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G3 (Bethesda). 2019 Jun 5;9(6):1933-1943. doi: 10.1534/g3.119.400162.

Tissue-Specific Split sfGFP System for Streamlined Expression of GFP Tagged Proteins in the Caenorhabditis elegans Germline.

Author information

1
Department of Biology, University of Iowa, Iowa City, IA 52240.
2
Department of Biology, University of Iowa, Iowa City, IA 52240 sarit-smolikove@uiowa.edu.

Abstract

Identifying protein localization is a useful tool in analyzing protein function. Using GFP-fusion tags, researchers can study the function of endogenous proteins in living tissue. However, these tags are considerably large, making them difficult to insert, and they can potentially affect the normal function of these proteins. To improve on these drawbacks, we have adopted the split sfGFP system for studying the localization of proteins in the Caenorhabditis elegans germline. This system divides the "super folder" GFP into 2 fragments, allowing researchers to use CRISPR/Cas9 to tag proteins more easily with the smaller subunit, while constitutively expressing the larger subunit from another locus. These two parts are able to stably interact, producing a functional GFP when both fragments are in the same cellular compartment. Our data demonstrate that the split sfGFP system can be adapted for use in C. elegans to tag endogenous proteins with relative ease. Strains containing the tags are homozygous viable and fertile. These small subunit tags produce fluorescent signals that matched the localization patterns of the wild-type protein in the gonad. Thus, our study shows that this approach could be used for tissue-specific GFP expression from an endogenous locus.

KEYWORDS:

AKIR-1; CRISPR; GFP1-10; GFP11; RPA-1; SYP-4; germline; meiosis; sfGFP; split GFP

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