Plant Regeneration via Somatic Embryogenesis in Mature Wild Olive Genotypes Resistant to the Defoliating Pathotype of Verticillium dahliae

Abstract

Regeneration capacity, via somatic embryogenesis, of four wild olive genotypes differing in their response to defoliating Verticillium dahliae (resistant genotypes StopVert, OutVert, Ac-18 and the susceptible one, Ac-15) has been evaluated. To induce somatic embryogenesis, methodologies previously used in wild or cultivated olive were used. Results revealed the importance of genotype, explant type, and hormonal balance in the induction process. Use of apical buds obtained from micropropagated shoots following a methodology used in cultivated olive (4 days induction in liquid 1/2 MS medium supplemented with 30 µM TDZ–0.54 µM NAA, followed by 8 weeks in basal 1/2 MS medium) was adequate to obtain somatic embryos in two genotypes, StopVert and Ac-18, with a 5.0 and 2.5% induction rates, respectively; however, no embryogenic response was observed in the other two genotypes. Embryogenic cultures were transferred to basal ECO medium supplemented with 0.5 µM 2iP, 0.44 µM BA, and 0.25 µM indole-3-butyric acid (IBA) for further proliferation. Somatic embryos from StopVert were maturated and germinated achieving a 35.4% conversion rate. An analysis of genetic stability on StopVert, using Simple Sequence Repeats (SSRs) and Random Amplified Polymorphic DNA (RAPDs) markers, was carried out in embryogenic callus, plants regenerated from this callus and two controls, micropropagated shoots used as explant source, and the original mother plant. Polymorphism was only observed in the banding pattern generated by RAPDs in 1 of the 10 callus samples evaluated, resulting in a variation rate of 0.07%. This is the first time in which plants have been regenerated via somatic embryogenesis in wild olive.

Somatic Embryogenesis from Mature Embryos of Olea europaea L. cv. ‘Galega Vulgar’ and Long-Term Management of Calli Morphogenic Capacity

“…Individual genotypes within the same species vary greatly in embryogenic capacity, reflecting substantial differences in the ability to activate key elements for the achievement of embryogenic competence [ 41 ] which shows the need to develop improved methodologies for each genotype. Results reported by Narváez and co-authors [ 39 ] emphasize this dependence, showing SE induction efficiency in two olive wild genotypes from the four initially considered, and embryos conversion from a single one.…”

“…As mentioned by Sanchéz-Romero [33], the conversion of olive somatic embryos into plants did not meet rigorous maturation requirements. Even when individualized in earlier developmental stages, an acceptable conversion rate is achieved [76], unlike other species where somatic embryos only germinate when they reach advanced stages of development [39,77]. Recalcitrant behavior has been described in embryos conversion when mature olive plant tissues are used as initial explants, and exceptions are only known in cv.…”

“…Recalcitrant behavior has been described in embryos conversion when mature olive plant tissues are used as initial explants, and exceptions are only known in cv. 'Canino' and 'Moraiolo' [34] and one wild olive genotype [39].…”

“…The first report is known from Capelo and co-authors [38], which demonstrated the possibility to use petioles/leaf tissues taken from plants established under greenhouse conditions (no juvenile or rejuvenated material), to induce SE response with efficient differentiation of somatic embryos. More recently, Narváez and co-authors [39] described a protocol focused on different olive wild genotypes differing in their response to defoliating Verticillium dahlia, in which was stated the use of shoot apex collected from in vitro plantlets as the most appropriated tissue to induce SE and further efficient embryos conversion.…”

“…The main factors affecting SE efficiency are the genotype of the donor plant, the type of explant taken for the establishment of in vitro cultures, the development and physiological stage, the growth conditions (mainly photoperiod and temperature), and the chemical composition of the culture media (including type and concentration of growth regulators) [ 40 , 41 ]. The genotype strongly affects the ability of a tissue to dedifferentiate and de novo acquire meristematic competence and to differentiate embryogenic structures [ 39 , 42 , 43 , 44 ]. Individual genotypes within the same species vary greatly in embryogenic capacity, reflecting substantial differences in the ability to activate key elements for the achievement of embryogenic competence [ 41 ] which shows the need to develop improved methodologies for each genotype.…”