The use of in vitro techniques for conserving plant biodiversity and protecting rare and endangered multipurpose plant species is considered as one of the most important ex-situ conservation policies. Development of an efficient in vitro regeneration protocol of Calligonum comosum is important and that has achieved to protect the endangered multipurpose medicinally important desert plant in the Kingdom of Bahrain. Nodal segments were used as explants source and the effect of various plant growth regulators (PGRs) were studied for responses and to regenerate the whole plants in modified Murashige and Skoog (MS) media through direct and indirect organogenesis via callus induction. 50% explants of C. comosum responded to initiate shoot in presence of 4.44 µM BAP with 2.68 µM NAA after four weeks of culture while 40% shoot initiation response was the highest value in presence of 9.29 µM KI with 5.37 µM NAA after 4 weeks of culture among the treatments of KI with NAA. The highest callus induction rate of 100% was found in media containing 9.29 µM KI and 5.37 µM NAA after four weeks. Multiple initial shoots those originated from nodal segments develop calli and showed organogenic differentiation of shoots in presence of BAP and IAA. The highest shoot multiplication frequency of 15 was observed while the shoots initiated in media contained 4.44 µM BAP with 2.85 µM IAA and were transferred to 8.56 µM IAA with 2.22 µM BAP. Shoot multiplication and shoot regeneration capacity was compared in different media and the highest performance of 234 shoots /explants after second multiplication was observed while shoots initiated in presence of 13.3 µM BAP and 5.71 µM IAA. As a precautionary approach to conserve the endangered medicinal plant species in the Kingdom of Bahrain, the application of in vitro culture is considered as an important alternative method in ex situ conservation strategy in the present study.