Drought is one of the major stress factors limiting crop production worldwide. Polyamines i.e. spermine play a pivotal role in abiotic stress tolerance of plants in general and drought tolerance in particular. As key regulators of transpiration, stomatal guard cells were measured to determine the involvement of spermine in drought responses. In Arabidopsis, the application of spermine, was found to induce stomatal closure. Spermine triggered changes in guard cell [Ca2+]cyt levels by Ca2+ influx as well as release of Ca2+ from internal stores. Intracellular pH play a crucial role in the spermine-triggered generation of Ca2+ signals which eventually ends up with the closure of stomata. To investigate the role of spermine in barley, transgenic barley lines, were generated. Drought stress experiments with transgenic lines showed significantly increased relative water content under extreme stress, going along with increased chlorophyll concentration and PSII efficiency. Furthermore, transgenic plants indicated better yield potential under environmental stress. Taken together, this study demonstrated in improving drought tolerance in barley.