STOMATA AND STRUCTURE OF TETRAPLOID APPLE LEAVES CULTURED IN-VITRO

Leaves of anther-derived tetraploid apple (Malus pumila Mill.) shoots were examined by low-temperature scanning electron microscopy (LT-SEM). Leaves were serrate and wide with an undulating adaxial surface due to convex epidermal cells, apparently without crystalline epicuticular wax. Stomata were absent from the adaxial surface, except for the marginal teeth which exhibited 40-60 stomata per leaf; they probably originated from residual mitotic activity. One third of abaxial stomata was occluded by the residual cuticle of the mother guard cell across the stomatal pore which ruptured when the stomata became functional. The stomatal index was 7·2 (± 1·6) with 60-75 stomata mm-2, i.e. abaxial stomata of tetraploid leaves expanded in vitro were less frequent than those in triploid leaves either cultured in vitro (475-575 stomata mm-2) or grown on the tree (320-390 stomata mm-2) where the stomatal index was 21 (± 4). Freeze-fracture transsections showed that the tetraploid in vitro leaves were composed of a layer of adaxial epidermal cells, followed by a single layer of palisade cells and four to five layers of spongy mesophyll cells and the abaxial layer of epidermal cells, in contrast to juvenile seedling-grown apple leaves in which the two layers of palisade cells comprised the majority (52-60%) of the leaf volume. The same morphological features, such as wide and less pointed leaves, reduced stomatal density and stomatal index, and increased stomatal size that were previously reported for tree-grown tetraploid leaves were also expressed in vitro. Thus, causes of the stomatal deformation in tissue-cultured Rosaceae are interpreted to be in part genetic and not purely environmental.