{"id":{"repo_id":"drake","oai_identifier":"oai:escholarshare.drake.edu:2092/320"},"canonical_url":"https://search.dev.ndltd.org/etd/drake/oai:escholarshare.drake.edu:2092/320","repository":{"repo_id":"drake","name":"Drake University","base_url":"https://escholarshare.drake.edu/server/oai/request"},"display":{"title":"Retroviral-Medicated Transfer of Suppressor tRNA Genes Into Human Cells","abstract":"Nonsense mutations in protein genes have been associated with a variety of human diseases. A useful therapeutic approach to correct such mutations may be to introduce genes expressing suppressor tRNA that recognize the premature stop codon, insert the appropriate amino acid, and allow translation of a full-length, functional protein. Human opal arginine suppressor tRNA genes that encode the structural portion of the tRNA and about 15 bases from the 3' flanking sequence were synthetically constructed using oligonucleotides. Retroviral vectors were selected for delivery to human target cells in order to stably integrate the suppressor tRNA into the target cells' genome. Recombinant retroviral vectors were used to transfect retrovirus packaging cells and produce recombinant viral supernatants. Recombinant viral supernatants were then used to transduce the target cells. Expression of suppressor tRNA in both the retrovirus packaging cells and in the target cells was confirmed by slot blot analysis. To test the functional activity of the expressed suppressor tRNA, target cells expressing a nonsense mutated GFP reporter gene (Arg73opal mhRGFP) were transduced with the recombinant viral supernatant. Suppression of the opal nonsense mutation in hRGFP was demonstrated by restoration of hRGFP fluorescence. This study suggests that retroviral vectors may be used to deliver suppressor tRNA genes to human target cells.","abstract_html":"Nonsense mutations in protein genes have been associated with a variety of human diseases. A useful therapeutic approach to correct such mutations may be to introduce genes expressing suppressor tRNA that recognize the premature stop codon, insert the appropriate amino acid, and allow translation of a full-length, functional protein. Human opal arginine suppressor tRNA genes that encode the structural portion of the tRNA and about 15 bases from the 3' flanking sequence were synthetically constructed using oligonucleotides. Retroviral vectors were selected for delivery to human target cells in order to stably integrate the suppressor tRNA into the target cells' genome. Recombinant retroviral vectors were used to transfect retrovirus packaging cells and produce recombinant viral supernatants. Recombinant viral supernatants were then used to transduce the target cells. Expression of suppressor tRNA in both the retrovirus packaging cells and in the target cells was confirmed by slot blot analysis. To test the functional activity of the expressed suppressor tRNA, target cells expressing a nonsense mutated GFP reporter gene (Arg73opal mhRGFP) were transduced with the recombinant viral supernatant. Suppression of the opal nonsense mutation in hRGFP was demonstrated by restoration of hRGFP fluorescence. This study suggests that retroviral vectors may be used to deliver suppressor tRNA genes to human target cells.","abstract_has_math":false,"creators":["Breithaupt, Barbara Janine"],"institution":"Drake University","degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2000,"date_issued":"2000-03","date_published":"2000-03","updated_at":"2026-07-27T19:19:04Z","subjects":["Recombinant proteins--Synthesis.","Gene therapy","Retroviruses."],"languages":["en_US"],"rights":[],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["2000 .B748"],"render_values":[{"text":"2000 .B748","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/2092/320","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["Breithaupt, Barbara Janine"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2006-02-28T17:03:44Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2006-02-28T17:03:44Z"]},{"key":"dc:date.issued","label":"Date","values":["2000-03"]},{"key":"dc:publisher","label":"Institution","values":["Drake University"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Recombinant proteins--Synthesis.","Gene therapy","Retroviruses."]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language.iso","label":"Language (ISO)","values":["en_US"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["2000 .B748"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["http://hdl.handle.net/2092/320"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["x, 51 leaves. Advisor: Charisse M. Buising."]},{"key":"dc:description.abstract","label":"Abstract","values":["Nonsense mutations in protein genes have been associated with a variety of human diseases. A useful therapeutic approach to correct such mutations may be to introduce genes expressing suppressor tRNA that recognize the premature stop codon, insert the appropriate amino acid, and allow translation of a full-length, functional protein. Human opal arginine suppressor tRNA genes that encode the structural portion of the tRNA and about 15 bases from the 3' flanking sequence were synthetically constructed using oligonucleotides. Retroviral vectors were selected for delivery to human target cells in order to stably integrate the suppressor tRNA into the target cells' genome. Recombinant retroviral vectors were used to transfect retrovirus packaging cells and produce recombinant viral supernatants. Recombinant viral supernatants were then used to transduce the target cells. Expression of suppressor tRNA in both the retrovirus packaging cells and in the target cells was confirmed by slot blot analysis. To test the functional activity of the expressed suppressor tRNA, target cells expressing a nonsense mutated GFP reporter gene (Arg73opal mhRGFP) were transduced with the recombinant viral supernatant. Suppression of the opal nonsense mutation in hRGFP was demonstrated by restoration of hRGFP fluorescence. This study suggests that retroviral vectors may be used to deliver suppressor tRNA genes to human target cells."]},{"key":"dc:format.mimetype","label":"Dc Format Mimetype","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Retroviral-Medicated Transfer of Suppressor tRNA Genes Into Human Cells"]}]}],"canonical_facts":{"dc:creator":["Breithaupt, Barbara Janine"],"dc:date.accessioned":["2006-02-28T17:03:44Z"],"dc:date.available":["2006-02-28T17:03:44Z"],"dc:date.issued":["2000-03"],"dc:description":["x, 51 leaves. Advisor: Charisse M. Buising."],"dc:description.abstract":["Nonsense mutations in protein genes have been associated with a variety of human diseases. A useful therapeutic approach to correct such mutations may be to introduce genes expressing suppressor tRNA that recognize the premature stop codon, insert the appropriate amino acid, and allow translation of a full-length, functional protein. Human opal arginine suppressor tRNA genes that encode the structural portion of the tRNA and about 15 bases from the 3' flanking sequence were synthetically constructed using oligonucleotides. Retroviral vectors were selected for delivery to human target cells in order to stably integrate the suppressor tRNA into the target cells' genome. Recombinant retroviral vectors were used to transfect retrovirus packaging cells and produce recombinant viral supernatants. Recombinant viral supernatants were then used to transduce the target cells. Expression of suppressor tRNA in both the retrovirus packaging cells and in the target cells was confirmed by slot blot analysis. To test the functional activity of the expressed suppressor tRNA, target cells expressing a nonsense mutated GFP reporter gene (Arg73opal mhRGFP) were transduced with the recombinant viral supernatant. Suppression of the opal nonsense mutation in hRGFP was demonstrated by restoration of hRGFP fluorescence. This study suggests that retroviral vectors may be used to deliver suppressor tRNA genes to human target cells."],"dc:format.mimetype":["application/pdf"],"dc:identifier.other":["2000 .B748"],"dc:identifier.uri":["http://hdl.handle.net/2092/320"],"dc:language.iso":["en_US"],"dc:publisher":["Drake University"],"dc:subject":["Recombinant proteins--Synthesis.","Gene therapy","Retroviruses."],"dc:title":["Retroviral-Medicated Transfer of Suppressor tRNA Genes Into Human Cells"],"dc:type":["Thesis"]},"updated_at":"2026-07-27T19:19:04Z"}