Host:Vector Systems for Gene Cloning in Pseudomonas

Bagdasarian, Michael; Timmis, Kenneth N.

doi:10.1007/978-3-642-68315-2_4

Cited by 141 publications

(96 citation statements)

References 47 publications

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“…Plasmids used were pWWO (TOL) (6), pWWO-161 [TOL::Tn401, which encodes resistance to ampicillin (Apr) and carbenicillin (Cbr)] (7), pWWO-39 (a xylA mutant of pWWO; in PaW39), pWWO-210 (a xylR mutant of pWWO; in PaW210), RP4 (8), pKT234, pKT210, pKT230, and pKT231. The latter three plasmids are cloning vectors derived from the broad host range plasmid RSF1010 and specify streptomycin and chloramphenicol resistance (Smr, Cmr), Smr and kanamycin resistance (Kmr), and Smr and Kmr, respectively (5,9,10 growth and thereby results in increased expression of cloned genes via gene dosage (unpublished data).…”

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confidence: 99%

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Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway.

Franklin¹,

Bagdasarian²,

Timmis³

1981

Proc. Natl. Acad. Sci. U.S.A.

502

281

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confidence: 99%

“…Media used for cultivation of E. coli (11) and P. putida (6) bacteria have been described, as have conditions for plasmid DNA isolation (11,12), transformation (5), and cleavage by restriction endonucleases, analysis by agarose gel electrophoresis, and ligation (13). Initial cloning of TOL DNA fragments was carried out in E. coli SK1592.…”

mentioning

confidence: 99%

Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway.

Franklin¹,

Bagdasarian²,

Timmis³

1981

Proc. Natl. Acad. Sci. U.S.A.

502

281

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“…The transformation of P. putida cells with plasmid DNA was as described by Bagdasarian & Timmis (1982) and the transformation of E. coli cells was as described by Hanahan (1983).…”

Section: Methodsmentioning

confidence: 99%

Transposon-mediated mobilization of chromosomally located catabolic operons of the CAM plasmid by TOL plasmid transposon Tn4652 and CAM plasmid transposon Tn3614

Mäe¹,

Heinaru

1994

Microbiology

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The CAM (camphor degradation) plasmid is integrated into the chromosome of Pseudomonas putida Paw-line strains and is not selftransferable as a plasmid via conjugation. Our results show that the mobilization of chromosomally located CAM and the integration of cam-operons into the chromosome of the new Cam+ transconjugants is a red-independent process mediated by transposons Tn4652 (17 kbp) and Tn3614 (7.2 kbp). Transposon Tn3674 is apparently identical to the left-hand and the right-hand sequences of the TOL plasmid pWW0 transposon Tn4654. The insertion of Tn407 inside the left-hand terminal IR of Tn4652 completely inhibited the mobilization of CAM. According to our data transposons Tn4652 and Tn3614 together with CAM plasmid catabolic operons are integrated into the chromosome. We propose that in pseudomonads the transposons Tn4652 and Tn3614 play a key role in the evolution and spread of new catabolic plasmids in nature.

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“…For the overproduction of P1 repressor, a P1 DNA subfragment of pBR325-P1:7 (8) and pBR325-P1:7c1.100 (obtained from N. Sternberg, Du Pont) was inserted into the expression vector pPLc28 (19) and pJF118EH (20), respectively. Other plasmids used were pBR325 (21), pKT101 (22), and the galK promoter selection vector pFD51 (23).…”

Section: Methodsmentioning

confidence: 99%

Multiple repressor binding sites in the genome of bacteriophage P1.

Velleman¹,

Dreiseikelmann²,

Schuster³

1987

Proc. Natl. Acad. Sci. U.S.A.

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After digestion of bacteriophage P1 DNA with EcoRI in the presence of P1 repressor, 6 repressor binding sites were identified in 5 of 26 EcoRI fragments. Binding sites were localized by the decreased mobility of DNA fragment-repressor complexes during electrophoresis and by DNase protection ("footprinting") analysis. The repressor binding sites, or operators, comprise a 17-base-pair-long consensus sequence lacking symmetrical elements. Three operators can be related to known genes, whereas the function of the others is still unknown. The mutant P1 bac, rendering ban expression constitutive, is identified as an operator-constitutive mutation of the ban operon.P1 is a temperate bacteriophage with a genome size of about 90 kilobases (kb). In the prophage state the proviral DNA is maintained as a plasmid, and the vegetative P1 functions are repressed. Repression is accomplished by a phage-specific repressor, the product of the cl gene, which is located at the far right side of the P1 genetic map in EcoRI restriction fragment 7 (P1:7) (1, 2). Partially purified P1 repressor binds in vitro to at least two regions near cl within BamHI fragment 9 that itself is located within P1:7 (3, 4).The binding sites close to the cl gene, however, are not the only region at which the P1 repressor acts. The latter can also repress in vivo the expression of the P1 ban gene, which is located in P1:3 (5, 6). Furthermore, P1:14 also contains a promoter repressible by the product of cl (7). Together these results reveal that the P1 cl repression system must differ from that of other temperate phages such as A, P2, and P22, in which only promoters adjacent to the repressor gene are repressed (7).During our studies on the regulation of phage P1 ban expression we have localized by indirect methods a region 5' upstream of the ban gene within P1:3 at which the P1 repressor acts (8). Highly purified P1 repressor protein binds to this region in vitro (H.S., unpublished data). Now in a systematic search for other repressor binding sites in the phage genome, EcoRI-digested P1 DNA is incubated-with repressor, and binding regions are identified by the decreased mobility of EcoRI fragment-repressor complexes during electrophoresis (9,10). Thus binding regions were detected in P1:7 and P1:14, as expected and, in addition, in P1:9 and P1:11. Moreover, the results of DNA sequence and DNase protection analyses reveal an asymmetric 17-base-pair (bp)-long consensus sequence for the P1 repressor binding site. MATERIALS AND METHODSBacteria, Phage, and Plasmids. Escherichia coli K-12 strains used included the following: C600, HB101 (recA13) (11), NY58 (dnaBI07, recA56) (12), JM101 (13), N100 (galK, recA) (14), DW101 (sup'), and DW103 (supD) (15). For the cloning of P1 DNA fragments containing the ban operon (8) the recipient bacteria C600, HB101, NY58, and JM101 carried the plasmid pKT101-P1:7 harboring the P1 repressor gene (8). Strains DW101 and DW103 were transformed by plasmid pBR325-P1:11 for marker rescue tests (2).Phage used were M13mp8/9 (16)...

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Host:Vector Systems for Gene Cloning in Pseudomonas

Cited by 141 publications

References 47 publications

Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway.

Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway.

Transposon-mediated mobilization of chromosomally located catabolic operons of the CAM plasmid by TOL plasmid transposon Tn4652 and CAM plasmid transposon Tn3614

Multiple repressor binding sites in the genome of bacteriophage P1.

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