Fig. S1

Fig. S1 (A) Targeting efficiency of Sox 7 and Sox18 morpholinos. Messenger RNA synthesized from Sox7 and Sox18 5′UTR–GFP fusion constructs was injected in the embryos and re-injected afterwards with the respective morpholino. mRNA and morpholinos were not coinjected simultaneously, as in the test tube, the binding of the morpholino to its target is greatly favored compared to the in vivo situation in the embryo. If this binding is not totally and quickly reversible in the embryo, injecting together would lead to a reduction of the fluorescence which does not reflect the targeting efficiency of the morpholino in vivo. In all situation, GFP expression was totally abolished when MO1 Sox7, MO1 Sox18 or MO2 Sox18 were injected at a concentration of 4 ng/egg with 300 pg of the corresponding gene's 5′UTR–GFP expression construct. (B) Targeting efficiency of MOE2i2 Sox7 morpholino. Splice-site-targeted SOX7 morpholino alters splicing in zebrafish. Upper part: Genomic structure of zebrafish SOX7 gene. The ORF is indicated by white boxes. Splice site targeted by e2i2 is shown as well as the location of the primers used for the PCR. Lower part: Reverse transcriptase polymerase chain reaction (RT-PCR) analysis of Sox7 mRNA structure in control (lane 2) and e2i2 morphants (lane 3) using BP458 and O76. In the e2i2 morphants, we can observe a reduction of intensity of the band at 285 pb corresponding to the spliced transcript together with the apparition of a 1359 bp band corresponding to a non-spliced transcript containing the second intron. Lane 1 is the Smart ladder of Eurogentec (Belgium).