We have developed a protocol for rapid generation of barley plants with introduced genes by employing techniques of tissue culture, plant regeneration and crossing. Compared to conventional protocols, the time between pollination of To plants and identification of phenotypic expression in the T(1) generation is reduced from around 17-9 weeks. All of the selected candidates after the tissue culture screening exhibited phenotypic expression. Furthermore, by utilizing ordinary pollen crossing, we demonstrated a route for introduction of genes to a recalcitrant barley elite cultivar. This Tissue Culture after Crossing (TCC) procedure significantly enhances the prospects for producing transgenic barley lines from desirable germplasm. We suggest that the TCC method will contribute to improve barley as a promising bioenergy crop. (C) 2011 Elsevier Ltd. All rights reserved.