Directed Evolution of Key Residues in Fluorescent Protein Inverses the Polarity of Voltage Sensitivity in the Genetically Encoded Indicator ArcLight.

ABSTRACT: Genetically encoded calcium indicators (GECIs) produce unprecedentedly large signals that have enabled routine optical recording of single neuron activity in vivo in rodent brain. Genetically encoded voltage indicators (GEVIs) offer a more direct measure of neuronal electrical status, however the signal-to-noise characteristics and signal polarity of the probes developed to date have precluded routine use in vivo. We applied directed evolution to target modulable areas of the fluorescent protein in GEVI ArcLight to create the first GFP-based GEVI (Marina) that exhibits a ΔF/ΔV with a positive slope relationship. We found that only three rounds of site-directed mutagenesis produced a family of "brightening" GEVIs with voltage sensitivities comparable to that seen in the parent probe ArcLigh

SUBMITTER: Platisa J 

PROVIDER: S-EPMC5355904 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature