霞草原生质体培养和植株再生

PLANT REGENERATION FROM PROTOPLASTS ISOLATED FROM SUSPENSION-CULTURED CELLS OF GYPSOPHILA OLDHAMINA MIQ

用霞草胚性悬浮细胞分离原生质体，以含０．２％琼脂糖的ＫＭ８ｐ培养基薄层漂浮培养，原生质体培养密度６×１０＾３－１×１０＾４／ｍｌ。培养３天再生细胞开始分裂，７天统计分裂频率最高达２５．４％，１０天形成小细胞团，并加降低渗透压的稀释培养基，每周一次，２０－２５天形成肉眼可见的小愈伤组织，植板率达３．５％。成肉眼可见的小愈伤组织Ｔ谠鲋撑嘌奔尤耄埃常埃矗セ钚蕴坑欣谏ぜ胺只Ｔ诤叮拢粒常?

Protoplasts of Gypsophila oldhamiana Miq. were enzymatically isolated from the suspension cultured cells. The protoplasts were cultured in the thin solid KM 8 P medium solidified by 0.2% agarose, which floating on the surface of liquid KM 8 P medium. In the media, the optimum plant growth regulators and their concentrations and combinations were 2, 4-dichloropheno-xyacetic acid (2,4-D) 0.5mg/L, naphthalene acetic acid (NAA) 0.8mg/L and zeatin 0.3mg/L, The optimum density of cultured protoplasts was ranging 6× 103/ml to 1×104/ml. The culture condition was 25× in dark. The first divisions of regenerated cells were observed at 3 days from initial culture and the division frequency reached to 25.4% at 7 days. Ten days after culture, a dilution medium which was half concentration in mannitol was added and the dilution was repeated at one week interval thereafter. The cultures were transferred to light condition with an intensity about 800-1000 1x. Micro-calli observed with naked eyes were formed after being cultured for 20-25 days. A 3.5% plating efficiency was achieved. When transferred on modified MS differentiation medium supplemented with indol butyric acid (ISA) 0.8mg/L and 6-benzylaminopurine(6-BA) 3.5 mg/L, calli were differentiated adventitious shoots. The differentiation frequency was up to 85%. Regenerated shoots were rooted in a half strength MS medium supplemented with NAA 0.5 mg/L and 6-BA 0.05 mg/L. The plantlets with normal morphology were transplanted into peat/soil mixture and growth vigorously under green house conditions.