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Figure 3. Mesodermal Conversion Induced by XFDL Knockdown in the Animal Cap(A–J) In vivo effects of XFDL-MO. Control embryos (A, E, G, and I) or embryos given 50 ng/cell of XFDL-MO (25 ng/cell of XFDL156-MO + 25 ng/cell of XFDL141-MO for the short minor splicing variant; see Supplemental Experimental Procedures; B, F, H, and J), two five-base-mismatched MOs (25 ng/cell of each; C), or 50 ng/cell of XFDL-MO + XFDL mRNA (100 pg/cell; D) by injection were harvested at the early gastrula stage and analyzed by in situ hybridization using the indicated probes. Sectioned embryos are shown in (E) and (F). Brackets (in panels A and B) and arrowheads (in panels E and F) indicate the expression areas of Xbra and VegT, respectively.(K) q-PCR analysis for the expression of mesodermal marker genes. XFDL-MO (50 ng/cell) was injected at the 8-cell stage. Animal caps were prepared at stage 8.5, cultured until siblings reached stage 10.5, and analyzed by q-PCR.(L) Chordin-induced neural differentiation was attenuated by XFDL-MO in the animal cap. Control (10 pg/cell; lane 1) or Chordin (10 pg/cell; lanes 2–4) mRNAs were injected with 12.5 ng/cell (lane 3) or 50 ng/cell (lane 4) of XFDL-MO. Explants were prepared at stage 8.5 and harvested at stage 11.5. Sox2 expression was analyzed by q-PCR. In (K) and (L), the expression of each gene in the control animal cap was defined as 1. |
