Protocol for CRISPR-based endogenous protein tagging in mammalian cells
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Updated: Jun 12, 2025
Endogenous Protein Tagging in Human Induced Pluripotent Stem Cells Using CRISPR/Cas9
Published on: August 25, 2018
Yu-Xi Xiao1, Jiarun Wei1, Jason Moffat2
1Program in Genetics and Genome Biology, The Hospital for Sick Children, Toronto, ON, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada.
Abstract:
Tracking the localization and proximal interaction partners of endogenous proteins provides valuable functional insight. Here, we present a protocol for CRISPR-based endogenous protein tagging in mammalian cells. We describe steps for endogenously tagging human TSC22D2 and MAP4, including designing Cas9 and Cas12a guides for knockin, modularized repair template design and cloning, and procedures for lipid transfection and electroporation. This protocol accommodates Cas nucleases in plasmid expression or ribonucleoprotein complex (RNP) formats. This "endo-tagging" approach offers flexibility and broad applicability. For complete details on the use and execution of this protocol, please refer to Xiao et al.1.
