Recurrent somatic embryogenesis and development of somatic embryos in Akebia trifoliata (Thunb.) Koidz (Lardizabalaceae)

Abstract

A simple and effective protocol was established for recurrent somatic embryogenesis and plant regeneration in Akebia trifoliata (Thunb.) Koidz. Somatic embryos directly formed from the root zone of the immature zygotic embryos cultured on MS medium devoid of plant growth regulator (PGR). The induction frequency of immature zygotic embryos was 52.5%, and the mean number of somatic embryos reached up to 19.5. Secondary somatic embryos arose on the primary embryos at a high frequency (95.8%), and this process maintained in a recurrent way on PGR-free MS medium. The highest mean number of somatic embryos also appeared in the primary embryos which can reach up to 44.7. While both of the embryogenic potential and mean number of embryos per explant displayed a gradual diminution with subculturing. The addition of 0.5 mg l−1 6-benzyladenine in the 1/2 MS regeneration medium can significantly improve the frequency of somatic embryos convert to plantlets. The microscopic analysis revealed that the development process of somatic embryos via the globular, heart, torpedo and cotyledonal stages in A. trifoliata, the histological analysis showed that somatic embryos directly initiated from root epidermal cells and there are no vascular connections between the somatic embryos and maternal tissue. Regenerated plantlets acclimated successfully to greenhouse conditions. Approximately 65% plantlets survived and displayed no morphological characteristics differences with seed-derived plants. The simple and effective protocols established in this study will promote large-scale clonal propagation and genetic improvement of A. trifoliata. Somatic embryogenesis has never been reported in Akebia trifoliata (Thunb.) Koidz. The study showed the origin and development process of somatic embryos in Akebia trifoliata (Thunb.) Koidz.