ZFN-induced mutagenesis at the cry2 locus in somatic cells. (A) Effects of ZFN mRNA injections on monarch embryos. The bar graph depicts the rate of hatching larvae (gray bars) and of embryos presenting development but dying before hatching (white bars), associated with egg injection of each of the ZFNs mRNA alone (linked to either of the obligate heterodimeric versions of the FokI nuclease domain) or both ZFNs mRNA at 0.5 μg/μL. For comparison, control embryos injected with water in similar conditions hatch at a rate of 19.5% with an additional 3.6% that develop but die before hatching (n = 82). Uninjected embryos hatch at a rate of ∼75%. (B) Agarose gel example of lesions observed in somatic cells of hatched larvae and dead embryos injected with both ZFN mRNAs, showing a high degree of targeting (green arrow), compared with an uninjected control. (C) Mutations observed in somatic cells of dead embryos, and dead or live larvae, from eggs injected at different doses of ZFN mRNAs.
