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Nat Commun. 2016 Mar 18;7:11046. doi: 10.1038/ncomms11046.

Versatile protein tagging in cells with split fluorescent protein.

Author information

1
Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, USA.
2
Tetrad Graduate Program, University of California, San Francisco, California 94143, USA.
3
Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California 94143, USA.
4
Department of Biochemistry and Biophysics, University of California, San Francisco, California 94143, USA.
5
Howard Hughes Medical Institute, San Francisco, California 94143, USA.

Abstract

In addition to the popular method of fluorescent protein fusion, live cell protein imaging has now seen more and more application of epitope tags. The small size of these tags may reduce functional perturbation and enable signal amplification. To address their background issue, we adapt self-complementing split fluorescent proteins as epitope tags for live cell protein labelling. The two tags, GFP11 and sfCherry11 are derived from the eleventh β-strand of super-folder GFP and sfCherry, respectively. The small size of FP11-tags enables a cost-effective and scalable way to insert them into endogenous genomic loci via CRISPR-mediated homology-directed repair. Tandem arrangement FP11-tags allows proportional enhancement of fluorescence signal in tracking intraflagellar transport particles, or reduction of photobleaching for live microtubule imaging. Finally, we show the utility of tandem GFP11-tag in scaffolding protein oligomerization. These experiments illustrate the versatility of FP11-tag as a labelling tool as well as a multimerization-control tool for both imaging and non-imaging applications.

PMID:
26988139
PMCID:
PMC4802074
DOI:
10.1038/ncomms11046
[Indexed for MEDLINE]
Free PMC Article

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