TY - GEN A1 - Baumann, Tobias A1 - Arndt, Katja Maren A1 - Müller, Kristian M. T1 - Directional cloning of DNA fragments using deoxyinosine-containing oligonucleotides and endonuclease V T2 - Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe N2 - Background: DNA fragments carrying internal recognition sites for the restriction endonucleases intended for cloning into a target plasmid pose a challenge for conventional cloning. Results: A method for directional insertion of DNA fragments into plasmid vectors has been developed. The target sequence is amplified from a template DNA sample by PCR using two oligonucleotides each containing a single deoxyinosine base at the third position from the 5' end. Treatment of such PCR products with endonuclease V generates 3' protruding ends suitable for ligation with vector fragments created by conventional restriction endonuclease reactions. Conclusions: The developed approach generates terminal cohesive ends without the use of Type II restriction endonucleases, and is thus independent from the DNA sequence. Due to PCR amplification, minimal amounts of template DNA are required. Using the robust Taq enzyme or a proofreading Pfu DNA polymerase mutant, the method is applicable to a broad range of insert sequences. Appropriate primer design enables direct incorporation of terminal DNA sequence modifications such as tag addition, insertions, deletions and mutations into the cloning strategy. Further, the restriction sites of the target plasmid can be either retained or removed. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 983 KW - cohesive ends KW - DNA cleavage KW - genetic vectors KW - modified primers KW - molecular methods KW - polymerase chain reaction KW - recombinant Escherichia coli KW - restriction enzymes Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-431085 SN - 1866-8372 IS - 983 ER - TY - JOUR A1 - Feiner, Rebecca Christine A1 - Teschner, Julian A1 - Teschner, Kathrin E. A1 - Radukic, Marco T. A1 - Baumann, Tobias A1 - Hagen, Sven A1 - Hannappel, Yvonne A1 - Biere, Niklas A1 - Anselmetti, Dario A1 - Arndt, Katja Maren A1 - Müller, Kristian Mark T1 - rAAV Engineering for Capsid-Protein Enzyme Insertions and Mosaicism Reveals Resilience to Mutational, Structural and Thermal Perturbations JF - International journal of molecular sciences N2 - Recombinant adeno-associated viruses (rAAV) provide outstanding options for customization and superior capabilities for gene therapy. To access their full potential, facile genetic manipulation is pivotal, including capsid loop modifications. Therefore, we assessed capsid tolerance to modifications of the structural VP proteins in terms of stability and plasticity. Flexible glycine-serine linkers of increasing sizes were, at the genetic level, introduced into the 587 loop region of the VP proteins of serotype 2, the best studied AAV representative. Analyses of biological function and thermal stability with respect to genome release of viral particles revealed structural plasticity. In addition, insertion of the 29 kDa enzyme beta-lactamase into the loop region was tested with a complete or a mosaic modification setting. For the mosaic approach, investigation of VP2 trans expression revealed that a Kozak sequence was required to prevent leaky scanning. Surprisingly, even the full capsid modification with beta-lactamase allowed for the assembly of capsids with a concomitant increase in size. Enzyme activity assays revealed lactamase functionality for both rAAV variants, which demonstrates the structural robustness of this platform technology. KW - adeno-associated-virus KW - beta-lactamase KW - inverted terminal repeat (ITR) KW - loop modification KW - capsid stability Y1 - 2019 U6 - https://doi.org/10.3390/ijms20225702 SN - 1422-0067 VL - 20 IS - 22 PB - MDPI CY - Basel ER - TY - JOUR A1 - Baumann, Tobias A1 - Arndt, Katja Maren A1 - Müller, Kristian M. T1 - Directional cloning of DNA fragments using deoxyinosine-containing oligonucleotides and endonuclease V JF - BMC biotechnology N2 - Background: DNA fragments carrying internal recognition sites for the restriction endonucleases intended for cloning into a target plasmid pose a challenge for conventional cloning. Results: A method for directional insertion of DNA fragments into plasmid vectors has been developed. The target sequence is amplified from a template DNA sample by PCR using two oligonucleotides each containing a single deoxyinosine base at the third position from the 5' end. Treatment of such PCR products with endonuclease V generates 3' protruding ends suitable for ligation with vector fragments created by conventional restriction endonuclease reactions. Conclusions: The developed approach generates terminal cohesive ends without the use of Type II restriction endonucleases, and is thus independent from the DNA sequence. Due to PCR amplification, minimal amounts of template DNA are required. Using the robust Taq enzyme or a proofreading Pfu DNA polymerase mutant, the method is applicable to a broad range of insert sequences. Appropriate primer design enables direct incorporation of terminal DNA sequence modifications such as tag addition, insertions, deletions and mutations into the cloning strategy. Further, the restriction sites of the target plasmid can be either retained or removed. KW - Cohesive ends KW - DNA cleavage KW - Genetic vectors KW - Modified primers KW - Molecular methods KW - Polymerase chain reaction KW - Recombinant Escherichia coli KW - Restriction enzymes Y1 - 2013 U6 - https://doi.org/10.1186/1472-6750-13-81 SN - 1472-6750 VL - 13 IS - 10 PB - BioMed Central CY - London ER - TY - JOUR A1 - Jedrusik-Bode, Monika A1 - Studencka, Maja A1 - Smolka, Christian A1 - Baumann, Tobias A1 - Schmidt, Henning A1 - Kampf, Jan A1 - Paap, Franziska A1 - Martin, Sophie A1 - Tazi, Jamal A1 - Müller, Kristian M. A1 - Krüger, Marcus A1 - Braun, Thomas A1 - Bober, Eva T1 - The sirtuin SIRT6 regulates stress granule formation in C. elegans and mammals JF - Journal of cell science N2 - SIRT6 is a NAD(+)-dependent deacetylase that modulates chromatin structure and safeguards genomic stability. Until now, SIRT6 has been assigned to the nucleus and only nuclear targets of SIRT6 are known. Here, we demonstrate that in response to stress, C. elegans SIR-2.4 and its mammalian orthologue SIRT6 localize to cytoplasmic stress granules, interact with various stress granule components and induce their assembly. Loss of SIRT6 or inhibition of its catalytic activity in mouse embryonic fibroblasts impairs stress granule formation and delays disassembly during recovery, whereas deficiency of SIR-2.4 diminishes maintenance of P granules and decreases survival of C. elegans under stress conditions. Our findings uncover a novel, evolutionary conserved function of SIRT6 in the maintenance of stress granules in response to stress. KW - C. elegans KW - G3BP KW - SIRT6 KW - Sirtuins KW - Stress KW - Stress granules Y1 - 2013 U6 - https://doi.org/10.1242/jcs.130708 SN - 0021-9533 SN - 1477-9137 VL - 126 IS - 22 SP - 5166 EP - + PB - Company of Biologists Limited CY - Cambridge ER - TY - THES A1 - Baumann, Tobias T1 - Stability and Interconnected protein properties studied with TEM ß-lactamase Y1 - 2013 CY - Potsdam ER - TY - JOUR A1 - Hagen, Sven A1 - Baumann, Tobias A1 - Wagner, Hanna J. A1 - Morath, Volker A1 - Kaufmann, Beate A1 - Fischer, Adrian A1 - Bergmann, Stefan A1 - Schindler, Patrick A1 - Arndt, Katja Maren A1 - Mueller, Kristian M. T1 - Modular adeno-associated virus (rAAV) vectors used for cellular virus-directed enzyme prodrug therapy JF - Scientific reports N2 - The pre-clinical and clinical development of viral vehicles for gene transfer increased in recent years, and a recombinant adeno-associated virus (rAAV) drug took center stage upon approval in the European Union. However, lack of standardization, inefficient purification methods and complicated retargeting limit general usability. We address these obstacles by fusing rAAV-2 capsids with two modular targeting molecules (DARPin or Affibody) specific for a cancer cell-surface marker (EGFR) while simultaneously including an affinity tag (His-tag) in a surface-exposed loop. Equipping these particles with genes coding for prodrug converting enzymes (thymidine kinase or cytosine deaminase) we demonstrate tumor marker specific transduction and prodrug-dependent apoptosis of cancer cells. Coding terminal and loop modifications in one gene enabled specific and scalable purification. Our genetic parts for viral production adhere to a standardized cloning strategy facilitating rapid prototyping of virus directed enzyme prodrug therapy (VDEPT). Y1 - 2014 U6 - https://doi.org/10.1038/srep03759 SN - 2045-2322 VL - 4 PB - Nature Publ. Group CY - London ER - TY - JOUR A1 - Kuekenshoener, Tim A1 - Hagemann, Urs B. A1 - Wohlwend, Daniel A1 - Raeuber, Christina A1 - Baumann, Tobias A1 - Keller, Sandro A1 - Einsle, Oliver A1 - Mueller, Kristian M. A1 - Arndt, Katja Maren T1 - Analysis of Selected and Designed Chimeric D- and L-alpha-Helix Assemblies JF - Biomacromolecules : an interdisciplinary journal focused at the interface of polymer science and the biological sciences N2 - D-Peptides have been attributed pharmacological advantages over regular L-peptides, yet design rules are largely unknown. Based on a designed coiled coil-like D/L heterotetramer, named L-Base/D-Acid, we generated a library offering alternative residues for interaction with the D-peptide. Phage display selection yielded one predominant peptide, named HelixA, that differed at 13 positions from the scaffold helix. In addition to the observed D-/L-heterotetramers, ratio-dependent intermediate states were detected by isothermal titration calorimetry. Importantly, the formation of the selected HelixA/D-Acid bundle passes through fewer intermediate states than L-Base/D-Acid. Back mutation of HelixA core residues to L-Base (HelixLL) revealed that the residues at e/g-positions are responsible for the different intermediates. Furthermore, a Val-core variant (PeptideVV) was completely devoid of binding D-Acid, whereas an Ile-core helix (HelixII) interacted with D-Acid in a significantly more specific complex than L-Base. Y1 - 2014 U6 - https://doi.org/10.1021/bm5006883 SN - 1525-7797 SN - 1526-4602 VL - 15 IS - 9 SP - 3296 EP - 3305 PB - American Chemical Society CY - Washington ER - TY - BOOK A1 - Zhang, Shuhao A1 - Plauth, Max A1 - Eberhardt, Felix A1 - Polze, Andreas A1 - Lehmann, Jens A1 - Sejdiu, Gezim A1 - Jabeen, Hajira A1 - Servadei, Lorenzo A1 - Möstl, Christian A1 - Bär, Florian A1 - Netzeband, André A1 - Schmidt, Rainer A1 - Knigge, Marlene A1 - Hecht, Sonja A1 - Prifti, Loina A1 - Krcmar, Helmut A1 - Sapegin, Andrey A1 - Jaeger, David A1 - Cheng, Feng A1 - Meinel, Christoph A1 - Friedrich, Tobias A1 - Rothenberger, Ralf A1 - Sutton, Andrew M. A1 - Sidorova, Julia A. A1 - Lundberg, Lars A1 - Rosander, Oliver A1 - Sköld, Lars A1 - Di Varano, Igor A1 - van der Walt, Estée A1 - Eloff, Jan H. P. A1 - Fabian, Benjamin A1 - Baumann, Annika A1 - Ermakova, Tatiana A1 - Kelkel, Stefan A1 - Choudhary, Yash A1 - Cooray, Thilini A1 - Rodríguez, Jorge A1 - Medina-Pérez, Miguel Angel A1 - Trejo, Luis A. A1 - Barrera-Animas, Ari Yair A1 - Monroy-Borja, Raúl A1 - López-Cuevas, Armando A1 - Ramírez-Márquez, José Emmanuel A1 - Grohmann, Maria A1 - Niederleithinger, Ernst A1 - Podapati, Sasidhar A1 - Schmidt, Christopher A1 - Huegle, Johannes A1 - de Oliveira, Roberto C. L. A1 - Soares, Fábio Mendes A1 - van Hoorn, André A1 - Neumer, Tamas A1 - Willnecker, Felix A1 - Wilhelm, Mathias A1 - Kuster, Bernhard ED - Meinel, Christoph ED - Polze, Andreas ED - Beins, Karsten ED - Strotmann, Rolf ED - Seibold, Ulrich ED - Rödszus, Kurt ED - Müller, Jürgen T1 - HPI Future SOC Lab – Proceedings 2017 T1 - HPI Future SOC Lab – Proceedings 2017 N2 - The “HPI Future SOC Lab” is a cooperation of the Hasso Plattner Institute (HPI) and industry partners. Its mission is to enable and promote exchange and interaction between the research community and the industry partners. The HPI Future SOC Lab provides researchers with free of charge access to a complete infrastructure of state of the art hard and software. This infrastructure includes components, which might be too expensive for an ordinary research environment, such as servers with up to 64 cores and 2 TB main memory. The offerings address researchers particularly from but not limited to the areas of computer science and business information systems. Main areas of research include cloud computing, parallelization, and In-Memory technologies. This technical report presents results of research projects executed in 2017. Selected projects have presented their results on April 25th and November 15th 2017 at the Future SOC Lab Day events. N2 - Das Future SOC Lab am HPI ist eine Kooperation des Hasso-Plattner-Instituts mit verschiedenen Industriepartnern. Seine Aufgabe ist die Ermöglichung und Förderung des Austausches zwischen Forschungsgemeinschaft und Industrie. Am Lab wird interessierten Wissenschaftlern eine Infrastruktur von neuester Hard- und Software kostenfrei für Forschungszwecke zur Verfügung gestellt. Dazu zählen teilweise noch nicht am Markt verfügbare Technologien, die im normalen Hochschulbereich in der Regel nicht zu finanzieren wären, bspw. Server mit bis zu 64 Cores und 2 TB Hauptspeicher. Diese Angebote richten sich insbesondere an Wissenschaftler in den Gebieten Informatik und Wirtschaftsinformatik. Einige der Schwerpunkte sind Cloud Computing, Parallelisierung und In-Memory Technologien. In diesem Technischen Bericht werden die Ergebnisse der Forschungsprojekte des Jahres 2017 vorgestellt. Ausgewählte Projekte stellten ihre Ergebnisse am 25. April und 15. November 2017 im Rahmen der Future SOC Lab Tag Veranstaltungen vor. T3 - Technische Berichte des Hasso-Plattner-Instituts für Digital Engineering an der Universität Potsdam - 130 KW - Future SOC Lab KW - research projects KW - multicore architectures KW - In-Memory technology KW - cloud computing KW - machine learning KW - artifical intelligence KW - Future SOC Lab KW - Forschungsprojekte KW - Multicore Architekturen KW - In-Memory Technologie KW - Cloud Computing KW - maschinelles Lernen KW - Künstliche Intelligenz Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-433100 SN - 978-3-86956-475-3 SN - 1613-5652 SN - 2191-1665 IS - 130 PB - Universitätsverlag Potsdam CY - Potsdam ER - TY - BOOK A1 - Rana, Kaushik A1 - Mohapatra, Durga Prasad A1 - Sidorova, Julia A1 - Lundberg, Lars A1 - Sköld, Lars A1 - Lopes Grim, Luís Fernando A1 - Sampaio Gradvohl, André Leon A1 - Cremerius, Jonas A1 - Siegert, Simon A1 - Weltzien, Anton von A1 - Baldi, Annika A1 - Klessascheck, Finn A1 - Kalancha, Svitlana A1 - Lichtenstein, Tom A1 - Shaabani, Nuhad A1 - Meinel, Christoph A1 - Friedrich, Tobias A1 - Lenzner, Pascal A1 - Schumann, David A1 - Wiese, Ingmar A1 - Sarna, Nicole A1 - Wiese, Lena A1 - Tashkandi, Araek Sami A1 - van der Walt, Estée A1 - Eloff, Jan H. P. A1 - Schmidt, Christopher A1 - Hügle, Johannes A1 - Horschig, Siegfried A1 - Uflacker, Matthias A1 - Najafi, Pejman A1 - Sapegin, Andrey A1 - Cheng, Feng A1 - Stojanovic, Dragan A1 - Stojnev Ilić, Aleksandra A1 - Djordjevic, Igor A1 - Stojanovic, Natalija A1 - Predic, Bratislav A1 - González-Jiménez, Mario A1 - de Lara, Juan A1 - Mischkewitz, Sven A1 - Kainz, Bernhard A1 - van Hoorn, André A1 - Ferme, Vincenzo A1 - Schulz, Henning A1 - Knigge, Marlene A1 - Hecht, Sonja A1 - Prifti, Loina A1 - Krcmar, Helmut A1 - Fabian, Benjamin A1 - Ermakova, Tatiana A1 - Kelkel, Stefan A1 - Baumann, Annika A1 - Morgenstern, Laura A1 - Plauth, Max A1 - Eberhard, Felix A1 - Wolff, Felix A1 - Polze, Andreas A1 - Cech, Tim A1 - Danz, Noel A1 - Noack, Nele Sina A1 - Pirl, Lukas A1 - Beilharz, Jossekin Jakob A1 - De Oliveira, Roberto C. L. A1 - Soares, Fábio Mendes A1 - Juiz, Carlos A1 - Bermejo, Belen A1 - Mühle, Alexander A1 - Grüner, Andreas A1 - Saxena, Vageesh A1 - Gayvoronskaya, Tatiana A1 - Weyand, Christopher A1 - Krause, Mirko A1 - Frank, Markus A1 - Bischoff, Sebastian A1 - Behrens, Freya A1 - Rückin, Julius A1 - Ziegler, Adrian A1 - Vogel, Thomas A1 - Tran, Chinh A1 - Moser, Irene A1 - Grunske, Lars A1 - Szárnyas, Gábor A1 - Marton, József A1 - Maginecz, János A1 - Varró, Dániel A1 - Antal, János Benjamin ED - Meinel, Christoph ED - Polze, Andreas ED - Beins, Karsten ED - Strotmann, Rolf ED - Seibold, Ulrich ED - Rödszus, Kurt ED - Müller, Jürgen T1 - HPI Future SOC Lab – Proceedings 2018 N2 - The “HPI Future SOC Lab” is a cooperation of the Hasso Plattner Institute (HPI) and industry partners. Its mission is to enable and promote exchange and interaction between the research community and the industry partners. The HPI Future SOC Lab provides researchers with free of charge access to a complete infrastructure of state of the art hard and software. This infrastructure includes components, which might be too expensive for an ordinary research environment, such as servers with up to 64 cores and 2 TB main memory. The offerings address researchers particularly from but not limited to the areas of computer science and business information systems. Main areas of research include cloud computing, parallelization, and In-Memory technologies. This technical report presents results of research projects executed in 2018. Selected projects have presented their results on April 17th and November 14th 2017 at the Future SOC Lab Day events. N2 - Das Future SOC Lab am HPI ist eine Kooperation des Hasso-Plattner-Instituts mit verschiedenen Industriepartnern. Seine Aufgabe ist die Ermöglichung und Förderung des Austausches zwischen Forschungsgemeinschaft und Industrie. Am Lab wird interessierten Wissenschaftler:innen eine Infrastruktur von neuester Hard- und Software kostenfrei für Forschungszwecke zur Verfügung gestellt. Dazu zählen Systeme, die im normalen Hochschulbereich in der Regel nicht zu finanzieren wären, bspw. Server mit bis zu 64 Cores und 2 TB Hauptspeicher. Diese Angebote richten sich insbesondere an Wissenschaftler:innen in den Gebieten Informatik und Wirtschaftsinformatik. Einige der Schwerpunkte sind Cloud Computing, Parallelisierung und In-Memory Technologien. In diesem Technischen Bericht werden die Ergebnisse der Forschungsprojekte des Jahres 2018 vorgestellt. Ausgewählte Projekte stellten ihre Ergebnisse am 17. April und 14. November 2018 im Rahmen des Future SOC Lab Tags vor. T3 - Technische Berichte des Hasso-Plattner-Instituts für Digital Engineering an der Universität Potsdam - 151 KW - Future SOC Lab KW - research projects KW - multicore architectures KW - in-memory technology KW - cloud computing KW - machine learning KW - artifical intelligence KW - Future SOC Lab KW - Forschungsprojekte KW - Multicore Architekturen KW - In-Memory Technologie KW - Cloud Computing KW - maschinelles Lernen KW - künstliche Intelligenz Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-563712 SN - 978-3-86956-547-7 SN - 1613-5652 SN - 2191-1665 IS - 151 PB - Universitätsverlag Potsdam CY - Potsdam ER -