Preparation of Plastoglobules from Arabidopsis Plastids for Proteomic Analysis and Other Studies

Abstract

Plastoglobules are particles specifically located inside different types of plastids. They mainly contain lipids and proteins and are physically attached to thylakoids. Proteomic studies have underlined the role of plastoglobules in diverse plastid metabolic pathways, such as those producing vitamin K, vitamin E, and carotenoids, and have implicated them in plant response to stress. This chapter describes the isolation of pure and intact plastoglobules from Arabidopsis leaves. The procedure starts with the isolation of intact chloroplasts by centrifugation on a Percoll gradient. Plastoglobules are then separated from the plastid membranes by flotation on a sucrose gradient. Finally, the purity of the plastoglobule fraction is verified by immunoblotting.