Fig. 1

Organ left-right patterning defects in embryos injected with cdon MO at the 4-cell stage (A) The expression of cdon was examined using whole mount in situ hybridization from 128-cell stage to 24hpf. Cdon was expressed at 128-cell stage (Aa1). White arrowhead showed the expression of cdon in DFCs at 80% epiboly stage (Aa2). Black arrowhead showed the expression of cdon in DFCs at bud stage (Aa3-a4). Cdon was also expressed in midline and presumptive neural crest (Aa5). Black arrow head showed the expression of cdon in epithelial cell in KV at 6ss (Aa6-a7). Expression of cdon was examined at 24 hpf (Aa8). (B) Embryos injected with cdon MO displayed liver LR defects. b1, normal liver in Tg (fabp10:GFP) transgenic controls (93.1%, n = 102); b2, normal liver in embryos injected with cdon MO (60.8%, n = 79); b3, liver bifida in embryos injected with cdon MO (25.3%, n = 79); b4, reversed liver in embryos injected with cdon MO (13.9%, n = 79). b5, normal liver in wild type controls (93.3%, n = 75); b6-b8, embryos injected with cdon MO were examined at 4dpf using in situ experiments (n = 84). (C) Percentages of normal liver, liver bifida, and reversed liver in embryos used as control, embryos injected with cdon MO and embryos co-injected with cdon MO and cdon mRNA, in respectively. Embryos injected with cdon MO show a statistically significant difference compared to controls, and embryos co-injected with cdon MO and cdon mRNA show a statistically significant difference compared to those injected with cdon MO alone. (D) Embryos injected with cdon MO displayed heart LR defects. d1, normal-loop in Tg(cmlc2:GFP) transgenic controls (97.4%, n = 77); d2, normal-loop in embryos injected with cdon MO (56.8%, n = 95); d3, no loop in embryos injected with cdon MO (16.8%, n = 95); d4, reversed-loop in embryos injected with cdon MO (26.4%, n = 95); d5, normal-loop in wild type controls (94.5%, n = 73); d6-d8, wild-type embryos injected with cdon MO at the 4-cell stage were examined for cardiac looping at 72 hpf by WISH against my17 (n = 83). The blue dashed line indicates the atrial edge. (E) Percentages of normal looping, no looping, and reversed looping of the heart in embryos being as control, embryos injected with cdon MO, and embryos co-injcted with cdon MO and cdon mRNA. Cdon mRNA injection partially rescued the heart LR defect in embryos injected with cdon MO (E, the right column showing). Embryos injected with cdon MO show a statistically significant difference compared to controls, and embryos co-injected with cdon MO and cdon mRNA show a statistically significant difference compared to those injected with cdon MO alone. Notice: in ?C? and ?E,? all the transgenic embryos and wild type embryos were used together to calculate the percentage. Statistical analysis was performed using Student?s t-test. ?*?p < 0.05, ?**? p < 0.01, ?***? p < 0.001. Notice: ?control? refers to wild-type embryos that were not injected with cdon MO.