Figure S2) CRISPR-Cas9 mediated editing of ache gene perturbs gut elongation. Xenopus embryos were injected with Cas9 mRNA/protein (A), ache gRNA (B), or Cas9 plus ache gRNA (C), and allowed to develop until stage 46. Relative to injection of Cas9 or ache gRNA alone (A, B), injection of Cas9 plus ache gRNA (C) disrupts intestinal lengthening. Intestine phenotypes resulted from injection of Cas9 plus ache gRNA at both the 1- and 8-cell stages; craniofacial and spinal deformities, similar to those observed with AChE-inhibitor exposure, were also observed in embryos injected at the 1-cell stage (not shown), though absent in 8-cell endoderm targeted injections. The graph (D) indicates the mean (± S.E.M) percentage of embryos in which the intestine is shortened (n=3 different experiments, both 1- and 8-cell injections). E) Genomic sequencing validates the efficacy of CRISPR-Cas9 mediated editing, revealing the presence of deleterious mutations in representative Cas9 plus ache gRNA-injected embryos. Scale bars = 1000 um.
Image published in: Pickett MA et al. (2017)
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