ArticleProceedings of the National Academy of Sciences of the United States of America2016
A scalable strategy for high-throughput GFP tagging of endogenous human proteins.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 155 papers.
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Who cites it
155 citing papers in PubMed.
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- A toolkit for programmable transcriptional engineering across eukaryotic kingdoms.bioRxiv : the preprint server for biology · 2026Article
- Detecting the Activation of Endogenous Small GTPases via Fluorescent Signals Utilizing a Split mNeonGreen: Small GTPase ActIvitY ANalyzing (SAIYAN) System.Bio-protocol · 2026Article
- WormTagDB: A Systematic Survey of Endogenously Tagged Proteins inbioRxiv : the preprint server for biology · 2025Article
- Genome-scale CRISPR screens identify PTGES3 as a direct modulator of androgen receptor function in advanced prostate cancer.Nature genetics · 2025Article
- Regulators of androgen receptor activity revealed by CRISPR interference screens.Nature genetics · 2025Article
- Site-specific genome engineering of primary human natural killer cells for programmable anti-tumor function.bioRxiv : the preprint server for biology · 2025Article
- Lamin A/C Expression in Hematopoietic Cells Declines During Human Aging and Constrains Atherosclerosis in Mice.Arteriosclerosis, thrombosis, and vascular biology · 2025Article
- Programmable epigenome editing by transient delivery of CRISPR epigenome editor ribonucleoproteins.Nature communications · 2025Article
95 more citing papers are in PubMed but not listed here.
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5 authors.
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Abstract
A central challenge of the postgenomic era is to comprehensively characterize the cellular role of the ∼20,000 proteins encoded in the human genome. To systematically study protein function in a native cellular background, libraries of human cell lines expressing proteins tagged with a functional sequence at their endogenous loci would be very valuable. Here, using electroporation of Cas9 nuclease/single-guide RNA ribonucleoproteins and taking advantage of a split-GFP system, we describe a scalable method for the robust, scarless, and specific tagging of endogenous human genes with GFP. Our approach requires no molecular cloning and allows a large number of cell lines to be processed in parallel. We demonstrate the scalability of our method by targeting 48 human genes and show that the resulting GFP fluorescence correlates with protein expression levels. We next present how our protocols can be easily adapted for the tagging of a given target with GFP repeats, critically enabling the study of low-abundance proteins. Finally, we show that our GFP tagging approach allows the biochemical isolation of native protein complexes for proteomic studies. Taken together, our results pave the way for the large-scale generation of endogenously tagged human cell lines for the proteome-wide analysis of protein localization and interaction networks in a native cellular context.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.