Abstract: Taking different transgenic barley as test materials for pot experiment, the lignin content, key enzyme activity, stem structure and Fourier transform infrared spectroscopy of different barley stems were studied to reveal the unintended changes related to lignin metabolism in transgenic barley. The results showed that the lignin content, parenchymal tissue, numbers of vascular bundles, guaiacyl lignin (G), syringyl lignin (S) and A(S)/A(G) of transgenic barley (T1, T2) stems had no significant changes in comparison with that of the wild type (WT) and non-transgenic barley (NT1). The unintended changes of the transgenic barley T1, such as the key enzyme activities of lignin metabolism (for PAL decreased by 66% and for POD increased significantly by 76%), the sclerenchymal tissue and the ρ-hydroxyphenyl lignin (H) ratio (decreased by 59%), may be mainly caused by somaclonal variation generated from tissue culture during genetic transformation.