This study aims to develop a method for isolating and purifying protoplasts/vacuoles from fresh leaves of the Cd hyperaccumulator plant species, Sedum alfredii. The results revealed that preheating cellulase and macer...This study aims to develop a method for isolating and purifying protoplasts/vacuoles from fresh leaves of the Cd hyperaccumulator plant species, Sedum alfredii. The results revealed that preheating cellulase and macerozyme at 50 ℃ for 5 rain significantly accel- erated the cell wall degradation. For the most optimal conditions for mesophyll protoplast isolation, the mixture of fresh leaves and cell lysates was followed by a 2-h-long vibration. The protoplast lysate for vacu- ole isolation was diluted, and 0.675 mmol/L was identi- fied as the most appropriate 3-[（3-cholamidopropyl） dimethylammonio]- 1 -propanesulfonic acid （CHAPS） level, in which S. alfredii large vacuoles are charac- terized by a high metal and malic acid content.展开更多
基金Project supported by the National Natural Science Foundation of China(Nos.31370040 and 41401366)the Zhejiang Provincial Natural Science Foundation of China(No.LR14C150001)
文摘This study aims to develop a method for isolating and purifying protoplasts/vacuoles from fresh leaves of the Cd hyperaccumulator plant species, Sedum alfredii. The results revealed that preheating cellulase and macerozyme at 50 ℃ for 5 rain significantly accel- erated the cell wall degradation. For the most optimal conditions for mesophyll protoplast isolation, the mixture of fresh leaves and cell lysates was followed by a 2-h-long vibration. The protoplast lysate for vacu- ole isolation was diluted, and 0.675 mmol/L was identi- fied as the most appropriate 3-[（3-cholamidopropyl） dimethylammonio]- 1 -propanesulfonic acid （CHAPS） level, in which S. alfredii large vacuoles are charac- terized by a high metal and malic acid content.
