Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it>
<p>Abstract</p> <p>Background</p> <p><it>Burkholderia gladioli </it>pathovar <it>cocovenenans </it>(BGC) is responsible for sporadic food-poisoning outbreaks with high morbidity and mortality in Asian countries. Little is known about the regulati...
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| Format: | Article |
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BMC
2010-11-01
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| Series: | BMC Research Notes |
| Online Access: | http://www.biomedcentral.com/1756-0500/3/308 |
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| author | Karkhoff-Schweizer RoxAnn R McMillan Ian Somprasong Nawarat Mongkolsuk Skorn Schweizer Herbert P |
| author_facet | Karkhoff-Schweizer RoxAnn R McMillan Ian Somprasong Nawarat Mongkolsuk Skorn Schweizer Herbert P |
| author_sort | Karkhoff-Schweizer RoxAnn R |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p><it>Burkholderia gladioli </it>pathovar <it>cocovenenans </it>(BGC) is responsible for sporadic food-poisoning outbreaks with high morbidity and mortality in Asian countries. Little is known about the regulation of virulence factor and toxin production in BGC, and studies in this bacterium have been hampered by lack of genetic tools.</p> <p>Findings</p> <p>Establishment of a comprehensive antibiotic susceptibility profile showed that BGC strain ATCC33664 is susceptible to a number of antibiotics including aminoglycosides, carbapenems, fluoroquinolones, tetracyclines and trimethoprim. In this study, we established that gentamicin, kanamycin and trimethoprim are good selection markers for use in BGC. Using a 10 min method for preparation of electrocompetent cells, the bacterium could be transformed by electroporation at high frequencies with replicative plasmids containing the pRO1600-derived origin of replication. These plasmids exhibited a copy number of > 100 in BGC. When co-conjugated with a transposase expressing helper plasmid, mini-Tn<it>7 </it>vectors inserted site- and orientation-specifically at a single <it>glmS</it>-associated insertion site in the BGC genome. Lastly, a <it>Himar1 </it>transposon was used for random transposon mutagenesis of BGC.</p> <p>Conclusions</p> <p>A series of genetic tools previously developed for other Gram-negative bacteria was adapted for use in BGC. These tools now facilitate genetic studies of this pathogen and allow establishment of toxin biosynthetic pathways and their genetic regulation.</p> |
| first_indexed | 2024-12-18T19:22:45Z |
| format | Article |
| id | doaj.art-f3591db125bf4530bacc99f8b4b6b48c |
| institution | Directory Open Access Journal |
| issn | 1756-0500 |
| language | English |
| last_indexed | 2024-12-18T19:22:45Z |
| publishDate | 2010-11-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Research Notes |
| spelling | doaj.art-f3591db125bf4530bacc99f8b4b6b48c2022-12-21T20:55:56ZengBMCBMC Research Notes1756-05002010-11-013130810.1186/1756-0500-3-308Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it>Karkhoff-Schweizer RoxAnn RMcMillan IanSomprasong NawaratMongkolsuk SkornSchweizer Herbert P<p>Abstract</p> <p>Background</p> <p><it>Burkholderia gladioli </it>pathovar <it>cocovenenans </it>(BGC) is responsible for sporadic food-poisoning outbreaks with high morbidity and mortality in Asian countries. Little is known about the regulation of virulence factor and toxin production in BGC, and studies in this bacterium have been hampered by lack of genetic tools.</p> <p>Findings</p> <p>Establishment of a comprehensive antibiotic susceptibility profile showed that BGC strain ATCC33664 is susceptible to a number of antibiotics including aminoglycosides, carbapenems, fluoroquinolones, tetracyclines and trimethoprim. In this study, we established that gentamicin, kanamycin and trimethoprim are good selection markers for use in BGC. Using a 10 min method for preparation of electrocompetent cells, the bacterium could be transformed by electroporation at high frequencies with replicative plasmids containing the pRO1600-derived origin of replication. These plasmids exhibited a copy number of > 100 in BGC. When co-conjugated with a transposase expressing helper plasmid, mini-Tn<it>7 </it>vectors inserted site- and orientation-specifically at a single <it>glmS</it>-associated insertion site in the BGC genome. Lastly, a <it>Himar1 </it>transposon was used for random transposon mutagenesis of BGC.</p> <p>Conclusions</p> <p>A series of genetic tools previously developed for other Gram-negative bacteria was adapted for use in BGC. These tools now facilitate genetic studies of this pathogen and allow establishment of toxin biosynthetic pathways and their genetic regulation.</p>http://www.biomedcentral.com/1756-0500/3/308 |
| spellingShingle | Karkhoff-Schweizer RoxAnn R McMillan Ian Somprasong Nawarat Mongkolsuk Skorn Schweizer Herbert P Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it> BMC Research Notes |
| title | Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it> |
| title_full | Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it> |
| title_fullStr | Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it> |
| title_full_unstemmed | Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it> |
| title_short | Methods for genetic manipulation of <it>Burkholderia gladioli </it>pathovar <it>cocovenenans</it> |
| title_sort | methods for genetic manipulation of it burkholderia gladioli it pathovar it cocovenenans it |
| url | http://www.biomedcentral.com/1756-0500/3/308 |
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