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J Bacteriol, June 1998, p. 3039-3048, Vol. 180, No. 12
Molecular Genetics Program, Wadsworth Center,
New York State Department of Health, and Department of Biomedical
Sciences, School of Public Health, State University of New York at
Albany, Albany, New York 12201-2002
Received 20 January 1998/Accepted 10 April 1998
We have examined the targeting preference of the bacterial
insertion element IS903 by determining the sites of
insertion of a large number of transposition events into the 55-kb
conjugative plasmid pOX38. Despite the large target size, all the
insertions were clustered in four small distinct regions associated
with conjugal DNA transfer. Within these regions, many different sites were used for insertion; however, there were a few sites that IS903 inserted into more than once. Alignment of the
insertion sites showed that there was no consensus sequence within the
9-bp target duplication but that there were preferred sequences located symmetrically on either side of the target. This is consistent with
target recognition by a dimer or multimer of transposase, with either
sequence-specific or structure-specific interactions on both sides of
the target. We show further that when one of these preferred regions
was cloned into a second conjugative plasmid, pUB307, it was still a
preferred target, implying that all the sequences necessary for target
selection are contained within this DNA segment. Also, we observed a
very strong preference for insertion in a single orientation in pUB307.
We examined the possibility that either DNA replication from the origin
of vegetative replication, oriV, or the origin of transfer,
oriT, might determine this orientation effect. We find that
reversing the direction of vegetative replication had no effect on the
orientation of transposon insertions; however, reversing the direction
of DNA transfer abolished the orientation effect. This supports the
idea that conjugal DNA transfer imparts a polarity on the target that
is sensed by the transposon.
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Target Choice and Orientation Preference of the
Insertion Sequence IS903
*
Corresponding author. Mailing address: Wadsworth
Center, David Axelrod Institute, 120 New Scotland Ave.,
Albany, NY 12201-2002. Phone: (518) 473-6079. Fax: (518) 486-7971. E-mail: keith.derbyshire{at}wadsworth.org.
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