Jesus Escudero - Academia.edu (original) (raw)

Papers by Jesus Escudero

Research paper thumbnail of T-DNA transfer in meristematic cells of maize provided with intracellular Agrobacterium

Plant Journal, 1996

Agrobacterium has been established as a tool for gene delivery to most dicotyledonous plant speci... more Agrobacterium has been established as a tool for gene delivery to most dicotyledonous plant species. However, it is not generally efficient in monocotyledonous plant species, especially not in Graminae. In maize, Agrobacterium-mediated DNA transfer has been detected but early developmental stages in the plant proved incompetent as recipients. This research tests whether the lack of competence in young immature embryos of maize could be overcome by providing Agrobacterium in the interior of the plant cell. A microinjection technique was used to target single meristematic cells and prove competence to Agrobacterium. This response is dependent on the maize plant genotype.

Research paper thumbnail of Transfer and Integration of T-DNA without Cell Injury in the Host Plant

Plant Cell, 1997

Agrobacterium colonizes plant cells via a gene transfer mechanism that results in plant tumorigen... more Agrobacterium colonizes plant cells via a gene transfer mechanism that results in plant tumorigenesis. Virulence (vi4 genes are transcriptionally activated in the bacteria by plant metabolites released from the wound site. Hence, it is believed that agrobacteria use injuries to facilitate their entrance into the host plant and that the wounded state is required for plant cell competence for Agrobacterium-mediated gene delivery. However, our experiments using vir gene-activated bacteria sprayed onto tobacco plantlets demonstrated that cells in unwounded plants could also be efficiently transformed. The condition of the plant cells was monitored using P-glucuronidase under the control of a wound-inducible promoter. lnfection of leaf tissue is light dependent, and it is drastically reduced when abscisic acid is exogenously applied to the plant. Under these experimental conditions, stomatal opening seems to be used by Agrobacterium to circumvent the physical barrier of the cuticle. These results thus show that the proposed cellular responses evoked by wounding in higher plants are not essential for Agrobacterium-mediated transformation.

Research paper thumbnail of Efficient and sensitive assay for T-DNA-dependent transient gene expression

Plant Molecular Biology Reporter, 1993

We describe here a very sensitive and reproducible method to detect the efficiency ofAgrobacteriu... more We describe here a very sensitive and reproducible method to detect the efficiency ofAgrobacterium-mediated T-DNA transfer. This method is based on a quantitative assay of β-glucuronidase activity produced in the plant cell upon transfer of T-DNA carrying a specialuidA gene construct. Analysis of the transfer efficiency of a transfer-proficient bacterium compared with that of the same bacterium diluted at different ratios with a transfer-defective bacterium shows a high sensitivity of the β-glucuronidase activity in the plant. Five orders of magnitude in T-DNA transfer efficiency can be covered when the activity is measured combining the fluorimetric MUG assay (for high activity) and the histochemical X-Gluc assay (very sensitive for low activity).

Research paper thumbnail of T-DNA Transfer to Maize Cells: Histochemical Investigation of beta- Glucuronidase Activity in Maize Tissues

Proceedings of The National Academy of Sciences, 1993

Agrobacterium tumefaciens is routinely used to engineer desirable genes into dicotyledonous plant... more Agrobacterium tumefaciens is routinely used to engineer desirable genes into dicotyledonous plants. However, the economically important graminaceous plant maize is refractory to tumor induction by inoculation with virulent strains of A. tumefaciens. Currently, the only clearcut evidence for transferred DNA (T-DNA) transport from Agrobacterium to maize comes from agroinfection. To study T-DNA transfer from Agrobacterium to maize cells in a virus-free system, we used here the β-glucuronidase (GUS; EC 3.2.1.31) gene as a marker. GUS expression was observed with high efficiency on shoots of young maize seedlings after cocultivation with Agrobacterium carrying the GUS gene. Agrobacterium virulence mutants, incapable of transferring T-DNA to dicot tissue, were shown to be deficient in eliciting GUS expression in maize. Hence, expression of the T-DNA-located GUS gene in maize cells is strictly dependent on Agrobacterium-mediated DNA transfer. Histochemical staining of maize shoots revealed GUS expression located mainly in the leaves and the coleoptile.

Research paper thumbnail of T-DNA transfer to maize plants

Molecular Biotechnology, 1999

Agrobacterium-mediated transformation is the method of choice to engineer desirable genes into pl... more Agrobacterium-mediated transformation is the method of choice to engineer desirable genes into plants. Here we describe a protocol for demonstrating T-DNA transfer from Agrobacterium into the economically important graminaceous plant maize. Expression of the T-DNA-located GUS gene was observed with high efficiency on shoots of young maize seedlings after cocultivation with Agrobacterium.

Research paper thumbnail of Intracellular Agrobacterium can Transfer DNA to the Cell Nucleus of the Host Plant

Proceedings of The National Academy of Sciences, 1995

Agrobacterium tumefaciens is a Gramnegative, soil-borne bacterium responsible for the crown gall ... more Agrobacterium tumefaciens is a Gramnegative, soil-borne bacterium responsible for the crown gall disease ofplants. The galls result from genetic transformation of plant cells by the bacteria. Genes located on the transferred DNA (T-DNA), which is part of the large tumor-inducing (Ti) plasmid of Agrobacterium, are integrated into host plant chromosomes and expressed. This transfer requires virulence

Research paper thumbnail of Matrimonio cor nico espa a marruecos 2011 2012

Lo primero que hay que hacer es solicitar el CERTIFICADO DE CAPACIDAD MATRIMONIAL (un papel que p... more Lo primero que hay que hacer es solicitar el CERTIFICADO DE CAPACIDAD MATRIMONIAL (un papel que permite que se realice el matrimonio), el cual se pide en el Registro Civil. Para ello: A) PEDIR CITA EN EL REGISTRO CIVIL (C/Pradillo 66) Tardan unos 20 días en dártela. Con los siguientes documentos: De la española Del marroquí fotocopia del DNI, Instancia y Declaración Jurada firmada por los 2 (te la dan en la 5º planta del R.C. cuando pides información), Certificado literal de nacimiento (en la segunda planta del RC, en el momento) fotocopia del pasaporte, Instancia y Declaración Jurada firmada por los 2, Certificado literal de nacimiento (sellado en Wilaya, Ministerio de Asuntos Exteriores, Consulado General De España) B) DÍA DE LA CITA EN EL REGISTRO CIVIL: Se acude con un testigo y con los siguientes documentos: De la española Del marroquí DNI (original y fotocopia), Instancia y Declaración Jurada firmada por los 2, Certificado literal de Nacimiento (la solicitas en la segunda planta del Registro Civil de Madrid, tiene validez de un año). Padrón Municipal de los 2 últimos años (se pide en cualquier oficina de atención al ciudadano, en el momento), "fe de vida y estado" (se pide en la planta baja del registro civil, en el momento)

Research paper thumbnail of T-DNA transfer in meristematic cells of maize provided with intracellular Agrobacterium

Plant Journal, 1996

Agrobacterium has been established as a tool for gene delivery to most dicotyledonous plant speci... more Agrobacterium has been established as a tool for gene delivery to most dicotyledonous plant species. However, it is not generally efficient in monocotyledonous plant species, especially not in Graminae. In maize, Agrobacterium-mediated DNA transfer has been detected but early developmental stages in the plant proved incompetent as recipients. This research tests whether the lack of competence in young immature embryos of maize could be overcome by providing Agrobacterium in the interior of the plant cell. A microinjection technique was used to target single meristematic cells and prove competence to Agrobacterium. This response is dependent on the maize plant genotype.

Research paper thumbnail of Transfer and Integration of T-DNA without Cell Injury in the Host Plant

Plant Cell, 1997

Agrobacterium colonizes plant cells via a gene transfer mechanism that results in plant tumorigen... more Agrobacterium colonizes plant cells via a gene transfer mechanism that results in plant tumorigenesis. Virulence (vi4 genes are transcriptionally activated in the bacteria by plant metabolites released from the wound site. Hence, it is believed that agrobacteria use injuries to facilitate their entrance into the host plant and that the wounded state is required for plant cell competence for Agrobacterium-mediated gene delivery. However, our experiments using vir gene-activated bacteria sprayed onto tobacco plantlets demonstrated that cells in unwounded plants could also be efficiently transformed. The condition of the plant cells was monitored using P-glucuronidase under the control of a wound-inducible promoter. lnfection of leaf tissue is light dependent, and it is drastically reduced when abscisic acid is exogenously applied to the plant. Under these experimental conditions, stomatal opening seems to be used by Agrobacterium to circumvent the physical barrier of the cuticle. These results thus show that the proposed cellular responses evoked by wounding in higher plants are not essential for Agrobacterium-mediated transformation.

Research paper thumbnail of Efficient and sensitive assay for T-DNA-dependent transient gene expression

Plant Molecular Biology Reporter, 1993

We describe here a very sensitive and reproducible method to detect the efficiency ofAgrobacteriu... more We describe here a very sensitive and reproducible method to detect the efficiency ofAgrobacterium-mediated T-DNA transfer. This method is based on a quantitative assay of β-glucuronidase activity produced in the plant cell upon transfer of T-DNA carrying a specialuidA gene construct. Analysis of the transfer efficiency of a transfer-proficient bacterium compared with that of the same bacterium diluted at different ratios with a transfer-defective bacterium shows a high sensitivity of the β-glucuronidase activity in the plant. Five orders of magnitude in T-DNA transfer efficiency can be covered when the activity is measured combining the fluorimetric MUG assay (for high activity) and the histochemical X-Gluc assay (very sensitive for low activity).

Research paper thumbnail of T-DNA Transfer to Maize Cells: Histochemical Investigation of beta- Glucuronidase Activity in Maize Tissues

Proceedings of The National Academy of Sciences, 1993

Agrobacterium tumefaciens is routinely used to engineer desirable genes into dicotyledonous plant... more Agrobacterium tumefaciens is routinely used to engineer desirable genes into dicotyledonous plants. However, the economically important graminaceous plant maize is refractory to tumor induction by inoculation with virulent strains of A. tumefaciens. Currently, the only clearcut evidence for transferred DNA (T-DNA) transport from Agrobacterium to maize comes from agroinfection. To study T-DNA transfer from Agrobacterium to maize cells in a virus-free system, we used here the β-glucuronidase (GUS; EC 3.2.1.31) gene as a marker. GUS expression was observed with high efficiency on shoots of young maize seedlings after cocultivation with Agrobacterium carrying the GUS gene. Agrobacterium virulence mutants, incapable of transferring T-DNA to dicot tissue, were shown to be deficient in eliciting GUS expression in maize. Hence, expression of the T-DNA-located GUS gene in maize cells is strictly dependent on Agrobacterium-mediated DNA transfer. Histochemical staining of maize shoots revealed GUS expression located mainly in the leaves and the coleoptile.

Research paper thumbnail of T-DNA transfer to maize plants

Molecular Biotechnology, 1999

Agrobacterium-mediated transformation is the method of choice to engineer desirable genes into pl... more Agrobacterium-mediated transformation is the method of choice to engineer desirable genes into plants. Here we describe a protocol for demonstrating T-DNA transfer from Agrobacterium into the economically important graminaceous plant maize. Expression of the T-DNA-located GUS gene was observed with high efficiency on shoots of young maize seedlings after cocultivation with Agrobacterium.

Research paper thumbnail of Intracellular Agrobacterium can Transfer DNA to the Cell Nucleus of the Host Plant

Proceedings of The National Academy of Sciences, 1995

Agrobacterium tumefaciens is a Gramnegative, soil-borne bacterium responsible for the crown gall ... more Agrobacterium tumefaciens is a Gramnegative, soil-borne bacterium responsible for the crown gall disease ofplants. The galls result from genetic transformation of plant cells by the bacteria. Genes located on the transferred DNA (T-DNA), which is part of the large tumor-inducing (Ti) plasmid of Agrobacterium, are integrated into host plant chromosomes and expressed. This transfer requires virulence

Research paper thumbnail of Matrimonio cor nico espa a marruecos 2011 2012

Lo primero que hay que hacer es solicitar el CERTIFICADO DE CAPACIDAD MATRIMONIAL (un papel que p... more Lo primero que hay que hacer es solicitar el CERTIFICADO DE CAPACIDAD MATRIMONIAL (un papel que permite que se realice el matrimonio), el cual se pide en el Registro Civil. Para ello: A) PEDIR CITA EN EL REGISTRO CIVIL (C/Pradillo 66) Tardan unos 20 días en dártela. Con los siguientes documentos: De la española Del marroquí fotocopia del DNI, Instancia y Declaración Jurada firmada por los 2 (te la dan en la 5º planta del R.C. cuando pides información), Certificado literal de nacimiento (en la segunda planta del RC, en el momento) fotocopia del pasaporte, Instancia y Declaración Jurada firmada por los 2, Certificado literal de nacimiento (sellado en Wilaya, Ministerio de Asuntos Exteriores, Consulado General De España) B) DÍA DE LA CITA EN EL REGISTRO CIVIL: Se acude con un testigo y con los siguientes documentos: De la española Del marroquí DNI (original y fotocopia), Instancia y Declaración Jurada firmada por los 2, Certificado literal de Nacimiento (la solicitas en la segunda planta del Registro Civil de Madrid, tiene validez de un año). Padrón Municipal de los 2 últimos años (se pide en cualquier oficina de atención al ciudadano, en el momento), "fe de vida y estado" (se pide en la planta baja del registro civil, en el momento)