Miracle-dz/UniFAR-FTUs huggingface dataset:
Analysis of the effects of spatial localisation of transgenes on expression of recombinant proteins in CHO-DG44 cells

Chinese hamster ovary cell lines are commonly used as host for production of recombinant protein both in research and in the biotech industry. Recombinant cell lines are generated through random integration of multi-cistronic plasmid vector containing the genes of interest and selection marker gene into the host genome. The recombinant cell lines require several rounds of limiting clonal dilution to isolate stable high expressing clones. Stable high expression of the genes of interest is a rare and desired trait in recombinant clonal cell lines. Despite being clonal, cell lines eventually become heterogeneous and lose productivity.The Mammalian genome resides as packaged chromatin in the nucleus, which is a highly organized structure with specialized spatial and functional compartments. Increasingly, evidence is pointing to the interaction between plasmid and nuclear architecture as a factor that affects expression and stability. This project is based on the hypothesis that certain locations within the three-dimensional structure of the nucleus are more favourable for stable high-level expression of recombinant genes, integration of transgene into these specific location will produce high expressing stable cell linesTo prove the hypothesis a set of far-red reporter gene expressing recombinant clonal cell lines were generated to use as model cell lines. These cell lines were characterized over 80 days of continuous culture to determine their expression and stability. Flow cytometry results showed that all cell lines showed heterogeneity and gradual loss of expression of the far-red gene. Majority of the cell lines loss 80% of their gene copy number by day 25 of continuous culture.The possibility of using telomeric repeat sequences and nuclear bodies as nuclear landmarks was explored in order to develop tools that can be used to study the localization and interactions of the integrated recombinant genes and the CHO genome in the nuclear environment. Telomeric repeat sequences were far too numerous and scattered to use as nuclear landmark. While PML nuclear bodies and Cajal bodies appeared to be randomly positioned.The possibility of cross-species chromosome painting using FACS sorted Chinese hamster chromosome as paints was investigated, results showed that two copies of chromosome 1 and a copy of chromosome 4 along with segments of other chromosomes 7 and X&Y are conserved in the CHO genome. This finding allowed us to further explore the localization of chromosomes in the interphase nuclei. Results showed that the CHO cell equivalent of hamster chromosome 4 does not have a preferred radial localization in the interphase nuclei. FISH data also showed that the site of reporter gene integration is variable and that the integrated plasmid does not have a preferred radial position within the nuclei. These findings indicated that the CHO nuclei in our cell lines do not have a common nuclear organization.
A New Halogenated Biindole and A New Apo-carotenone from Green Alga Chaetomorpha basiretorsa Setchell

A new halogenated biindole and a new apo-carotenone have been isolated from the ethanolic extract of the green alga Chaetomorpha basiretorsa Sethcell. On the basis of chemical and spectroscopic methods including 2D NMR technique, their structures have been elucidated as 4,4′-dichloro-5,5′-dibromo-7,7′-dimethoxy-2,2′-bi- 1H-indole and l ′S*,4′R*-8-(4′-hydroxy-2′,6′,6′-trimethylcyclohex-2-enyl)-6-methyloct-3E,5E,7E-trien-2-one, respectively.
High water roost selection by waders: maximizing feeding opportunities or avoiding predation?

During the highest spring tides the intertidal sediment flats of estuaries are fully inundated at high water, and waders have no choice but to move to supratidal roosts, e.g. on open farmland, saltpans or beaches. However, in many estuaries during the lowest neap or intermediate tides there are sectors of upper intertidal sediment flats that remain exposed even at the peak of high water, and so waders have the choice of roosting either there or in supratidal sites. In the Tagus Estuary, Portugal, as elsewhere, waders use both types of roosts during high water. Our main objective was to understand what makes waders opt for one of these two types of available roosts. We monitored wader use of saltpans and intertidal roosts from spring to neap tides, and measured foraging and alarm behaviour, prey availability and disturbance by predators. Most of the wader species studied chose intertidal (mudflat) roosts whenever these were available, and only roosted in saltpans during the peak of spring tides. We hypothesized that this preference was explained either by an attempt (i) to continue feeding into the high water period, or (ii) to minimize predation risk. Extending feeding time into the high water period did not seem to be very relevant for roost choice because both prey availability and foraging activity were low in both types of roosts. However, predator disturbance was several times higher in the saltpans than in the intertidal roosts, suggesting that this factor may be the determinant in the choice of roost type.
A Novel γ-Secretase Assay Based on Detection of the Putative C-terminal Fragment-γ of Amyloid β Protein Precursor*

Alzheimer's disease is characterized by the deposits of the 4-kDa amyloid β peptide (Aβ). The Aβ protein precursor (APP) is cleaved by β-secretase to generate a C-terminal fragment, CTFβ, which in turn is cleaved by γ-secretase to generate Aβ. Alternative cleavage of the APP by α-secretase at Aβ16/17 generates the C-terminal fragment, CTFα. In addition to Aβ, endoproteolytic cleavage of CTFα and CTFβ by γ-secretase should yield a C-terminal fragment of 57–59 residues (CTFγ). However, CTFγ has not yet been reported in either brain or cell lysates, presumably due to its instability in vivo. We detected thein vitro generation of Aβ as well as an ∼6-kDa fragment from guinea pig brain membranes. We have provided biochemical and pharmacological evidence that this 6-kDa fragment is the elusive CTFγ, and we describe an in vitro assay for γ-secretase activity. The fragment migrates with a synthetic peptide corresponding to the 57-residue CTFγ fragment. Three compounds previously identified as γ-secretase inhibitors, pepstatin-A, MG132, and a substrate-based difluoroketone (t-butoxycarbonyl-Val-Ile-(S)-4-amino-3-oxo-2,2-difluoropentanoyl-Val-Ile-OMe), reduced the yield of CTFγ, providing additional evidence that the fragment arises from γ-secretase cleavage. Consistent with reports that presenilins are the elusive γ-secretases, subcellular fractionation studies showed that presenilin-1, CTFα, and CTFβ are enriched in the CTFγ-generating fractions. The in vitroγ-secretase assay described here will be useful for the detailed characterization of the enzyme and to screen for γ-secretase inhibitors.
NUMERICAL ANALYSIS OF AXIALLY LOADED COLD-FORMED STEEL BUILT-UP BOX SECTION

Built up box section is a doubly symmetrical section in which two C-channel connected at the flange using self-drilling screw. They are generally used in the construction industry to obtain higher rigidity, when a single section is not sufficient. Despite being widely used in the construction industry, there are limited studies on built-up box section. Moreover, the current design code does not provide clear design guideline for the calculation of built-up box section. Therefore, research on the built-up box section is important. This paper presents a finite element model to predict the compressive strength and simulate the behavior of cold-formed steel built-up box section. The finite element model was developed using ABAQUS CAE/6.14. Comparison of the finite element and experimental results showed good correlation. The model well predicted the behavior of the built-up box column.
Self-organisation and evolution of social systems

Preface Introduction 1. From unicellular to multicellular organisation: the development of the social amoebae Dicyostelium discoideum C. Weijer 2. Selection and self-organisation of collective decision in social insects: trail, network and migration J. L. Deneubourg 3. The interplay of intracolonial genotype variance and self-organisation of dominance hierarchies in honeybees Crewe Moritz 4. Traffic rules of fish schools: a review of agent-based approaches J. K. Parrish and S. V. Viscido 5. A process-orientated approach to the social behaviour of primates C. K. Hemelrijk 6. Order and noise in primate societies B. Thierry 7. Self-organisation in language B. de Boer 8. Minority opinion spreading in democratic random debate S. Galam 9. Natural selection and complex systems: a complex interaction D. S. Wilson 10. Interlocking of self-organisation and evolution P. Hogeweg.
Engineering dioxygenases by laboratory evolution: a comparison of evolutionary search strategies

Due to the unique and difficult chemistry they perform, the aromatic ring-hydroxylating dioxygenases are of interest as industrial catalysts. Unfortunately, an application-specific array of problems limits their utility. To address these problems through laboratory evolution, I developed methods for high-throughput screening of tens of thousands of dioxygenase variants. These methods rely on a phenol detection reagent (Gibbs reagent) and can be applied to liquid cultures or to growing bacterial colonies expressing variant enzymes. Recombination of genes encoding homologous enzymes ("family shuffling") has emerged as a promising tool for evolutionary protein engineering. Using the dioxygenases as a model system, I have investigated the value of recombination as a search strategy for laboratory evolution. Chimeric dioxygenase libraries constructed by DNA shuffling are first evaluated for biases that limit sequence diversity using a probe hybridization approach in lieu of sequencing. This analysis shows that crossovers preferentially occur in regions with high sequence identity and that certain parent sequences can be preferred at particular gene positions. High-throughput functional screening allowed characterization of substrate specificity for hundreds of dioxygenase chimeras. These data are coupled with sequence data to reveal sequence-function relationships and demonstrate the utility of recombination as a tool for functional genomics. One region of sequence is shown to be a primary determinant of substrate specificity for the enzymes studied. Furthermore, several sets of variant enzymes with similar functionality are shown to have sequence similarities. Recombination and random mutagenesis are compared as search strategies for generating functionally-diverse dioxygenases. I screened similarly sized libraries of chimeric and mutant dioxygenases for variants with altered substrate specificity or activity toward n-hexylbenzene, which is not accepted by the parent enzymes. Both recombination and random mutagenesis gave rise to enzymes with altered substrate specificity, although such enzymes were more frequent in the chimeric libraries and more distinct specificities were found in the chimeric libraries. Only chimeras were active toward n-hexylbenzene. These results support the view that recombination is an effective search strategy for evolving substrate specificity, and may be more effective than random mutagenesis.
Chemical composition of leaf tissues of Sultana vines grown in nutrient solutions deficient in macro-elements

Sultana vines, grown for one month after budburst in complete nutrient solution, were then grown in solutions deficient in N, P, K, Ca, Mg, or S for periods ranging from one to two months. Shoot and root weights were obtained from vines at five sampling times. The first time was associated with nitrogen deficiency symptoms in the nitrogen deficient treatment and the second time with potassium, calcium, magnesium, and sulphur deficiency symptoms in the respective deficient treatments. In the phosphorus deficient treatment foliar symptoms were not observed but shoot growth was reduced. Shoot lengths were measured weekly. Contents of P, K, Ca, and Mg in basal and recently matured laminae and petioles, and of N in laminae are discussed in relation to the choice of sampling tissue for diagnostic purposes. It is concluded that the most satisfactory single tissue for diagnosing deficiencies of P, K, Ca, or Mg is basal petioles. Concentrations (per cent dry matter) in basal petioles of 0.1 % P, 0.8% K, 0.5% Ca, and 0.2% Mg were associated with either the onset of deficiency symptoms or of reduced shoot growth.
Two new species and a new subspecies of Tetraclitella (Cirripedia: Thoracica) from the Cainozoic of Australia and New Zealand and a consideration of the significance of tubiferous walls

A tubiferous cirripede Tetraclitella judiciae sp. nov., previously known only from two isolated and incomplete shell wall plates identified as Tetraclitella sp. cf. T. purpurascens (Wood, 1815), is described from the early Miocene of Victoria, Australia; a further taxon, Tetraclitella purpurascens miocenica subsp. nov., occurs in the late Miocene-Pliocene of Victoria, and specimens from the early Miocene of New Zealand, previously recorded as Tetraclitella sp. cf. T. purpurascens (Wood, 1815) are redesignated as Tetraclitella nodicostata sp. nov. Tetraclitella is the first cirripede genus known to have had tubiferous walls. Incorporation of chitinous stringers within the shell wall of early tetraclitids (e.g. Epopella) may have facilitated the development of the tubiferous shell wall, which permitted sessile barnacles to maximise the shell strength to calcite ratio: in doing so, these chitinous stringers not only reduced the diversion of energy required to extract calcium carbonate from seawater, but improved the effectiveness of the shell wall in resisting predators. It is also argued here that the presence of chitin within the shell increased resistance to both corrasion and corrosion, the latter becoming an increasing problem for calcareous shelled organisms following a drop in the pH of seawater after the PalaeoceneEocene Thermal Maximum.
Cyclooxygenase (COX)-2 Inhibitor Celecoxib Abrogates TNF-Induced NF-κB Activation through Inhibition of Activation of IκBα Kinase and Akt in Human Non-Small Cell Lung Carcinoma: Correlation with Suppression of COX-2 Synthesis1

The cyclooxygenase 2 (COX-2) inhibitor celecoxib (also called celebrex), approved for the treatment of colon carcinogenesis, rheumatoid arthritis, and other inflammatory diseases, has been shown to induce apoptosis and inhibit angiogenesis. Because NF-κB plays a major role in regulation of apoptosis, angiogenesis, carcinogenesis, and inflammation, we postulated that celecoxib modulates NF-κB. In the present study, we investigated the effect of this drug on the activation of NF-κB by a wide variety of agents. We found that celecoxib suppressed NF-κB activation induced by various carcinogens, including TNF, phorbol ester, okadaic acid, LPS, and IL-1β. Celecoxib inhibited TNF-induced IκBα kinase activation, leading to suppression of IκBα phosphorylation and degradation. Celecoxib suppressed both inducible and constitutive NF-κB without cell type specificity. Celecoxib also suppressed p65 phosphorylation and nuclear translocation. Akt activation, which is required for TNF-induced NF-κB activation, was also suppressed by this drug. Celecoxib also inhibited the TNF-induced interaction of Akt with IκBα kinase (IKK). Celecoxib abrogated the NF-κB-dependent reporter gene expression activated by TNF, TNF receptor, TNF receptor-associated death domain, TNF receptor-associated factor 2, NF-κB-inducing kinase, and IKK, but not that activated by p65. The COX-2 promoter, which is regulated by NF-κB, was also inhibited by celecoxib, and this inhibition correlated with suppression of TNF-induced COX-2 expression. Besides NF-κB, celecoxib also suppressed TNF-induced JNK, p38 MAPK, and ERK activation. Thus, overall, our results indicate that celecoxib inhibits NF-κB activation through inhibition of IKK and Akt activation, leading to down-regulation of synthesis of COX-2 and other genes needed for inflammation, proliferation, and carcinogenesis.
Dynamic and thermodynamic processes driving the January 2014 precipitation record in southern UK

HAL is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers. L’archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d’enseignement et de recherche français ou étrangers, des laboratoires publics ou privés. Revisiting dynamic and thermodynamic processes driving the January 2014 precipitation record in southern UK Boutheina Oueslati, Pascal Yiou, Aglaé Jézéquel
Response of Current Winter Wheat Cultivars Grown in Turkey to Immature Embryo Culture

Selecting the explant genotypes is crucial step in in-vitro culture and Agrobacterium-mediated transformationsystem due to its host range specificity. Poor tissue culture performance is restricting the number of wheat genotypes that can bestably transformed. Response of fourteen cultivars of winter wheat (Triticum aestivum L.) to immature embryo culture, callusproduction and plant regeneration was evaluated. A significant (p < 0.05) genotypic difference in callus induction frequency wasobserved ranging between 45% (cv. Surak) and 99.54% (cv. Bobwhite) of the cultured explants. Plant regeneration through somaticembryogenesis was observed from calli of all 15 cultivars after a 4-week callus induction period. Genotypes with the highestnumber of plantles per cultured embryo were Sivas-111/33 (8.66), Kirac-66 (7.36), Haymana 79 (6.77) and Conkesme (6.56). Fourwinter wheat genotypes, ‘Sivas-111/33, Kirac-66, Haymana 79, and Conkesme’, have been selected for initial Agrobacteriummediated transformation experiments due to their high regeneration potentials.
Genetic diversity parameters in two experimental chicken populations designed for mapping quantitative trait loci.

Financial support by FAPESP (grant number 04/02080-2), USP/ESALQ, Department of Genetics, Av: Padua Dias, 11, 13400-918, Piracicaba/SP, Brazil, Embrapa Suinos e Aves, BR 153, km 110, Concordia/SC, 89700-000, Brazil, UNESP/FMVZ, Botucatu/SP, Brazil, University of Azores, DCA/CBA, Terceira, Azores Portugal, AgResearch Limited/Grasslands Research Centre, Palmerston North, New Zealand, Unb/FAV, Brasilia/DF, Brazil, USP/ESALQ, Department of Animal Science, Piracicaba/SP, Brazil
Elevational variation in diversity and composition of land-snail faunas in a Tanzanian forest

We sampled terrestrial molluscs at fifteen elevations between 400 and 2000 m in Mwanihana Forest Reserve, Tanzania and recorded 84 taxa. Four diversity measures – species density (DP), species richness (D20) and observed (SO) and interpolated (SI) alpha diversity – were measured; beta diversity and abundance were also investigated. Mean elevational range was 470 m. DP, D20, alpha diversity and mollusc abundance increased with elevation, but alpha diversity peaked at 1695 m (SO) or 1500 m (SI) and declined at higher elevations. Maximum beta diversity was at 1000 m. Soil pH was negatively correlated with elevation and DP. Cluster analysis divided the sites into lowland (400–900 m) and highland (>1000 m) groups. Axis 1 of a canonical correspondence analysis was associated with altitude and suggested a faunal discontinuity at 1000 m. Variation within the highland (>1000 m) and lowland faunal sets was related to elevation and forest structure respectively. The findings indicate that mollusc diversity peaks at intermediate elevations. This may be related to the combined effects of low rainfall conditions at low elevations and increasing effects of soil leaching at high elevations, both of which may limit mollusc diversity and abundance. Diversity at intermediate altitudes may be further elevated because of faunal mixing of lowland and highland groups. Resume Nous avons echantillonne des mollusques terrestres a 15 elevations entre 400 m et 2000 m dans la Reserve Forestiere de Mwanihana en Tanzanie et enregistre 84 taxons. Quatre mesures de diversite– la densite des especes (DP), l'abondance des especes (D20) et la diversite alpha, la diversite beta et l'abondance observees (SO) et interpolees (SI) furent aussi enquetes. La portee d’elevation moyenne fut 470 m. DP, D20, la diversite alpha et l'abondance des mollusques augmenterent avec l’elevation, mais la diversite alpha culmina a 1695 m (SO) ou 1500 m (SI) et diminua aux elevations plus hautes. Le niveau maximum de diversite beta fut a 1000 m. Le pH du sol fit une correlation negative avec le niveau d’elevation et DP. Les sites furent repartis en groupes de basse altitude (400–900 m) et haute altitude (>1000 m) a travers une analyse par faisceau. Le premier axe d'une analyse canonique des correspondances (CCA) fut associea l'altitude et impliqua une discontinuite de la faune a 1000 m. Des fluctuations dans les groupes de faune de haute altitude (>1000 m) et de basse altitude furent liees a l’elevation et la structure forestiere respectivement. Les conclusions demontrent que la diversite des mollusques atteint des pics aux elevations intermediaires. Ceci peut etre lie aux effets joints d'une pluviosite faible dans les basse altitudes et le relevement des effets du lessivage du sol aux hautes elevations, qui peuvent tous les deux limiter la diversite et l'abondance des mollusques. La diversite aux altitudes intermediaires peut etre elevee davantage a cause du melange des faunes dans les groupes de basse altitude et de haute altitude.