| Summary: | This study was carried out to establish an efficient regeneration protocol for mass propagation, uniform and clean planting materials of FD 010 (UniSZA) Ficus deltoidea Jack var. kunstleri (King) Corner. Aseptic study on leaves explant of F. deltoidea var. kunstleri taken from the field was carried out to establish an efficient sterilization protocol. It was found that the sterilization treatment could increase the in vitro survival percentage of field-derived explant rom 0% to 78.88%. It is also suggested tp pretreat the field plants by spraying with 2% Benlate once a week for at least one month before the explants are taken to the laboratory to get better survival percentage of explants up to 100%. The sterilization method for field-derived explants of F. deltoidea is as follows: wash the explants with a few drops of Teepol and place under running tap water fo 30 minutes. Rinse one time with distilled H2O and soak with 2% Benlate for about 30 minutes. Explants were rinsed with distilled H2O for three to five times or until the cloudy color of Benlate is gone. In the laminar air flow cabinet, the explants were soaked in 70% EtOH for one to two minutes followed by 0.1% HgCl2 for 5 minutes. Then the explants were soaked in 25% Clorox plus a few drops of Tween 20 for another 30 minutes. Finally, the explants were rinsed with sterile distilled H2O for 5 times. For every sterilant substitution from EtOH to HgCl2 or Clorox, explants should be rinsed for at least two times with sterile distilled H2O to eliminate traces of sterilants that might stick on the explants. In the study on callus induction and callus quality improvement of FD 010 (UniSZA) F. deltoidea var. kunstleri, it was found that the most suitable explant for the callus induction in a shorter time was the young leaves taken from stage one to three. The four different media that were screened were; Murashige and Skoog (1962) (MS), Bourgain and Nitsch (1967) (BN), Lloyd and Mc Cown (1980) (LM) and Modified Murashige and Skoog (Cb). MS medium with addition of 0.8 mg/L 2,4-D and 0.7 mg/L KIN was found to be the best because it gave good callus proliferation with friable and pale yellow morphology which are believed to be embryogenic. Many manipulations were carried out for the improvement of callus quality such as treatment with different modified MS media and different concentration of sucrose, coconut water, plant growth regulators (PGRs), silver nitrate (AgNO3), casein hydrolysates (CH) an glutamine (GLU). The results showed that Line FD 010 (UniSZA) F. deltoidea var. kunstleri was not able to form embryoids, despite all attempts to induce somatic embryogenesis and we are inclined to classify it as very recalcitrant. However, some of the results showed a glimpse of hope of an embryogenic tissue in the callus that was cultured on MS medium supplemented with 0.4 mg/L NAA, 0.8 mg/L BAP, 5% (g/L) sucrose, 10% (v,v) coconut water, 3 mg/L AgNO3, 400 mg/L GLU and 800 mg/L CH and MS medium supplemented with 2mg/L BAP, 0.2 mg/L IBA, 5% (g/L) sucrose, 10% (v,v) coconut water, 3mg/L AgNO3, 400mg/L CH, 400 mg/L GLU and 100 mg/L proline. It was also found that the use of 5% sucrose, 10% coconut water and 3mg/L AgNO3 were very helpful in maintaining and proliferating the callus. Other substances such as GLU and CH could also be used for the improvement of callus quality. Plantlet production through node culture was also attempted. The effects of different concentration of BAP ( 0.0, 0.2, 0.5, 1.0, 2.0, 4.0 and 8.0 mg/L) and IBA (0.0, 0.2, 0.5, 1.0 and 2.0 mg/L) were evaluated for shoot and root induction. It was found that 1 mg/L BAP was the most efficient on shoot proliferation, while 2 mg/L IBA was considered the most efficient for the rooting of shoots. The qualitative study of phytochemical constituents showed that all the phytochemicals tested (tannins, phlobatannins, saponins, flavonoids, steroids, terpenoids, alkaloids, anthraquinones and polyphenols) were present in both leaves and callus of FD 010 (UniSZA) F. deltoidea var. kunstleri except for cardiac glycosides that was only present in leaves but not in callus. The quantitative analysis showed that the callus do have the same phytochemicals like the intact plant but in loer amount. The quantitative analysis of five major groups of phytochemical contituents (phenols, flavonoids, tannins, alkaloids, saponins) showed that leaves contain (0.28+0.01, 0.58+0.08, 10.39+0.15, 1.75+0.12, 4.14+0.26) and callus contains (0.25+0.01, 0.44+0.03, 9.98+0.32, 0.72+0.12, 1.85+0.18) respectively. The findings of this study hopefully coul be used as a source of information and act as a baseline for more intensive Research & Development Program on this particular species. The callus pathway through somatic embryogenesis will be needed for genetic engineering and transformations studies in order to obtain new cultivars while the node culture will be a faster and easier alternative for the micropropagation of this species. The study of phytochemical constituents of leaves and callus is important to know and prove the existance of certain phytochemicals that are found in this plant. It couls be used to explain scientifically the use of this plant in the traditional treatment and reveal the potential of the callus, which may be used in the pharmaceutical industry.
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