Studies On The Establishment Of Efficient Regeneration System And Genetic Transformation In Phyllanthus Emblica L.

Aonla or Indian gooseberry (Phyllanthus emblica L.) is an important fruit indigenous to South Asia, particularly to central and south India. The fruit has always been recognized for its nutritional and medicinal values. China is rich in Aonla resources. The improvement of Aonla horticultural traits is very important to expand Aonla's culture. But the improvement is limited through conventional breeding, because of a short knowledge to its complex genetic background. Recently, genetic engineering has opened a new platform to modify fruits. Many possible horticultural characters are expected to be genetically engineered into Aonla in the near future. An efficient tissue culture system is therefore thought to be crucial to the success of plant genetic engineering in Agrobacterium-based transformation protocols. In the study some factors that influenced the efficiency of the regeneration of the infected tissues had been examed. The main results observed are as follows:1. An efficient plant regeneration protocol for Phyllanthus emblica.L. was achieved via organogenesis from callus tissues formed on hypocotyl explants in vitro. In this paper, effects ofexplants type, medium formations, different combinations of growth regulators, sucrosetconcentrations, AgNO₃ concentrations and dark treatment on callus, shoot and root formation wereexamined. After 5 weeks of culture maximum shoot regeneration frequency of 100% with 11.7 shoots per explant was obtained on 1/2 MS medium containing 0.5μM NAA, 5μM BA, 40g/L sucrose, 1mg/L AgNO₃, inoculated in the dark for 14 days. Excised shoots rooted when they were transferred to half-strength of MS medium containing 10μM IBA after 5 weeks and developed into healthy plantlets, which generated a rooting rate of 29.2%. The results of this study will facilitate the application of genetic transformation methods in Phyllanthus emblica.L.2. Effects of antibiotics on in-vitro regeneration and organogenesis of Phyllanthus emblica.L. from hypocotyl explants were examined. Kan inhibited significantly in-vitro regeneration and organogenesis. Callus and shoot formation from hypocotyl explants was completely inhibited in presence of 20mg/L Kan. Results also showed that all treatments of Cef and Carb inhibited shoot differentiation. Relatively, the effect of Cef was better than that of Carb. When 300mg/L Cef was present in the regeneration medium, both shoot formation rate and mean number of shoots perexplant were 70.8% and 5.3shoots/explant from hypocotyl explants. It was considered that Cef 300mg/L was suitable for Phyllanthus emblica.L. to elimination of Agrobacterium-tumefaciens during transgenic shoot induction;kan 20mg/L was suitable in the selection of transgenic versus false-positive shoots from hypocotyl explants.In the study the genetic transformation of Phyllanthus emblica.L. were first investigated and optimal concentration of antibiotic on hypocotyl explants of Phyllanthus emblica.L. was established. The results of this study will facilitate the application of genetic engineering methods to improve the horticultural traits of Phyllanthus emblica.L.