Genome scale ordered RNA structure (GORS) is found throughout the genome of many single stranded positive sense RNA viruses, including plant viruses. It was hypothesised that GORS may function to help evade RNAi either by preventing the generation of siRNAs or by stopping RNAi-mediated cleavage of the target. This project used Potato Leafroll Virus (PLRV) to investigate the function of GORS in plant viruses. The RNA structure of a 750nt region of the genome was modified to have less, more or the same energy as the WT sequence. The physical structure of these sequences was shown to be different using two distinct methods. Viral infectivity was tested and although all four viruses were able to replicate and spread to distal leaves, the WT virus was always able to outcompete the variant viruses in competition assays. This suggests GORS provides a distinct selective advantage to the WT virus. The effect on the siRNA response was tested using a dedicated siRNA assay. In plants, this showed that the WT sequence was more resistant to degradation by siRNAs than the variant sequences in the presence of their specific inducers. The WT inducer was also not able to cause suppression of the other targets indicating that this inducer failed to produce siRNAs or that they were not effective. The siRNA populations generated during infections were sequenced and the profiles compared. This showed that all four viruses stimulated the production of siRNAs but the location of siRNA hotspots differed. It is therefore hypothesised that GORS may function to evade the RNAi response by directing the generation of less effective siRNAs. The data presented in this thesis not only informs current work on GORS and RNA structure in viral genomes but also has wider implications for research on siRNAs and food biosecurity.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:582274 |
| Date | January 2012 |
| Creators | Hornsey, Crystal A. |
| Publisher | University of Warwick |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://wrap.warwick.ac.uk/56672/ |
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