無窮花 原形質體 融合에 의한 體細胞 雜種 誘起 Ⅰ. 原形質體 分離에 있어 關聯要因調査

Somatic hybridization by protoplast fusion in hibiscus syriacus and H. rosa-sinensis Ⅰ. Factors affected to protoplast isolation

Mesophyll and callus tissues of Hibiscus syriacus and H. rosa-sinensis obtained through in vitro culture of immature embryos were used for protoplast isolation, and several factors influencing the yield were investigated in this experiment. Protoplast isolation from mesophyll tissues required preplasmolysis in CPW 13M medium for 3 or 5 hours under dark conditions. Treatment of mesophyll tissues to 1.0mg/ℓ 2,4-D solution for 12 hours prior to preplasmolysis increased yield of protoplast in H. syriacus and 3.0mg/ℓ 2,4-D for 24 hrs in H. rosa-sinensis. 0.7M mannitol in CPW medium was the most effective as osmoticum and protoplast yield was increased in the enzyme mixture of 3-5% Cellulase, 0.5% Macerozyme and 0.2% Driselase. Physiological age of mesophyll tissue in vitro influenced significantly on the protoplast yield:young tissue grown for less than 5-week was more productive than 8-week old ones for high protoplast yield. Optimum amount of mesophyll and callus tissues in a definite 5 ml of enzyme mixture was 400 and 500 mg, respectively and optimum digestion time for healthy protoplast yield ranged from 5 to 7 hours.