Generation of Conditional Knockout Zebrafish Using an Invertible Gene-Trap Cassette

Conditional knockout (cKO) is a genetic technique to inactivate gene expression in specific tissues or cell types in a temporally regulated manner. cKO analysis is essential to investigate gene function while avoiding the confounding effects of global gene deletion. Genetic techniques enabling cKO analysis were developed in mice based on culturable embryonic stem cells that were not generally available in zebrafish, which hampered precise analysis of genetic mechanisms of organ development and regeneration. However, recent advances in genome editing technologies have resolved this limitation, providing a platform for the generation of cKO models in any organism. Here we describe a detailed protocol for the generation of cKO zebrafish using a Cre-dependent genetic switch.