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See detailDevelopment and applications of CRISPR-Cas9 and RNAi for rice and wheat agronomic traits improvement
Sun, Yongwei ULg

Doctoral thesis (2017)

Genome editing technologies enable precise modifications of DNA sequences in vivo and offer a great promise for harnessing plant genes in crop improvement. The precise manipulation of plant genomes relies ... [more ▼]

Genome editing technologies enable precise modifications of DNA sequences in vivo and offer a great promise for harnessing plant genes in crop improvement. The precise manipulation of plant genomes relies on the induction of DNA double-strand breaks (DSBs) by sequence-specific nucleases (SSNs) to initiate DNA repair reactions that are based on either non-homologous end joining (NHEJ) or homology-directed repair (HDR). NHEJ can result in frame-shift mutations that often create genetic knockouts. These knockout lines are useful for functional and reverse genetic studies but also have applications in agriculture. HDR has a variety of applications as it can be used for gene replacement, gene stacking, and for creating various fusion proteins. An overview of development and applications of precise GT in plants using SSNs systems is presented in Chapter I. Cereals high in amylose content (AC) and resistant starch (RS) offer potential health benefits. Starch branching enzyme (SBE) plays a major role in determining the fine structure and physical properties of starch. Here, we use CRISPR/Cas9 technology to generate targeted mutagenesis in SBEI and SBEIIb in rice. The frequencies of obtained homozygous or bi-allelic mutant lines with indels in SBEI and SBEIIb in T0 generation were from 26.7 to 40%. SbeII mutants showed higher proportion of long chains presented in debranched amylopectin, significantly increased AC and RS content to as higher as 25.0% and 9.8%, respectively. The details of results are presented in Chapter IV Complete knockouts and loss-of-function mutations are very valuable in defining gene functions, their applications in crop improvement are somewhat limited because many agriculturally important traits are conferred by point mutations or a change of gene expression levels. Development of a technology that enables gene replacement rather than gene inactivation will greatly facilitate plant breading. In this study, we report an efficient method to introduce multiple discrete point mutations in the rice ALS gene using CRISPR/Cas9-mediated homologous recombination. We not only generated homozygous herbicide resistance rice plants in one generation, but also demonstrated that the presented strategy is feasible and effective in precise gene replacement by using CRISPR/Cas9 system to facilitate crop genetic improvement. The details of results are presented in Chapter V Aphids are major agricultural pests which cause significant yield losses of wheat each year in China. Present attempts to improve the aphid resistance of wheat through conventional breeding in China are being confronted by slow progress due to lack of aphid resistant wheat germplasm, the complexity of plant-aphid interactions and the rapid development of resistant pest biotypes. An overview of the engineering of plants for aphid resistance is presented in chapter II. We identify a gene (23028) related to aphid ingestion and digestion from the aphid, S. avenae. Aphids fed on plant material expressing double-stranded RNA (dsRNA) specific to 23028 and C002 show a decline in growth, reproduction and survival rates. The details of results presented in the chapter VI. [less ▲]

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See detailGENETIC DIVERSITY AND FACULTATIVE SYMBIONTS INFECTION PATTERNS OF TWO RHOPALOSIPHUM APHIDS
Guo, Jianqing ULg

Doctoral thesis (2017)

Aphids, during the long-term evolution, kept their genetic traits for continuation of the species while some variations caused by selection, migration, mutation, and genetic drift occurred meanwhile to ... [more ▼]

Aphids, during the long-term evolution, kept their genetic traits for continuation of the species while some variations caused by selection, migration, mutation, and genetic drift occurred meanwhile to adapt to novel environments. The differentiation takes place not only among species but could also be observed within species that finally contributes to the population genetic diversity. However, the migrant behavior of aphid can enhances the gene flow among populations, as a result, decreasing the genetic variation. Additionally, insect such as aphids usually hold the mutualistic relationships with endosymbionts which may affect the adaptive characteristics of the host aphid. Both Corn leaf aphid Rhopalosiphum maidis (F.) (Hemiptera: Aphididae) and Bird cherry-oat aphid Rhopalosiphum padi (L.) (Hemiptera: Aphididae) are worldwidely severe pests that attack cereal crops such as maize, barley, wheat, sorghum and oats. They suck the phloem and can transmit viruses including Maize dwarf mosaic virus (MDMV) and Barley yellow dwarf virus (BYDV), the main virus of maize and barley, which may result in serious damage for the yield and quality of the host plant. Moreover, one aphid species usually exerts genetic differentiation among populations which is related to factors such as geographic conditions, host plants and life history. Hence, it is necessary to have a clear understanding of the genetic structure and variation among geographic populations to further explore the historical expanding routes and occurrence of these pests and design efficient control strategies. Targeting the genetic divergence among populations of two Rhopalosiphum aphids, 32 populations of R. padi and 38 populations of R. maidis were collected on maize from most regions of China as well as some countries in Europe. The sequencing results of two mitochondrial (COI and COII) and one nuclear (EF-1α) genes indicated that a high level of gene flow existed among the populations of R. maidis in terms of geographic locations, proving the migratory capacity of this aphid. With regard to R. padi, high genetic diversity were found between the populations from China and Europe based on combined COI-COII, however, the genetic diversification among Chinese populations and European populations were low, which illustrated that the length of geographic distance may play an important role in genetic diversity. The infection patterns of seven facultative symbionts (Serratia symbiotica, Hamiltonella defensa, Regiella insecticola, Rickettsia, Spiroplasma, Wolbachia and Arsenophonus) in R. padi and R. maidis were investigated as well. Both aphids were found to have wide symbiotic associations with variant symbionts. Unlike Acyrthosiphum pisim (H.) which is frequently infected with S. symbiotica, R. insecticola, Rickettsia and Spiroplasma were dominant facultative symbionts in these two Rhopalosiphum aphids. The results also indicated that the geographic conditions such as latitude of collecting locations may have an effect on the distribution of facultative symbionts, implying the adaptive function of the facultative symbionts to various environmental conditions on their host aphids. Considering the population diversity within species, we also tested the survival and reproduction rates of two R. maidis populations (i.e. Beijing and Mangshi) on barley and maize of small (10 cm) and high (50 cm) size. Direct observation method was used and none of the populations could survive on maize of small size whereas they were capable to develop on maize of high size, but with much lower growing speeds compared with barley on which both populations showed the best performances. In addition, Beijing population was found to perform significantly better than Mangshi population which may be related to the infection of facultative symbionts. We hypothesize that the lethality on small maize may result from the secondary metabolites synthesized to be more abundant in young maize seedlings. [less ▲]

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See detailStrategies of host defense and viral counter defense: analysis of Fusarium graminearium responses to FgHV1 infection
Wang, Shuangchao ULg

Doctoral thesis (2017)

Viruses that selectively infect fungi are named fungal viruses or mycoviruses. Fungal viruses exist in all major groups of fungi and are important in both fundamental research and biological control of ... [more ▼]

Viruses that selectively infect fungi are named fungal viruses or mycoviruses. Fungal viruses exist in all major groups of fungi and are important in both fundamental research and biological control of plant pathogens. Fusarium graminearum hypovirus 1 (FgHV1) was the first virus isolated from F. graminearum belonging to the hypoviridae family. FgHV1 was supposed to encode two open reading frames, which had high sequence identities with Cryphonectria hypovirus 1 and 2 (CHV1 and CHV2). F. graminearum strain infected with FgHV1 showed lower growth rate and reduced conidia production, but with no pathogenicity change. To elucidate the host defense and viral counter defense mechanism, we analyzed the F. graminearum stress responses to FgHV1 infection from several aspects. In the first part, we preliminarily elucidated the RNA silencing mechanism of the F. graminearum/hypovirus system from a small RNA (sRNA) perspective. The length distributions of F. graminearum sRNA were altered by FgHV1 infection, especially 24-nt sRNA. Extensive FgHV1-derived sRNAs were detected, yielding the first high resolution map of sRNA from mycoviruses. In particular, the RNA silencing-related genes FgDicer1 and FgRdRp5 were predicted targets of FgHV1- and FgHV2-derived siRNAs, possibly revealing a novel anti-RNA silencing strategy employed by mycoviruses. In the second part, we performed genome-wide expression analysis to reveal stress-related genes with expression changes in response to FgHV1 infection. A total of 248 differentially expressed genes were identified. Cellular redox regulation related genes were the most affected categories in F. graminearum challenged with FgHV1. Then we verified that FgHV1 encoded protein 20 (p20) could induce the H2O2 accumulation and hypersensitive response in Nicotiana benthamiana leaves. In the last part, our experiments confirmed that p20 functioned as a RNA silencing suppressor (RSS) with the agroinfiltration-mediated technique, making p20 the third RSS identified in mycoviruses. By incorporating single strand sRNA, p20 could suppress host RNA silencing. Globally, the defense responses of F. graminearum to FgHV1 infection were studied all-around and in-depth. On the other hand, FgHV1 developed counter defense strategies to protect from invasion resistance. [less ▲]

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See detail"888" Tweets on the Art and Science of Internet-Based Investing
Choffray, Jean-Marie ULg

E-print/Working paper (2017)

Here we are! Exactly “888” Tweets, regularly updated, on The Art and Science of Investing to “show you the way”, to share some observations on the evolution of the world, the behavior of the financial ... [more ▼]

Here we are! Exactly “888” Tweets, regularly updated, on The Art and Science of Investing to “show you the way”, to share some observations on the evolution of the world, the behavior of the financial markets, and to illustrate the key heuristics that I learned over the last twenty years. As, without any doubt, I am no Warren Buffett, feel free to disagree and develop you own “theory” of investing. Painful and solitary work... Success is at that price! And, based on what I did - and did not! - achieve, it shouldn’t be hard to do much better... That’s my wish for you! [less ▲]

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See detailUntangling glycaemia and mortality in critical care
Uyttendaele, Vincent ULg; Dickson, Jennifer L.; Shaw, Geoffrey M. et al

in Critical Care (2017), 21(1), 152

Background: Hyperglycaemia is associated with adverse outcomes in the intensive care unit, and initial studies suggested outcome benefits of glycaemic control (GC). However, subsequent studies often ... [more ▼]

Background: Hyperglycaemia is associated with adverse outcomes in the intensive care unit, and initial studies suggested outcome benefits of glycaemic control (GC). However, subsequent studies often failed to replicate these results, and they were often unable to achieve consistent, safe control, raising questions about the benefit or harm of GC as well as the nature of the association of glycaemia with mortality and clinical outcomes. In this study, we evaluated if non-survivors are harder to control than survivors and determined if glycaemic outcome is a function of patient condition and eventual outcome or of the glycaemic control provided. Methods: Clinically validated, model-based, hour-to-hour insulin sensitivity (SI) and its hour-to-hour variability (%ΔSI) were identified over the first 72 h of therapy in 145 patients (119 survivors, 26 non-survivors). In hypothesis testing, we compared distributions of SI and %ΔSI in 6-hourly blocks for survivors and non-survivors. In equivalence testing, we assessed if differences in these distributions, based on blood glucose measurement error, were clinically significant. Results: SI level was never equivalent between survivors and non-survivors (95% CI of percentage difference in medians outside ±12%). Non-survivors had higher SI, ranging from 9% to 47% higher overall in 6-h blocks, and this difference became statistically significant as glycaemic control progressed. %ΔSI was equivalent between survivors and non-survivors for all 6-hourly blocks (95% CI of difference in medians within ±12%) and decreased in general over time as glycaemic control progressed. Conclusions: Whereas non-survivors had higher SI levels, variability was equivalent to that of survivors over the first 72 h. These results indicate survivors and non-survivors are equally controllable, given an effective glycaemic control protocol, suggesting that glycaemia level and variability, and thus the association between glycaemia and outcome, are essentially determined by the control provided rather than by underlying patient or metabolic condition. [less ▲]

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See detailLe contrôle du respect des droits fondamentaux de ses condamnés par la Cour pénale internationale : théorie et pratique naissante
Deprez, Christophe ULg

Conference (2017, June 23)

Cette présentation PowerPoint a servi de support à une communication orale dans le cadre de l'édition 2017 de la Conférence des Assistants en droit de Belgique (ACCA), organisée le 23 juin 2017 par l ... [more ▼]

Cette présentation PowerPoint a servi de support à une communication orale dans le cadre de l'édition 2017 de la Conférence des Assistants en droit de Belgique (ACCA), organisée le 23 juin 2017 par l'Université de Liège. [less ▲]

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See detailGreen Biotechnology in Belgium: The Ghent Innovation District.
Segers, Jean Pierre ULg

E-print/Working paper (2017)

This contribution explores the long-standing evolution of the green biotechnology cluster in the bioRegion of Flanders (Belgium), in particular in the Ghent innovation district.

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See detailLa comparaison en Belgique
Delville, Michel ULg

Conference (2017, June 22)

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See detailUHPLC-ESI-MS/MS quantitation of natural estrogens in aqueous matrices as pyridine-3-sulfonyl derivatives
Glineur, Alex ULg; Ronkart, Sébastien; Nott, Kathérine et al

Conference (2017, June 22)

Natural estrogens (estrone: E1, 17-β-estradiol : E2, estriol : E3) and synthetic estrogen (17-α-ethinylestradiol: EE2) are powerful endocrine disruptors. They may have deleterious effects on aquatic ... [more ▼]

Natural estrogens (estrone: E1, 17-β-estradiol : E2, estriol : E3) and synthetic estrogen (17-α-ethinylestradiol: EE2) are powerful endocrine disruptors. They may have deleterious effects on aquatic wildlife and also humans even at extremely low concentrations. For this reason, these molecules have been included in a watch list from the European Commission regarding emerging aquatic pollutants. The maximum detection limits are set at 0.035 and 0.40 ng/L for EE2 and E1, E2 respectively. Reaching such low levels of concentration of estrogenic compounds is a challenge, even using state-of-the-art analytical methods. In this study, we developed a UHPLC-ESI-MS/MS method allowing the quantification of E1, E2, E3 and EE2 residues in aqueous matrices. Studies commonly used ESI in negative mode albeit the poor ionization efficiency of native estrogenic compounds in this mode. In this study, the molecules were derivatised using a sulfonyl chloride reagent (pyridine-3-sulfonyl, P-3-S). The resulting response in the positive mode was significantly enhanced. Similarly to other dansyl derivatives, the product ion spectra of the P-3-S derivatives indicate ions originating from the derivatization reagent moiety. Moreover, several other ions were included in the product ion spectra of the P-3-S derivatives. Their specificity was assessed by a qualitative approach implying the analysis of different types of water samples (groundwater, surface water). Some product ions were found to be noticeably better for quantification and confirmation of the analytes. The developed analytical method was validated according to the NF T90-210 norm which is suitable to assess the performances of a method in the water quality field. The limits of quantification were 0.04, 0.05, 0.10 and 0.02 ng/L in groundwater and 0.89, 0.79,4.42 and 0.29 ng/L in surface water for E1, E2, E3 and EE2 respectively. [less ▲]

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See detailImprovement of the ChIP-seq technique for histone posttranslational modifications through enhanced laboratory protocols and data processing methods
Laczik, Miklos ULg

Doctoral thesis (2017)

Massively parallel sequencing has rapidly become the dominant technique in various omics studies as it provides an unequalled amount of high quality quantitative data for a reasonable cost that is ... [more ▼]

Massively parallel sequencing has rapidly become the dominant technique in various omics studies as it provides an unequalled amount of high quality quantitative data for a reasonable cost that is decreasing ever since the first next generation sequencers appeared. Subsequently, chromatin immunoprecipitation coupled with massively parallel sequencing (ChIP-seq) is the prevailing method of choice for investigating protein–DNA interactions in a genome-wide manner. Bioinformatics tools are also evolving quickly to meet the increasing demands of processing huge amounts of ChIP-seq data and to open the way for novel techniques and insights. However areas still exist that could benefit from improved wet-lab and dry-lab methods. One such area is data visualisation and interpretation; another is the ChIP-seq study of histone posttranslational modifications, especially the research of inactive histone marks which tend to produce diffuse broad enrichments instead of point-source peaks. Achieving proper enrichment and unbiased analysis in such histone mark studies proves to be a great challenge. In this doctoral thesis we show how we addressed these issues on both the level of bioinformatics and the level of sample processing methods. We present our innovative analysis tools we developed to this end, among others a highly customisable, feature rich viewer for next generation sequencing data visualisation, and an analysis pipeline specifically aimed to handle broad enrichments from ChIP-seq studies of (inactive) histone marks. We propose specific software and specific peak calling settings to detect a range of histone modifications accurately, and we describe the way to determine the optimal settings. Along the pipeline we also present a protocol designed to enhance enrichments and facilitate peak detection in broad peak studies typical of inactive histone marks. We demonstrate how this method affects various enrichment types and propose potential applications that could benefit from it. Furthermore we show diverse achievements with the analysis pipeline, including the interpretation of the aforementioned wet-lab method, and the development of an automated ChIP-seq protocol optimised for low cell numbers. [less ▲]

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See detailField border flowering strips as a source of valuable compounds
Paul, Aman ULg

Doctoral thesis (2017)

Intensification of agricultural practices has caused irreversible damage to environment during the last few years. Several consumers are now deviating towards healthier diets produced from eco-friendly ... [more ▼]

Intensification of agricultural practices has caused irreversible damage to environment during the last few years. Several consumers are now deviating towards healthier diets produced from eco-friendly and sustainable agricultural systems. In this regard, the possibilities of utilizing edible biomass originating from sustainable agricultural practices have gained recent attention. The underutilized edible plants, especially their seeds could be one of the interesting alternates, as some of these seeds are not only nutritious, but could also be produced using sustainable practices. Similarly, edible insects represent another category of biomass which are rich in nutrients and could be produced sustainably. The seeds from underutilized edible plants and the edible insects could be simultaneously harvested using a sustainable agricultural system involving field border flowering strips. Field border flowering strip is a part of agricultural landscape that is reserved for herbaceous vegetation. These strips are popularly grown throughout the world to enhance biodiversity. The main objective of this thesis is to utilize seeds from some of the plants grown as field border flowering strips and insects that find refuge in these plants for the provision of food and health promoting substances. From the literature reviewed in chapter two, it was observed that: (1). Most plants that are grown in field border flowering strips are edible, and their aerial parts have been extensively analyzed for chemical composition. However, there is a scarcity of literature evaluating chemical composition/food utilization of the seeds from plants that are grown as field border flowering strips. So, the primary objective of this thesis is to investigate the nutritional and health promoting potential of the seeds from some plants that are grown in these strips. (2). A number of grasshopper species find refuge in field border flowering strips. Several grasshopper species are considered edible throughout the world and they are interesting source of nutrients. So the secondary objective of this thesis is to screen some edible grasshopper species that are present in field border flowering strips, analyze their nutritional value, and investigate possibilities to establish their commercial rearing for ensuring year-long availability of edible biomass. The research strategy adopted to achieve the objectives of this thesis is mentioned in chapter three. This chapter includes details about the selection of raw materials (both plant seeds and insects), and subsequent analysis. Chapter four contains the detail about the materials and methods used for analysis during this study. Chapter five includes details about the investigations on edible insects. Chorthippus parallelus Zetterstedt species grasshoppers were shortlisted for detailed investigation due to their high densities in field border flowering strips. This insect species was analyzed for proximate composition, amino acid profile, fatty acid profile and mineral profile. Moreover the toxicity of these insects was also evaluated using two different models. Results indicated that these insects could be consumed as an alternate source of proteins (69%) and omega-3 fatty acid rich lipids (10%). Rearing trials done during this study indicates that commercial rearing could be developed to produce sufficient and safe biomass for human consumption. The selection of seeds from three plant species (Achillea millefolium L., Anthriscus sylvestris (L). Hoffm. and Prunella vulgaris L.), for detailed analysis on the basis of lipid content and fatty acid profile has been mentioned in chapter six. Chapter seven, eight and nine include the details about the composition and anti-oxidant activity of A. millefolium, A. sylvestris and P. vulgaris seeds, respectively. Proximate composition, lipid profile, amino acid profile, mineral profile, lignocellulosic profile, phenolic profile and phytate content of the three plant seeds were investigated during this study. Two new phenolic acids were discovered originating from P. vulgaris seeds. These compounds were named amolsamic acid A and amolsamic acid B. Discovery of these compounds was the true highlight of this thesis. All the three plant seeds were found to contain substantial level of total phenolics (0.8-2.6%) and interesting phenolic profiles (dominated by chlorogenic acid, rosmarinic acids and related compounds). Keeping this in mind, the detailed anti-oxidant activity (including anti-radical scavenging, horseradish peroxidase response modulation, cellular anti-oxidant, myeloperoxidase response modulation and anti-lipid peroxidation activity) of their respective seed extracts was also analyzed. Results obtained during this study indicate that A. millefolium, A. sylvestris and P. vulgaris seeds not only contain interesting level of nutrients, but their extracts also exhibit significant anti-radical scavenging, horseradish peroxidase response modulation, cellular anti-oxidant (IC50 values order: P. vulgaris>A. sylvestris>A. millefolium) and myeloperoxidase response modulation activity (IC50 values order: A. sylvestris>A. millefolium>P. vulgaris for both direct and SIEFED assay). The main conclusions (chapter ten) of this PhD dissertation are: (1). C. parallelus insects could be viewed as an alternative source of nutrients to diversify human diets. The preliminary rearing studies done during this study indicate that commercial rearing could be developed for generation of substantial (and safe) biomass to support human consumption. (2). A. millefolium, A. sylvestris and P. vulgaris seeds could be included in food formulations (or consumed as whole) as a source of proteins, lipids, minerals and phenolics. P. vulgaris seeds could also be used for the extraction of two new phenolic constituents (amolsamic acid A and amolsamic acid B). The first investigations involving A. millefolium, A. sylvestris and P. vulgaris seeds realized during this study, indicate that seed extract (or whole seeds) from all three plants could possibly be consumed for the prevention of neutrophil and myeloperoxidase mediated damage in human body. [less ▲]

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See detailSeasonal Transport in Mars’ Mesosphere revealed by Nitric Oxide Nightglow
Stiepen, Arnaud ULg

Conference (2017, June 21)

We analyze the ultraviolet nightglow in the atmosphere of Mars through Nitric Oxide (NO) δ and γ bands emissions. On the dayside thermosphere of Mars, solar extreme ultraviolet radiation partly ... [more ▼]

We analyze the ultraviolet nightglow in the atmosphere of Mars through Nitric Oxide (NO) δ and γ bands emissions. On the dayside thermosphere of Mars, solar extreme ultraviolet radiation partly dissociates CO2 and N2 molecules. O(3P) and N(4S) atoms are carried by the day-to-night hemispheric transport. They preferentially descend in the nightside mesosphere in the winter hemisphere, where they can radiatively recombine to form NO(C2Π). The excited molecules promptly relax by emitting photons in the UV δ bands and in the γ bands through cascades via the A2Σ, v’ = 0 state. These emissions are thus indicators of the N and O atom fluxes transported from the dayside to Mars’ nightside and the winter descending circulation pattern from the nightside thermosphere to the mesosphere (e.g. Bertaux et al., 2005 ; Bougher et al., 1990 ; Cox et al., 2008 ; Gagné et al., 2013 ; Gérard et al., 2008 ; Stiepen et al., 2015, 2017). Observations of these emissions have been accumulated on a large dataset of nightside disk images and vertical profiles obtained at the limb by the Imaging Ultraviolet Spectrograph (IUVS, McClintock et al., 2015) instrument when the Mars Atmosphere and Volatile EvolutioN (MAVEN) spacecraft is at its apoapsis and its periapsis phases along its orbit, respectively. We present discussion on the variability in the brightness and altitude and of the emission with season, geographical position and local time and possible interpretation for local and global changes in the mesosphere dynamics. IUVS images and limb scans reveal unexpected complex structure of the emission. The brightest emission is observed close to the winter pole. The emission is also surprisingly more intense in some longitude, revealing possible impact of tides and waves in Mars’ nightside mesosphere. Observations also reveal spots and streaks, indicating irregularities in the wind circulation pattern. The disk images and limb profiles are compared to the LMD-MGCM model (Gonzàlez-Galindo et al., 2009 ; Lopez-Valverde et al., 2011) to focus on the seasonal, local time and geographical influences on the NO Nightglow emission. We will also provide a statistical study of the regions of enhanced brightness (i.e. splotches and streaks) and discuss possible interpretation from the comparison to the GCM. [less ▲]

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