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The genetic transformation of monocot grasses is a resource intensive process, the quality and efficiency of which is dependent in part upon the method of DNA introduction, as well as the ability to effectively separate transformed from wildtype tissue.  Agrobacterium -mediated transformation of  Brachypodium  has relied mainly on  Agrobacterium tumefaciens  strain AGL1. Currently the antibiotic hygromycin B has been the selective agent of choice for robust identification of transgenic calli in  Brachypodium distachyon  and  Brachypodium sylvaticum  but few other chemicals have been shown to work as well for selection of transgenic  Brachypodium  cells in tissue culture. This study demonstrates that  Agrobacterium rhizogenes  strain 18r12v and paromomycin selection can be successfully used for the efficient generation of transgenic  B. distachyon  and  B. sylvaticum . Additionally we observed that the transformation rates were similar to or higher than those obtained with  A .  tumefaciens  strain AGL1 and hygromycin selection. The  A .  rhizogenes  strain 18r12v harboring the pARS1 binary vector and paromomycin selection is an effective means of generating transgenic  Brachypodium  plants. This novel approach will facilitate the transgenic complementation of T-DNA knockout mutants of  B. distachyon  which were created using hygromycin selection, as well as aid the implementation of more complex genome manipulation strategies which require multiple rounds of transformation.

Use of Agrobacterium rhizogenes Strain 18r12v and Paromomycin Selection for Transformation of Brachypodium distachyon and Brachypodium sylvaticum