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AuthorKarl, Michael Maximiliandc.contributor.author
Date of accession2019-04-04T14:24:49Zdc.date.accessioned
Available in OPARU since2019-04-04T14:24:49Zdc.date.available
Year of creation2018dc.date.created
Date of first publication2019-04-04dc.date.issued
AbstractEven though Clostridium aceticum is the first described acetogen, no reproducible protocol for genetic modification has been published in a peer-reviewed scientific article. Furthermore, C. aceticum possesses cytochromes with still unknown function. In general, cytochromes are involved in energy metabolism and are therefore promising targets for metabolic engineering approaches in order to overcome energetic constrains ever present in acetogens. Consequently, a transformation protocol for C. aceticum cells was developed during this study. Restriction analysis identified the REase Cac1496I as the main obstacle for electrotransformation and probably conjugation. Consequently, the pMK80 plasmids without respective recognition sites were constructed showing resistance to Cac1496I and MvaI digestion. Electrotransformation applying the Molitor protocol in combination with the pMK83 plasmid resulted in recombinant C. aceticum strains. This protocol is also suitable to electrotransform the phylogenetically related acetogen Clostridium formicaceticum. Oxford Nanopore Technology (ONT) sequencing generates long DNA reads, which can be used in combination with accurate Illumina reads to produce high quality and closed genome assemblies. Since C. formicaceticum was genetically modified for the first time during this study and Clostridium coskatii is an industrial relevant acetogen, both organisms were resequenced using the MinION device. The complete genome sequence of C. formicaceticum and C. coskatii were resolved using ONT R9.4 and Illumina MiSeq reads. Therefore, different assembling tools were employed and evaluated giving detailed assembly statistics. Categorisation of annotated CDS into COGs was conducted. Assembly of the C. coskatii genome revealed a second closed contig. Coverage, GC content, and BLAST comparison clearly indicated the presence of a native plasmid, which was ultimately confirmed by isolation and analytic digestion.dc.description.abstract
Languageendc.language.iso
PublisherUniversität Ulmdc.publisher
LicenseStandarddc.rights
Link to license texthttps://oparu.uni-ulm.de/xmlui/license_v3dc.rights.uri
KeywordSequenzanalysedc.subject
KeywordDNAdc.subject
KeywordClostridium coskatiidc.subject
KeywordClostridium formicaceticumdc.subject
KeywordDesoxyribonucleic aciddc.subject
Dewey Decimal GroupDDC 540 / Chemistry & allied sciencesdc.subject.ddc
Dewey Decimal GroupDDC 570 / Life sciencesdc.subject.ddc
LCSHClostridium; Biotechnologydc.subject.lcsh
LCSHNucleotide sequencedc.subject.lcsh
LCSHNucleotides; Analysisdc.subject.lcsh
LCSHGenetic transformationdc.subject.lcsh
LCSHElectroporationdc.subject.lcsh
LCSHPlasmidsdc.subject.lcsh
TitleInsights into old and new acetogens: transformation barriers and genomicsdc.title
Resource typeDissertationdc.type
Date of acceptance2018-12-13dcterms.dateAccepted
RefereeDürre, Peterdc.contributor.referee
RefereeEikmanns, Bernharddc.contributor.referee
DOIhttp://dx.doi.org/10.18725/OPARU-13161dc.identifier.doi
PPN1663518874dc.identifier.ppn
URNhttp://nbn-resolving.de/urn:nbn:de:bsz:289-oparu-13218-3dc.identifier.urn
GNDClostridiumdc.subject.gnd
GNDClostridium aceticumdc.subject.gnd
GNDSequenzanalyse <Chemie>dc.subject.gnd
GNDDNS-Sequenzdc.subject.gnd
GNDGentransferdc.subject.gnd
GNDElektroporationdc.subject.gnd
GNDPlasmiddc.subject.gnd
GNDDNSdc.subject.gnd
FacultyFakultät für Naturwissenschaftenuulm.affiliationGeneral
InstitutionInstitut für Mikrobiologie und Biotechnologieuulm.affiliationSpecific
Grantor of degreeFakultät für Naturwissenschaftenuulm.thesisGrantor
DCMI TypeTextuulm.typeDCMI
CategoryPublikationenuulm.category
Bibliographyuulmuulm.bibliographie


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