Maturation of somatic embryos of Persian walnut (Juglans regia L.) was evaluated on media containing various concentrations of Gelrite® (0.2, 0.3, 0.4 and 0.5%) and polyethylene glycol (0.0, 3.0, 5.0 and 7.5%) in the presence of 7.56 µmol abscisic acid and 3.0% sucrose. The highest rate of cotyledonary (70.0%) and normal (50.0%) somatic embryos was obtained using 0.3% Gelrite® without polyethylene glycol. Media with the least amount of Gelrite® (0.2%) induced the most secondary embryogenesis. Increasing the concentration of Gelrite® decreased the number of mature somatic embryos. The combination of Gelrite® and polyethylene glycol was the least effective for maturation of somatic embryos. Among polyethylene glycol concentrations, 7.5% polyethylene glycol produced the highest number of cotyledonary embryos and 3.0% polyethylene glycol produced the highest number of normal embryos (white-opaque embryos with two cotyledons and visible root and shoot meristems). Treatments were compared to medium with 0.2% Gelrite® and lacking polyethylene glycol (control). The 0.5% Gelrite® and 7.5% polyethylene glycol treatment produced the highest number of abnormal somatic embryos. In another experiment, different pre-germination treatments were tested to enhance somatic embryo germination. Germination response was evaluated in terms of the percentages of embryos exhibiting root-only elongation, shoot-only elongation, or both (conversion to plantlets). A cold pre-treatment (4°C for four weeks) in combination with plant growth regulators resulted in the highest rate of conversion to plantlets (55.0%). One month storage of somatic embryos at 4°C between maturation and germination treatments gave the lowest conversion frequency but did promote root-only germination. Addition of plant growth regulators (including 2.32 µmol kinetin (Kin), 2.22 µmol N6-Benzyladenine and 5.77 µmol gibberellic acid) to the germination medium for 4 weeks in the absence of a cold treatment was less effective for conversion to plantlets than growth regulator use in combination with cold, but the use in the absence of cold increased the frequency of shoot-only elongation. Plant growth regulator treatments decreased the frequency of root-only elongation regardless of cold treatment.
Key words: Gelrite®, plant growth regulator, polyethylene glycol, temperature, somatic embryogenesis.
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