Tn7 transposition in vitro proceeds through an excised transposon intermediate generated by staggered breaks in DNA.
We have developed a cell-free system in which the bacterial transposon Tn7 inserts at high frequency into its preferred target site in the Escherichia coli chromosome, attTn7; Tn7 transposition in vitro requires ATP and Tn7-encoded proteins. Tn7 transposes via a cut and paste mechanism in which the element is excised from the donor DNA by staggered double-strand breaks and then inserted into attTn7 by the joining of 3' transposon ends to 5' target ends. Neither recombination intermediates nor products are observed in the absence of any protein component or DNA substrate. Thus, we suggest that Tn7 transposition occurs in a nucleoprotein complex containing several proteins and the substrate DNAs and that recognition of attTn7 within this complex provokes strand cleavages at the Tn7 ends.
Bainton
R
R
Department of Biochemistry and Biophysics, George W. Hooper Foundation, University of California, San Francisco, California 94143.
Gamas
P
P
Craig
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NL
D000255:Adenosine Triphosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D015139:Blotting, Southern; D002474:Cell-Free System; D002876:Chromosomes, Bacterial; D004261:DNA Replication; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods; D011995:Recombination, Genetic
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D000255:Adenosine Triphosphate