Objective Establishment a Sleeping Beauty (SB)transposase-expressing transgenic mice for study of the genetic modification mediated by transposon system in mice. Methods Using the UBC promoter to drive the expression of SB transposase. The transgenic mice were created by microinjection. The gene type of transgenic line was identified by PCR. The expressing level in testis was determined by Western Blot and immunohistochemistry (IHC) staining. Results Four lines of transposase transgenic mice were obtained by microinjection and three can be germline. One mouse line with higher expression level of transposase in the testis was obtained. Conclusion One transgenic mice model with Sleeping Beauty transposase –expressing was successfully established. This model will greatly contribute to the research of genetic modification mediated by transposon in mouse.