The carbapenemase gene bla(KPC), which is rapidly spreading worldwide, is located on a Tn3-based transposon, Tn4401. In a transposition-conjugation assay, Tn4401 was able to mobilize bla(KPC-2) gene at a frequency of 4.4 × 10(-6)/recipient cell. A 5-bp target site duplication was evidenced upon each insertion without target site specificity. This study demonstrated that Tn4401 is an active transposon capable of mobilizing bla(KPC) genes at high frequency.