Hua, W., Zhu, J., Shang, Y., Wang, J., Jia, Q., Lin, F. and Yang, J. 2013. Establishment of a highly efficient regeneration method from the scraped broken embryo of mature barley seed. Can. J. Plant Sci. 93: 1029-1035. An efficient and reproducible plant regeneration system for mature barley would represent a significant innovation for barley tissue culture and genetic transformations to optimize barley crop traits. To address this need, we used scraped broken embryos (SBEs) from dried barley seeds to establish a highly efficient regeneration system. Mature SBEs cultured in an induction medium with 2 mg L^-1 2,4-D offered the best callus formation (92.99% induction efficiency) and no germination. Furthermore, calli arising from SBEs had the highest regeneration frequency (69.72%) in FHG medium with 1 mg L^-1 6-benzylaminopurine. We then compared three different cultivars: Golden Promise, Zaoshu3, and Zhepi8 for callogenesis and regeneration using an established protocol. Zhepi8 had the greatest callus induction (92.78%) and regeneration (68.24%). Our innovative methods for mature barley plant reproduction will contribute to improvements in genetic barley transformation as an alternate regeneration system.