Fig. 4.

Scoring metamorphosis defects in CRISPR larvae. (A) Example of a juvenile at 45 hpf in the “negative control” population, showing the retracted tail and body rotation that occurs during metamorphosis. Foxc>H2B::mCherry (red) labels the cells of the oral siphon and the papillae, the latter of which are transformed into the stolon of the juvenile. Nuclei counterstained by DAPI (blue). (B) Scoring of Foxc>H2B::mCherry+ individuals upon papilla-specific CRISPR/Cas9-mediated mutagenesis of the selected candidate genes. “Tailed juveniles” are individuals that have undergone body rotation but not tail retraction. Pou4 CRISPR served as the “positive control”, eliminating the papilla neurons that trigger metamorphosis (see text for citations). Of the four genes tested, only Vamp1/2/3 CRISPR appeared to result in substantial loss of tail retraction and body rotation, though not as penetrant as the Pou4 CRISPR. n=100 for each gene. (C) Representative example of Vamp1/2/3 CRISPR larva, showing intact tail at 45 hpf.