Respuesta morfogenética en Agave angustifolia al gradiente de auxina-citocinina durante el desarrollo de embriones somáticos indirectos

Abstract

Agave angustifolia Haw. is a highly valuable species for the Mexican mezcal agroindustry. However, the increasing demand contrasts with the scarcity of high-quality germplasm and the low efficiency of conventional propagation methods. In this context, somatic embryogenesis emerges as a promising biotechnological strategy for the mass regeneration of elite plants under controlled conditions. The objective of this study was to evaluate the effect of three auxins (2,4-dichlorophenoxyacetic acid [2,4-D], indole-3-acetic acid [IAA], and naphthaleneacetic acid [NAA]) combined with N6-benzyladenine (BA) on the induction of somatic embryogenesis in A. angustifolia, using zygotic embryo axes as explants. A completely randomized experimental design was applied to assess 27 hormonal combinations, derived from the factorial interaction of three concentrations of auxins (13.57, 18.10 and 22.62 µM of 2,4-D; 17.13, 22.83 and 28.54 µM of IAA; 16.11, 21.48 and 26.85 µM of NAA) and three of BA (4.44, 8.88 and 13.32 µM). The induction medium for proembryogenic masses consisted of 25 % MS supplemented with 60 g L⁻¹ sucrose and L2 vitamins. Each zygotic embryo axis was considered an experimental unit. After 60 days of start of cultivation, the resulting calli were transferred to a histodifferentiation MS medium containing 0.45 µM of 2,4-D, 0.57 µM of IAA, or 0.54 µM of NAA, as appropriate, and without BA. All cultures were kept in darkness for an additional 30– 60 days to promote embryonic differentiation. Combinations of 2,4-D + BA induced callus formation in 71% of the explants, with a predominance of embryogenic structures. The most effective combination was 22.62 µM of 2,4-D + 13.32 µM of BA, yielding between 1 and 36 somatic embryos per explant and an embryogenic efficiency of 17.6 ± 7.1. Combinations of NAA + BA mainly promoted rhizogenesis (87%) with no embryo formation, while IAA + BA combinations were ineffective (≤ 10%), resulting in oxidized tissues with no morphogenic development. Plantlets regenerated from somatic embryos germinated properly and exhibited a 100 % survival rate when acclimated under ex vitro conditions. The combination of 22.62 µM of 2,4-D and 13.32 µM of BA proved to be the most efficient for inducing somatic embryogenesis in A. angustifolia. Among the tested auxins, 2,4-D showed a superior effect compared to NAA and IAA, establishing itself as the key regulator for inducing cellular totipotency. The high ex vitro acclimatization rate confirms the practical viability of the proposed protocol, with broad potential applications in conservation, genetic improvement, and sustainable agave production.