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STEM_31_sm_suppinformationfigure1.tif20091KSupporting Information Figure S1. In vivo differentiation demonstrates the pluripotency of human iPS cells. One million human iPS cells were injected into the testis of immunocompromised mice, and eight weeks later teratoma tumors were removed and subjected to histology by hematoxylin and eosin staining. Arrows indicate regions where cells typical of the indicated germ layers were found. Note that while human iPS cells always generated multipotent teratomas, injection of up to ten million parental fibroblasts never produced a single tumor of any kind (data not shown).
STEM_31_sm_suppinformationfigure2.tif1247KSupporting Information Figure S2. HSF1 and human iPS appear to have different propensities for neural specification. Differentiation of embryoid bodies (EBs) made from HSF1 and human iPS cells drives a portion of them down a neural lineage as measured by expression of Sox3. Quantification of the percentage of EBs with at least 90% of cells expressing Sox3 highlights a difference in neuralization between HSF1 and human iPS1 and 2 after 2 weeks of differentiation (* denotes p≤0.0003 for each human iPS cell line versus HSF1)(a). The reduced efficiency of human IPS cells in forming neural progenitors was similarly observed when three different lines (hiPS1, 2, and 18) along with HSF1 were directly differentiated into neural rosettes without EB formation. These results suggest that the differences in neural formation are not simply due to variations in EB formation by HSF1 and these human iPS cell lines. Two weeks after replacing the standard hESC maintenance media (KO serum replacer, Invitrogen) to neural/ventralization media (as described in the text, Fig 1), the number of colonies that displayed neural rosette formation was quantified as a percentage of the total (* denotes p≤0.0004 for each human iPS line versus HSF1)(b). Data were collected from at least two independent experiments and represent the average across an n ≥ 6. Error bars represent standard error of the mean. A two-tailed t-test was performed based on unequal variance to generate p values for each human iPS cell line versus HSF1.
STEM_31_sm_suppinformationfigure3.tif11013KSupporting Information Figure S3. HSF1, hiPS1, and hiPS18 all generate mature, electrophysiologically active motor neurons. Rosettes derived from HSF1, hiPS1, and hiPS18 were differentiated for at least 50 days and transfected with the Hb9::GFP reporter. Whole cell patch clamp recordings from Hb9::GFP expressing HSF1 and human iPS-derived cells (Right) show repetitive firing after stimulation (Left). Results shown are representative of recordings made from at least 7 cells derived from HSF1 and hiPS1 and hiPS18.

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