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Updated: Jun 17, 2025
Efficient Genome Editing of Mice by CRISPR Electroporation of Zygotes
Published on: December 16, 2022
Distance-dependent effects on CRISPR/Cas9-mediated genome editing in Schizosaccharomyces pombe compromise efficiency
Reine U Protacio1, Emory G Malone1, Wayne P Wahls1
1Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States.
Abstract:
Discrete DNA sites position meiotic recombination at hotspots. We sought to create four different, 15 bp long, candidate regulatory DNA sites within the ura4 reporter gene. Each effort employed a fission yeast-optimized CRISPR system (SpEDIT), optimal guide RNA, and one of four homologous recombination templates with 10 to 15 bp substitutions. Remarkably, every Ura - transformant analyzed had template-directed, PAM-disabling bp substitutions near (5-6 bp away from) the DSB but no DNA site-generating substitutions at distance (42-56 bp). An unsought novel allele, ura4-P127* , has two substitutions (C379T, C380A) that create a stop codon, rendering strains unable to grow without uracil.
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01:59CRISPR
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