Department of Botany, Louisiana State University, Baton Rouge, Louisiana 70803.
Plant Physiol. 1991 Dec;97(4):1576-80. doi: 10.1104/pp.97.4.1576.
A simple procedure that yields highly purified intact chloroplasts from Chlamydomonas reinhardtii is described. This procedure involves breakage of cell wall-deficient cells by passing them through a narrow bore syringe needle. The intact chloroplasts are then purified from the crude homogenate by differential centrifugation and Percoll gradient centrifugation. This procedure generates relatively high yields of chloroplasts capable of CO(2) fixation. These chloroplasts were characterized by electron microscopy, marker enzyme analysis, and ferricyanide exclusion. Transmission electron microscopy indicates that these chloroplasts retain their pyrenoids and eyespots. Scanning electron microscopy confirms that the characteristic cup shape of C. reinhardtii chloroplasts persists in vitro. This rapid, inexpensive procedure produces chloroplasts that should be useful for researchers studying the biochemistry and cell biology of C. reinhardtii chloroplasts.
介绍了一种从莱茵衣藻中获得高度纯化完整叶绿体的简单方法。该方法通过将细胞壁缺陷的细胞穿过细口径注射器针头来进行细胞破碎。然后通过差速离心和 Percoll 梯度离心从粗匀浆中纯化完整的叶绿体。该方法可以产生相对较高产量的能够固定 CO2 的叶绿体。这些叶绿体通过电子显微镜、标记酶分析和铁氰化物排斥进行了表征。透射电子显微镜表明，这些叶绿体保留了它们的淀粉核和眼点。扫描电子显微镜证实，莱茵衣藻叶绿体的特征杯状在体外仍然存在。这种快速、廉价的方法产生的叶绿体应该对研究莱茵衣藻叶绿体的生物化学和细胞生物学的研究人员有用。
