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Density Gradient Centrifugation for Enrichment and Identification of GFP-tagged Chitosomal Microvesicles of Filamentous Fungi   

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Original research article

A brief version of this protocol appeared in:
Molecular Microbiology
Feb 2015

Abstract

Density gradient centrifugation has been utilized to characterize the subcellular distribution of physiologically relevant enzymes in yeasts and filamentous fungi (Leal-Morales et al., 1988; Martínez et al., 1989; Kamada et al., 1991). This approach is now potentiated by protein tagging and live imaging techniques, which make possible to relate a single protein with, for example, a discrete population of intracellular vesicles and their in vivo dynamics (Verdín et al., 2009; Fajardo-Somera et al., 2013; Sánchez-León et al., 2015). Here, we describe the density gradient centrifugation and fractionation analysis of cell-free homogenates of a Neurospora crassa (N. crassa) strain that expresses CHS-6 chitin synthase fused to the green fluorescent protein (Riquelme et al., 2007).

Keywords: Neurospora crassa, Chitosomes, Vesicles, Fractionation

Copyright: © 2015 The Authors; exclusive licensee Bio-protocol LLC.
How to cite: Verdín, J., Sánchez-León, E., Fajardo-Somera, R., Morales, C. A., Bartnicki-García, S. and Riquelme, M. (2015). Density Gradient Centrifugation for Enrichment and Identification of GFP-tagged Chitosomal Microvesicles of Filamentous Fungi. Bio-protocol 5(19): e1611. DOI: 10.21769/BioProtoc.1611.
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