Influences of aging and cloning methods on the capacity for somatic embryogenesis of a mature Hevea brasiliensis genotype.

We compared embryogenic capacities of integument explants excised from three sources of the Hevea brasiliensis (Müll. Arg.) mature genotype PB 260. The three sources were 17-year-old (BT 86) and 7-year-old (BT 96) budded trees and 7-year-old emblings (EM 96). The highest proportions of embryogenic calluses obtained from the total number of integument explants initially used were from trees of EM 96 origin, followed by BT 96 trees, with explants from BT 86 trees producing the lowest number of embryogenic calluses. Further initiation of embryogenic callus lines from the primary somatic embryos derived from the three sources was successful only for EM 96. Somatic embryo cultures from BT 86 and BT 96 sources produced only friable calluses that could not be further amplified. Overall, somatic embryo explants derived from EM 96 responded over a wider range of 3,4-dichlorophenoxyacetic acid and kinetin concentrations than the somatic embryo explants from BT 86 and BT 96 origins. The effects of chronologic, ontogenetic and physiologic aging on explant capacity for somatic embryogenesis and on the overall efficiency of the process in H. brasiliensis are discussed.

Effect of exogenous calcium on post-thaw growth recovery and subsequent plant regeneration of cryopreserved embryogenic calli of Hevea brasiliensis (Müll. Arg.).

A reliable cryopreservation technique was developed for friable embryogenic callus lines of Hevea brasiliensis. The study showed that reducing the CaCl(2) concentration of the pre-culture medium from 9 mM to 1 or 0 mM CaCl(2) before cryopreservation promoted post-thaw callus growth, 1 mM being the optimum CaCl(2) concentration for embryo regeneration. Post-thaw callus proliferation decreased in line with the increase of plated callus weight. The effect of cryopreservation was assessed on 39 independent lines showing that cryopreservation did not affect embryogenic and plant regeneration for a majority of lines. The decrease in CaCl(2) concentration of the pre-culture medium led to a drop in callus calcium content indicating a direct link between the CaCl(2) concentration of the pre-culture medium and the endogenous calcium content of the calli. It also highlighted the implication of tissue calcium content in cryotolerance. Callus water status and the different ways by which calcium could prevent cryoinjury is also discussed.