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Application of RNA Interference for the Study of Lethal Genes and Dynamic Processes

dc.contributor.advisorBucher, Gregor Prof. Dr.
dc.contributor.authorUlrich, Julia
dc.titleApplication of RNA Interference for the Study of Lethal Genes and Dynamic Processesde
dc.contributor.refereeBucher, Gregor Prof. Dr.
dc.description.abstractengRNA interference (RNAi) is a highly conserved cellular mechanism and in the last years it has become to a major tool for functional analyses of genes in different species. Moreover, RNAi has recently been suggested as a novel and promising approach for pest control. Transgenic plants can be engineered to express double stranded RNAs (dsRNAs) targeting essential genes of a pest species. Upon feeding, the dsRNAs induce gene silencing in the pest, resulting in its death. However, the main challenge of RNAi-mediated plant protection is the identification of efficient RNAi target genes. In most pest species, the screening for RNAi target genes by a whole-animal-high-throughput-approach is not feasible due to missing genomic tools and limited breeding capacity. Therefore, the first aim of this thesis was to use Tribolium castaneum (Tc) as a screening platform in order to identify the most efficient RNAi target genes. By employing the data from the iBeetle RNAi screen, some novel and highly efficient RNAi target genes were identified that induced organism death most rapidly after knockdown. The orthologs of these genes are hence excellent candidates for RNAi based pest control methods in other pest insects. Based on this set of RNAi targets, Gene Ontology term (GO term) combinations were identified that are predictive for efficient RNAi target genes and which detect proteasomal genes as prime targets. Further, I could show that the efficiency of RNAi mediated pest control cannot be increased by synergistic action in double knockdowns. Finally, an off target analysis revealed that protein sequence conservation does not strongly correlate with the number of potential off target sites, indicating that it will be difficult to design dsRNAs without potential off-target sites in non-target organisms. In the second part of this thesis, I aimed to establish a method to regulate the RNAi response. Ubiquitous gene silencing can sometimes lead to pleiotropic effects, hampering the identification of specific phenotypes. Temporal and/or spatial regulation of RNAi can circumvent such effects. The viral RNAi suppressor protein CrPV1A can be used for this purpose. The expression of CrPV1A effectively blocks the RNAi mechanism and does not impair the development or viability of Tribolium. Probably, it also does not interfere with the microRNA (miRNA) pathway. Furthermore, temporal activation of CrPV1A is able to terminate the RNAi response of a previously silenced gene, resulting in a rescued RNAi phenotype. Temporally controlled RNAi by CrPV1A activation is therefore an effective approach to investigate genetic interactions of further genes.de
dc.contributor.coRefereeWimmer, Ernst A. Prof. Dr.
dc.contributor.thirdRefereeSchuh, Reinhard Prof. Dr.
dc.contributor.thirdRefereeDosch, Roland Dr.
dc.contributor.thirdRefereeVorbrüggen, Gerd Dr.
dc.contributor.thirdRefereeJackson, Daniel John Prof. Dr.
dc.subject.engRNA interferencede
dc.subject.engpest controlde
dc.subject.engTribolium castaneumde
dc.subject.engproteasomal genesde
dc.subject.engRNAi suppressorde
dc.affiliation.instituteBiologische Fakultät für Biologie und Psychologiede
dc.subject.gokfullBiologie (PPN619462639)de



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