Faster evaluation of induced floral sterility in transgenic early flowering poplar

A major concern over the use of transgenic trees is the potential for transgene dispersal through pollen and seeds. The incorporation of sterility inducing genes into transgenic lines of trees has been proposed to reduce or even avoid gene flow of transgenes into non-transgenic interbreeding species. The evaluation of strategies for the induction of sterility in transgenic forest tree species has been hindered by their long vegetative phases. In this study an early flowering 35

A saturated consensus linkage map of Picea abies including AFLP, SSR, STS, 5S rDNA and morphological markers

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Efficient evaluation of a gene containment system for poplar through early flowering induction

[EN] Key message The early flowering system HSP::AtFT allowed a fast evaluation of a gene containment system based on the construct PsEND1::barnase-barstar for poplar. Transgenic lines showed disturbed pollen development and sterility. Vertical gene transfer through pollen flow from transgenic or non-native plant species into their crossable natural relatives is a major concern. Gene containment approaches have been proposed to reduce or even avoid gene flow among tree species. However, evaluation of genetic containment strategies for trees is very difficult due to the long-generation times. Early flowering induction would allow faster evaluation of genetic containment in this case. Although no reliable methods were available for the induction of fertile flowers in poplar, recently, a new early flowering approach was developed. In this study, early flowering poplar lines containing the gene construct PsEND1::barnase-barstar were obtained. The PsEND1 promoter was chosen due to its early expression pattern, its versality and efficiency for generation of male-sterile plants fused to the barnase gene. RT-PCRs confirmed barnase gene activity in flowers, and pollen development was disturbed, leading to sterile flowers. The system developed in this study represents a valuable tool for gene containment studies in forest tree species.Open Access funding provided by Projekt DEAL. This work was funded with a scholarship by the Deutscher Akademischer Austauschdienst (DAAD). We thank S. Bein, D. Ebbinghaus, and A. Worm for helpful technical assistance in the laboratory, and the greenhouse staff (M. Hunger, G. Wiemann, R. Ebbinghaus, and M. Spauszus) for plant cultivation.Briones, MV.; Hoenicka, H.; Cañas Clemente, LA.; Beltran Porter, JP.; Hanelt, D.; Sharry, S.; Fladung, M. (2020). Efficient evaluation of a gene containment system for poplar through early flowering induction. 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Optimización de un sistema de modificación génetica de álamos para la incorporación de genes inductores de floración temprana y androesterilidad

Las especies de árboles forestales alcanzan la madurez reproductiva después de muchos años o incluso décadas de crecimiento juvenil. Los tratamientos especiales, basados en hormonas vegetales, inhibidores del crecimiento, manipulación del ambiente de crecimiento o métodos físicos, han promovido el desarrollo de flores en plantas juveniles de varias especies de árboles forestales, pero no en álamo. Sólo la ingeniería genética ha permitido que la floración temprana sea inducida en plantas juveniles de álamo. La liberación al medio de álamos genéticamente modificados (GM) tiene limitantes, tanto desde el punto de vista ambiental, como comercial y de percepción pública. La obtención de álamos GM estériles facilitarían la liberación de los mismos al medio de forma segura. El objetivo general de este trabajo fue optimizar un sistema de modificación génetica de Populus sp. para la incorporación, en simultáneo, de genes de floración temprana y androesterilidad, acelerando el tiempo de obtención de líneas transgénicas dobles. Se logró ajustar un método eficiente de regeneración de plantas in vitro de clones recalcitrantes de Populus deltoides y clones del híbrido Populus x canadensis. Se obtuvieron líneas transgénicas dobles de P. tremula L, conteniendo en simultaneo los genes inductores de la floración temprana y la androesterilidad. También se obtuvieron líneas transgénicas simples de P. deltoides y P. x euramericana, portadoras del gen inductor de la floración temprana. Todas las líneas obtenidas regeneraron en plantas completas donde pudo confirmarse, con técnicas moleculares, la presencia de los transgenes. Se evaluó el desarrollo de la floración precoz inducida por el constructo HSP::AtFT , al mismo tiempo que la androesterilidad, inducida por la expresión del constructo XY::barnase en plantas crecidas en invernáculo de Populus tremula L. transgénicas. Este estudio representa un gran avance en la transformación del P. deltoides y P. euramericana siendo éstos los primeros álamos de estas dos especies con la característica de floración temprana. Confirmamos que el sistema HSP::AtFT en álamos combina una inducción de la floración temprana con un normal crecimiento vegetativo y que el sistema XY::barnase es efectivo para obtener alámos masculinos transgénicos estériles. Disponer de este protocolo eficiente de transformación con genes de androesterilidad permitirá en un futuro modificar genéticamente clones de álamo de interés local para introducir, junto con esta característica, caracteres de importancia silvicultural, de forma controlada. También se podrán desarrollar clones de álamos con floración temprana para incorporar a los programas de mejora genética convencional.Instituto de Biotecnología y Biología MolecularLaboratorio de Investigaciones en Mader

FLP/FRT Recombination from Yeast: Application of a Two Gene Cassette Scheme as an Inducible System in Plants

Phytosensors are plants that are genetically engineered for sensing and reporting the presence of a specific contaminant, including agriculturally important biological agents. Phytosensors are constructed by transforming plants to contain specific biotic- or abiotic-inducible promoters fused to a reporter gene. When such transgenic plants encounter the target biotic or abiotic agent, the specific inducible promoter is triggered and subsequently drives the expression of the reporter gene, which produces a signal for detection. However, several systems lack robustness, rapid induction and promoter strength. Here, we tested the FLP/FRT recombination system in a construct containing a two gene cassette organization and examined its potential in transgenic Arabidopsis and tobacco plants using a β-glucuronidase (GUS) reporter. In this model system, a heat-shock inducible promoter was employed to control the expression of the FLP recombinase gene. Upon heat induction and subsequent active FLP-mediated excision event, the GUS gene was placed in close proximity to the 35S promoter resulting in an active GUS reporter expression. Our results demonstrate that the two gene cassette scheme of inducible FLP/FRT recombination system is functional in tobacco and Arabidopsis, providing additional insights into its possible application in phytosensing such as creating strong readout capabilities