Instability of genomic methylation patterns in human embryonic stem (ES) and induced pluripotent stem (iPS) cells. DNA was analyzed for demethylation at the sequences named on the left. Controls were the human mammary carcinoma cell line Hs578T, which has a severely demethylated genome, and the MDA175 cell line, which is not detectably demethylated. Note that all human ES and iPS cells showed demethylation of a satellite relative to MDA175 and that trypsin-passaged H9 and CHB1 human ES and D10 cells and high-passage 1774-2 iPS cells showed demethylation of LINE1 promoters and satellite 3 DNA. All human ES and iPS lines were maintained under the same cell culture conditions, without the use of trypsin except in the case of the indicated H9 cell line, for five or more passages.