Figure 6

(A–D) Morphometric analysis of TL/Wt zebrafish embryos injected with nppcl/gRNA/Cas9 RNA. Zebrafish mutants co-injected with nppcl-gRNA and Cas9 RNA were reared alongside embryos injected with Cas9 RNA only and collected at 24 hpf, 48 hpf and 72 hpf, dechorionated and fixed with 4% (w/v) PFA. Embryos were subsequently mounted and 20x magnification phase-contrast images captured using an Axiovert 135 inverted microscope (Carl Zeiss Ltd., Cambridge, UK) and DC500 color camera. Phase-contrast images were Z-stacked using Adobe Photoshop (Adobe, USA) and morphometric measurements collected using ImageJ (http://imagej.nih.gov/in/). (A–C) Measurements were taken for: embryo length (n = 12), head width (n = 12) and eye diameter (n = 12). (D) Representative live images for were captured using a Nikon SMZ1500 stereomicroscope and a Nikon ds-2mv color camera, objective magnification was 10× and variable stage magnification at 3×. Data shown are box-and-whisker plots of medians (with min-max values), analyzed by a paired t-test (2-tailed); * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, significantly different from control.