Agrobacterium-mediated transformation of Botrytis cinerea, simple purification of monokaryotic transformants and rapid conidia-based identification of the transfer-DNA host genomic DNA flanking sequences
The Agrobacterium tumefaciens-mediated transfer of foreign DNA to the phytopathogenic fungus Botrytis cinerea was investigated. Fifteen stable transformants per 10(6) conidia were consistently produced. Monokaryons were purified in a single step and their molecular analysis demonstrated the random i...
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| Veröffentlicht in: | Current genetics 2003-11, Vol.44 (3), p.164-171 |
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| creator | Rolland, S Jobic, C Fevre, M Bruel, C |
| description | The Agrobacterium tumefaciens-mediated transfer of foreign DNA to the phytopathogenic fungus Botrytis cinerea was investigated. Fifteen stable transformants per 10(6) conidia were consistently produced. Monokaryons were purified in a single step and their molecular analysis demonstrated the random integration of predominantly single or tandem copies of the foreign DNA into their genome. Thermal asymmetric interlaced PCR performed directly on conidia led to the rapid identification of the genomic DNA sequences that flanked the integration sites of the transfer-DNA. Transcriptional fusions of green fluorescent protein and beta-glucuronidase-encoding genes to the promoter of the secreted proteolytic enzyme ACP1 were realised to validate the system. We provide herein observations of B. cinerea hyphae producing green fluorescent protein or beta-glucuronidase under growth conditions similar to those known to induce transcription of the acp1 gene. |
| doi_str_mv | 10.1007/s00294-003-0438-8 |
| format | Article |
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Fifteen stable transformants per 10(6) conidia were consistently produced. Monokaryons were purified in a single step and their molecular analysis demonstrated the random integration of predominantly single or tandem copies of the foreign DNA into their genome. Thermal asymmetric interlaced PCR performed directly on conidia led to the rapid identification of the genomic DNA sequences that flanked the integration sites of the transfer-DNA. Transcriptional fusions of green fluorescent protein and beta-glucuronidase-encoding genes to the promoter of the secreted proteolytic enzyme ACP1 were realised to validate the system. We provide herein observations of B. cinerea hyphae producing green fluorescent protein or beta-glucuronidase under growth conditions similar to those known to induce transcription of the acp1 gene.</description><identifier>ISSN: 0172-8083</identifier><identifier>EISSN: 1432-0983</identifier><identifier>DOI: 10.1007/s00294-003-0438-8</identifier><identifier>PMID: 12937946</identifier><language>eng</language><publisher>United States: Springer Nature B.V</publisher><subject>acp1 gene ; ACP1 promoter ; ACP1 protein ; Agrobacterium tumefaciens ; Agrobacterium tumefaciens - genetics ; Ascomycota - genetics ; Bacteria ; Blotting, Southern ; Blotting, Western ; Botrytis - genetics ; Botrytis cinerea ; Cells ; conidia ; Deoxyribonucleic acid ; DNA ; DNA Primers ; DNA, Bacterial - genetics ; DNA, Bacterial - metabolism ; gene expression ; gene expression regulation ; gene transfer ; genetic recombination ; genetic transformation ; Glucuronidase ; green fluorescent protein ; Green Fluorescent Proteins ; Growth conditions ; Luminescent Proteins ; Microscopy, Fluorescence ; Plasmids - genetics ; polymerase chain reaction ; Polymerase Chain Reaction - methods ; promoter regions ; reporter genes ; Sclerotinia sclerotiorum ; thermal asymmetric interlaced polymerase chain reaction ; Transformation, Genetic - genetics</subject><ispartof>Current genetics, 2003-11, Vol.44 (3), p.164-171</ispartof><rights>Springer-Verlag 2003</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c445t-f965ffe324d8e3a34a17b9e9a58bc1d97e543d0b2d04ad5b93ea9d0471cfdd8c3</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/12937946$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Rolland, S</creatorcontrib><creatorcontrib>Jobic, C</creatorcontrib><creatorcontrib>Fevre, M</creatorcontrib><creatorcontrib>Bruel, C</creatorcontrib><title>Agrobacterium-mediated transformation of Botrytis cinerea, simple purification of monokaryotic transformants and rapid conidia-based identification of the transfer-DNA host genomic DNA flanking sequences</title><title>Current genetics</title><addtitle>Curr Genet</addtitle><description>The Agrobacterium tumefaciens-mediated transfer of foreign DNA to the phytopathogenic fungus Botrytis cinerea was investigated. Fifteen stable transformants per 10(6) conidia were consistently produced. Monokaryons were purified in a single step and their molecular analysis demonstrated the random integration of predominantly single or tandem copies of the foreign DNA into their genome. Thermal asymmetric interlaced PCR performed directly on conidia led to the rapid identification of the genomic DNA sequences that flanked the integration sites of the transfer-DNA. Transcriptional fusions of green fluorescent protein and beta-glucuronidase-encoding genes to the promoter of the secreted proteolytic enzyme ACP1 were realised to validate the system. We provide herein observations of B. cinerea hyphae producing green fluorescent protein or beta-glucuronidase under growth conditions similar to those known to induce transcription of the acp1 gene.</description><subject>acp1 gene</subject><subject>ACP1 promoter</subject><subject>ACP1 protein</subject><subject>Agrobacterium tumefaciens</subject><subject>Agrobacterium tumefaciens - genetics</subject><subject>Ascomycota - genetics</subject><subject>Bacteria</subject><subject>Blotting, Southern</subject><subject>Blotting, Western</subject><subject>Botrytis - genetics</subject><subject>Botrytis cinerea</subject><subject>Cells</subject><subject>conidia</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>DNA Primers</subject><subject>DNA, Bacterial - genetics</subject><subject>DNA, Bacterial - metabolism</subject><subject>gene expression</subject><subject>gene expression regulation</subject><subject>gene transfer</subject><subject>genetic recombination</subject><subject>genetic transformation</subject><subject>Glucuronidase</subject><subject>green fluorescent protein</subject><subject>Green Fluorescent Proteins</subject><subject>Growth conditions</subject><subject>Luminescent Proteins</subject><subject>Microscopy, Fluorescence</subject><subject>Plasmids - genetics</subject><subject>polymerase chain reaction</subject><subject>Polymerase Chain Reaction - methods</subject><subject>promoter regions</subject><subject>reporter genes</subject><subject>Sclerotinia sclerotiorum</subject><subject>thermal asymmetric interlaced polymerase chain reaction</subject><subject>Transformation, Genetic - genetics</subject><issn>0172-8083</issn><issn>1432-0983</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2003</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>BENPR</sourceid><recordid>eNqFks2OFCEUhStG44yjD-BGiQtXopeC6oJlO_4mE13orAkFlx5muqAHqMU8oy8lne74t3FBIDffPecCp-ueMnjNAMY3BaBXggJwCoJLKu91p0zwnoKS_H53CmzsqQTJT7pHpVwDsF6q8WF3wnrFRyVWp92P9SanydiKOSwzndEFU9GRmk0sPuXZ1JAiSZ68TTXf1VCIDREzmlekhHm3RbJbcvDB_gLnFNONyXepBvuHTqyFmOhINrvgiE0xNCs6mdLcgsNY_xKpV3jsxUzffVmTq1Qq2WBMc1PdF_zWxJsQN6Tg7YLRYnncPfBmW_DJcT_rLj-8_37-iV58_fj5fH1BrRBDpV6tBu-R98JJ5IYLw8ZJoTKDnCxzasRBcAdT70AYN0yKo1HtPDLrnZOWn3UvD7q7nJp1qXoOxeK2DYRpKXpkXK742P8XZKqHlWCsgS_-Aa_TkmO7hJYwrnpgw9AgdoBsTqVk9HqXw9weWjPQ-zzoQx50y4Pe50HL1vPsKLxM7W9_dxwD0IDnB8CbpM0mh6IvvzU_DgzaarP9BB7Cv7Q</recordid><startdate>20031101</startdate><enddate>20031101</enddate><creator>Rolland, S</creator><creator>Jobic, C</creator><creator>Fevre, M</creator><creator>Bruel, C</creator><general>Springer Nature B.V</general><scope>FBQ</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>7QL</scope><scope>7SS</scope><scope>7TK</scope><scope>7TM</scope><scope>7X7</scope><scope>7XB</scope><scope>88A</scope><scope>88E</scope><scope>8AO</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>P64</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>RC3</scope><scope>7X8</scope></search><sort><creationdate>20031101</creationdate><title>Agrobacterium-mediated transformation of Botrytis cinerea, simple purification of monokaryotic transformants and rapid conidia-based identification of the transfer-DNA host genomic DNA flanking sequences</title><author>Rolland, S ; Jobic, C ; Fevre, M ; Bruel, C</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c445t-f965ffe324d8e3a34a17b9e9a58bc1d97e543d0b2d04ad5b93ea9d0471cfdd8c3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2003</creationdate><topic>acp1 gene</topic><topic>ACP1 promoter</topic><topic>ACP1 protein</topic><topic>Agrobacterium tumefaciens</topic><topic>Agrobacterium tumefaciens - genetics</topic><topic>Ascomycota - genetics</topic><topic>Bacteria</topic><topic>Blotting, Southern</topic><topic>Blotting, Western</topic><topic>Botrytis - genetics</topic><topic>Botrytis cinerea</topic><topic>Cells</topic><topic>conidia</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>DNA Primers</topic><topic>DNA, Bacterial - genetics</topic><topic>DNA, Bacterial - metabolism</topic><topic>gene expression</topic><topic>gene expression regulation</topic><topic>gene transfer</topic><topic>genetic recombination</topic><topic>genetic transformation</topic><topic>Glucuronidase</topic><topic>green fluorescent protein</topic><topic>Green Fluorescent Proteins</topic><topic>Growth conditions</topic><topic>Luminescent Proteins</topic><topic>Microscopy, Fluorescence</topic><topic>Plasmids - genetics</topic><topic>polymerase chain reaction</topic><topic>Polymerase Chain Reaction - methods</topic><topic>promoter regions</topic><topic>reporter genes</topic><topic>Sclerotinia sclerotiorum</topic><topic>thermal asymmetric interlaced polymerase chain reaction</topic><topic>Transformation, Genetic - genetics</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Rolland, S</creatorcontrib><creatorcontrib>Jobic, C</creatorcontrib><creatorcontrib>Fevre, M</creatorcontrib><creatorcontrib>Bruel, C</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Neurosciences Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Biology Database (Alumni Edition)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>Technology Research Database</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest One Sustainability</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>Engineering Research Database</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>ProQuest Biological Science Collection</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biological Science Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Current genetics</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Rolland, S</au><au>Jobic, C</au><au>Fevre, M</au><au>Bruel, C</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Agrobacterium-mediated transformation of Botrytis cinerea, simple purification of monokaryotic transformants and rapid conidia-based identification of the transfer-DNA host genomic DNA flanking sequences</atitle><jtitle>Current genetics</jtitle><addtitle>Curr Genet</addtitle><date>2003-11-01</date><risdate>2003</risdate><volume>44</volume><issue>3</issue><spage>164</spage><epage>171</epage><pages>164-171</pages><issn>0172-8083</issn><eissn>1432-0983</eissn><abstract>The Agrobacterium tumefaciens-mediated transfer of foreign DNA to the phytopathogenic fungus Botrytis cinerea was investigated. Fifteen stable transformants per 10(6) conidia were consistently produced. Monokaryons were purified in a single step and their molecular analysis demonstrated the random integration of predominantly single or tandem copies of the foreign DNA into their genome. Thermal asymmetric interlaced PCR performed directly on conidia led to the rapid identification of the genomic DNA sequences that flanked the integration sites of the transfer-DNA. Transcriptional fusions of green fluorescent protein and beta-glucuronidase-encoding genes to the promoter of the secreted proteolytic enzyme ACP1 were realised to validate the system. We provide herein observations of B. cinerea hyphae producing green fluorescent protein or beta-glucuronidase under growth conditions similar to those known to induce transcription of the acp1 gene.</abstract><cop>United States</cop><pub>Springer Nature B.V</pub><pmid>12937946</pmid><doi>10.1007/s00294-003-0438-8</doi><tpages>8</tpages></addata></record> |
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| subjects | acp1 gene ACP1 promoter ACP1 protein Agrobacterium tumefaciens Agrobacterium tumefaciens - genetics Ascomycota - genetics Bacteria Blotting, Southern Blotting, Western Botrytis - genetics Botrytis cinerea Cells conidia Deoxyribonucleic acid DNA DNA Primers DNA, Bacterial - genetics DNA, Bacterial - metabolism gene expression gene expression regulation gene transfer genetic recombination genetic transformation Glucuronidase green fluorescent protein Green Fluorescent Proteins Growth conditions Luminescent Proteins Microscopy, Fluorescence Plasmids - genetics polymerase chain reaction Polymerase Chain Reaction - methods promoter regions reporter genes Sclerotinia sclerotiorum thermal asymmetric interlaced polymerase chain reaction Transformation, Genetic - genetics |
| title | Agrobacterium-mediated transformation of Botrytis cinerea, simple purification of monokaryotic transformants and rapid conidia-based identification of the transfer-DNA host genomic DNA flanking sequences |
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