{"id":{"repo_id":"vt","oai_identifier":"oai:vtechworks.lib.vt.edu:10919/44396"},"canonical_url":"https://search.dev.ndltd.org/etd/vt/oai:vtechworks.lib.vt.edu:10919/44396","repository":{"repo_id":"vt","name":"Virginia Tech","base_url":"https://vtechworks.lib.vt.edu/oai/request"},"display":{"title":"Cloning of a region in the Brucella abortus chromosome necessary for o-side chain biosynthesis","abstract":"As a first step in characterizing the genes involved in O-side chain synthesis in Brucella abortus strain 2308, a portion of the genomic DNA was cloned from a rough mutant created by Tn5 (KnR) mutagenesis. This mutant was rough based on the lack of reactivity by either whole cells or extracted LPS to an O-side chain monoclonal antibody (BRU-38). A 30 kb XbaI genomic fragment (including Tn5) from the rough strain was subcloned into a sequencing vector to create pJM6. When B. abortus 2308 was electroporated with pJM6, KnR clones were unable to react with BRU-38; a Southern analysis of these clones revealed Tn5 in the 30 kb XbaI genomic fragment. Various regions of the 30kb fragment were subcloned and tested for their ability to complement specific rfa and rfb mutants of Escherichia coli and Salmonella typhimurium. One particular DNA fragment complemented an rfbD mutation in E. coli as judged by agglutination with E. coli anti-O (0:85) serum. The same DNA fragment failed to cause E. coli rfbD to react with either BRU-38 or B. abortus anti-O polyclonal antisera. The B. abortus 30 kb XbaI fragment contains a gene which has been identified by comple-mentation as containing the equivalent of the rfbD gene encoding dTDP-rhamnose synthetase in E. coli. Since Brucella is not known to have rhamnose in its core this enzyme may have a different function in Brucella LPS synthesis.","abstract_html":"As a first step in characterizing the genes involved in O-side chain synthesis in <i>Brucella abortus</i> strain 2308, a portion of the genomic DNA was cloned from a rough mutant created by Tn5 (KnR) mutagenesis. This mutant was rough based on the lack of reactivity by either whole cells or extracted LPS to an O-side chain monoclonal antibody (BRU-38). A 30 kb <i>Xba</i>I genomic fragment (including Tn5) from the rough strain was subcloned into a sequencing vector to create pJM6. When <i>B. abortus</i> 2308 was electroporated with pJM6, KnR clones were unable to react with BRU-38; a Southern analysis of these clones revealed Tn5 in the 30 kb <i>Xba</i>I genomic fragment. Various regions of the 30kb fragment were subcloned and tested for their ability to complement specific <i>rfa</i> and <i>rfb</i> mutants of <i>Escherichia coli</i> and <i>Salmonella typhimurium</i>. One particular DNA fragment complemented an <i>rfbD</i> mutation in <i>E. coli</i> as judged by agglutination with <i>E. coli</i> anti-O (0:85) serum. The same DNA fragment failed to cause <i>E. coli rfbD</i> to react with either BRU-38 or <i>B. abortus</i> anti-O polyclonal antisera. The <i>B. abortus</i> 30 kb <i>Xba</i>I fragment contains a gene which has been identified by comple-mentation as containing the equivalent of the <i>rfbD</i> gene encoding dTDP-rhamnose synthetase in <i>E. coli</i>. Since <i>Brucella</i> is not known to have rhamnose in its core this enzyme may have a different function in <i>Brucella</i> LPS synthesis.","abstract_has_math":false,"creators":["McQuiston, John R."],"institution":"Virginia Tech","degree_name":"Master of Science","degree_level":"masters","degree_discipline":"Veterinary Medical Sciences","degree_department":"Veterinary Medical Sciences","school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":1992,"date_issued":"1992","date_published":"1992","updated_at":"2026-07-22T22:20:26Z","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-08222009-040429"],"render_values":[{"text":"etd-08222009-040429","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/10919/44396","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.department","label":"Department","values":["Veterinary Medical Sciences"]},{"key":"dc:creator","label":"Author","values":["McQuiston, John R."]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2014-03-14T21:43:29Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2014-03-14T21:43:29Z","2009-08-22"]},{"key":"dc:date.issued","label":"Date","values":["1992"]},{"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":["Veterinary Medical Sciences"]},{"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-08222009-040429"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["http://hdl.handle.net/10919/44396"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["As a first step in characterizing the genes involved in O-side chain synthesis in Brucella abortus strain 2308, a portion of the genomic DNA was cloned from a rough mutant created by Tn5 (KnR) mutagenesis. This mutant was rough based on the lack of reactivity by either whole cells or extracted LPS to an O-side chain monoclonal antibody (BRU-38). A 30 kb XbaI genomic fragment (including Tn5) from the rough strain was subcloned into a sequencing vector to create pJM6. When B. abortus 2308 was electroporated with pJM6, KnR clones were unable to react with BRU-38; a Southern analysis of these clones revealed Tn5 in the 30 kb XbaI genomic fragment. Various regions of the 30kb fragment were subcloned and tested for their ability to complement specific rfa and rfb mutants of Escherichia coli and Salmonella typhimurium. One particular DNA fragment complemented an rfbD mutation in E. coli as judged by agglutination with E. coli anti-O (0:85) serum. The same DNA fragment failed to cause E. coli rfbD to react with either BRU-38 or B. abortus anti-O polyclonal antisera. The B. abortus 30 kb XbaI fragment contains a gene which has been identified by comple-mentation as containing the equivalent of the rfbD gene encoding dTDP-rhamnose synthetase in E. coli. Since Brucella is not known to have rhamnose in its core this enzyme may have a different function in Brucella LPS synthesis."]},{"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":["Cloning of a region in the Brucella abortus chromosome necessary for o-side chain biosynthesis"]}]}],"canonical_facts":{"dc:contributor.department":["Veterinary Medical Sciences"],"dc:creator":["McQuiston, John R."],"dc:date.accessioned":["2014-03-14T21:43:29Z"],"dc:date.available":["2014-03-14T21:43:29Z","2009-08-22"],"dc:date.issued":["1992"],"dc:description.abstract":["As a first step in characterizing the genes involved in O-side chain synthesis in Brucella abortus strain 2308, a portion of the genomic DNA was cloned from a rough mutant created by Tn5 (KnR) mutagenesis. This mutant was rough based on the lack of reactivity by either whole cells or extracted LPS to an O-side chain monoclonal antibody (BRU-38). A 30 kb XbaI genomic fragment (including Tn5) from the rough strain was subcloned into a sequencing vector to create pJM6. When B. abortus 2308 was electroporated with pJM6, KnR clones were unable to react with BRU-38; a Southern analysis of these clones revealed Tn5 in the 30 kb XbaI genomic fragment. Various regions of the 30kb fragment were subcloned and tested for their ability to complement specific rfa and rfb mutants of Escherichia coli and Salmonella typhimurium. One particular DNA fragment complemented an rfbD mutation in E. coli as judged by agglutination with E. coli anti-O (0:85) serum. The same DNA fragment failed to cause E. coli rfbD to react with either BRU-38 or B. abortus anti-O polyclonal antisera. The B. abortus 30 kb XbaI fragment contains a gene which has been identified by comple-mentation as containing the equivalent of the rfbD gene encoding dTDP-rhamnose synthetase in E. coli. Since Brucella is not known to have rhamnose in its core this enzyme may have a different function in Brucella LPS synthesis."],"dc:description.degree":["Master of Science"],"dc:format.medium":["BTD"],"dc:format.mimetype":["application/pdf"],"dc:identifier.other":["etd-08222009-040429"],"dc:identifier.uri":["http://hdl.handle.net/10919/44396"],"dc:language.iso":["en"],"dc:publisher":["Virginia Tech"],"dc:rights":["In Copyright"],"dc:rights.uri":["http://rightsstatements.org/vocab/InC/1.0/"],"dc:title":["Cloning of a region in the Brucella abortus chromosome necessary for o-side chain biosynthesis"],"dc:type":["Thesis"],"dc:type.dcmitype":["Text"],"thesis:degree_discipline":["Veterinary Medical Sciences"],"thesis:degree_level":["masters"],"thesis:degree_name":["Master of Science"],"thesis:institution_name":["Virginia Polytechnic Institute and State University"]},"updated_at":"2026-07-22T22:20:26Z"}