Isolation and characterization of IS1 circles.
Transposase encoded by insertion sequence IS1 is produced from two out-of-phase reading frames by translational frameshifting that occurs in a run of adenines. An IS1 mutant with a single adenine insertion in the run of adenines efficiently produces transposase, resulting in generation of miniplasmids by deletion for a region adjacent to IS1 from a plasmid carrying the IS1 mutant. Here, we found that besides miniplasmids, cells harboring the plasmid contained minicircles without the region required for replication. Cloning and DNA sequencing of the minicircles revealed that most of them were IS1 circles consisting of the entire IS1 sequence and a sequence, 5-9 bp in length, which intervenes between terminal inverted repeats, IRL and IRR, of IS1. Analysis of more IS1 circles isolated by polymerase chain reaction revealed that the intervening sequence was derived from the region flanking either IRL or IRR in the parental plasmid, suggesting that IS1 circles are generated by an excision event from the parental plasmid. The IS1 circles may be formed due to the cointegration reaction occurring within the parental plasmid carrying IS1.
Sekine
Y
Y
Institute of Molecular and Cellular Biosciences, The University of Tokyo, Bunkyo-ku, Japan.
Eisaki
N
N
Kobayashi
K
K
Ohtsubo
E
E
D003001:Cloning, Molecular; D004254:DNA Nucleotidyltransferases / Q000235:genetics*; D004251:DNA Transposable Elements / Q000235:genetics*; D004270:DNA, Circular / Q000737:chemistry / Q000235:genetics*; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D018965:Frameshifting, Ribosomal; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000737:chemistry / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D017422:Sequence Analysis, DNA; D019895:Transposases
D004251:DNA Transposable Elements; D004270:DNA, Circular; D004254:DNA Nucleotidyltransferases; D019895:Transposases