PEGG: A computational pipeline for rapid design of prime editing guide RNAs and sensor libraries

Abstract

AbstractPrime editing leverages Cas proteins fused to reverse transcription enzymes and prime editing guide RNAs (pegRNAs) to programmably engineer diverse genetic alterations without DNA double strand breaks or exogenous donor templates. Prime editing is powerful, but the process of pegRNA design remains a challenge that limits its widespread adoption by the community. Although a number of computational pegRNA design tools have been developed, their capability for rapid and systematic design of pegRNAs for variant engineering and high-throughput genetic screens is limited. Here, we describePrimeEditingGuideGenerator (PEGG): a computational pipeline for rapid design of pegRNAs and paired ‘sensor’ pegRNA libraries. PEGG is a user-friendly Python package that generates and visualizes pegRNA designs for installing desired mutations while integrating user-defined properties like variable length of primer binding site and reverse transcription template regions, and outputs ready-to-order pegRNA oligos and libraries containing optimized sequence adapters for pooled cloning. PEGG (including documentation) can be accessed athttps://pegg.readthedocs.io.