12. Fluorescent Reporter Methods
By: Harald Hutter4
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The identification and cloning of the green fluorescent protein (GFP) from jellyfish marks the beginning of a new era of fluorescent
reporters. In Caenorhabditis elegans, genetically encoded markers like the fluorescent proteins of the GFP family became the reporter of choice for gene expression
studies and protein localization. The small size and transparency of the worm allows the visualization of in vivo dynamics,
which increases the number of potential applications for fluorescent reporters tremendously. In combination with subcellular
tags, GFP can be used to label subcellular structures like synapses allowing novel approaches to study developmental processes
like synapse formation. Other fluorescent labels like small organic dyes, which are in widespread use in cell culture systems,
are rarely used in C. elegans owing to difficulties in applying these labels through the impenetrable cuticle or eggshell of the animal. A notable exception
is the use of lipophilic dyes, which are taken up by certain sensory neurons in the intact animal and can be introduced into
the embryo after puncturing of the egg shell. This chapter covers the use of fluorescent dyes and fluorescent proteins in
C. elegans. Emphasis is placed on microscopic techniques including wide field and confocal microscopy as well as time-lapse recordings.
The use of fluorescent proteins as transgenic markers and image processing of fluorescence images are briefly discussed.
Affiliation(s): (4) Department of Biological Sciences, Simon Fraser University, Burnaby, British Columbia, Canada
Book Title: C. elegans: Methods and Applications
Series: Methods in Molecular Biology | Volume: 351 | Pub. Date: Aug-15-2006 | Page Range: 155-173 | DOI: 10.1385/1-59745-151-7:155
Subject: Genetics/Genomics
Key Words: GFP - CFP - YFP - DsRed - DiI - confocal microscopy - C. elegans - time lapse - fluorescent reporter - embryo
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