Plant & Animal Genomes XVI Conference Plant & Animal Genomes XVI ConferenceJanuary 12-16, 2008
Town & Country Convention Center
San Diego, CA
Brachypodium distachyon is a promising model system for temperate grasses structural and functional genomics because of its physical, genetic and genome attributes. A 4x shotgun sequence of the community standard line Bd21 has been produced in August 2007. Here we describe two very efficient Agrobacterium-mediated transformation systems for the inbred lines Bd21-3 (USDA) and Bd21 (John Innes Centre). Both systems rely on Agrobacterium-mediated transformation of embryogenic calli derived from immature embryos. Key parameters included the use of the green fluorescent protein (GFP) for visual screening of transgenic tissues and plants, the utilisation of different desiccation treatments during co-culture as well as supplementing culture media prior and during the regeneration of transgenic plants with CuSO4. Transformation efficiencies of 40-50% can be obtained for both Bd21-3 and Bd21 genotypes enabling the production of large population of T-DNA insertion lines. An adaptor-anchor strategy has been developed to retrieve Flanking Sequence Tags (FSTs) of T-DNA inserts from Bd21, and sequences are available in online databases. A parallel strategy is being developed for Bd21-3 in a project funded by the US Department of Energy. The development of a collaborative network of laboratories focussed on the production of much larger populations of T-DNA mutants using the methods presented here will be discussed.