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. 1972 Nov;12(11):1573–1582. doi: 10.1016/S0006-3495(72)86183-6

Breakage of Parental DNA Strands in Haemophilus influenzae by 313 nm Radiation after Replication in the Presence of 5-Bromodeoxyuridine

Kenneth L Beattie
PMCID: PMC1484201  PMID: 4539331

Abstract

Haemophilus influenzae was labeled with thymidine-3H (dThd), then grown in the presence of 5-bromodeoxyuridine (BrdUrd), and then irradiated with 313 nm light (a wavelength that selectively photolyzes DNA containing 5-bromouracil [BrUra]). Irradiation with 313 nm light induced breaks in the 3H-labeled strands in cells grown with BrdUrd at a much higher frequency than in 14C-labeled DNA of cells not exposed to BrdUrd. Breakage of the 3H-labeled strands was about 0.6% as efficient as that of fully BrUra-substituted DNA. During growth in the presence of BrdUrd, susceptibility to 313 nm-induced breakage of the 3H-labeled DNA strands increased, reaching a maximum in about one generation, and it decreased to zero during subsequent growth for one generation in medium containing dThd instead of BrdUrd. Heat denaturation of DNA extracted from dThd-3H-labeled cells grown in the presence of BrdUrd eliminated 313 nm-induced breakage of the 3H-labeled strands. It is concluded that breakage of the 3H-labeled DNA strands resulted from reaction with photoproducts in the base-paired, BrUra-containing strands, rather than from photolysis of BrdUrd incorporated into parental strands. It may be possible to utilize the phenomenon of interstrand breakage in physical studies of DNA replication.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beattie K. L., Setlow J. K. Transformation-defective strains of Haemophilus influenzae. Nat New Biol. 1971 Jun 9;231(23):177–179. doi: 10.1038/newbio231177a0. [DOI] [PubMed] [Google Scholar]
  2. Boling M. E., Setlow J. K. Dependence of Vegetative Recombination Among Haemophilus influenzae Bacteriophage on the Host Cell. J Virol. 1969 Sep;4(3):240–243. doi: 10.1128/jvi.4.3.240-243.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bram S. Secondary structure of DNA depends on base composition. Nat New Biol. 1971 Aug 11;232(2):174–176. doi: 10.1038/newbio232174a0. [DOI] [PubMed] [Google Scholar]
  4. Carrier W. L., Setlow R. B. Paper strip method for assaying gradient fractions containing radioactive macromolecules. Anal Biochem. 1971 Oct;43(2):427–432. doi: 10.1016/0003-2697(71)90272-7. [DOI] [PubMed] [Google Scholar]
  5. Herriott R. M., Meyer E. Y., Vogt M., Modan M. Defined medium for growth of Haemophilus influenzae. J Bacteriol. 1970 Feb;101(2):513–516. doi: 10.1128/jb.101.2.513-516.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hotz G., Reuschl H. Damage to deoxyribose molecules and to U-gene reactivation in UV-irradiated 5-bromouracil-DNA of phage T4 Bor as influenced by cysteamine. Mol Gen Genet. 1967;99(1):5–11. doi: 10.1007/BF00306453. [DOI] [PubMed] [Google Scholar]
  7. 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]
  8. Ishihara H., Wang S. Y. Photochemistry of 5-bromo-1,3-dimethyluracil in aqueous solution. Biochemistry. 1966 Jul;5(7):2302–2307. doi: 10.1021/bi00871a019. [DOI] [PubMed] [Google Scholar]
  9. Köhnlein W., Hutchinson F. ESR-studies of normal and 5-bromouracil-substituted DNA of Bacillus subtilis after irradiation with ultraviolet light. Radiat Res. 1969 Sep;39(3):745–757. [PubMed] [Google Scholar]
  10. Lehmann A. R. Postreplication repair of DNA in ultraviolet-irradiated mammalian cells. J Mol Biol. 1972 May 28;66(3):319–337. doi: 10.1016/0022-2836(72)90418-4. [DOI] [PubMed] [Google Scholar]
  11. Ley R. D., Setlow R. B. Repair replication in Escherichia coli as measured by the photolysis of bromodeoxyuridine. Biophys J. 1972 Apr;12(4):420–431. doi: 10.1016/S0006-3495(72)86094-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. McGrath R. A., Williams R. W. Reconstruction in vivo of irradiated Escherichia coli deoxyribonucleic acid; the rejoining of broken pieces. Nature. 1966 Oct 29;212(5061):534–535. doi: 10.1038/212534a0. [DOI] [PubMed] [Google Scholar]
  13. RUPP W. D., PRUSOFF W. H. PHOTOCHEMISTRY OF IODOURACIL. II. EFFECTS OF SULFUR COMPOUNDS, ETHANOL AND OXYGEN. Biochem Biophys Res Commun. 1965 Jan 18;18:158–164. [PubMed] [Google Scholar]
  14. 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]
  15. Setlow J. K., Brown D. C., Boling M. E., Mattingly A., Gordon M. P. Repair of deoxyribonucleic acid in Haemophilus influenzae. I. X-ray sensitivity of ultraviolet-sensitive mutants and their behavior as hosts to ultraviolet-irradiated bacteriophage and transforming deoxyribonucleic acid. J Bacteriol. 1968 Feb;95(2):546–558. doi: 10.1128/jb.95.2.546-558.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Wells R. D., Larson J. E., Grant R. C., Shortle B. E., Cantor C. R. Physicochemical studies on polydeoxyribonucleotides containing defined repeating nucleotide sequences. J Mol Biol. 1970 Dec 28;54(3):465–497. doi: 10.1016/0022-2836(70)90121-x. [DOI] [PubMed] [Google Scholar]

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