TY - JOUR A1 - Bencurova, Elena A1 - Akash, Aman A1 - Dobson, Renwick C.J. A1 - Dandekar, Thomas T1 - DNA storage-from natural biology to synthetic biology JF - Computational and Structural Biotechnology Journal N2 - Natural DNA storage allows cellular differentiation, evolution, the growth of our children and controls all our ecosystems. Here, we discuss the fundamental aspects of DNA storage and recent advances in this field, with special emphasis on natural processes and solutions that can be exploited. We point out new ways of efficient DNA and nucleotide storage that are inspired by nature. Within a few years DNA-based information storage may become an attractive and natural complementation to current electronic data storage systems. We discuss rapid and directed access (e.g. DNA elements such as promotors, enhancers), regulatory signals and modulation (e.g. lncRNA) as well as integrated high-density storage and processing modules (e.g. chromosomal territories). There is pragmatic DNA storage for use in biotechnology and human genetics. We examine DNA storage as an approach for synthetic biology (e.g. light-controlled nucleotide processing enzymes). The natural polymers of DNA and RNA offer much for direct storage operations (read-in, read-out, access control). The inbuilt parallelism (many molecules at many places working at the same time) is important for fast processing of information. Using biology concepts from chromosomal storage, nucleic acid processing as well as polymer material sciences such as electronical effects in enzymes, graphene, nanocellulose up to DNA macramé , DNA wires and DNA-based aptamer field effect transistors will open up new applications gradually replacing classical information storage methods in ever more areas over time (decades). KW - DNA KW - RNA KW - data storage KW - natural processing KW - synthetic biology Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-349971 SN - 2001-0370 VL - 21 ER - TY - THES A1 - Esch, Mandy T1 - Novel Nucleic Acid Sensors for the Rapid Detection of Cryptosporidium Parvum T1 - Neue Nukleinsäure-Sensoren für die Detektion von Cryptosporidium parvum N2 - Recent advances in the development of immunoassays and nucleic acid assays have improved the performance and increased the sensitivity of sensors that are based on biochemical recognition. The new approaches taken by researchers include detecting pathogens by detecting their nucleic acids, using new nontoxic reporter entities for generating signals, and downscaling and miniaturizing sensors to micromigration and microfluidic formats. This dissertation connects some of these successful approaches, thereby leading to the development of novel nucleic acid sensors for rapid and easy detection of pathogens. The author's goal was to develop diagnostic tools that enable investigators to detect pathogens rapidly and on site. While the sensors can be used to detect any pathogen, the author first customized them for detecting particularly Cryptosporidium parvum, a pathogen whose detection is important, yet presents many challenges. Chapter 2 of this thesis presents a novel test-strip for the detection of C. parvum. The test-strip is designed to detect nucleic acids rather than proteins or other epitopes. While test strips are commonly used for sensors based on immunological recognition, this format is very new in applications in which nucleic acids are detected. Further, to indicate the presence or absence of a specific target on the test strip, dye-entrapped, oligonucleotide-tagged liposomes are employed. Using liposomes as reporter particles has advantages over using other reporter labels, because the cavity that the phospholipidic membranes of the liposomes form can be filled with up to 106 dye molecules. By using heterobifunctional linkers liposomes can be tagged with oligonucleotides, thereby enabling their use in nucleic acid hybridization assays. The developed test-strip provides an internal control. The limit of detection is 2.7 fmol/mL with a sample volume of 30 mL. In chapter 3 the detection of nucleic acids by means of oligonucleotide-tagged liposomes is scaled down to a microfluidic assay format. Because the application of biosensors to microfluidic formats is very new in the field of analytical chemistry, the first part of this chapter is devoted to developing the design and the method to fabricate the microchip devices. The performance of the microchips is then optimized by investigating the interactions of nucleic acids and liposomes with the material the chips consist of and by passivating the surface of the chips with blocking reagents. The developed microfluidic chip enabled us to reduce the sample volume needed for one assay to 12.5 mL. The limit of detection of this assay was determined to be 0.4 fmol/mL. Chapters 4 and 5 expand on the development of the microfluidic assay. A prototype microfluidic array that is able to detect multiple analytes in a single sample simultaneously is developed. Using such an array will enable investigators to detect pathogens that occur in the same environment, for example, C. parvum and Giardia duodenalis by conducting a single test. The array's ability to perform multiple sample analysis is shown by detecting different concentrations of target nucleic acids. Further, the author developed a microfluidic chip in which interdigitated microelectrode arrays (IDAs) that consist of closely spaced microelectrodes are integrated. The IDAs facilitate electrochemical detection of cryptosporidial RNA. Electrochemical detection schemes offer benefits of technical simplicity, speed, and sensitivity. In this project liposomes are filled with electrochemically active molecules and are then utilized to generate electrochemical signals. Chapter 6 explores the feasibility of liposomes for enhancing signals derived from nucleic acid hybridization in surface plasmon resonance (SPR) spectroscopy. SPR spectroscopy offers advantages because nucleic acid hybridization can be monitored in real time and under homogeneous conditions because no washing steps are required. SPR spectroscopy is very sensitive and it can be expected that, in the future, SPR will be integrated into microfluidic nucleic acid sensors. N2 - Jüngste Fortschritte in der Entwicklung von Immuno- und Nucleinsäure- Assays haben die Arbeitsleistung und die Spezifität von Sensoren, die auf biochemischer Erkennung basieren (Biosensoren), verbessert. Neu entwickelte Methoden umfassen die Detektion von Pathogenen durch die Detektion ihrer RNA oder DNA, das Benutzen von neuen nicht-toxischen Reporter Molekülen, um Signale in Sensoren zu erzeugen, und die Verkleinerung und Miniaturisierung von Sensoren zu Mikromigrations- und Mikrofluid Formaten. Die in dieser Dissertation entwickelten Sensoren, die der Detektion von Pathogenen dienen, verbinden einige der neu entwickelten Methoden. Das Ziel der Autorin war es, Sensoren zu entwickeln, die es ermöglichen, Pathogene an Ort und Stelle zu detektieren. Die entwickelten Sensoren können zur Detektion von einer Reihe von Pathogenen benutzt werden. In dieser Dissertation sind sie für die spezifische Detektion von Cryptosporidium parvum entwickelte worden. Kapitel 2 der Dissertation präsentiert einen neuen Teststreifen für die Detektion von C. parvum. Der Teststreifen detektiert die RNA von C. parvum, die als Reaktion auf einen Hitzeschock produziert wird. Das Teststreifen-Format ist üblich für Sensoren, die auf immunologischer Erkennung basieren. Es ist jedoch neu für Anwendungen in denen RNA oder DNA detektiert werden sollen. Die An- oder Abwesenheit eines bestimmten Ziel Moleküls wird durch Liposomen, die Oligonukleotide auf der Aussenseite ihrer Membranen enthalten und mit Farbstoff gefüllt sind, angedeutet. Die Experimente zeigten, dass die mit dem entwickelten Test-Streifen kleinste detektierbare Konzentration von RNA in einem 30 mL Probenvolumen 2.7 fmol/mL ist. In Kapitel 3 ist die Signalerzeugung durch Liposomen in ein Mikrofliess-System integriert. Da die Entwicklung von Mikrofliess-Systemen ein sehr neues Forschungsgebiet ist, befasst sich ein Teil dieses Kapitels mit dem Design und der Herstellung des Microchips. Die Untersuchung von Interaktionen von Nukleinsäuren und Liposomen mit dem Material aus dem der Chip hergestellt ist und die Passivierung dieses Materials ist dabei ein Schwerpunkt. Das Probenvolumen, dass zur Detektion mit dem entwickelten Mikrofliess-Sensor nötig ist, konnte auf 12.5 mL reduziert werden. Die kleinste detektierbare Konzentration von Nucleinsäuren ist 5 fmol/mL. In Kapitel 4 und 5 erweitert die Autorin die Entwicklung des Mikrofliess-Sensors aus Kapitel 3. Das Detektionsformat ist auf ein Array, das für die gleichzeitige Detektion von mehreren Pathogenen benutzt werden kann, angewandt. Eine Methode zum Herstellen eines Arrays-Prototypen ist entwickelt. Ferner, stellte die Autorin verzahnte Mikroelektroden her und benutzte diese um die elektrochemische Detektion der RNA von C. parvum zu ermöglichen. In Kapitel 6 ist die Anwendbarkeit von Liposomen zur Erhöhung von Signalen von Nukleinsäure-Hybridisierungen in Surface Plasmon Resonance Spectroscopy (SPR) untersucht. KW - Cryptosporidium KW - RNS KW - Biosensor KW - Nucleinsäure-Sensoren KW - RNA KW - Cryptosporidium parvum KW - Mikrofliess-System KW - Liposomen KW - Teststreifen KW - Nuleic Acids Sensors KW - RNA KW - Cryptosporidium parvum KW - Microfluidic Chip KW - Liposomes KW - Test-Strip Y1 - 2001 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-323 ER - TY - JOUR A1 - Bakari-Soale, Majeed A1 - Ikenga, Nonso Josephat A1 - Scheibe, Marion A1 - Butter, Falk A1 - Jones, Nicola G. A1 - Kramer, Susanne A1 - Engstler, Markus T1 - The nucleolar DExD/H protein Hel66 is involved in ribosome biogenesis in Trypanosoma brucei JF - Scientific Reports N2 - The biosynthesis of ribosomes is a complex cellular process involving ribosomal RNA, ribosomal proteins and several further trans-acting factors. DExD/H box proteins constitute the largest family of trans-acting protein factors involved in this process. Several members of this protein family have been directly implicated in ribosome biogenesis in yeast. In trypanosomes, ribosome biogenesis differs in several features from the process described in yeast. Here, we have identified the DExD/H box helicase Hel66 as being involved in ribosome biogenesis. The protein is unique to Kinetoplastida, localises to the nucleolus and its depletion via RNAi caused a severe growth defect. Loss of the protein resulted in a decrease of global translation and accumulation of rRNA processing intermediates for both the small and large ribosomal subunits. Only a few factors involved in trypanosome rRNA biogenesis have been described so far and our findings contribute to gaining a more comprehensive picture of this essential process. KW - infection KW - parasite evolution KW - parasite genetics KW - RNA Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-263872 VL - 11 IS - 1 ER - TY - JOUR A1 - Förster, Frank A1 - Beisser, Daniela A1 - Grohme, Markus A. A1 - Liang, Chunguang A1 - Mali, Brahim A1 - Siegl, Alexander Matthias A1 - Engelmann, Julia C. A1 - Shkumatov, Alexander V. A1 - Schokraie, Elham A1 - Müller, Tobias A1 - Schnölzer, Martina A1 - Schill, Ralph O. A1 - Frohme, Marcus A1 - Dandekar, Thomas T1 - Transcriptome analysis in tardigrade species reveals specific molecular pathways for stress adaptations JF - Bioinformatics and biology insights N2 - Tardigrades have unique stress-adaptations that allow them to survive extremes of cold, heat, radiation and vacuum. To study this, encoded protein clusters and pathways from an ongoing transcriptome study on the tardigrade \(Milnesium\) \(tardigradum\) were analyzed using bioinformatics tools and compared to expressed sequence tags (ESTs) from \(Hypsibius\) \(dujardini\), revealing major pathways involved in resistance against extreme environmental conditions. ESTs are available on the Tardigrade Workbench along with software and databank updates. Our analysis reveals that RNA stability motifs for \(M.\) \(tardigradum\) are different from typical motifs known from higher animals. \(M.\) \(tardigradum\) and \(H.\) \(dujardini\) protein clusters and conserved domains imply metabolic storage pathways for glycogen, glycolipids and specific secondary metabolism as well as stress response pathways (including heat shock proteins, bmh2, and specific repair pathways). Redox-, DNA-, stress- and protein protection pathways complement specific repair capabilities to achieve the strong robustness of \(M.\) \(tardigradum\). These pathways are partly conserved in other animals and their manipulation could boost stress adaptation even in human cells. However, the unique combination of resistance and repair pathways make tardigrades and \(M.\) \(tardigradum\) in particular so highly stress resistant. KW - RNA KW - expressed sequence tag KW - cluster KW - protein familiy KW - adaption KW - tardigrada KW - transcriptome Y1 - 2012 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-123089 N1 - This is an open access article. Unrestricted non-commercial use is permitted provided the original work is properly cited. VL - 6 ER -