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. 1977 Apr;4(4):969–987. doi: 10.1093/nar/4.4.969

Selective breakage of DNA alongside 5-bromodeoxyuridine nucleotide residues by high temperature hydrolysis.

G W Grigg
PMCID: PMC342498  PMID: 866201

Abstract

The substitution of thymine mucleotides (pT) in oligodeoxynucleotides by bromouracil nucleotides (pBU) changes the properties of the oligonucleotides in two ways: (1) It alters their mobility during DEAE-Cellulose homochromatography1. (2) It substantially enhances their sensitivity to high temperature hydrolysis under mildly alkaline conditions (pH 8.9). The resultant breaks occur adjacent to pBU residues and leave terminal phosphates on the breakage products. With more extreme conditions some loss of terminal phosphates can occur. Heating at 100 degrees for 16 hr at pH 8.9 produces cleavage at about half of the pBU residues with minimal loss of terminal phosphates. The properties described here may explain the thermal sensitivity of bacteria grown in 5BU2 and may have a use in DNA sequencing technology.

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Selected References

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  1. Bick M. D., Davidson R. L. Total substitution of bromodeoxyuridine for thymidine in the DNA of a bromodeoxyuridine-dependent cell line. Proc Natl Acad Sci U S A. 1974 May;71(5):2082–2086. doi: 10.1073/pnas.71.5.2082. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Boyce R. P. Production of additional sites of deoxyribonucleic acid breakdown in bromouracil containing Escherichia coli exposed to ultra-violet light. Nature. 1966 Feb 12;209(5024):688–691. doi: 10.1038/209688a0. [DOI] [PubMed] [Google Scholar]
  3. Brownlee G. G., Sanger F. Chromatography of 32P-labelled oligonucleotides on thin layers of DEAE-cellulose. Eur J Biochem. 1969 Dec;11(2):395–399. doi: 10.1111/j.1432-1033.1969.tb00786.x. [DOI] [PubMed] [Google Scholar]
  4. GREER S. Studies on ultraviolet irradiation of Escherichia coli containing 5-bromouracil in its DNA. J Gen Microbiol. 1960 Jun;22:618–634. doi: 10.1099/00221287-22-3-618. [DOI] [PubMed] [Google Scholar]
  5. GREER S., ZAMENHOF S. Studies on depurination of DNA by heat. J Mol Biol. 1962 Mar;4:123–141. doi: 10.1016/s0022-2836(62)80046-1. [DOI] [PubMed] [Google Scholar]
  6. Galibert F., Sedat J., Ziff E. Direct determination of DNA nucleotide sequences: structure of a fragment of bacteriophage phiX172 DNA. J Mol Biol. 1974 Aug 15;87(3):377–407. doi: 10.1016/0022-2836(74)90093-x. [DOI] [PubMed] [Google Scholar]
  7. Henderson E. E., Strauss B. Differences in the incorporation of bromodeoxyuridine by human lymphoblastoid cell lines. Cell. 1975 Aug;5(4):381–387. doi: 10.1016/0092-8674(75)90057-4. [DOI] [PubMed] [Google Scholar]
  8. Hutchinson F., Hales H. B. Mechanism of the sensitization of bacterial transforming DNA to ultraviolet light by the incorporation of 5-bromouracil. J Mol Biol. 1970 May 28;50(1):59–69. doi: 10.1016/0022-2836(70)90103-8. [DOI] [PubMed] [Google Scholar]
  9. KOZINSKI A. W., SZYBALSKI W. Dispersive transfer of the parental DNA molecule to the progeny of phage phiX-174. Virology. 1959 Oct;9:260–274. doi: 10.1016/0042-6822(59)90119-9. [DOI] [PubMed] [Google Scholar]
  10. Korenberg J. R., Freedlender E. F. Giemsa technique for the detection of sister chromatid exchanges. Chromosoma. 1974;48(4):355–360. doi: 10.1007/BF00290992. [DOI] [PubMed] [Google Scholar]
  11. Lion M. B. Search for a mechanism for the increased sensitivity of 5-bromouracil-substituted DNA to ultraviolet radiation. II. Single-strand breaks in the DNA of irradiated 5-bromouracil-substituted T3 coliphage. Biochim Biophys Acta. 1970 May 21;209(1):24–33. doi: 10.1016/0005-2787(70)90657-x. [DOI] [PubMed] [Google Scholar]
  12. Regan J. D., Setlow R. B., Ley R. D. Normal and defective repair of damaged DNA in human cells: a sensitive assay utilizing the photolysis of bromodeoxyuridine. Proc Natl Acad Sci U S A. 1971 Apr;68(4):708–712. doi: 10.1073/pnas.68.4.708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Sanger F., Coulson A. R. A rapid method for determining sequences in DNA by primed synthesis with DNA polymerase. J Mol Biol. 1975 May 25;94(3):441–448. doi: 10.1016/0022-2836(75)90213-2. [DOI] [PubMed] [Google Scholar]
  14. Sanger F., Donelson J. E., Coulson A. R., Kössel H., Fischer D. Determination of a nucleotide sequence in bacteriophage f1 DNA by primed synthesis with DNA polymerase. J Mol Biol. 1974 Dec 5;90(2):315–333. doi: 10.1016/0022-2836(74)90376-3. [DOI] [PubMed] [Google Scholar]
  15. Sanger F., Donelson J. E., Coulson A. R., Kössel H., Fischer D. Use of DNA polymerase I primed by a synthetic oligonucleotide to determine a nucleotide sequence in phage fl DNA. Proc Natl Acad Sci U S A. 1973 Apr;70(4):1209–1213. doi: 10.1073/pnas.70.4.1209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ziff E. B., Sedat J. W., Galibert F. Determination of the nucleotide sequence of a fragment of bacteriophage phiX 174 DNA. Nat New Biol. 1973 Jan 10;241(106):34–37. doi: 10.1038/newbio241034a0. [DOI] [PubMed] [Google Scholar]

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