Commercial scale genetic transformation of mature seed embryo explants in maize
A novel, efficient maize genetic transformation system was developed using -mediated transformation of embryo explants from mature seeds. Seeds from field grown plants were sterilized and crushed to isolate embryo explants consisting of the coleoptile, leaf primordia, and shoot apical meristem which...
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| Published in | Frontiers in plant science Vol. 13; p. 1056190 |
|---|---|
| Main Authors | , , , , , , , , , , , , |
| Format | Journal Article |
| Language | English |
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Frontiers Media S.A
29.11.2022
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| Abstract | A novel, efficient maize genetic transformation system was developed using
-mediated transformation of embryo explants from mature seeds. Seeds from field grown plants were sterilized and crushed to isolate embryo explants consisting of the coleoptile, leaf primordia, and shoot apical meristem which were then purified from the ground seed bulk preparation. The infection of relevant tissues of seed embryo explants (SEEs) by
was improved by the centrifugation of the explants. Transgenic plants were obtained by multiple bud induction on high cytokinin media, followed by plant regeneration on hormone-free medium. Three different selectable markers (
, and
) were successfully used for producing transgenic plants. Stable integration of transgenes in the maize genome was demonstrated by molecular analyses and germline transmission of the inserted transgenes to the next generation was confirmed by pollen segregation and progeny analysis. Phenotypic evidence for chimeric transgenic tissue was frequently observed in initial experiments but was significantly reduced by including a second bud induction step with optimized cytokinin concentration. Additional improvements, including culturing explants at an elevated temperature during bud induction led to the development of a revolutionary system for efficient transgenic plant production and genome editing. To our knowledge, this is the first report of successful transgenic plant regeneration through
-mediated transformation of maize mature SEEs. This system starts with mature seed that can be produced in large volumes and the SEEs explants are storable. It has significant advantages in terms of scalability and flexibility over methods that rely on immature explants. |
|---|---|
| AbstractList | A novel, efficient maize genetic transformation system was developed using
Agrobacterium
-mediated transformation of embryo explants from mature seeds. Seeds from field grown plants were sterilized and crushed to isolate embryo explants consisting of the coleoptile, leaf primordia, and shoot apical meristem which were then purified from the ground seed bulk preparation. The infection of relevant tissues of seed embryo explants (SEEs) by
Agrobacterium
was improved by the centrifugation of the explants. Transgenic plants were obtained by multiple bud induction on high cytokinin media, followed by plant regeneration on hormone-free medium. Three different selectable markers (
cp4 epsps, aadA
, and
nptII
) were successfully used for producing transgenic plants. Stable integration of transgenes in the maize genome was demonstrated by molecular analyses and germline transmission of the inserted transgenes to the next generation was confirmed by pollen segregation and progeny analysis. Phenotypic evidence for chimeric transgenic tissue was frequently observed in initial experiments but was significantly reduced by including a second bud induction step with optimized cytokinin concentration. Additional improvements, including culturing explants at an elevated temperature during bud induction led to the development of a revolutionary system for efficient transgenic plant production and genome editing. To our knowledge, this is the first report of successful transgenic plant regeneration through
Agrobacterium
-mediated transformation of maize mature SEEs. This system starts with mature seed that can be produced in large volumes and the SEEs explants are storable. It has significant advantages in terms of scalability and flexibility over methods that rely on immature explants. A novel, efficient maize genetic transformation system was developed using Agrobacterium-mediated transformation of embryo explants from mature seeds. Seeds from field grown plants were sterilized and crushed to isolate embryo explants consisting of the coleoptile, leaf primordia, and shoot apical meristem which were then purified from the ground seed bulk preparation. The infection of relevant tissues of seed embryo explants (SEEs) by Agrobacterium was improved by the centrifugation of the explants. Transgenic plants were obtained by multiple bud induction on high cytokinin media, followed by plant regeneration on hormone-free medium. Three different selectable markers (cp4 epsps, aadA, and nptII) were successfully used for producing transgenic plants. Stable integration of transgenes in the maize genome was demonstrated by molecular analyses and germline transmission of the inserted transgenes to the next generation was confirmed by pollen segregation and progeny analysis. Phenotypic evidence for chimeric transgenic tissue was frequently observed in initial experiments but was significantly reduced by including a second bud induction step with optimized cytokinin concentration. Additional improvements, including culturing explants at an elevated temperature during bud induction led to the development of a revolutionary system for efficient transgenic plant production and genome editing. To our knowledge, this is the first report of successful transgenic plant regeneration through Agrobacterium-mediated transformation of maize mature SEEs. This system starts with mature seed that can be produced in large volumes and the SEEs explants are storable. It has significant advantages in terms of scalability and flexibility over methods that rely on immature explants. A novel, efficient maize genetic transformation system was developed using -mediated transformation of embryo explants from mature seeds. Seeds from field grown plants were sterilized and crushed to isolate embryo explants consisting of the coleoptile, leaf primordia, and shoot apical meristem which were then purified from the ground seed bulk preparation. The infection of relevant tissues of seed embryo explants (SEEs) by was improved by the centrifugation of the explants. Transgenic plants were obtained by multiple bud induction on high cytokinin media, followed by plant regeneration on hormone-free medium. Three different selectable markers ( , and ) were successfully used for producing transgenic plants. Stable integration of transgenes in the maize genome was demonstrated by molecular analyses and germline transmission of the inserted transgenes to the next generation was confirmed by pollen segregation and progeny analysis. Phenotypic evidence for chimeric transgenic tissue was frequently observed in initial experiments but was significantly reduced by including a second bud induction step with optimized cytokinin concentration. Additional improvements, including culturing explants at an elevated temperature during bud induction led to the development of a revolutionary system for efficient transgenic plant production and genome editing. To our knowledge, this is the first report of successful transgenic plant regeneration through -mediated transformation of maize mature SEEs. This system starts with mature seed that can be produced in large volumes and the SEEs explants are storable. It has significant advantages in terms of scalability and flexibility over methods that rely on immature explants. A novel, efficient maize genetic transformation system was developed using Agrobacterium-mediated transformation of embryo explants from mature seeds. Seeds from field grown plants were sterilized and crushed to isolate embryo explants consisting of the coleoptile, leaf primordia, and shoot apical meristem which were then purified from the ground seed bulk preparation. The infection of relevant tissues of seed embryo explants (SEEs) by Agrobacterium was improved by the centrifugation of the explants. Transgenic plants were obtained by multiple bud induction on high cytokinin media, followed by plant regeneration on hormone-free medium. Three different selectable markers (cp4 epsps, aadA, and nptII) were successfully used for producing transgenic plants. Stable integration of transgenes in the maize genome was demonstrated by molecular analyses and germline transmission of the inserted transgenes to the next generation was confirmed by pollen segregation and progeny analysis. Phenotypic evidence for chimeric transgenic tissue was frequently observed in initial experiments but was significantly reduced by including a second bud induction step with optimized cytokinin concentration. Additional improvements, including culturing explants at an elevated temperature during bud induction led to the development of a revolutionary system for efficient transgenic plant production and genome editing. To our knowledge, this is the first report of successful transgenic plant regeneration through Agrobacterium-mediated transformation of maize mature SEEs. This system starts with mature seed that can be produced in large volumes and the SEEs explants are storable. It has significant advantages in terms of scalability and flexibility over methods that rely on immature explants.A novel, efficient maize genetic transformation system was developed using Agrobacterium-mediated transformation of embryo explants from mature seeds. Seeds from field grown plants were sterilized and crushed to isolate embryo explants consisting of the coleoptile, leaf primordia, and shoot apical meristem which were then purified from the ground seed bulk preparation. The infection of relevant tissues of seed embryo explants (SEEs) by Agrobacterium was improved by the centrifugation of the explants. Transgenic plants were obtained by multiple bud induction on high cytokinin media, followed by plant regeneration on hormone-free medium. Three different selectable markers (cp4 epsps, aadA, and nptII) were successfully used for producing transgenic plants. Stable integration of transgenes in the maize genome was demonstrated by molecular analyses and germline transmission of the inserted transgenes to the next generation was confirmed by pollen segregation and progeny analysis. Phenotypic evidence for chimeric transgenic tissue was frequently observed in initial experiments but was significantly reduced by including a second bud induction step with optimized cytokinin concentration. Additional improvements, including culturing explants at an elevated temperature during bud induction led to the development of a revolutionary system for efficient transgenic plant production and genome editing. To our knowledge, this is the first report of successful transgenic plant regeneration through Agrobacterium-mediated transformation of maize mature SEEs. This system starts with mature seed that can be produced in large volumes and the SEEs explants are storable. It has significant advantages in terms of scalability and flexibility over methods that rely on immature explants. |
| Author | Rivlin, Anatoly Kumpf, Jennifer Subbarao, Shubha Vaghchhipawala, Zarir Armstrong, Charles Martinell, Brian Ye, Xudong Shrawat, Ashok Saltarikos, M Annie Moeller, Lorena Chen, Yurong Williams, Edward Somers, David |
| AuthorAffiliation | 1 Plant Biotechnology, Bayer Crop Science , W. St. Louis, MO , United States 2 Agracetus Campus, Monsanto Company , Middleton, WI , United States 3 Mystic Research, Monsanto Company , Mystic, CT , United States |
| AuthorAffiliation_xml | – name: 1 Plant Biotechnology, Bayer Crop Science , W. St. Louis, MO , United States – name: 3 Mystic Research, Monsanto Company , Mystic, CT , United States – name: 2 Agracetus Campus, Monsanto Company , Middleton, WI , United States |
| Author_xml | – sequence: 1 givenname: Xudong surname: Ye fullname: Ye, Xudong organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 2 givenname: Ashok surname: Shrawat fullname: Shrawat, Ashok organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 3 givenname: Edward surname: Williams fullname: Williams, Edward organization: Agracetus Campus, Monsanto Company, Middleton, WI, United States – sequence: 4 givenname: Anatoly surname: Rivlin fullname: Rivlin, Anatoly organization: Agracetus Campus, Monsanto Company, Middleton, WI, United States – sequence: 5 givenname: Zarir surname: Vaghchhipawala fullname: Vaghchhipawala, Zarir organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 6 givenname: Lorena surname: Moeller fullname: Moeller, Lorena organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 7 givenname: Jennifer surname: Kumpf fullname: Kumpf, Jennifer organization: Mystic Research, Monsanto Company, Mystic, CT, United States – sequence: 8 givenname: Shubha surname: Subbarao fullname: Subbarao, Shubha organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 9 givenname: Brian surname: Martinell fullname: Martinell, Brian organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 10 givenname: Charles surname: Armstrong fullname: Armstrong, Charles organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 11 givenname: M Annie surname: Saltarikos fullname: Saltarikos, M Annie organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States – sequence: 12 givenname: David surname: Somers fullname: Somers, David organization: Mystic Research, Monsanto Company, Mystic, CT, United States – sequence: 13 givenname: Yurong surname: Chen fullname: Chen, Yurong organization: Plant Biotechnology, Bayer Crop Science, W. St. Louis, MO, United States |
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| Cites_doi | 10.1104/pp.110.4.1097 10.1111/pbi.13837 10.1007/BF02319006 10.1038/342837a0 10.1139/g02-120 10.1007/s00299-013-1519-x 10.1016/j.pbi.2020.06.007 10.1080/13102818.2014.907654 10.1093/jexbot/51.351.1713 10.1007/BF02822763 10.1023/A:1018321428210 10.1038/nbt0990-833 10.1104/pp.95.2.426 10.1007/s00299-003-0748-9 10.1007/s11240-006-9157-4 10.1038/s41587-020-0703-0 10.1007/BF00199967 10.1146/annurev.arplant.55.031903.141633 10.1038/s41477-022-01173-3 10.1023/B:MOLB.0000038005.73265.61 10.1038/nbt0795-677 10.1007/s00299-007-0398-4 10.1007/s11248-010-9458-6 10.1371/journal.pone.0200972 10.1007/BF00269560 10.2307/3869124 10.1007/s00299-006-0273-8 10.1007/s11032-021-01225-0 10.1038/nbt0888-923 10.1007/BF00199966 10.1105/tpc.16.00196 10.1002/ps.4652 10.1016/0168-9525(89)90101-7 10.1007/s00299-002-0513-5 10.1038/nbt0696-745 10.1007/BF02712670 10.3389/fpls.2020.572319 10.1007/BF00285191 10.1105/tpc.16.00124 10.1079/IVP2004616 10.1007/s11248-008-9169-4 10.1007/BF00034948 10.1007/s00425-006-0237-9 10.1079/IVP2002291 10.3389/fpls.2020.575283 10.1007/s00299-005-0058-5 10.1007/BF02822775 10.1007/s00299-015-1906-6 |
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| Copyright | Copyright © 2022 Ye, Shrawat, Williams, Rivlin, Vaghchhipawala, Moeller, Kumpf, Subbarao, Martinell, Armstrong, Saltarikos, Somers and Chen. Copyright © 2022 Ye, Shrawat, Williams, Rivlin, Vaghchhipawala, Moeller, Kumpf, Subbarao, Martinell, Armstrong, Saltarikos, Somers and Chen 2022 Ye, Shrawat, Williams, Rivlin, Vaghchhipawala, Moeller, Kumpf, Subbarao, Martinell, Armstrong, Saltarikos, Somers and Chen |
| Copyright_xml | – notice: Copyright © 2022 Ye, Shrawat, Williams, Rivlin, Vaghchhipawala, Moeller, Kumpf, Subbarao, Martinell, Armstrong, Saltarikos, Somers and Chen. – notice: Copyright © 2022 Ye, Shrawat, Williams, Rivlin, Vaghchhipawala, Moeller, Kumpf, Subbarao, Martinell, Armstrong, Saltarikos, Somers and Chen 2022 Ye, Shrawat, Williams, Rivlin, Vaghchhipawala, Moeller, Kumpf, Subbarao, Martinell, Armstrong, Saltarikos, Somers and Chen |
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| Keywords | Agrobacterium Zea mays meristem genotype-flexible maize transformation seed embryo explants (SEEs) organogenesis |
| Language | English |
| License | Copyright © 2022 Ye, Shrawat, Williams, Rivlin, Vaghchhipawala, Moeller, Kumpf, Subbarao, Martinell, Armstrong, Saltarikos, Somers and Chen. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
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| Notes | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Edited by: Phanikanth Jogam, Kakatiya University, India Reviewed by: Vijay Sheri, East Carolina University, United States; Sahil Mehta, University of Delhi, India; Dhirendra Fartyal, Agricutural Research Organization, Israel These authors have contributed equally to this work This article was submitted to Technical Advances in Plant Science, a section of the journal Frontiers in Plant Science |
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| Snippet | A novel, efficient maize genetic transformation system was developed using
-mediated transformation of embryo explants from mature seeds. Seeds from field... A novel, efficient maize genetic transformation system was developed using Agrobacterium -mediated transformation of embryo explants from mature seeds. Seeds... A novel, efficient maize genetic transformation system was developed using Agrobacterium-mediated transformation of embryo explants from mature seeds. Seeds... |
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| SubjectTerms | Agrobacterium maize transformation meristem organogenesis Plant Science seed embryo explants (SEEs) Zea mays |
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| Title | Commercial scale genetic transformation of mature seed embryo explants in maize |
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