Summary
Recombination is extensively used in order to move alleles between replicons. The exchange of wild-type chromosomal and mutant plasmid-borne alleles is a two-step process entailing the formation of a cointegrate between the entire plasmid and the chromosome, followed by resolution of such cointegrates to give a mutant chromosome and a plasmid carrying the wild-type chromosomal sequence. Often the cointegrate and the resolved forms cannot be distinguished phenotypically. To enable the direct isolation of the resolved products we have developed a positive selection technique. Cells containing a cointegrated plasmid R1 were constructed by transduction using a P1 lysate prepared from cells harbouring a plasmid comprising a mutant chromosomal allele and the so-called Ω fragment which carries an aad (aminoglycoside adenylyltransferase) gene. P1 transduction from the cointegrate strain into an SmD recipient allowed direct selection for the resolved complex, since transduction of the aad gene is lethal to an SmD strain.
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Abbreviations
- kb:
-
kilobase pairs
- bp:
-
base pairs
- Ap:
-
ampicillin
- Sp:
-
spectinomycin
- Sm:
-
streptomycin
- SmD:
-
streptomycin dependence
- superscripts (Γ):
-
refer to antibiotic resistance
References
Benveniste R, Yamada T, Davies J (1970) Enzymatic adenylylation of streptomycin and spectinomycin by R-factor-resistant Escherichia coli. Infect Immun 1:109–119
Bertani G (1951) Studies on lysogenesis. The mode of phage liberation by lysogenic Escherichia coli. J Bacteriol 62:293–300
Buhk, H-J, Messer W (1983) The replication origin region of Escherichia coli: nucleotide sequence and functional units. Gene 24:265–279
Chak KF, de Lencastre H, Liu H-M, Piggot PJ (1982) Facile in vivo transfer of mutations between the Bacillus subtilis chromosome and a plasmid harbouring homologous DNA. J Gen Microbiol 128:2813–2816
Frey J, Krisch HM (1985) Ω mutagenesis in Gram-negative bacteria: a selectable interposon which is strongly polar in a wide range of bacterial species. Gene 36:143–150
Herman RK (1965) Reciprocal recombination of chromosome and F-merogenote in Escherichia coli. J Bacteriol 90:1664–1668
Hofemeister J, Böhme H (1968) Interaction between R-factors and the host cells. I. Lethal interference between R-factor determined streptomycin resistance and streptomycin dependence in Proteus mirabilis. Mol Gen Genet 103:258–265
Iglesias A, Trautner TA (1983) Plasmid transformation in Bacillus subtilis: symmetry of gene conversion in transformation with a hybrid plasmid containing chromosomal DNA. Mol Gen Genet 189:73–76
Koppes L, Nordström K (1986) Insertion of an R1 plasmid into the origin of replication of the E. coli chromosome: random timing of replication of the hybrid chromosome. Cell 44: 117–124
Liljeström P, Pirhonen M, Palva ET (1985) In vivo transfer of chromosomal mutations onto multicopy plasmids by transduction with bacteriophage P1. Gene 40:241–246
Park C, Hazelbauer GL (1986) Transfer of chromosomal mutations to plasmids via Hfr-mediated conduction. J Bacteriol 165: 312–314
Rownd RH, Miki T, Appelbaum ER, Miller JR, Finkelstein M, Barton CR (1978) Dissociation, amplification and reassociation of composite R-plasmid DNA. In: Schlessinger D (ed) Microbiology — 1978. American Society for Microbiology, Washington DC, pp 33–37
Ruusala T, Andersson D, Ehrenberg M, Kurland CG (1984) Hyper-accurate ribosomes inhibit growth. EMBO J 3: 2575–2580
von Meyenburg K, Hansen FG, Nielsen LD, Riise E (1978) Origin of replication, oriC, of the Escherichia coli chromosome on specialized transducing phages λasn. Mol Gen Genet 160: 287–295
Whitehouse HLK (1982) Genetic recombination. Understanding the mechanisms. Wiley, Chichester, New York, Brisbane, Toronto, pp 162–165
Yamada T, Tipper D, Davies J (1968) Enzymatic inactivation of streptomycin by R factor-resistant Escherichia coli. Nature 219:288–291
Yamaguchi K, Tomizawa J-I (1980) Establishment of Escherichia coli cells with an integrated high copy number plasmid. Mol Gen Genet 178:525–533
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Communicated by R. Devoret
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Merryweather, A., Bernander, R. & Nordström, K. Direct selection for the exchange of alleles between a plasmid and the Escherichia coli chromosome. Mol Gen Genet 210, 381–384 (1987). https://doi.org/10.1007/BF00327186
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DOI: https://doi.org/10.1007/BF00327186