Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1975 Oct;2(10):1911–1929. doi: 10.1093/nar/2.10.1911

Hybridization of labeled RNA to DNA in agarose gels.

T M Shinnick, E Lund, O Smithies, F R Blattner
PMCID: PMC343556  PMID: 1103090

Abstract

Specific DNA restriction endonuclease fragments can be identified after electrophoresis in agarose gels by hybridization in the gel (in situ) to radioactive homologous RNA. RNA-DNA hybrids are detected by autoradiography of the gel. Comparison of band patterns of the autoradiogram and the ethidium bromide stained gel allows the identification of the DNA fragment which is complementary to the RNA probe. The technique is rapid, easy and inexpensive. It is sensitive enough to detect individual genes in a mixture of fragments produced by restriction enzyme digestion of complex cellular DNA. We have used this technique to determine which of the Hin III and Eco R1 fragments of phi80d3ilv+su+7 and E. coli DNAs contain the 5S, 16S and 23S ribosomal RNA (rRNA) genes of E. coli.

Full text

PDF
1913

Images in this article

1918Image
on p.1918
1920Image
on p.1920
1921Image
on p.1921
1924Image
on p.1924
1926Image
on p.1926

Selected References

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

  1. Blattner F. R., Dahlberg J. E. RNA synthesis startpoints in bacteriophage lambda: are the promoter and operator transcribed? Nat New Biol. 1972 Jun 21;237(77):227–232. doi: 10.1038/newbio237227a0. [DOI] [PubMed] [Google Scholar]
  2. Danna K., Nathans D. Specific cleavage of simian virus 40 DNA by restriction endonuclease of Hemophilus influenzae. Proc Natl Acad Sci U S A. 1971 Dec;68(12):2913–2917. doi: 10.1073/pnas.68.12.2913. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ehresmann C., Stiegler P., Fellner P., Ebel J. P. The determination of the primary structure of the 16S ribosomal RNA of Escherichia coli. 2. Nucleotide sequences of products from partial enzymatic hydrolysis. Biochimie. 1972;54(7):901–967. doi: 10.1016/s0300-9084(72)80007-5. [DOI] [PubMed] [Google Scholar]
  4. Fry M., Artman M. Deoxyribonucleic acid-ribonucleic acid hybridization. Annealing and quantitative recovery of intact ribosomal ribonucleic acid molecules from hybrids. Biochem J. 1969 Nov;115(2):287–294. doi: 10.1042/bj1150287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. Hedgpeth J., Goodman H. M., Boyer H. W. DNA nucleotide sequence restricted by the RI endonuclease. Proc Natl Acad Sci U S A. 1972 Nov;69(11):3448–3452. doi: 10.1073/pnas.69.11.3448. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. 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]
  8. Ikemura T., Dahlberg J. E. Small ribonucleic acids of Escherichia coli. I. Characterization by polyacrylamide gel electrophoresis and fingerprint analysis. J Biol Chem. 1973 Jul 25;248(14):5024–5032. [PubMed] [Google Scholar]
  9. Kelly T. J., Jr, Smith H. O. A restriction enzyme from Hemophilus influenzae. II. J Mol Biol. 1970 Jul 28;51(2):393–409. doi: 10.1016/0022-2836(70)90150-6. [DOI] [PubMed] [Google Scholar]
  10. Kossman C. R., Stamato T. D., Pettijohn D. E. Tandem synthesis of the 16S and 23S ribosomal RNA sequences of Escherichia coli. Nat New Biol. 1971 Nov 24;234(47):102–104. doi: 10.1038/newbio234102a0. [DOI] [PubMed] [Google Scholar]
  11. Otsubo E., Soll L., Deonier R. C., Lee H. J., Davidson N. Electron microscope heteroduplex studies of sequence relations among plasmids of Escherichia coli. VIII. The structure of bacteriophage phi 80d-3ilv+su+7, including the mapping of the ribosomal RNA genes. J Mol Biol. 1974 Nov 15;89(4):631–646. doi: 10.1016/0022-2836(74)90040-0. [DOI] [PubMed] [Google Scholar]
  12. Tanaka T., Weisblum B. Construction of a colicin E1-R factor composite plasmid in vitro: means for amplification of deoxyribonucleic acid. J Bacteriol. 1975 Jan;121(1):354–362. doi: 10.1128/jb.121.1.354-362.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES