Agrobacterium-mediated genetic transformation of Coffea arabica (L.) is greatly enhanced by using established embryogenic callus cultures

Ribas Alessandra, Dechamp Eveline, Champion Anthony, Bertrand Benoît, Combes Marie-Christine, Verdeil Jean-Luc, Lapeyre Fabienne, Lashermes Philippe, Etienne Hervé. 2011. Agrobacterium-mediated genetic transformation of Coffea arabica (L.) is greatly enhanced by using established embryogenic callus cultures. BMC Plant Biology, 11 (92), 15 p.

Journal article ; Article de revue à facteur d'impact Revue en libre accès total
[img] Published version - Anglais
Use under authorization by the author or CIRAD.

Télécharger (2MB)

Quartile : Q1, Sujet : PLANT SCIENCES

Abstract : Background: Following genome sequencing of crop plants, one of the main challenges today is determining the function of all the predicted genes. When gene validation approaches are used for woody species, the main obstacle is the low recovery rate of transgenic plants from elite or commercial cultivars. Embryogenic calli have frequently been the target tissue for transformation, but the difficulty in producing or maintaining embryogenic tissues is one of the main problems encountered in genetic transformation of many woody plants, including Coffea arabica. Results: We identified the conditions required for successful long-term proliferation of embryogenic cultures in C. arabica and designed a highly efficient and reliable Agrobacterium tumefaciens-mediated transformation method based on these conditions. The transformation protocol with LBA1119 harboring pBin 35S GFP was established by evaluating the effect of different parameters on transformation efficiency by GFP detection. Using embryogenic callus cultures, co-cultivation with LBA1119 OD600 = 0.6 for five days at 20 °C enabled reproducible transformation. The maintenance conditions for the embryogenic callus cultures, particularly a high auxin to cytokinin ratio, the age of the culture (optimum for 7-10 months of proliferation) and the use of a yellow callus phenotype, were the most important factors for achieving highly efficient transformation (> 90%). At the histological level, successful transformation was related to the number of proembryogenic masses present. All the selected plants were proved to be transformed by PCR and Southern blot hybridization. Conclusion: Most progress in increasing transformation efficiency in coffee has been achieved by optimizing the production conditions of embryogenic cultures used as target tissues for transformation. This is the first time that a strong positive effect of the age of the culture on transformation efficiency was demonstrated. Our results make Agrobacterium-mediated transformation of embryogenic cultures a viable and useful tool both for coffee breeding and for the functional analysis of agronomically important genes. (Résumé d'auteur)

Mots-clés Agrovoc : Coffea arabica, Agrobacterium tumefaciens

Classification Agris : F30 - Plant genetics and breeding
F02 - Plant propagation

Champ stratégique Cirad : Axe 1 (2005-2013) - Intensification écologique

Auteurs et affiliations

  • Ribas Alessandra
  • Dechamp Eveline, CIRAD-BIOS-UMR RPB (FRA)
  • Champion Anthony, IRD (FRA)
  • Bertrand Benoît, CIRAD-BIOS-UMR RPB (FRA)
  • Combes Marie-Christine, IRD (FRA)
  • Verdeil Jean-Luc, CIRAD-BIOS-UMR AGAP (FRA)
  • Lapeyre Fabienne, CIRAD-BIOS-UMR AGAP (FRA)
  • Lashermes Philippe, IRD (FRA)
  • Etienne Hervé, CIRAD-BIOS-UMR RPB (FRA)

Autres liens de la publication

Source : Cirad - Agritrop (

View Item (staff only) View Item (staff only)

[ Page générée et mise en cache le 2020-09-25 ]