Abstract
ABSTRACT Effects of culture medium supplements and osmotic stress treatment on embryogenic callus induction andsomatic embryogenesis were investigated in order to optimize tissue culture conditions of alfalfa( Medicagosativa L.). SH medium containing 5mg/L 2,4-D and 0.2mg/L kinetin was optimal for embryogenic callusinduction from cotyledon tissue of alfalfa. Somatic embryos were formed when the embryogenic callus wascultured on SH medium supplemented with 1mg/L 2,4-D and 2mg/L BA. Supplementation of 5mM L-pro-line and 1g/L casein hydrolysate into the regeneration medium further increased plant regenerationfrequency. Osmotic stress treatment of callus appeared to improve the frequency of somatic embryo for-mation, but the frequency of somatic embryo formation differed by the osmotic stress treatment using dif-ferent osmotic stressors. The highest plant regeneration frequency of 30.7% was observed when embryogeniccallus was treated with 0.7M sucrose for 18h. Efficient regeneration system established in this study will beuseful for molecular breeding of alfalfa through genetic transformation.
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