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. 2014 Oct 6;5:5042. doi: 10.1038/ncomms6042

Figure 2. Dax1-knockdown ESCs retain multilineage differentiation potential.

Figure 2

(a) qRT–PCR analysis of gene expression in the indicated lines after 9 days of EB differentiation. All data are normalized to Gapdh and shown relative to WT ESCs (set at 1.0). Data are represented as mean±s.d.; n=3. (b) Enhanced ExEn differentiation is observed in Dax1 KD ESCs. Monolayer cultures were treated with RA (0.1 μM) for 4 days and costained with Gata6 and DAPI. Scale bar, 100 μm. Proportions of Gata6+ cells are shown in the bar graph next to the images. Data are represented as mean±s.e.m. (n=3). *P<0.05. P values were calculated using Student’s t-test. (c) Flow cytometric analysis of mesoderm induction, as measured by the Flk1+ population, from Luc KD (control) and Dax1 KD ESCs. Numbers in quadrants indicate the percentage of each population. (d) Immunofluorescent analyses of neuroectoderm (Nestin, red) and TE (Cdx2, red) induction from Luc KD (control) and Dax1 KD ESCs. Scale bar, 100 μm. Proportions of Nestin+ cells and Cdx2+ cells are shown in the bar graph next to the images. Data are represented as mean±s.e.m. (n=3). (e) Neuroectoderm, mesoderm and ExEn-derived structures are present in teratomas from Dax1 KD ESCs. Hematoxylin and eosin-stained tissue sections are shown. Scale bar, 50 μm.