Fig. 4

Expression of mutation reporters for chd7, hace1 and pycr1a mutations quantifies their frameshift efficiency. (A) Representative images of 16-18 hpf (∼16-somite stage) zebrafish embryos injected with the indicated mutation reporter mRNA from constructs made as described in Fig. 3 or uninjected. Fluorescence images of superfolder GFP (sfGFP) and TagRFP and merged images are shown. (B) Areas in the somite regions of these embryos were quantified by measuring fluorescence intensities of both sfGFP and TagRFP, adjusted for background in the corresponding channel, and the ratio of these intensities was calculated to account for injection variability and used for plotting the data and statistical analysis. For chd7 mutation reporters, the difference between wild-type (n=15) and 2-bp deletion mutant (n=12) mutation reporters was large and very significant (P=2.034e−15). Embryos injected with hace1 wild-type (n=10) and 8-bp insertion mutant (n=12) cDNA fragments did not show any significant difference. The pycr1a wild-type (n=41) and 71-bp deletion mutant (n=28) reporters had a small but significant difference (P=7.776e−13). ***P<0.001. The injections were performed twice on different days with very similar results and embryos from one of the injections were subjected to imaging and statistical analysis of fluorescent protein intensities ratios. Student's two-tailed t-test was performed for each experiment.