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| Format: | Artículo científico |
| Langue: | en |
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Universidad Autónoma de Baja California
2015
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| Accès en ligne: | https://www.redalyc.org/articulo.oa?id=48036868002 |
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| _version_ | 1866814300033646592 |
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| author | Mario Fernando Ortiz-Matamoros |
| author_facet | Mario Fernando Ortiz-Matamoros |
| contents | Transient transformation of cul tured photosynthetic dinoflagell ates (Symbiodinium spp.) with plant-targeted vectors Mario Fernando Ortiz-Matamoros Marco A Villanueva Tania Islas-Flores Ciencias de la Tierra GFP AtRACK1C Symbiodinium transformation Dinoflagellates Reproducible and reliable genetic transformation methods are a key tool for understanding the physiology and cell biology of Symbiodinium. Nevertheless, transformation methods previously applied to ce lls such as microalgae, including those util izing glass beads, have not been tested on th ese microorganisms. Here, we report a simp le, transient transformation met hod, which allowed plasmid inco rporation into three distinct clades of cultured Symbiodinium cells with the plant-targeted plasmid pCB302 harboring sequenc es encoding a fusion of green fluorescent protein ( gfp ) with RACK1C from Arabidopsis thaliana ( AtRACK1C ). Accessibility of the plasmid to the resistant cell wall and through the plasma membrane of t he dinoflagellates was achieved through vigorous shaking in the presence of glass beads and po lyethylene glycol. A resistance gene to the herbicide Basta allowed approp riate selection in the photosynth etic cells. The transformation frequency per every 10 6 cells was 107±7 transformants for Symbiodinium kawagutii , 74 ± 8 for Symbiodinium microadriaticum ssp. microadriaticum, and 65±5 for Symbiodinium sp. Mf11. Moreover, Symbiodinium pulchrorum cultures were successfully trans formed with a different vector (pCAMBIA- FA B D 2 - gfp ) under the same conditions, furt her validating our procedure. The observation of green fluorescent emission from the cell cytoplasm in all performed t ransformations indicated that the procedure allowed the heterologous plasmids to enter and undergo expression in the Symbiodinium cells. The success of this transient transformation method opens inter esting possibilities for functional genomics studies in Symbiodinium spp. 2015 artículo científico 0185-3880 https://www.redalyc.org/articulo.oa?id=48036868002 en http://www.redalyc.org/revista.oa?id=480 Ciencias Marinas application/pdf Universidad Autónoma de Baja California Ciencias Marinas (México) Num.1 Vol.41 |
| format | Artículo científico |
| id | redalyc_48036868002 |
| language | en |
| publishDate | 2015 |
| publisher | Universidad Autónoma de Baja California |
| spellingShingle | Transient transformation of cul tured photosynthetic dinoflagell ates (Symbiodinium spp.) with plant-targeted vectors Mario Fernando Ortiz-Matamoros Ciencias de la Tierra GFP AtRACK1C Symbiodinium transformation Dinoflagellates Transient transformation of cul tured photosynthetic dinoflagell ates (Symbiodinium spp.) with plant-targeted vectors Mario Fernando Ortiz-Matamoros Marco A Villanueva Tania Islas-Flores Ciencias de la Tierra GFP AtRACK1C Symbiodinium transformation Dinoflagellates Reproducible and reliable genetic transformation methods are a key tool for understanding the physiology and cell biology of Symbiodinium. Nevertheless, transformation methods previously applied to ce lls such as microalgae, including those util izing glass beads, have not been tested on th ese microorganisms. Here, we report a simp le, transient transformation met hod, which allowed plasmid inco rporation into three distinct clades of cultured Symbiodinium cells with the plant-targeted plasmid pCB302 harboring sequenc es encoding a fusion of green fluorescent protein ( gfp ) with RACK1C from Arabidopsis thaliana ( AtRACK1C ). Accessibility of the plasmid to the resistant cell wall and through the plasma membrane of t he dinoflagellates was achieved through vigorous shaking in the presence of glass beads and po lyethylene glycol. A resistance gene to the herbicide Basta allowed approp riate selection in the photosynth etic cells. The transformation frequency per every 10 6 cells was 107±7 transformants for Symbiodinium kawagutii , 74 ± 8 for Symbiodinium microadriaticum ssp. microadriaticum, and 65±5 for Symbiodinium sp. Mf11. Moreover, Symbiodinium pulchrorum cultures were successfully trans formed with a different vector (pCAMBIA- FA B D 2 - gfp ) under the same conditions, furt her validating our procedure. The observation of green fluorescent emission from the cell cytoplasm in all performed t ransformations indicated that the procedure allowed the heterologous plasmids to enter and undergo expression in the Symbiodinium cells. The success of this transient transformation method opens inter esting possibilities for functional genomics studies in Symbiodinium spp. 2015 artículo científico 0185-3880 https://www.redalyc.org/articulo.oa?id=48036868002 en http://www.redalyc.org/revista.oa?id=480 Ciencias Marinas application/pdf Universidad Autónoma de Baja California Ciencias Marinas (México) Num.1 Vol.41 |
| title | Transient transformation of cul tured photosynthetic dinoflagell ates (Symbiodinium spp.) with plant-targeted vectors |
| topic | Ciencias de la Tierra GFP AtRACK1C Symbiodinium transformation Dinoflagellates |
| url | https://www.redalyc.org/articulo.oa?id=48036868002 |