XB-IMG-81362
Xenbase Image ID: 81362
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Fig. 1. Maternal Dnd1 is vegetally localized and required for axis specification. (A) Western blot showing vegetal localization of endogenous maternal Dnd1 protein. Manually bisected animal and vegetal halves of oocytes were analyzed by immunoprecipitation/western blotting. Each group contains 50 animal or vegetal halves. β-tubulin served as the loading control. (B) Immunofluorescence showing colocalization of Dnd1 and Nanos1 in the germ plasm at the vegetal pole at the eight-cell stage (indicated by arrowheads). (C) Dnd1 knockdown efficiency. Oocytes were injected with the AS-oligo against dnd1, cultured for 24 hours, and treated with progesterone to induce germinal vesicle breakdown (GVBD). Uninjected controls and injected oocytes were harvested at 24 hours or 40 hours (after GVBD) after microinjection of the AS-oligo, and analyzed by immunoprecipitation/western blotting. Each group contains 50 oocytes or eggs. β-tubulin served as the loading control. (D,E) Morphology of uninjected controls (D; note rare examples of class II) and AS-oligo-injected host-transfer embryos (E) at the tadpole stage. (F) Summary of the phenotypes of control and AS-oligo-injected host-transfer embryos. (G,H) Cross-section of uninjected (G) and AS-oligo-injected (H) host-transfer embryos at the tadpole stage. H represents the most severe ventralization phenotype. (I) RT-PCR results showing effects of Dnd1 knockdown on the expression of NCAM, type II collagen (col II), muscle actin (m-actin), epidermal keratin (e-keratin), alas and α-globin at stage 20. (J) Real-time RT-PCR showing the expression of Wnt target genes (Xnr3 and siamois), organizer-specific genes (noggin and chordin), and ventral markers (msx1, sizzled, vent1 and vent2), and dnd1 in control and AS-oligo-injected host-transfer embryos (stage 10). All samples were normalized to odc levels. (K) Whole-mount in situ hybridization showing the expression of chordin and vent2 in control and AS-oligo-injected host-transfer embryos at stage 10.5. Arrows point to the dorsalmost region of control embryos, a region positive for chordin expression, but negative for vent2 expression. Image published in: Mei W et al. (2013) Copyright © 2013. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License.
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