Isolation of chloroplasts for in organelle protein degradation assay

Abstract

A method for isolating intact chloroplasts from mature and senescent Arabidopsis thaliana leaves is described that utilizes two subsequent Percoll gradients. The isolated chloroplasts can be incubated in the dark to track in organelle protein degradation. To remove chloroplasts that lysed during the incubation period, a second Percoll gradient is performed prior to SDS-PAGE and immunoblot analysis.