Abstract：
In vitro reconstitution studies enable the controllable and stepwise investigation of complicated biochemical processes. In yeast and mammals, in vitro reconstitution of COPII vesicles marked a pivotal point in characterizing the endoplasmic reticulum-to-Golgi anterograde trafficking route and revealed how vesicles mediate the selective and reliable transportation among topologically equivalent compartments. By providing the necessary physiological conditions in a cell-free environment, it enables the dissection of essential components required for the vesicle formation. To enrich and purify the small amount in vivo membrane-bounded compartments, it simplifies the evaluation of vesicle regulation by distinct external stimuli or upstream signals. Here, we describe the preparation of plant microsomes and cytosol for the reconstitution of plant COPII vesicles. Purified vesicles can be used for further biochemical or microscopical analyses.
摘要：
体外重建研究可以对复杂的生化过程进行可控和逐步的研究。在酵母和哺乳动物中，COPII囊泡的体外重建标志着表征内质网至高尔基体顺行运输途径的关键点，并揭示了囊泡如何介导拓扑等效区室之间的选择性和可靠运输。通过在无细胞环境中提供必要的生理条件，它能够解剖囊泡形成所需的基本成分。为了富集和纯化体内少量的膜结合区室，它简化了通过不同的外部刺激或上游信号对囊泡调节的评估。这里,我们描述了用于重建植物COPII囊泡的植物微粒体和细胞质的制备。纯化的囊泡可用于进一步的生物化学或显微镜分析。
