CRISPR/Cas9-mediated genome modification in the mollusc, Crepidula fornicata.

Abstract

The discovery and application of the CRISPR/Cas9 genome editing method has greatly enhanced the ease with which transgenic manipulation can occur. We applied this technology to the mollusc, Crepidula fornicata, and have successfully created transgenic embryos expressing mCherry fused to endogenous β-catenin. Specific integration of the fluorescent reporter was achieved by homologous recombination with a β-catenin-specific donor DNA containing the mCherry coding sequence. This fluorescent gene knock-in strategy permits in vivo observations of β-catenin expression during embryonic development and represents the first demonstration of CRISPR/Cas9-mediated transgenesis in the Lophotrochozoa superphylum. The CRISPR/Cas9 method is a powerful and economical tool for genome modification and presents an option for analysis of gene expression in not only major model systems, but also in those more diverse species that may not have been amenable to the classic methods of transgenesis. This approach will allow one to generate transgenic lines of snails for future studies.