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Development of a bovine adenovirus type 2-based gene delivery vector /

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dc.contributor.author Ojkic, Davor. en_US
dc.date.accessioned 2009-05-21T13:25:00Z
dc.date.available 2009-05-21T13:25:00Z
dc.date.issued 1997-05-21T13:25:00Z
dc.identifier.uri http://hdl.handle.net/10464/1245
dc.description.abstract ABSTRACT Recombinant adenoviruses are currently under intense investigation as potential gene delivery and gene expression vectors with applications in human and veterinary medicine. As part of our efforts to develop a bovine adenovirus type 2 (BAV2) based vector system, the nucleotide sequence of BAV2 was determined. Sixty-six open reading frames (ORFs) were found with the potential to encode polypeptides that were at least 50 amino acid (aa) residue long. Thirty-one of the BAV2 polypeptide sequences were found to share homology to already identified adenovirus proteins. The arrangement of the genes revealed that the BAV2 genomic organization closely resembles that of well-characterized human adenoviruses. In the course of this study, continuous propagation of BAV2 over many generations in cell culture resulted in the isolation of a BAV2 spontaneous mutant in which the E3 region was deleted. Restriction enzyme, sequencing and PCR analyses produced concordant results that precisely located the deletion and revealed that its size was exactly 1299 bp. The E3-deleted virus was plaque-purified and further propagated in cell culture. It appeared that the replication of such a virus lacking a portion of the E3 region was not affected, at least in cell culture. Attempts to rescue a recombinant BAV2 virus with the bacterial kanamycin resistance gene in the E3 region yielded a candidate as verified with extensive Southern blotting and PCR analyses. Attempts to purify the recombinant virus were not successful, suggesting that such recombinant BAV2 was helper-dependent. Ten clones containing full-length BAV2 genomes in a pWE15 cosmid vector were constructed. The infectivity of these constructs was tested by using different transfection methods. The BAV2 genomic clones did appear to be infectious only after extended incubation period. This may be due to limitations of various transfection methods tested, or biological differences between virus- and E. co//-derived BAV2 DNA. en_US
dc.language.iso eng en_US
dc.publisher Brock University en_US
dc.subject Adenoviruses. en_US
dc.subject DNA viruses. en_US
dc.subject Genetic vectors. en_US
dc.title Development of a bovine adenovirus type 2-based gene delivery vector / en_US
dc.type Electronic Thesis or Dissertation en_US
dc.degree.name M.Sc. Biological Sciences en_US
dc.degree.level Masters en_US
dc.contributor.department Department of Biological Sciences en_US
dc.degree.discipline Faculty of Mathematics and Science en_US


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