We stained H1 ES cells and H1-derived iN cells at 21 days after induction for EGFP (to identify cells with viral transduction), the stem-cell markers Sox2, Oct4, and Nanog, and the glial cell marker GFAP (to mark mouse astrocytes added to the iN cell culture for promotion of synapse formation). At the level of immunolabeling, expression of stem-cell markers was abolished in iN cells, consistent with a conversion of H1 ES cells into iN cells (Fig. 2A). Quantitative RT-PCR analyses revealed that iN cells expressed increased levels of endogenous Ngn2 as well as of two neuronal markers, NeuN and MAP2, whose levels were elevated ~100-fold (Fig. 2B). In addition, we observed an even larger induction of the expression of the transcription factors Brn2 and FoxG1, which are markers for excitatory cortical neurons (Fig. 2B).
