The experiment observed the estrous cycle of twenty-five mice with two vagina smear methods .The mice whose estrous cycle were regular were cheese to handle with superovulation method. According to the different periods (estrus,post-estrus,interestrus ),these mice classified into three test groups, and then other thirteen mice that were unknown estrous cycle were contrast group. The results showed that : (1) The method 2 was better than method 1, the method 2 had little effect to the estrous cycle of the mice. (2) The superovulation results of the interestrus mice were the best of all other periods. The rates of the valuable embryos and embryos quantity were significantly better than the estrus,post-estrus and unknown estrous cycle.