Fig. 1. Temporal dynamics of the primitive streak and mesoderm progenitors in β-catenin-activated hES cells. (A) The Nβ-cateninER cells were cultured in defined medium with or without 4OHT (days 0-3), and the representative morphology at the indicated time periods was shown. Upper or lower panels shows low or high magnification images, respectively. Scale bars: 100 µm. (B) Immunostaining analysis of the day-3 Nβ-cateninER-activated cells. Cells were cultured with or without 4OHT for 3 days, and subjected to immunostaining with indicated antibodies, and nuclei were counterstained with diaminopimelic acid (DAPI). Scale bar: 100 µm. (C) The Nβ-cateninER cells were cultured with (days 1-3) or without (day 0) 4OHT for the indicated time periods, and subjected to immunostaining with anti-E-cadherin and anti-N-cadherin antibodies. Nuclei were counterstained with DAPI. Scale bar: 100 µm. (D) Quantitative real-time PCR (qPCR) analysis on RNA isolated from undifferentiated Nβ-cateninER cells (day 0) and β-catenin-activated cells harvested at daily intervals (days 1-5) was performed using specific primers for the genes indicated. (E) Immunoblot analysis of cell lysates from Nβ-cateninER cells. Cells were cultured for the indicated time periods with 4OHT, and cell lysates were subjected to SDS-PAGE and probed with specific antibodies as indicated.