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The correct subcellular distribution of proteins establishes the complex morphology and function of neurons. Fluorescence microscopy techniques are invaluable to investigate subcellular protein distribution, but they suffer from the limited ability to efficiently and reliably label endogenous proteins with fluorescent probes. We developed ORANGE: Open Resource for the Application of Neuronal Genome Editing, which mediates targeted genomic integration of epitope tags in rodent dissociated neuronal culture, in organotypic slices, and in vivo. ORANGE includes a knock-in library for in-depth investigation of endogenous protein distribution, viral vectors, and a detailed two-step cloning protocol to develop knock-ins for novel targets. Using ORANGE with (live-cell) superresolution microscopy, we revealed the dynamic nanoscale organization of endogenous neurotransmitter receptors and synaptic scaffolding proteins, as well as previously uncharacterized proteins. Finally, we developed a mechanism to create multiple knock-ins in neurons, mediating multiplex imaging of endogenous proteins. Thus, ORANGE enables quantification of expression, distribution, and dynamics for virtually any protein in neurons at nanoscale resolution.
Pubmed ID: 32275651 RIS Download
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Multi paradigm numerical computing environment and fourth generation programming language developed by MathWorks. Allows matrix manipulations, plotting of functions and data, implementation of algorithms, creation of user interfaces, and interfacing with programs written in other languages, including C, C++, Java, Fortran and Python. Used to explore and visualize ideas and collaborate across disciplines including signal and image processing, communications, control systems, and computational finance.
View all literature mentionsNon-profit plasmid repository dedicated to helping scientists around the world share high-quality plasmids. Facilitates archiving and distributing DNA-based research reagents and associated data to scientists worldwide. Repository contains over 65,000 plasmids, including special collections on CRISPR, fluorescent proteins, and ready-to-use viral preparations. There is no cost for scientists to deposit plasmids, which saves time and money associated with shipping plasmids themselves. All plasmids are fully sequenced for validation and sequencing data is openly available. We handle the appropriate Material Transfer Agreements (MTA) with institutions, facilitating open exchange and offering intellectual property and liability protection for depositing scientists. Furthermore, we curate free educational resources for the scientific community including a blog, eBooks, video protocols, and detailed molecular biology resources.
View all literature mentionsSoftware package as distribution of ImageJ and ImageJ2 together with Java, Java3D and plugins organized into coherent menu structure. Used to assist research in life sciences.
View all literature mentionsSoftware tool for automated microscope acquisition, device control, and image analysis. Used for integrating dissimilar fluorescent microscope hardware and peripherals into a single custom workstation, while providing all the tools needed to perform analysis of acquired images. Offers user friendly application modules for analysis such as cell signaling, cell counting, and protein expression.
View all literature mentionsA national mouse monoclonal antibody generating resource for biochemical and immunohistochemical applications in mammalian brain. NeuroMabs are generated from mice immunized with synthetic and recombinant immunogens corresponding to components of the neuronal proteome as predicted from genomic and other large-scale cloning efforts. Comprehensive biochemical and immunohistochemical analyses of human, primate and non-primate mammalian brain are incorporated into the initial NeuroMab screening procedure. This yields a subset of mouse mAbs that are optimized for use in brain (i.e. NeuroMabs): for immunocytochemical-based imaging studies of protein localization in adult, developing and pathological brain samples, for biochemical analyses of subunit composition and post-translational modifications of native brain proteins, and for proteomic analyses of native brain protein networks. The NeuroMab facility was initially funded with a five-year U24 cooperative grant from NINDS and NIMH. The initial goal of the facility for this funding period is to generate a library of novel NeuroMabs against neuronal proteins, initially focusing on membrane proteins (receptors/channels/transporters), synaptic proteins, other neuronal signaling molecules, and proteins with established links to disease states. The scope of the facility was expanded with supplements from the NIH Blueprint for Neuroscience Research to include neurodevelopmental targets, the NIH Roadmap for Medical Research to include epigenetics targets, and NIH Office of Rare Diseases Research to include rare disease targets. These NeuroMabs will then be produced on a large scale and made available to the neuroscience research community on an inexpensive basis as tissue culture supernatants or purified immunoglobulin by Antibodies Inc. The UC Davis/NIH NeuroMab Facility makes NeuroMabs available directly to end users and is unable to accommodate sales to distributors for third party distribution. Note, NeuroMab antibodies are now offered through antibodiesinc.
View all literature mentionsCommercial vendor and service provider of laboratory reagents and antibodies. Supplier of scientific instrumentation, reagents and consumables, and software services.
View all literature mentionsAmerican chemical, life science and biotechnology company owned by Merck KGaA. Merger of Sigma Chemical Company and Aldrich Chemical Company. Provides organic and inorganic chemicals, building blocks, reagents, advanced materials and stable isotopes for chemical synthesis, medicinal chemistry and materials science, antibiotics, buffers, carbohydrates, enzymes, forensic tools, hematology and histology, nucleotides, proteins, peptides, amino acids and their derivatives.
View all literature mentionsVector graphics software to create digital graphics, illustrations, and typography for several types of media: print, web, interactive, video, and mobile.
View all literature mentionsCell line HEK293T is a Transformed cell line with a species of origin Homo sapiens (Human)
View all literature mentionsThis monoclonal targets alpha-Tubulin antibody produced in mouse
View all literature mentionsThis monoclonal targets Bassoon mAb (SAP7F407)
View all literature mentionsThis monoclonal targets HA
View all literature mentionsThis monoclonal targets FLAG
View all literature mentionsThis monoclonal targets PSD 95
View all literature mentionsThis monoclonal targets Shank2
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