Transgene integration complexity and expression stability following biolistic or Agrobacterium-mediated transformation of sugarcane
Sugarcane (Saccharum spp. hybrids) accounts for 80% of the table sugar produced worldwide and is also a prime feedstock for biofuel production. However, very few studies are available for directly comparing Agrobacterium tumefaciens-mediated transfer of T-DNA (AMT) and biolistic transfer of minimal...
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| Veröffentlicht in: | In vitro cellular & developmental biology. Plant 2015-12, Vol.51 (6), p.603-611 |
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| creator | Wu, Hao Awan, Faisal Saeed Vilarinho, Aloisio Zeng, Qianchun Kannan, Baskaran Phipps, Tenisha McCuiston, Jamie Wang, Wenling Caffall, Kerry Altpeter, Fredy |
| description | Sugarcane (Saccharum spp. hybrids) accounts for 80% of the table sugar produced worldwide and is also a prime feedstock for biofuel production. However, very few studies are available for directly comparing Agrobacterium tumefaciens-mediated transfer of T-DNA (AMT) and biolistic transfer of minimal expression cassettes (BLT MC) regarding transgene complexity and expression stability. In this study, the transformation efficiency, transgene integration pattern, expression level, and expression stability were compared in the commercially important sugarcane cultivar CP88-1762. A total of 312 transgenic lines derived from AMT and 250 lines derived from BLT MC were identified by PCR from genomic DNA using nptII-specific primers. Lines were analyzed with both qPCR (TaqMan®) and NPTII ELISA to determine the nptII transgene copy number and expression level. The results of Southern blot analysis on selected lines were highly correlated to the qPCR results. There were no significant differences between the two transformation systems for transformation efficiency, frequency of single copy integration, or level and stability of transgene expression when carried out with the same expression cassette, tissue culture, and selection procedure in 12 independent experiments. These findings suggested that both BLT MC and AMT provide suitable platforms for generation of elite sugarcane events. |
| doi_str_mv | 10.1007/s11627-015-9710-0 |
| format | Article |
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However, very few studies are available for directly comparing Agrobacterium tumefaciens-mediated transfer of T-DNA (AMT) and biolistic transfer of minimal expression cassettes (BLT MC) regarding transgene complexity and expression stability. In this study, the transformation efficiency, transgene integration pattern, expression level, and expression stability were compared in the commercially important sugarcane cultivar CP88-1762. A total of 312 transgenic lines derived from AMT and 250 lines derived from BLT MC were identified by PCR from genomic DNA using nptII-specific primers. Lines were analyzed with both qPCR (TaqMan®) and NPTII ELISA to determine the nptII transgene copy number and expression level. The results of Southern blot analysis on selected lines were highly correlated to the qPCR results. There were no significant differences between the two transformation systems for transformation efficiency, frequency of single copy integration, or level and stability of transgene expression when carried out with the same expression cassette, tissue culture, and selection procedure in 12 independent experiments. These findings suggested that both BLT MC and AMT provide suitable platforms for generation of elite sugarcane events.</description><identifier>ISSN: 1054-5476</identifier><identifier>EISSN: 1475-2689</identifier><identifier>DOI: 10.1007/s11627-015-9710-0</identifier><language>eng</language><publisher>New York: Springer US</publisher><subject>Agrobacterium ; Biofuels ; biolistics ; Biomedical and Life Sciences ; BIOTECHNOLOGY ; Cell Biology ; Cultivars ; Deoxyribonucleic acid ; Developmental Biology ; DNA ; enzyme-linked immunosorbent assay ; feedstocks ; Flowers & plants ; fuel production ; gene expression ; Genes ; genetically modified organisms ; Genomes ; Hybrids ; in vitro studies ; Life Sciences ; Methods ; Plant Breeding/Biotechnology ; Plant Genetics and Genomics ; Plant Sciences ; Plasmids ; quantitative polymerase chain reaction ; Rodents ; Saccharum ; Southern blotting ; Sugarcane ; tissue culture ; transfer DNA ; transgenes ; white sugar</subject><ispartof>In vitro cellular & developmental biology. Plant, 2015-12, Vol.51 (6), p.603-611</ispartof><rights>2015 Society for In Vitro Biology</rights><rights>The Society for In Vitro Biology 2015</rights><rights>Copyright Society for In Vitro Biology Dec 2015</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c432t-5e975eaa3ae7f86b84497952b02d4d54b396612da864dfff16d23e69ee1547db3</citedby><cites>FETCH-LOGICAL-c432t-5e975eaa3ae7f86b84497952b02d4d54b396612da864dfff16d23e69ee1547db3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.jstor.org/stable/pdf/24596488$$EPDF$$P50$$Gjstor$$H</linktopdf><linktohtml>$$Uhttps://www.jstor.org/stable/24596488$$EHTML$$P50$$Gjstor$$H</linktohtml><link.rule.ids>314,776,780,799,27901,27902,41464,42533,51294,57992,58225</link.rule.ids></links><search><creatorcontrib>Wu, Hao</creatorcontrib><creatorcontrib>Awan, Faisal Saeed</creatorcontrib><creatorcontrib>Vilarinho, Aloisio</creatorcontrib><creatorcontrib>Zeng, Qianchun</creatorcontrib><creatorcontrib>Kannan, Baskaran</creatorcontrib><creatorcontrib>Phipps, Tenisha</creatorcontrib><creatorcontrib>McCuiston, Jamie</creatorcontrib><creatorcontrib>Wang, Wenling</creatorcontrib><creatorcontrib>Caffall, Kerry</creatorcontrib><creatorcontrib>Altpeter, Fredy</creatorcontrib><title>Transgene integration complexity and expression stability following biolistic or Agrobacterium-mediated transformation of sugarcane</title><title>In vitro cellular & developmental biology. Plant</title><addtitle>In Vitro Cell.Dev.Biol.-Plant</addtitle><description>Sugarcane (Saccharum spp. hybrids) accounts for 80% of the table sugar produced worldwide and is also a prime feedstock for biofuel production. However, very few studies are available for directly comparing Agrobacterium tumefaciens-mediated transfer of T-DNA (AMT) and biolistic transfer of minimal expression cassettes (BLT MC) regarding transgene complexity and expression stability. In this study, the transformation efficiency, transgene integration pattern, expression level, and expression stability were compared in the commercially important sugarcane cultivar CP88-1762. A total of 312 transgenic lines derived from AMT and 250 lines derived from BLT MC were identified by PCR from genomic DNA using nptII-specific primers. Lines were analyzed with both qPCR (TaqMan®) and NPTII ELISA to determine the nptII transgene copy number and expression level. The results of Southern blot analysis on selected lines were highly correlated to the qPCR results. There were no significant differences between the two transformation systems for transformation efficiency, frequency of single copy integration, or level and stability of transgene expression when carried out with the same expression cassette, tissue culture, and selection procedure in 12 independent experiments. These findings suggested that both BLT MC and AMT provide suitable platforms for generation of elite sugarcane events.</description><subject>Agrobacterium</subject><subject>Biofuels</subject><subject>biolistics</subject><subject>Biomedical and Life Sciences</subject><subject>BIOTECHNOLOGY</subject><subject>Cell Biology</subject><subject>Cultivars</subject><subject>Deoxyribonucleic acid</subject><subject>Developmental Biology</subject><subject>DNA</subject><subject>enzyme-linked immunosorbent assay</subject><subject>feedstocks</subject><subject>Flowers & plants</subject><subject>fuel production</subject><subject>gene expression</subject><subject>Genes</subject><subject>genetically modified organisms</subject><subject>Genomes</subject><subject>Hybrids</subject><subject>in vitro studies</subject><subject>Life Sciences</subject><subject>Methods</subject><subject>Plant Breeding/Biotechnology</subject><subject>Plant Genetics and Genomics</subject><subject>Plant Sciences</subject><subject>Plasmids</subject><subject>quantitative polymerase chain reaction</subject><subject>Rodents</subject><subject>Saccharum</subject><subject>Southern blotting</subject><subject>Sugarcane</subject><subject>tissue culture</subject><subject>transfer DNA</subject><subject>transgenes</subject><subject>white sugar</subject><issn>1054-5476</issn><issn>1475-2689</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>BENPR</sourceid><recordid>eNp9UU1r3TAQNKWFpkl-QA6lgp6VSrI-7GMI_QgEemhyFrK1MnrY0utKjybn_PHauJSeetqFmdnZnW2aK86uOWPmU-FcC0MZV7Q3nFH2qjnj0igqdNe_XnumJFXS6LfNu1IOjDHOuDlrXh7QpTJBAhJThQldjTmRMS_HGZ5ifSYueQJPR4RSNqRUN8R5A0Ke5_wrpokMMc-x1DiSjORmwjy4sQLG00IX8NFV8KRuPiHjshvkQMppcji6BBfNm-DmApd_6nnz-OXzw-03ev_9693tzT0dZSsqVdAbBc61Dkzo9NBJ2ZteiYEJL72SQ9trzYV3nZY-hMC1Fy3oHoCvh_uhPW8-7nOPmH-eoFR7yCdMq6XlnVFtq5jqVhbfWSPmUhCCPWJcHD5bzuyWtd2ztmvWdsvaslUjdk1ZuWkC_Gfyf0Tvd9Gh1Ix_XYRUvZbdtsiHHQ8uWzdhLPbxh2Bcb79bj5ftb3d9mFA</recordid><startdate>20151201</startdate><enddate>20151201</enddate><creator>Wu, Hao</creator><creator>Awan, Faisal Saeed</creator><creator>Vilarinho, Aloisio</creator><creator>Zeng, Qianchun</creator><creator>Kannan, Baskaran</creator><creator>Phipps, Tenisha</creator><creator>McCuiston, Jamie</creator><creator>Wang, Wenling</creator><creator>Caffall, Kerry</creator><creator>Altpeter, Fredy</creator><general>Springer US</general><general>Springer</general><general>Springer Nature B.V</general><scope>FBQ</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>4T-</scope><scope>4U-</scope><scope>7X2</scope><scope>7X7</scope><scope>7XB</scope><scope>88A</scope><scope>88I</scope><scope>8AF</scope><scope>8AO</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>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0K</scope><scope>M0S</scope><scope>M2P</scope><scope>M7P</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>Q9U</scope><scope>S0X</scope></search><sort><creationdate>20151201</creationdate><title>Transgene integration complexity and expression stability following biolistic or Agrobacterium-mediated transformation of sugarcane</title><author>Wu, Hao ; Awan, Faisal Saeed ; Vilarinho, Aloisio ; Zeng, Qianchun ; Kannan, Baskaran ; Phipps, Tenisha ; McCuiston, Jamie ; Wang, Wenling ; Caffall, Kerry ; Altpeter, Fredy</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c432t-5e975eaa3ae7f86b84497952b02d4d54b396612da864dfff16d23e69ee1547db3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Agrobacterium</topic><topic>Biofuels</topic><topic>biolistics</topic><topic>Biomedical and Life Sciences</topic><topic>BIOTECHNOLOGY</topic><topic>Cell Biology</topic><topic>Cultivars</topic><topic>Deoxyribonucleic acid</topic><topic>Developmental Biology</topic><topic>DNA</topic><topic>enzyme-linked immunosorbent assay</topic><topic>feedstocks</topic><topic>Flowers & plants</topic><topic>fuel production</topic><topic>gene expression</topic><topic>Genes</topic><topic>genetically modified organisms</topic><topic>Genomes</topic><topic>Hybrids</topic><topic>in vitro studies</topic><topic>Life Sciences</topic><topic>Methods</topic><topic>Plant Breeding/Biotechnology</topic><topic>Plant Genetics and Genomics</topic><topic>Plant Sciences</topic><topic>Plasmids</topic><topic>quantitative polymerase chain reaction</topic><topic>Rodents</topic><topic>Saccharum</topic><topic>Southern blotting</topic><topic>Sugarcane</topic><topic>tissue culture</topic><topic>transfer DNA</topic><topic>transgenes</topic><topic>white sugar</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Wu, Hao</creatorcontrib><creatorcontrib>Awan, Faisal Saeed</creatorcontrib><creatorcontrib>Vilarinho, Aloisio</creatorcontrib><creatorcontrib>Zeng, Qianchun</creatorcontrib><creatorcontrib>Kannan, Baskaran</creatorcontrib><creatorcontrib>Phipps, Tenisha</creatorcontrib><creatorcontrib>McCuiston, Jamie</creatorcontrib><creatorcontrib>Wang, Wenling</creatorcontrib><creatorcontrib>Caffall, Kerry</creatorcontrib><creatorcontrib>Altpeter, Fredy</creatorcontrib><collection>AGRIS</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Docstoc</collection><collection>University Readers</collection><collection>Agricultural Science Collection</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Biology Database (Alumni Edition)</collection><collection>Science Database (Alumni Edition)</collection><collection>STEM Database</collection><collection>ProQuest Pharma Collection</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>Agricultural & Environmental Science Collection</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection (ProQuest)</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</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>Agricultural Science Database</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Science Database</collection><collection>Biological Science Database</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central Basic</collection><collection>SIRS Editorial</collection><jtitle>In vitro cellular & developmental biology. Plant</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Wu, Hao</au><au>Awan, Faisal Saeed</au><au>Vilarinho, Aloisio</au><au>Zeng, Qianchun</au><au>Kannan, Baskaran</au><au>Phipps, Tenisha</au><au>McCuiston, Jamie</au><au>Wang, Wenling</au><au>Caffall, Kerry</au><au>Altpeter, Fredy</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Transgene integration complexity and expression stability following biolistic or Agrobacterium-mediated transformation of sugarcane</atitle><jtitle>In vitro cellular & developmental biology. Plant</jtitle><stitle>In Vitro Cell.Dev.Biol.-Plant</stitle><date>2015-12-01</date><risdate>2015</risdate><volume>51</volume><issue>6</issue><spage>603</spage><epage>611</epage><pages>603-611</pages><issn>1054-5476</issn><eissn>1475-2689</eissn><abstract>Sugarcane (Saccharum spp. hybrids) accounts for 80% of the table sugar produced worldwide and is also a prime feedstock for biofuel production. However, very few studies are available for directly comparing Agrobacterium tumefaciens-mediated transfer of T-DNA (AMT) and biolistic transfer of minimal expression cassettes (BLT MC) regarding transgene complexity and expression stability. In this study, the transformation efficiency, transgene integration pattern, expression level, and expression stability were compared in the commercially important sugarcane cultivar CP88-1762. A total of 312 transgenic lines derived from AMT and 250 lines derived from BLT MC were identified by PCR from genomic DNA using nptII-specific primers. Lines were analyzed with both qPCR (TaqMan®) and NPTII ELISA to determine the nptII transgene copy number and expression level. The results of Southern blot analysis on selected lines were highly correlated to the qPCR results. There were no significant differences between the two transformation systems for transformation efficiency, frequency of single copy integration, or level and stability of transgene expression when carried out with the same expression cassette, tissue culture, and selection procedure in 12 independent experiments. These findings suggested that both BLT MC and AMT provide suitable platforms for generation of elite sugarcane events.</abstract><cop>New York</cop><pub>Springer US</pub><doi>10.1007/s11627-015-9710-0</doi><tpages>9</tpages></addata></record> |
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| ispartof | In vitro cellular & developmental biology. Plant, 2015-12, Vol.51 (6), p.603-611 |
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| subjects | Agrobacterium Biofuels biolistics Biomedical and Life Sciences BIOTECHNOLOGY Cell Biology Cultivars Deoxyribonucleic acid Developmental Biology DNA enzyme-linked immunosorbent assay feedstocks Flowers & plants fuel production gene expression Genes genetically modified organisms Genomes Hybrids in vitro studies Life Sciences Methods Plant Breeding/Biotechnology Plant Genetics and Genomics Plant Sciences Plasmids quantitative polymerase chain reaction Rodents Saccharum Southern blotting Sugarcane tissue culture transfer DNA transgenes white sugar |
| title | Transgene integration complexity and expression stability following biolistic or Agrobacterium-mediated transformation of sugarcane |
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