| 2,4-D and a wounding treatment, alone and in combination, were used for callus initiation from micropropagated leaflets of Rosa hybrida L. cv John Davis. The only initiation treatment that resulted in callus formation was the combination of 2,4-D (5 mg/ml) and wounding treatment. Histological examination of the leaflets from the callus initiation treatments revealed potentially embryogenic cells were present in week-old calli. Examination of calli clusters and embryogenic tissue allowed the identification and characterization of non-embryogenic and embryogenic cells. Somatic embryos were initiated from the embryogenic tissue and were fixed and examined using light and fluorescence microscopy. The somatic embryos followed similar stages of development as have been described in the literature, e.g. globular, heart, torpedo and cotyledonary stages. The protoderm was the first tissue to differentiate. It was observed in the globular stage. The beginnings of the procambium, ground tissue and cotyledons were observed in the late globular/early heart-shaped stage. The root apical meristem (RAM) was developed by the cotyledonary stage. Shoot apical meristem (SAM) development was rarely observed. The lowest level of Abscisic Acid (ABA) tested, (0.1 mg/L), combined with highest level of sucrose tested, (40 g/L), resulted in the most mature somatic embryos. Maturation treatments including PEG1500 (polyethylene glycol 1500) yielded lower numbers of mature somatic embryos. Histological examination of mature rose zygotic embryos showed that the tissues of zygotic embryos are more regular, yet similar to somatic embryos. |
