Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1993 Dec 11;21(24):5720–5726. doi: 10.1093/nar/21.24.5720

Heterodimeric DNA-binding dyes designed for energy transfer: stability and applications of the DNA complexes.

S C Benson 1, R A Mathies 1, A N Glazer 1
PMCID: PMC310540  PMID: 8284220

Abstract

Spectroscopic studies of the complexes of double-stranded (ds) DNA with the polymethylene-amine linked heterodimers thiazole orange-thiazole blue, thiazole orange-ethidium, and fluorescein-ethidium, in each case show efficient energy transfer from donor to acceptor chromophores (Benson, S.C., Singh, P. and Glazer, A.N. (1993) accompanying manuscript). A quantitative assay of the stability of such complexes during gel electrophoresis is presented. The off-rate of dye from complexes formed at an initial dsDNA bp:dye ratio > or = 10:1 follows strict first-order kinetics. The t0.5 values for the dissociation of a series of related dyes provide a quantitative criterion for the design of DNA-binding fluorophores. Complexes of dsDNA with the monomeric propidium and cyanine dyes, [1-(9-amino-4,7-diazanonyl)-3,8-diamino-6-phenyl-phenanthridinium bromide trihydrobromide] and (N,N'-tetramethyl-1,3-propanediamino)propyl thiazole orange [4-[3-methyl-2,3-dihydro-(benzo-1,3-thiazole)-2-methylidenyl]-1-(4 ,4,8-trimethyl-4,8-diazanonyl)-quinolinium diiodide], are much more stable than those with their widely used counterparts, ethidium and thiazole orange. Applications of the new dyes in post-staining of gels and in the multiplex detection of DNA restriction fragments are presented.

Full text

PDF
5720

Images in this article

5722Image
on p.5722
5723Image
on p.5723
5724Image
on p.5724

Selected References

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

  1. Glazer A. N., Peck K., Mathies R. A. A stable double-stranded DNA-ethidium homodimer complex: application to picogram fluorescence detection of DNA in agarose gels. Proc Natl Acad Sci U S A. 1990 May;87(10):3851–3855. doi: 10.1073/pnas.87.10.3851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Goodwin P. M., Johnson M. E., Martin J. C., Ambrose W. P., Marrone B. L., Jett J. H., Keller R. A. Rapid sizing of individual fluorescently stained DNA fragments by flow cytometry. Nucleic Acids Res. 1993 Feb 25;21(4):803–806. doi: 10.1093/nar/21.4.803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Lavery R., Pullman B. The dependence of the surface electrostatic potential of B-DNA on environmental factors. J Biomol Struct Dyn. 1985 Feb;2(5):1021–1032. doi: 10.1080/07391102.1985.10507618. [DOI] [PubMed] [Google Scholar]
  4. LePecq J. B., Paoletti C. A fluorescent complex between ethidium bromide and nucleic acids. Physical-chemical characterization. J Mol Biol. 1967 Jul 14;27(1):87–106. doi: 10.1016/0022-2836(67)90353-1. [DOI] [PubMed] [Google Scholar]
  5. Manning G. S. The molecular theory of polyelectrolyte solutions with applications to the electrostatic properties of polynucleotides. Q Rev Biophys. 1978 May;11(2):179–246. doi: 10.1017/s0033583500002031. [DOI] [PubMed] [Google Scholar]
  6. Quesada M. A., Rye H. S., Gingrich J. C., Glazer A. N., Mathies R. A. High-sensitivity DNA detection with a laser-excited confocal fluorescence gel scanner. Biotechniques. 1991 May;10(5):616–625. [PubMed] [Google Scholar]
  7. Rodríguez-Núez A., Camiña F., Lojo S., Rodríguez-Segade S., Castro-Gago M. Concentrations of nucleotides, nucleosides, purine bases and urate in cerebrospinal fluid of children with meningitis. Acta Paediatr. 1993 Oct;82(10):849–852. doi: 10.1111/j.1651-2227.1993.tb17625.x. [DOI] [PubMed] [Google Scholar]
  8. Rye H. S., Dabora J. M., Quesada M. A., Mathies R. A., Glazer A. N. Fluorometric assay using dimeric dyes for double- and single-stranded DNA and RNA with picogram sensitivity. Anal Biochem. 1993 Jan;208(1):144–150. doi: 10.1006/abio.1993.1020. [DOI] [PubMed] [Google Scholar]
  9. Rye H. S., Quesada M. A., Peck K., Mathies R. A., Glazer A. N. High-sensitivity two-color detection of double-stranded DNA with a confocal fluorescence gel scanner using ethidium homodimer and thiazole orange. Nucleic Acids Res. 1991 Jan 25;19(2):327–333. doi: 10.1093/nar/19.2.327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Rye H. S., Yue S., Quesada M. A., Haugland R. P., Mathies R. A., Glazer A. N. Picogram detection of stable dye-DNA intercalation complexes with two-color laser-excited confocal fluorescence gel scanner. Methods Enzymol. 1993;217:414–431. doi: 10.1016/0076-6879(93)17080-o. [DOI] [PubMed] [Google Scholar]
  11. Rye H. S., Yue S., Wemmer D. E., Quesada M. A., Haugland R. P., Mathies R. A., Glazer A. N. Stable fluorescent complexes of double-stranded DNA with bis-intercalating asymmetric cyanine dyes: properties and applications. Nucleic Acids Res. 1992 Jun 11;20(11):2803–2812. doi: 10.1093/nar/20.11.2803. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES