As one of the most limiting AAs, methionine (Met) and its peptide form have been investigated decades for the ability to promote milk protein synthesis in vitro. This study aimed to investigate the effects of methionyl-methionine (Met-Met) on prolificacy and milk performance in mice. 1) To study the first pass effect of Met, 48 pregnant mice were randomly divided into 6 groups with intraperitoneal injection of 0, 5, 15, 25, 35 and 45% Met daily (based on 5g /d dry matter intake), from embryonic d 1 to 17. The control group was fed with Met supplementation diet, the other groups were fed with Met free diets. 2) Then 56 pregnant mice were assigned to 7 groups with intraperitoneal injection of 0, 5, 15, 25, 35 and 45% Met-Met replaced of 35% Met daily. At embryonic d 17, all mice were slaughtered to collect mammary gland for the underlying mechanisms of milk protein synthesis. Data were analyzed using the ANOVA procedure of IBM SPSS statistics 20.The results showed that (1) 35% Met supplementation increased total number of fetuses and placental weight compared with 5, 15% Met treatments. However, placental weights were decreased significantly when mice fed free-Met diet; (2) 25% Met-Met supplementation increased total number of fetuses compared with 45% Met-Met supplementation. Additionally, placental weights were increased when mice supplemented with 25% Met-Met compared with 0, 5 and 45% Met-Met. Furthermore, the mRNA abundance of β-casein, mammalian target of rapamycin (mTOR), Janus kinase 2 (JAK2) and signal transducer and activator of transcription 5 (STAT5) were increased in 25% Met-Met compared with the control and 35% free Met supplementation. In conclusion, Met-Met restored prolificacy and promoted milk protein synthesis by activating mTOR and JAK2-STAT5 signaling pathways.