Development of a modular system for detection of genetically modified organisms in food based on ligation-dependent probe amplification

The application of a method based on ligation-dependent probe amplification to the simultaneous detection of different genetically modified organisms in food is described. The ligation reaction of target-specific probes with characteristic lengths and universal primer binding sites is followed by PCR amplification using fluorescein-labeled primers. The separation and the detection of DNA fragments are achieved by capillary electrophoresis via laser-induced fluorescence. The approach allows the simultaneous detection of several targets corresponding to different levels of specificity in a one-tube assay. Synthetic oligonucleotides were designed to detect (1) reference genes in the genome from maize, soya and rapeseed, (2) the CaMV 35S-promotor as screening element, (3) the construct-specific 35S-pat junction, and (4) the event-specific regions of the transgenic maize line MON 810 and of Roundup Ready soya. Specificity and sensitivity (GMO content 0.1%) of the approach were shown for all targets. This novel analytical strategy represents a flexible, modular system for the surveillance of GMO-derived products that can be readily complemented by further target sequences of interest.     

Free and bound phenolic compounds in leaves of pak choi (Brassica campestris L. ssp. chinensis var. communis) and Chinese leaf mustard (Brassica juncea Coss)

Eleven pak choi cultivars and two leaf mustard cultivars grown under field conditions in China were investigated for the free polyphenol content in their outer and inner leaves, as well as in their leaf blades and leaf stalks. In most cases, there were no significant differences between the hydroxycinnamic acid derivative and flavonoid derivative contents in the outer and inner leaves for the 13 cultivars. However, the contents of blades and stalks differed: hydroxycinnamic acids and flavonoids were present in greater amounts in the leaf blade than in the leaf stalk. Trace or small amounts of flavonoids were detected in the pak choi and leaf mustard stalks. Additionally, the bound phenolic contents of two pak choi cultivars and two leaf mustard cultivars were investigated. The concentrations of cell wall-bound phenolic compounds were higher in the leaf blade than in the leaf stalk under field conditions in China. These compounds represent only a minor portion of the total phenolic contents (flavonoids and hydroxycinnamic acids) in leaf stalks (0.81–1.18%) and leaf blades (0.05–0.08%) from fresh plant material. The storage of plant samples from four Chinese cabbage cultivars resulted, in most cases, in an increase of phenolic content, within six days, at 4 °C and 20 °C. The increase might have been triggered by post-harvest plant stresses, which stimulate the biosynthesis of polyphenols.     

Intelligent download and cache management for interactive non-linear video

Since the invention of digital video significant progress has been made in reducing the amount of data needed to be transferred in the World Wide Web while improving viewing experience. However the paradigm of linear behavior has not changed at all. While the feature set of traditional digital video may be sufficient for some applications, there are several use cases where a significantly improved way of interacting with the content is highly desirable. It is possible to organize a video in an interactive and non-linear way. Additional information (for example high resolution images) can be added to any scene of the video. The non-linearity of the video flow and the implementation of additional content not found in traditional videos may lead to an increased download volume and/or a playback with many breaks for downloading missing elements. This paper describes a player framework for interactive non-linear videos. We developed the framework and its associated algorithms to simulate the playback of non-linear video content. It minimizes interruption when the sequence of scenes is directly influenced by interaction, while the traditional viewing experience is not altered. The evaluation showed that it is possible to reach shorter startup times using our strategies than using the strategies of other players. Furthermore, we demonstrated that a prefetching of elements does not lead to an increased download volume in every case. In contrast, it can even decrease the download volume if the right delete strategy is selected. It can be noted that the knowledge of the structure of interactive non-linear videos can be used to minimize startup times at the beginning of scenes while the download volume is not increased.     

Herstellung und Charakterisierung von zellstrukturiertem Apfelmaterial mit unterschiedlichem Pectingehalt

Zusammenfassung Ausgehend von Apfelparenchymgewebe wurden alkoholunlösliche Substanzen als hellfarbige Materialien mit cellularer Struktur und unterschiedlichem Pectingehalt hergestellt. Die Materialien haben unabhängig vom Pectingehalt vergleichbare funktionelle Eigenschaften und besitzen ein hohes Wasserbindevermögen über 50 g/g. Jedoch wird das Wasserbindevermögen bei den pectinarmen Materialien durch den Schritt der Pectinextraktion herabgesetzt. Die Herstellung des zellstrukturierten Materials mit niedrigem Pectingehalt kann über den Schritt einer einstufigen sauren Pectinextraktion mit der Isolierung von hochverestertem, hochmolekularem Pectin verknüpft werden. Beim Autoklavieren von rehydratisierten zellstrukturierten Materialien werden beträchtliche Mengen an wasserlöslichem, hoch verestertem, niedermolekularem Pectin freigesetzt. Das Autoklavieren und nachfolgende Aufbewahren der Proben bei Raumtemperatur führt zu sinkenden Werten der Helligkeit und der Fließgrenze.

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Preparation and characterization of materials with a cellular structure and different pectin contents from apple

Alcohol-insoluble substances in the form of light (colourless) materials with a cellular structure and two different pectin contents have been prepared from apple parenchyma. Except for the water-binding capacity, the pectin content was found to have only a small effect on the functional properties of the materials. The water-binding capacity of the materials was decreased by a step of pectin extraction. However, in all cases the water-binding capacity was higher than 50 g/g. It was found that the depectinization of the materials with a cellular structure by acid one-step pectin extraction can be combined with the isolation of a highly methoxylated, high-molecular-mass pectin. Considerable amounts of water-soluble, highly methoxylated but low-molecular-mass pectins are released from rehydrated materials with a cellular structure when subjected to autoclave treatment. Both autoclaving and storage at room temperature decrease the lightness and the flow limit value of the rehydrated materials.

     

N-terminal myristoylation is required for membrane localization of cGMP-dependent protein kinase type II

The apical membrane of intestinal epithelial cells harbors a unique isozyme of cGMP-dependent protein kinase (cGK type II) which acts as a key regulator of ion transport systems, including the cystic fibrosis transmembrane conductance regulator (CFTR)-chloride channel. To explore the mechanism of cGK II membrane-anchoring, recombinant cGK II was expressed stably in HEK 293 cells or transiently in COS-1 cells. In both cell lines, cGK II was found predominantly in the particulate fraction. Immunoprecipitation of solubilized cGK II did not reveal any other tightly associated proteins, suggesting a membrane binding motif within cGK II itself. The primary structure of cGK II is devoid of hydrophobic transmembrane domains; cGK II does, however, contain a penultimate glycine, a potential acceptor for a myristoyl moiety. Metabolic labeling showed that cGK II was indeed able to incorporate [3H]myristate. Moreover, incubation of cGK II-expressing 293 cells with the myristoylation inhibitor 2-hydroxymyristic acid (1 mM) significantly increased the proportion of cGK II in the cytosol from 10 +/- 5 to 35 +/- 4%. Furthermore, a nonmyristoylated cGK II Gly2 --> Ala mutant was localized predominantly in the cytosol after transient expression in COS-1 cells. The absence of the myristoyl group did not affect the specific enzyme activity or the Ka for cGMP and only slightly enhanced the thermal stability of cGK II. These results indicate that N-terminal myristoylation fulfills a crucial role in directing cGK II to the membrane.     

Der Einfluß von Bakterien auf die Eisenkorrosion in Brack- und Meerwasser

Influences of bacteria on iron corrosion in brackish and sea waters The corrosion of steel in seawaters is subject to seasonal variations; the corrosion maxma being in spring and autumn. The variations are not attribuable to changes in water temperature, salt concentration or oxygen content but are due to the co-action of biotic and abiotic factors, the former givin rise to temperature dependent variations of corrosion. The activity of bacteria becomes bovious only at temperatures above 10 °C; at these temperatures metablolism is activated and the products of matabolism may attack iron (or bac-     

Nanoliter Scale Parallel Liquid–Liquid Extraction for High-Throughput Purification on a Droplet Microarray

In the current drug discovery process, the synthesis of compound libraries is separated from biological screenings both conceptually and technologically. One of the reasons is that parallel on-chip high-throughput purification of synthesized compounds is still a major challenge. Here, on-chip miniaturized high-throughput liquid–liquid extraction in volumes down to 150 nL with efficiency comparable to or better than large-scale extraction utilizing separation funnels is demonstrated. The method is based on automated and programmable merging of arrays of aqueous nanoliter droplets with organic droplets. Multi-step extraction performed simultaneously or with changing conditions as well as handling of femtomoles of compounds are demonstrated. In addition, the extraction efficiency is analyzed with a fast optical readout as well as matrix-assisted laser desorption ionization-mass spectrometry on-chip detection. The new massively parallel and miniaturized purification method adds another important tool to the chemBIOS concept combining chemical combinatorial synthesis with biological screenings on the same miniaturized droplet microarray platform, which will be essential to accelerate drug discovery.     

New DNA polymerase IIIC inhibitors: 3-subtituted anilinouracils with potent antibacterial activity in vitro and in vivo

The development of resistance has rendered several antibiotics clinically ineffective, and there is an urgent medical need for potent and safe antibacterials with a novel and valid mode of action. To avoid cross-resistance, they should preferably inhibit targets that are not addressed by established antibiotics. In this respect, 6-anilinouracils represent a promising lead structure. They target the Gram-positive DNA polymerase IIIC, a target that is associated with a bactericidal mode of action. Moreover, they have no cross-resistance to marketed antibiotics. This paper describes the synthesis and biological characterization of structurally novel anilinouracils, some of which display potent in vivo efficacy in murine models of bacterial septicemia.