Document Type

Article

Publication Date

September 2019

Publication Title

Communications Biology

Volume

2

Issue Number

1

DOI

10.1038/s42003-019-0589-x

ISSN

23993642

Keywords

Fluorescence imaging, Fluorescent proteins, Genetic engineering, Neural circuits

Disciplines

Biology

Abstract

Self-associating split fluorescent proteins (FPs) are split FPs whose two fragments spontaneously associate to form a functional FP. They have been widely used for labeling proteins, scaffolding protein assembly and detecting cell-cell contacts. Recently developments have expanded the palette of self-associating split FPs beyond the original split GFP1-10/11. However, these new ones have suffered from suboptimal fluorescence signal after complementation. Here, by investigating the complementation process, we have demonstrated two approaches to improve split FPs: assistance through SpyTag/SpyCatcher interaction and directed evolution. The latter has yielded two split sfCherry3 variants with substantially enhanced overall brightness, facilitating the tagging of endogenous proteins by gene editing. Based on sfCherry3, we have further developed a new red-colored trans-synaptic marker called Neuroligin-1 sfCherry3 Linker Across Synaptic Partners (NLG-1 CLASP) for multiplexed visualization of neuronal synapses in living C. elegans, demonstrating its broad applications.

Comments

This article was published in Communications Biology, volume 2, issue 1, 2019, and can also be found online here. © The Author(s) 2019

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This work is licensed under a Creative Commons Attribution 4.0 License.

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