One‐step generation of mice with gene editing by Tol2 transposon‐dependent gRNA delivery

Conditional knockout mice are valuable tools for examining the functions of targeted genes in a time‐ and space‐specific manner. Here, we generated gene‐edited mice by using the Tol2 transposon to introduce guide RNA (gRNA) into fertilized eggs obtained by crossing LSL (loxP‐stop‐loxP)‐CRISPR‐associated 9 (Cas9) mice, which express Cas9 in a Cre‐dependent manner, with CAG‐CreER mice. Transposase mRNA and plasmid DNA, which contained a gRNA sequence for the gene encoding tyrosinase flanked by the transposase recognition sequence, were injected together into fertilized eggs. As a result, the transcribed gRNA cleaved the target genome in a Cas9‐dependent manner. Using this method, it is possible to generate conditional genome‐edited mice more easily in a shorter period of time. Conditional knockout mice are valuable tools for exploring the functions of genes of interest. We generated gene‐editing mice by using Tol2 transposon to introduce guide RNA into fertilized eggs obtained by crossing LSL (loxP‐stop‐loxP)‐Cas9 mice with CAG‐CreER mice. Using this method, it is possible to generate conditional gene‐edited mice more easily in a shorter period of time.