Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1973 Dec;70(12 Pt 1-2):3660–3664. doi: 10.1073/pnas.70.12.3660

Molecular Nature of Mammalian Cell DNA in Alkaline Sucrose Gradients

Joseph R Simpson *, William A Nagle *, Michael D Bick †, James A Belli *,‡
PMCID: PMC427301  PMID: 4519655

Abstract

Mammalian cell DNA that exhibited anomalous sedimentation in alkaline sucrose gradients was examined directly by electron microscopy. Its appearance was that of duplex DNA. In addition, some duplex DNA was observed under conditions in which the sedimentation anomaly was no longer apparent. Persistence of double-stranded DNA under denaturing conditions suggests caution in the interpretation of changes in the molecular weight or conformation of DNA based solely on analysis of sedimentation profiles.

Keywords: duplex DNA, electron microscopy, anomalous sedimentation, DNA complex, DNA conformation

Full text

PDF
3660

Images in this article

3662Image
on p.3662

Selected References

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

  1. Ahnström G., Erixon K. Radiation induced strand breakage in DNA from mammalian cells. Strand separation in alkaline solution. Int J Radiat Biol Relat Stud Phys Chem Med. 1973 Mar;23(3):285–289. doi: 10.1080/09553007314550311. [DOI] [PubMed] [Google Scholar]
  2. Alberts B. M. Characterization of a naturally occurring, cross-linked fraction of DNA. II. Origin of the cross-linkage. J Mol Biol. 1968 Mar 14;32(2):405–421. doi: 10.1016/0022-2836(68)90018-1. [DOI] [PubMed] [Google Scholar]
  3. Alberts B. M., Doty P. Characterization of a naturally occurring, cross-linked fraction of DNA. 1. Nature of the cross-linkage. J Mol Biol. 1968 Mar 14;32(2):379–403. doi: 10.1016/0022-2836(68)90017-x. [DOI] [PubMed] [Google Scholar]
  4. Belli J. A., Cooper S., Brown J. A. Sedimentation properties of mammalian-cell DNA: evidence that non-specific molecular aggregation does not occur during cell lysis. Int J Radiat Biol Relat Stud Phys Chem Med. 1972 Jun;21(6):603–606. doi: 10.1080/09553007214550701. [DOI] [PubMed] [Google Scholar]
  5. Belli J. A. Daughter cell repair by mammalian cells in culture after potentially lethal radiation damage. Nat New Biol. 1971 Sep 8;233(36):47–48. doi: 10.1038/newbio233047a0. [DOI] [PubMed] [Google Scholar]
  6. Belli J. A., Shelton M. Potentially lethal radiation damage: repair by mammalian cells in culture. Science. 1969 Aug 1;165(3892):490–492. doi: 10.1126/science.165.3892.490. [DOI] [PubMed] [Google Scholar]
  7. Cole R. S. Repair of DNA containing interstrand crosslinks in Escherichia coli: sequential excision and recombination. Proc Natl Acad Sci U S A. 1973 Apr;70(4):1064–1068. doi: 10.1073/pnas.70.4.1064. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Comings D. E. The rationale for an ordered arrangement of chromatin in the interphase nucleus. Am J Hum Genet. 1968 Sep;20(5):440–460. [PMC free article] [PubMed] [Google Scholar]
  9. Edwards P. A., Shooter K. V. Sedimentation characteristics of DNA multiply crosslinked by a difunctional alkylating agent, mustard gas. Biopolymers. 1971 Nov;10(11):2079–2082. doi: 10.1002/bip.360101105. [DOI] [PubMed] [Google Scholar]
  10. Elkind M. M., Chang-Liu C. M. Repair of a DNA complex from x-irradiated Chinese hamster cells. Int J Radiat Biol Relat Stud Phys Chem Med. 1972 Jul;22(1):75–90. doi: 10.1080/09553007214550801. [DOI] [PubMed] [Google Scholar]
  11. Elkind M. M., Kamper C. Two forms of repair of DNA in mammalian cells following irradiation. Biophys J. 1970 Mar;10(3):237–245. doi: 10.1016/S0006-3495(70)86296-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Elkind M. M. Sedimentation of DNA released from Chinese hamster cells. Biophys J. 1971 Jun;11(6):502–520. doi: 10.1016/S0006-3495(71)86231-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. GEIDUSCHEK E. P. "Reversible" DNA. Proc Natl Acad Sci U S A. 1961 Jul 15;47:950–955. doi: 10.1073/pnas.47.7.950. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. GOLDER R. H., MARTIN-GUZMAN G., JONES J., GOLDSTEIN N. O., ROTENBERG S., RUTMAN R. J. EXPERIMENTAL CHEMOTHERAPY STUDIES. 3. PROPERTIES OF DNA FROM ASCITES CELLS TREATED IN VIVO WITH NITROGEN MUSTARD. Cancer Res. 1964 Jul;24:964–968. [PubMed] [Google Scholar]
  15. HOTTA Y., BASSEL A. MOLECULAR SIZE AND CIRCULARITY OF DNA IN CELLS OF MAMMALS AND HIGHER PLANTS. Proc Natl Acad Sci U S A. 1965 Feb;53:356–362. doi: 10.1073/pnas.53.2.356. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Humphrey R. M., Steward D. L., Sedita B. A. DNA-strand breaks and rejoining following exposure of synchronized Chinese hamster cells to ionizing radiations. Mutat Res. 1968 Nov-Dec;6(3):459–465. doi: 10.1016/0027-5107(68)90063-8. [DOI] [PubMed] [Google Scholar]
  17. Lawley P. D., Brookes P. Interstrand cross-linking of DNA by difunctional alkylating agents. J Mol Biol. 1967 Apr 14;25(1):143–160. doi: 10.1016/0022-2836(67)90285-9. [DOI] [PubMed] [Google Scholar]
  18. Lehmann A. R., Ormerod M. G. Artefact in the measurement of the molecular weight of pulse labelled DNA. Nature. 1969 Mar 15;221(5185):1053–1056. doi: 10.1038/2211053b0. [DOI] [PubMed] [Google Scholar]
  19. Lett J. T., Caldwell I., Dean C. J., Alexander P. Rejoining of x-ray induced breaks in the DNA of leukaemia cells. Nature. 1967 May 20;214(5090):790–792. doi: 10.1038/214790a0. [DOI] [PubMed] [Google Scholar]
  20. Lett J. T., Klucis E. S., Sun C. On the size of the DNA in the mammalian chromosome. Structural subunits. Biophys J. 1970 Mar;10(3):277–292. doi: 10.1016/S0006-3495(70)86300-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lett J. T., Sun C. The production of strand breaks in mammalian DNA by X-rays: at different stages in the cell cycle. Radiat Res. 1970 Dec;44(3):771–787. [PubMed] [Google Scholar]
  22. McBurney M. W., Graham F. L., Whitmore G. F. Anomalous sedimentation of high molecular weight denatured mammalian DNA. Biochem Biophys Res Commun. 1971 Jul 2;44(1):171–177. doi: 10.1016/s0006-291x(71)80174-2. [DOI] [PubMed] [Google Scholar]
  23. McBurney M. W., Graham F. L., Whitmore G. F. Sedimentation analysis of DNA from irradiated and unirradiated L-cells. Biophys J. 1972 Apr;12(4):369–383. doi: 10.1016/S0006-3495(72)86090-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. Moroson H., Furlan M. An improvement in alkaline sucrose density gradient sedimentation of mammalian cell DNA. Radiat Res. 1970 Dec;44(3):713–726. [PubMed] [Google Scholar]
  26. Ormerod M. G., Lehmann A. R. Artefacts arising from the sedimentation of high molecular weight DNA on sucrose gradients. Biochim Biophys Acta. 1971 Oct;247(3):369–372. doi: 10.1016/0005-2787(71)90021-9. [DOI] [PubMed] [Google Scholar]
  27. Ormerod M. G., Lehmann A. R. The release of high molecular weight DNA from a mammalian cell (L-5178Y). Attachment of the DNA to the nuclear membrane. Biochim Biophys Acta. 1971 Jan 28;228(2):331–343. [PubMed] [Google Scholar]
  28. Sasaki M. S., Norman A. DNA fibres from human lymphocyte nuclei. Exp Cell Res. 1966 Nov-Dec;44(2):642–645. doi: 10.1016/0014-4827(66)90474-5. [DOI] [PubMed] [Google Scholar]
  29. Shipley W. U., Elkind M. M. DNA damage and repair following irradiation: the effect of 5-bromodeoxyuridine in cultured Chinese hamster cells. Radiat Res. 1971 Oct;48(1):86–94. [PubMed] [Google Scholar]
  30. Simpson R. T. Modification of chromatin by trypsin. The role of proteins in maintainance of deoxyribonucleic acid conformation. Biochemistry. 1972 May 23;11(11):2003–2008. doi: 10.1021/bi00761a002. [DOI] [PubMed] [Google Scholar]
  31. Stonington O. G., Pettijohn D. E. The folded genome of Escherichia coli isolated in a protein-DNA-RNA complex. Proc Natl Acad Sci U S A. 1971 Jan;68(1):6–9. doi: 10.1073/pnas.68.1.6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Studier F. W. Effects of the conformation of single-stranded DNA on renaturation and aggregation. J Mol Biol. 1969 Apr;41(2):199–209. doi: 10.1016/0022-2836(69)90385-4. [DOI] [PubMed] [Google Scholar]
  33. Veatch W., Okada S. Radiation-induced breaks of DNA in cultured mammalian cells. Biophys J. 1969 Mar;9(3):330–346. doi: 10.1016/S0006-3495(69)86390-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Worcel A., Burgi E. On the structure of the folded chromosome of Escherichia coli. J Mol Biol. 1972 Nov 14;71(2):127–147. doi: 10.1016/0022-2836(72)90342-7. [DOI] [PubMed] [Google Scholar]
  35. Yamamoto N., Naito T., Shimkin M. B. Mechanism of inactivation of DNA and RNA bacteriophages by alkylating agents in vitro. Cancer Res. 1966 Nov;26(11):2301–2306. [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES