Fig. 4: P. canaliculata zygotes can be collected, micro-injected with exogenous mRNA and cultured ex ovo.

A Schematic representation of ex ovo culture protocol steps. Clutches are collected, crushed and centrifuged. The supernatant is collected and used to make drops where the embryos can be cultured. Everything is covered with paraffin oil to avoid evaporation. B Images of embryos cultured at 0 days post fertilization (dpf) and developing ex ovo. The images are representative of data collected through 6 independent experiments with more than 10 embryos in each experiment. C Schematic representation of the setup used for P. canaliculata zygote micro-injection (see Supplementary Fig. 5B, C). D Embryonic survival rate after micro-injection using different micro-injection protocols. Each dot represents an injection session with 50–100 embryos. E Percentage of embryos Dextran-positive, used as read-out of successfully injected embryos. Each dot represents an injection session with 50–100 embryos. F Confocal images of 3 dpf embryos injected with GFP mRNA or Lyn-tdTomato mRNA. GFP localizes in the cytoplasm, the membrane targeted tdTomato localizes in the cell membranes and the uninjected controls show some autofluorescence with a stereotypical localization in the red channel. Data are represented as mean ± SEM. The images are representative of data collected through two independent rounds of injections with more than 20 embryos each.