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Different friable Hevea callus lines from the same genotype can display different embryogenic and regenerative potentials. These lines can not be distinguished on the basis of macroscopic criteria. A histological and molecular study was undertaken to characterize the differences existing between five callus lines with different potentials. The genes differentially expressed during induction were analyzed using the differential display (DD-RT) technique. Twenty-eight cDNAs were found to be differentially expressed during induction in the embryogenic regenerating line (ER). Embryogenic nodules were formed earlier in the ER lines than in the embryogenic non-regenerating lines and were completely absent in the non-embryogenic line. Of these 28 cDNAs, five could be used to distinguish between calli prior to induction, thereby enabling an early diagnosis of friable Hevea callus embryogenic potential.

Abstract: The effect of different carbohydrates was tested on early somatic embryogenesis of Hevea brasiliensis. Sucrose was replaced with maltose, fructose or glucose. Somatic embryo production was significantly higher with maltose. With maltose, the initial yellow colour of the calli turned orange, and dry matter production after 28 days' culture was half that obtained with sucrose. Maltose also reduced the soluble sugar content by around 60%, whereas in the presence of sucrose, the soluble sugar content of calli increased. On maltose medium, embryonic cells were initiated after 2 weeks, whereas calli obtained on media containing sucrose mostly consisted of parenchymatous cells. The embryogenic cells rapidly formed into globular embryos. The origin of the `maltose' effect is discussed: is it carbohydrate stress or a signal molecule? The effect of maltose was checked on 6 Hevea callus lines maintained in the laboratory.

The mineral composition of Hevea brasiliensis shoots was analyzed at different stages of the microcutting procedure from mother plants grown in greenhouses to rooted microcuttings at the acclimatization stage. Compared to the mineral status of the mother plants, a mineral imbalance was noticed in shoots right from the in vitro establishment phase. Phosphorus (P), potassium (K), sodium (Na), chloride (Cl), and zinc (Zn) contents increased, while calcium (Ca), magnesium (Mg), iron (Fe), and copper (Cu) diminished, resulting in a substantial modification to the mineral balance. The N: P, N: K, Mg: P and K: Na ratios were very low, whereas the K: Mg, K: P, and K: Ca ratios were very high. During the multiplication phase, the N: P ratio returned virtually to its initial value but the other ratios remained modified. A mineral analysis of shoots after three years multiplication in vitro revealed a very low N: P ratio, a very high K: Ca ratio and a normal K: Na ratio. The same results were obtained right from the initial culture cycles on shoots developed in vitro from mother plants with a high multiplication potential. These results are discussed and lead us to consider the N: P, K: Ca, and K: Na ratios as potential markers of Hevea brasiliensis multiplication ability in vitro. During the rooting phase, the Mg: P and K: Ca ratios returned to their normal values, whereas the other ratios returned to normal values on formation of the second leaf flush ex-vitro.

Nodal cuttings (primary nodal explants) were used to initiate microcutting cultures ofHevea brasiliensisandTheobroma cacao. The physiological stage of growth reached by the branches from which the explants were taken had a considerable influence on culture initiation and response. In both species, the buds of the nodal explants taken from dormant branches (stages 11, 12 and 13 inTheobroma; stage D inHevea) revealed greater opening abilityin vitroand shoot elongation than buds taken from branches in a phase of active meristem growth inTheobroma(stages F1 and F2) or during a foliar growth phase inHevea(stage C). InHevea, the branch growth phase effect persisted into the multiplication phase when shoots developedin vitrowere used as the source of secondary nodal explants. However, it disappeared during subcultures of primary nodal explants. In both species, the position of the nodes on the branch with respect to the apex also had an effect on culture initiation. The buds of nodes furthest from the apex were more likely to develop into shoots. InHevea, the position of secondary nodal explants obtained from the primary shoot was seen to vary in effect depending on growth regulator applications during primary culture. These results are analyzed in terms of variations in the balance between endogenous growth promoters/inhibitors in the buds, depending on morphogenic stages and the bud position on the branch.