 °C. This trend is consistent with Arrhenius parameters for cell injury in the literature that are significantly higher for clonogenics (45-50 °C) vs. membrane dye assays (60-70 °C). Future opportunities to monitor cell injury by spectroscopic measurement of protein denaturation are discussed.

  8. Development of novel short-term heating angioplasty: thermal denaturation dynamics of collagen in artery wall

    Science.gov (United States)

    Shimazaki, N.; Tokunaga, H.; Katou, Y.; Hayashi, T.; Arai, T.

    2009-02-01

    We have studied to develop the new thermal angioplasty methodology, photo-thermo dynamic balloon angioplasty (PTDBA), which provides artery dilatation with short-term (collagen in artery media may be the important factor to attain sufficient artery dilatation for the PTDBA. In order to predict the optimum heating condition i.e. the balloon temperature and heating duration, we investigated the thermal denaturation dynamics of artery collagen in ex vivo. The extracted fresh porcine carotid artery was used. The temperature-dependent light scattering property and mechanical property of the artery specimen were simultaneously measured during artery temperature rising by specially made setup to assess the denaturation of arterial collagen. The change rate of the backscattered light intensity from the artery specimen; I(T)/I0 with 633nm was measured to evaluate the artery scattering property change with the thermal denaturation. The artery specimen was heated from 25°C to 80°C with constant temperature rising rate of 3°C/min. The measured I(T)/I0 was suddenly increased over 48°C. This boundary temperature might be the initiation temperature of the arterial collagen denaturation. We defined the variation of the I(T)/I0 as the collagen denaturation ratio, and calculated the reactive enthalpy by the chemical equilibrium theory. Since the calculated enthalpy was similar to the enthalpy in literature report, the variety of I(T)/I0 during the temperature rising might be attributed to the collagen conformational change due to the denaturation. In terms of the artery internal force measurement, the artery force was decreased with increasing of the artery temperature up to 65°C (i.e. softening), and increased over 65°C (i.e. shrinkage). We confirmed that the changes of the backscattered light (at 633nm in wavelength) from the artery might represent the artery collagen thermal denaturation degree.

  9. Effect of high-pressure treatment on denaturation of bovine lactoferrin and lactoperoxidase.

    Science.gov (United States)

    Mazri, C; Sánchez, L; Ramos, S J; Calvo, M; Pérez, M D

    2012-02-01

    Lactoferrin and lactoperoxidase are whey proteins with biological properties that may provide health benefits to consumers. These properties are vulnerable to potentially denaturing conditions during processing. High-pressure treatment is an appealing alternative to the traditional heat processing of foods because it exerts an antimicrobial effect without changing the sensory and nutritional quality of foods. In this work, the effect of high-pressure treatment on the denaturation of lactoferrin and lactoperoxidase present in skim milk and whey, and as isolated proteins in buffer, was studied over a pressure range of 450 to 700 MPa at 20°C. Denaturation of lactoferrin was measured by the loss of reactivity with their specific antibodies using a sandwich ELISA. Denaturation of lactoperoxidase was determined by measuring the loss of enzymatic activity using a spectrophotometric technique. No substantial inactivation of lactoperoxidase was observed in any treatment assayed. The concentration of the residual immunoreactive lactoferrin after each pressure treatment was determined, and the data were subjected to kinetic analysis to obtain D and Z values. Denaturation of lactoferrin increased with pressure and holding time, and D values were lower when lactoferrin was treated in whey than in milk, and lower in both whey and milk than in phosphate buffer. Thus, protein is denatured more slowly in buffer and in milk than in whey. Denaturation of lactoferrin in the 3 media was found to follow a reaction order of n=1.5. Volumes of activation of about -34.77, -24.35, and -24.09 mL/mol were obtained for lactoferrin treated in skim milk, whey, and buffer, respectively, indicating a decrease in protein volume under pressure.

  10. MicroRNA-23b Inhibits the Proliferation and Migration of Heat-Denatured Fibroblasts by Targeting Smad3.

    Directory of Open Access Journals (Sweden)

    Xipeng Zhang

    Full Text Available Skin grafting with the preservation of denatured dermis is a novel strategy for the treatment of burn-injured skin. Denatured dermis has the ability to restore to the morphology and function of normal skin, but the underlying molecular mechanism is elusive. MicroRNAs (miRNA are small noncoding RNAs and regulate normal physiology as well as disease development. In this study, we assessed the potential role of miRNA-23b (miR-23b in the regulation of cell proliferation and migration of heat-denatured fibroblasts and identified the underlying mechanism.The expression of miR-23b in denatured dermis and heat-denatured fibroblasts was detected by quantitative real-time polymerase chain reaction (RT-PCR. The effects of miR-23b on cell proliferation and migration of heat-denatured fibroblasts were assessed by transient transfection of miR-23b mimics and inhibitor. The target gene of miR-23b and the downstream pathway were further investigated.miR-23b was downregulated in denatured dermis and heat-denatured fibroblasts. Downregulation of miR-23b dramatically promoted the proliferation and migration of heat-denatured fibroblasts. Subsequent analyses demonstrated that Smad3 was a direct and functional target of miR-23b in heat-denatured fibroblasts, which was validated by the dual luciferase reporter assay. Moreover, immunohistochemistry analysis showed that denatured dermis from rats displayed enhanced staining of Smad3. In addition, miR-23b modulated denatured dermis by activating the Notch1 and TGF-β signaling pathways.Our findings suggest that downregulation of miR-23b contributes to the recovery of denatured dermis, which may be valuable for treatment of skin burns.

  11. Hybrid Baryons

    CERN Document Server

    Page, P R

    2003-01-01

    We review the status of hybrid baryons. The only known way to study hybrids rigorously is via excited adiabatic potentials. Hybrids can be modelled by both the bag and flux-tube models. The low-lying hybrid baryon is N 1/2^+ with a mass of 1.5-1.8 GeV. Hybrid baryons can be produced in the glue-rich processes of diffractive gamma N and pi N production, Psi decays and p pbar annihilation.

  12. Comprehensive immunohistochemical analysis of Her-2/neu oncoprotein overexpression in breast cancer: HercepTest (Dako) for manual testing and Her-2/neuTest 4B5 (Ventana) for Ventana BenchMark automatic staining system with correlation to results of fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Mayr, Doris; Heim, Sibylle; Werhan, Cedric; Zeindl-Eberhart, Evelyn; Kirchner, Thomas

    2009-03-01

    Overexpression of Her-2/neu-oncoprotein is used as marker for Herceptin therapy. To investigate the sensitivity and specificity of automatic immunohistochemistry (Benchmark, Ventana), we compared the results to the manual testing (Dako) in 130 breast carcinomas and validated the results by fluorescence in situ hybridization (FISH). Manual and automatic immunohistochemistry of Her-2/neu-oncoprotein using two different antibodies (HercepTest, Her-2/neuTest 4B5) was analyzed. FISH was performed in all cases with uncertain or strong overexpression in either immunohistochemical stainings or with different immunohistochemical results. Same immunohistochemical results were seen in 73.8%. Two cases with overexpression, detected with Her-2/neuTest 4B5 and confirmed by FISH, showed no overexpression using HercepTest. From 21 cases with 2+ by Her-2/neuTest 4B5, 15 cases had no gene amplification (two of them with 3+ HercepTest); three cases showed a gene amplification (one of them with failing overexpression by HercepTest); two other cases were polysomic; one could not be analyzed. Ventana immunohistochemistry seems to be of same reliability like Dako with a little better concordance to FISH in our study.

  13. Parasites modify sub-cellular partitioning of metals in the gut of fish

    Energy Technology Data Exchange (ETDEWEB)

    Oyoo-Okoth, Elijah, E-mail: elijaoyoo2009@gmail.com [Division of Environmental Health, School of Environmental Studies, Moi University, P.O. Box 3900, Eldoret (Kenya); Department of Aquatic Ecology and Ecotoxicology, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, P.O. Box 9424/1090 GE (Netherlands); Admiraal, Wim [Department of Aquatic Ecology and Ecotoxicology, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, P.O. Box 9424/1090 GE (Netherlands); Osano, Odipo [Division of Environmental Health, School of Environmental Studies, Moi University, P.O. Box 3900, Eldoret (Kenya); Kraak, Michiel H.S. [Department of Aquatic Ecology and Ecotoxicology, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, P.O. Box 9424/1090 GE (Netherlands); Gichuki, John; Ogwai, Caleb [Kenya Marine and Fisheries Research Institute, P.O. Box 1881, Kisumu (Kenya)

    2012-01-15

    Infestation of fish by parasites may influence metal accumulation patterns in the host. However, the subcellular mechanisms of these processes have rarely been studied. Therefore, this study determined how a cyprinid fish (Rastrineobola argentea) partitioned four metals (Cd, Cr, Zn and Cu) in the subcellular fractions of the gut in presence of an endoparasite (Ligula intestinalis). The fish were sampled along four sites in Lake Victoria, Kenya differing in metal contamination. Accumulation of Cd, Cr and Zn was higher in the whole body and in the gut of parasitized fish compared to non-parasitized fish, while Cu was depleted in parasitized fish. Generally, for both non-parasitized and parasitized fish, Cd, Cr and Zn partitioned in the cytosolic fractions and Cu in the particulate fraction. Metal concentrations in organelles within the particulate fractions of the non-parasitized fish were statistically similar except for Cd in the lysosome, while in the parasitized fish, Cd, Cr and Zn were accumulated more by the lysosome and microsomes. In the cytosolic fractions, the non-parasitized fish accumulated Cd, Cr and Zn in the heat stable proteins (HSP), while in the parasitized fish the metals were accumulated in the heat denatured proteins (HDP). On the contrary, Cu accumulated in the HSP in parasitized fish. The present study revealed specific binding of metals to potentially sensitive sub-cellular fractions in fish in the presence of parasites, suggesting interference with metal detoxification, and potentially affecting the health status of fish hosts in Lake Victoria.

  14. Alcohol-induced structural transitions in the acid-denatured Bacillus licheniformis α-amylase

    Directory of Open Access Journals (Sweden)

    Adyani Azizah Abd Halim

    2017-01-01

    Full Text Available Alcohol-induced structural changes in the acid-denatured Bacillus licheniformis α-amylase (BLA at pH 2.0 were studied by far-ultra violet circular dichroism, intrinsic, three-dimensional and 8-anilino-1-naphthalene sulfonic acid (ANS fluorescence, acrylamide quenching and thermal denaturation. All the alcohols used in this study produced partial refolding in the acid-denatured BLA as evident from the increased mean residue ellipticity at 222 nm, increased intrinsic fluorescence and decreased ANS fluorescence. The order of effectiveness of these alcohols to induce a partially folded state of BLA was found to be: 2,2,2-trifluoroethanol/tert-butanol > 1-propanol/2-propanol > 2-chloroethanol > ethanol > methanol. Three-dimensional fluorescence and acrylamide quenching results obtained in the presence of 5.5 M tert-butanol also suggested formation of a partially folded state in the acid-denatured BLA. However, 5.5 M tert-butanol-induced state of BLA showed a non-cooperative thermal transition. All these results suggested formation of a partially folded state of the acid-denatured BLA in the presence of these alcohols. Furthermore, their effectiveness was found to be guided by their chain length, position of methyl groups and presence of the substituents.

  15. Drying and denaturation characteristics of whey protein isolate in the presence of lactose and trehalose.

    Science.gov (United States)

    Haque, M Amdadul; Chen, Jie; Aldred, Peter; Adhikari, Benu

    2015-06-15

    The denaturation kinetics of whey protein isolate (WPI), in the presence and absence of lactose and trehalose, was quantified in a convective air-drying environment. Single droplets of WPI, WPI-lactose and WPI-trehalose were dried in conditioned air (2.5% RH, 0.5m/s air velocity) at two temperatures (65°C and 80°C) for 500s. The initial solid concentration of these solutions was 10% (w/v) in all the samples. Approximately 68% of WPI was denatured when it was dried in the absence of sugars. Addition of 20% trehalose prevented the irreversible denaturation of WPI at both temperatures. Thirty percent lactose was required to prevent denaturation of WPI at 65°C and the same amount of lactose protected only 70% of WPI from denaturation at 80°C. The secondary structures of WPI were found to be altered by the drying-induced stresses, even in the presence of 20% trehalose and 30% lactose.

  16. Denaturation of milk proteins and their influence on the yield of fresh cheese

    Directory of Open Access Journals (Sweden)

    Ana Mejía-López

    2017-03-01

    Full Text Available To determine the denaturation of milk proteins by the effects of heat treatment on pasteurization and to establish their influence on the yield of the fresh cheese manufactured, 20 laboratory- scale controlled trials and 40 plant productions were made. Crude and treated milk was used at 65 ° C for 30 minutes, 72 ° C for 15 seconds and boiled for 2 seconds, and the protein was quantified in milk to calculate percent denaturation. In the cheese the moisture content was determined and the amount of cheese obtained was quantified. The data were processed by Tukey's mean analysis (p> 0.05. The results at the laboratory level showed that the increase in temperature caused higher denaturation of the proteins, a higher yield and an increase in moisture in the cheese compared to that obtained with raw milk. However, statistically the results showed that the heat treatment does influence the denaturation of the proteins but not the performance of the cheese. The results obtained in the factory investigation revealed that at 65 and 72 ° C the yield decreases relative to the production with raw milk, but statistically does not present significant differences in the yield, concluding that the pasteurization at different temperatures denature the protein But does not influence the performance of fresh processed cheese.

  17. A versatile protein microarray platform enabling antibody profiling against denatured proteins.

    Science.gov (United States)

    Wang, Jie; Barker, Kristi; Steel, Jason; Park, Jin; Saul, Justin; Festa, Fernanda; Wallstrom, Garrick; Yu, Xiaobo; Bian, Xiaofang; Anderson, Karen S; Figueroa, Jonine D; LaBaer, Joshua; Qiu, Ji

    2013-06-01

    We aim to develop a protein microarray platform capable of presenting both natural and denatured forms of proteins for antibody biomarker discovery. We will further optimize plasma screening protocols to improve detection. We developed a new covalent capture protein microarray chemistry using HaloTag fusion proteins and ligand. To enhance protein yield, we used HeLa cell lysate as an in vitro transcription translation (IVTT) system. Escherichia coli lysates were added to the plasma blocking buffer to reduce nonspecific background. These protein microarrays were probed with plasma samples and autoantibody responses were quantified and compared with or without denaturing buffer treatment. We demonstrated that protein microarrays using the covalent attachment chemistry endured denaturing conditions. Blocking with E. coli lysates greatly reduced the background signals and expression with IVTT based on HeLa cell lysates significantly improved the antibody signals on protein microarrays probed with plasma samples. Plasma samples probed on denatured protein arrays produced autoantibody profiles distinct from those probed on natively displayed proteins. This versatile protein microarray platform allows the display of both natural and denatured proteins, offers a new dimension to search for disease-specific antibodies, broadens the repertoire of potential biomarkers, and will potentially yield clinical diagnostics with greater performance. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Comparison of chemical and thermal protein denaturation by combination of computational and experimental approaches. II

    Science.gov (United States)

    Wang, Qian; Christiansen, Alexander; Samiotakis, Antonios; Wittung-Stafshede, Pernilla; Cheung, Margaret S.

    2011-11-01

    Chemical and thermal denaturation methods have been widely used to investigate folding processes of proteins in vitro. However, a molecular understanding of the relationship between these two perturbation methods is lacking. Here, we combined computational and experimental approaches to investigate denaturing effects on three structurally different proteins. We derived a linear relationship between thermal denaturation at temperature Tb and chemical denaturation at another temperature Tu using the stability change of a protein (ΔG). For this, we related the dependence of ΔG on temperature, in the Gibbs-Helmholtz equation, to that of ΔG on urea concentration in the linear extrapolation method, assuming that there is a temperature pair from the urea (Tu) and the aqueous (Tb) ensembles that produces the same protein structures. We tested this relationship on apoazurin, cytochrome c, and apoflavodoxin using coarse-grained molecular simulations. We found a linear correlation between the temperature for a particular structural ensemble in the absence of urea, Tb, and the temperature of the same structural ensemble at a specific urea concentration, Tu. The in silico results agreed with in vitro far-UV circular dichroism data on apoazurin and cytochrome c. We conclude that chemical and thermal unfolding processes correlate in terms of thermodynamics and structural ensembles at most conditions; however, deviations were found at high concentrations of denaturant.

  19. Polar or apolar--the role of polarity for urea-induced protein denaturation.

    Directory of Open Access Journals (Sweden)

    Martin C Stumpe

    2008-11-01

    Full Text Available Urea-induced protein denaturation is widely used to study protein folding and stability; however, the molecular mechanism and driving forces of this process are not yet fully understood. In particular, it is unclear whether either hydrophobic or polar interactions between urea molecules and residues at the protein surface drive denaturation. To address this question, here, many molecular dynamics simulations totalling ca. 7 micros of the CI2 protein in aqueous solution served to perform a computational thought experiment, in which we varied the polarity of urea. For apolar driving forces, hypopolar urea should show increased denaturation power; for polar driving forces, hyperpolar urea should be the stronger denaturant. Indeed, protein unfolding was observed in all simulations with decreased urea polarity. Hyperpolar urea, in contrast, turned out to stabilize the native state. Moreover, the differential interaction preferences between urea and the 20 amino acids turned out to be enhanced for hypopolar urea and suppressed (or even inverted for hyperpolar urea. These results strongly suggest that apolar urea-protein interactions, and not polar interactions, are the dominant driving force for denaturation. Further, the observed interactions provide a detailed picture of the underlying molecular driving forces. Our simulations finally allowed characterization of CI2 unfolding pathways. Unfolding proceeds sequentially with alternating loss of secondary or tertiary structure. After the transition state, unfolding pathways show large structural heterogeneity.

  20. Experimental and Modelling Study of the Denaturation of Milk Protein by Heat Treatment

    Science.gov (United States)

    Qian, Fang; Sun, Jiayue; Cao, Di; Tuo, Yanfeng; Jiang, Shujuan; Mu, Guangqing

    2017-01-01

    Heat treatment of milk aims to inhibit the growth of microbes, extend the shelf-life of products and improve the quality of the products. Heat treatment also leads to denaturation of whey protein and the formation of whey protein-casein polymer, which has negative effects on milk product. Hence the milk heat treatment conditions should be controlled in milk processing. In this study, the denaturation degree of whey protein and the combination degree of whey protein and casein when undergoing heat treatment were also determined by using the Native-PAGE and SDS-PAGE analysis. The results showed that the denaturation degree of whey protein and the combination degree of whey protein with casein extended with the increase of the heat-treated temperature and time. The effects of the heat-treated temperature and heat-treated time on the denaturation degree of whey protein and on the combination degree of whey protein and casein were well described using the quadratic regression equation. The analysis strategy used in this study reveals an intuitive and effective measure of the denaturation degree of whey protein, and the changes of milk protein under different heat treatment conditions efficiently and accurately in the dairy industry. It can be of great significance for dairy product proteins following processing treatments applied for dairy product manufacturing. PMID:28316470

  1. Antibodies with specificity for native and denatured forms of ovalbumin differ in reactivity between enzyme-linked immunosorbent assays

    DEFF Research Database (Denmark)

    Holm, B.E.; Bergmann, A.C.; Hansen, Paul Robert

    2015-01-01

    In this study, polyclonal and monoclonal antibodies to native and denatured chicken ovalbumin (OVA) were produced to compare their dependency on continuous and three-dimensional epitopes. These antibodies were characterized with respect to reactivity to native and denatured OVA by enzyme...... to native OVA reacted strongly with native and denatured OVA in both assays, but did not react with the overlapping peptides. Polyclonal antibodies to denatured OVA reacted strongly with both OVA forms and with several of the overlapping peptides. Monoclonal antibodies to native OVA reacted preferentially...... with three-dimensional epitopes on native OVA and not with denatured OVA. Monoclonal antibodies to denatured OVA showed reactivity to both OVA forms. Two of these monoclonal antibodies, HYB 94-06 and 94-07, showed reactivity to overlapping peptides and their epitopes were identified as flexible structures...

  2. Fishing Access Areas

    Data.gov (United States)

    Vermont Center for Geographic Information — The Vermont Fish & Wildlife Department maintains developed fishing access areas. These sites provide public access to waters in Vermont for shore fishing...

  3. Identification of nitrite-reducing bacteria using sequential mRNA fluorescence in situ hybridization and fluorescence-assisted cell sorting.

    Science.gov (United States)

    Mota, Cesar R; So, Mark Jason; de los Reyes, Francis L

    2012-07-01

    Sequential mRNA fluorescence in situ hybridization (mRNA FISH) and fluorescence-assisted cell sorting (SmRFF) was used for the identification of nitrite-reducing bacteria in mixed microbial communities. An oligonucleotide probe labeled with horseradish peroxidase (HRP) was used to target mRNA of nirS, the gene that encodes nitrite reductase, the enzyme responsible for the dissimilatory reduction of nitrite to nitric oxide. Clones for nirS expression were constructed and used to provide proof of concept for the SmRFF method. In addition, cells from pure cultures of Pseudomonas stutzeri and denitrifying activated sludge were hybridized with the HRP probe, and tyramide signal amplification was performed, conferring a strongly fluorescent signal to cells containing nirS mRNA. Flow cytometry-assisted cell sorting was used to detect and physically separate two subgroups from a mixed microbial community: non-fluorescent cells and an enrichment of fluorescent, nitrite-reducing cells. Denaturing gradient gel electrophoresis (DGGE) and subsequent sequencing of 16S ribosomal RNA (rRNA) genes were used to compare the fragments amplified from the two sorted subgroups. Sequences from bands isolated from DGGE profiles suggested that the dominant, active nitrite reducers were closely related to Acidovorax BSB421. Furthermore, following mRNA FISH detection of nitrite-reducing bacteria, 16S rRNA FISH was used to detect ammonia-oxidizing and nitrite-oxidizing bacteria on the same activated sludge sample. We believe that the molecular approach described can be useful as a tool to help address the longstanding challenge of linking function to identity in natural and engineered habitats.

  4. Inhibition of thermal induced protein denaturation of extract/fractions of Withania somnifera and isolated withanolides.

    Science.gov (United States)

    Khan, Murad Ali; Khan, Haroon; Rauf, Abdul; Ben Hadda, Taibi

    2015-01-01

    This study describes the in vitro inhibition of protein denaturation of extract/fractions of Withania somnifera and isolated withanolides including 20β hydroxy-1-oxo(22R)-witha-2,5,24 trienolide (1), (20R,22R-14α,20α)-dihydroxy-1-oxowitha-2,5,16,24 tetraenolide (2). The results showed that the extract/fractions of the plant evoked profound inhibitory effect on thermal-induced protein denaturation. The chloroform fraction caused the most dominant attenuation of 68% at 500 μg/mL. The bioactivity-guided isolation from chloroform fraction led to the isolation of compounds 1 and 2 that showed profound protein inhibition with 78.05% and 80.43% effect at 500 μg/mL and thus strongly complimented the activity of extract/fractions. In conclusion, extract/fractions of W. somnifera possessed strong inhibition of protein denaturation that can be attributed to these isolated withanolides.

  5. Nonsurgical transurethral radiofrequency collagen denaturation: results at three years after treatment.

    Science.gov (United States)

    Elser, Denise M; Mitchell, Gretchen K; Miklos, John R; Nickell, Kevin G; Cline, Kevin; Winkler, Harvey; Wells, W Glen

    2011-01-01

    Objective. To assess treatment efficacy and quality of life in women with stress urinary incontinence 3 years after treatment with nonsurgical transurethral radiofrequency collagen denaturation. Methods. This prospective study included 139 women with stress urinary incontinence due to bladder outlet hypermobility. Radiofrequency collagen denaturation was performed using local anesthesia in an office setting. Assessments included incontinence quality of life (I-QOL) and urogenital distress inventory (UDI-6) instruments. Results. In total, 139 women were enrolled and 136 women were treated (mean age, 47 years). At 36 months, intent-to-treat analysis (n = 139) revealed significant improvements in quality of life. Mean I-QOL score improved 17 points from baseline (P = .0004), while mean UDI-6 score improved (decreased) 19 points (P = .0005). Conclusions. Transurethral collagen denaturation is a low-risk, office-based procedure that results in durable quality-of-life improvements in a significant proportion of women for as long as 3 years.

  6. Stabilizing Effect of Various Polyols on the Native and the Denatured States of Glucoamylase

    Directory of Open Access Journals (Sweden)

    Mohammed Suleiman Zaroog

    2013-01-01

    Full Text Available Different spectral probes were employed to study the stabilizing effect of various polyols, such as, ethylene glycol (EG, glycerol (GLY, glucose (GLC and trehalose (TRE on the native (N, the acid-denatured (AD and the thermal-denatured (TD states of Aspergillus niger glucoamylase (GA. Polyols induced both secondary and tertiary structural changes in the AD state of enzyme as reflected from altered circular dichroism (CD, tryptophan (Trp, and 1-anilinonaphthalene-8-sulfonic acid (ANS fluorescence characteristics. Thermodynamic analysis of the thermal denaturation curve of native GA suggested significant increase in enzyme stability in the presence of GLC, TRE, and GLY (in decreasing order while EG destabilized it. Furthermore, CD and fluorescence characteristics of the TD state at 71°C in the presence of polyols showed greater effectiveness of both GLC and TRE in inducing native-like secondary and tertiary structures compared to GLY and EG.

  7. Gel electrophoresis of DNA partially denatured at the ends: what are the dominant conformations?

    Science.gov (United States)

    Sean, David; Slater, Gary W

    2013-03-01

    Gel electrophoresis of a partially denatured dsDNA fragment is studied using Langevin Dynamics computer simulations. For simplicity, the denatured ssDNA sections are placed at the ends of the fragment in a symmetrical fashion. A squid-like conformation is found to sometimes cause the fragment to completely block in the gel. In fact, this conformation is the principal cause of the steep reduction in mobility observed in the simulations. As the field is increased, it is found that the occurrence of this conformation dominates the migration dynamics. Although the squid conformation seems to be more stable at high fields, the field can eventually force the fragments to thread through the gel pores regardless. We qualitatively explore the behavior of this squid-like conformation across a range of fields and degrees of denaturation, and we discuss the relevance of our findings for TGGE. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. EQUIVALENCY OF FOULING THICKNESS WITH DENATURED Β-LG IN HEATING OF MILK

    Directory of Open Access Journals (Sweden)

    Heriberto Molina-Pérez

    2015-03-01

    Full Text Available This work presents an approach to match the foaling thickness of dairy food and the concentration of denatured β-lg (β-lactoglobuline by a mathematical model. This includes, on the one hand, the dynamic simulation of fouling and, on the other hand, the generation of denatured β-lg under a kinetic model. In both cases a transient energy balance is developed, including the rigorous calculation of the global coefficient, and the properties by the Choi-Okos model. The solution was obtained by a fourth order Runge-Kutta written in Excel’s Macro language Visual Basic. The equivalence concluded with a model obtained by a non-linear multiple regression that relates the concentration of denatured β-lg and the foaling thickness. This methodology is applicable to analysis equipment cleaning in which kinetic cleaning has equality by reduction of the foaling thickness.

  9. Statistical mechanics of the denatured state of a protein using replica-averaged metadynamics.

    Science.gov (United States)

    Camilloni, Carlo; Vendruscolo, Michele

    2014-06-25

    The characterization of denatured states of proteins is challenging because the lack of permanent structure in these states makes it difficult to apply to them standard methods of structural biology. In this work we use all-atom replica-averaged metadynamics (RAM) simulations with NMR chemical shift restraints to determine an ensemble of structures representing an acid-denatured state of the 86-residue protein ACBP. This approach has enabled us to reach convergence in the free energy landscape calculations, obtaining an ensemble of structures in relatively accurate agreement with independent experimental data used for validation. By observing at atomistic resolution the transient formation of native and non-native structures in this acid-denatured state of ACBP, we rationalize the effects of single-point mutations on the folding rate, stability, and transition-state structures of this protein, thus characterizing the role of the unfolded state in determining the folding process.

  10. Nonsurgical Transurethral Radiofrequency Collagen Denaturation: Results at Three Years after Treatment

    Directory of Open Access Journals (Sweden)

    Denise M. Elser

    2011-01-01

    Full Text Available Objective. To assess treatment efficacy and quality of life in women with stress urinary incontinence 3 years after treatment with nonsurgical transurethral radiofrequency collagen denaturation. Methods. This prospective study included 139 women with stress urinary incontinence due to bladder outlet hypermobility. Radiofrequency collagen denaturation was performed using local anesthesia in an office setting. Assessments included incontinence quality of life (I-QOL and urogenital distress inventory (UDI-6 instruments. Results. In total, 139 women were enrolled and 136 women were treated (mean age, 47 years. At 36 months, intent-to-treat analysis (n=139 revealed significant improvements in quality of life. Mean I-QOL score improved 17 points from baseline (P=.0004, while mean UDI-6 score improved (decreased 19 points (P=.0005. Conclusions. Transurethral collagen denaturation is a low-risk, office-based procedure that results in durable quality-of-life improvements in a significant proportion of women for as long as 3 years.

  11. Effects of the protein denaturant guanidinium chloride on aqueous hydrophobic contact-pair interactions.

    Science.gov (United States)

    Macdonald, Ryan D; Khajehpour, Mazdak

    2015-01-01

    Guanidinium chloride (GdmCl) is one of the most common protein denaturants. Although GdmCl is well known in the field of protein folding, the mechanism by which it denatures proteins is not well understood. In fact, there are few studies looking at its effects on hydrophobic interactions. In this work the effect of GdmCl on hydrophobic interactions has been studied by observing how the denaturant influences model systems of phenyl and alkyl hydrophobic contact pairs. Contact pair formation is monitored through the use of fluorescence spectroscopy, i.e., measuring the intrinsic phenol fluorescence being quenched by carboxylate ions. Hydrophobic interactions are isolated from other interactions through a previously developed methodology. The results show that GdmCl does not significantly affect hydrophobic interactions between small moieties such as methyl groups and phenol; while on the other hand, the interaction of larger hydrophobes such as hexyl and heptyl groups with phenol is significantly destabilized.

  12. BayFish: Bayesian inference of transcription dynamics from population snapshots of single-molecule RNA FISH in single cells

    National Research Council Canada - National Science Library

    Mariana Gomez-Schiavon; Liang-Fu Chen; Anne E West; Nicolas E Buchler

    2017-01-01

    Single-molecule RNA fluorescence in situ hybridization (smFISH) provides unparalleled resolution in the measurement of the abundance and localization of nascent and mature RNA transcripts in fixed, single cells...

  13. Quantitative fluorescent in-situ hybridization: a hypothesized competition mode between two dominant bacteria groups in hydrogen-producing anaerobic sludge processes.

    Science.gov (United States)

    Huang, C-L; Chen, C-C; Lin, C-Y; Liu, W-T

    2009-01-01

    Two hydrogen-producing continuous flow stirred tank reactors (CSTRs) fed respectively with glucose and sucrose were investigated by polymerase chain reaction-denatured gradient gel electrophoresis (PCR-DGGE) and fluorescent in-situ hybridization (FISH). The substrate was fed in a continuous mode decreased from hydraulic retention time (HRT) 10 hours to 6, 5, 4, 3, and 2 hours. Quantitative fluorescent in-situ hybridization (FISH) observations further demonstrated that two morphotypes of bacteria dominated both microbial communities. One was long rod bacteria which can be targeted either by Chis150 probe designed to hybridize the gram positive low G + C bacteria or the specific oligonucleotide probe Lg10-6. The probe Lg10-6, affiliated with Clostridium pasteurianum, was designed and then checked with other reference organisms. The other type, unknown group, which cannot be detected by Chis150 was curved rod bacteria. Notably, the population ratios of the two predominant groups reflected the different operational performance of the two reactors, such as hydrogen producing rates, substrate turnover rates and metabolites compositions. Therefore, a competition mode of the two dominant bacteria groups was hypothesized. In the study, 16S rRNA-based gene library of hydrogen-producing microbial communities was established. The efficiency of hydrogen yields was correlated with substrates (glucose or sucrose), HRT, metabolites compositions (acetate, propionate, butyrate and ethanol), thermal pre-treatment (seed biomass was heated at 100 degrees C for 45 minutes), and microbial communities in the bioreactor, not sludge sources (municipal sewage sludge, alcohol-processing sludge, or bean-processing sludge). The designed specific oligonucleotide probe Lg10-6 also provides us a useful and fast molecular tool to screen hydrogen-producing microbial communities in the future research.

  14. Do bacteria, not fish, produce 'fish kairomone'?

    NARCIS (Netherlands)

    Ringelberg, J.; Van Gool, E.

    1998-01-01

    Fish-associated chemicals enhance phototactic downward swimming in Daphnia. If perch were treated with the antibiotic ampicillin, this enhancement was significantly decreased. Therefore, not fish, but bacteria associated with fish, seem to produce this kairomone. [KEYWORDS: Diel vertical migration;

  15. Impact of organic modifier and temperature on protein denaturation in hydrophobic interaction chromatography.

    Science.gov (United States)

    Bobaly, Balázs; Beck, Alain; Veuthey, Jean-Luc; Guillarme, Davy; Fekete, Szabolcs

    2016-11-30

    The goal of this study was to better understand the chromatographic conditions in which monoclonal antibodies (mAbs) of broad hydrophobicity scale and a cysteine conjugated antibody-drug conjugate (ADCs), namely brentuximab-vedotin, could denaturate. For this purpose, some experiments were carried out in HIC conditions using various organic modifier in natures and proportions, different mobile phase temperatures and also different pHs. Indeed, improper analytical conditions in hydrophobic interaction chromatography (HIC) may create reversed-phase (RP) like harsh conditions and therefore protein denaturation. In terms of organic solvents, acetonitrile (ACN) and isopropanol (IPA) were tested with proportions ranging from 0 to 40%. It appeared that IPA was a less denaturating solvent than ACN, but should be used in a reasonable range (10-15%). Temperature should also be kept reasonable (below 40°C), to limit denaturation under HIC conditions. However, the combined increase of temperature and organic content induced denaturation of protein biopharmaceuticals in all cases. Indeed, above 30-40°C and 10-15% organic modifier in mobile phase B, heavy chain (HC) and light chain (LC) fragments dissociated. Mobile phase pH was also particularly critical and denaturation was significant even under moderately acidic conditions (pH of 5.4). Today, HIC is widely used for measuring drug-to-antibody ratio (DAR) of ADCs, which is a critical quality attribute of such samples. Here, we demonstrated that the estimation of average DAR can be dependent on the amount of organic modifier in the mobile phase under HIC conditions, due to the better recovery of the most hydrophobic proteins in presence of organic solvent (IPA). So, special care should be taken when measuring the average DAR of ADCs in HIC. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Effects of Glucides on Thermal Denaturation and Coagulation of Whey Proteins Studied by Ultraviolet Spectroscopy

    Science.gov (United States)

    Mongo Antoine, Etou; Abena, A. A.; Gbeassor, M.; Chaveron, H.

    The thermal coagulation of whey proteins concentrates was inhibited by various glucides. The disaccharides, saccharose and lactose, were most effective and the amino sugar, glucosamine, least effective in this respect. Ultraviolet absorption and light-scattering measurements on thermal denaturation and coagulation of both unfractionated and individual whey proteins (α-lactalbumin, ß-lactoglobulin and bovine serum albumin) showed that saccharose promotes the denaturation of these proteins but inhibits their subsequent coagulation. These results are interpreted in terms of the effect of saccharose on the hydrophobic interactions between solvent and protein.

  17. Surprisingly high stability of barley lipid transfer protein, LTP1, towards denaturant, heat and proteases

    DEFF Research Database (Denmark)

    Lindorff-Larsen, Kresten; Winther, J R

    2001-01-01

    Barley LTP1 belongs to a large family of plant proteins termed non-specific lipid transfer proteins. The in vivo function of these proteins is unknown, but it has been suggested that they are involved in responses towards stresses such as pathogens, drought, heat, cold and salt. Also, the proteins...... have been suggested as transporters of monomers for cutin synthesis. We have analysed the stability of LTP1 towards denaturant, heat and proteases and found it to be a highly stable protein, which apparently does not denature at temperatures up to 100 degrees C. This high stability may be important...

  18. Refolding of Denatured/Reduced Lysozyme Using Weak-Cation Exchange Chromatography

    Institute of Scientific and Technical Information of China (English)

    Yan WANG; Bo Lin GONG; Xin Du GENG

    2003-01-01

    Oxidative refolding of the denatured/reduced lysozyme was investigated by using weak-cation exchange chromatography (WCX). The stationary phase of WCX binds to the reduced lysozyme and prevented it from forming intermolecular aggregates. At the same time urea and ammonium sulfate were added to the mobile phase to increase the elution strength for lysozyme. Ammonium sulfate can more stabilize the native protein than a common eluting agent, sodium chloride. Refolding of lysozyme by using this WCX is successfully. It was simply carried out to obtain a completely and correctly refolding of the denatured lysozyme at high concentration of 20.0 mg/mL.

  19. Effect of ethanol denaturant on gasoline RVP (revised). Topical report, June 21, 1993--December 31, 1993

    Energy Technology Data Exchange (ETDEWEB)

    Wu, L.; Timpe, R.C.

    1993-12-01

    The Clean Air Act (CAA) Amendments of 1990 require further reduction in gasoline Reid vapor pressure (RVP) to reduce pollution. This research focused on characterizing the effect of ethanol denaturant and water on the RVP of the final ethanol-blended fuel. Anectdotal stories tell of up to a 0.5-psi effect of ethanol denaturant on the RVP of the finished ethanol-blended gasoline. Additionally, earlier Energy & Environmental Research Center (EERC) data indicated water could have a significant effect on the RVP. It was necessary to scientifically verify these effects using acceptable laboratory protocols.

  20. The Fishing Cat

    Institute of Scientific and Technical Information of China (English)

    孙雅飞; 乐伟国

    2008-01-01

    @@ 一、故事内容 A cat goes fishing every day. He wants to eat fish, but he can't catch any fish. One day, he goes to the river as usual. Suddenly, a fish comes out. He catches the fish and putsthe fish in the basket. He's very happy, but he forgest to put the lid on the basket.

  1. Fish Vaccine Development and Use to Prevent Streptococcal Diseases

    Science.gov (United States)

    An important pathogen of tilapia, hybrid striped bass and trout raised in intensive aquaculture is Streptococcus sp., a cause of severe economic losses in the fish farming industry. Infected fish experience severe to moderate mortality due to Streptococcus iniae and/or S. agalactiae. The diseased ...

  2. Hybrid vehicles

    Energy Technology Data Exchange (ETDEWEB)

    West, J.G.W. [Electrical Machines (United Kingdom)

    1997-07-01

    The reasons for adopting hybrid vehicles result mainly from the lack of adequate range from electric vehicles at an acceptable cost. Hybrids can offer significant improvements in emissions and fuel economy. Series and parallel hybrids are compared. A combination of series and parallel operation would be the ideal. This can be obtained using a planetary gearbox as a power split device allowing a small generator to transfer power to the propulsion motor giving the effect of a CVT. It allows the engine to run at semi-constant speed giving better fuel economy and reduced emissions. Hybrid car developments are described that show the wide range of possible hybrid systems. (author)

  3. Fish gelatin.

    Science.gov (United States)

    Boran, Gokhan; Regenstein, Joe M

    2010-01-01

    Gelatin is a multifunctional ingredient used in foods, pharmaceuticals, cosmetics, and photographic films as a gelling agent, stabilizer, thickener, emulsifier, and film former. As a thermoreversible hydrocolloid with a narrower gap between its melting and gelling temperatures, both of which are below human body temperature, gelatin provides unique advantages over carbohydrate-based gelling agents. Gelatin is mostly produced from pig skin, and cattle hides and bones. Some alternative raw materials have recently gained attention from both researchers and the industry not just because they overcome religious concerns shared by Jews and Muslims but also because they provide, in some cases, technological advantages over mammalian gelatins. Fish skins from a number of fish species are among the other sources that have been comprehensively studied as sources for gelatin production. Fish skins have a significant potential for the production of high-quality gelatin with different melting and gelling temperatures over a much wider range than mammalian gelatins, yet still have a sufficiently high gel strength and viscosity. Gelatin quality is industrially determined by gel strength, viscosity, melting or gelling temperatures, the water content, and microbiological safety. For gelatin manufacturers, yield from a particular raw material is also important. Recent experimental studies have shown that these quality parameters vary greatly depending on the biochemical characteristics of the raw materials, the manufacturing processes applied, and the experimental settings used for quality control tests. In this review, the gelatin quality achieved from different fish species is reviewed along with the experimental procedures used to determine gelatin quality. In addition, the chemical structure of collagen and gelatin, the collagen-gelatin conversion, the gelation process, and the gelatin market are discussed.

  4. Fish hemoglobins

    Directory of Open Access Journals (Sweden)

    P.C. de Souza

    2007-06-01

    Full Text Available Vertebrate hemoglobin, contained in erythrocytes, is a globular protein with a quaternary structure composed of 4 globin chains (2 alpha and 2 beta and a prosthetic group named heme bound to each one. Having myoglobin as an ancestor, hemoglobin acquired the capacity to respond to chemical stimuli that modulate its function according to tissue requirements for oxygen. Fish are generally submitted to spatial and temporal O2 variations and have developed anatomical, physiological and biochemical strategies to adapt to the changing environmental gas availability. Structurally, most fish hemoglobins are tetrameric; however, those from some species such as lamprey and hagfish dissociate, being monomeric when oxygenated and oligomeric when deoxygenated. Fish blood frequently possesses several hemoglobins; the primary origin of this finding lies in the polymorphism that occurs in the globin loci, an aspect that may occasionally confer advantages to its carriers or even be a harmless evolutionary remnant. On the other hand, the functional properties exhibit different behaviors, ranging from a total absence of responses to allosteric regulation to drastic ones, such as the Root effect.

  5. Heat denaturation of soy glycinin. Structural characteristics in relation to aggregation and gel formation.

    NARCIS (Netherlands)

    Lakemond, C.M.M.

    2001-01-01

    key words: soy protein; glycinin; thermal stability; pH; ionic strength;genetic variant; solubility; gelationThe main aim of this thesis was to study structural changes of soy glycinin at different conditions (pH and ionic strength) during thermal denaturation and their effect on aggregation and gel

  6. Joule Heating and Thermal Denaturation of Proteins in Nano-ESI Theta Tips

    Science.gov (United States)

    Zhao, Feifei; Matt, Sarah M.; Bu, Jiexun; Rehrauer, Owen G.; Ben-Amotz, Dor; McLuckey, Scott A.

    2017-10-01

    Electro-osmotically induced Joule heating in theta tips and its effect on protein denaturation were investigated. Myoglobin, equine cytochrome c, bovine cytochrome c, and carbonic anhydrase II solutions were subjected to electro-osmosis in a theta tip and all of the proteins were denatured during the process. The extent of protein denaturation was found to increase with the applied square wave voltage and electrolyte concentration. The solution temperature at the end of a theta tip was measured directly by Raman spectroscopy and shown to increase with the square wave voltage, thereby demonstrating the effect of Joule heating through an independent method. The electro-osmosis of a solution comprised of myoglobin, bovine cytochrome c, and ubiquitin demonstrated that the magnitude of Joule heating that causes protein denaturation is positively correlated with protein melting temperature. This allows for a quick determination of a protein's relative thermal stability. This work establishes a fast, novel method for protein conformation manipulation prior to MS analysis and provides a temperature-controllable platform for the study of processes that take place in solution with direct coupling to mass spectrometry. [Figure not available: see fulltext.

  7. Manifestations of native topology in the denatured state ensemble of Rhodopseudomonas palustris cytochrome c'.

    Science.gov (United States)

    Dar, Tanveer A; Schaeffer, R Dustin; Daggett, Valerie; Bowler, Bruce E

    2011-02-15

    To provide insight into the role of local sequence in the nonrandom coil behavior of the denatured state, we have extended our measurements of histidine-heme loop formation equilibria for cytochrome c' to 6 M guanidine hydrochloride. We observe that there is some reduction in the scatter about the best fit line of loop stability versus loop size data in 6 M versus 3 M guanidine hydrochloride, but the scatter is not eliminated. The scaling exponent, ν(3), of 2.5 ± 0.2 is also similar to that found previously in 3 M guanidine hydrochloride (2.6 ± 0.3). Rates of histidine-heme loop breakage in the denatured state of cytochrome c' show that some histidine-heme loops are significantly more persistent than others at both 3 and 6 M guanidine hydrochloride. Rates of histidine-heme loop formation more closely approximate random coil behavior. This observation indicates that heterogeneity in the denatured state ensemble results mainly from contact persistence. When mapped onto the structure of cytochrome c', the histidine-heme loops with slow breakage rates coincide with chain reversals between helices 1 and 2 and between helices 2 and 3. Molecular dynamics simulations of the unfolding of cytochrome c' at 498 K show that these reverse turns persist in the unfolded state. Thus, these portions of the primary structure of cytochrome c' set up the topology of cytochrome c' in the denatured state, predisposing the protein to fold efficiently to its native structure.

  8. ASP53, a thermostable protein from Acacia erioloba seeds that protects target proteins against thermal denaturation

    CSIR Research Space (South Africa)

    Mtwisha, L

    2007-02-01

    Full Text Available stages of protein thermal denaturation. ASP53 decreased the rate of loss of alcohol dehydrogenase activity at 55°C, decreased the rate of temperature-dependent loss of secondary structure of haemoglobin and completely inhibited the temperature...

  9. Protecting role of cosolvents in protein denaturation by SDS: a structural study

    Directory of Open Access Journals (Sweden)

    Wouters Johan

    2008-06-01

    Full Text Available Abstract Background Recently, we reported a unique approach to preserve the activity of some proteins in the presence of the denaturing agent, Sodium Dodecyl Sulfate (SDS. This was made possible by addition of the amphipathic solvent 2,4-Methyl-2-PentaneDiol (MPD, used as protecting but also as refolding agent for these proteins. Although the persistence of the protein activity in the SDS/MPD mixture was clearly established, preservation of their structure was only speculative until now. Results In this paper, a detailed X-ray study addresses the pending question. Crystals of hen egg-white lysozyme were grown for the first time in the presence of MPD and denaturing concentrations of SDS. Depending on crystallization conditions, tetragonal crystals in complex with either SDS or MPD were collected. The conformation of both structures was very similar to the native lysozyme and the obtained complexes of SDS-lysozyme and MPD-lysozyme give some insights in the interplay of protein-SDS and protein-MPD interactions. Conclusion This study clearly established the preservation of the enzyme structure in a SDS/MPD mixture. It is hypothesized that high concentrations of MPD would change the properties of SDS and lower or avoid interactions between the denaturant and the protein. These structural data therefore support the hypothesis that MPD avoids disruption of the enzyme structure by SDS and can protect proteins from SDS denaturation.

  10. Denaturing gradient gel electrophoresis analysis to study bacterial community structure in pockets of periodontitis patients

    NARCIS (Netherlands)

    Zijnge, V.; Harmsen, H.J.M.; Kleinfelder, J.W.; Rest, M.E. van der; Degener, J.E.; Welling, G.W.

    2003-01-01

    Bacteria are involved in the onset and progression of periodontitis. A promising molecular technique, denaturing gradient gel electrophoresis (DGGE), to study microbial population dynamics in the subgingival pocket is presented. Twenty-three samples were taken from the subgingival pockets of nine pa

  11. Investigating the fermentation of cocoa by correlating denaturing gradient gel electrophoresis profiles and near infrared spectra

    DEFF Research Database (Denmark)

    Nielsen, Dennis Sandris; Snitkjær, Pia; van der Berg, Franciscus Winfried J

    2008-01-01

    of the beans and the chemical processes inside the beans have been carried out previously. Recently it has been shown that Denaturing Gradient Gel Electrophoresis (DGGE) offers an efficient tool for monitoring the microbiological changes taking place during the fermentation of cocoa. Near Infrared (NIR...

  12. A simple remedy against artifactual double bands in denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Janse, I.; Bok, J.M.; Zwart, G.

    2004-01-01

    Denaturant gradient gel electrophoresis (DGGE) is a widely used method for mutation analysis and for studies of microbial diversity. Particular combinations of target gene fragments and primers may give rise to erroneous DGGE profiles. We report on a very straightforward means to eliminate the

  13. Thermal denaturation of protein studied by terahertz time-domain spectroscopy

    Science.gov (United States)

    Fu, Xiuhua; Li, Xiangjun; Liu, Jianjun; Du, Yong; Hong, Zhi

    2012-12-01

    In this study, the absorption spectra of native or thermal protein were measured in 0.2-1.4THz using terahertz time-domain spectroscopy (THz-TDS) system at room temperature, their absorption spectra and the refractive spectra were obtained. Experimental results indicate that protein both has strong absorption but their characteristics were not distinct in the THz region, and the absorption decreased during thermal denatured state. In order to prove protein had been denatured, we used Differential scanning calorimeter (DSC) measured their denatured temperature, from their DSC heating traces, collagen Td=101℃, Bovine serum albumin Td=97℃. While we also combined the Fourier transform infrared spectrometer (FTIR) to investigate their secondary and tertiary structure before and after denatuation, but the results did not have the distinct changes. We turned the absorption spectra and the refractive spectra to the dielectric spectra, and used the one-stage Debye model simulated the terahertz dielectric spectra of protein before and after denaturation. This research proved that the terahertz spectrum technology is feasible in testing protein that were affected by temperature or other factors which can provide theoretical foundation in the further study about the THz spectrum of protein and peptide temperature stability.

  14. Mesoscopic modeling of DNA denaturation rates: Sequence dependence and experimental comparison

    Energy Technology Data Exchange (ETDEWEB)

    Dahlen, Oda, E-mail: oda.dahlen@ntnu.no; Erp, Titus S. van, E-mail: titus.van.erp@ntnu.no [Department of Chemistry, Norwegian University of Science and Technology (NTNU), Høgskoleringen 5, Realfagbygget D3-117 7491 Trondheim (Norway)

    2015-06-21

    Using rare event simulation techniques, we calculated DNA denaturation rate constants for a range of sequences and temperatures for the Peyrard-Bishop-Dauxois (PBD) model with two different parameter sets. We studied a larger variety of sequences compared to previous studies that only consider DNA homopolymers and DNA sequences containing an equal amount of weak AT- and strong GC-base pairs. Our results show that, contrary to previous findings, an even distribution of the strong GC-base pairs does not always result in the fastest possible denaturation. In addition, we applied an adaptation of the PBD model to study hairpin denaturation for which experimental data are available. This is the first quantitative study in which dynamical results from the mesoscopic PBD model have been compared with experiments. Our results show that present parameterized models, although giving good results regarding thermodynamic properties, overestimate denaturation rates by orders of magnitude. We believe that our dynamical approach is, therefore, an important tool for verifying DNA models and for developing next generation models that have higher predictive power than present ones.

  15. Affinity chromatography of chaperones based on denatured proteins: Analysis of cell lysates of different origin.

    Science.gov (United States)

    Marchenko, N Yu; Sikorskaya, E V; Marchenkov, V V; Kashparov, I A; Semisotnov, G V

    2016-03-01

    Molecular chaperones are involved in folding, oligomerization, transport, and degradation of numerous cellular proteins. Most of chaperones are heat-shock proteins (HSPs). A number of diseases of various organisms are accompanied by changes in the structure and functional activity of chaperones, thereby revealing their vital importance. One of the fundamental properties of chaperones is their ability to bind polypeptides lacking a rigid spatial structure. Here, we demonstrate that affinity chromatography using sorbents with covalently attached denatured proteins allows effective purification and quantitative assessment of their bound protein partners. Using pure Escherichia coli chaperone GroEL (Hsp60), the capacity of denatured pepsin or lysozyme-based affinity sorbents was evaluated as 1 mg and 1.4 mg of GroEL per 1 ml of sorbent, respectively. Cell lysates of bacteria (E. coli, Thermus thermophilus, and Yersinia pseudotuberculosis), archaea (Halorubrum lacusprofundi) as well as the lysate of rat liver mitochondria were analyzed using affinity carrier with denatured lysozyme. It was found that, apart from Hsp60, other proteins with a molecular weight of about 100, 50, 40, and 20 kDa are able to interact with denatured lysozyme.

  16. Reversible Dimerization of Acid-Denatured ACBP Controlled by Helix A4

    DEFF Research Database (Denmark)

    Fieber, Wolfgang; Kragelund, Birthe Brandt; Meldal, Morten Peter;

    2005-01-01

    of dimers and revealed a cooperative stabilization of helix A4 in this process. This emphasizes its special role in the structure formation in the denatured state of ACBP. No dimers are formed in the presence of guanidine hydrochloride, which underlines the fundamental difference between the nature...

  17. Sufficient minimal model for DNA denaturation: Integration of harmonic scalar elasticity and bond energies.

    Science.gov (United States)

    Singh, Amit Raj; Granek, Rony

    2016-10-14

    We study DNA denaturation by integrating elasticity - as described by the Gaussian network model - with bond binding energies, distinguishing between different base pairs and stacking energies. We use exact calculation, within the model, of the Helmholtz free-energy of any partial denaturation state, which implies that the entropy of all formed "bubbles" ("loops") is accounted for. Considering base pair bond removal single events, the bond designated for opening is chosen by minimizing the free-energy difference for the process, over all remaining base pair bonds. Despite of its great simplicity, for several known DNA sequences our results are in accord with available theoretical and experimental studies. Moreover, we report free-energy profiles along the denaturation pathway, which allow to detect stable or meta-stable partial denaturation states, composed of bubble, as local free-energy minima separated by barriers. Our approach allows to study very long DNA strands with commonly available computational power, as we demonstrate for a few random sequences in the range 200-800 base-pairs. For the latter, we also elucidate the self-averaging property of the system. Implications for the well known breathing dynamics of DNA are elucidated.

  18. Denaturing Effects of Urea and Guanidine Hydrochloride on Hyperthermophilic Esterase from Aeropyrum pernix K1

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The changes in the activity and the conformation of the hyperthermophilic esterase derived from aerobic thermophilic Aeropyrumpernix K1 (APE1547) were studied during denaturation by guanidine hydrochloride (GdnHCl)and urea. The denaturation course of APE1547 was followed by the steady-state and time resolved fluorescence methods. An increase in the denaturant concentration in the denatured system can significantly enhance the inactivation and unfolding of APE1547. The enzyme can be completely inactivated with a urea concentration of 2. 7 mol/L or a GdnHCl concentration of 7.5 mol/L. The fluorescence emission maximum of the enzyme protein red shifts in magnitude to a maximum value(355 nm) when the concentration of GdnHCl is 5.1 mol/L. The experimental results indicate that APE1547 has a high resistance to urea. Unfolding of APE1547 in GdnHCl(4.2-6.0 mol/L) was shown to be an irreversible process. The present results indicate that the ion pairs in this protein may be a key factor for the stability of this esterase.

  19. Sufficient minimal model for DNA denaturation: Integration of harmonic scalar elasticity and bond energies

    Science.gov (United States)

    Singh, Amit Raj; Granek, Rony

    2016-10-01

    We study DNA denaturation by integrating elasticity — as described by the Gaussian network model — with bond binding energies, distinguishing between different base pairs and stacking energies. We use exact calculation, within the model, of the Helmholtz free-energy of any partial denaturation state, which implies that the entropy of all formed "bubbles" ("loops") is accounted for. Considering base pair bond removal single events, the bond designated for opening is chosen by minimizing the free-energy difference for the process, over all remaining base pair bonds. Despite of its great simplicity, for several known DNA sequences our results are in accord with available theoretical and experimental studies. Moreover, we report free-energy profiles along the denaturation pathway, which allow to detect stable or meta-stable partial denaturation states, composed of bubble, as local free-energy minima separated by barriers. Our approach allows to study very long DNA strands with commonly available computational power, as we demonstrate for a few random sequences in the range 200-800 base-pairs. For the latter, we also elucidate the self-averaging property of the system. Implications for the well known breathing dynamics of DNA are elucidated.

  20. Lysozyme Thermal Denaturation and Self-Interaction: Four Integrated Thermodynamic Experiments for the Physical Chemistry Laboratory

    Science.gov (United States)

    Schwinefus, Jeffrey J.; Schaefle, Nathaniel J.; Muth, Gregory W.; Miessler, Gary L.; Clark, Christopher A.

    2008-01-01

    As part of an effort to infuse our physical chemistry laboratory with biologically relevant, investigative experiments, we detail four integrated thermodynamic experiments that characterize the denaturation (or unfolding) and self-interaction of hen egg white lysozyme as a function of pH and ionic strength. Students first use Protein Explorer to…

  1. Triggered isothermal PCR by denaturation bubble-mediated strand exchange amplification.

    Science.gov (United States)

    Shi, Chao; Shang, Fanjin; Zhou, Meiling; Zhang, Pansong; Wang, Yifan; Ma, Cuiping

    2016-10-04

    Here, we introduced the concept of strand exchange amplification (SEA) mediated by denaturation bubbles. Similar to traditional PCR, it only employed a DNA polymerase and a pair of common primers to realize a three-step cycle process, but the entire SEA reaction was performed at a single temperature.

  2. Comparison of the fine structure of mitochondrial DNA from Saccharomyces cerevisiae and S. carlsbergensis: electron microscopy of partially denatured molecules

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C

    1976-01-01

    Denaturation-maps of mitochondrial DNA from Saccharomyces cerevisiae and S. carlsbergensis have been derived from electron microscopic observations of partially denatured complete circular molecules and large fragments of these circles. The mitochondrial DNA from the two species differ by 6...

  3. Reversible denaturation of Brazil nut 2S albumin (Ber e1) and implication of structural destabilization on digestion by pepsin

    NARCIS (Netherlands)

    Koppelman, S.J.; Nieuwenhuizen, W.F.; Gaspari, M.; Knippels, L.M.J.; Penninks, A.H.; Knol, E.F.; Hefle, S.L.; Jongh, H.H.J.de

    2005-01-01

    The high resistance of Brazil nut 2S albumin, previously identified as an allergen, against proteolysis by pepsin was examined in this work. Although the denaturation temperature of this protein exceeds the 110 °C at neutral pH, at low pH a fully reversible thermal denaturation was observed at ∼82 °

  4. Structural properties of cyanase. Denaturation, renaturation, and role of sulfhydryls and oligomeric structure in catalytic activity.

    Science.gov (United States)

    Little, R M; Anderson, P M

    1987-07-25

    Cyanase is an inducible enzyme in Escherichia coli that catalyzes bicarbonate-dependent decomposition of cyanate to give ammonia and bicarbonate. The enzyme is composed of 8-10 identical subunits (Mr = 17,008). The objective of this study was to clarify some of the structural properties of cyanase for the purpose of understanding the relationship between oligomeric structure and catalytic activity. Circular dichroism studies showed that cyanase has a significant amount of alpha-helix and beta-sheet structure. The one sulfhydryl group per subunit does not react with 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) unless cyanase is denatured. Denaturation is apparently complete in 10 M urea or 6 M guanidine hydrochloride, but is significantly reduced in 10 M urea by the presence of azide (analog of cyanate) and is incomplete in 8 M urea. Denatured cyanase could be renatured and reactivated (greater than 85%) by removal of denaturants. Reactivation was greatly facilitated by the presence of certain anions, particularly bicarbonate, and by high ionic strength and protein concentration. The catalytic activity of renatured cyanase was associated only with oligomer. Cyanase that had been denatured in the presence of DTNB to give a cyanase-DTNB derivative could also be renatured at 26 degrees C to give active cyanase-DTNB oligomer. The active oligomeric form of the cyanase-DTNB derivative could be converted reversibly to inactive dimer by lowering the temperature to 4 degrees C or by reduction of the ionic strength and removal of monoanions. These results provide evidence that free sulfhydryl groups are not required for catalytic activity and that catalytic activity may be dependent upon oligomeric structure.

  5. [Some properties of complexes formed by small heat shock proteins with denatured actin].

    Science.gov (United States)

    Pivovarova, A V; Chebotareva, N A; Guseev, N B; Levitskiĭ, D I

    2008-01-01

    We applied different methods to analyze the effects of the recombinant wild-type small heat shock protein with an apparent molecular mass of 27 kD (Hsp27-wt) and its S15,78,82D mutant (Hsp27-3D), which mimics the naturally occurring phosphorylation of this protein, on the thermal denaturation and aggregation of F-actin. It has been shown that, at the weight ratio of Hsp27/actin equal to 1/4, both Hsp27-wt and Hsp27-3D do not affect the thermal unfolding of F-actin but effectively prevent the aggregation of F-actin by forming soluble complexes with denatured actin. The formation of these complexes occurs upon heating and accompanies the F-actin thermal denaturation. It is known that Hsp27-wt forms high-molecular-mass oligomers, whereas Hsp27-3D forms small dimers or tetramers. However, the complexes formed by Hsp27-wt and Hsp27-3D with denatured actin did not differ in their size, as measured by dynamic light scattering, and demonstrated the same hydrodynamic radius of 17-18 nm. On the other hand, the sedimentation coefficients of these complexes were distributed within the range 10-45 S in the case of Hsp27-3D and 18-60 S in the case of Hsp27-wt. Thus, the ability of Hsp27 to form soluble complexes with denatured actin does not significantly depend on the initial oligomeric state of Hsp27.

  6. Evaluación de los desechos de pescado frescos y ensilados en la alimentación de híbridos de Clarias gariepinus X C. macrocephalus (Fresh fishing offals and silage evaluation for hybrid of Clarias gariepinus X C. macrocephalus feeding

    Directory of Open Access Journals (Sweden)

    Llanes Iglesias, José E

    2007-09-01

    Full Text Available ResumenSe evaluó el uso de los desechos frescos de pescado y sus respectivos ensilados por vías biológica y bioquímica, como única fuente de proteína animal en dietas húmedas para alevines de híbridos de Clarias gariepinus X C. macrocephalus (4,53+ 0,05g, las que fueron comparadas con un Alimento Comercial (20% de harina de desechos de pescado durante 60 días. Los resultados mostraron que los mejores indicadores de crecimiento y utilización de la proteína fueron obtenidos con los desechos frescos, los que difirieron significativamente (P85%, fue similar entre las dietas. Los ensilados y la harina de pescado no difirieron significativamente en los indicadores evaluados, siendo una alternativa para la producción de alimentos acuícola para países en vía de desarrollo. AbstractThis study evaluated the use of fresh fishes wastes and their respectives silages by biologic and quimic methodogies like only one source of animal protein in moist diets for hybrid of Clarias gariepinus X C. macrocephalus (4,53+ 0,05g. These were compared with a comercial foods (20% of fish wastes meal, during 60 days. The results showed that better growth and use of protein were obtened with fresh waste, what were differed significatively (P<0.05, to silage and fish meal. Beside, the use of food conversión was similar between the diets. The silage and fish meal did not differ significatively in the indicators evaluated.

  7. Epidermal Growth Factor Receptor (EGFR gene copy number (GCN correlates with clinical activity of irinotecan-cetuximab in K-RAS wild-type colorectal cancer: a fluorescence in situ (FISH and chromogenic in situ hybridization (CISH analysis

    Directory of Open Access Journals (Sweden)

    Scartozzi Mario

    2009-08-01

    Full Text Available Abstract Background K-RAS wild type colorectal tumors show an improved response rate to anti-EGFR monoclonal antibodies. Nevertheless 70% to 40% of these patients still does not seem to benefit from this therapeutic approach. FISH EGFR GCN has been previously demonstrated to correlate with clinical outcome of colorectal cancer treated with anti-EGFR monoclonal antibodies. CISH also seemed able to provide accurate EGFR GCN information with the advantage of a simpler and reproducible technique involving immunohistochemistry and light microscopy. Based on these findings we investigated the correlation between both FISH and CISH EGFR GCN and clinical outcome in K-RAS wild-type colorectal cancer treated with irinotecan-cetuximab. Methods Patients with advanced K-RAS wild-type, colorectal cancer receiving irinotecan-cetuximab after failure of irinotecan-based chemotherapy were eligible. A cut-off value for EGFR GCN of 2.6 and 2.12 for FISH and CISH respectively was derived from ROC curve analysis. Results Forty-four patients were available for analysis. We observed a partial remission in 9 (60% and 2 (9% cases with a FISH EGFR GCN ≥ 2.6 and Conclusion FISH and CISH EGFR GCN may both represent effective tools for a further patients selection in K-RAS wild-type colorectal cancer treated with cetuximab.

  8. Fish Tales

    Energy Technology Data Exchange (ETDEWEB)

    McLerran, L.

    2010-07-06

    This talk is about fishing and the friendships that have resulted in its pursuit. It is also about theoretical physics, and the relationship of imagination and fantasy to the establishment of ideas about nature. Fishermen, like theoretical physicists, are well known for their inventive imaginations. Perhaps neither are as clever as sailors, who conceived of the mermaid. If one doubts the power of this fantasy, one should remember the ghosts of the many sailors who drowned pursuing these young nymphs. An extraordinary painting by J. Waterhouse is shown as Fig. 1. The enchantment of a mermaid must reflect an extraordinary excess of imagination on the part of the sailor, perhaps together with an impractical turn of mind. A consummated relationship with a mermaid is after all, by its very nature a fantasy incapable of realization. To a theoretical physicist, she is symbolic of many ideas we develop. There are many truths known to fisherman in which one might also find parallels to the goals of scientists: (1) A fish is the only animal that keeps growing after its death; (2) Nothing makes a fish bigger than almost being caught; (3) ''...of all the liars among mankind, the fisherman is the most trustworthy.'' (William Sherwood Fox, in Silken Lines and Silver Hooks); and (4) Men and fish are alike. They both get into trouble when they open their mouths. These quotes may be interpreted as reflecting skepticism regarding the honesty of fisherman, and probably do not reflect adequate admiration for a creative imagination. Is it fair to criticize a person for believing a falsehood that he or she sincerely believes to be true? The fisherman simultaneously invents the lie, and believes in it himself. The parallel with theoretical physics is perhaps only approximate, although we physicists may invent stories that we come to believe, on some rare occasions our ideas actually correspond to a more or less true descriptions of nature. These minor philosophical

  9. Fish Immunoglobulins

    Science.gov (United States)

    Mashoof, Sara; Criscitiello, Michael F.

    2016-01-01

    The B cell receptor and secreted antibody are at the nexus of humoral adaptive immunity. In this review, we summarize what is known of the immunoglobulin genes of jawed cartilaginous and bony fishes. We focus on what has been learned from genomic or cDNA sequence data, but where appropriate draw upon protein, immunization, affinity and structural studies. Work from major aquatic model organisms and less studied comparative species are both included to define what is the rule for an immunoglobulin isotype or taxonomic group and what exemplifies an exception. PMID:27879632

  10. Fishing activities

    Science.gov (United States)

    Oberle, Ferdinand; Puig, Pere; Martin, Jacobo; Micallef, Aaron; Krastel, Sebastian; Savini, Alessandra

    2018-01-01

    Unlike the major anthropogenic changes that terrestrial and coastal habitats underwent during the last centuries such as deforestation, river engineering, agricultural practices or urbanism, those occurring underwater are veiled from our eyes and have continued nearly unnoticed. Only recent advances in remote sensing and deep marine sampling technologies have revealed the extent and magnitude of the anthropogenic impacts to the seafloor. In particular, bottom trawling, a fishing technique consisting of dragging a net and fishing gear over the seafloor to capture bottom-dwelling living resources has gained attention among the scientific community, policy makers and the general public due to its destructive effects on the seabed. Trawling gear produces acute impacts on biota and the physical substratum of the seafloor by disrupting the sediment column structure, overturning boulders, resuspending sediments and imprinting deep scars on muddy bottoms. Also, the repetitive passage of trawling gear over the same areas creates long-lasting, cumulative impacts that modify the cohesiveness and texture of sediments. It can be asserted nowadays that due to its recurrence, mobility and wide geographical extent, industrial trawling has become a major force driving seafloor change and affecting not only its physical integrity on short spatial scales but also imprinting measurable modifications to the geomorphology of entire continental margins.

  11. Deep Fish.

    Science.gov (United States)

    Ishaq, Omer; Sadanandan, Sajith Kecheril; Wählby, Carolina

    2017-01-01

    Zebrafish ( Danio rerio) is an important vertebrate model organism in biomedical research, especially suitable for morphological screening due to its transparent body during early development. Deep learning has emerged as a dominant paradigm for data analysis and found a number of applications in computer vision and image analysis. Here we demonstrate the potential of a deep learning approach for accurate high-throughput classification of whole-body zebrafish deformations in multifish microwell plates. Deep learning uses the raw image data as an input, without the need of expert knowledge for feature design or optimization of the segmentation parameters. We trained the deep learning classifier on as few as 84 images (before data augmentation) and achieved a classification accuracy of 92.8% on an unseen test data set that is comparable to the previous state of the art (95%) based on user-specified segmentation and deformation metrics. Ablation studies by digitally removing whole fish or parts of the fish from the images revealed that the classifier learned discriminative features from the image foreground, and we observed that the deformations of the head region, rather than the visually apparent bent tail, were more important for good classification performance.

  12. 荧光原位杂交技术在检测胎儿染色体亚显微结构异常中的应用%Application of the fluorescent in situ hybridization (FISH) on the prenatal diagnosis of the subtle chromosomal aberrations

    Institute of Scientific and Technical Information of China (English)

    尹爱华; 刘舒; 潘小英; 傅文婷; 王继成; 卢建; 杨洁霞

    2012-01-01

    Objective To study the diagnostic value of the fluorescent in situ hybridization on detection of subtle chromosomal aberrations in cultured amniotic fluid cells. Methods Amniocenteses were performed in a pregnant women of 18 gestational weeks, and metaphase FISH was followed because the results of amniotic fluid karyotype and ultrasound were disagree with each other. The metaphase chromosomes were hybridized in situ with the human centromere probes of chromosome 18,X,Y and site specific probe of chromosome 13,21. The treated slides were examined and taken photos under the carl zeiss fluoromicroscope. Result Subtle translocation of chromosome Y was detected by FISH. Conclusion Our result indicated that it is very helpful for the diagnosis of subtle chromosomal aberrations by combining FISH with traditional karyotype analysis.%目的 探讨荧光原位杂交(FISH)技术在诊断培养后的羊水细胞染色体亚显微结构异常中的应用价值.方法 对1例18孕周,经典细胞遗传学羊水染色体核型分析结果与B超检查结果有不符合的胎儿,应用FISH的18号、X、Y染色体着丝粒探针和13、21号染色体位点特异性探针,对培养后的羊水中期细胞标本进行检测.结果 共分析了22个独立细胞克隆的分裂象,发现胎儿染色体存在两种核型嵌合,结果记为:mos 45,X[20]/46,XY[2];FISH检测发现此胎儿核型存在Y染色体亚显微小片段易位.结论 FISH技术结合传统细胞遗传学核型分析,对于诊断染色体亚显微结构异常非常重要.

  13. Investigation of Collective Behaviour and Electrocommunication in the Weakly Electric Fish, Mormyrus rume, through a biomimetic Robotic Dummy Fish.

    Science.gov (United States)

    Donati, Elisa; Worm, Martin; Mintchev, Stefano; van der Wiel, Marleen; Benelli, Giovanni; von der Emde, Gerhard; Stefanini, Cesare

    2016-12-01

    A robotic fish has been developed to create a mixed bio-hybrid system made up of weakly electric fish and a mobile dummy fish. Weakly electric fish are capable of interacting with each other via sequences of self-generated electric signals during electrocommunication. Here we present the design of an artificial dummy fish, which is subsequently tested in behavioural experiments. The robot consists of two parts: a flexible tail that can move at different frequencies and amplitudes, performing a carangiform oscillation, and a rigid head containing the motor for the tail oscillation. The dummy fish mimics the weakly electric fish Mormyrus rume in morphology, size and electric signal generation. In order to study electrical interactions, the dummy fish is equipped with ten electrodes that record electric signals of nearby real fish and generate electric dipole fields around itself that are similar to those produced by real fish in both waveform and sequence. Behavioural experiments demonstrate that the dummy fish is able to recruit both single individuals and groups of M. rume from a shelter into an exposed area. The development of an artificial dummy fish may help to understand fundamental aspects of collective behaviour in weakly electric fish and the properties necessary to initiate and sustain it in closed-loop feedback experiments based on electrocommunication.

  14. Comprehensive phylogenetic analysis of all species of swordtails and platies (Pisces: Genus Xiphophorus uncovers a hybrid origin of a swordtail fish, Xiphophorus monticolus, and demonstrates that the sexually selected sword originated in the ancestral lineage of the genus, but was lost again secondarily

    Directory of Open Access Journals (Sweden)

    Kang Ji Hyoun

    2013-01-01

    Full Text Available Abstract Background Males in some species of the genus Xiphophorus, small freshwater fishes from Meso-America, have an extended caudal fin, or sword – hence their common name “swordtails”. Longer swords are preferred by females from both sworded and – surprisingly also, non-sworded (platyfish species that belong to the same genus. Swordtails have been studied widely as models in research on sexual selection. Specifically, the pre-existing bias hypothesis was interpreted to best explain the observed bias of females in presumed ancestral lineages of swordless species that show a preference for assumed derived males with swords over their conspecific swordless males. However, many of the phylogenetic relationships within this genus still remained unresolved. Here we construct a comprehensive molecular phylogeny of all 26 known Xiphophorus species, including the four recently described species (X. kallmani, X. mayae, X. mixei and X. monticolus. We use two mitochondrial and six new nuclear markers in an effort to increase the understanding of the evolutionary relationships among the species in this genus. Based on the phylogeny, the evolutionary history and character state evolution of the sword was reconstructed and found to have originated in the common ancestral lineage of the genus Xiphophorus and that it was lost again secondarily. Results We estimated the evolutionary relationships among all known species of the genus Xiphophorus based on the largest set of DNA markers so far. The phylogeny indicates that one of the newly described swordtail species, Xiphophorus monticolus, is likely to have arisen through hybridization since it is placed with the southern platyfish in the mitochondrial phylogeny, but with the southern swordtails in the nuclear phylogeny. Such discordance between these two types of markers is a strong indication for a hybrid origin. Additionally, by using a maximum likelihood approach the possession of the sexually

  15. Comprehensive phylogenetic analysis of all species of swordtails and platies (Pisces: Genus Xiphophorus) uncovers a hybrid origin of a swordtail fish, Xiphophorus monticolus, and demonstrates that the sexually selected sword originated in the ancestral lineage of the genus, but was lost again secondarily.

    Science.gov (United States)

    Kang, Ji Hyoun; Schartl, Manfred; Walter, Ronald B; Meyer, Axel

    2013-01-29

    Males in some species of the genus Xiphophorus, small freshwater fishes from Meso-America, have an extended caudal fin, or sword - hence their common name "swordtails". Longer swords are preferred by females from both sworded and - surprisingly also, non-sworded (platyfish) species that belong to the same genus. Swordtails have been studied widely as models in research on sexual selection. Specifically, the pre-existing bias hypothesis was interpreted to best explain the observed bias of females in presumed ancestral lineages of swordless species that show a preference for assumed derived males with swords over their conspecific swordless males. However, many of the phylogenetic relationships within this genus still remained unresolved. Here we construct a comprehensive molecular phylogeny of all 26 known Xiphophorus species, including the four recently described species (X. kallmani, X. mayae, X. mixei and X. monticolus). We use two mitochondrial and six new nuclear markers in an effort to increase the understanding of the evolutionary relationships among the species in this genus. Based on the phylogeny, the evolutionary history and character state evolution of the sword was reconstructed and found to have originated in the common ancestral lineage of the genus Xiphophorus and that it was lost again secondarily. We estimated the evolutionary relationships among all known species of the genus Xiphophorus based on the largest set of DNA markers so far. The phylogeny indicates that one of the newly described swordtail species, Xiphophorus monticolus, is likely to have arisen through hybridization since it is placed with the southern platyfish in the mitochondrial phylogeny, but with the southern swordtails in the nuclear phylogeny. Such discordance between these two types of markers is a strong indication for a hybrid origin. Additionally, by using a maximum likelihood approach the possession of the sexually selected sword trait is shown to be the most likely

  16. Effects of urea denatureation and pH on the ability of porcine myoglobin to undergo reduction.

    Science.gov (United States)

    Zhu, L G; Brewer, M S

    2003-04-01

    To determine the effects of globin moiety denaturation and pH on the ability of metmyoglobin (MetMb) to undergo reduction, MetMb isolated from porcine hearts was denatured in 8.5M urea. Both native and denatured MetMb solutions were serially reduced with Na(2)S(2)O(4) (0, 7.5, 15, 18.75, 22.5, 26.25, 30, 30.75, and 45 umol). Reduction was conducted at pH 5, 5.2, 5.4, 5.6, 6, 6.2, 6.4, 6.6, and 7. After reduction, absorbance was determined at 635 nm and the percent of the original MetMb which was reduced was calculated. The average percent MetMb reduced from the native and denatured forms was 35 and 25%, respectively. pH significantly influenced the percentage of MetMb reduced, especially when pH was <6. If the MetMb was denatured prior to reduction, the influence of pH on its ability to undergo reduction was slight. The percentage of denatured MetMb reduced was higher at pH 7 than at all other pHs. High pH enhanced the ability of MetMb to undergo reduction; while low pH decreases it. Low pH may have denatured the native globin moiety.

  17. Bounded hybrid superiority in an avian hybrid zone: effects of mate, diet, and habitat choice.

    Science.gov (United States)

    Good, T P; Ellis, J C; Annett, C A; Pierotti, R

    2000-10-01

    differences in hatching and fledging success, with females paired to hybrid males showing better success compared to females paired to glaucous-winged gull males. Hybrids showed better hatching and fledging success in the north because hybrids are more like western gulls than glaucous-winged gulls in foraging behavior, taking a higher percentage of fish in their diet, which enhances chick growth and survival. This is believed to be the first documentation of bounded hybrid superiority that delineates the mechanisms that underlie hybrid superiority.

  18. Molecular cytogenetics of pituitary adenomas, assessed by FISH technique.

    Science.gov (United States)

    Kontogeorgos, George

    2004-01-01

    Fluorescent in situ hybridization (FISH) represents a moden molecular pathology technique, alternative to conventional cytogenetics (karyotyping). In addition to metaphase spreads, it can be applied directly to interphase nuclei. The latter makes the FISH technique powerful for pathologists for it integrates molecular genetics and classic cytogenetics and brings them together to a single framework for morphologic evaluation. Interphase FISH can be applied to imprints from fresh tissue or to paraffin sections after proteinase K digestion. Centromeric, telomeric and locus DNA-sequence specific probes can be used to identify aneuploidy or gene mutations. Several protocols combine molecular cytogenetics with classic karyotyping. Other sophisticated, FISH-based protocols have been introduced. Among them, comparative genomic hybridization is very important for it can detect non-balanced chromosomal aberrations of uncultured tumor cells and provide overall genomic information in a single experiment. This review presents the principles and applications of FISH technique for the investigation of the cytogenetic background of pituitary adenomas.

  19. Fishing amplifies forage fish population collapses.

    Science.gov (United States)

    Essington, Timothy E; Moriarty, Pamela E; Froehlich, Halley E; Hodgson, Emma E; Koehn, Laura E; Oken, Kiva L; Siple, Margaret C; Stawitz, Christine C

    2015-05-26

    Forage fish support the largest fisheries in the world but also play key roles in marine food webs by transferring energy from plankton to upper trophic-level predators, such as large fish, seabirds, and marine mammals. Fishing can, thereby, have far reaching consequences on marine food webs unless safeguards are in place to avoid depleting forage fish to dangerously low levels, where dependent predators are most vulnerable. However, disentangling the contributions of fishing vs. natural processes on population dynamics has been difficult because of the sensitivity of these stocks to environmental conditions. Here, we overcome this difficulty by collating population time series for forage fish populations that account for nearly two-thirds of global catch of forage fish to identify the fingerprint of fisheries on their population dynamics. Forage fish population collapses shared a set of common and unique characteristics: high fishing pressure for several years before collapse, a sharp drop in natural population productivity, and a lagged response to reduce fishing pressure. Lagged response to natural productivity declines can sharply amplify the magnitude of naturally occurring population fluctuations. Finally, we show that the magnitude and frequency of collapses are greater than expected from natural productivity characteristics and therefore, likely attributed to fishing. The durations of collapses, however, were not different from those expected based on natural productivity shifts. A risk-based management scheme that reduces fishing when populations become scarce would protect forage fish and their predators from collapse with little effect on long-term average catches.

  20. Sport Fishing Regulations

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — The regulations for sport fishing on St. Vincent National Wildlife Refuge are outlined in this document. Fishing is only permitted from sunrise to sunset, and only...

  1. Fish Springs pond snail

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — Communication scenario between the branch of Listing and Recovery, Fish and Wildlife Enhancement, and Fish Springs National Wildlife Refuge (NWR), in regards to the...

  2. Fish tapeworm infection

    Science.gov (United States)

    Fish tapeworm infection is an intestinal infection with the tapeworm parasite found in fish. ... The fish tapeworm ( Diphyllobothrium latum ) is the largest parasite that infects humans. Humans become infected when they eat raw ...

  3. Got a Sick Fish?

    Science.gov (United States)

    ... Welfare Veterinary Careers Public Health Got a sick fish? Fish with disease can show a variety of signs. If you notice your pet fish having any unusual disease signs, contact your veterinarian ...

  4. On the influence of the mixture of denaturants on protein structure stability: A molecular dynamics study

    Science.gov (United States)

    Shao, Qiang; Wang, Jinan; Zhu, Weiliang

    2014-09-01

    Mixtures of osmolytes and/or inorganic salts are present in the cell. Therefore, the understanding of the interplay of mixed osmolyte molecules and inorganic salts and their combined effects on protein structure is of fundamental importance. A novel test is presented to investigate the combined effects of urea and a chaotropic inorganic salt, potassium iodide (KI), on protein structure by using molecular dynamics simulation. It is found that the coexistence of KI and urea does not affect their respective distribution in solution. The solvation of KI salt in urea solution makes the electrostatic interactions of urea more favorable, promoting the hydrogen bonding between urea (and water) to protein backbone. The interactions from K+ and hydrogen bonding from urea and water to protein backbone work as the driving force for protein denaturation. The collaborative behavior of urea and KI salt thus enhances the denaturing ability of urea and KI mixed solution.

  5. Protective role of salt in catalysis and maintaining structure of halophilic proteins against denaturation

    Directory of Open Access Journals (Sweden)

    Rajeshwari eSinha

    2014-04-01

    Full Text Available Search for new enzymes of industrial relevance, bestowed with novel properties continues to be a desirable pursuit in enzyme research. Halophilism is the unusual existence of life in saline/ hypersaline habitats and haloenzymes, are the proteins from such origin, naturally endowed with unique structural features which enable them to sustain functionality under high salt. Driven by industrial requirements, halophilic enzymes have been explored for their stability and catalytic abilities under harsh operational conditions. These have been documented to withstand high temperature, pH, organic solvents, and chaotropic agents. However, this stability is modulated by salt. Understanding the basis of salt mediated protection amidst a denaturing milieu will add significantly to the existing knowledge about structure function relationships in halophilic proteins. Exploring their protein architecture may provide template for rationale design of stable enzymes. The article encompasses the current level of understanding about haloadaptations in halophiles and structural basis of their stability against classical denaturants.

  6. pH-sensitive polymer-assisted refolding of urea-denatured fibroblast growth factor

    Institute of Scientific and Technical Information of China (English)

    Zhi Feng Huang; Shan Shan Wang; Chun Yan Ni; Shu Lin Yang; Xiao Kun Li; Susanna S.J.Leong

    2009-01-01

    A pH-responsive polymer Eudragit S-100 has been found to assist in correct folding of FGF-2(fibroblast growth factor-2)denatured with 8 mol/L urea and 10 mmol/L dithiothreitol at pH 7.2.The refolding of FGF-2 was performed by directly diluting denatured FGF-2 into a refolding buffer containing Eudragit S-100.The ability of Eudragit S-100 to enhance protein refolding level was investigated using MTT method,fluorescence emission spectroscopy and reversc phase HPLC.On the other hand,the result shows the ability of Eudragit S-100 to enhance the refolding level of protein is due to the interaction between Eudragit S-100 and positively charged FGF-2.

  7. Theoretical aspects of pressure and solute denaturation of proteins: A Kirkwood-buff-theory approach

    Science.gov (United States)

    Ben-Naim, Arieh

    2012-12-01

    A new approach to the problem of pressure-denaturation (PD) and solute-denaturation (SD) of proteins is presented. The problem is formulated in terms of Le Chatelier principle, and a solution is sought in terms of the Kirkwood-Buff theory of solutions. It is found that both problems have one factor in common; the excluded volumes of the folded and the unfolded forms with respect to the solvent molecules. It is shown that solvent-induced effects operating on hydrophilic groups along the protein are probably the main reason for PD. On the other hand, the SD depends on the preferential solvation of the folded and the unfolded forms with respect to solvent and co-solvent molecules.

  8. ZnO nanoparticles assist the refolding of denatured green fluorescent protein.

    Science.gov (United States)

    Pandurangan, Muthuraman; Zamany, Ahmad Jawid; Kim, Doo Hwan

    2016-04-01

    Proteins are essential for cellular and biological processes. Proteins are synthesized and fold into the native structure to become active. The inability of a protein molecule to remain in its native conformation is called as protein misfolding, and this is due to several environmental factors. Protein misfolding and aggregation handle several human diseases. Protein misfolding is believed to be one of the causes of several disorders such as cancer, degenerative diseases, and metabolic pathologies. The zinc oxide (ZnO) nanoparticle was significantly promoted refolding of thermally denatured green fluorescent protein (GFP). In the present study, ZnO nanoparticles interaction with GFP was investigated by ultraviolet-visible spectrophotometer, fluorescence spectrophotometer, and dynamic light scattering. Results suggest that the ZnO nanoparticles significantly assist the refolding of denatured GFP. Copyright © 2015 John Wiley & Sons, Ltd.

  9. Small angle neutron scattering studies on protein denaturation induced by different methods

    Indian Academy of Sciences (India)

    S Chodankar; V K Aswal; J Kohlbrecher; R Vavrin; A G Wagh

    2008-11-01

    Small angle neutron scattering (SANS) has been used to study conformational changes in protein bovine serum albumin (BSA) as induced by varying temperature and in the presence of protein denaturating agents urea and surfactant. BSA has pro-late ellipsoidal shape and is found to be stable up to 60°C above which it denaturates and subsequently leads to aggregation. The protein solution exhibits a fractal structure at temperatures above 64°C, with fractal dimension increasing with temperature. BSA protein is found to unfold in the presence of urea at concentrations greater than 4 M and acquires a random coil Gaussian chain conformation. The conformation of the unfolded protein in the presence of surfactant has been determined directly using contrast variation SANS measurements by contrast matching surfactant molecules. The protein acquires a random coil Gaussian conformation on unfolding with its radius of gyration increasing with increase in surfactant concentration

  10. Teaching what one does not know: strangeness and denaturation in (autobiographical narrations

    Directory of Open Access Journals (Sweden)

    Jorge Luiz da Cunha

    2014-01-01

    Full Text Available The thematic focus in this text are the estrangement/denaturation processes in (autobiographical narrations. The aim of this study was to reflect on the possibility to promote estrangement/denatura - tion in (autobiographical writings made by teenagers in the space/ time of the classroom environment. The methodological proposal consisted on developing (autobiographical writings by students from sociology classes in High School. A total of 138 teenagers from a public school, attending the first school trimester in the year 2013, have participated in the study. The concepts of estrangement/de - naturation are located in the anthropology field and, the work with (autobiographical narrations is located in the socio-clinic perspec - tives and of biographization processes. The results indicate that (autobiographical narrations provide estrangements/denaturation and go towards teaching what one does not know. We can, then, conclude that this possibility, as an educational act, may generate knowledge suspension to self-inventiveness.

  11. PROTECTIVE EFFECT OF PROLACTIN INDUCED PROTEIN ON ZINC α2-GLYCOPROTEIN AGAINST VARIOUS DENATURANTS

    Directory of Open Access Journals (Sweden)

    Md. Imtaiyaz Hassan

    2012-12-01

    Full Text Available Zinc α2-glycoprotein (ZAG and Prolactin induced protein (PIP are considered as important elements for fertility and biomarker for prostate and breast carcinomas. The stabilities of ZAG alone and its naturally occurring complex with PIP were compared. A significant difference in CD signal was recorded for native ZAG and ZAG-PIP complex against pH-, GdnHCl- and temperature-induced denaturation. These finding suggests that PIP plays a protective role for ZAG against several denaturants. PIP contributes to the hydrophobic as well as electrostatic interactions on ZAG for the complex formation. Moreover, the observed changes in far-UV spectra between ZAG and ZAG-PIP complex in the presence of PEG support the hydrophobic nature of the forces governing the formation of complex. This pH dependent study provides evidence that formation of the complex is a natural event required for physiological function.

  12. Surface characterization of proteins using multi-fractal property of heat-denatured aggregates

    Science.gov (United States)

    Lahiri, Tapobrata; Mishra, Hrishikesh; Sarkar, Subrata; Misra, Krishna

    2008-01-01

    Multi-fractal property of heat-denatured protein aggregates (HDPA) is characteristic of its individual form. The visual similarity between digitally generated microscopic images of HDPA with that of surface-image of its individual X-ray structures in protein databank (PDB) displayed using Visual Molecular Dynamics (VMD) viewer is the basis of the study. We deigned experiments to view the fractal nature of proteins at different aggregate scales. Intensity based multi-fractal dimensions (ILMFD) extracted from various planes of digital microscopic images of protein aggregates were used to characterize HDPA into different classes. Moreover, the ILMFD parameters extracted from aggregates show similar classification pattern to digital images of protein surface displayed by VMD viewer using PDB entry. We discuss the use of irregular patterns of heat-denatured aggregate proteins to understand various surface properties in native proteins. PMID:18795110

  13. Acid Denaturation Inducing Self-Assembly of Curcumin-Loaded Hemoglobin Nanoparticles

    Directory of Open Access Journals (Sweden)

    Kaikai Wang

    2015-12-01

    Full Text Available Hemoglobin is a promising drug carrier but lacks extensive investigation. The chemical conjugation of hemoglobin and drugs is costly and complex, so we have developed curcumin-loaded hemoglobin nanoparticles (CCM-Hb-NPs via self-assembly for the first time. Using the acid-denaturing method, we avoid introducing denaturants and organic solvents. The nanoparticles are stable with uniform size. We have conducted a series of experiments to examine the interaction of hemoglobin and CCM, including hydrophobic characterization, SDS-PAGE. These experiments substantiate that this self-assembly process is mainly driven by hydrophobic forces. Our nanoparticles achieve much higher cell uptake efficiency and cytotoxicity than free CCM solution in vitro. The uptake inhibition experiments also demonstrate that our nanoparticles were incorporated via the classic clathrin-mediated endocytosis pathway. These results indicate that hemoglobin nanoparticles formed by self-assembly are a promising drug delivery system for cancer therapy.

  14. Comparison between conformational change and inactivation rates of aminoacylase during denaturation in urea solutions

    Institute of Scientific and Technical Information of China (English)

    王洪睿; 王希成; 张彤; 周海梦

    1995-01-01

    The kinetic method of the substrate reaction in the presence of mactivator previously described by Tsou has been applied to the determination of inactivation rates of aminoacylase during denaturation in urea solutions. The protective effect of substrate on the inactivation of aminoacylase by urea has been investigated. Simultaneously, the comparison between conformational change and inactivation rates of enzyme in the urea solutions of different concentrations has been studied. Results obtained show that the inactivation rate constants of the enzyme are larger than the rate constants of conformational changes. The present results show that the active site of metal enzyme-aminoacylase is also located in a limited and flexible region of the molecule that is more sensitive to denaturants than the enzyme as a whole.

  15. Formation of Native and Non-native Interactions in Ensembles of Denatured ACBP Molecules from Paramagnetic Relaxation Enhancement Studies

    DEFF Research Database (Denmark)

    Kristjansdottir, S.; Lindorff-Larsen, Kresten; Fieber, W.;

    2005-01-01

    in the denatured states with those in the transition state for folding we also provided new insights into the mechanism of formation of the native state of this protein. Keywords: protein folding; denatured state; NMR; molecular dynamics; structural studies Abbreviations: ACBP, acyl coenzyme A binding protein; Gu...... of the residual structure in the denatured state of ACBP under these different conditions has enabled us to infer that regions in the N and C-terminal parts of the protein sequence have a high tendency to interact in the unfolded state under physiological conditions. By comparing the structural features...

  16. Micro-CT Imaging of Denatured Chitin by Silver to Explore Honey Bee and Insect Pathologies

    Science.gov (United States)

    Butzloff, Peter R.

    2011-01-01

    Background Chitin and cuticle coatings are important to the environmental and immune defense of honey bees and insect pollinators. Pesticides or environmental effects may target the biochemistry of insect chitin and cuticle coating. Denaturing of chitin involves a combination of deacetylation, intercalation, oxidation, Schweiger-peeling, and the formation of amine hydrochloride salt. The term “denatured chitin” calls attention to structural and property changes to the internal membranes and external carapace of organisms so that some properties affecting biological activities are diminished. Methodology/Principal Findings A case study was performed on honey bees using silver staining and microscopic computer-tomographic x-ray radiography (micro-CT). Silver nitrate formed counter-ion complexes with labile ammonium cations and reacted with amine hydrochloride. Silver was concentrated in the peritrophic membrane, on the abdomen, in the glossa, at intersegmental joints (tarsi), at wing attachments, and in tracheal air sacs. Imaged mono-esters and fatty acids from cuticle coating on external surfaces were apparently reduced by an alcohol pretreatment. Conclusions/Significance The technique provides 3-dimensional and sectional images of individual honey bees consistent with the chemistries of silver reaction and complex formation with denatured chitin. Environmental exposures and influences such as gaseous nitric oxide intercalant, trace oxidants such as ozone gas, oligosachharide salt conversion, exposure to acid rain, and chemical or biochemical denaturing by pesticides may be studied using this technique. Peritrophic membranes, which protect against food abrasion, microorganisms, and permit efficient digestion, were imaged. Apparent surface damage to the corneal lenses of compound eyes by dilute acid exposure consistent with chitin amine hydrochloride formation was imaged. The technique can contribute to existing insect pathology research, and may provide an

  17. Thermal denaturation of beta-galactosidase and of two site-specific mutants.

    Science.gov (United States)

    Edwards, R A; Jacobson, A L; Huber, R E

    1990-12-11

    The thermal denaturation of wild-type beta-galactosidase and two beta-galactosidases with substitutions at the active site was studied by kinetics, differential scanning calorimetry, electrophoresis, molecular exclusion chromatography, and circular dichroism. From the results, a model is developed for thermal denaturation of beta-galactosidase which includes the reversible dissociation of ligands, reversible formation of an inactive tetramer, irreversible dissociation of the inactive tetramer to inactive monomers, and subsequent aggregation of inactive monomers to dimers and larger aggregates. Under some conditions, partial reversibility of the activity loss could be demonstrated, and several intermediates in the thermal denaturation process were trapped by quenching and observed by electrophoresis and molecular exclusion chromatography. The ligands Mg2+ and phenylethyl thio-beta-D-galactoside increase the stability of beta-galactosidase to heat denaturation by shifting the ligand binding equilibrium according to Le Chatelier's principle, thus decreasing the concentration of the ligand-free tetramer which can proceed to subsequent steps. Circular dichroism results indicated that beta-galactosidase is dominated by beta-sheet with lower amounts of alpha-helix. Large changes in secondary structure begin to occur only after activity has been lost. Single amino acid changes at the active site can have significant effects on thermal stability of beta-galactosidases. Some of the effects result from increased thermal stability of the ligand-free enzyme itself. Other effects result from changes in ligand binding, but the magnitude of the resulting changes in stability is not related to the strength of ligand binding in a simple fashion.

  18. Detection of Rifampin Resistance in Mycobacterium tuberculosis by Double Gradient-Denaturing Gradient Gel Electrophoresis

    Science.gov (United States)

    Scarpellini, Paolo; Braglia, Sergio; Carrera, Paola; Cedri, Maura; Cichero, Paola; Colombo, Alessia; Crucianelli, Rosella; Gori, Andrea; Ferrari, Maurizio; Lazzarin, Adriano

    1999-01-01

    We applied double gradient-denaturing gradient gel electrophoresis (DG-DGGE) for the rapid detection of rifampin (RMP) resistance from rpoB PCR products of Mycobacterium tuberculosis isolates and clinical samples. The results of this method were fully concordant with those of DNA sequencing and susceptibility testing analyses. DG-DGGE is a valid alternative to the other methods of detecting mutations for predicting RMP resistance. PMID:10508043

  19. A kinetic study of jack-bean urease denaturation by a new dithiocarbamate bismuth compound

    Science.gov (United States)

    Menezes, D. C.; Borges, E.; Torres, M. F.; Braga, J. P.

    2012-10-01

    A kinetic study concerning enzymatic inhibitory effect of a new bismuth dithiocarbamate complex on jack-bean urease is reported. A neural network approach is used to solve the ill-posed inverse problem arising from numerical treatment of the subject. A reaction mechanism for the urease denaturation process is proposed and the rate constants, relaxation time constants, equilibrium constants, activation Gibbs free energies for each reaction step and Gibbs free energies for the transition species are determined.

  20. Combining an Optical Resonance Biosensor with Enzyme Activity Kinetics to Understand Protein Adsorption and Denaturation

    OpenAIRE

    Wilson, Kerry A.; Finch, Craig A.; Anderson, Phillip; Vollmer, Frank; Hickman, James J.

    2014-01-01

    Understanding protein adsorption and resultant conformation changes on modified and unmodified silicon dioxide surfaces is a subject of keen interest in biosensors, microfluidic systems and for medical diagnostics. However, it has been proven difficult to investigate the kinetics of the adsorption process on these surfaces as well as understand the topic of the denaturation of proteins and its effect on enzyme activity. A highly sensitive optical whispering gallery mode (WGM) resonator was us...

  1. Multifocal peritoneal splenosis in Tc-99m-labeled heat-denatured red blood cell scintigraphy

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Min Ki; Hwang, Kyung Hoon; Choe, Won Sick [Gachon University Gil Medical Center, Incheon (Korea, Republic of)

    2006-06-15

    A 44-year-old man with a past medical history of splenectomy came to hospital because of epigastric pain abdominopelvic computed tomography(CT) showed a soft tissue mass and multifocal variable-sized nodules as well as finding suggestive of cholecystitis. Subsequently, he underwent Tc-99m-labeled heat- denatured red blood cell(RBC) scintigraphy to evaluate the mass and nodules. The scintigraphy confirmed multifocal peritoneal splenosis in the abdominopelvic cavity.

  2. Surface characterization of proteins using multi-fractal property of heat-denatured aggregates

    OpenAIRE

    Lahiri, Tapobrata; Mishra, Hrishikesh; Sarkar, Subrata; Misra, Krishna

    2008-01-01

    Multi-fractal property of heat-denatured protein aggregates (HDPA) is characteristic of its individual form. The visual similarity between digitally generated microscopic images of HDPA with that of surface-image of its individual X-ray structures in protein databank (PDB) displayed using Visual Molecular Dynamics (VMD) viewer is the basis of the study. We deigned experiments to view the fractal nature of proteins at different aggregate scales. Intensity based multi-fractal dimensions (ILMFD)...

  3. Thermal stability of chemically denatured green fluorescent protein (GFP) A preliminary study

    Energy Technology Data Exchange (ETDEWEB)

    Nagy, Attila; Malnasi-Csizmadia, Andras; Somogyi, Bela; Lorinczy, Denes

    2004-02-09

    Green fluorescent protein (GFP) is a light emitter in the bioluminescence reaction of the jellyfish Aequorea victoria. The protein consist of 238 amino acids and produces green fluorescent light ({lambda}{sub max}=508 nm), when irradiated with near ultraviolet light. The fluorescence is due to the presence of chromophore consisting of an imidazolone ring, formed by a post-translational modification of the tripeptide -Ser{sup 65}-Tyr{sup 66}-Gly{sup 67}-, which buried into {beta}-barrel. GFP is extremely compact and heat stable molecule. In this work, we present data for the effect of chemical denaturing agent on the thermal stability of GFP. When denaturing agent is applied, global thermal stability and the melting point of the molecule is decreases, that can be monitored with differential scanning calorimetry. The results indicate, that in 1-6 M range of GuHCl the melting temperature is decreasing continuously from 83 to 38 deg. C. Interesting finding, that the calculated calorimetric enthalpy decreases with GuHCl concentration up to 3 M (5.6-0.2 kJ mol{sup -1}), but at 4 M it jumps to 8.4 and at greater concentration it is falling down to 1.1 kJ mol{sup -1}. First phenomena, i.e. the decrease of melting point with increasing GuHCl concentration can be easily explained by the effect of the extended chemical denaturation, when less and less amount of heat required to diminish the remaining hydrogen bonds in {beta}-barrel. The surprising increase of calorimetric enthalpy at 4 M concentration of GuHCl could be the consequence of a dimerization or a formation of stable complex between GFP and denaturing agent as well as a precipitation at an extreme GuHCl concentration. We are planning further experiments to elucidate fluorescent consequence of these processes.

  4. Micro-CT imaging of denatured chitin by silver to explore honey bee and insect pathologies.

    Directory of Open Access Journals (Sweden)

    Peter R Butzloff

    Full Text Available BACKGROUND: Chitin and cuticle coatings are important to the environmental and immune defense of honey bees and insect pollinators. Pesticides or environmental effects may target the biochemistry of insect chitin and cuticle coating. Denaturing of chitin involves a combination of deacetylation, intercalation, oxidation, Schweiger-peeling, and the formation of amine hydrochloride salt. The term "denatured chitin" calls attention to structural and property changes to the internal membranes and external carapace of organisms so that some properties affecting biological activities are diminished. METHODOLOGY/PRINCIPAL FINDINGS: A case study was performed on honey bees using silver staining and microscopic computer-tomographic x-ray radiography (micro-CT. Silver nitrate formed counter-ion complexes with labile ammonium cations and reacted with amine hydrochloride. Silver was concentrated in the peritrophic membrane, on the abdomen, in the glossa, at intersegmental joints (tarsi, at wing attachments, and in tracheal air sacs. Imaged mono-esters and fatty acids from cuticle coating on external surfaces were apparently reduced by an alcohol pretreatment. CONCLUSIONS/SIGNIFICANCE: The technique provides 3-dimensional and sectional images of individual honey bees consistent with the chemistries of silver reaction and complex formation with denatured chitin. Environmental exposures and influences such as gaseous nitric oxide intercalant, trace oxidants such as ozone gas, oligosachharide salt conversion, exposure to acid rain, and chemical or biochemical denaturing by pesticides may be studied using this technique. Peritrophic membranes, which protect against food abrasion, microorganisms, and permit efficient digestion, were imaged. Apparent surface damage to the corneal lenses of compound eyes by dilute acid exposure consistent with chitin amine hydrochloride formation was imaged. The technique can contribute to existing insect pathology research, and may

  5. Supplement Analysis for the Wildlife Mitigation Program EIS (DOE/EIS-0246/SA-27) - Abbot Creek Fish Barrier Project (Hungry Horse Mitigation / Habitat Improvements)

    Energy Technology Data Exchange (ETDEWEB)

    Yarde, Richard [Bonneville Power Administration (BPA), Portland, OR (United States)

    2002-06-28

    BPA proposes to fund a fishery enhancement project where a fish passage barrier will be installed in Abbot Creek to remove introduced rainbow trout and prevent hybridization with westslope cutthroat trout. Montana Fish, Wildlife & Parks (MFWP) will operate a fish trap downstream of the barrier for 6-10 consecutive years to manually remove the rainbow trout and hybrid spawners from the population. Removal of rainbow trout and hybrids from the stream will eradicate the existing hybrid population spawning in Abbot Creek and ultimately reduce the threat of hybridization in the Flathead River system. Pending completion of a successful disease screening and authorization from MFWP Fish Health Committee, live fish captured in the fish trap will be transported to a nearby close-basin lake for use in MFWP’s Urban Fishing Program.

  6. Comparison of membrane electroporation and protein denature in response to pulsed electric field with different durations.

    Science.gov (United States)

    Huang, Feiran; Fang, Zhihui; Mast, Jason; Chen, Wei

    2013-05-01

    In this paper, we compared the minimum potential differences in the electroporation of membrane lipid bilayers and the denaturation of membrane proteins in response to an intensive pulsed electric field with various pulse durations. Single skeletal muscle fibers were exposed to a pulsed external electric field. The field-induced changes in the membrane integrity (leakage current) and the Na channel currents were monitored to identify the minimum electric field needed to damage the membrane lipid bilayer and the membrane proteins, respectively. We found that in response to a relatively long pulsed electric shock (longer than the membrane intrinsic time constant), a lower membrane potential was needed to electroporate the cell membrane than for denaturing the membrane proteins, while for a short pulse a higher membrane potential was needed. In other words, phospholipid bilayers are more sensitive to the electric field than the membrane proteins for a long pulsed shock, while for a short pulse the proteins become more vulnerable. We can predict that for a short or ultrashort pulsed electric shock, the minimum membrane potential required to start to denature the protein functions in the cell plasma membrane is lower than that which starts to reduce the membrane integrity.

  7. DENATURATION OF NATIVE AND DEGLYCOSYLATED α-GALACTOSIDASES FROM Penicillium canescens BY GUANIDINE HYDROCHLORIDE

    Directory of Open Access Journals (Sweden)

    Borzova N. V.

    2015-08-01

    Full Text Available The aim of this work was the study of native and galactosidases from Penicillium canescens under denaturing conditions caused by guanidine hydrochloride. Calculation of kinetics and constants of enzymes inactivation was carried out on using experimental kinetic curves of enzyme denaturation. We observed significant differences in the kinetics of inactivation of native and deglycosylated α-galactosidases from P. canescens caused by guanidine hydrochloride. Native enzyme was stable within the selected range of guanidine hydrochloride concentrations (from 0.1 to 3.0 M, retaining no less than 50% of the initial enzyme activity for 3 days. Deglycosylated enzyme preparations were less stable and they lost their activity within 5–30 minutes, when they were treated with guanidine hydrochloride in concentrations above 1 M. Dissociation rate constant of native and deglycosylated forms of the enzyme differed by 10 to 100 folds. It was shown that subunit interactions play a major role in the process of inactivation of the enzyme, and the carbohydrate component is essential for stabilizing of subunit bonds and maintaining conformational stability of the enzyme under denaturing conditions of chemical agents.

  8. Thermal and mechanical denaturation properties of a DNA model with three sites per nucleotide

    CERN Document Server

    Florescu, Ana-Maria; 10.1063/1.3626870

    2011-01-01

    In this paper, we show that the coarse grain model for DNA, which has been proposed recently by Knotts, Rathore, Schwartz and de Pablo (J. Chem. Phys. 126, 084901 (2007)), can be adapted to describe the thermal and mechanical denaturation of long DNA sequences by adjusting slightly the base pairing contribution. The adjusted model leads to (i) critical temperatures for long homogeneous sequences that are in good agreement with both experimental ones and those obtained from statistical models, (ii) a realistic step-like denaturation behaviour for long inhomogeneous sequences, and (iii) critical forces at ambient temperature of the order of 10 pN, close to measured values. The adjusted model furthermore supports the conclusion that the thermal denaturation of long homogeneous sequences corresponds to a first-order phase transition and yields a critical exponent for the critical force equal to sigma=0.70. This model is both geometrically and energetically realistic, in the sense that the helical structure and th...

  9. Single domain antibodies are specially suited for quantitative determination of gliadins under denaturing conditions.

    Science.gov (United States)

    Doña, Vanina; Urrutia, Mariela; Bayardo, Mariela; Alzogaray, Vanina; Goldbaum, Fernando Alberto; Chirdo, Fernando G

    2010-01-27

    Food intended for celiac patients' consumption must be analyzed for the presence of toxic prolamins using high detectability tests. Though 60% ethanol is the most commonly used solvent for prolamins extraction, 2-mercaptoethanol (2-ME) and guanidinium chloride (GuHCl) can be added to increase protein recovery. However, ethanol and denaturing agents interfere with antigen recognition when conventional antibodies are used. In the present work, a new method for gliadins quantification is shown. The method is based on the selection of llama single domain antibody fragments able to operate under denaturing conditions. Six out of 28 VHH-phages obtained retained their binding capacity in 15% ethanol. Selected clones presented a long CDR3 region containing two additional cysteines that could be responsible for the higher stability. One of the clones (named VHH26) was fully operative in the presence of 15% ethanol, 0.5% 2-ME, and 0.5 M GuHCl. Capture ELISA using VHH26 was able to detect gliadins in samples shown as negatives by conventional ELISA. Therefore, this new strategy appears as an excellent platform for quantitative determination of proteins or any other immunogenic compound, in the presence of denaturing agents, when specific recognition units with high stability are required.

  10. Remarkable activation of enzymes in nonaqueous media by denaturing organic cosolvents

    Energy Technology Data Exchange (ETDEWEB)

    Almarsson, O.; Klibanov, A.M. [Massachusetts Inst. of Tech., Cambridge, MA (United States). Dept. of Chemistry

    1996-01-05

    The rates of transesterification reactions catalyzed by the protease subtilisin Carlsberg suspended in various anhydrous solvents at 30 C can be increased more than 100-fold by the addition of denaturing organic cosolvents (dimethyl sulfoxide or formamide); in water, the same cosolvents exert no enzyme activation. At 4 C, the activation effect on the lyophilized protease is even higher, reaching 1,000-fold. Marked enhancement of enzymatic activity in anhydrous solvents by formamide is also observed for two other enzymes, {alpha}-chymotrypsin and Rhizomucor miehei lipase, and is manifested in two transesterification reactions. In addition to lyophilized subtilisin, crosslinked crystals of subtilisin are also amenable to the dramatic activation by the denaturing cosolvents. In contrast, subtilisin solubilized in anhydrous media by covalent modification with poly(ethylene glycol) exhibits only modest activation. These observations are rationalized in terms of a mechanistic hypothesis based on an enhanced protein flexibility in anhydrous milieu brought about by the denaturing organic cosolvents. The latter exert their lubricating effect largely at the interfaces between enzyme molecules in a solid preparation, thus easing the flexibility constraints imposed by protein-protein contacts.

  11. Effects of high pressure freezing (HPF) on denaturation of natural actomyosin extracted from prawn (Metapenaeus ensis).

    Science.gov (United States)

    Cheng, Lina; Sun, Da-Wen; Zhu, Zhiwei; Zhang, Zhihang

    2017-08-15

    Effects of protein denaturation caused by high pressure freezing, involving Pressure-Factors (pressure, time) and Freezing-Factors (temperature, phase transition, recrystallization, ice crystal types), are complicated. In the current study, the conformation and functional changes of natural actomyosin (NAM) under pressure assisted freezing (PAF, (100,150,300,400,500MPa)P-20°C/25min), pressure shift freezing (PSF, (200MPa)P-20°C/25min), and immersion freezing ((0.1MPa)P-20°C/5min) after pressure was released to 0.1MPa, as compared to normal immersion freezing process (IF, (0.1MPa)P-20°C/30min). Results indicated that PSF ((200MPa)P-20°C/30min) could reduce the denaturation of frozen NAM and a pressure of 300MPa was the critical point to induce such a denaturation. During the periods of B→D in PSF or B→C→D in PAF, the generation and growth of ice crystals played an important role on changing the secondary and tertiary structure of the treated NAM. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Fourier transform infrared spectroscopy provides an evidence of papain denaturation and aggregation during cold storage.

    Science.gov (United States)

    Rašković, Brankica; Popović, Milica; Ostojić, Sanja; Anđelković, Boban; Tešević, Vele; Polović, Natalija

    2015-01-01

    Papain is a cysteine protease with wide substrate specificity and many applications. Despite its widespread applications, cold stability of papain has never been studied. Here, we used differential spectroscopy to monitor thermal denaturation process. Papain was the most stabile from 45 °C to 60 °C with ΔG°321 of 13.9±0.3 kJ/mol and Tm value of 84±1 °C. After cold storage, papain lost parts of its native secondary structures elements which gave an increase of 40% of intermolecular β-sheet content (band maximum detected at frequency of 1621 cm(-1) in Fourier transform infrared (FT-IR) spectrum) indicating the presence of secondary structures necessary for aggregation. The presence of protein aggregates after cold storage was also proven by analytical size exclusion chromatography. After six freeze-thaw cycles around 75% of starting enzyme activity of papain was lost due to cold denaturation and aggregation of unfolded protein. Autoproteolysis of papain did not cause significant loss of the protein activity. Upon the cold storage, papain underwent structural rearrangements and aggregation that correspond to other cold denatured proteins, rather than autoproteolysis which could have the commercial importance for the growing polypeptide based industry.

  13. Stabilization of Human Serum Albumin against Urea Denaturation by Diazepam and Ketoprofen.

    Science.gov (United States)

    Manoharan, Pralad; Wong, Yin H; Tayyab, Saad

    2015-01-01

    Stabilizing effect of diazepam and ketoprofen, Sudlow's site II markers on human serum albumin (HSA) against urea denaturation was studied using fluorescence spectroscopy. The two-step, three-state urea transition of HSA was transformed into a single-step, two-state transition with the abolishment of the intermediate state along with a shift of the transition curve towards higher urea concentrations in the presence of diazepam or ketoprofen. Interestingly, a greater shift in the transition curve of HSA was observed in the presence of ketoprofen compared to diazepam. A comparison of the intrinsic fluorescence and three-dimensional fluorescence spectra of HSA and partially-denatured HSAs, obtained in the absence and the presence of diazepam or ketoprofen suggested significant retention of native-like conformation in the partially-denatured states of HSA in the presence of Sudlow's site II markers. Taken together, all these results suggested stabilization of HSA in the presence of diazepam or ketoprofen, being greater in the presence of ketoprofen.

  14. Evaluation of gasoline-denatured ethanol as a carbon source for denitrification.

    Science.gov (United States)

    Kazasi, Anna; Boardman, Gregory D; Bott, Charles B

    2013-06-01

    In this study concerning denitrification, the performance of three carbon sources, methanol (MeOH), ethanol (EtOH) and gasoline-denatured ethanol (dEtOH), was compared and evaluated on the basis of treatment efficiency, inhibition potential and cost. The gasoline denaturant considered here contained mostly aliphatic compounds and little of the components that typically boost the octane rating, such as benzene, toluene, ethylbenzene and xylenes. Results were obtained using three lab-scale SBRs operated at SRT of 12.0 +/- 0.9 days. After biomass was acclimated, denitrification rates with dEtOH were similar to those of EtOH (201 +/- 50 and 197 +/- 28 NO3-N/g MLVSS x d, respectively), and higher than those of MeOH (165 +/- 49 mg NO3-N/g MLVSS x d). The denaturant did not affect biomass production, nitrification or denitrification. Effluent soluble COD concentrations were always less than the analytical detection limit. Although the cost of dEtOH ($2.00/kg nitrate removed) was somewhat higher than that of methanol ($1.63/kg nitrate removed), the use of dEtOH is very promising and utilities will have to decide if it is worth paying a little extra to take advantage of its benefits.

  15. AFM visualization at a single-molecule level of denaturated states of proteins on graphite.

    Science.gov (United States)

    Barinov, Nikolay A; Prokhorov, Valery V; Dubrovin, Evgeniy V; Klinov, Dmitry V

    2016-10-01

    Different graphitic materials are either already used or believed to be advantageous in biomedical and biotechnological applications, e.g., as biomaterials or substrates for sensors. Most of these applications or associated important issues, such as biocompatibility, address the problem of adsorption of protein molecules and, in particular the conformational state of the adsorbed protein molecule on graphite. High-resolution AFM demonstrates highly oriented pyrolytic graphite (HOPG) induced denaturation of four proteins of blood plasma, such as ferritin, fibrinogen, human serum albumin (HSA) and immunoglobulin G (IgG), at a single molecule level. Protein denaturation is accompanied by the decrease of the heights of protein globules and spreading of the denatured protein fraction on the surface. In contrast, the modification of HOPG with the amphiphilic oligoglycine-hydrocarbon derivative monolayer preserves the native-like conformation and provides even more mild conditions for the protein adsorption than typically used mica. Protein unfolding on HOPG may have universal character for "soft" globular proteins. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Denaturation Kinetics of Whey Protein Isolate Solutions and Fouling Mass Distribution in a Plate Heat Exchanger

    Directory of Open Access Journals (Sweden)

    Marwa Khaldi

    2015-01-01

    Full Text Available Few investigations have attempted to connect the mechanism of dairy fouling to the chemical reaction of denaturation (unfolding and aggregation occurring in the bulk. The objective of this study is to contribute to this aspect in order to propose innovative controls to limit fouling deposit formation. Experimental investigations have been carried out to observe the relationship between the deposit mass distribution generated in plate heat exchangers (PHE by a whey protein isolate (WPI mainly composed of β-lactoglobulin (β-Lg and the ratio between the unfolding and aggregation rate constants. Experiments using a PHE were carried out at a pilot scale to identify the deposit distribution of a model fouling solution with different calcium contents. In parallel, laboratory experiments were performed to determine the unfolding/aggregation rate constants. Data analysis showed that (i β-Lg denaturation is highly dependent on the calcium content, (ii for each fouling solution, irrespective of the imposed temperature profile, the deposit mass in each channel and the ratio between the unfolding and aggregation rate constants seem to be well correlated. This study demonstrates that both the knowledge of the thermal profile and the β-Lg denaturation rate constants are required in order to predict accurately the deposit distribution along the PHE.

  17. Fluorescent in situ hybridization as an adjunct to conventional cytogenetics.

    Science.gov (United States)

    Mark, H F

    1994-01-01

    Fluorescent in situ hybridization (FISH) is a molecular cytogenetic technique that exploits the availability of recombinant deoxyribonucleic acid (DNA) technology. In metaphase FISH, a specific nucleic acid sequence (probe) is bound to the homologous segment on a metaphase chromosome in a fixed preparation on a glass slide. The presence of a region-specific DNA sequence in a nondividing cell can be detected using interphase FISH. Interphase cytogenetics via FISH can be performed on fixed cells harvested during a routine culture, on tissue sections and on many cytologic specimens. Specific examples of clinical and research applications are discussed to illustrate the utility of FISH in the detection of constitutional and acquired chromosomal abnormalities.

  18. Fish mycobacteriosis (Tuberculosis)

    Science.gov (United States)

    Parisot, T.J.; Wood, J.W.

    1959-01-01

    The etiologic agent for the bacterial disease, "fish tuberculosis" (more correctly "mycobacteriosis"), was first observed in carp in 189& from a pond in France. Subsequently similar agents have been isolated from or observed in fish in fresh water, salt water, and brackish water, in fish in aquaria, hatcheries, and natural habitat~ (wild populations of fish). The disease has been recognized as an important infection among hatchery reared salmonid fishes on the West Coast of the United States, and in aquarium fishes such as the neon tetra, the Siamese fighting fish, and in salt water fish held in zoological displays.

  19. Molecular Hybridization of Iodinated 4S, 5S, and 18S + 28S RNA to Salamander Chromosomes

    National Research Council Canada - National Science Library

    Pedro E. León

    1976-01-01

    4S, 5S, and 18S + 28S RNA from the newt Taricha granulosa granulosa were iodinated in vitro with carrier-free 125I and hybridized to the denatured chromosomes of Taricha granulosa and Batrachoseps wrighti. Iodinated 18S...

  20. No Fishing Now,More Fish Later

    Institute of Scientific and Technical Information of China (English)

    2011-01-01

    Fishing ban for ecological purposes starts on the Pearl River Since April1,a two-month ban on fishing has been imposed on the Pearl River valley in south China.It is the first fishing ban in this area with the purpose of preserving biodiversity in China’s third longest

  1. Microbiological spoilage of fish and fish products

    DEFF Research Database (Denmark)

    Gram, Lone; Huss, Hans Henrik

    1996-01-01

    Spoilage of fresh and lightly preserved fish products is caused by microbial action. This paper reviews the current knowledge in terms of the microbiology of fish and fish products with particular emphasis on identification of specific spoilage bacteria and the qualitative and quantitative...

  2. 膀胱移行细胞癌各分期分级中荧光原位杂交检测敏感性比较%Evaluation on significance of fluorescence in situ hybridization (FISH) for detection of bladder cancer

    Institute of Scientific and Technical Information of China (English)

    徐秀红; 杜雨; 张道新; 田野

    2011-01-01

    Objective In order to look for a better non - invasive method for diagnosing bladder cancer, the fluorescence in situ hybridization ( FISH ) technique had been applied in diagnosis of bladder cancer. Methods FISH and cytological analysis were performed simultaneously on urine - exfoliated cells from 138 patients with hematuria. Chromosomal abnormalities and gene mutation were determined by FISH with probes for chromosomes 3,7, 17 and pl6. The sensitivity of FISH and cytological examination in diagnosing transitional cell carcinoma of bladder was studied and compared. Results Big and irregular cell nuclei and position effect variegation were seen in some cells ( P < 0. 05 ). In stage Tl bladder cancer, the sensitivity of FISH and cytological analysis was 83 . 6 % and 31.3% respectively, while in stage T2 - T4 , their sensitivity was 100 % and 55. 6 % respectively. Their sensitivity was 36. 3 % and 9.1% respectively with grade Gl versus 95. 2 % and 42. 9 % respectively with grade G2, and the sensitivity was 100 % and 50 % respectively with grade G3 ( P < 0. 05 ) in a comparable manner. There was significant difference in total sensitivity for diagnosis and staging of transitional cell carcinoma of bladder between FISH and cytological examination ( P < 0. 05 ). Conclusion FISH is more sensitive than cytological examination, hence it may be applied as a valuable method for diagnosing transitional cell carcinoma of bladder.%目的 评价荧光原位杂交(FISH)在诊断膀胱移行细胞癌各分期分级中的敏感性.方法 应用荧光标记的3、7、17号染色体及p16基因探针检测138例血尿患者尿液中脱落细胞的染色体及基因畸变情况,根据诊断标准计算FISH诊断膀胱移行细胞癌的敏感性.同时留取上述患者尿液送病理科行尿找瘤细胞检查,并比较两种方法对膀胱移行细胞癌的诊断效能.结果 FISH图像中细胞核大且不规则者更易观察到染色体基因异常情况,并可见具有较多染

  3. Lyophilization-induced protein denaturation in phosphate buffer systems: monomeric and tetrameric beta-galactosidase.

    Science.gov (United States)

    Pikal-Cleland, K A; Carpenter, J F

    2001-09-01

    During freezing in phosphate buffers, selective precipitation of a less soluble buffer component and subsequent pH shifts may induce protein denaturation. Previous reports indicate significantly more inactivation and secondary structural perturbation of monomeric and tetrameric beta-galactosidase (beta-gal) during freeze-thawing in sodium phosphate (NaP) buffer as compared with potassium phosphate (KP) buffer. This observation was attributed to the significant pH shifts (from 7.0 to as low as 3.8) observed during freezing in the NaP buffer (1). In the current study, we investigated the impact of the additional stress of dehydration after freezing on the recovery of active protein on reconstitution and the retention of the native structure in the dried state. Freeze-drying monomeric and tetrameric beta-gal in either NaP or KP buffer resulted in significant secondary structural perturbations, which were greatest for the NaP samples. However, similar recoveries of active monomeric protein were observed after freeze-thawing and freeze-drying, indicating that most dehydration-induced unfolding was reversible on reconstitution of the freeze-dried protein. In contrast, the tetrameric protein was more susceptible to dehydration-induced denaturation as seen by the greater loss in activity after reconstitution of the freeze-dried samples relative to that measured after freeze-thawing. To ensure optimal protein stability during freeze-drying, the protein must be protected from both freezing and dehydration stresses. Although poly(ethylene glycol) and dextran are preferentially excluded solutes and should confer protection during freezing, they were unable to prevent lyophilization-induced denaturation. In addition, Tween did not foster maintenance of native protein during freeze-drying. However, sucrose, which hydrogen bonds to dried protein in the place of lost water, greatly reduced freezing- and drying-induced denaturation, as observed by the high retention of native

  4. Protein denaturation due to the action of surfactants: a study by SAXS and ITC

    Energy Technology Data Exchange (ETDEWEB)

    Oseliero Filho, Pedro Leonidas; Oliveira, Cristiano Luis Pinto de [Universidade de Sao Paulo (USP), SP (Brazil); Pedersen, Jan Skov; Otzen, Daniel Erik [University of Aarhus (Denmark)

    2012-07-01

    Full text: Proteins are the major constituent of biological systems along with carbohydrates, lipids and nucleic acids (DNA and RNA). According to their structure and composition, proteins perform several functions in the organism, starting from the macroscopic level, with participation on the olfaction of animals, down to the cellular level, allocated in the membrane and making the connection between extra and intracellular environment. The function of a protein (which may be enzymatic, hormonal, structural, energetic, transport etc) is related to several factors including its structure (primary, secondary, tertiary or quaternary). Denaturation occurs when the secondary structure and/or tertiary is lost, which is almost always followed by the loss of the associated biological function. Temperature, pH and the action of surfactants influence the process of the denaturation. The influence of surfactants to the protein structure and function is the aim of this work. Therefore we are using an isolated protein, {alpha}-lactalbumin, that is found in the milk and whose function is related to the synthesis of galactose. The purpose is to characterize, in a thermodynamic-structural point of view, the denaturation of alpha-lactalbumin in the presence of surfactants anionic (sodium dodecyl sulfate - SDS), cationic (tetradecyltrimethylammonium bromide - TTAB), zwitterionic (2-diheptanoyl-sn-glycero-3- phosphocholine - DHPC) and nonionic (decyl-{beta}-D-Maltopyranoside - DM). The isothermal titration calorimetry (ITC) technique, which provides information of structural changes from changes in energy, represents the starting point for the study, while the technique of small angle X-ray scattering (SAXS) provides information about the structural characteristics of surfactant-protein complexes formed at each step of the denaturation process. The data analysis is in the initial stage, but it was possible to obtain general parameters related to the complex formed from the

  5. Antibodies with specificity for native and denatured forms of ovalbumin differ in reactivity between enzyme-linked immunosorbent assays

    DEFF Research Database (Denmark)

    Holm, B. E.; Bergmann, Ann Christina; Hansen, Paul Robert

    2015-01-01

    In this study, polyclonal and monoclonal antibodies to native and denatured chicken ovalbumin (OVA) were produced to compare their dependency on continuous and three-dimensional epitopes. These antibodies were characterized with respect to reactivity to native and denatured OVA by enzyme......-linked immunosorbent assay (ELISA) employing surface-bound OVA and streptavidin-capture ELISA to determine whether effects of different coating influence antibody specificity and with respect to epitope specificity by peptide ELISA, using overlapping peptides, covering the complete OVA sequence. Polyclonal antibodies...... to native OVA reacted strongly with native and denatured OVA in both assays, but did not react with the overlapping peptides. Polyclonal antibodies to denatured OVA reacted strongly with both OVA forms and with several of the overlapping peptides. Monoclonal antibodies to native OVA reacted preferentially...

  6. Hybrid Metaheuristics

    CERN Document Server

    2013-01-01

    The main goal of this book is to provide a state of the art of hybrid metaheuristics. The book provides a complete background that enables readers to design and implement hybrid metaheuristics to solve complex optimization problems (continuous/discrete, mono-objective/multi-objective, optimization under uncertainty) in a diverse range of application domains. Readers learn to solve large scale problems quickly and efficiently combining metaheuristics with complementary metaheuristics, mathematical programming, constraint programming and machine learning. Numerous real-world examples of problems and solutions demonstrate how hybrid metaheuristics are applied in such fields as networks, logistics and transportation, bio-medical, engineering design, scheduling.

  7. Fluorescence in situ hybridization for the tissue detection of bacterial pathogens associated with porcine infections

    DEFF Research Database (Denmark)

    Jensen, Henrik Elvang; Jensen, Louise Kruse; Barington, Kristiane

    2015-01-01

    sequences within intact cells. FISH allows direct histological localization of the bacteria in the tissue and thereby a correlation between the infection and the histopathological changes present. This chapter presents protocols for FISH identification of bacterial pathogens in fixed deparaffinized tissue......Fluorescence in situ hybridization (FISH) is an efficient technique for the identification of specific bacteria in tissue of both experimental and spontaneous infections. The method detects specific sequences of nucleic acids by hybridization of fluorescently labeled probes to complementary target...

  8. Fluorescence In Situ Hybridization for the Tissue Detection of Bacterial Pathogens Associated with Porcine Infections

    DEFF Research Database (Denmark)

    Elvang Jensen, Henrik; Jensen, Louise Kruse; Barington, Kristiane

    2015-01-01

    sequences within intact cells. FISH allows direct histological localization of the bacteria in the tissue and thereby a correlation between the infection and the histopathological changes present. This chapter presents protocols for FISH identification of bacterial pathogens in fixed deparaffinized tissue......Fluorescence in situ hybridization (FISH) is an efficient technique for the identification of specific bacteria in tissue of both experimental and spontaneous infections. The method detects specific sequences of nucleic acids by hybridization of fluorescently labeled probes to complementary target...

  9. Plastic fish

    CERN Multimedia

    Antonella Del Rosso

    2015-01-01

    In terms of weight, the plastic pollution in the world’s oceans is estimated to be around 300,000 tonnes. This plastic comes from both land-based and ocean-based sources. A lecture at CERN by chemist Wolfgang Trettnak addressed this issue and highlighted the role of art in raising people’s awareness.   Artwork by Wolfgang Trettnak. Packaging materials, consumer goods (shoes, kids’ toys, etc.), leftovers from fishing and aquaculture activities… our oceans and beaches are full of plastic litter. Most of the debris from beaches is plastic bottles. “PET bottles have high durability and stability,” explains Wolfgang Trettnak, a chemist by education and artist from Austria, who gave a lecture on this topic organised by the Staff Association at CERN on 26 May. “PET degrades very slowly and the estimated lifetime of a bottle is 450 years.” In addition to the beach litter accumulated from human use, rivers bring several ki...

  10. Imaging high-intensity focused ultrasound-induced tissue denaturation by multispectral photoacoustic method: an ex vivo study.

    Science.gov (United States)

    Sun, Yao; O'Neill, Brian

    2013-03-10

    We present an ex vivo study for the first time, to the best of our knowledge, in multispectral photoacoustic imaging (PAI) of tissue denaturation induced by high-intensity focused ultrasound (HIFU) in this paper. Tissue of bovine muscle was thermally treated in a heated water bath and by HIFU, and then was imaged using a multispectral photoacoustic approach. Light at multiple optical wavelengths between 700 and 900 nm was delivered to the treated bovine muscle tissue to excite the photoacoustic signal. Apparent tissue denaturation has been observed in multispectral photoacoustic images after being treated in a water bath and by HIFU. It is interesting that the denaturation is more striking at shorter optical wavelength photoacoustic images than at longer optical wavelength photoacoustic images. Multispectral photoacoustic images of the tissue denaturation were further analyzed and the photoacoustic spectrums of the denaturized tissue were calculated in this paper. This study suggests that a multispectral PAI approach might be a promising tool to evaluate tissue denaturation induced by HIFU treatment.

  11. Denatured-state energy landscapes of a protein structural database reveal the energetic determinants of a framework model for folding.

    Science.gov (United States)

    Wang, Suwei; Gu, Jenny; Larson, Scott A; Whitten, Steven T; Hilser, Vincent J

    2008-09-19

    Position-specific denatured-state thermodynamics were determined for a database of human proteins by use of an ensemble-based model of protein structure. The results of modeling denatured protein in this manner reveal important sequence-dependent thermodynamic properties in the denatured ensembles as well as fundamental differences between the denatured and native ensembles in overall thermodynamic character. The generality and robustness of these results were validated by performing fold-recognition experiments, whereby sequences were matched with their respective folds based on amino acid propensities for the different energetic environments in the protein, as determined through cluster analysis. Correlation analysis between structure and energetic information revealed that sequence segments destined for beta-sheet in the final native fold are energetically more predisposed to a broader repertoire of states than are sequence segments destined for alpha-helix. These results suggest that within the subensemble of mostly unstructured states, the energy landscapes are dominated by states in which parts of helices adopt structure, whereas structure formation for sequences destined for beta-strand is far less probable. These results support a framework model of folding, which suggests that, in general, the denatured state has evolutionarily evolved to avoid low-energy conformations in sequences that ultimately adopt beta-strand. Instead, the denatured state evolved so that sequence segments that ultimately adopt alpha-helix and coil will have a high intrinsic structure formation capability, thus serving as potential nucleation sites.

  12. Hybrid intermediaries

    OpenAIRE

    Cetorelli, Nicola

    2014-01-01

    I introduce the concept of hybrid intermediaries: financial conglomerates that control a multiplicity of entity types active in the "assembly line" process of modern financial intermediation, a system that has become known as shadow banking. The complex bank holding companies of today are the best example of hybrid intermediaries, but I argue that financial firms from the "nonbank" space can just as easily evolve into conglomerates with similar organizational structure, thus acquiring the cap...

  13. Hybrid composites

    CSIR Research Space (South Africa)

    Jacob John, Maya

    2009-04-01

    Full Text Available effect was observed for the elongation at break of the hybrid composites. The impact strength of the hybrid composites increased with the addition of glass fibres. The tensile and impact properties of thermoplastic natural rubber reinforced short... panels made from conventional structural materials. Figure 3 illustrates the performance of cellular biocomposite panels against conventional systems used for building and residential construction, namely a pre- cast pre-stressed hollow core concrete...

  14. Three Kinds of Fish

    DEFF Research Database (Denmark)

    Høst, Jeppe Engset

    2012-01-01

    There are three kinds of fish. Fish you were given, fish you bought and fish you lease. This might sound a bit odd, but it is nevertheless the basis for the activities of Danish commercial fishers since the introduction of transferable fishing concessions (TFCs) in 2007. In the current 2012 reform...... of market based systems are wild speculation, concentration and monopolization of fishing access and subsequent leasing with fishing communities and new entrants very likely being worse off (see for example the chapter “From fishing rights to financial derivatives” is this volume or Olson 2011; Sumaila 2010...... will examine five Danish fishing operations and discuss how they have reacted in different ways to the newly introduced system of transferable fishing concessions. By introducing TFCs as a solution to fleet overcapacity, the EU Commission will also be introducing a system where buying, selling and leasing...

  15. Embryonic and genetic manipulation in fish

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Fishes, the biggest and most diverse community in vertebrates are good experimental models for studies of cell and developmental biology by many favorable charac teristics. Nuclear transplantation in fish has been thor oughly studied in China since 1960s. Fish nuclei of em bryonic cells from different genera were transplanted into enucleated eggs generating nucleo-cytoplasmic hybrids of adults. Most importantly, nuclei of cultured goldfish kid ney cells had been reprogrammed in enucleated eggs to support embryogenesis and ontogenesis of a fertile fish. This was the first case of cloned fish with somatic cells. Based on the technique of microinjection, recombinant MThGH gene has been transferred into fish eggs and the first batch of transgenic fish were produced in 1984. The behavior of foreign gene was characterized and the onset of the foreign gene replication occurred between the blas tula to gastrula stages and random integration mainly oc curred at later stages of embryogenesis. This eventually led to the transgenlc mosaicism. The MThGH-transferred common carp enhanced growth rate by 2-4 times in the founder juveniles and doubled the body weight in the adults. The transgenic common carp were more efficient in utilizing dietary protein than the controls. An “all-fish” gene construct CAgcGH has been made by splicing the common carpβ-actin gene (CA) promoter onto the grass carp growth hormone gene (gcGH) coding sequence. The CAgcGH-transferred Yellow River Carp have also shown significantly fast-growth trait. Combination of techniques * Corresponding author, Fax: 0086-27-87876624 E-mail: zyzhu@ihb.ac.cn of fish cell culture, gene transformation with cultured cells and nuclear transplantation should be able to generate ho mogeneous strain of valuable transgenic fish to fulfil human requirement in 21st century.

  16. Multiple unfolding intermediates of human placental alkaline phosphatase in equilibrium urea denaturation.

    Science.gov (United States)

    Hung, H C; Chang, G G

    2001-12-01

    Alkaline phosphatase is an enzyme with a typical alpha/beta hydrolase fold. The conformational stability of the human placental alkaline phosphatase was examined with the chemical denaturant urea. The red shifts of fluorescence spectra show a complex unfolding process involving multiple equilibrium intermediates indicating differential stability of the subdomains of the enzyme. None of these unfolding intermediates were observed in the presence of 83 mM NaCl, indicating the importance of ionic interactions in the stabilization of the unfolding intermediates. Guanidinium chloride, on the other hand, could stabilize one of the unfolding intermediates, which is not a salt effect. Some of the unfolding intermediates were also observed in circular dichroism spectroscopy, which clearly indicates steady loss of helical structure during unfolding, but very little change was observed for the beta strand content until the late stage of the unfolding process. The enzyme does not lose its phosphate-binding ability after substantial tertiary structure changes, suggesting that the substrate-binding region is more resistant to chemical denaturant than the other structural domains. Global analysis of the fluorescence spectral change demonstrated the following folding-unfolding process of the enzyme: N I(1) I(2) I(3) I(4) I(5) D. These discrete intermediates are stable at urea concentrations of 2.6, 4.1, 4.7, 5.5, 6.6, and 7.7 M, respectively. These intermediates are further characterized by acrylamide and/or potassium iodide quenching of the intrinsic fluorescence of the enzyme and by the hydrophobic probes, 1-anilinonaphthalene-8-sulfonic acid and 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid. The stepwise unfolding process was interpreted by the folding energy landscape in terms of the unique structure of the enzyme. The rigid central beta-strand domain is surrounded by the peripheral alpha-helical and coil structures, which are marginally stable toward a chemical

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

    Science.gov (United States)

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

    2005-09-01

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

  18. A study of the thermal denaturation of the S-layer protein from Lactobacillus salivarius

    Science.gov (United States)

    Lighezan, Liliana; Georgieva, Ralitsa; Neagu, Adrian

    2012-09-01

    Surface layer (S-layer) proteins display an intrinsic self-assembly property, forming monomolecular crystalline arrays, identified in outermost structures of the cell envelope in many organisms, such as bacteria and archaea. Isolated S-layer proteins also possess the ability to recrystallize into regular lattices, being used in biotechnological applications, such as controlling the architecture of biomimetic surfaces. To this end, the stability of the S-layer proteins under high-temperature conditions is very important. In this study, the S-layer protein has been isolated from Lactobacillus salivarius 16 strain of human origin, and purified by cation-exchange chromatography. Using circular dichroism (CD) spectroscopy, we have investigated the thermal denaturation of the S-layer protein. The far- and near-UV CD spectra have been collected, and the temperature dependence of the CD signal in these spectral domains has been analyzed. The variable temperature results show that the secondary and tertiary structures of the S-layer protein change irreversibly due to the heating of the sample. After the cooling of the heated protein, the secondary and tertiary structures are partially recovered. The denaturation curves show that the protein unfolding depends on the sample concentration and on the heating rate. The secondary and tertiary structures of the protein suffer changes in the same temperature range. We have also detected an intermediate state in the protein denaturation pathway. Our results on the thermal behavior of the S-layer protein may be important for the use of S-layer proteins in biotechnological applications, as well as for a better understanding of the structure and function of S-layer proteins.

  19. Stabilizing effect of biochar on soil extracellular enzymes after a denaturing stress.

    Science.gov (United States)

    Elzobair, Khalid A; Stromberger, Mary E; Ippolito, James A

    2016-01-01

    Stabilizing extracellular enzymes may maintain enzymatic activity while protecting enzymes from proteolysis and denaturation. A study determined whether a fast pyrolysis hardwood biochar (CQuest™) would reduce evaporative losses, subsequently stabilizing soil extracellular enzymes and prohibiting potential enzymatic activity loss following a denaturing stress (microwaving). Soil was incubated in the presence of biochar (0%, 1%, 2%, 5%, or 10% by wt.) for 36 days and then exposed to microwave energies (0, 400, 800, 1600, or 3200 J g(-1) soil). Soil enzymes (β-glucosidase, β-d-cellobiosidase, N-acetyl-β-glucosaminidase, phosphatase, leucine aminopeptidase, β-xylosidase) were analyzed by fluorescence-based assays. Biochar amendment reduced leucine aminopeptidase and β-xylosidase potential activity after the incubation period and prior to stress exposure. The 10% biochar rate reduced soil water loss at the lowest stress level (400 J microwave energy g(-1) soil). Enzyme stabilization was demonstrated for β-xylosidase; intermediate biochar application rates prevented a complete loss of this enzyme's potential activity after soil was exposed to 400 (1% biochar treatment) or 1600 (5% biochar treatment) J microwave energy g(-1) soil. Remaining enzyme potential activities were not affected by biochar, and activities decreased with increasing stress levels. We concluded that biochar has the potential to reduce evaporative soil water losses and stabilize certain extracellular enzymes where activity is maintained after a denaturing stress; this effect was biochar rate and enzyme dependent. While biochar may reduce the potential activity of certain soil extracellular enzymes, this phenomenon was not universal as the majority of enzymes assayed in this study were unaffected by exposure to biochar.

  20. Detection of enzymes active on various beta-1,3-glucans after denaturing polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Trudel, J; Grenier, J; Asselin, A

    1998-07-01

    Enzymes were assayed for glucanase activity after denaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in gels containing beta-1,3-glucans embedded as substrate. Lentinan, curdlan, paramylon, baker's yeast alkali-insoluble glucan, baker's yeast alkali-soluble glucan and carboxymethyl (CM)-pachyman were compared to oligomeric laminarin, which is the usual substrate for assaying beta-1,3-glucanase activities. Detecting enzyme activities by aniline blue fluorescent staining was also compared with the staining of released reducing sugars by 2,3,5-triphenyltetrazolium chloride (TTC). For the nonreduced proteins, the Driselase extract exhibited one major band at 32.5 kDa and one less intense band at 23 kDa for most substrates with the two detection procedures. No Lyticase enzyme was detected in either detection procedures for all tested substrates. For barley enzymes, no activity was revealed after aniline blue staining while one undescribed 19 kDa glucanase activity was best shown after TTC staining with curdlan, paramylon and CM-pachyman as substrates. In the case of reduced proteins, the Lyticase extract yielded three bands (33, 36 and 46 kDa) on several substrates with both detection procedures. This was the same for the barley leaf extract (32, 36 and 39 kDa). The Driselase extract showed one 42 kDa band. Many enzymes active on beta-1,3-glucans are thus best revealed when proteins are denatured and reduced and when protein renaturation after SDS-PAGE involves a pH 8.0 treatment and the inclusion of 1 mM cysteine in buffers. However, some enzymes are only detected when proteins are denatured without reduction. Finally, the use of various polymeric beta-1,3-glucan substrates different from oligomeric laminarin is necessary to detect new types of enzymes such as the 19 kDa barley glucanase.

  1. Screening for mutations in the uroporphyrinogen decarboxylase gene using denaturing gradient gel electrophoresis

    DEFF Research Database (Denmark)

    Christiansen, L; Ged, C; Hombrados, I

    1999-01-01

    The two porphyrias, familial porphyria cutanea tarda (fPCT) and hepatoerythropoietic porphyria (HEP), are associated with mutations in the gene encoding the enzyme uroporphyrinogen decarboxylase (UROD). Several mutations, most of which are private, have been identified in HEP and fPCT patients......, confirming the heterogeneity of the underlying genetic defects of these diseases. We have established a denaturing gradient gel electrophoresis (DGGE) assay for mutation detection in the UROD gene, enabling the simultaneous screening for known and unknown mutations. The established assay has proved able...

  2. Light and Heat Induced Denaturation of Photosystem Ⅱ Core Antenna Complex CP47

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Light and heat induced denaturation of CP47, the core antenna complex of photosystem Ⅱ purified from spinach, were investigated using absorption and circular dichroism spectra.Light caused the destruction of chlorophyll a and excitonic interaction of chlorophyll a in CP47, while the protein secondary structure was not apparently changed.Heat induced the destruction of protein secondary structure and excitonic interaction of chlorophyll a, but the chlorophyll a molecule was not damaged.The results suggest that both the chlorophyll a molecular structure and the protein native conformation are necessary for excitonic interaction of chlorophyll a and the energy transfer function of the chlorophyll a binding protein.

  3. Optical Tweezers Analysis of Double-Stranded DNA Denaturation in the Presence of Urea

    Science.gov (United States)

    Zhu, Chunli; Li, Jing

    2016-09-01

    Urea is a kind of denaturant prone to form hydrogen bonds with the electronegative centers of the nitrogenous bases, threatening the stability of hydrogen bonds between DNA base pairs. In this paper, the stability and stiffness of DNA double helix influenced by urea are investigated at single-molecule level using optical tweezers. Experimental results show that DNA's double helix stability and stiffness both decrease with increasing urea concentration. In addition, the re-forming of ruptured hydrogen bonds between the base pairs is blocked by urea as the tension on DNA is released.

  4. Two-dimensional salt and temperature DNA denaturation analysis using a magnetoresistive sensor

    DEFF Research Database (Denmark)

    Rizzi, Giovanni; Dufva, Martin; Hansen, Mikkel Fougt

    2017-01-01

    . The results demonstrate that concentration melting provides an attractive alternative to temperature melting in on-chip DNA denaturation experiments and further show that the magnetoresistive platform is attractive due to its low cross-sensitivity to temperature and liquid composition....... temperature as well as salt concentration. The salt concentration melting curves were found to be more reliable than temperature melting curves. We performed a two-dimensional mapping of the melting profiles of a target to probes targeting its wild type (WT) and mutant type (MT) variants in the temperature...

  5. Glassy behavior of denatured DNA films studied by differential scanning calorimetry.

    Science.gov (United States)

    Valle-Orero, Jessica; Garden, Jean-Luc; Richard, Jacques; Wildes, Andrew; Peyrard, Michel

    2012-04-12

    We use differential scanning calorimetry (DSC) to study the properties of DNA films, made of oriented fibers, heated above the thermal denaturation temperature of the double helical form. The films show glassy properties that we investigate in two series of experiments, a slow cooling at different rates followed by a DSC scan upon heating and aging at a temperature below the glass transition. Introducing the fictive temperature to characterize the glass allows us to derive quantitative information on the relaxations of the DNA films, in particular to evaluate their enthalpy barrier. A comparison with similar aging studies on PVAc highlights some specificities of the DNA samples.

  6. Hydrolysates of Fish Skin Collagen: An Opportunity for Valorizing Fish Industry Byproducts.

    Science.gov (United States)

    Blanco, María; Vázquez, José Antonio; Pérez-Martín, Ricardo I; Sotelo, Carmen G

    2017-05-05

    During fish processing operations, such as skinning and filleting, the removal of collagen-containing materials can account for up to 30% of the total fish byproducts. Collagen is the main structural protein in skin, representing up to 70% of dry weight depending on the species, age and season. It has a wide range of applications including cosmetic, pharmaceutical, food industry, and medical. In the present work, collagen was obtained by pepsin extraction from the skin of two species of teleost and two species of chondrychtyes with yields varying between 14.16% and 61.17%. The storage conditions of the skins appear to influence these collagen extractions yields. Pepsin soluble collagen (PSC) was enzymatically hydrolyzed and the resultant hydrolysates were ultrafiltrated and characterized. Electrophoretic patterns showed the typical composition of type I collagen, with denaturation temperatures ranged between 23 °C and 33 °C. In terms of antioxidant capacity, results revealed significant intraspecific differences between hydrolysates, retentate, and permeate fractions when using β-Carotene and DPPH methods and also showed interspecies differences between those fractions when using DPPH and ABTS methods. Under controlled conditions, PSC hydrolysates from Prionace glauca, Scyliorhinus canicula, Xiphias gladius, and Thunnus albacares provide a valuable source of peptides with antioxidant capacities constituting a feasible way to efficiently upgrade fish skin biomass.

  7. 转基因水稻中外源基因的荧光原位杂交(FISH)分析%DETECTION AND ANALYSIS OF ALIEN GENES IN TRANSGENIC RICE BY FLUORESCENCE IN SITU HYBRIDIZATION

    Institute of Scientific and Technical Information of China (English)

    金危危; 李霞; 李宗芸; 宁顺斌; 凌定厚; 宋运淳

    2001-01-01

    利用荧光原位杂交(FISH),在供试8个转基因水稻株系的染色体上检出了转入的外源基因barnase-ps1片段.其中两株系各检出了两个不同的信号位点,而其他株系均只检出一个信号位点.所有染色体着丝粒区都没有杂交信号.结合Southern杂交及表达分析发现,大多数株系中,整合在染色体端部区的barnase基因表现为高水平表达,而靠近着丝粒区的则表现为低水平表达,表明表达水平与转基因整合位置可能存在一定程度的相关性.表达水平与拷贝数无明显相关.本文还利用多色荧光原位杂交(Multi-color FISH)技术,进行了barnaseps1片段与报道基因pHctinG的共杂交,多数情况下,barnase-ps1片段与报道基因整合在染色体的同一位置.

  8. Effect of heating strategies on whey protein denaturation--Revisited by liquid chromatography quadrupole time-of-flight mass spectrometry.

    Science.gov (United States)

    Akkerman, M; Rauh, V M; Christensen, M; Johansen, L B; Hammershøj, M; Larsen, L B

    2016-01-01

    Previous standards in the area of effect of heat treatment processes on milk protein denaturation were based primarily on laboratory-scale analysis and determination of denaturation degrees by, for example, electrophoresis. In this study, whey protein denaturation was revisited by pilot-scale heating strategies and liquid chromatography quadrupole time-of-flight mass spectrometer (LC/MC Q-TOF) analysis. Skim milk was heat treated by the use of 3 heating strategies, namely plate heat exchanger (PHE), tubular heat exchanger (THE), and direct steam injection (DSI), under various heating temperatures (T) and holding times. The effect of heating strategy on the degree of denaturation of β-lactoglobulin and α-lactalbumin was determined using LC/MC Q-TOF of pH 4.5-soluble whey proteins. Furthermore, effect of heating strategy on the rennet-induced coagulation properties was studied by oscillatory rheometry. In addition, rennet-induced coagulation of heat-treated micellar casein concentrate subjected to PHE was studied. For skim milk, the whey protein denaturation increased significantly as T and holding time increased, regardless of heating method. High denaturation degrees were obtained for T >100°C using PHE and THE, whereas DSI resulted in significantly lower denaturation degrees, compared with PHE and THE. Rennet coagulation properties were impaired by increased T and holding time regardless of heating method, although DSI resulted in less impairment compared with PHE and THE. No significant difference was found between THE and PHE for effect on rennet coagulation time, whereas the curd firming rate was significantly larger for THE compared with PHE. Micellar casein concentrate possessed improved rennet coagulation properties compared with skim milk receiving equal heat treatment.

  9. Comparative study of denaturation of whey protein isolate (WPI) in convective air drying and isothermal heat treatment processes.

    Science.gov (United States)

    Haque, M Amdadul; Aldred, Peter; Chen, Jie; Barrow, Colin J; Adhikari, Benu

    2013-11-15

    The extent and nature of denaturation of whey protein isolate (WPI) in convective air drying environments was measured and analysed using single droplet drying. A custom-built, single droplet drying instrument was used for this purpose. Single droplets having 5±0.1μl volume (initial droplet diameter 1.5±0.1mm) containing 10% (w/v) WPI were dried at air temperatures of 45, 65 and 80°C for 600s at constant air velocity of 0.5m/s. The extent and nature of denaturation of WPI in isothermal heat treatment processes was measured at 65 and 80°C for 600s and compared with those obtained from convective air drying. The extent of denaturation of WPI in a high hydrostatic pressure environment (600MPa for 600s) was also determined. The results showed that at the end of 600s of convective drying at 65°C the denaturation of WPI was 68.3%, while it was only 10.8% during isothermal heat treatment at the same medium temperature. When the medium temperature was maintained at 80°C, the denaturation loss of WPI was 90.0% and 68.7% during isothermal heat treatment and convective drying, respectively. The bovine serum albumin (BSA) fraction of WPI was found to be more stable in the convective drying conditions than β-lactoglobulin and α-lactalbumin, especially at longer drying times. The extent of denaturation of WPI in convective air drying (65 and 80°C) and isotheral heat treatment (80°C) for 600s was found to be higher than its denaturation in a high hydrostatic pressure environment at ambient temperature (600MPa for 600s).

  10. Air leak seal for lung dissection plane with diode laser irradiation: monitoring heat-denature with auto-fluorescence

    Science.gov (United States)

    Gotoh, Maya; Arai, Tsunenori

    2008-02-01

    We studied the monitoring of heat-denature by autofluorescence spectrum from lung dissection plane during laser air leak sealing procedure. In order to seal the air leakage from lung in thoracotomy, we proposed novel laser sealing method with the combination of the diode laser (810nm wavelength) irradiation and indocyanine green staining (peak absorption wavelength: 805 nm). This sealing method is expected to preserve the postoperative ventilatory capacity and achieve minimally invasive surgery. We previously reported that this laser sealing only requires thin sealing margin (less than 300 μm in thickness) compared with that of the suturing or stapling. The most serious issue on the laser air leak sealing might be re-air-leakage due to rigid surface layer caused by excessive heat-denature, such as carbonization. We should achieve laser air leak sealing minimizing the degree of heat denature. Dissection planes of isolated porcine lung with /without the diode laser irradiation were prepared as samples. We measured the auto-fluorescence from these samples using a spectrometer. When the diode laser was irradiated with 400J/cm2, the surface of diode laser irradiated lung was fully carbonized. The ration of auto-fluorescence emission of 450nm / 500 nm, with 280 nm excitation wavelength was decreased less tha 50 % of initial value. That of 600 nm / 500 nm was increased over 700 % of initial value. The decreasing of the 450 nm auto-fluorescence intensity might be attributed to the heat-denaturing of the interstitial collagen in lung. However, increasing of the 600 nm didn't specify the origins, we suppose it might be originated from heat-denature substance, like carbonization. We could establish the useful monitoring for lung heat-denaturing with simple methodology. We think the auto-fluorescence measurement can be helpful not only for understanding the sealing mechanism, but also for controlling the degree of heat-denaturing during the procedure.

  11. Molecular monitoring of the intestinal flora by denaturing high performance liquid chromatography.

    Science.gov (United States)

    Goldenberg, Oliver; Herrmann, Stefanie; Marjoram, Gina; Noyer-Weidner, Mario; Hong, George; Bereswill, Stefan; Göbel, Ulf B

    2007-01-01

    Gut flora analysis is hampered by the complexity of the intestinal microbiota and by inherent limitations of culture-based approaches. Therefore, culture-independent molecular methods based upon 16S rRNA gene analysis were applied successfully for the analysis of complex microbial communities. However, generally accepted and validated profiling methods such as denaturing and temperature gradient gel electrophoresis (DGGE/TGGE) are still laborious and time consuming. Thus, we adapted the separation of amplified bacterial 16S rRNA gene fragments by denaturing high performance liquid chromatography (DHPLC) using the WAVE Microbial Analysis System as a rapid and convenient means to display complex intestinal bacterial communities and to monitor changes in the gut flora. The separation of 16S rRNA gene fragments amplified from reference strains representing main gut bacterial populations and from human stool samples revealed that DHPLC analysis effectively detects bacterial groups predominant in the human gut flora. The investigation of faecal samples from hospitalized patients before, during and after antibiotic therapy showed that PCR-based DHPLC can be used to monitor gut flora changes. Results from DHPLC analysis were comparable with DGGE profiles generated from the same samples, demonstrating that the adapted DHPLC protocol is well suited for the analysis of complex microbial communities.

  12. Effect of sulfoxides on the thermal denaturation of hen lysozyme: A calorimetric and Raman study

    Science.gov (United States)

    Torreggiani, A.; Di Foggia, M.; Manco, I.; De Maio, A.; Markarian, S. A.; Bonora, S.

    2008-11-01

    A multidisciplinary study of the thermal denaturation of lysozyme in the presence of three sulfoxides with different length in hydrocarbon chain (DMSO, DESO, and DPSO) was carried out by means of DSC, Raman spectroscopy, and SDS-PAGE techniques. In particular, the Td and Δ H values obtained from the calorimetric measurements showed that lysozyme is partially unfolded by sulfoxides but most of the conformation holds native state. The sulfoxide denaturing ability increases in the order DPSO > DESO > DMSO. Moreover, only DMSO and DESO have a real effect in preventing the heat-induced inactivation of the protein and their maximum heat-protective ability is reached when the DMSO and DESO amount is ⩾25% w/w. The sulfoxide ability to act as effective protective agents against the heat-induced inactivation was confirmed by the protein analysis. The enzymatic activity, as well as the SDS-PAGE analysis, suggested that DESO, having a low hydrophobic character and a great ability to stabilise the three-dimensional water structure, is the most heat-protective sulfoxide. An accurate evaluation of the heat-induced conformational changes of the lysozyme structure before and after sulfoxide addition was obtained by the analysis of the Raman spectra. The addition of DMSO or DESO in low concentration resulted to sensitively decrease the heat-induced structural modifications of the protein.

  13. Ultrafast folding of WW domains without structured aromatic clusters in the denatured state.

    Science.gov (United States)

    Ferguson, N; Johnson, C M; Macias, M; Oschkinat, H; Fersht, A

    2001-11-06

    Ultrafast-folding proteins are important for combining experiment and simulation to give complete descriptions of folding pathways. The WW domain family comprises small proteins with a three-stranded antiparallel beta-sheet topology. Previous studies on the 57-residue YAP 65 WW domain indicate the presence of residual structure in the chemically denatured state. Here we analyze three minimal core WW domains of 38-44 residues. There was little spectroscopic or thermodynamic evidence for residual structure in either their chemically or thermally denatured states. Folding and unfolding kinetics, studied by using rapid temperature-jump and continuous-flow techniques, show that each domain folds and unfolds very rapidly in a two-state transition through a highly compact transition state. Folding half-times were as short as 17 micros at 25 degrees C, within an order of magnitude of the predicted maximal rate of loop formation. The small size and topological simplicity of these domains, in conjunction with their very rapid two-state folding, may allow us to reduce the difference in time scale between experiment and theoretical simulation.

  14. Increasing protein stability: Importance of ΔCp and the denatured state

    Science.gov (United States)

    Fu, Hailong; Grimsley, Gerald; Scholtz, J Martin; Pace, C Nick

    2010-01-01

    Increasing the conformational stability of proteins is an important goal for both basic research and industrial applications. In vitro selection has been used successfully to increase protein stability, but more often site-directed mutagenesis is used to optimize the various forces that contribute to protein stability. In previous studies, we showed that improving electrostatic interactions on the protein surface and improving the β-turn sequences were good general strategies for increasing protein stability, and used them to increase the stability of RNase Sa. By incorporating seven of these mutations in RNase Sa, we increased the stability by 5.3 kcal/mol. Adding one more mutation, D79F, gave a total increase in stability of 7.7 kcal/mol, and a melting temperature 28°C higher than the wild-type enzyme. Surprisingly, the D79F mutation lowers the change in heat capacity for folding, ΔCp, by 0.6 kcal/mol/K. This suggests that this mutation stabilizes structure in the denatured state ensemble. We made other mutants that give some insight into the structure present in the denatured state. Finally, the thermodynamics of folding of these stabilized variants of RNase Sa are compared with those observed for proteins from thermophiles. PMID:20340133

  15. Culture-independent analysis of probiotic products by denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Temmerman, R; Scheirlinck, I; Huys, G; Swings, J

    2003-01-01

    In order to obtain functional and safe probiotic products for human consumption, fast and reliable quality control of these products is crucial. Currently, analysis of most probiotics is still based on culture-dependent methods involving the use of specific isolation media and identification of a limited number of isolates, which makes this approach relatively insensitive, laborious, and time-consuming. In this study, a collection of 10 probiotic products, including four dairy products, one fruit drink, and five freeze-dried products, were subjected to microbial analysis by using a culture-independent approach, and the results were compared with the results of a conventional culture-dependent analysis. The culture-independent approach involved extraction of total bacterial DNA directly from the product, PCR amplification of the V3 region of the 16S ribosomal DNA, and separation of the amplicons on a denaturing gradient gel. Digital capturing and processing of denaturing gradient gel electrophoresis (DGGE) band patterns allowed direct identification of the amplicons at the species level. This whole culture-independent approach can be performed in less than 30 h. Compared with culture-dependent analysis, the DGGE approach was found to have a much higher sensitivity for detection of microbial strains in probiotic products in a fast, reliable, and reproducible manner. Unfortunately, as reported in previous studies in which the culture-dependent approach was used, a rather high percentage of probiotic products suffered from incorrect labeling and yielded low bacterial counts, which may decrease their probiotic potential.

  16. Urea-induced denaturation of apolipoprotein serum amyloid A reveals marginal stability of hexamer

    Science.gov (United States)

    Wang, Limin; Colón, Wilfredo

    2005-01-01

    Serum Amyloid A (SAA) is an acute phase reactant protein that is predominantly found bound to high-density lipoprotein in plasma. Upon inflammation, the plasma concentration of SAA can increase dramatically, occasionally leading to the development of amyloid A (AA) amyloidosis, which involves the deposition of SAA amyloid fibrils in major organs. We previously found that the murine isoform SAA2.2 exists in aqueous solution as a hexamer containing a central channel. Here we show using various biophysical and biochemical techniques that the SAA2.2 hexamer can be totally dissociated into monomer by ~2 M urea, with the concerted loss of its α-helical structure. However, limited trypsin proteolysis experiments in urea showed a conserved digestion profile, suggesting the preservation of major backbone topological features in the urea-denatured state of SAA2.2. The marginal stability of hexameric SAA2.2 and the presence of residual structure in the denatured monomeric protein suggest that both forms may interconvert in vivo to exert different functions to meet the various needs during normal physiological conditions and in response to inflammatory stimuli. PMID:15937280

  17. Urea denatured state ensembles contain extensive secondary structure that is increased in hydrophobic proteins

    Science.gov (United States)

    Nick Pace, C; Huyghues-Despointes, Beatrice M P; Fu, Hailong; Takano, Kazufumi; Scholtz, J Martin; Grimsley, Gerald R

    2010-01-01

    The goal of this article is to gain a better understanding of the denatured state ensemble (DSE) of proteins through an experimental and computational study of their denaturation by urea. Proteins unfold to different extents in urea and the most hydrophobic proteins have the most compact DSE and contain almost as much secondary structure as folded proteins. Proteins that unfold to the greatest extent near pH 7 still contain substantial amounts of secondary structure. At low pH, the DSE expands due to charge–charge interactions and when the net charge per residue is high, most of the secondary structure is disrupted. The proteins in the DSE appear to contain substantial amounts of polyproline II conformation at high urea concentrations. In all cases considered, including staph nuclease, the extent of unfolding by urea can be accounted for using the data and approach developed in the laboratory of Wayne Bolen (Auton et al., Proc Natl Acad Sci 2007; 104:15317–15323). PMID:20198681

  18. Approach to analysis of single nucleotide polymorphisms by automated constant denaturant capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Bjoerheim, Jens; Abrahamsen, Torveig Weum; Kristensen, Annette Torgunrud; Gaudernack, Gustav; Ekstroem, Per O

    2003-05-15

    Melting gel techniques have proven to be amenable and powerful tools in point mutation and single nucleotide polymorphism (SNP) analysis. With the introduction of commercially available capillary electrophoresis instruments, a partly automated platform for denaturant capillary electrophoresis with potential for routine screening of selected target sequences has been established. The aim of this article is to demonstrate the use of automated constant denaturant capillary electrophoresis (ACDCE) in single nucleotide polymorphism analysis of various target sequences. Optimal analysis conditions for different single nucleotide polymorphisms on ACDCE are evaluated with the Poland algorithm. Laboratory procedures include only PCR and electrophoresis. For direct genotyping of individual SNPs, the samples are analyzed with an internal standard and the alleles are identified by co-migration of sample and standard peaks. In conclusion, SNPs suitable for melting gel analysis based on theoretical thermodynamics were separated by ACDCE under appropriate conditions. With this instrumentation (ABI 310 Genetic Analyzer), 48 samples could be analyzed without any intervention. Several institutions have capillary instrumentation in-house, thus making this SNP analysis method accessible to large groups of researchers without any need for instrument modification.

  19. APPLICATION OF RE-ESTABLISHING DOSE-RESPONSE CURVES BY FLUORESCENCE IN SITU HYBRIDIZATION (FISH)%荧光原位杂交技术在剂量重建中的应用

    Institute of Scientific and Technical Information of China (English)

    段志凯; 张淑贤; 郭万龙; 吴启庆; 郭鲜花; 时爱丽; 王斌生

    2008-01-01

    采用不同剂量60Co γ射线照射两名正常人外周血淋巴细胞后,应用荧光原位杂交(FISH)技术检测染色体易位率,并拟合出剂量效应关系曲线为:Y=0.01316+0.07569D+0.01583D2.通过盲法对实验结果验证,验证结果表明拟合的曲线可用于剂量重建.FISH技术用来检测染色体稳定性畸变,既快速又准确,适合于染色体易位的检测,有望成为剂量重建的理想技术方法.

  20. 荧光原位杂交(FISH)技术在转基因小鼠检测中的应用%Application of the technique of fluorescence in situ hybridization in detecting transgenic mice

    Institute of Scientific and Technical Information of China (English)

    谢建云; 邵伟娟; 潘漪清; 高诚

    2002-01-01

    制备转基因小鼠特有的DIG标记探针,应用染色体荧光原位杂交(FISH)技术,对转基因小鼠外周血细胞的遗传性状进行分析,检测其合子类型,以挑选纯合子小鼠用于保种.结果杂合型小鼠61%的细胞有一个荧光信号,而纯合型小鼠约24%的细胞有两个荧光信号,48%的细胞有一个荧光信号,野生型小鼠无荧光信号.因此染色体荧光原位杂交技术可作为检测转基因小鼠遗传特性的一种方法.

  1. Exact Solution to the Extended Zwanzig Model for Quasi-Sigmoidal Chemically Induced Denaturation Profiles: Specific Heat and Configurational Entropy

    Directory of Open Access Journals (Sweden)

    G. E. Aguilar-Pineda

    2014-01-01

    Full Text Available Temperature and chemically induced denaturation comprise two of the most characteristic mechanisms to achieve the passage from the native state N to any of the unstructured states Dj in the denatured ensemble in proteins and peptides. In this work we present a full analytical solution for the configurational partition function qs of a homopolymer chain poly-X in the extended Zwanzig model (EZM for a quasisigmoidal denaturation profile. This solution is built up from an EZM exact solution in the case where the fraction α of native contacts follows exact linear dependence on denaturant’s concentration ζ; thus an analytical solution for L in the case of an exact linear denaturation profile is also provided. A recently established connection between the number ν of potential nonnative conformations per residue and temperature-independent helical propensity ω complements the model in order to identify specific proteinogenic poly-X chains, where X represents any of the twenty naturally occurring aminoacid residues. From qs, equilibrium thermodynamic potentials like entropy and average internal energy 〈E〉 and thermodynamic susceptibilities like specific heat C are calculated for poly-valine (poly-V and poly-alanine (poly-A chains. The influence of the rate at which native contacts denature as function of ζ on thermodynamic stability is also discussed.

  2. Effects of thermally induced denaturation on technological-functional properties of whey protein isolate-based films.

    Science.gov (United States)

    Schmid, M; Krimmel, B; Grupa, U; Noller, K

    2014-09-01

    This study examined how and to what extent the degree of denaturation affected the technological-functional properties of whey protein isolate (WPI)-based coatings. It was observed that denaturation affected the material properties of WPI-coated films significantly. Surface energy decreased by approximately 20% compared with native coatings. Because the surface energy of a coating should be lower than that of the substrate, this might result in enhanced wettability characteristics between WPI-based solution and substrate surface. Water vapor barrier properties increased by about 35% and oxygen barrier properties increased by approximately 33%. However, significant differences were mainly observed between coatings made of fully native WPI and ones with a degree of denaturation of 25%. Higher degrees of denaturation did not lead to further improvement of material properties. This observation offers cost-saving potential: a major share of denatured whey proteins may be replaced by fully native ones that are not exposed to energy-intensive heat treatment. Furthermore, native WPI solutions can be produced with higher dry matter content without gelatinizing. Hence, less moisture has to be removed through drying, resulting in reduced energy consumption.

  3. CLINICAL STUDY OF CHROMOSOMAL ABNORMALITY IN MISSED ABORTION TISSUE DETECTED BY FLUORESCENCE IN SITU HYBRIDIZATION TECHNIQUE%FISH 技术检测稽留流产组织中染色体异常的临床研究

    Institute of Scientific and Technical Information of China (English)

    王春珺; 于晶峰

    2014-01-01

    目的:探讨稽留流产胎儿组织细胞中染色体异常总发生率及异常种类和各种异常的发生率。方法:采用18号、X 和 Y 染色体着丝粒探针及13、16、21、22号染色体单一序列探针,对105例稽留流产胎儿组织进行FISH 检测。结果:49.5%的稽留流产是由胎儿染色体异常引起的;染色体异常的前3位为三倍体,16号染色体三体,X 单体,分别占染色体异常总发生率的32.7%、15.4%和15.4%;高龄组(≥35岁)和非高龄组(0.05);妊娠≤12wk和妊娠>12wk者染色体异常率分别为60.9%和31.7%,差异有统计学意义(P0. 05). The detection rate of chromosomal abnormality in the cases of ≤12 gestational weeks was 60. 9%,while the detection rate of chromosomal abnormality in the cases of >12 gestational weeks was 31. 7%,there was significant difference(P<0. 05). Conclusion:Most missed abortion are caused by genetic gene defect, FISH technique can detect aborted fetuses, find chromosomal abnormality rapidly and accurately,provide data for genetic counseling of subsequent pregnancy.

  4. Genetic identification of F1 and post-F1 serrasalmid juvenile hybrids in Brazilian aquaculture.

    Directory of Open Access Journals (Sweden)

    Diogo Teruo Hashimoto

    Full Text Available Juvenile fish trade monitoring is an important task on Brazilian fish farms. However, the identification of juvenile fish through morphological analysis is not feasible, particularly between interspecific hybrids and pure species individuals, making the monitoring of these individuals difficult. Hybrids can be erroneously identified as pure species in breeding facilities, which might reduce production on farms and negatively affect native populations due to escapes or stocking practices. In the present study, we used a multi-approach analysis (molecular and cytogenetic markers to identify juveniles of three serrasalmid species (Colossoma macropomum, Piaractus mesopotamicus and Piaractus brachypomus and their hybrids in different stocks purchased from three seed producers in Brazil. The main findings of this study were the detection of intergenus backcrossing between the hybrid ♀ patinga (P. mesopotamicus×P. brachypomus×♂ C. macropomum and the occurrence of one hybrid triploid individual. This atypical specimen might result from automixis, a mechanism that produces unreduced gametes in some organisms. Moreover, molecular identification indicated that hybrid individuals are traded as pure species or other types of interspecific hybrids, particularly post-F1 individuals. These results show that serrasalmid fish genomes exhibit high genetic heterogeneity, and multi-approach methods and regulators could improve the surveillance of the production and trade of fish species and their hybrids, thereby facilitating the sustainable development of fish farming.

  5. PNA-COMBO-FISH: From combinatorial probe design in silico to vitality compatible, specific labelling of gene targets in cell nuclei.

    Science.gov (United States)

    Müller, Patrick; Rößler, Jens; Schwarz-Finsterle, Jutta; Schmitt, Eberhard; Hausmann, Michael

    2016-07-01

    Recently, advantages concerning targeting specificity of PCR constructed oligonucleotide FISH probes in contrast to established FISH probes, e.g. BAC clones, have been demonstrated. These techniques, however, are still using labelling protocols with DNA denaturing steps applying harsh heat treatment with or without further denaturing chemical agents. COMBO-FISH (COMBinatorial Oligonucleotide FISH) allows the design of specific oligonucleotide probe combinations in silico. Thus, being independent from primer libraries or PCR laboratory conditions, the probe sequences extracted by computer sequence data base search can also be synthesized as single stranded PNA-probes (Peptide Nucleic Acid probes). Gene targets can be specifically labelled with at least about 20 PNA-probes obtaining visibly background free specimens. By using appropriately designed triplex forming oligonucleotides, the denaturing procedures can completely be omitted. These results reveal a significant step towards oligonucleotide-FISH maintaining the 3d-nanostructure and even the viability of the cell target. The method is demonstrated with the detection of Her2/neu and GRB7 genes, which are indicators in breast cancer diagnosis and therapy.

  6. Improving DNA capture on micro-arrays by integrated repeated denaturing

    NARCIS (Netherlands)

    Servoli, E.; Feitsma, H.M.; Kaptheijns, B.; Van der Zaag, P.J.; Wimberger-Friedl, R.

    2012-01-01

    Hybridization of nucleic acids to microarrays is a crucial step forseveral biological and biomedical applications. However, the poor efficiency and resulting long incubation times are major drawbacks. Inaddition to diffusion limitation, back-hybridization to complementary strands in solution is show

  7. Improving DNA capture on micro-arrays by integrated repeated denaturing

    NARCIS (Netherlands)

    Servoli, E.; Feitsma, H.M.; Kaptheijns, B.; Van der Zaag, P.J.; Wimberger-Friedl, R.

    2012-01-01

    Hybridization of nucleic acids to microarrays is a crucial step forseveral biological and biomedical applications. However, the poor efficiency and resulting long incubation times are major drawbacks. Inaddition to diffusion limitation, back-hybridization to complementary strands in solution is show

  8. Genetic population structure of turbot ( Scophthalmus maximus L.) supports the presence of multiple hybrid zones for marine fishes in the transition zone between the Baltic Sea and the North Sea

    DEFF Research Database (Denmark)

    Eg Nielsen, Einar; Nielsen, P.H.; Meldrup, Dorte

    2004-01-01

    Genetic population structure of turbot (Scophthalmus maximus L.) in the Northeast Atlantic was investigated using eight highly variable microsatellite loci. In total 706 individuals from eight locations with temporal replicates were assayed, covering an area from the French Bay of Biscay...... Sea, suggesting high gene flow among populations in these areas. In contrast, there was a sharp cline in genetic differentiation going from the low saline Baltic Sea to the high saline North Sea. The data were explained best by two divergent populations connected by a hybrid zone; however......, a mechanical mixing model could not be ruled out. A significant part of the genetic variance could be ascribed to variation among years within locality. Nevertheless, the population structure was relatively stable over time, suggesting that the observed pattern of genetic differentiation is biologically...

  9. Umatilla - Rough Fish Eradication

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — In order to enhance environmental conditions in the McCormack Slough on Umatilla NWR, the population of rough fish, including common carp (Cyprinus carpio) and...

  10. Textbook of fish health

    National Research Council Canada - National Science Library

    Post, George

    1987-01-01

    Written to fill the great need for a fish disease textbook for college students, this new edition contains the latest information and important discoveries related to those fish diseases that affect man economically...

  11. Scorpion fish sting

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/002849.htm Scorpion fish sting To use the sharing features on this page, please enable JavaScript. Scorpion fish are members of the family Scorpaenidae, which ...

  12. Pittsburgh Fish Fry Locations

    Data.gov (United States)

    Allegheny County / City of Pittsburgh / Western PA Regional Data Center — Lenten Fish Fry records for the Greater Pittsburgh region. Data is collected before and during the Lenten fish fry season each year by Code for Pittsburgh. Data is...

  13. 49 CFR 173.218 - Fish meal or fish scrap.

    Science.gov (United States)

    2010-10-01

    ... 49 Transportation 2 2010-10-01 2010-10-01 false Fish meal or fish scrap. 173.218 Section 173.218... Fish meal or fish scrap. (a) Except as provided in Column (7) of the HMT in § 172.101 of this subchapter, fish meal or fish scrap, containing at least 6%, but not more than 12% water, is authorized...

  14. Medusivorous fishes, a review

    NARCIS (Netherlands)

    Ates, R.M.L.

    1988-01-01

    A preliminary review is presented of fish species having consumed pelagic Cnidaria (Scyphozoa and Hydrozoa) as well as Ctenophora. Quantitative data are scarce. Knowledge of morphological and physiological adaptations of fishes foraging on gelatinous plankton is almost non-existent. Many fish specie

  15. Medusivorous fishes, a review

    OpenAIRE

    Ates, R.M.L.

    1988-01-01

    A preliminary review is presented of fish species having consumed pelagic Cnidaria (Scyphozoa and Hydrozoa) as well as Ctenophora. Quantitative data are scarce. Knowledge of morphological and physiological adaptations of fishes foraging on gelatinous plankton is almost non-existent. Many fish species consume medusae and some reasons to suspect that there are even more that do so, are discussed.

  16. Medusivorous fishes, a review

    NARCIS (Netherlands)

    Ates, R.M.L.

    1988-01-01

    A preliminary review is presented of fish species having consumed pelagic Cnidaria (Scyphozoa and Hydrozoa) as well as Ctenophora. Quantitative data are scarce. Knowledge of morphological and physiological adaptations of fishes foraging on gelatinous plankton is almost non-existent. Many fish

  17. Denaturation and Oxidative Stability of Hemp Seed (Cannabis sativa L.) Protein Isolate as Affected by Heat Treatment.

    Science.gov (United States)

    Raikos, Vassilios; Duthie, Garry; Ranawana, Viren

    2015-09-01

    The present study investigated the impact of heat treatments on the denaturation and oxidative stability of hemp seed protein during simulated gastrointestinal digestion (GID). Heat-denatured hemp protein isolate (HPI) solutions were prepared by heating HPI (2 mg/ml, pH 6.8) to 40, 60, 80 and 100 °C for 10 min. Heat-induced denaturation of the protein isolates was monitored by polyacrylamide gel electrophoresis. Heating HPI at temperatures above 80 °C significantly reduced solubility and led to the formation of large protein aggregates. The isolates were then subjected to in vitro GID and the oxidative stability of the generated peptides was investigated. Heating did not significantly affect the formation of oxidation products during GID. The results suggest that heat treatments should ideally remain below 80 °C if heat stability and solubility of HPI are to be preserved.

  18. FISH analysis in Prader-Willi and Angelman syndrome patients

    Energy Technology Data Exchange (ETDEWEB)

    Bettio, D.; Rizzi, N.; Giardino, D. [Centro Auxologico Italiano, Milan (Italy)] [and others

    1995-03-27

    We report on a combined high resolution cytogenetic and fluorescent in situ hybridization study (FISH) on 15 Prader-Willi syndrome (PWS) and 14 Angelman syndrome (AS) patients. High resolution banding showed a microdeletion in the 15q11-q13 region in 7 out of 15 PWS patients, and FISH analysis of the D15S11 and SNRPN cosmids demonstrated absence of the critical region in three additional cases. Likewise 8 out of 14 AS patients were found to be deleted with FISH, using the GABRB3 specific cosmid, whereas only 4 of them had a cytogenetically detectable deletion. 19 refs., 3 figs., 1 tab.

  19. FISH and FICTION to detect chromosomal aberrations in lymphomas.

    Science.gov (United States)

    Giefing, Maciej; Siebert, Reiner

    2013-01-01

    Fluorescence In Situ Hybridization (FISH) is a powerful and robust technique allowing the visualization of target sequences like genes in interphase nuclei. It is widely used in routine diagnostics to identify cancer specific aberrations including lymphoma associated translocations or gene copy number changes in single tumor cells. By combining FISH with immunophenotyping-a technique called Fluorescence Immunophenotyping and Interphase Cytogenetic as a Tool for Investigation Of Neoplasia (FICTION)-it is moreover possible to identify a cell population of interest. Here we describe standard protocols for FISH and FICTION as used in our laboratory in diagnosis and research.

  20. Inactivation of pectin methylesterase by immobilized trypsins from cunner fish and bovine pancreas.

    Science.gov (United States)

    Li, Dan; Matos, Madyu; Simpson, Benjamin K

    2013-01-01

    Immobilized cunner fish trypsin was used to inactivate pectin methylesterase (PME). The effects of different reaction conditions (e.g., incubation time, PME concentration, and temperature) on PME inactivation and kinetics of inactivation were investigated. Temperature, incubation time, and PME concentration significantly affected the extent of PME inactivation. Generally, higher temperature, longer incubation time, and low PME concentration caused more PME inactivation. The immobilized fish trypsin had higher capacity to inactivate PME than immobilized bovine trypsin. The inactivation efficiency of the immobilized fish trypsin was about 20% higher than that of its bovine counterpart. However, PME inactivated by both trypsins regained partial activity during storage at 4°C, with immobilized fish trypsin-treated PME regaining more of its original activity than the immobilized bovine trypsin-treated PME. Heat-denatured PME was hydrolyzed more extensively by immobilized fish trypsin than by its bovine counterpart. The rate constants increased, whereas the D-values decreased with temperature for both immobilized fish and bovine trypsins. The inactivation rate constants of immobilized fish trypsin at all the temperatures investigated (i.e., 15-35°C) were higher than those of immobilized bovine trypsin. Furthermore, the activation energy (Ea ) of PME inactivation by immobilized fish trypsin was lower than that of immobilized bovine trypsin.

  1. COmbined Binary RAtio fluorescence in situ hybridiziation (COBRA-FISH): development and applications.

    Science.gov (United States)

    Raap, A K; Tanke, H J

    2006-01-01

    The ability to probe for the location of DNA sequences in morphologically preserved chromosomes and nuclei by fluorescence in situ hybridization (FISH) provided for cytogenetics a quantum leap forward in resolution and ease of detection of chromosomal aberrations. COBRA-FISH, an acronym for COmbined Binary RAtio-FISH is a multicolor FISH methodology, which enables recognition of all human chromosome arms on the basis of color, thus greatly facilitating cytogenetic analysis. It also permits gene and viral integration site mapping in the context of chromosome arm painting. Here we review the principle, practice and applications of COBRA-FISH.

  2. Biological function evaluation and effects of laser micro-pore burn-denatured acellular dermal matrix.

    Science.gov (United States)

    Zhang, Youlai; Zeng, Yuanlin; Xin, Guohua; Zou, Lijin; Ding, Yuewei; Duyin, Jiang

    2017-08-18

    In the field of burns repairs, many problems exist in the shortage of donor skin, the expense of allograft or xenograft skin, temporary substitution and unsatisfactory extremity function after wound healing. Previous studies showed that burn-denatured skin could return to normal dermis formation and function. This study investigates the application of laser micro-pore burn-denatured acellular dermis matrix (DADM) from an escharotomy in the repair of burn wounds and evaluates the biological properties and wound repair effects of DADM in implantation experiments in Kunming mice. Specific-pathogen-free (SPF) Kunming mice were used in this study. A deep II° burn wound was created on the dorsum of the mice by an electric heated water bath. The full-thickness wound tissue was harvested. The necrotic tissue and subcutaneous tissue were removed. The denatured dermis was preserved and treated with 0.25% trypsin, 0.5% Triton X-100. The DADM was drilled by laser micro-pore. The biological properties and grafting effects of laser micro-pore burn-DADM were evaluated by morphology, cytokine expression levels and subcutaneous implantation experiments in Kunming mice. We found statistical significance (Plaser micro-pore burn-DADM (experimental group) compared to the control group (no laser micro-pore burn-DADM). Cytokine expression level was different in the dermal matrixes harvested at various time points after burn (24h, 48h, 72h and infected wound group). Comparing the dermal matrix from 24h burn tissue to infected wound tissue, the expression level of IL-6, MMP-24, VE-cadherin and VEGF were decreased. We found no inflammatory cells infiltration in the dermal matrix were observed in both experimental and control groups (24h burn group), while the obviously vascular infiltration and fiber fusion were observed in the experimental group after subcutaneous implantation experiments. There was better bio-performance, low immunogenicity and better dermal incorporation after treated

  3. DNA breakage detection-fish (DBD-FISH): effect of unwinding time.

    Science.gov (United States)

    Vázquez-Gundín, F; Gosálvez, J; de la Torre, J; Fernández, J L

    2000-09-20

    DBD-FISH is a new procedure that allows detection and quantification of DNA breakage in situ within specific DNA target sites. Cells embedded in an agarose matrix on a slide are treated in an alkaline unwinding solution to transform DNA breaks into single-stranded DNA (ssDNA). After removal of proteins, DNA probes are hybridized and detected. DNA breaks increase the ssDNA and relax supercoiling of DNA loops, so more probe hybridizes, thereby increasing the surface area and fluorescence intensity of the FISH signal. The probe selects the chromatin area to be analysed. In order to restrict the extension of unwound ssDNA to a region closer to the origin of the DNA break, human leukocytes were processed for DBD-FISH with a whole genome probe, after a 10 Gy dose of X-rays, for various unwinding times: 5, 2 min and 30s. Two cell populations were detected after 30s, but not with the 5 or 2 min unwinding times. One cell group had small to medium haloes corresponding to the relaxation of DNA supercoiling after DAPI staining, and strong DBD-FISH labelling of induced DNA breaks, whereas the other cell group showed big haloes of DNA loop unfolding and an absence of DBD-FISH labelling. The latter group was similar to cells processed by DBD-FISH without the unwinding step. Thus, they should correspond to cells unaffected by the alkaline unwinding solution, possibly because very brief unwinding times do not allow the diffusion of the alkali into the cells deep within the gel, thus biasing the results. Taking this into account, 2 min seems to be the minimum unwinding time required for an accurate detection of a signal by DBD-FISH.

  4. Anisakid nematodes as possible markers to trace fish products

    Directory of Open Access Journals (Sweden)

    Vincenzo Ferrantelli

    2015-03-01

    Full Text Available In this work a total of 949 fish samples were analysed for the identification of nematode larvae belonging to the Anisakidae family. Biomolecular application for the identification of Anisakidae larvae can be an optimal instrument for the traceability of fish products, described on the Reg. EC 178/2002. Results confirm a correlation between geographical distribution of fishes and presence of specific Anisakid larvae. FAO 37 zone (Mediterranean sea showed a prevailing distribution of Anisakis pegreffii and a minimal presence of A. simplex s.s. in hybrid form with Anisakis pegreffii. FAO 27 zone showed a prevailing distribution of A. simplex s.s. in fish like Brosme (Brosme brosme and infestation prevalence of Pseudoterranova krabbei and P. decipiens s.s. in Gadus morhua. Obtained results validate the hypothesis that molecular biology methods for identifying Anisakidae larvae are effective traceability markers of fish products.

  5. FISH applications for genomics and plant breeding strategies in tomato and other Solanaceous crops

    NARCIS (Netherlands)

    Szinay, D.; Bai, Y.; Visser, R.G.F.; Jong, de J.H.

    2010-01-01

    This paper describes the use of advanced fluorescence in situ hybridization (FISH) technologies for genomics and breeding of tomato and related Solanum species. The first part deals with the major determinants of FISH technology: (1) spatial resolution, which depends on the diffraction limit of the

  6. Molecular Cytogenetic Analysis of Spontaneous Interspecific Hybrid Between Oryza sativa and Oryza minuta

    Directory of Open Access Journals (Sweden)

    Chuan-deng YI

    2008-12-01

    Full Text Available Genomic in situ hybridization (GISH is a powerful tool to characterize parental chromosomes in interspecific hybrids, including the behaviour of autosynapsis and chromosome pairing. It was used to distinguish the chromosomes of Oryza sativa from wild species in a spontaneous interspecific hybrid and to investigate the chromosome pairing at metaphase I in meiosis of the hybrid in this study. The hybrid was a triploid with 36 chromosomes according to the chromosome number investigated in mitosis of root tips. During metaphase I of meiosis in the hybrid, less chromosome pairing was observed and most of the chromosomes existed as univalent. Based on GISH and FISH (Fluorescent in situ hybridization analyses, the chromosomes of the hybrid were composed of genomes A, B and C. Thus, it was believed that the hybrid was the result of natural hybridization between cultivated rice and wild species O. minuta which was planted in experimental fields.

  7. Light flux density threshold at which protein denaturation is induced by synchrotron radiation circular dichroism beamlines.

    Science.gov (United States)

    Miles, A J; Janes, Robert W; Brown, A; Clarke, D T; Sutherland, J C; Tao, Y; Wallace, B A; Hoffmann, S V

    2008-07-01

    New high-flux synchrotron radiation circular dichroism (SRCD) beamlines are providing important information for structural biology, but can potentially cause denaturation of the protein samples under investigation. This effect has been studied at the new CD1 dedicated SRCD beamline at ISA in Denmark, where radiation-induced thermal damage effects were observed, depending not only on the radiation flux but also on the focal spot size of the light. Comparisons with similar studies at other SRCD facilities worldwide has lead to the estimation of a flux density threshold under which SRCD beamlines should be operated when samples are to be exposed to low-wavelength vacuum ultraviolet radiation for extended periods of time.

  8. Fuel utilization improvement in PWRs using the denatured /sup 233/U-Th cycle

    Energy Technology Data Exchange (ETDEWEB)

    Jones, H.M.; Schwenk, G.A.; Toops, E.C.; Yotinen, V.O.

    1980-06-01

    A number of changes in PWR core design and/or operating strategy were evaluated to assess the fuel utilization improvement achievable by their implementation in a PWR using thorium-based fuel and operating in a recycle mode. The reference PWR for this study was identical to the B and W Standard Plant except that the fuel pellets were of denatured (/sup 233/U//sup 238/U-Th)O/sub 2/. An initial scoping study identified the three most promising improvement concepts as (1) a very tight lattice, (2) thorium blankets, and (3) ThO/sub 2/ rods placed in available guide tubes. A conceptual core design incorporating these changes was then developed, and the fuel utilization of this modified design was compared with that of the reference case.

  9. Denatured G-protein coupled receptors as immunogens to generate highly specific antibodies.

    Science.gov (United States)

    Talmont, Franck; Moulédous, Lionel; Boué, Jérôme; Mollereau, Catherine; Dietrich, Gilles

    2012-01-01

    G-protein coupled receptors (GPCRs) play a major role in a number of physiological and pathological processes. Thus, GPCRs have become the most frequent targets for development of new therapeutic drugs. In this context, the availability of highly specific antibodies may be decisive to obtain reliable findings on localization, function and medical relevance of GPCRs. However, the rapid and easy generation of highly selective anti-GPCR antibodies is still a challenge. Herein, we report that highly specific antibodies suitable for detection of GPCRs in native and unfolded forms can be elicited by immunizing animals against purified full length denatured recombinant GPCRs. Contrasting with the currently admitted postulate, our study shows that an active and well-folded GPCR is not required for the production of specific anti-GPCR antibodies. This new immunizing strategy validated with three different human GPCR (μ-opioid, κ-opioid, neuropeptide FF2 receptors) might be generalized to other members of the GPCR family.

  10. New Primers for Denaturing Gradient Gel Electrophoresis Analysis of Nitrate-Reducing Bacterial Community in Soil

    Institute of Scientific and Technical Information of China (English)

    R.PASTORELLI; R.PICCOLO; S.SIMONCINI; S.LANDI

    2013-01-01

    The narG gene is frequently used as a molecular marker for bacterial nitrate-reducing community analysis.In this study,a new set of primers targeting the narG gene was designed and applied to semi-nested polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) assay.The potential of the new primers was verified on DNA directly extracted from soils from five different experimental sites distributed in Central and Southern Italy.Specificity of the primers was determined by excision,amplification,and sequencing of bands resolved by DGGE.A phylogenetic analysis showed the correlation between the sequences retrieved from the soils studied and the narG sequences from β and γ-Proteobacteria.These primers expanded the existing molecular tools for ecological study on the size and diversity of nitrate-reducing bacterial community in soil.

  11. An approach for protein to be completely reversible to thermal denaturation even at autoclave temperatures.

    Science.gov (United States)

    Iwakura, M; Nakamura, D; Takenawa, T; Mitsuishi, Y

    2001-08-01

    Reversibility of protein denaturation is a prerequisite for all applications that depend on reliable enzyme catalysis, particularly, for using steam to sterilize enzyme reactors or enzyme sensor tips, and for developing protein-based devices that perform on-off switching of the protein function such as enzymatic activity, ligand binding and so on. In this study, we have successfully constructed an immobilized protein that retains full enzymatic activity even after thermal treatments as high as 120 degrees C. The key for the complete reversibility was the development of a new reaction that allowed a protein to be covalently attached to a surface through its C-terminus and the protein engineering approach that was used to make the protein compatible with the new attachment chemistry.

  12. Fish allergy: in review.

    Science.gov (United States)

    Sharp, Michael F; Lopata, Andreas L

    2014-06-01

    Globally, the rising consumption of fish and its derivatives, due to its nutritional value and divergence of international cuisines, has led to an increase in reports of adverse reactions to fish. Reactions to fish are not only mediated by the immune system causing allergies, but are often caused by various toxins and parasites including ciguatera and Anisakis. Allergic reactions to fish can be serious and life threatening and children usually do not outgrow this type of food allergy. The route of exposure is not only restricted to ingestion but include manual handling and inhalation of cooking vapors in the domestic and occupational environment. Prevalence rates of self-reported fish allergy range from 0.2 to 2.29 % in the general population, but can reach up to 8 % among fish processing workers. Fish allergy seems to vary with geographical eating habits, type of fish processing, and fish species exposure. The major fish allergen characterized is parvalbumin in addition to several less well-known allergens. This contemporary review discusses interesting and new findings in the area of fish allergy including demographics, novel allergens identified, immunological mechanisms of sensitization, and innovative approaches in diagnosing and managing this life-long disease.

  13. Do Fish Resist?

    Directory of Open Access Journals (Sweden)

    Dinesh Joseph Wadiwel

    2016-03-01

    Full Text Available There have been a number of scientific studies on the question of whether fish feel pain. Some have suggested that some fish indeed do feel pain and that this has significant welfare implications (2003. Others have argued that fish do not have the brain development necessary to feel pain. In terms of number of animals killed, the slaughter of sea animals for human consumption significantly exceeds that of any land animals that we use for food, and sea animal slaughter practices frequently lack any basic welfare protections. If fish can be shown to feel pain—or more importantly, if humans can agree that fish feel pain—then this would place a significant question mark over many contemporary fishing practices.  This article substitutes the question 'Do Fish Feel Pain?' with an alternative: 'Do Fish Resist?' It explores the conceptual problems of understanding fish resistance, and the politics of epistemology that surrounds and seeks to develop a conceptual framework for understanding fish resistance to human capture by exploring the development of fishing technologies - the hook, the net and contemporary aquaculture.

  14. Fish under exercise.

    Science.gov (United States)

    Palstra, Arjan P; Planas, Josep V

    2011-06-01

    Improved knowledge on the swimming physiology of fish and its application to fisheries science and aquaculture (i.e., farming a fitter fish) is currently needed in the face of global environmental changes, high fishing pressures, increased aquaculture production as well as increased concern on fish well-being. Here, we review existing data on teleost fish that indicate that sustained exercise at optimal speeds enhances muscle growth and has consequences for flesh quality. Potential added benefits of sustained exercise may be delay of ovarian development and stimulation of immune status. Exercise could represent a natural, noninvasive, and economical approach to improve growth, flesh quality as well as welfare of aquacultured fish: a FitFish for a healthy consumer. All these issues are important for setting directions for policy decisions and future studies in this area. For this purpose, the FitFish workshop on the Swimming Physiology of Fish ( http://www.ub.edu/fitfish2010 ) was organized to bring together a multidisciplinary group of scientists using exercise models, industrial partners, and policy makers. Sixteen international experts from Europe, North America, and Japan were invited to present their work and view on migration of fishes in their natural environment, beneficial effects of exercise, and applications for sustainable aquaculture. Eighty-eight participants from 19 different countries contributed through a poster session and round table discussion. Eight papers from invited speakers at the workshop have been contributed to this special issue on The Swimming Physiology of Fish.

  15. Development and characterization of interspecific hybrids between Oryza sativa and O. latifolia by in situ hybridization

    Institute of Scientific and Technical Information of China (English)

    YI ChuanDeng; TANG ShuZhu; ZHOU Yong; LIANG GuoHua; GONG ZhiYun; GU MingHong

    2008-01-01

    Oryza sativa and O. latifolia belong to the AA and CCDD genomes of Oryza, respectively. In this study, interspecific hybrids of these species were obtained using the embryo rescue technique. Hybrid pani-cle traits, such as long awns, small grain, exoteric large purple stigma, grain shattering and dispersed panicles, resemble that of the paternal parent, O. latifolia, whereas there is obvious heterosis in such respects as plant height, tillering ability and vegetative vigor. Chromosome pairing and the genomic components of the hybrid were subsequently investigated using genomic in situ hybridization (GISH) and fluorescent in situ hybridization (FISH) analysis. Based on the mitotic metaphase chromosome numbers of the root tips investigated, the hybrid is a triploid with 36 chromosomes. The genomic con-stitution of the hybrid is ACD. In the meiotic metaphase I of the hybrid pollen mother cell, poor chro-mosome pairing was identified and most of the chromosomes were univalent, which resulted in com-plete male sterility in the hybrid.

  16. Zoonoses associated with fish.

    Science.gov (United States)

    Boylan, Shane

    2011-09-01

    The taxonomic group that composes the fishes is the most diverse group of vertebrates worldwide. The challenges of unique physiologies, a foreign environment, and many unknowns attract a passionate group of biologists and veterinarians. Economically, fishes have become vital as food, bait, and companion animals. Fishermen and fish handlers (processing plants) represent the historical human population exposed to fish zoonoses, but growth in aquaculture and aquarium hobbyists have led to an increase in published fish-borne zoonotic cases starting in the late 1950s that bloomed in the 1980s. Human physicians, particularly dermatologists and infectious disease specialists, are now more aware of fish-borne zoonoses, but they can be assisted with diagnosis when informed patients give more detailed histories with fish/water exposure.

  17. Effect of freezing and thawing rates on denaturation of proteins in aqueous solutions.

    Science.gov (United States)

    Cao, Enhong; Chen, Yahuei; Cui, Zhanfeng; Foster, Peter R

    2003-06-20

    The freeze denaturation of model proteins, LDH, ADH, and catalase, was investigated in absence of cryoprotectants using a microcryostage under well-controlled freezing and thawing rates. Most of the experimental data were obtained from a study using a dilute solution with an enzyme concentration of 0.025 g/l. The dependence of activity recovery of proteins on the freezing and thawing rates showed a reciprocal and independent effect, that is, slow freezing (at a freezing rate about 1 degrees C/min) and fast thawing (at a thawing rate >10 degrees C/min) produced higher activity recovery, whereas fast freezing with slow thawing resulted in more severe damage to proteins. With minimizing the freezing concentration and pH change of buffer solution by using a potassium phosphate buffer, this phenomenon could be ascribed to surface-induced denaturation during freezing and thawing process. Upon the fast freezing (e.g., when the freezing rate >20 degrees C/min), small ice crystals and a relatively large surface area of ice-liquid interface are formed, which increases the exposure of protein molecules to the ice-liquid interface and hence increases the damage to the proteins. During thawing, additional damage to proteins is caused by recrystallization process. Recrystallization exerts additional interfacial tension or shear on the entrapped proteins and hence causes additional damage to the latter. When buffer solutes participated during freezing, the activity recovery of proteins after freezing and thawing decreased due to the change of buffer solution pH during freezing. However, the patterns of the dependence on freezing and thawing rates of activity recovery did not change except for that at extreme low freezing rates (solutions could be reduced by changing the buffer type and composition and by optimizing the freezing-thawing protocol.

  18. Microbial Community Structure of Korean Cabbage Kimchi and Ingredients with Denaturing Gradient Gel Electrophoresis.

    Science.gov (United States)

    Hong, Sung Wook; Choi, Yun-Jeong; Lee, Hae-Won; Yang, Ji-Hee; Lee, Mi-Ai

    2016-06-28

    Kimchi is a traditional Korean fermented vegetable food, the production of which involves brining of Korean cabbage, blending with various other ingredients (red pepper powder, garlic, ginger, salt-pickled seafood, etc.), and fermentation. Recently, kimchi has also become popular in the Western world because of its unique taste and beneficial properties such as antioxidant and antimutagenic activities, which are derived from the various raw materials and secondary metabolites of the fermentative microorganisms used during production. Despite these useful activities, analysis of the microbial community present in kimchi has received relatively little attention. The objective of this study was to evaluate the bacterial community structure from the raw materials, additives, and final kimchi product using the culture-independent method. Specifically, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) was used to analyze the 16S rRNA partial sequences of the microflora. One primer set for bacteria, 341F(GC)-518R, reliably produced amplicons from kimchi and its raw materials, and these bands were clearly separated on a 35-65% denaturing gradient gel. Overall, 117 16S rRNA fragments were identified by PCR-DGGE analysis. Pediococcus pentosaceus, Leuconostoc citreum, Leuconostoc gelidum, and Leuconostoc mesenteroides were the dominant bacteria in kimchi. The other strains identified were Tetragenococcus, Pseudomonas, Weissella, and uncultured bacterium. Comprehensive analysis of these microorganisms could provide a more detailed understanding of the biologically active components of kimchi and help improve its quality. PCR-DGGE analysis can be successfully applied to a fermented food to detect unculturable or other species.

  19. Size is a major determinant of dissociation and denaturation behaviour of reconstituted high-density lipoproteins.

    Science.gov (United States)

    Gianazza, Elisabetta; Eberini, Ivano; Sirtori, Cesare R; Franceschini, Guido; Calabresi, Laura

    2002-08-15

    Lipid-free apolipoprotein A-I (apoA-I) and A-I(Milano) (A-I(M)) were compared for their denaturation behaviour by running across transverse gradients of a chaotrope, urea, and of a ionic detergent, SDS. For both apo A-I and monomeric apoA-I(M) in the presence of increasing concentrations of urea the transition from high to low mobility had a sigmoidal course, whereas for dimeric A-I(M)/A-I(M) a non-sigmoidal shape was observed. The co-operativity of the unfolding process was lower for dimeric A-I(M)/A-I(M) than for apoA-I or for monomeric apoA-I(M). A slightly higher susceptibility to denaturation was observed for dimeric A-I(M)/A-I(M) than for monomeric apoA-I(M). A similar behaviour of A-I(M)/A-IM versus apoA-I(M) was observed in CD experiments. Large- (12.7/12.5 nm) and small- (7.8 nm) sized reconstituted high-density lipoproteins (rHDL) containing either apoA-I or A-I(M)/A-I(M) were compared with respect to their protein-lipid dissociation behaviour by subjecting them to electrophoresis in the presence of urea, of SDS and of a non-ionic detergent, Nonidet P40. A higher susceptibility to dissociation of small-sized versus large-sized rHDL, regardless of the apolipoprotein component, was observed in all three instances. Our data demonstrate that the differential plasticity of the various classes of rHDL is a function of their size; the higher stability of 12.5/12.7 nm rHDL is likely connected to the higher number of protein-lipid and lipid-lipid interactions in larger as compared with smaller rHDL.

  20. Thermal, chemical and pH induced unfolding of turmeric root lectin: modes of denaturation.

    Directory of Open Access Journals (Sweden)

    Himadri Biswas

    Full Text Available Curcuma longa rhizome lectin, of non-seed origin having antifungal, antibacterial and α-glucosidase inhibitory activities, forms a homodimer with high thermal stability as well as acid tolerance. Size exclusion chromatography and dynamic light scattering show it to be a dimer at pH 7, but it converts to a monomer near pH 2. Circular dichroism spectra and fluorescence emission maxima are virtually indistinguishable from pH 7 to 2, indicating secondary and tertiary structures remain the same in dimer and monomer within experimental error. The tryptophan environment as probed by acrylamide quenching data yielded very similar data at pH 2 and pH 7, implying very similar folding for monomer and dimer. Differential scanning calorimetry shows a transition at 350.3 K for dimer and at 327.0 K for monomer. Thermal unfolding and chemical unfolding induced by guanidinium chloride for dimer are both reversible and can be described by two-state models. The temperatures and the denaturant concentrations at which one-half of the protein molecules are unfolded, are protein concentration-dependent for dimer but protein concentration-independent for monomer. The free energy of unfolding at 298 K was found to be 5.23 Kcal mol-1 and 14.90 Kcal mol-1 for the monomer and dimer respectively. The value of change in excess heat capacity upon protein denaturation (ΔCp is 3.42 Kcal mol-1 K-1 for dimer. The small ΔCp for unfolding of CLA reflects a buried hydrophobic core in the folded dimeric protein. These unfolding experiments, temperature dependent circular dichroism and dynamic light scattering for the dimer at pH 7 indicate its higher stability than for the monomer at pH 2. This difference in stability of dimeric and monomeric forms highlights the contribution of inter-subunit interactions in the former.

  1. Aqueous Solutions of the Ionic Liquid 1-butyl-3-methylimidazolium Chloride Denature Proteins

    Energy Technology Data Exchange (ETDEWEB)

    Baker, Gary A [ORNL; Heller, William T [ORNL

    2009-01-01

    As we advance our understanding, ionic liquids (ILs) are finding ever broader scope within the chemical sciences including, most recently, pharmaceutical, enzymatic, and bioanalytical applications. With examples of enzymatic activity reported in both neat ILs and in IL/water mixtures, enzymes are frequently assumed to adopt a quasi-native conformation, even if little work has been carried out to date toward characterizing the conformation, dynamics, active-site perturbation, cooperativity of unfolding transitions, free energy of stabilization, or aggregation/oligomerization state of enzymes in the presence of an IL solvent component. In this study, human serum albumin and equine heart cytochrome c were characterized in aqueous solutions of the fully water-miscible IL 1-butyl-3-methylimidazolium chloride, [bmim]Cl, by small-angle neutron and X-ray scattering. At [bmim]Cl concentrations up to 25 vol.%, these two proteins were found to largely retain their higher-order structures whereas both proteins become highly denatured at the highest IL concentration studied here (i.e., 50 vol.% [bmim]Cl). The response of these proteins to [bmim]Cl is analogous to their behavior in the widely studied denaturants guanidine hydrochloride and urea which similarly lead to random coil conformations at excessive molar concentrations. Interestingly, human serum albumin dimerizes in response to [bmim]Cl, whereas cytochrome c remains predominantly in monomeric form. These results have important implications for enzymatic studies in aqueous IL media, as they suggest a facile pathway through which biocatalytic activity can be altered in these nascent and potentially green electrolyte systems.

  2. Hybrid microelectronic technology

    Science.gov (United States)

    Moran, P.

    Various areas of hybrid microelectronic technology are discussed. The topics addressed include: basic thick film processing, thick film pastes and substrates, add-on components and attachment methods, thin film processing, and design of thick film hybrid circuits. Also considered are: packaging hybrid circuits, automating the production of hybrid circuits, application of hybrid techniques, customer's view of hybrid technology, and quality control and assurance in hybrid circuit production.

  3. Genome analysis of 7 Kengyilia (Triticeae Poaceae) species with FISH and GISH

    Science.gov (United States)

    Genome composition of and genetic relationships among seven Kengyilia species were assessed using a technique of sequential FISH (fluorescence in situ hybridization) and GISH (genomic in situ hybridization). Five of these 7 species, K. kokonorica, K. rigidula, K. hirsula, K. grandiglumis, and K. th...

  4. Southeast Alaska ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains biological resource data for estuarine, benthic, and pelagic fish in Southeast Alaska. Vector polygons in this data set represent locations of...

  5. Columbia River ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains sensitive biological resource data for marine, estuarine, anadromous, and freshwater fish species in Columbia River. Vector polygons in this...

  6. North Slope, Alaska ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains sensitive biological resource data for marine, estuarine, anadromous, and freshwater fish species for the North Slope of Alaska. Vector...

  7. American Samoa ESI: FISH (Fish Polygons)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains sensitive biological resource data for reef, pelagic, benthic, and estuarine fish species in American Samoa. Vector polygons in this data set...

  8. 雌核发育二倍体鲫鲤及其他倍性鱼cdc2基因cDNA全序列克隆及表达%The cloning of cdc2 cDNAs and a comparative study of its expression in different ploidy fishes including the diploid gynogenetic hybrid of red crucian carp × common carp

    Institute of Scientific and Technical Information of China (English)

    陶敏; 刘少军; 钟欢; 周毅; 宋灿; 张纯; 刘筠

    2013-01-01

    Cdc2(Cyclin Dependent Kinase,namely CDK1)encoded by cdc2 gene and CyclinB combination regulates G2/M transition. To find out molecular mechanism that diploid hybrid fish could produce diploid gamete,the full length cDNAs of cdcl in the third gynogenetic generation(G3) ,red crucian carp( Carassius auratus red var. ) ,triploid crucian carp and allotetraploid were obtained by PCR and rapid amplification of cDNA ends. Our data showed that all the cDNAs of cdcl gene in the four different ploidy fishes encode a protein of 302 amino acids containing a domain (PSTAVRE) which combine Cyclins. A high homology of 97. 6% of the Cdc2 protein can be drawn by comparing the amino acid sequences in these four fishes, which indicates the higher conservative function and evolution of Cdc2 protein in these four fishes. A comparative expression pattern of cdcl in early-stage gonads of G3 and different ploidy fishes was carried out by Realtime PCR using specific primers against the same sequences of coding regions in the four fishes. The results showed that the expression of cdcl in the ovary of G3 was higher than those of red crucian carp and triploid crucian carp, while lower than that of allotetraploid, which, at the molecular level, indicates existence of polyploid oogonia in early-stage gonads of G3. The higher expression of cdcl in G3 suggests that consecutive S-phase replication may occur without intervening mitosis, which might be related to the formation mechanisms for the diploid eggs generated by diploid hybrids.%为研究雌核发育二倍体鲫鲤产生二倍体卵子的分子机制,实验采用PCR和cDNA末端快速分离法,克隆获得了雌核发育二倍体鲫鲤第三代(G3)、二倍体红鲫、三倍体湘云鲫和四倍体鲫鲤的细胞周期相关基因——cdc2基因cDNA全序列.结果显示,4种不同倍性鱼cdc2基因均编码含有302个氨基酸蛋白,而且编码的蛋白都含有与其他CDK激酶相当保守的序列PSTAVRE;同源性分析发现,4

  9. Sugar Cane Genome Numbers Assumption by Ribosomal DNA FISH Techniques

    NARCIS (Netherlands)

    Thumjamras, S.; Jong, de H.; Iamtham, S.; Prammanee, S.

    2013-01-01

    Conventional cytological method is limited for polyploidy plant genome study, especially sugar cane chromosomes that show unstable numbers of each cultivar. Molecular cytogenetic as fluorescent in situ hybridization (FISH) techniques were used in this study. A basic chromosome number of sugar cane

  10. Fish elevator and method of elevating fish

    Science.gov (United States)

    Truebe, Jonathan; Drooker, Michael S.

    1984-01-01

    A means and method for transporting fish from a lower body of water to a higher body of water. The means comprises a tubular lock with a gated entrance below the level of the lower body of water through which fish may enter the lock and a discharge passage above the level of the upper body of water. The fish raising means in the lock is a crowder pulled upward by a surface float as water from the upper body of water gravitationally flows into the closed lock filling it to the level of the upper body. Water is then pumped into the lock to raise the level to the discharge passage. The crowder is then caused to float upward the remaining distance through the water to the level of the discharge passage by the introduction of air into a pocket on the underside of the crowder. The fish are then automatically discharged from the lock into the discharge passage by the out of water position of the crowder. The movement of the fish into the discharge passage is aided by the continuous overflow of water still being pumped into the lock. A pipe may be connected to the discharge passage to deliver the fish to a selected location in the upper body of water.

  11. Comprehensive TP53-denaturing gradient gel electrophoresis mutation detection assay also applicable to archival paraffin-embedded tissue

    NARCIS (Netherlands)

    Hayes, V M; Bleeker, W; Verlind, E; Timmer, T; Karrenbeld, A; Plukker, J T; Marx, M P; Hofstra, R M; Buys, C H

    1999-01-01

    A comprehensive mutation detection assay is described for the entire coding region and all splice site junctions of TP53. The assay is based on denaturing gradient gel electrophoresis, which follows either multiplex polymerase chain reaction (PCR) applied to DNA extracted from fresh or frozen tissue

  12. Denaturation and in Vitro Gastric Digestion of Heat-Treated Quinoa Protein Isolates Obtained at Various Extraction pH

    NARCIS (Netherlands)

    Ruiz, Geraldine Avila; Opazo-Navarrete, Mauricio; Meurs, Marlon; Minor, Marcel; Sala, Guido; Boekel, van Martinus; Stieger, Markus; Janssen, Anja E.M.

    2016-01-01

    The aim of this study was to determine the influence of heat processing on denaturation and digestibility properties of protein isolates obtained from sweet quinoa (Chenopodium quinoa Willd) at various extraction pH values (8, 9, 10 and 11). Pretreatment of suspensions of protein isolates at 60,

  13. Heat-induced denaturation and aggregation of ovalbumin at neutral pH described by irreversible first-order kinetics

    NARCIS (Netherlands)

    Weijers, M.; Barneveld, P.A.; Cohen Stuart, M.A.; Visschers, R.W.

    2003-01-01

    The heat-induced denaturation kinetics of two different sources of ovalbumin at pH 7 was studied by chromatography and differential scanning calorimetry. The kinetics was found to be independent of protein concentration and salt concentration, but was strongly dependent on temperature. For highly

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

    DEFF Research Database (Denmark)

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

    2005-01-01

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

  15. Denaturation and in Vitro Gastric Digestion of Heat-Treated Quinoa Protein Isolates Obtained at Various Extraction pH

    NARCIS (Netherlands)

    Ruiz, Geraldine Avila; Opazo-Navarrete, Mauricio; Meurs, Marlon; Minor, Marcel; Sala, Guido; Boekel, van Tiny; Stieger, Markus; Janssen, Anja E.M.

    2016-01-01

    The aim of this study was to determine the influence of heat processing on denaturation and digestibility properties of protein isolates obtained from sweet quinoa (Chenopodium quinoa Willd) at various extraction pH values (8, 9, 10 and 11). Pretreatment of suspensions of protein isolates at 60,

  16. Reaction of Native and Denatured Brucella abortus (S19 Proteins with Antibody Using Affinity Chromatography and Immunoblotting

    Directory of Open Access Journals (Sweden)

    R. Karimi

    2005-01-01

    Full Text Available Western blotting or immunoblotting commonly use for study of reaction between antigens and antibodies. Denaturation of many proteins in immunoblotting can affect greatly the reactivity of antibodies and outcome of the procedure.In this study proteins of Brucella abortus (S19 was extracted by a mild method and reaction of the extracted proteins with serum of infected human and goat and immunized rabbit compared by affinity chromatography and immunoblotting. Gamma globulin (mostly IgG fraction of the sera was precipitated by half saturation of ammonium sulfate and linked to activated sepharose 4B. The extracted proteins were loaded on the affinity column. Attached proteins was eluted by low pH and analyzed by SDS-PAGE. Reaction of the total extract and eluted fractions with IgG fraction of sera was evaluated by Western blotting.Upon the results of affinity chromatography and immunoblotting, Brucella proteins can be classified in four groups: 1- The proteins that adsorbed to the affinity column and react with IgG in westernblotting. 2- Proteins that react with IgG in native state but no in denatured state. 3- Proteins that do not react with IgG in native state but react in denatured state. 4- Proteins that do not react with IgG in native and denatured state.

  17. High-intensity focused ultrasound monitoring using harmonic motion imaging for focused ultrasound (HMIFU) under boiling or slow denaturation conditions.

    Science.gov (United States)

    Hou, Gary Y; Marquet, Fabrice; Wang, Shutao; Apostolakis, Iason-Zacharias; Konofagou, Elisa E

    2015-07-01

    Harmonic motion imaging for focused ultrasound (HMIFU) is a recently developed high-intensity focused ultrasound (HIFU) treatment monitoring method that utilizes an amplitude-modulated therapeutic ultrasound beam to induce an oscillatory radiation force at the HIFU focus and estimates the focal tissue displacement to monitor the HIFU thermal treatment. In this study, the performance of HMIFU under acoustic, thermal, and mechanical effects was investigated. The performance of HMIFU was assessed in ex vivo canine liver specimens (n = 13) under slow denaturation or boiling regimes. A passive cavitation detector (PCD) was used to assess the acoustic cavitation activity, and a bare-wire thermocouple was used to monitor the focal temperature change. During lesioning with slow denaturation, high quality displacements (correlation coefficient above 0.97) were observed under minimum cavitation noise, indicating the tissue initial-softening-then- stiffening property change. During HIFU with boiling, HMIFU monitored a consistent change in lesion-to-background displacement contrast (0.46 ± 0.37) despite the presence of strong cavitation noise due to boiling during lesion formation. Therefore, HMIFU effectively monitored softening-then-stiffening during lesioning under slow denaturation, and detected lesioning under boiling with a distinct change in displacement contrast under boiling in the presence of cavitation. In conclusion, HMIFU was shown under both boiling and slow denaturation regimes to be effective in HIFU monitoring and lesioning identification without being significantly affected by cavitation noise.

  18. Evaluation of several techniques to modify denatured muscle tissue to obtain a scaffold for peripheral nerve regeneration

    NARCIS (Netherlands)

    Meek, MF; den Dunnen, WFA; Schakenraad, JM; Robinson, PH

    The aim of this study was to (1) evaluate the effect of several preparation techniques of denatured muscle tissue to obtain an open three-dimensional structure, and (2) test if this scaffold is suitable for peripheral nerve regeneration. Four samples (A-D) of muscle tissue specimens were evaluated

  19. The unfolding/denaturation of immunogammaglobulin of isotype 2b and its F-ab and F-c fragments

    NARCIS (Netherlands)

    Vermeer, AWP; Norde, W; van Amerongen, A

    2000-01-01

    The unfolding and further denaturation of IgG and its F-ab and F-c fragments were studied both on a macroscopic and molecular level, using differential scanning calorimetry and circular dichroism spectroscopy, respectively. It was shown that the structural integrity of the F-ab and F-c units was ret

  20. Sequential Events in the Irreversible Thermal Denaturation of Human Brain-Type Creatine Kinase by Spectroscopic Methods

    Directory of Open Access Journals (Sweden)

    Yan-Song Gao

    2010-06-01

    Full Text Available The non-cooperative or sequential events which occur during protein thermal denaturation are closely correlated with protein folding, stability, and physiological functions. In this research, the sequential events of human brain-type creatine kinase (hBBCK thermal denaturation were studied by differential scanning calorimetry (DSC, CD, and intrinsic fluorescence spectroscopy. DSC experiments revealed that the thermal denaturation of hBBCK was calorimetrically irreversible. The existence of several endothermic peaks suggested that the denaturation involved stepwise conformational changes, which were further verified by the discrepancy in the transition curves obtained from various spectroscopic probes. During heating, the disruption of the active site structure occurred prior to the secondary and tertiary structural changes. The thermal unfolding and aggregation of hBBCK was found to occur through sequential events. This is quite different from that of muscle-type CK (MMCK. The results herein suggest that BBCK and MMCK undergo quite dissimilar thermal unfolding pathways, although they are highly conserved in the primary and tertiary structures. A minor difference in structure might endow the isoenzymes dissimilar local stabilities in structure, which further contribute to isoenzyme-specific thermal stabilities.