A modular chemigenetic calcium indicator for multiplexed in vivo functional imaging

Molecular biologySynthetic DNA oligonucleotides were purchased from Twist Bioscience and Integrated DNA Technologies. Q5 High-Fidelity DNA Polymerase (New England Biolabs) was used for all PCR amplifications. Isothermal assembly reactions were performed with a NEBuilder HiFi kit (New England Biolabs). Small-scale DNA isolation was performed with the QIAprep Spin Miniprep Kit (Qiagen). Vector backbones were acquired from the following sources: pRSET was acquired from Life Technologies; pGP-AAV-syn-jGCaMP7f-WPRE was a gift from D. Kim (GENIE project, Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA) and the GENIE project (Addgene plasmid 104488); pAAV-CaMKIIa-EGFP was a gift from B. Roth (Department of Pharmacology, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, NC, USA; Addgene plasmid 50469); p10xUAS-IVS-Syn21-Voltron-p10 was from a previously deposited Addgene plasmid (Addgene plasmid 119041); pTol2-HuC(elavl3)-CaMPARI2 was from...