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Titlebook: Membrane Trafficking; Methods and Protocol Jingshi Shen Book 2022 The Editor(s) (if applicable) and The Author(s), under exclusive license

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Measuring Plasma Membrane Recycling Using Microscopic and Biochemical Approachesis exclusively required to transport these cargoes through various intracellular routes for their delivery to the site of action. Among these, recycling of cargoes to the plasma membrane is a crucial pathway for the efficient functioning of the cell. Hence, endosomal cargo recycling assays are cruci
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Book 2022tro. biochemical and biophysical assays. Covering virtually all the major trafficking branches, the book delves into the exocytic pathway, which focuses on cargo transport from the ER to the Golgi, through the Golgi cisternae, and to the plasma membrane and the extracellular space; the endocytic pat
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Generation of Endogenously Tagged Membrane Trafficking Regulators Using CRISPR Genome Editingthe advent of CRISPR technologies, epitope tags and fluorescent proteins can now be introduced into endogenous proteins in almost any cell type that are proliferating in culture. This chapter describes approaches for inserting tags at the endogenous loci of genes, with the vesicle tethering protein complex, exocyst, as an example.
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Imaging Insulin Granule Dynamics in Human Pancreatic β-Cells Using Total Internal Reflection Fluoresocking and fusion dynamics and can also map its regulators with high spatiotemporal resolution. Here, we describe the basic principles and practical implementation of a fast dual-color TIRF microscope, detailing a how-to guide on imaging and analysis of insulin granule dynamics in human β-cells.
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Localizing Proteins on Single Trafficking Organelles in 3D with Semisynthetic Gold Labeling and Platund that this combined method can label and localize proteins with isotropic high precision to generate quantitative maps of protein positions in and around trafficking organelles at the inner plasma membrane of mammalian cells.
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Determine the Function of the Exocyst in Vesicle Tethering by Ectopic Targeting recruits the other exocyst subunits and secretory vesicles to a target membrane, suggesting that it plays a pivotal role in vesicle tethering. This approach is likely appropriate for studying other tethering complexes at their specific stages of trafficking and may also be used in other eukaryotic cells such as the cultured mammalian cells.
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