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. 2014 Feb 20;2(3):351–365. doi: 10.1016/j.stemcr.2014.01.005

Figure 2.

Figure 2

Characterization of iPSCs Generated from Tetracycline-Inducible Oct4 Cells

(A) Clonal induced pluripotent stem cell (iPSC) lines generated by tetracycline-induced Oct4 expression in combination with virally delivered Sox2, Klf4, and Myc. Phase-contrast overlay with Oct4-GFP reporter fluorescence and ALP staining. The scale bars represent 50 μm.

(B) Immunological stainings for NANOG and SSEA-1 (red), overlaid with nuclear Hoechst staining (blue). The scale bars represent 50 μm.

(C) DNA methylation of Oct4 and Nanog promoters in iPSCs and controls. Empty and full circles represent unmethylated and methylated CpG cytosine residues, respectively.

(D) Immunocytochemistry for specific marker proteins (red) showed in vitro differentiation of iPSC clones into all three germ layers. Nuclei were counterstained with Hoechst (blue). The scale bars represent 50 μm.

(E) PCR genotyping for the GFP transgene demonstrated that iPSCs could contribute to organs representative of all germ layers in 14.5 dpc chimeric embryos. Germline contribution was proven by the presence of Oct4-GFP-positive cells in the fetal gonads. The black scale bars represent 5 mm. The white scale bars represent 250 μm.

Abbreviations: TUBB3, β-III-tubulin (ectoderm; neurons); ACTA2, α-II-actin (mesoderm; smooth muscle cells); SOX17, SRY-box-containing 17 (definitive endoderm); br, brain; he, heart; li, liver. See also Tables S5 and S6.