Abstract
Editing the Drosophila genome is incredibly useful for gene functional analysis. However, compared to gene knockouts, precise gene editing is difficult to achieve. Prime editing, a recently described CRISPR/Cas9-based technique, has the potential to make precise editing simpler and faster, and produce less errors than traditional methods. Initially described in mammalian cells, prime editing is functional in Drosophila somatic and germ cells. Here, we outline steps to design, generate, and express prime editing components in transgenic flies. Furthermore, we highlight a crossing scheme to produce edited fly stocks in less than 3 months.
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Acknowledgments
The authors thank Gabriel Birchak for discussions and S2R+ work, Rich Binari for general lab assistance and help with fly, the TRiP and Drosophila RNAi Screening Center for help generating transgenic flies, Ram Viswanatha for general discussions, Gillian Millburn for discussions on pegRNA transgene nomenclature, Cathryn Murphy for general lab assistance, Cooper Cavers for help isolating transgenic flies, Jorden Rabasco for help with molecular cloning, Andrew Anzalone for advice with synthetic pegRNAs, and Ben Ewen-Campen, Jonathan Zirin, and Thai LaGraff for cloning help. J.A.B. was supported by the Damon Runyon Foundation (DRG-2258-16) and a “Training Grant in Genetics” T32 Ruth Kirschstein-National Research Service Award institutional research training grant funded through the NIH/National Institute of General Medical Sciences (T32GM007748). This work was also supported by NIH Grant P41GM132087. N.P. is an Investigator of the Howard Hughes Medical Institute.
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Bosch, J.A., Perrimon, N. (2022). Prime Editing for Precise Genome Engineering in Drosophila. In: Dahmann, C. (eds) Drosophila. Methods in Molecular Biology, vol 2540. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2541-5_5
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DOI: https://doi.org/10.1007/978-1-0716-2541-5_5
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