Generation of Zebrafi sh Models by CRISPR/Cas9 Genome Editing

Abstract

The CRISPR/Cas system identified in archaea has been adopted and optimized for genome editing purposes in zebrafi sh. In vitro transcribed guide RNA and Cas9 mRNA are microinjected into fertilized zebrafi sh embryos to edit the zebrafish genome. Here, we describe how to design a gRNA, a fast method for in vitro transcription of gRNA from oligonucleotides, microinjection into fertilized zebrafi sh embryos, and a PCR -based restriction fragment length assay to identify mutations at the gRNA target site.