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Fig. 4 Homozygous col22a1−/− embryos show increased percentage of hemorrhages when raised at increased temperature. Embryos were stained with o-Dianisidine at 3 dpf. (A) The heterozygous col22a1+/− embryo shows small scattered blood cells in the head, while a large hemorrhage is apparent within the brain in the col22a1−/− embryo (arrow, B). (C) Compared with the WT and col22a1+/− embryos, a significantly higher percentage of col22a1−/− mutant embryos exhibit hemorrhages after heat stress. *P<0.005, determined by chi-square test. Embryo numbers were combined from three independent experiments (two for col22a1+/− embryos). The scoring numbers from each experiment are shown in Table S1. (D-F) Examples of hemorrhages (arrows) at different locations in heat-stressed col22a1−/− embryos.