Artificial transposable elements in the study of the ends of IS 1

Abstract

We have constructed artificial IS 1-based transposons by attaching synthetic oligodeoxynucleotides, corresponding to the sequence of the ends of IS 1, to a selectable DNA segment [‘Ω’ fragment; Prentki and Krisch, Gene 29 (1984) 303–313]. These transposons were used to examine the sequence requirements at the ends for IS 1 transposition. We show here that a 24- to 28-bp sequence from the left or right ends of IS 1 is capable of transposition when present at both ends of the Ω fragment in the correct orientation. Transposition activity requires the presence of an intact IS 1 in cis on the same plasmid molecule. In trans, however, neither resident genomic copies of IS 1, nor copies carried by a compatible, high-copy-number plasmid present in the same cell, complement the artificial transposons efficiently. Transposition frequencies in the presence of a cis-complementing IS 1 are, however, similar to those of the naturally occurring IS 1-based transposon, Tn9. In addition, transposition results in a 9-bp duplication in the target DNA molecule as is usually the case for insertion of the intact IS 1. Using this system, we have obtained evidence indicating that the activity of a synthetic IS 1 end is not determined exclusively by its sequence, but can be strongly enhanced by a second, wild-type end used in the transposition event. The data also show that single base pair mutations can exhibit a cumulative effect in reducing transposition activity.