Photoswitching Proteins

This volume provides cutting edge protocols for using optogenetic and opto-chemical techniques across a wide spectrum of applications. Chapters guide readers through methods in neuro-optogenetics, including methods to dissect local and long-range connectivity ex vivo, investigate G-protein coupled receptors, study astrocyte-neuron networks in vitro and in vivo, employ upconversion nanoparticles for transcranial optogenetic stimulation, and use Adeno-associated virus (AAV) for delivery of optogenetic constructs. At the same time, this book covers a range of methods for non-neuronal optogenetics, including various applications of light-oxygen-voltage domains or cryptochromes for control of nucleocytoplasmic transport, transgene expression and chromatin recruitment. The book further presents protocols for traceless light-control of gene expression with melanopsin, light-inducible cAMP production, use of light-regulated receptor tyrosine kinases (Opto-RTKs), synthesis of light-dependent biomaterials, and highlights emerging tools at the intersection of optogenetics and CRISPR. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Authoritative and cutting-edge, Photoswitching Proteins: Methods and Protocols aims to provide a comprehensive list of methods and tools to investigate highly dynamic pathways and behaviors in cells and animals using optogenetics.

Includes cutting-edge methods and protocols Provides step-by-step detail essential for reproducible results Contains key notes and implementation advice from the experts

Inhalt

Studying Neuronal Function Ex Vivo using Optogenetic Stimulation and Patch Clamp.- Optogenetic Techniques for Probing G Protein-Coupled Receptor Signaling.- Melanopsin for Time-Controlling Activation of Astrocyte-Neuron Networks.- Near-infrared Deep Brain Stimulation in Living Mice.- Lab-scale Production of Recombinant Adeno-associated Viruses (AAV) for Expression of Optogenetic Elements.- AAV-mediated Gene Delivery to Foveal Cones.- Engineering Optogenetic Protein Analogs.- Optogenetic Control of Nucleocytoplasmic Protein Transport.- CRISPR Labeling Light-inducible.- Optogenetic Control of Gene Expression using Cryptochrome 2 and a Light-activated Degron.- Optogenetic Downregulation of Protein Levels to Control Programmed Cell Death in Mammalian Cells with a Dual Blue-Light Switch.- Tracing Reversible Light-induced Chromatin Binding with Near-infrared Fluorescent Proteins.- Construction of a Multiwell Light-induction Platform for Traceless Control of Gene Expression in Mammalian Cells.- Dual-activation of cAMP Production through Photo-Stimulation or Chemical Stimulation.- Synthesis of a Light-controlled Phytochrome-based Extracellular Matrix with Reversibly Adjustable Mechanical Properties.- Design and Application of Light-regulated Receptor Tyrosine Kinases.- All-optical Miniaturized Co-culture Assay of Voltage-gated Ca2+ Channels.- Optogenetics and CRISPR: A New Relationship Built to Last.