Gene Editing in Channel Catfish via Double Electroporation of Zinc-Finger Nucleases

The traditional approach for gene editing with zinc-finger nucleases (ZFNs) in fish has been microinjection of mRNA. Here, we develop and describe an alternative protocol in which ZFN plasmids are electroporated to channel catfish, Ictalurus punctatus, sperm, and embryos. Briefly, plasmids were propagated to supply a sufficient quantity for electroporation. Sperm cells were prepared in saline solution, electroporated with plasmids, and then used for fertilization. Embryos were incubated with the plasmids before performing electroporation just prior to first cell division. Plasmids were then transcribed and translated by embryonic cells to produce ZFNs for gene editing, resulting in mutated fry.