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Fig. 1. ascl1 is a maternal gene in Xenopus embryos and ectopic ASCL1 represses mesendoderm formation. (A) Expression of ascl1 at different stages of Xenopus development as analysed by qPCR and represented as the relative expression levels (means±s.d.) to that at stage 10 after being normalized to odc. (B-E) Whole-mount in situ hybridization for ascl1 in oocytes and embryos at maternal stages of development. (B) Bisection view of a full-grown oocyte at stage (st) VI. GV, germinal vesicle. (C) Bisection view of a full-grown st VI oocyte in situ hybridized with ascl1 sense probe. (D,E) Animal (D) or lateral (E) view of an embryo at the mid-blastula stage (st8) in situ hybridized with ascl1. (F) RT-PCR analysis of the relative distribution of ascl1, vegt and pou5f3.3 along the animal-vegetal axis of blastulae at stage 8.5, as indicated in E. AC, animal cap; MZ, marginal zone; VM, vegetal mass. (G) qPCR comparison of gene expression in animal cap explants injected with VegT (300 pg) and mRNA encoding β-gal or Ascl1 (500 pg) in the animal pole at the two-cell stage. Animal caps were dissected at stage 8.5 followed by qPCR analysis at the sibling stage 10.5. The expression level of each individual gene (mean±s.d.) was normalized to odc. (H) ascl1 (500 pg) was injected into the vegetal pole, followed by injection of Xnr1 (100 pg). Expression of mesendoderm genes was analysed by WISH at stage 10.5 (vegetal view). Scale bar: 1 mm. |
