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1. Exploring DNA quality of single cells for genome analysis with simultaneous whole-genome amplification

Author

Christiane Bäumer, Evelyn Fisch, Holger Wedler, Frank Reinecke, and Christian Korfhage

Subjects
Medicine, Science
Abstract

Abstract Single cell genome analysis methods are powerful tools to define features of single cells and to identify differences between them. Since the DNA amount of a single cell is very limited, cellular DNA usually needs to be amplified by whole-genome amplification before being subjected to further analysis. A single nucleus only contains two haploid genomes. Thus, any DNA damage that prevents amplification results in loss of damaged DNA sites and induces an amplification bias. Therefore, the assessment of single cell DNA quality is urgently required. As of today, there is no simple method to determine the quality of a single cell DNA in a manner that will still retain the entire cellular DNA for amplification and downstream analysis. Here, we describe a method for whole-genome amplification with simultaneous quality control of single cell DNA by using a competitive spike-in DNA template.

Published
2018
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2. Genome sequencing and analysis of the versatile cell factory Aspergillus niger CBS 513.88

Author

Marc J. E. C. van der Maarel, Marco A. van den Berg, Steven Geysens, Etienne Danchin, Rachel M. van der Kaaij, Hein Stam, Jibin Sun, Kaj Albermann, Maurien M.A. Olsthoorn, Arnold J. M. Driessen, Frans M. Klis, Monika Schmoll, Gerald Hofmann, K. Pal, Thomas Guillemette, Bernard Henrissat, Johannes H. de Winde, David B. Archer, Coenie Goosen, Pedro M. Coutinho, Richard Albang, Michael Cornell, Jens Nielsen, Johannes Andries Roubos, Harrie J. Kools, Peter J.J. Van De Vondervoort, Cornelis Maria Jacobus Sagt, Noël Nicolaas Maria Elisabeth Van Peij, Jannick Dyrløv Bendtsen, Alard Van Dijk, Marga Herweijer, Martin Mortimer, Ursula Rinas, An-Ping Zeng, Roland Contreras, Holger Wedler, Stephen G. Oliver, Cees A. M. J. J. van den Hondel, David W. Ussery, Gert S.P. Groot, Jaap Visser, Mikael Rørdam Andersen, Arthur F. J. Ram, Hildegard Henna Menke, René T. J. M. van der Heijden, Paul S. Dyer, Piet W. J. de Groot, Han A. B. Wösten, Alfons J. M. Debets, Jacques A.E. Benen, János Varga, Lubbert Dijkhuizen, Christian P. Kubicek, Peter J. T. Dekker, Albert J. J. van Ooyen, Johannes Petrus Theodorus Wilhelmus Van Den Hombergh, Ronald P. de Vries, Rogier Meulenberg, Jürgen Lauber, Mark X. Caddick, Geoffrey Turner, Xin Lu, Patricia Ann van Kuyk, Wouter Vervecken, Peter J. Schaap, Piet W.M. van Dijck, Stefaan Breestraat, Herman Jan Pel, Christophe d'Enfert, DSM, Delft University of Technology (TU Delft), School of Biology, University of Nottingham, UK (UON), Technical University of Denmark [Lyngby] (DTU), Wageningen University and Research Centre (WUR), Department of Molecular Biology and Biotechnology, University of Sheffield [Sheffield], Utrecht University [Utrecht], Biomax Informatics AG, CLC bio, Partenaires INRAE, School of Biological Sciences, University of Liverpool, Universiteit Gent = Ghent University [Belgium] (UGENT), Flanders Institute for Biotechnology, University of Manchester [Manchester], Architecture et fonction des Macromolécules Biologiques - UMR 6098 (AFMB), Université de Provence - Aix-Marseille 1-Centre National de la Recherche Scientifique (CNRS), Laboratory of Genetics, Wageningen University and Research [Wageningen] (WUR), University of Groningen, Biologie et Pathogénicité fongiques, Institut Pasteur [Paris]-Institut National de la Recherche Agronomique (INRA), Institut Pasteur [Paris], Centre for Carbohydrate Bioprocessing, VU University Amsterdam, Unité de recherche Pathologie végétale et phytobactériologie, Institut National de la Recherche Agronomique (INRA), Leiden University, Radboud University Medical Center [Nijmegen], Vienna University of Technology, Qiagen GmbH, Center for Microbial Biotechnology (CMB), University of Szeged, Helmholtz Centre for Infection Research (HZI), Center for Biological Sequence Analysis, Fungal Biodiversity Centre, Fungal Genetics and Technology Consultancy, Ministère de la Recherche : programme ACI-BCMS, Enzywall, Sonderforschungsbereich 578 (SFB578) of the Deutsche Forschungsgemeinschaft, Germany, Groningen Biomolecular Sciences and Biotechnology, Host-Microbe Interactions, Molecular Microbiology, Danmarks Tekniske Universitet = Technical University of Denmark (DTU), Universiteit Gent = Ghent University (UGENT), Biologie et Pathogénicité fongiques (BPF), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris] (IP), Institut Pasteur [Paris] (IP), Vrije Universiteit Amsterdam [Amsterdam] (VU), Universiteit Leiden, University of Szeged [Szeged], Vrije universiteit = Free university of Amsterdam [Amsterdam] (VU), and Sub Molecular Microbiology

Subjects
[SDV.BIO]Life Sciences [q-bio]/Biotechnology, INDUSTRIAL APPLICATION, autolysis, dna, Applied Microbiology and Biotechnology, Genome, Microbiologie, Putative gene, ENZYME TECHNOLOGY, TRANSCRIPTION FACTOR, Plant Proteins, degradation, 2. Zero hunger, chemistry.chemical_classification, 0303 health sciences, biology, filamentous fungi, nidulans, Chromosome Mapping, unfolded protein response, PE&RC, SEQUENCE ANALYSIS, GENE FUNCTION, Biochemistry, Molecular Medicine, SECRETION, saccharomyces-cerevisiae, Laboratory of Genetics, Chromosomes, Fungal, Genome, Fungal, Biotechnology, reconstruction, Bioinformatics, Molecular Sequence Data, Biomedical Engineering, Bioengineering, Stress, Laboratorium voor Erfelijkheidsleer, Microbiology, DNA sequencing, 03 medical and health sciences, 030304 developmental biology, Synteny, VLAG, Base Sequence, ASPERGILLUS NIGER, 030306 microbiology, GENOMIC, Protein, Aspergillus niger, Sequence Analysis, DNA, fumigatus, biology.organism_classification, Major facilitator superfamily, Open reading frame, Enzyme, chemistry, Cluster, Cellular energy metabolism [UMCN 5.3], metabolism
Abstract

Contains fulltext : 51722.pdf (Publisher’s version ) (Closed access) The filamentous fungus Aspergillus niger is widely exploited by the fermentation industry for the production of enzymes and organic acids, particularly citric acid. We sequenced the 33.9-megabase genome of A. niger CBS 513.88, the ancestor of currently used enzyme production strains. A high level of synteny was observed with other aspergilli sequenced. Strong function predictions were made for 6,506 of the 14,165 open reading frames identified. A detailed description of the components of the protein secretion pathway was made and striking differences in the hydrolytic enzyme spectra of aspergilli were observed. A reconstructed metabolic network comprising 1,069 unique reactions illustrates the versatile metabolism of A. niger. Noteworthy is the large number of major facilitator superfamily transporters and fungal zinc binuclear cluster transcription factors, and the presence of putative gene clusters for fumonisin and ochratoxin A synthesis.

Published
2007
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3. The full-ORF clone resource of the German cDNA Consortium

Author

Ingo Schupp, Helmut Blöcker, Dagmar Heubner, Stefan Wiemann, Annemarie Poustka, Karl Köhrer, Birgit Ottenwälder, Heiko Rosenfelder, Holger Wedler, Ruth Wellenreuther, Anny Duda, Alexander Mehrle, Christian Schmidt, Claudia Schuster, Andreas Hoerlein, Andre Bahr, Stephanie Bechtel, Ute Ernst, and Guenter Michel

Subjects
Quality Control, DNA, Complementary, lcsh:QH426-470, lcsh:Biotechnology, DNA-Directed DNA Polymerase, Biology, Genome, Models, Biological, Polymerase Chain Reaction, Open Reading Frames, User-Computer Interface, lcsh:TP248.13-248.65, Complementary DNA, Germany, Databases, Genetic, Genetics, Humans, Computer Simulation, ORFS, Cloning, Molecular, Cooperative Behavior, ORFeome, DNA Primers, Cloning, Recombination, Genetic, Genome, Human, Methodology Article, Sequence Analysis, DNA, lcsh:Genetics, Open reading frame, Codon, Terminator, Human genome, DNA microarray, Biotechnology
Abstract

Background With the completion of the human genome sequence the functional analysis and characterization of the encoded proteins has become the next urging challenge in the post-genome era. The lack of comprehensive ORFeome resources has thus far hampered systematic applications by protein gain-of-function analysis. Gene and ORF coverage with full-length ORF clones thus needs to be extended. In combination with a unique and versatile cloning system, these will provide the tools for genome-wide systematic functional analyses, to achieve a deeper insight into complex biological processes. Results Here we describe the generation of a full-ORF clone resource of human genes applying the Gateway cloning technology (Invitrogen). A pipeline for efficient cloning and sequencing was developed and a sample tracking database was implemented to streamline the clone production process targeting more than 2,200 different ORFs. In addition, a robust cloning strategy was established, permitting the simultaneous generation of two clone variants that contain a particular ORF with as well as without a stop codon by the implementation of only one additional working step into the cloning procedure. Up to 92 % of the targeted ORFs were successfully amplified by PCR and more than 93 % of the amplicons successfully cloned. Conclusion The German cDNA Consortium ORFeome resource currently consists of more than 3,800 sequence-verified entry clones representing ORFs, cloned with and without stop codon, for about 1,700 different gene loci. 177 splice variants were cloned representing 121 of these genes. The entry clones have been used to generate over 5,000 different expression constructs, providing the basis for functional profiling applications. As a member of the recently formed international ORFeome collaboration we substantially contribute to generating and providing a whole genome human ORFeome collection in a unique cloning system that is made freely available in the community.

Published
2007

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