{"id":{"repo_id":"vt","oai_identifier":"oai:vtechworks.lib.vt.edu:10919/43112"},"canonical_url":"https://search.dev.ndltd.org/etd/vt/oai:vtechworks.lib.vt.edu:10919/43112","repository":{"repo_id":"vt","name":"Virginia Tech","base_url":"https://vtechworks.lib.vt.edu/oai/request"},"display":{"title":"The use of alkaline gel electrophoresis to analyze hydrogen peroxide-caused DNA damage and repair in Escherichia coli","abstract":"Reactive forms of oxygen such as hydrogen peroxide cause single-strand breaks in DNA. Most of the methods for estimating such breakage and subsequent repair are designed for eucaryotic cells and methods for use with bacteria are needed. Accordingly, a method based on alkaline gel electrophoresis of DNA was developed and tested with isogenic strains of Escherichia coli deficient in one or more DNA repair enzymes, viz., recA (recA), exonuclease III (xthA), DNA polymerase I (po/A) or DNA polymerase I plus exonuclease I] (polA-xthA). For DNA analysis of single-strand breaks, samples from a cell suspension were removed at 2 min intervals following an initial 15 min exposure to 20 mmol l⁻¹ hydrogen peroxide. Catalase was added and the cells were embedded in blocks of low-melting point agarose and lysed to liberate their DNA . After alkaline gel electrophoresis, photographs of the gels were taken and the lengths of the distributions of DNA fragments were measured with a scanner and computer. The wild type and recA strain showed only a moderate increase in the length of the DNA distribution whereas the remaining strains all showed a large increase in the length of the distributions. The lengths of the distributions were correlated with cell survival at the same concentration of H₂O₂ and with the importance of particular DNA repair enzymes. Alkaline gel electrophoresis appears to be a relatively simple method for analyzing the level of H₂O₂-caused DNA damage and repair in E. coli.","abstract_html":"Reactive forms of oxygen such as hydrogen peroxide cause single-strand breaks in DNA. Most of the methods for estimating such breakage and subsequent repair are designed for eucaryotic cells and methods for use with bacteria are needed. Accordingly, a method based on alkaline gel electrophoresis of DNA was developed and tested with isogenic strains of Escherichia coli deficient in one or more DNA repair enzymes, viz., recA (recA), exonuclease III (xthA), DNA polymerase I (po/A) or DNA polymerase I plus exonuclease I] (polA-xthA). For DNA analysis of single-strand breaks, samples from a cell suspension were removed at 2 min intervals following an initial 15 min exposure to 20 mmol l⁻¹ hydrogen peroxide. Catalase was added and the cells were embedded in blocks of low-melting point agarose and lysed to liberate their DNA . After alkaline gel electrophoresis, photographs of the gels were taken and the lengths of the distributions of DNA fragments were measured with a scanner and computer. The wild type and recA strain showed only a moderate increase in the length of the DNA distribution whereas the remaining strains all showed a large increase in the length of the distributions. The lengths of the distributions were correlated with cell survival at the same concentration of H₂O₂ and with the importance of particular DNA repair enzymes. Alkaline gel electrophoresis appears to be a relatively simple method for analyzing the level of H₂O₂-caused DNA damage and repair in E. coli.","abstract_has_math":false,"creators":["Zirkle, Ross Eric"],"institution":"Virginia Tech","degree_name":"Master of Science","degree_level":"masters","degree_discipline":"Biology","degree_department":"Biology","school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":1995,"date_issued":"1995","date_published":"1995","updated_at":"2026-07-22T22:20:38Z","subjects":[],"languages":["en"],"rights":["In Copyright"],"rights_urls":["http://rightsstatements.org/vocab/InC/1.0/"],"identifier_entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-06112009-063020"],"render_values":[{"text":"etd-06112009-063020","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/10919/43112","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.department","label":"Department","values":["Biology"]},{"key":"dc:creator","label":"Author","values":["Zirkle, Ross Eric"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2014-03-14T21:37:49Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2014-03-14T21:37:49Z","2009-06-11"]},{"key":"dc:date.issued","label":"Date","values":["1995"]},{"key":"dc:publisher","label":"Institution","values":["Virginia Tech"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.dcmitype","label":"Dc Type Dcmitype","values":["Text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Biology"]},{"key":"thesis:degree_level","label":"Degree Level","values":["masters"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Master of Science"]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["Virginia Polytechnic Institute and State University"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language.iso","label":"Language (ISO)","values":["en"]},{"key":"dc:rights","label":"Dc Rights","values":["In Copyright"]},{"key":"dc:rights.uri","label":"Rights URI","values":["http://rightsstatements.org/vocab/InC/1.0/"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-06112009-063020"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["http://hdl.handle.net/10919/43112"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Reactive forms of oxygen such as hydrogen peroxide cause single-strand breaks in DNA. Most of the methods for estimating such breakage and subsequent repair are designed for eucaryotic cells and methods for use with bacteria are needed. Accordingly, a method based on alkaline gel electrophoresis of DNA was developed and tested with isogenic strains of Escherichia coli deficient in one or more DNA repair enzymes, viz., recA (recA), exonuclease III (xthA), DNA polymerase I (po/A) or DNA polymerase I plus exonuclease I] (polA-xthA). For DNA analysis of single-strand breaks, samples from a cell suspension were removed at 2 min intervals following an initial 15 min exposure to 20 mmol l⁻¹ hydrogen peroxide. Catalase was added and the cells were embedded in blocks of low-melting point agarose and lysed to liberate their DNA . After alkaline gel electrophoresis, photographs of the gels were taken and the lengths of the distributions of DNA fragments were measured with a scanner and computer. The wild type and recA strain showed only a moderate increase in the length of the DNA distribution whereas the remaining strains all showed a large increase in the length of the distributions. The lengths of the distributions were correlated with cell survival at the same concentration of H₂O₂ and with the importance of particular DNA repair enzymes. Alkaline gel electrophoresis appears to be a relatively simple method for analyzing the level of H₂O₂-caused DNA damage and repair in E. coli."]},{"key":"dc:description.degree","label":"Dc Description Degree","values":["Master of Science"]},{"key":"dc:format.medium","label":"Dc Format Medium","values":["BTD"]},{"key":"dc:format.mimetype","label":"Dc Format Mimetype","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["The use of alkaline gel electrophoresis to analyze hydrogen peroxide-caused DNA damage and repair in Escherichia coli"]}]}],"canonical_facts":{"dc:contributor.department":["Biology"],"dc:creator":["Zirkle, Ross Eric"],"dc:date.accessioned":["2014-03-14T21:37:49Z"],"dc:date.available":["2014-03-14T21:37:49Z","2009-06-11"],"dc:date.issued":["1995"],"dc:description.abstract":["Reactive forms of oxygen such as hydrogen peroxide cause single-strand breaks in DNA. Most of the methods for estimating such breakage and subsequent repair are designed for eucaryotic cells and methods for use with bacteria are needed. Accordingly, a method based on alkaline gel electrophoresis of DNA was developed and tested with isogenic strains of Escherichia coli deficient in one or more DNA repair enzymes, viz., recA (recA), exonuclease III (xthA), DNA polymerase I (po/A) or DNA polymerase I plus exonuclease I] (polA-xthA). For DNA analysis of single-strand breaks, samples from a cell suspension were removed at 2 min intervals following an initial 15 min exposure to 20 mmol l⁻¹ hydrogen peroxide. Catalase was added and the cells were embedded in blocks of low-melting point agarose and lysed to liberate their DNA . After alkaline gel electrophoresis, photographs of the gels were taken and the lengths of the distributions of DNA fragments were measured with a scanner and computer. The wild type and recA strain showed only a moderate increase in the length of the DNA distribution whereas the remaining strains all showed a large increase in the length of the distributions. The lengths of the distributions were correlated with cell survival at the same concentration of H₂O₂ and with the importance of particular DNA repair enzymes. Alkaline gel electrophoresis appears to be a relatively simple method for analyzing the level of H₂O₂-caused DNA damage and repair in E. coli."],"dc:description.degree":["Master of Science"],"dc:format.medium":["BTD"],"dc:format.mimetype":["application/pdf"],"dc:identifier.other":["etd-06112009-063020"],"dc:identifier.uri":["http://hdl.handle.net/10919/43112"],"dc:language.iso":["en"],"dc:publisher":["Virginia Tech"],"dc:rights":["In Copyright"],"dc:rights.uri":["http://rightsstatements.org/vocab/InC/1.0/"],"dc:title":["The use of alkaline gel electrophoresis to analyze hydrogen peroxide-caused DNA damage and repair in Escherichia coli"],"dc:type":["Thesis"],"dc:type.dcmitype":["Text"],"thesis:degree_discipline":["Biology"],"thesis:degree_level":["masters"],"thesis:degree_name":["Master of Science"],"thesis:institution_name":["Virginia Polytechnic Institute and State University"]},"updated_at":"2026-07-22T22:20:38Z"}