Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce
Background: Lettuce is a globally important leafy vegetable and a model plant for biotechnology due to its adaptability to tissue culture and stable genetic transformation. Lettuce is also crucial for functional genomics research in the Asteraceae which includes species of great agronomical importan...
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| Format: | Artículo |
| Language: | Inglés |
| Published: |
2018
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| Online Access: | http://hdl.handle.net/20.500.12123/2147 https://www.sciencedirect.com/science/article/pii/S0717345817300611 https://doi.org/10.1016/j.ejbt.2017.10.002 |
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| author | Darqui, Flavia Soledad Radonic, Laura Mabel Lopez, Nilda Ester Hopp, Horacio Esteban Lopez Bilbao, Marisa Gisela |
| author_browse | Darqui, Flavia Soledad Hopp, Horacio Esteban Lopez Bilbao, Marisa Gisela Lopez, Nilda Ester Radonic, Laura Mabel |
| author_facet | Darqui, Flavia Soledad Radonic, Laura Mabel Lopez, Nilda Ester Hopp, Horacio Esteban Lopez Bilbao, Marisa Gisela |
| author_sort | Darqui, Flavia Soledad |
| collection | INTA Digital |
| description | Background: Lettuce is a globally important leafy vegetable and a model plant for biotechnology due to its adaptability to tissue culture and stable genetic transformation. Lettuce is also crucial for functional genomics research in the Asteraceae which includes species of great agronomical importance. The development of
transgenic events implies the production of a large number of shoots that must be differentiated between transgenic and non-transgenic through the activity of the selective agent, being kanamycin the most popular.
Results: In this work we adjusted the selection conditions of transgenic seedlings to avoid any escapes, finding that threshold concentration of kanamycin was 75 mg/L. To monitor the selection system, we studied the morphological response of transgenic and non-transgenic seedlings in presence of kanamycin to look for a
visual morphological marker. Several traits like shoot length, primary root length, number of leaves, fresh weight, and appearance of the aerial part and development of lateral roots were affected in non-transgenic seedlings after 30 d of culture in selective media. However, only lateral root development showed an early, qualitative and reliable association with nptII presence, as corroborated by PCR detection. Applied in successive transgenic progenies, this method of selection combined with morphological follow-up allowed selecting the
homozygous presence of nptII gene in 100% of the analyzed plants from T2 to T5.
Conclusions: This protocol allows a simplified scaling-up of the production of multiple homozygous transgenic progeny lines in the early generations avoiding expensive and time-consuming molecular assays. |
| format | Artículo |
| id | INTA2147 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2018 |
| publishDateRange | 2018 |
| publishDateSort | 2018 |
| record_format | dspace |
| spelling | INTA21472018-03-28T14:54:37Z Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce Darqui, Flavia Soledad Radonic, Laura Mabel Lopez, Nilda Ester Hopp, Horacio Esteban Lopez Bilbao, Marisa Gisela Lechugas Plantas Transgénicas Hortalizas de Hoja Lettuces Transgenic Plants Leaf Vegetables Background: Lettuce is a globally important leafy vegetable and a model plant for biotechnology due to its adaptability to tissue culture and stable genetic transformation. Lettuce is also crucial for functional genomics research in the Asteraceae which includes species of great agronomical importance. The development of transgenic events implies the production of a large number of shoots that must be differentiated between transgenic and non-transgenic through the activity of the selective agent, being kanamycin the most popular. Results: In this work we adjusted the selection conditions of transgenic seedlings to avoid any escapes, finding that threshold concentration of kanamycin was 75 mg/L. To monitor the selection system, we studied the morphological response of transgenic and non-transgenic seedlings in presence of kanamycin to look for a visual morphological marker. Several traits like shoot length, primary root length, number of leaves, fresh weight, and appearance of the aerial part and development of lateral roots were affected in non-transgenic seedlings after 30 d of culture in selective media. However, only lateral root development showed an early, qualitative and reliable association with nptII presence, as corroborated by PCR detection. Applied in successive transgenic progenies, this method of selection combined with morphological follow-up allowed selecting the homozygous presence of nptII gene in 100% of the analyzed plants from T2 to T5. Conclusions: This protocol allows a simplified scaling-up of the production of multiple homozygous transgenic progeny lines in the early generations avoiding expensive and time-consuming molecular assays. Inst. de Biotecnología Fil: Darqui, Flavia Soledad. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Radonic, Laura Mabel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina Fil: Lopez, Nilda Ester. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina Fil: Hopp, Horacio Esteban. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina Fil: López Bilbao, Marisa Gisela. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina 2018-03-28T14:36:25Z 2018-03-28T14:36:25Z 2018-01 info:eu-repo/semantics/article info:ar-repo/semantics/artículo info:eu-repo/semantics/publishedVersion http://hdl.handle.net/20.500.12123/2147 https://www.sciencedirect.com/science/article/pii/S0717345817300611 0717-3458 https://doi.org/10.1016/j.ejbt.2017.10.002 eng info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-sa/4.0/ Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0) application/pdf Electronic Journal of Biotechnology 31 : 1-9 (January 2018) |
| spellingShingle | Lechugas Plantas Transgénicas Hortalizas de Hoja Lettuces Transgenic Plants Leaf Vegetables Darqui, Flavia Soledad Radonic, Laura Mabel Lopez, Nilda Ester Hopp, Horacio Esteban Lopez Bilbao, Marisa Gisela Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce |
| title | Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce |
| title_full | Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce |
| title_fullStr | Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce |
| title_full_unstemmed | Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce |
| title_short | Simplified methodology for large scale isolation of homozygous transgenic lines of lettuce |
| title_sort | simplified methodology for large scale isolation of homozygous transgenic lines of lettuce |
| topic | Lechugas Plantas Transgénicas Hortalizas de Hoja Lettuces Transgenic Plants Leaf Vegetables |
| url | http://hdl.handle.net/20.500.12123/2147 https://www.sciencedirect.com/science/article/pii/S0717345817300611 https://doi.org/10.1016/j.ejbt.2017.10.002 |
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