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  • 1
    ISSN: 1432-0517
    Keywords: Key words GMO detection ; PCR ; Food labelling ; Quality assurance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract  The detection of genetically modified organisms (GMOs) by the polymerase chain reaction (PCR) is a complex multiparameter problem. Therefore, a number of critical issues in respect to quality control need to be considered. For practical purposes, the PCR process itself can be divided into three subprocesses: template isolation and reaction setup (pre-PCR), PCR reaction and detection of amplification products, and data evaluation (post-PCR). Crucial factors for the pre-PCR process are the following: homogeneity of the sample to be analysed, performance of template isolation and purification in terms of yield and purity, standardized process for the estimation of concentrations of genomic DNA and all reagents used in the reaction. For the PCR itself, crucial factors to be controlled are: setup of reactions, batch to batch variations of reagents, temperature-time programs used for the PCR amplification, and the performance of different types of hardware (e.g. different brands of thermocyclers). The crucial factor for the post-PCR process is the detection of the amplification products of the PCR. The tremendous sensitivity of PCR methods requires a careful and consequent separation of the three processes in terms of hardware, laboratory space and sample handling. The avoidance of contamination is one of the most critical factors. The goal of quality assurance measures must be to ensure appropriate results at maximum sensitivity. The complexity of any PCR system used for the detection of GMOs leads to the requirement of a careful validation process for any laboratory using such methods. For qualitative analyses crucial validation parameters are: specificity, selectivity, repeatability, intermediate precision, reproducibility, limit of detection and robustness.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1434-4475
    Keywords: Keywords. Molecular calculations; ab initio Calculations; Conformational analysis; Levcromakalim; Potassium channel opener.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Description / Table of Contents: Zusammenfassung.  Die Konformationen des Kaliumkanalöffners Levcromakalim werden mit Hilfe quantenchemischer Molekülrechnungen untersucht, um die energetisch günstigsten Strukturen zu bestimmen. Der Einfluß der intramolekularen Wasserstoffbrücke auf die Konformationen und die Dynamik des Moleküls wird analysiert. Die Geometrien verschiedener rigider Analoga werden mit den konformativen Möglichkeiten des Levcromakalims verglichen, um Ähnlichkeiten der Molekülgestalt festzustellen.
    Notes: Summary.  The conformations of the potassium channel opener levcromakalim are analyzed with the aid of quantum chemical calculations in order to determine the energetically most favourable structures. The influence of intramolecular hydrogen bonding on the conformations and on the rotational potential of the molecule is investigated. The structures of different conformationally restricted analogs are compared with the energetically accessible conformational space of levcromakalim with emphasis on similarities in molecular shape.
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  • 3
    Publication Date: 2014-11-12
    Description: To understand function and stability of ecosystems it is crucial to gain insights into their species composition, particulary in the face of global warming. Next Generation Sequencing (NGS) is the method of choice for getting fast overviews of species diversity in a high number of samples. Currently, there are lively discussions about bioinformatic techniques to enhance the quality of sequencing outputs and how to post process these data in order to estimate the “real” diversity as precisely as possible. In this study, we analyzed the protist composition of three water samples, collected in the Fram Strait in 2010. We compared different potential sequencing error corrected and uncorrected datasets, which were generated with widely used open-source software: QIIME, mothur and PhyloAssigner. Relative abundance of protist phyla was hardly affected by the choice of the software, quality filtering and error correction. However, the outputs differed strongly in relative abundance of diatom genera and were not comparable to dominant diatoms observed with light microscopy. Our main findings are beneficial for the enhancement of study design, data preparation and interpretation and gives insights into the optimization potential of NGS experiments in general.
    Repository Name: EPIC Alfred Wegener Institut
    Type: Conference , NonPeerReviewed , info:eu-repo/semantics/conferenceObject
    Format: application/pdf
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  • 4
    Publication Date: 2015-06-15
    Description: In the ongoing discussion of the distribution of protists, whether they are globally distributed or endemic to one or both of the polar regions is the subject of heated debate. In this study, we compared next-generation sequencing data from the Arctic and the Southern oceans to reveal the extent of similarities and dissimilarities between the protist communities in the polar regions. We found a total overlap of operational taxonomic units (OTUs) between the two regions of 11.2%. On closer inspection of different taxonomic groups, the overlap ranged between 5.5% (haptophytes) and 14.5% (alveolates). Within the different groups, the proportion of OTUs occurring in both regions greatly differed between the polar regions. On the one hand, the overlap between these two regions is remarkable, given the geographical distance between them. On the other hand, one could expect a greater overlap of OTUs between these regions on account of the similar environmental conditions. The overlap suggests a connection between the polar regions for at least certain species or that the evolutionary divergence has been slow, relative to the timescales of isolation. The different proportions of common OTUs among the groups or regions may be a result of different life cycle strategies or environmental adaptations.
    Repository Name: EPIC Alfred Wegener Institut
    Type: Article , NonPeerReviewed
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  • 5
    Publication Date: 2017-10-24
    Description: Here we present a pilot study demonstrating, that preservation with mercury chloride allows the application of PCR-based molecular methods for the characterization of marine protist communities collected with moored long-term sediment traps. They can provide information on pelagic protist communities by collecting sinking plankton from the upper water column all year-round, even in remote polar oceans. Assessment of small protist species from the nano- and picoplankton fractions in sedimented material by microscopy is extremely challenging or almost impossible. Hence, comprehensive studies of variability in protist community composition in moored long-term sediment traps are scarce. Considering that marine nano- and picoeukaryotes are ecologically very important, new approaches are urgently needed to investigate protists in the smallest size-fractions of moored long-term sediment trap samples. We applied the quick and cost-effective Terminal Restriction Length Polymorphism (T-RFLP) on a set of selected samples that were collected between 2000 and 2010 in September at a depth of ~300 m in the area of the “LTER (Long-Term Ecological Research) site HAUSGARTEN” in the eastern Fram Strait (Arctic). The results of these analyses suggest a change in the trapped protist community after 2002 in this area. A comparison of 18S sequences obtained via 454-pyrosequencing from samples collected in the water column and mercury chloride preserved sediment traps in 2009 and 2010 suggests, that sediment traps might reflect the pelagic eukaryotic microbial biodiversity qualitatively. Furthermore, we have indication that preservation with mercury chloride does not severely change the nucleotide composition of 18S rRNA genes in long-term sediment traps. Overall, we suggest that preservation with mercury chloride is a key to open the door for molecular genetic analyses of long-term sediment trap samples, and that PCR-based molecular methods have a strong potential to become an important tool for comprehensive taxonomic analyses of protist- and bacterial communities in moored long-term sediment traps.
    Repository Name: EPIC Alfred Wegener Institut
    Type: Article , NonPeerReviewed
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  • 6
    Publication Date: 2018-06-01
    Description: Next‐generation sequencing is a common method for analysing microbial community diversity and composition. Configuring an appropriate sequence processing strategy within the variety of tools and methods is a nontrivial task and can considerably influence the resulting community characteristics. We analysed the V4 region of 18S rRNA gene sequences of marine samples by 454‐pyrosequencing. Along this process, we generated several data sets with QIIME, mothur, and a custom‐made pipeline based on DNAStar and the phylogenetic tree‐based PhyloAssigner. For all processing strategies, default parameter settings and punctual variations were used. Our results revealed strong differences in total number of operational taxonomic units (OTUs), indicating that sequence preprocessing and clustering had a major impact on protist diversity estimates. However, diversity estimates of the abundant biosphere (abundance of ≥1%) were reproducible for all conducted processing pipeline versions. A qualitative comparison of diatom genera emphasized strong differences between the pipelines in which phylogenetic placement of sequences came closest to light microscopy‐based diatom identification. We conclude that diversity studies using different sequence processing strategies are comparable if the focus is on higher taxonomic levels, and if abundance thresholds are used to filter out OTUs of the rare biosphere.
    Repository Name: EPIC Alfred Wegener Institut
    Type: Article , NonPeerReviewed
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  • 7
    Publication Date: 2017-05-18
    Description: nformation on recent diversity and biogeography of Arctic marine protists with adequate temporal and spatial resolution is urgently needed to better understand consequences of environmental change for marine ecosystems. Here, we introduce a molecular-based observation strategy for high resolution assessment of marine protists in space and time, even in remote areas such as the Arctic Ocean. The observation strategy involves molecular analyses (e.g. Next Generation Sequencing (NGS) or quantitative PCR) of samples, collected with a set of complementary methods such as a newly developed automated under-way sampling device, CTD-casts and moored sediment traps. This integrated approach allows generating detailed information on marine protist community composition or abundance with adequate resolution. Currently, the observation strategy is organized at four major levels. At level 1, samples are collected at high spatial and temporal resolution based on under-way sampling with the remote-controlled automated filtration system AUTOFIM (developed in the COSYNA-project), and sampling at fixed stations based on CTD-casts and moored sediment traps. Resulting samples can either be preserved for later laboratory analyses, or directly subjected to molecular surveillance of key species aboard the ship, e.g. via quantitative polymerase chain reaction (level 2). Preserved samples are analyzed at the next observational levels in the laboratory (level 3 and 4). This involves at level 3 molecular fingerprinting methods for a quick and reliable overview of differences in protist community composition. Finally, selected samples can be used to generate a detailed analysis of taxonomic protist composition via the latest Next Generation Sequencing Technology (NGS) at level 4. An overall integrated dataset of all results provides comprehensive information on the diversity and biogeography of protists, including all related size classes. In the future, the observation strategy for Arctic marine protists will be part of the Molecular Microbial Observatory envisioned for the Arctic observatory FRAM (Frontiers in Arctic Monitoring).
    Repository Name: EPIC Alfred Wegener Institut
    Type: Conference , NonPeerReviewed
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  • 8
    ISSN: 1520-510X
    Source: ACS Legacy Archives
    Topics: Chemistry and Pharmacology
    Type of Medium: Electronic Resource
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  • 9
    ISSN: 1520-510X
    Source: ACS Legacy Archives
    Topics: Chemistry and Pharmacology
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  • 10
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    Universität Göttingen
    In:  In: Global and regional controls on biogenic sedimentation. I. Reef evolution. Research reports. , ed. by Reitner, J., Neuweiler, F. and Gunkel, F. Göttinger Arbeiten zur Geologie und Paläontologie, Sb 2 . Universität Göttingen, Göttingen, pp. 19-22.
    Publication Date: 2017-02-02
    Type: Book chapter , NonPeerReviewed
    Format: text
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