Fig. 1

Genetic targeting ofsox2andsox3. (A) Sequences for targeting vectors for sox2 (TALEN) and sox3 (sgRNA) and resulting lesions in sox2 (allele x50) and sox3 (alleles x52 and x53). The deletion in x50 leads to loss of an SfoI restriction site (magenta), which was used for genotyping, whereas sox3 indels were identified by allele-specific PCR primers (see Materials&Methods). (B-L) Live embryos at 26 hpf showing general morphology (B-D), cranial development (E-G), tail development (K-L) and the otic vesicle (H-J, lateral views with anterior to the left; Scale bar, 50 µm) in control, sox2-/- and sox3-/- mutants. Excess cells at the ventral tip of the tail in sox2-/- embryos are indicated (arrow). (M) Box-and-whisker plot of surface area of otic vesicle, normalized to control embryos, in control embryos and sox2-/- and sox3-/- mutants. Green line indicates mean. Asterisk indicates statistically significant difference relative to control embryos (*** P<0.001, Tukey's HSD test). (N) Quantitative real time PCR measurements of fold changes in sox2 and sox3 mRNA levels in sox2-/- and sox3-/- mutants at 36 hpf normalized to wild-type control embryos. Error bars represent standard error of the means. Asterisks indicate statistically significant differences relative to controls (* P<0.05, t-tests).