橡胶树PR107原生质体的分离和培养研究

Protoplast Isolation and Culture of Hevea brasiliensis Müll. Arg. PR107

以橡胶树PR107花药愈伤组织建立的胚性悬浮细胞为材料,进行原生质体的酶解分离和培养研究。结果表明:在酶组合为纤维素酶1.5%+果胶酶0.15%+离析酶0.5%的酶解处理下,继代培养第6天、直径小于0.5 mm悬浮细胞团获得最高产量的原生质体,达到2.2×107个/mL PCV;原生质体在看护培养基上培养1周后观察到细胞开始发生分裂,培养2周后细胞分裂形成含8-10个细胞的小细胞团。看护培养45 d后原生质体持续分裂并发育成肉眼可见的小愈伤组织。

Embryogenic suspension cells derived from anther calli of PR107 was used as material for protoplast isolation and culture. Results showed that the highest yield of protoplast at 2.2× 107/mL PCV was isolated from suspension cell clusters less than 0.5 mm in diameter after 6 d of subculture when incubated in enzyme solution containing 1.5 % cellulose, 0.15 % pectinase and 0.5 % macerozyme. Protoplasts started to divide after one week of culture on feeder layer. Cell colony containing 8-10 cells was obtained after two weeks and continued to divide and develop into visible microcalli after 45 d of culture.