Enhancing frequency of regeneration of somatic embryos of avocado (Persea americana Mill.) using semi-permeable cellulose acetate membranes

Regeneration of avocado via somatic embryogenesis is difficult due to poor embryo maturation, resulting in low frequencies of germination. In this study, the influence of semi-permeable cellulose acetate membranes and culture media, containing high levels of sucrose along with coconut water, on maturation and germination of somatic embryos of avocado have been evaluated. The culture of embryogenic calli on top of cellulose acetate membranes significantly increased the number of mature, white-opaque embryos that were recovered after 5 weeks of culture. These embryos showed a much more normal appearance and better quality compared with the control embryos, although the embryo size was significantly reduced. To increase the embryo size and to complete maturation, several two-step maturation treatments were tested. The culture of white-opaque somatic embryos in a modified MS medium with B5 macronutrients gelled with 10 g L^?1 agar (B5m10A medium) over a 5-week period, followed by 5 additional weeks in B5m10A with 45 g L^?1 sucrose and 20 % coconut water, yielded the best results, reducing the percentage of necrotic embryos and the number of calli formed. The beneficial effects of this maturation treatment were enhanced when using embryos that were pre-matured on cellulose acetate membranes. Following this two-step maturation treatment, the germination rate of the control somatic embryos, which were not cultured on cellulose membranes, was lower than 10 %, but it significantly improved when the embryos had been pre-matured on cellulose acetate membranes for 5 weeks, reaching a germination rate close to 40 %. The water availability was significantly reduced when somatic embryos were cultured on cellulose membranes, and after this pre-maturation treatment, the white-opaque embryos showed lower water potential and ABA content compared with the control embryos. These results suggest that culturing over cellulose membranes causes a controlled embryo desiccation that enhances the recovery of plants.