Summary
Interest in the fate of long palindromic DNA sequences in E. coli has been kindled by the observation that their inviability is overcome in recBC sbcB strains and that these hosts permit the construction of DNA libraries containing long palindromic sequences present in the human genome. In this paper we show that a reduction in the level of intracellular supercoiled DNA occurs as the result of the presence of a 530 bp palindrome in bacteriophage lambda. This reduction occurs in Rec+ and recA strains but not in strains lacking exonucleases V and I (recBC sbcB). However, the DNA must be active (not repressed) for this reduction to be observed, since it is not seen in a Rec+ host lysogenic for phage lambda. These results argue against two hypotheses: firstly, that the palindrome causes inviability solely by interfering with packaging, and secondly, that it dose so solely by interfering with recombination. Conversely, these results suggest that a feature of active monomeric DNA (probably its replication) is involved in inviability.
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Bergsma DJ, Olive DM, Hartzell SW, Byrne BJ, Subramanian KN (1982) Cyclization of linear chimeric plasmids in vivo by a novel end-to-end joining reaction or by intramolecular recombination: one of the products contains a 147-bp perfect palindrome stable in E. coli. Gene 20:157–167
Boissy R, Astell CR (1985) An Escherichia coli recBC sbcB recF host permits the deletion resistant propagation of plasmid clones containing the 5′ terminal palindrome of minute virus of mice. Gene 35:179–185
Bolivar F, Betlach MC, Heyneker H, Shine J, Rodriguez R, Boyer H (1977) Origin of replication of pBR345 plasmid DNA. Proc Natl Acad Sci USA 74:5265–5269
Collins J (1981) The instability of palindromic DNA. Cold Spring Harbor Symp Quant Biol 45:409–416
Collins J, Volckaert G, Nevers P (1982) Precise and nearly-precise excision of the symmetrical inverted repeats of Tn5: common features of recA-independent deletion events in E. coli. Gene 19:139–146
Courey AJ, Wang JC (1983) Cruciform formation in a negatively supercoiled DNA may be kinetically forbidden under physiological conditions. Cell 33:817–829
Davis RW, Simon M, Davidson N (1971) Electron Microscope heteroduplex methods for mapping regions of base sequence homology in nucleic acids. Methods Enzymol 21:413–428
Echols H (1971) Regulation of lytic development. In: Hershey A (ed) Bacteriophage lambda. Cold Spring Harbour Laboratory, New York, pp 247–270
Enquist LW, Skalka A (1973) Replication of bacteriophage lambda DNA dependent on the function of host and viral genes. J Mol Biol 75:185–212
Gellert M, Mizuuchi K, O'Dea MH, Ohmori H, Tomizawa J (1979) DNA gyrase and DNA supercoiling. Cold Spring Harbor Symp Quant Biol 43:35–40
Gellert M, O'Dea MH, Mizuuchi K (1983) Slow cruciform transitions in palindromic DNA. Proc Natl Acad Sci USA 80:5545–5549
Greaves DR, Patient PK, Lilley DMJ (1985) Facile cruciform formation from a Xenopus globin gene. J Mol Biol 185:461–478
Hagan CE, Warren GI (1982) Lethality of palindromic DNA and its use in the selection of recombinant plasmids. Gene 19:147–151
Hagan CE, Warren GJ (1983) Viability of palindromic DNA is restored by deletions occuring at low but variable frequency in plasmids of E. coli. Gene 24:317–326
Kaiser D (1971) Lambda DNA replication. In Hershey AD (ed) The Bacteriophage Lambda. Cold Spring Harbor Laboratory, New York, pp 195–210
Kellenberger E, Edgar RS (1971) Structure and assembly of phage particles. In: Hershey A (ed) Bacteriophage Lambda. Cold Spring Harbor Laboratory, New York, pp 271–295
Leach DRF, Stahl FW (1983) Viability of lambda phages carrying a perfect palindrome in the absence of recombination nucleases. Nature 305:448–451
Lilley DMJ (1981) In vivo consequences of plasmid topology. Nature 292:380–382
Mizzuchi K, Mizuuchi M, Gellert M (1982) Cruciform structures in palindromic DNA are favoured by DNA supercoiling. J Mol Biol 156:229–244
Panayotatos N, Wells RD (1981) Cruciform structures in supercoiled DNA. Nature 289:466–470
Reuben R, Gefter M, Enquist L, Skalka A (1974) New method for large-scale preparation of covalently closed λ DNA molecules. J Virol 14:1104–1107
Sinden RR, Broyles SS, Pettijohn DE (1983) Perfect palindromic lac operator DNA sequence exists as a stable cruciform structure in supercoiled DNA in vitro but not in vivo. Proc Natl Acad Sci USA 80:1797–1801
Smith GE, Summers MD (1980) The bidirectional transfer of DNA and RNA to nitrocellulose and diazobenzyloxymethyl-paper. Anal Biochem 109:123–129
Stahl FW, McMilin KD, Stahl M, Malone RE, Nozu Y (1972) A role for recombination in the production of “free-loader” lambda bacteriophage particles. J Mol Biol 68:57–67
Taylor A, Smith GR (1980) Unwinding and rewinding of DNA by the RecBC enzyme. Cell 22:447–457
Taylor AF, Smith GR (1985) Substrate specificity of the DNA unwinding activity of the RecBC enzyme of E. coli. J Mol Biol 185:431–443
Warren GJ, Green RL (1985) Comparison of physical and genetic properties of palindromic DNA sequences. J Bacteriol 161:1103–1111
Wyman AR, Wolfe LB, Botstein D (1985) Propagation of some human DNA sequences in bacteriophage lambda vectors requires mutant E. coli hosts. Proc Natl Acad Sci USA 82:2880–2884
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Communicated by G.R. Smith
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Leach, D., Lindsey, J. In vivo loss of supercoiled DNA carrying a palindromic sequence. Molec. Gen. Genet. 204, 322–327 (1986). https://doi.org/10.1007/BF00425517
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DOI: https://doi.org/10.1007/BF00425517