Improved efficiency of somatic embryogenesis from zygotic embryos in Hyoscyamus niger by seed water-soaking

We describe an efficient procedure to obtain somatic embryos from mature zygotic embryos of Hyoscyamus niger (black henbane). It has several advantages over previous regeneration methods, which are: the use of mature seeds, an average 80% somatic embryogenesis rate and a high (eight-fold higher than the control) plant regeneration frequency. The critical step in this protocol was soaking of the seeds in sterile distilled water for 16 h, prior to excision and culture of zygotic embryos, on MS basal medium supplemented with 2% sucrose, 2 g/l myo-inositol, 0.5 g/l 2-(N-morpholino) ethanesulfonic acid and 1 mg/l α-naphthalene acetic acid (NAA). The regenerated somatic seedlings were fertile and were morphologically uniform. This procedure is simple, rapid and effective for high frequency of plant regeneration via somatic embryogenesis. Moreover, this new method should facilitate the development of strategies to routinely transform recalcitrant plant species, including henbane.