Abstract: For utilization of the mouse embryonic stem (ES) cells for various purposes,it is desirable that the cell lines are established from various sources such as inbred and outbred mouse strains. So far,however,most ES cell lines used in genetic manipulation have been derived from the 129/Sv strain. In this paper,we have established ES cell lines from blastocysts of the CD-1,C57BL/6J and 129/Sv×C57BL/6J mice besides 129/Sv mouse with a fibroblast cell as feeder cells,and supplemented with 1,000 unit/mL LIF in the culture medium. Twelve ES cell lines were obtained from CD-1,129/Sv, C57BL/6J and 129/Sv×C57BL/6J mouse blastocysts. No ES cell line from KM embryos suggests that KM strain might have inhibitory genetic factor(s) for the ES cell formation. Some ES cell lines could be obtained from hybrids at high efficiency suggested a heterosis effect can be expected for establishing ES cell lines in mice. These ES cell lines produced chimeric mice and contributed to the germ-line. The results indicate that genetic background is an important factor in isolating ES cells.