**8. Expression levels of GALK1, GALK2, and OTC**

Human fetal and adult hepatocytes express galactokinase and OTC, and would survive in HSM containing galactose and ornithine. If hiPS cells express similar levels of these enzymes, HSM could not be applied for selection of differentiated hepatocytes. Therefore, we compared the expression levels of galactokinase and OTC in hiPS cells with those in human fetal and adult livers. The hiPS cell line 201B7 (RIKEN Cell Bank, Tsukuba, Japan) was cultured feederfree in ReproFF medium (Reprocell, Yokohama, Japan) in dishes coated with a thin layer of Matrigel (Becton Dickinson, Franklin Lakes, NJ). Two galactokinase isoforms, GALK1 (GenBank: NM\_000154) and GALK2 (BC107153), have been identified in humans. The expression levels of GALK1, GALK2, and OTC in the 201B7 cells and fetal and adult livers were compared [6]. The expression levels of these enzymes in the 201B7 cells constituted 22.2% ± 5.0%, 14.2% ± 1.1%, and 1.2% ± 0.2% (mean ± standard deviation) of those in the adult liver, respectively, and the OTC expression was also significantly lower in the 201B7 cells than in the fetal liver. We then cultured 201B7 cells in HSM to assess their survival rates.
