Leonetti, M.D. et al. Proc. Natl. Acad. Sci. USA 113, E3501–E3508 (2016).

Over ten years ago, Erin O'Shea and colleagues tagged 75% of the yeast proteome with GFP, allowing for image-based analysis of protein localization on a global scale. Now, Leonetti et al. describe a CRISPR/Cas9-based method that will facilitate GFP tagging of the roughly 20,000 proteins encoded in the human genome. In their method, purified Cas9 and a desired single guide RNA are electroporated into cells along with a double-stranded DNA that encodes a short fragment of GFP and flanking sequence homologous to the target region. Cas9 editing at the desired target site leads to integration of this DNA and tagging of the protein of interest. The cells themselves stably express the complementary portion of GFP, allowing for formation of fluorescent GFP on the tagged protein. Using this strategy, the team successfully tagged 48 human genes.