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Nat Chem Biol DOI:10.1038/s41589-018-0004-9

A robotic multidimensional directed evolution approach applied to fluorescent voltage reporters.

Publication TypeJournal Article
Year of Publication2018
AuthorsPiatkevich, KD, Jung, EE, Straub, C, Linghu, C, Park, D, Suk, H-J, Hochbaum, DR, Goodwin, D, Pnevmatikakis, E, Pak, N, Kawashima, T, Yang, C-T, Rhoades, JL, Shemesh, O, Asano, S, Yoon, Y-G, Freifeld, L, Saulnier, JL, Riegler, C, Engert, F, Hughes, T, Drobizhev, M, Szabo, B, Ahrens, MB, Flavell, SW, Sabatini, BL, Boyden, ES
JournalNat Chem Biol
Date Published2018 04

We developed a new way to engineer complex proteins toward multidimensional specifications using a simple, yet scalable, directed evolution strategy. By robotically picking mammalian cells that were identified, under a microscope, as expressing proteins that simultaneously exhibit several specific properties, we can screen hundreds of thousands of proteins in a library in just a few hours, evaluating each along multiple performance axes. To demonstrate the power of this approach, we created a genetically encoded fluorescent voltage indicator, simultaneously optimizing its brightness and membrane localization using our microscopy-guided cell-picking strategy. We produced the high-performance opsin-based fluorescent voltage reporter Archon1 and demonstrated its utility by imaging spiking and millivolt-scale subthreshold and synaptic activity in acute mouse brain slices and in larval zebrafish in vivo. We also measured postsynaptic responses downstream of optogenetically controlled neurons in C. elegans.


Alternate JournalNat. Chem. Biol.
PubMed ID29483642
PubMed Central IDPMC5866759
Grant ListU01 NS099691 / NS / NINDS NIH HHS / United States
R01 DA029639 / DA / NIDA NIH HHS / United States
R24 MH106075 / MH / NIMH NIH HHS / United States
R01 MH103910 / MH / NIMH NIH HHS / United States
R01 EY023173 / EY / NEI NIH HHS / United States
R01 GM104948 / GM / NIGMS NIH HHS / United States
R01 NS087950 / NS / NINDS NIH HHS / United States
DP1 NS087724 / NS / NINDS NIH HHS / United States
R43 MH109332 / MH / NIMH NIH HHS / United States
R01 NS104892 / NS / NINDS NIH HHS / United States