Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1976 Jan;17(1):227–238. doi: 10.1128/jvi.17.1.227-238.1976

Restriction Enzyme Digests of Rapidly Renaturing Fragments of Vaccinia Virus DNA

Frank M DeFilippes 1
PMCID: PMC515407  PMID: 1206800

Abstract

Vaccinia virus DNA fragments that have been denatured by alkali and then neutralized contain a fraction that rapidly reforms duplex structures. The fraction is enriched by fractionating on hydroxyapatite columns and serves as a substrate for digestion by two restriction endonucleases isolated from Hemophilus parainfluenzae, Hpa I and HPa II. The patterns obtained by gel electrophoresis of the digested fragments show the presence of three major bands after Hpa I digestion and four major bands after Hpa II digestion. The DNA that is isolated from some of these bands quickly reforms duplex regions after alkaline denaturation. The size of the DNA segments in the major bands has been estimated to be in the range of 0.44 × 106 to 3.2 × 106 daltons. The fragments which rapidly reform duplex chains after denaturation are sensitive to single-strand-specific nucleases. These results are consistent with a model of vaccinia virus DNA which has a covalent link connecting complementary chains.

Full text

PDF
227

Images in this article

232Image
on p.232
233Image
on p.233

Selected References

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

  1. 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]
  2. Berns K. I., Silverman C. Natural occurrence of cross-linked vaccinia virus deoxyribonucleic acid. J Virol. 1970 Mar;5(3):299–304. doi: 10.1128/jvi.5.3.299-304.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Danna K. J., Sack G. H., Jr, Nathans D. Studies of simian virus 40 DNA. VII. A cleavage map of the SV40 genome. J Mol Biol. 1973 Aug 5;78(2):363–376. doi: 10.1016/0022-2836(73)90122-8. [DOI] [PubMed] [Google Scholar]
  4. DeFilippes F. M. A new method for isolation of a restriction enzyme from Hemophilus parainfluenzae. Biochem Biophys Res Commun. 1974 Jun 4;58(3):586–596. doi: 10.1016/s0006-291x(74)80460-2. [DOI] [PubMed] [Google Scholar]
  5. Defilippes F. M. In vitro RNA synthesis from unique pieces of simian virus 40 DNA produced by a restriction endonuclease. Biochim Biophys Acta. 1972 Jun 22;272(1):125–129. doi: 10.1016/0005-2787(72)90040-8. [DOI] [PubMed] [Google Scholar]
  6. Fairbanks G., Jr, Levinthal C., Reeder R. H. Analysis of C14-labeled proteins by disc electrophoresis. Biochem Biophys Res Commun. 1965 Aug 16;20(4):393–399. doi: 10.1016/0006-291x(65)90589-9. [DOI] [PubMed] [Google Scholar]
  7. Geshelin P., Berns K. I. Characterization and localization of the naturally occurring cross-links in vaccinia virus DNA. J Mol Biol. 1974 Oct 5;88(4):785–796. doi: 10.1016/0022-2836(74)90399-4. [DOI] [PubMed] [Google Scholar]
  8. Goodman N. C., Gulati S. C., Redfield R., Spiegelman S. Room-temperature chromatography of nucleic acids on hydroxylapatite columns in the presence of formamide. Anal Biochem. 1973 Mar;52(1):286–299. doi: 10.1016/0003-2697(73)90351-5. [DOI] [PubMed] [Google Scholar]
  9. Griffin B. E., Fried M., Cowie A. Polyoma DNA: a physical map. Proc Natl Acad Sci U S A. 1974 May;71(5):2077–2081. doi: 10.1073/pnas.71.5.2077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Griffin B. E., Fried M., Cowie A. Polyoma DNA: a physical map. Proc Natl Acad Sci U S A. 1974 May;71(5):2077–2081. doi: 10.1073/pnas.71.5.2077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Helling R. B., Goodman H. M., Boyer H. W. Analysis of endonuclease R-EcoRI fragments of DNA from lambdoid bacteriophages and other viruses by agarose-gel electrophoresis. J Virol. 1974 Nov;14(5):1235–1244. doi: 10.1128/jvi.14.5.1235-1244.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. JOKLIK W. K. The preparation and characteristics of highly purified radioactively labelled poxvirus. Biochim Biophys Acta. 1962 Aug 20;61:290–301. doi: 10.1016/0926-6550(62)90091-9. [DOI] [PubMed] [Google Scholar]
  13. MARMUR J., GROSSMAN L. Ultraviolet light induced linking of deoxyribonucleic acid strands and its reversal by photoreactivating enzyme. Proc Natl Acad Sci U S A. 1961 Jun 15;47:778–787. doi: 10.1073/pnas.47.6.778. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Moss B., Salzman N. P. Sequential protein synthesis following vaccinia virus infection. J Virol. 1968 Oct;2(10):1016–1027. doi: 10.1128/jvi.2.10.1016-1027.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Pogo B. G., Dales S. Two deoxyribonuclease activities within purified vaccinia virus. Proc Natl Acad Sci U S A. 1969 Jul;63(3):820–827. doi: 10.1073/pnas.63.3.820. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sharp P. A., Sugden B., Sambrook J. Detection of two restriction endonuclease activities in Haemophilus parainfluenzae using analytical agarose--ethidium bromide electrophoresis. Biochemistry. 1973 Jul 31;12(16):3055–3063. doi: 10.1021/bi00740a018. [DOI] [PubMed] [Google Scholar]
  17. Vogt V. M. Purification and further properties of single-strand-specific nuclease from Aspergillus oryzae. Eur J Biochem. 1973 Feb 15;33(1):192–200. doi: 10.1111/j.1432-1033.1973.tb02669.x. [DOI] [PubMed] [Google Scholar]
  18. Wilson D. A., Thomas C. A., Jr Hydroxyapatite chromatography of short double-helical DNA. Biochim Biophys Acta. 1973 Dec 21;331(3):333–340. doi: 10.1016/0005-2787(73)90019-1. [DOI] [PubMed] [Google Scholar]
  19. Wilson D. A., Thomas C. A., Jr Palindromes in chromosomes. J Mol Biol. 1974 Mar 25;84(1):115–138. doi: 10.1016/0022-2836(74)90216-2. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES