Analysis of IS21-mediated mobilization of plasmid pACYC184 by R68.45 in Escherichia coli

Riess, G.; Masepohl, Bernd; Puehler, Alfred

doi:10.1016/0147-619x(83)90063-x

Cited by 19 publications

(6 citation statements)

References 15 publications

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“…R68.45, which contains a tandem duplication of insertion sequence IS21, was originally isolated on the basis of its enhanced ability to promote chromosomal transfer (11). R68.45 has been shown to mobilize E. coli plasmids to recA+ and recA E. coli recipients (20,21). The molecular basis for such transfer involves the formation of a cointegrate intermediate during inverse transposition of IS21 (25).…”

Section: Resultsmentioning

confidence: 99%

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Use of a xylE marker gene to monitor survival of recombinant Pseudomonas putida populations in lake water by culture on nonselective media

Winstanley

Morgan

Pickup

et al. 1991

Appl Environ Microbiol

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We constructed equivalent IncP plasmids, pLV1016 and pLV1017, to provide conjugative alternative systems. Detection of the xylE gene carried by marker plasmids was found to be a valid indicator to use for studying the survival of released populations by culturing on nonselective media. These plasmids were used to study the survival of populations of Pseudomonas putida in both sterile and untreated lake water. The effects of inoculum size, the metabolic burden imposed on the cell by the unregulated expression of xylE, and an auxotrophic mutation carried by the host strain were studied. We also assessed the reproducibility and hence the predictability of the survival of released populations. Model systems with a single lake water sample and model systems with three different lake water samples, taken from the same site in consecutive months, were used to analyze variability between replicates and to assess differences caused by host strain or water sample. A large variability was found depending on which water sample was used. These findings imply that it will be difficult to predict accurately the survival of released populations in the natural environment.

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Section: Resultsmentioning

confidence: 99%

“…The molecular basis for such transfer involves the formation of a cointegrate intermediate during inverse transposition of IS21 (25). Resolution of the cointegrates in recA+ recipients occurs at a low frequency, with R68.45 sometimes losing one copy of IS21 and becoming indistinguishable from plasmid R68 (alias RP1, RP4, and RK2) (21).…”

Section: Resultsmentioning

confidence: 99%

Use of a xylE marker gene to monitor survival of recombinant Pseudomonas putida populations in lake water by culture on nonselective media

Winstanley

Morgan

Pickup

et al. 1991

Appl Environ Microbiol

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“…This arrangement enhances the transpositional activity ofIS2l by creating a new promoter reading from one IS2l sequence into the second (56,102,104,111). The tandem IS2l repeat acts as a composite transposon so that the entire intervening R68 plasmid is inserted into the target, either another plasmid or the chromosome (102,103,106,134). R68.45 is able to mobilize chromosomal genes in a wide variety of gramnegative bacteria.…”

Section: Transfer Of Chromosomal Genesmentioning

confidence: 98%

Broad Host Range Conjugative and Mobilizable Plasmids in Gram-Negative Bacteria

Guiney

1993

Bacterial Conjugation

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“…IS21 is relatively large (2131 bp); it is bordered by 11 bp inverted repeats and contains two genes, istA and istB, which are organized as an operon . The prototype plasmid carrying (IS21)2, R68.45, forms cointegrates with other replicons at high frequencies (10-3 to 10-5) and, by transient integration into the bacterial chromosome, promotes effective chromosome transfer in a wide range of bacteria (Haas and Holloway, 1976;Riess et al, 1983;Reimmann et al, 1988;Haas and Reimmann, 1989). IS21 tandem duplications [(IS21)2] arise spontaneously on plasmids in Pseudomonas aeruginosa and Escherichia coli.…”

Section: Introductionmentioning

confidence: 99%

The istA gene of insertion sequence IS21 is essential for cleavage at the inner 3′ ends of tandemly repeated IS21 elements in vitro.

Reimmann

Haas

1990

The EMBO Journal

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The bacterial 2.1 kb insertion sequence IS21 occurs as a tandem repeat [=(IS21)2] on the broad host range plasmid R68.45. In (IS21)2, the two IS21 elements are separated by 3 bp termed junction sequence. Plasmids carrying (IS21)2 form cointegrates with other replicons at high frequencies. The two IS21 genes, istA and istB, were found to be necessary for cointegrate formation in vivo. Since the outer ends of (IS21)2 are dispensable for cointegrate formation, we favor a transposition model according to which a plasmid carrying (IS21)2 is cleaved at the junction sequence; the opened plasmid is then inserted into a target replicon. Here we show that Escherichia coli cell extracts, which contained over‐produced IstA protein, nicked a supercoiled (IS21)2 plasmid precisely at the inner 3′ termini of IS21; the resulting staggered cut generated 5′ protrusions. The istA gene, but not the istB gene, was required for in vitro cleavage of an IS21‐IS21 junction. Because of this cleavage and our previous findings (generation of 4 bp target duplications and loss of the junction sequence after cointegrate formation in vivo) we propose that plasmids with (IS21)2 produce cointegrates by a mechanism which involves joining of the inner 3′ ends of IS21 to the 5′ ends of the target.

show abstract

Analysis of IS21-mediated mobilization of plasmid pACYC184 by R68.45 in Escherichia coli

Cited by 19 publications

References 15 publications

Use of a xylE marker gene to monitor survival of recombinant Pseudomonas putida populations in lake water by culture on nonselective media

Use of a xylE marker gene to monitor survival of recombinant Pseudomonas putida populations in lake water by culture on nonselective media

Broad Host Range Conjugative and Mobilizable Plasmids in Gram-Negative Bacteria

The istA gene of insertion sequence IS21 is essential for cleavage at the inner 3′ ends of tandemly repeated IS21 elements in vitro.

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