Survivin is a crucial node molecule involved in apoptosis, cell division and drug discovery. Up-regulation of survivin in the tissues of oral submucous fibrosis (OSF) and oral squamous cell carcinoma (OSCC) originated from OSF has already been demonstrated. Survivin Thr34 phosphorylation is involved in the inhibition of apoptosis and cell division. To determine the potential involvement of survivin Thr34 phosphorylation in carcinogenesis of OSF, 40 OSFs, 42 OSCCs originated from OSF and 10 normal tissues from surgical specimens were studied. Immunohistochemistry showed that the positive staining rate of the survivin phosphorylation on Thr34 in OSCC originated from OSF group was significantly higher than that in OSF group (P<0.01), and none in the normal oral mucosa specimens. Survivin phosphorylation on Thr34 is predominantly located in the nucleus, which account for its function in apoptosis at cell division. Western blotting analysis showed increasing expression of survivin Thr34 phosphorylation, cyclin B1 and p34cdc2 in carcinogenesis of OSF. Furthermore, p34cdc2-cyclin B1 kinase was confirmed to phosphorylate survivin on Thr34 in carcinogenesis of OSF by immunoprecipitation and immunoblot. These results suggest that the phosphorylation of survivin on Thr34 critically regulate survivin and plays an important role during the malignant transformation of OSF, which will provide an indication to early diagnosis and therapy in carcinogenesis of OSF.