The plant Golgi apparatus is composed of numerous stacks of cisterna, designated as cis, medial, and trans Golgi cisternae; these stacks move within the cytoplasm along the actin cytoskeleton. Cis cisternae receive secretory products from endoplasmic reticulum (ER) and they subsequently progress through the stack to the trans cisternae, where they are sorted to other destinations, including cell wall, plasma membrane (PM), vacuoles, and chloroplasts. In addition, the plant Golgi apparatus plays a role of glycosylating proteins as well as synthesizing cell wall polysaccharides, such as hemicelluloses and pectins. This protocol describes procedures for imaging fluorescently-tagged proteins localized to the plant Golgi apparatus of Arabidopsis seedlings using confocal laser microscopy (CLSM), total internal reflection fluorescence microscope (TIRF), and immunogold labeling of high-pressure frozen/freeze substituted samples by transmission electron microscopy (TEM). We particularly focus on long-term time lapse imaging and protein localization in subdomains within the Golgi. This protocol can be also used for other organelles, tissues, and plant species.
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