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Fig. 1 (A) Schematic depicting the mutagenesis systems. Cas9 protein expressed from the buc (oocytes) or ziwi (all germ cells) promoter. Target-specific guides are expressed from the U6 promoter, and GFP is expressed from the ziwi promoter to mark the germline. Mutagenesis system vectors express Cas9 as a single transcript with GFP following a T2A cleavage signal. In addition, GFP is expressed from an independent heart promoter, cardiac myosin light chain 2 (cmlc2), to permit earlier selection of transgenic animals. (B) Schematic of vectors and gateway-mediated recombination to generate OMS and GMS mutagenesis plasmids. (C) Schematic of mutagenesis strategy.