‘石牌’广藿香叶肉原生质体的分离及纯化

以‘石牌’广藿香无菌苗叶片为材料,对原生质体分离、纯化方法以及影响因素进行了研究。结果表明:以继代培养12～22 d的无菌苗顶芽下第3对展开叶片为材料,用0.5%果胶酶、0.2%离析酶和1.5%纤维素酶作用8 h,渗透压调节剂为11%甘露醇,‘石牌’广藿香叶肉原生质体产量达1.85×107个/g fw,活力达89%;原生质体纯化以12%聚蔗糖(Ficoll)漂浮法效果最佳。

Isolation and Purification of Mesophyll Protoplasts of Pogostem cablin cv. shipaiensis

Mesophyll protoplasts were isolated and purified from leaves of in vitro propagated Polgostemon cablin(Blanco) Benth.cv.shipaiensis,and different factors affecting protoplasts yield and activity were investigated.The results showed that young leaves of seedling subcultured for 12～ 22 days in the enzyme solution consisting of 0.5%(w/v) pectolyase Y-23,0.2%(w/v) Macerozyme R-10 and 1.5%(w/v) cellulase R-10,in 11%(w/v) mannitol buffered with 0.1% MES and 0.02% CaCl2 for 8 h,the yield was 1.85×107 protoplasts per gram leaves(fresh weight),the viability was above 89%.