A protocol for Agrobacterium-mediated transformation of Brachypodium distachyon community standard line Bd21

摘要

Brachypodium distachyon is a novel model system for structural and functional genomics studies of temperate grasses because of itsbiological and genetic attributes. Recently, the genome sequence of the community standard line Bd21 has been released and theavailability of an efficient transformation system is critical for the discovery and validation of the function of Brachypodium genes.Here, we provide an improved procedure for the facile and efficient Agrobacterium-mediated transformation of line Bd21. The protocolrelies on the transformation of compact embryogenic calli derived from immature embryos using visual and chemical screening oftransformed tissues and plants. The combination of green fluorescent protein expression and hygromycin resistance enables earlyidentification of transformation events and drastically reduces the quantity of tissue to be handled throughout the selection process.Approximately eight independent fully developed transgenic Bd21 plants can be produced from each immature embryo, enabling thegeneration of thousands of T-DNA lines. The process—from wild-type seeds to transgenic T1 seeds—takes —8 months to complete.