Tn5 in Vitro Transposition

This communication reports the development of an efficient in vitro transposition system for Tn 5 . A key component of this system was the use of hyperactive mutant transposase. The inactivity of wild type transposase is likely to be related to the low frequency of in vivo transposition. The in vitro experiments demonstrate the following: the only required macromolecules for most of the steps in Tn 5 transposition are the transposase, the specific 19-bp Tn 5 end sequences, and target DNA; transposase may not be able to self-dissociate from product DNAs; Tn 5 transposes by a conservative “cut and paste” mechanism; and Tn 5 release from the donor backbone involves precise cleavage of both 3′ and 5′ strands at the ends of the specific end sequences.