Agrobacterium tumefaciens-mediated barley transformation

Genetically transformed barley was produced by co-cultivating immature embryo explants with Agrobacterium tumefaciens carrying a binary vector coding for chimaeric bacterial genes, bar and gus, and selecting for bialaphos-resistant cultures from which plants were regenerated. Integration of both genes was confirmed by gel blot hybridization analysis of DNA from the transformed plants and their progenies. From 1282 embryos, plants were recovered for 54 independently transformed lines, giving a transformation efficiency of 4.2%. Transgene numbers in the different lines ranged from single copy insertion to at least ten copies. Sixteen out of 18 plants grown to maturity were fully fertile. Both marker genes, bar and gus, were expressed and co-segregated in the T-1 progeny plants. In the majority of cases, the genes showed Mendelian segregation predicted for transgene insertion at a single locus. In one family with multiple transgene insertions, molecular analysis of T-1 and T-2 plants suggested that the T-DNA had inserted at two unlinked loci.