Objectives Papers Grants Patents
Dr. Luis Cistué*, Dr. Ana M. Castillo, Dr. M. Pilar Vallés
Begoña Echávarri, Miguel Sanz, M. Jesús Espiau
* Research Group Leader
General objective
Our main objective is the optimization of doubled haploid lines (DH) production via gametic embryogenesis (anther and/or isolated microspore culture) of barley, rice, and both durum and bread wheat. We are focused on the improvement of different physiological factors and on genetic studies of mechanisms involved in the process. Furthermore, we are also using barley gametic embryogenesis in combination with procedures that generate new variability, such as mutagenesis and genetic transformation.
Specific objectives
1. Barley, rice, bread and durum wheat doubled haploid (DH) production.
L. Cistué, M.P. Vallés, A.M. Castillo
Doubled haploid production is undertaken by anther and/or isolated microspore in vitro culture, from F1s of high agronomic value, selected by private companies or public research institutions. The DH lines produced are introduced into plant breeding programs of cereals for the selection of new varieties, or are used for genetic analysis of agronomic characters. Particularly, a cytogenetic study and agronomic effects of the reciprocal translocation present in the six-rowed barley cultivar Albacete (the most widely grown variety in semiarid conditions) is being studied. Production of recombinant chromosome substitution lines (RCSL) of ‘Albacete’ in the widely grown variety ‘Plaisant’ is also proposed.
2. Optimization of barley, bread and durum wheat gametic embryogenesis.
A.M. Castillo, L. Cistué
The main factors hindering the production of DH lines are: the low response of anthers to the stress pretreatment and embryogenic capacity, the high percentage of albinism and the high mortality rate of haploid plants after application of a chromosome duplication treatment. Optimization of the in vitro microspores and anther culture for double haploid production will be sought, by generating severe osmotic, chemical and temperature stress as pre-treatments, and by synchronization of microspores cell cycle. Furthermore, we are studying the effect of colchicine on the embryogenesis and chromosome duplication capacities, when applied in different steps of the gametic embryogenesis process. We are developing efficient protocols for different cultivars of durum wheat, species with a very low rate of induction and a very high rate of albinism, as well as for barley and bread wheat cultivars of great agronomic value which, unfortunately, are normally recalcitrant to gametic embryogenesis.
3. Genetic analysis of barley gametic embryogenesis. Identification of QTLs and candidate genes associated to the process.
M.P. Vallés, A.M. Castillo
The genotype is one of the main factors limiting (conditioning) DH production. Knowledge of the genetic control of the process will facilitate the design of new strategies for its optimization. Identification and location of QTLs associated to embryogenesis and albinism is being undertaken. Validation of these QTLs in different cultivars of agronomic value will allow the prediction of the gametic embryogenesis response of genotypes prior to its utilization, and therefore leading to an optimization of economic resources for DH production. Furthermore, a genomic approach is being carried out for the identification of candidate genes associated to the microspores induction process, using barley microarray chips.
4. Generation of new variability by genetic transformation and the combination of mutagenesis and in vitro selection.
L. Cistué, M.P. Vallés, A.M. Castillo
Barley is still one the most recalcitrant cereals for genetic transformation. We have developed a protocol for transformation of different cultivars of barley. The transgenic lines, containing genes involved in metabolic pathway of fructans biosynthesis, will be taken up for advanced generations (T2, T3). Molecular characterization and gene expression studies of these plants will be performed. The mutation of gametic cells in combination with in vitro selection allows the rapid identification of DH mutants, and therefore with the character fixed. We have developed a mutagenesis system by the application of sodium azide to barley microspores. Validation of this mutagenic system, together with further in vitro selection of the microspores for boron tolerance will be undertaken.
Representative research papers
SCI papers:
Castillo AM, Cistué L, Sanz JM, Romagosa I, Vallés MP (2001) Low responsiveness of six-row genotypes to androgenesis in barley does not have a pleiotropic nature. Genome 44: 936-940.
Castillo AM, Cistué L, Vallés MP, Sanz JM, Romagosa I, Molina-Cano JL (2001) Efficient production of androgenic doubled-haploid mutants in barley by the application of sodium azide to anther and microspore cultures. Plant Cell Reports 20: 105-111.
Castillo AM, Vallés MP, Cistué L (2000) Comparison of anther and isolated microspore cultures in barley. Effects of culture density and regeneration medium. Euphytica 113: 1-8.
Prada D, Ullrich SE, Molina-Cano JL, Cistué L, Clancy JA, Romagosa I (2004) Genetic control of dormancy in a Triumpn/Morex cross in barley. Theoretical Applied Genetics 109: 62-70
Pulido A, Castillo AM, Vallés MP, Olmedilla A (2001) Early stages of pollen embryogenesis in barley anther cultures induced by pretreatment with mannitol. International Journal of Developmental Biology 45: 55-56.
Pulido A, Bakos F, Castillo AM, Vallés MP, Barnabas B, Olmedilla A (2005) Comparison of cytological and ultrastructural changes induced in anther and isolated-microspore cultures in barley: presence of Fe deposits in isolated-microspore cultures. Journal of Structural Biology 149/2:170-181
Ramirez C, Testillano PS, Castillo AM, Vallés MP, Coronado MJ, Cistué L, Risueño MC (2001) The early microspore embryogenesis pathway in barley is accompanied by concrete ultrastructural and expression changes. International Journal of Developmental Biology 45: 57-58.
Cistué L, Echávarri B, Batlle F, Soriano M, Castillo A, Vallés MP, Romagosa I (2005) Segregation distortion for agronomic traits in barley doubledhaploids lines. Plant Breeding 124: 546-550.
Books chapters:
Castillo AM, Vallés MP, Cistué L (2001) Improvements of barley androgenesis for plant breeding. En: Biotechnological Approaches for Utilization of Gametic Cells, (Bohanec B, ed.) pp 15-21. European COST Action 824: Gametic Embryogenesis, Brussels. ISBN: 92-894-0225-3
Cistué L, Vallés MP, Echávarri B, Sanz JM, Castillo AM (2004). Barley anther culture. Doubled Haploid Production in Crop Plants. A Manual: 29-35. Maluszynski M, Kasha K, Foster B (Eds.) FAO/IAEA División (Viena, Austria). ISBN 1-4020-1544-5.
Cistué L, Vallés MP, Echávarri B, Sanz JM, Castillo AM (2004) Production of barley doubled haploid by anther and microspore culture. In vitro applications in crop improvements- recent progress. In: A. Mujib et al (Eds.) Oxford & IBH Publishing Co. Pvt. Ltd, New Delhi, pp:1-18. ISBN: 1-57808-300-1.
Current research grants and contracts (active)
Fructanos de cebada para agroalimentación: mejora biotecnológica y caracterización molecular y funcional (Responsible: R. Morcuende - IRNASA). Junta deCastilla y León, CSI01A05. Responsible (EEAD): Vallés.
Estudio molecular, citológico y fenotípico de la translocación recíproca de la variedad de cebada Albacete. Obtención de líneas de sustitución (RCSL). Optimización de sistemas de androgénesis. MEC, PN, AGL2002-04139-C02-02. Responsible: Cistué.
Producción de lineas doblehaploides de cebada. Localización de QTLs asociados a los factores limitantes de la androgénesis y validación del efecto de los QTLs en materiales de interés para mejora. MEC, PN, AGL2004-03396. Responsible: Vallés.
Desarrollo de materiales y técnicas para el análisis integrado de la respuesta de la cebada (Hordeum vulgare L.) al estrés abiótico. MEC, PN,AGL2005-07195-C02-01. Responsible: Cistué.
Mapping adaptation of barley to drought environmentes (MABDE). EU, LIFE/001/1412. Responsible: Cistué.
Producción de doble haploides de trigo duro y trigo blando a partir de cruzamientos F1 entre variedades españolas. PETRI, PTR1995-0732-OP. Responsible: Cistué.
Emergent Research Group, acknowledged by the Aragón Regional Government. Aplicación de cultivos celulares y desarrollo de técnicas de biotecnología para mejora vegetal (A45). DGA. Responsible: Cistué.
Contract for "Producción de doble-haploides de trigo duro y trigo blando a partir de cruzamientos F1 entre cultivares españoles" (vinculado a PTR95-0732-OP). Semillas Batlle S.A. (Barcelona). Responsible: Cistué.
INICIO
ATRÁS
ADELANTE
IMPRIMIR