Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco Protoplasts
In view of the recent finding that different T-DNAs tend to ligate and integrate as repeats at single chromosomal positions, the frequency of transformation and cotransformation was determined during cocultivation of Arabidopsis thaliana root explants and Nicotiana tabacum protoplasts with two Agrob...
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
The American Phytopathological Society
1998-06-01
|
| Series: | Molecular Plant-Microbe Interactions |
| Subjects: | |
| Online Access: | https://apsjournals.apsnet.org/doi/10.1094/MPMI.1998.11.6.449 |
| _version_ | 1831672635313881088 |
|---|---|
| author | Sylvie De Buck Anni Jacobs Marc Van Montagu Ann Depicker |
| author_facet | Sylvie De Buck Anni Jacobs Marc Van Montagu Ann Depicker |
| author_sort | Sylvie De Buck |
| collection | DOAJ |
| description | In view of the recent finding that different T-DNAs tend to ligate and integrate as repeats at single chromosomal positions, the frequency of transformation and cotransformation was determined during cocultivation of Arabidopsis thaliana root explants and Nicotiana tabacum protoplasts with two Agrobacterium strains. The transformation frequency of unselected A. thaliana shoots was lower than 1% whereas that of cocultivated tobacco protoplasts was approximately 18%. The cotransformation frequencies, defined as the frequencies with which cells transformed with a first T-DNA contained a second unselected T-DNA, were approximately 40% reproducible, irrespective of the selection, the transformation frequency, and the plant system used. Extrapolation of these results suggests that at least two independently transferred T-DNAs were present in 64% of the transformed plant cells. Molecular analysis of cocultivated N. tabacum shoots regenerated on nonselective medium showed that only a few transformants had a silenced (2/46) or truncated (1/46) T-DNA. Therefore, most integrated T-DNAs expressed their selectable or screenable markers in primary transgenic plants. Remarkably, 10 to 30% of the selected A. thaliana shoots or progenies lost the T-DNA marker they were selected on. As these regenerants contained the unselected T-DNA with a high frequency (17%), these selected plants might result from the expression of unstable, transiently expressed T-DNAs. In conclusion, a significant part of the T-DNAs is lost from the transformed cells. |
| first_indexed | 2024-12-19T23:56:04Z |
| format | Article |
| id | doaj.art-1c9b4f475a794665b313ea903a98c1f1 |
| institution | Directory Open Access Journal |
| issn | 0894-0282 1943-7706 |
| language | English |
| last_indexed | 2024-12-19T23:56:04Z |
| publishDate | 1998-06-01 |
| publisher | The American Phytopathological Society |
| record_format | Article |
| series | Molecular Plant-Microbe Interactions |
| spelling | doaj.art-1c9b4f475a794665b313ea903a98c1f12022-12-21T20:00:59ZengThe American Phytopathological SocietyMolecular Plant-Microbe Interactions0894-02821943-77061998-06-0111644945710.1094/MPMI.1998.11.6.449Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco ProtoplastsSylvie De BuckAnni JacobsMarc Van MontaguAnn DepickerIn view of the recent finding that different T-DNAs tend to ligate and integrate as repeats at single chromosomal positions, the frequency of transformation and cotransformation was determined during cocultivation of Arabidopsis thaliana root explants and Nicotiana tabacum protoplasts with two Agrobacterium strains. The transformation frequency of unselected A. thaliana shoots was lower than 1% whereas that of cocultivated tobacco protoplasts was approximately 18%. The cotransformation frequencies, defined as the frequencies with which cells transformed with a first T-DNA contained a second unselected T-DNA, were approximately 40% reproducible, irrespective of the selection, the transformation frequency, and the plant system used. Extrapolation of these results suggests that at least two independently transferred T-DNAs were present in 64% of the transformed plant cells. Molecular analysis of cocultivated N. tabacum shoots regenerated on nonselective medium showed that only a few transformants had a silenced (2/46) or truncated (1/46) T-DNA. Therefore, most integrated T-DNAs expressed their selectable or screenable markers in primary transgenic plants. Remarkably, 10 to 30% of the selected A. thaliana shoots or progenies lost the T-DNA marker they were selected on. As these regenerants contained the unselected T-DNA with a high frequency (17%), these selected plants might result from the expression of unstable, transiently expressed T-DNAs. In conclusion, a significant part of the T-DNAs is lost from the transformed cells.https://apsjournals.apsnet.org/doi/10.1094/MPMI.1998.11.6.449gene silencingT-DNA transfertransgene expression |
| spellingShingle | Sylvie De Buck Anni Jacobs Marc Van Montagu Ann Depicker Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco Protoplasts Molecular Plant-Microbe Interactions gene silencing T-DNA transfer transgene expression |
| title | Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco Protoplasts |
| title_full | Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco Protoplasts |
| title_fullStr | Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco Protoplasts |
| title_full_unstemmed | Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco Protoplasts |
| title_short | Agrobacterium tumefaciens Transformation and Cotransformation Frequencies of Arabidopsis thaliana Root Explants and Tobacco Protoplasts |
| title_sort | agrobacterium tumefaciens transformation and cotransformation frequencies of arabidopsis thaliana root explants and tobacco protoplasts |
| topic | gene silencing T-DNA transfer transgene expression |
| url | https://apsjournals.apsnet.org/doi/10.1094/MPMI.1998.11.6.449 |
| work_keys_str_mv | AT sylviedebuck agrobacteriumtumefacienstransformationandcotransformationfrequenciesofarabidopsisthalianarootexplantsandtobaccoprotoplasts AT annijacobs agrobacteriumtumefacienstransformationandcotransformationfrequenciesofarabidopsisthalianarootexplantsandtobaccoprotoplasts AT marcvanmontagu agrobacteriumtumefacienstransformationandcotransformationfrequenciesofarabidopsisthalianarootexplantsandtobaccoprotoplasts AT anndepicker agrobacteriumtumefacienstransformationandcotransformationfrequenciesofarabidopsisthalianarootexplantsandtobaccoprotoplasts |