Highly Efficient, Rapid and Co-CRISPR-Independent Genome Editing in Caenorhabditis elegans

Abstract

We describe a rapid and highly efficient method to generate point mutations in Caenorhabditis elegans using direct injection of CRISPR-Cas9 ribonucleoproteins. This versatile method does not require sensitized genetic backgrounds or co-CRISPR selection-based methods, and represents a single strategy that can be used for creating genomic point mutations, regardless of location. As proof of principle, we show that knock-in mutants more faithfully report variant-associated phenotypes as compared to transgenic overexpression. Data for nine knock-in mutants across five genes are presented that demonstrate high editing efficiencies (60%), a reduced screening workload (24 F1 progeny), and a rapid timescale (4–5 d). This optimized method simplifies genome engineering and is readily adaptable to other model systems.

Keywords

• CRISPR
• Cas9
• ribonucleoprotein
• Caenorhabditis elegans
• disease variants