Handling of long DNA - applications and polymer physics

This thesis provides a proof of principle that deterministic lateral displacement (DLD) can be used to spatially separate DNA molecules by size. In order to achieve high separation quality, the pressure range, i.e. flow velocity has to be chosen carefully. The experiments were conducted in DLD devices fabricated in PDMS using soft lithography methods. Fluorescently labeled DNA molecules were transported through the device by pressure driven flow. The separation results were determined from and the separation result was recorded optically at the outlet. Both tested devices, with a critical diameter Dc of 0.64 μm and 0.75 μm respectively, showed capable of separating <10 kbp from 48.5 kbp DNA molecules with a separation quality of ~94%.DNA is the blueprint for life and contains all necessary information to ’build’ an organism. Therefore, DNA might be considered as the most important biologically relevant molecule. However, even though DNA is such a substantial molecule, the research methods and techniques regarding DNA are often surprisingly limited which is due to the complicated handling of the long molecule. This is also the case for DNA sorting methods, i.e the process of separating different DNA molecules from each other dependent on characteristic factors like weight or length. Sorting is especially important as pre-processing method for other established analysis techniques. Currently, the length-based sorting of DNA molecules is predominantly done using electrophoresis, for which a huge DNA sample volume with roughly a million molecules is required. Microfluidics is a relatively new, expanding field in biophysics which deals with fluids on the microliter and micrometer scale. Microfluidics are contrasted by macrofluidics which is our ’normal’ perception of fluids. An easy example which points out the difference is the surface tension of fluids. In every day life we experience it only as a side effect, but when dealing with cellular volumes, typically 1 μl which is one millionth of a liter, the surface tension wins over gravity. One application of microfluidics is the deterministic lateral displacement method, also called DLD, which was introduced by Richard Huang et al. in 2004. It is based on a micrometer-sized channel in which small obstacles are placed. If a fluid which also contains small particles flows through this obstacle-larded channel, the track of the particles vary dependent on their size: while very small particles follow the fluid flow and pass straight through the device, slightly larger particles get deflected to one of the channel walls. Thus, differently sized particles can be distinguished at the end of the channel as they get spatially separated. During the following master thesis, the sorting of DNA molecules dependent on their length was investigated. For this, the described DLD method was applied to DNA molecules in solution. It was found that DLD - if used under correct conditions - is an appropriate method to sort DNA molecules dependent on their length. This finding can be used in sample preparation and processing steps for DNA related experiments. Furthermore, since this is a microfluidic approach it allows DNA length separation for extremely small sample sizes, with only a couple of hundreds of molecules. Especially considering advancing single molecule analysis techniques and personalized medicine approaches, a cut-down sample volume will be crucial in future developments

A machine learning based 24-h-technique for an area-wide rainfall retrieval using MSG SEVIRI data over Central Europe

The aim of the present study was to develop a 24-h-technique for the process-related and quantitative estimation of precipitation in connection with extra-tropical cyclones in the mid-latitudes based on MSG SEVIRI data using the machine learning algorithm random forest. The algorithms and approaches needed were successfully developed and implemented within three working packages: (WP1) The cloud property retrieval SLALOM, first developed for Terra MODIS, was successfully transferred and adapted to the specific requirements of the SEVIRI system and an extensive validation study was carried out. The cloud optical properties retrieved by SLALOM, namely cloud effective radius and cloud optical thickness that were needed for satellitebased rainfall estimation in WP2 and WP3, were compared against the well known and validated NASA MODIS cloud property product (MODIS 06) as well as the cloud optical depth product (2B-TAU) of CloudSat. The suitability of SLALOM has been shown over the North Atlantic and over the European continent (chapter 3). (WP2) A new 24-h-technique for rainfall rate assignment was developed for MSG SEVIRI using the machine learning algorithm random forest as fundamental prediction algorithm. Based on the precipitation processes in connection with extra-tropical cyclones, rainfall rates were assigned to advectivestratiform and convective precipitating areas by means of individual RF models. As predictor variables for the RF models satellite-based information on cloud top height, cloud top temperature, cloud phase and cloud water path were chosen. The different illumination conditions (daytime, twilight and night-time) were taken into account with a proper SEVIRI spectral channel selection as surrogates for theses cloud physical parameters. The development was realised in three steps: First, an extensive tuning study was carried out to customise each of the RF models. Secondly, the RF models were trained using the optimum model parameter values found in the tuning study. Finally, the final RF models were used to predict rainfall rates using an independent validation data set and the results were validated against co-located rainfall rates observed by the RADOLAN RW product of the DWD. The outstanding validation results during all times of the day confirmed the ability of RF as tool for the rainfall rate assignment technique from MSG SEVIRI data (chapter 4). (WP3) A new coherent daytime, twilight and night-time rainfall retrieval was developed for MSG SEVIRI. The technique aims to retrieve rainfall rates for precipitation events in connection with extra-tropical cyclones in the midlatitudes in a continuous manner resulting in a 24 hour prediction. Based on the dominant precipitation processes, the proposed rainfall retrieval consists of three steps which are applied consecutively by means of individual RF models to get the final product: (i) Identification of precipitating cloud areas. (ii) Separation of precipitating areas into predominately convective and advective-stratiform cloud regions. (iii) Individual process-oriented assignment of rainfall rates to these cloud areas. Again, the relationship between cloud top temperature, cloud top height, cloud water path and cloud phase was used to retrieve information about precipitation and according to the illumination conditions, a suitable selection of the predictor variables were taken into account as input to the RF models (chapter 5). The newly developed rainfall retrieval technique was tested in an extensive validation study over Germany using the radar-based RADOLAN RW product as reference data. The validation results show reliable performance of the new technique concerning rain area detection, rain process separation as well as rainfall rate assignment during all times of the day which enables the estimation of precipitation for 24 hours of a day. Hereby, the twilight applicability of the technique as well as good rainfall rate prediction performances even on an hourly basis are particularly remarkable and set this study apart from other rainfall retrievals. For the first time, a 24-h precipitation monitoring becomes possible for precipitating clouds of not only convective but also of advective-stratiform character, opening many areas of application

Lipid droplet-based storage fat metabolism in Drosophila.

The fruit fly Drosophila melanogaster is an emerging model system in lipid metabolism research. Lipid droplets are omnipresent and dynamically regulated organelles found in various cell types throughout the complex life cycle of this insect. The vital importance of lipid droplets as energy resources and storage compartments for lipoanabolic components has recently attracted research attention to the basic enzymatic machinery, which controls the delicate balance between triacylglycerol deposition and mobilization in flies. This review aims to present current insights in experimentally supported and inferred biological functions of lipogenic and lipolytic enzymes as well as regulatory proteins, which control the lipid droplet-based storage fat turnover in Drosophila

A point-based event phonology for the phonetics-phonology interface

Wagner P, Kühnlein P. A point-based event phonology for the phonetics-phonology interface. In: Proceedings of the Joint Conference on Formal Grammar, Head Driven Phrase Structure Grammar, and Categorial Grammar. Saarbrücken, Germany; 1998: 1-10.Recent rethinking of the Phonetics/Phonology-Interface has come to drop the old notion of a syntactic relationship between both research domains and replaced it by a semantic one. However, it remained relatively unclear how this relationship ought to look like. Progress in diagrammatic reasoning provides a fruitful approach towards a mutually constraining interface description. But to utilise this tool we first have to argue that it is reasonable to suppose a common ontology of a certain variant for both fiels of research. This is so, bacause the heterogenous logics-approach attempted here relies on the sharing of models

tRNA sequences can assemble into a replicator

Can replication and translation emerge in a single mechanism via self-assembly? The key molecule, transfer RNA (tRNA), is one of the most ancient molecules and contains the genetic code. Our experiments show how a pool of oligonucleotides, adapted with minor mutations from tRNA, spontaneously formed molecular assemblies. They replicated information autonomously using only reversible hybridization under thermal oscillations. A pool of cross-complementary hairpins self-selected by agglomeration and sedimented under gravity. The metastable DNA hairpins bound to a template, consisting of one half of the hairpin assembly, and then interconnected by hybridization. Thermal oscillations separated replicates from their templates and drove an exponential, cross-catalytic replication. The molecular assembly could encode and replicate binary sequence information and reach a fidelity of 90 % per nucleotide. This mechanism of a replicating self-assembly of tRNA-like sequences indicates that the translation to proteins could be linked closer to molecular replication than previously thought