ORANGE: A CRISPR/Cas9-based Genome Editing Toolbox for Epitope Tagging of Endogenous Proteins in Neurons

April 22, 2020 | Biology

This week's Lab.Equipment Biology news comes from Public Library of Science Biology:

by Jelmer Willems, Arthur P. H. de Jong, Nicky Scheefhals, Eline Mertens, Lisa A. E. Catsburg, Rogier B. Poorthuis, Fred de Winter, Joost Verhaagen, Frank J. Meye, Harold D. MacGillavry

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 revea... Read the whole article at the PLOS Biology Blog.

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