Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1986 May 12;14(9):3883–3894. doi: 10.1093/nar/14.9.3883

There exists a distinct stage during mammalian DNA synthesis immediately after joining of replication intermediates.

U Lönn, S Lönn
PMCID: PMC339822  PMID: 3012484

Abstract

We describe an approach, using alkaline cell lysis and digestion with nuclease S1, which permits to distinguish between newly ligated DNA and the DNA of mature chromatin. When cells with steady-state labelled DNA (mature DNA) are analyzed, the results show labelled "nucleosomal-sized" DNA. However, when DNA of cells pulse-labelled with thymidine for 45 seconds is examined one can detect only large DNA. The newly ligated DNA is not reduced to "nucleosomal-sized" DNA by nuclease S1. When the large DNA is denatured in formamide one can detect 10 kb DNA fragments. Furthermore in pulse-chase experiments there appear, after formamide-treatment, increasing amounts of "nucleosomal-sized" DNA with a parallel decrease in the amount of 10 kb DNA fragments. Hence the newly ligated, large, DNA differs from mature DNA and represents a distinct stage during DNA replication.

Full text

PDF
3883

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. Ando T. A nuclease specific for heat-denatured DNA in isolated from a product of Aspergillus oryzae. Biochim Biophys Acta. 1966 Jan 18;114(1):158–168. doi: 10.1016/0005-2787(66)90263-2. [DOI] [PubMed] [Google Scholar]
  3. Annunziato A. T., Seale R. L. Chromatin replication, reconstitution and assembly. Mol Cell Biochem. 1983;55(2):99–112. doi: 10.1007/BF00673705. [DOI] [PubMed] [Google Scholar]
  4. Cockell M., Rhodes D., Klug A. Location of the primary sites of micrococcal nuclease cleavage on the nucleosome core. J Mol Biol. 1983 Oct 25;170(2):423–446. doi: 10.1016/s0022-2836(83)80156-9. [DOI] [PubMed] [Google Scholar]
  5. Cusick M. E., DePamphilis M. L., Wassarman P. M. Dispersive segregation of nucleosomes during replication of simian virus 40 chromosomes. J Mol Biol. 1984 Sep 15;178(2):249–271. doi: 10.1016/0022-2836(84)90143-8. [DOI] [PubMed] [Google Scholar]
  6. DePamphilis M. L., Wassarman P. M. Replication of eukaryotic chromosomes: a close-up of the replication fork. Annu Rev Biochem. 1980;49:627–666. doi: 10.1146/annurev.bi.49.070180.003211. [DOI] [PubMed] [Google Scholar]
  7. Hand R. Eucaryotic DNA: organization of the genome for replication. Cell. 1978 Oct;15(2):317–325. doi: 10.1016/0092-8674(78)90001-6. [DOI] [PubMed] [Google Scholar]
  8. McKnight S. L., Miller O. L., Jr Electron microscopic analysis of chromatin replication in the cellular blastoderm Drosophila melanogaster embryo. Cell. 1977 Nov;12(3):795–804. doi: 10.1016/0092-8674(77)90278-1. [DOI] [PubMed] [Google Scholar]
  9. Méchali M., Kearsey S. Lack of specific sequence requirement for DNA replication in Xenopus eggs compared with high sequence specificity in yeast. Cell. 1984 Aug;38(1):55–64. doi: 10.1016/0092-8674(84)90526-9. [DOI] [PubMed] [Google Scholar]
  10. Russev G., Tsanev R. Distribution of histones in alkali-denatured chromatin studied by isopycnic centrifugation in alkaline metrizamide density gradients. Nucleic Acids Res. 1976 Mar;3(3):697–707. doi: 10.1093/nar/3.3.697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Rydberg B. The rate of strand separation in alkali of DNA of irradiated mammalian cells. Radiat Res. 1975 Feb;61(2):274–287. [PubMed] [Google Scholar]
  12. Shenk T. E., Rhodes C., Rigby P. W., Berg P. Biochemical method for mapping mutational alterations in DNA with S1 nuclease: the location of deletions and temperature-sensitive mutations in simian virus 40. Proc Natl Acad Sci U S A. 1975 Mar;72(3):989–993. doi: 10.1073/pnas.72.3.989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Silber J. R., Loeb L. A. S1 nuclease does not cleave DNA at single-base mis-matches. Biochim Biophys Acta. 1981 Dec 28;656(2):256–264. doi: 10.1016/0005-2787(81)90094-0. [DOI] [PubMed] [Google Scholar]
  14. Stinchcomb D. T., Struhl K., Davis R. W. Isolation and characterisation of a yeast chromosomal replicator. Nature. 1979 Nov 1;282(5734):39–43. doi: 10.1038/282039a0. [DOI] [PubMed] [Google Scholar]
  15. Struhl K. The new yeast genetics. 1983 Sep 29-Oct 5Nature. 305(5933):391–397. doi: 10.1038/305391a0. [DOI] [PubMed] [Google Scholar]
  16. Taylor J. H. Origins of replication and gene regulation. Mol Cell Biochem. 1984;61(2):99–109. doi: 10.1007/BF00222489. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES