PT  - JOURNAL ARTICLE
AU  - Siyu Feng
AU  - Aruna Varshney
AU  - Doris Coto Villa
AU  - Cyrus Modavi
AU  - John Kohler
AU  - Fatima Farah
AU  - Nebat Ali
AU  - Joachim Dieter Mueller
AU  - Miri VanHoven
AU  - Bo Huang
TI  - Bright split red fluorescent proteins with enhanced complementation efficiency for the tagging of endogenous proteins and visualization of synapses
AID  - 10.1101/454041
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 454041
4099  - http://biorxiv.org/content/early/2018/10/25/454041.short
4100  - http://biorxiv.org/content/early/2018/10/25/454041.full
AB  - Self-associating split fluorescent proteins (FPs) have been widely used for labeling proteins, scaffolding protein assembly and detecting cell-cell contacts. Newly developed self-associating split FPs, however, have suffered from suboptimal fluorescence signal. 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 animals, demonstrating its broad applications.