Abstract Background Date palm (Phoenix dactylifera L.) is a traditional crop in arid and semi-arid areas. Its vegetative propagation can be achieved by offshoots, but possible number of offshoots in mother palm trees is limited. Micropropagation is a highly recommended strategy for obtaining date palm elite cultivars using shoot tip and immature inflorescences. In this study, micropropagation procedure using inflorescence explants of Medjool cv. is described. For culture initiation, explants from different spathe lengths were cultivated on Murashige and Skoog medium (MS) supplemented with picloram at 1.0 and 2.0 mg/l combined with 2iP at 0.5 mg/l alone and with both 2iP and BA at 0.25 mg/l for 24 weeks. The obtained direct globular embryos were transferred to maturation media with 0.1 mg/l picloram alone or combined with both 2iP and ABA separately and together for further development. Additionally, multiplication and rooting media were optimized by different cytokinins and auxins for high frequency of plantlet production. Acclimatization of in vitro plantlets was also investigated. Results The highest percentage of globular embryo formation was noticed with explants isolated from spathe lengths ranging from 10 to 15 cm. Addition of BA to initiation media with picloram encouraged a significant effect on embryonic culture formation percentage. Incorporation of ABA and 2iP to maturation medium was an effective factor for individual or multiple embryo emergence. Acclimatization of in vitro plantlets having 3–4 roots was successfully accomplished. Irrigation with the full strength solution (MS) encouraged the highest growth vigor degree, leaf number/plant, leaf width, root number, and root thickness degree of ex vitro plants. Conclusion This research provides an advanced regeneration system for large-scale production of date palm from immature inflorescences of Medjool cv. It opens up the prospects of using picloram with different growth regulators for rapid micropropagation of date palm.
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