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Induced pluripotent stem cells hold great promise for regenerative medicine but point mutations have been identified in these cells and have raised serious concerns about their safe use. We generated nucleartransfer ES cells (ntESCs) from both MEFs and tail-tip fibroblasts (TTFs) and by whole genome sequencing found fewer mutations compared with iPSCs generated by retroviral gene transduction. Furthermore, TTF-derived ntESCs showed only a very small number of point mutations, approximately 80% less than the number observed in iPSCs generated using retrovirus. Base substitution profile analysis confirmed this greatly reduced number of point mutations. The point mutations in iPS cells are therefore not a Yamanaka factor-specific phenomenon but are intrinsic to genome reprogramming. Moreover, the dramatic reduction in point mutations in ntESCs suggests that most are not essential for genome reprogramming. Our results suggest that it is feasible to reduce the point mutation frequency in induced pluripotent stem cells by optimizing various genome reprogramming conditions. STEM CELLS 2017; 00:000–000 SIGNIFICANCE STATEMENT iPS cells hold great promise for regenerative medicine but point mutations have been identified in these cells and have raised serious concerns about their safe use. We conducted whole genome sequencing of nucleartransferred ES cells derived from MEFs or tail-tip fibroblasts (TTFs). We thereby succeeded in establishing ntES cell lines with far fewer point mutations. Our present results thus show that point mutations in iPS cells are therefore not a Yamanaka factor-specific phenomenon but are intrinsic to genome reprogramming. Importantly, our results suggest that a greatly reduced point mutation frequency in induced pluripotent stem cells is feasible by optimizing various genome reprogramming conditions. Whole genome sequencing of ntES cells www.StemCells.com ©AlphaMed Press 2017 2