High-throughput optimized prime editing mediated endogenous protein tagging for pooled imaging of protein localization.

ABSTRACT: The subcellular organization of proteins carries important information on cellular state and gene function, yet currently there are no technologies that enable accurate measurement of subcellular protein localizations at scale. Here we develop an approach for pooled endogenous protein tagging using prime editing, which coupled with an optical readout and sequencing, provides a snapshot of proteome organization in a manner akin to perturbation-based CRISPR screens. We constructed a pooled library of 17,280 pegRNAs designed to exhaustively tag 60 endogenous proteins spanning diverse localization patterns and explore a large space of genomic and pegRNA design parameters. Pooled measurements of tagging efficiency uncovered both genomic and pegRNA features associated with increased efficiency,

SUBMITTER: Sanchez HM 

PROVIDER: S-EPMC11429766 | biostudies-literature | 2024 Sep

REPOSITORIES: biostudies-literature