Research Institute for Biochemical Regulation, School of Agriculture, Nagoya University, Chikusa, Nagoya 464, Japan.
Plant Physiol. 1985 Apr;77(4):963-8. doi: 10.1104/pp.77.4.963.
A method was developed for the isolation of vacuoles from the mesophyll protoplasts of spinach leaf, employing the discontinuous Ficoll density gradient centrifugation technique. Isolated vacuole preparations were judged to be free from other organellar fractions based on the assays of marker enzyme activities of individual organelles.Using this isolation method, a time-dependent translocation of (14)C-labeled photosynthates into vacuoles was determined. In contrast to a significant transport of (14)C organic acids such as malate and citrate within 10 to 15 minutes (14)C neutral sugars and amino acids were barely transported into vacuoles during 40 minutes incubation, in spite of the fact that a relatively large amount of these compounds are found in the vacuoles. It was also found that a majority of [(14)C]sucrose remains in the cytosol, apparently not actively moving into the vacuoles. Overall results appear to suggest that vacuoles are not actively engaged in photosynthetic carbon metabolism in spinach leaf protoplasts.
一种从菠菜叶片的质体原生质体中分离液泡的方法，采用不连续的 Ficoll 密度梯度离心技术。根据各细胞器的标记酶活性测定，分离的液泡制剂被判断为不含其他细胞器部分。使用这种分离方法，测定了（14）C 标记的光合作用产物向液泡中的时间依赖性转移。与（14）C 有机酸盐（如苹果酸和柠檬酸）在 10 至 15 分钟内的显著转运相反，在 40 分钟的孵育过程中，（14）C 中性糖和氨基酸几乎不转运到液泡中，尽管这些化合物在液泡中含量相对较大。还发现，大多数[（14）C]蔗糖仍留在细胞质中，显然没有主动进入液泡。总体结果表明，液泡在菠菜叶片原生质体中并没有积极参与光合作用碳代谢。
