Fig. 5

Effects of ZFcerkl silencing in eye development.

(A) Two morpholinos targeting an acceptor (acMO) and donor splice site (doMO) of ZFcerkl were used. The morpholino targeting site is depicted by a discontinuous line. To assess the knockdown effect, a RT-PCR analysis of control and ZFcerkl morphants injected with 8 ng of MO was performed (primers used are depicted with arrows). Silencing with acMO was almost complete, when compared with cerkl expression in 72 hpf control MO-injected embryos (coMO), whereas in doMO animals the WT spliced isoform was decreased by 35%. β-Actin was used for normalization. (B) acMO-injected embryos displayed either small eye phenotype (named mild phenotype) or very small eye, small head, and body and curved tail (named severe phenotype). White double-head arrows denote diameter of the eyes. * denotes small and curved head in severe-phenotype morphants. (C) Phenotype frequency of morphants. The cerkl-knockdown phenotype was rescued when human CERKL mRNA was co-injected with acMO. n, number of individuals. (D) Eye size (in diameter) was measured in at least ten independent embryos from each group. Data were analyzed by t-test and are presented as mean ± SEM. ** P<0.001. Mean eye size was 253.4 µm for control, 172.7 μm for acMO and 239 μm for doMO morphants.