To test whether truncated midkine (tMK), an alternative spliced form of exon 3, induces the transformation of mammalian cells, tMK cDNA was transfected into SW-13 cells. Although, the growth of MK transfectant (SW-13/MK) cells was close to those of the parent SW-13 and vector transfectant (SW-13/empty) cells, the growth of tMK transfectant (SW-13/tMK) cells was significantly promoted compared with that of the above three cells. Both SW-13/tMK and SW-13/MK formed colonies in 0.35% soft agar, indicating that tMK and MK induce mammalian cell transformation. SW-13/tMK frequently formed solid tumor after its subcutaneous injection into nude mice. Additionally, in SW-13/tMK and SW-13/MK-injected mice, advanced visible tumors were detected compared with that in the case of SW-13/empty-injected mice as control. These findings indicate that tMK induces mammalian cell transformation and promotes tumor establishment in vivo.