Objective To establish a hygromyein-resistant transgenie BDF1 model for preparation of feeder layers of hygromyein-resistant ES cells clones. Methods Hygremyein B phosphotransferase gene fragments were mleroinjected into the zygote male pronuelei of BDF1 mice. Altogether 169 zygotes were injected and 129 zygote cells were transplanted into the oviduct of aceeptor pregnant female mice. Results Genome DNA from mice tail were detected by PCR and Southern blot showing 9 of 37 were positives transgenie mice. One of the founders was analysed by RT-PCR and found that hyg was expressed in the kidney, muscle and spleen. Conclusion A hygromyein-resistant transgenie BDF1 mouse model has been successfully established. This model will provide favorable fundamental conditions for gene knock-out studies.