Characterization of Herpes Simplex Virus Type 1 Portal Mutants
dc.contributor.author | Allen, Jacob | |
dc.date.accessioned | 2024-07-30T20:04:27Z | |
dc.date.available | 2024-07-30T20:04:27Z | |
dc.identifier.uri | http://hdl.handle.net/10898/13915 | |
dc.description | 2024 | |
dc.description.abstract | The Herpesviridae family is comprised of nine ubiquitous viruses capable of causing primary and latent infection in humans. Herpes Simplex Virus Type 1 (HSV-1) is an alpha-herpesvirus that can manifest as herpes labialis, ocular infection, and encephalitis. The HSV-1 capsid contains a dodecameric assembly of pUL6 monomers at one vertex forming a portal for DNA to be translocated into the capsid. The portal contains several potential functional domains (stem, clip, b-hairpin, wing, crown, and wall) that coordinate the DNA encapsidation process during viral replication. In this study, we aimed to examine the contribution(s) of these domains to DNA packaging. A library of mutant UL6 genes containing in-frame insertions of 15 base-pairs throughout the gene as well as specific mutations to the region encoding the pUL6 -hairpin was generated. In addition, specific mutations to amino acid 541- 558 B-Hairpin domain included a full knockout, a scramble of residues and a substitution of charged residues (KRNQ) to alanine. Thirty-seven random insertions throughout UL6 gene were identified and the location of each insertion confirmed by DNA sequencing. Each mutant gene was cloned into the HSV-1 Strain-17 bacterial artificial chromosome (BAC) using a heat-shock controlled, homologous recombination event in SW102 E. coli. Mutant BACs were transfected into African green monkey kidney (Vero) and 31 (HSV-1 UL6 complementing) cell lines to generate mutant virus stocks. Western blot analysis of virus infected cell lysates was performed to confirm expression of mutant pUL6 proteins. In addition, viral replication kinetics were assessed in a replication assay. | |
dc.publisher | Mercer University | |
dc.subject | Molecular biology | |
dc.subject | Virology | |
dc.subject | Herpes, Portal Protein | |
dc.title | Characterization of Herpes Simplex Virus Type 1 Portal Mutants | |
dc.type | dissertation | en_US |
dc.date.updated | 2024-07-23T22:03:01Z | |
dc.language.rfc3066 | en | |
refterms.dateFOA | 2024-07-30T20:04:28Z | |
dc.contributor.department | School of Medicine | |
dc.description.advisor | Visalli, Robert | |
dc.description.committee | Bentz, Gretchen | |
dc.description.committee | Drummond, James | |
dc.description.degree | M.S. |
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