Effect of low 2,4-D concentration on enhancing indirect embryogenesis and genetic stability in date palm (Phoenix dactylifera L.)

This investigation systematically evaluated the effect of low (10 mg/L) and high (50 mg/L) concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D), combined with 3 mg/L 2iP, on callus induction from shoot tip explants of Barhi date palm cv. The induced calli were used to establish an indirect somatic embryogenesis (ISE) system on MS medium supplemented with different plant growth regulator (PGR) formulations. Genetic stability was assessed using inter-simple sequence repeat (ISSR) markers. Callus induced with low 2,4-D (C1) concentration exhibited significantly (p < 0.05) superior morphogenic competence, yielding a higher fresh weight (FW) of callus at both the induction (0.290 g/jar) and regeneration (0.475 g/jar) stages, more somatic embryos (100% frequency, 8/explant), and enhanced regeneration capacity compared to callus induced by high 2,4-D (C2). Regenerants from C1 displayed superior bud germination (6.2/explant), shoot proliferation (7/explant) and elongation (6.7 cm), as well as root induction (5.6/shoot) and elongation (6.3 cm). Plantlets were successfully acclimatized, achieving an 88.5% survival rate. ISSR analysis confirmed 97.5% of genetic fidelity among all regenerants. These results indicate that a lower 2,4-D concentration (10 mg/L) enhances callus vigor and subsequent organogenesis, facilitating a highly efficient and genetically stable ISE protocol for date palm micropropagation.