Cyclic somatic embryogenesis and scheme for multiplication of Quercus ilex L.

This work studied the induction of somatic embryogenesis in several holm oak explants. Subsequently, several aspects were studied such as those that affect somatic embryo development and maturation. Quercus ilex L. is the most important tree for the micorrhization with Tuber melanosporum Vitt. Firstly, several Q.ilex L explants were taken in order to induce somatic embryogenesis, such as hipocotyls, mature and juvenile plant leaves and acorns that were gathered during the summer. Several plant growth regulators (PGRs) e.g. naphthalenacetic acid (NAA), 6 benzyladenine (BA) and 2,4-dichlorophenoxy acetic acid (2,4-D) and different culture media were assayed. The immature zygotic embryos were cultured in darkness during 30 days on MS medium containing Gamborg's (1966) (G) as macronutrients. Next, the explants were transferred to a basal medium supplemented with 0.5 muM BA and 0.5 muM NAA, and cultured for 30 days in light. Somatic embryogenesis appeared in all media tested, including the control, except the one containing 3 muM 2,4-D. Somatic embryos appeared in a friable and yellowish embryogenic callus, or directly in the meristem axis. Later, the embryogenic callus was transferred to a medium free of PGRs, and secondary embryogenesis appeared, producing a large number of clonal plants. The macronutrient formula of Schenk and Hildebrandt (1972) (SH) gave better outputs for the generation of secondary embryogenesis and vigor than macronutrients formulas such as Murashige and Skoog (1.962) (MS), 1/2 MS, Woody Plant Medium (Lloyd and McCown, 1980) (WPM), 2SH, 1/2 SH, G and 1/2G. The presence of BA in the medium was not necessary for recurrent embryogenesis generation. The macronutrient formula of 1/2SH gave better outputs in holm oak somatic embryo maturation than all other formulas tested, as well as a lower percentage of secondary embryogenesis.