Interaction of diamidino-2-phenylindole (DAPI) with natural and synthetic nucleic acids

Manzini, Giorgio; Barcellona, Maria Luisa; Avitabile, M; Quadrifoglio, Franco

doi:10.1093/nar/11.24.8861

Cited by 152 publications

(111 citation statements)

References 32 publications

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“…In accordance with other literature reports (Manzini et al 1983(Manzini et al , 1985Szabo et al 1986;Gratton 1989, 1990), no``apparent''¯uorescence increase was observed for polyd(GC)/DAPI complexes at high P/D ratios (from 1000 to about 250), but rather there was a 40% decrease in the¯uorescence intensity, becoming equal to that of free DAPI in solution at a P/D ratio of 250 with a maximum emission at 462 nm (Fig. 2).…”

Section: Steady-state Measurementssupporting

confidence: 92%

4',6-Diamidino-2-phenylindole (DAPI) interacts with rare structures of GC polymers

Barcellona

Chen

Müller

et al. 2001

European Biophysics Journal

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Section: Steady-state Measurementssupporting

confidence: 92%

4',6-Diamidino-2-phenylindole (DAPI) interacts with rare structures of GC polymers

Barcellona

Chen

Müller

et al. 2001

European Biophysics Journal

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“…It has been previously shown that DAPI binds tightly to AT-rich regions (Manzini et al, 1983;Wilson et al, 1990) and it maintains a specific angle with respect to the helical axis (Barcellona and Gratton, 1996a,b) (Fig. 7).…”

Section: Methodsmentioning

confidence: 85%

Polarized fluorescence correlation spectroscopy of DNA‐DAPI complexes

Barcellona

Gammon

Hazlett

et al. 2004

Microscopy Res & Technique

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We discuss the use of fluorescence correlation spectroscopy for the measurement of relatively slow rotations of large macromolecules in solution or attached to other macromolecular structures. We present simulations and experimental results to illustrate the range of rotational correlation times and diffusion times that the technique can analyze. In particular, we examine various methods to analyze the polarization fluctuation data. We have found that by first constructing the polarization function and then calculating the autocorrelation function, we can obtain the rotational motion of the molecule with very little interference from the lateral diffusion of the macromolecule, as long as the rotational diffusion is significantly faster than the lateral diffusion. Surprisingly, for common fluorophores the autocorrelation of the polarization function is relatively unaffected by the photon statistics. In our instrument, two-photon excitation is used to define a small volume of illumination where a few molecules are present at any instant of time. The measurements of long DNA molecules labeled with the fluorescent probe DAPI show local rotational motions of the polymers in addition to translation motions of the entire polymer. For smaller molecules such as EGFP, the viscosity of the solution must be increased to bring the relaxation due to rotational motion into the measurable range. Overall, our results show that polarized fluorescence correlation spectroscopy can be used to detect fast and slow rotational motion in the time scale from microsecond to second, a range that cannot be easily reached by conventional fluorescence anisotropy decay methods.

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“…Energy transfer between identical molecules do not affect the lifetime of the excited state if all molecules are spectroscopically equivalent, but it can affect the decay of the emission anisotropy. In the case of DAPI bound to DNA, there are at least two different binding modalities, one in which DAPI is bound to AT sites and gives a fluorescent species and a second modality of binding in which DAPI is interacting also with GC pairs and gives a nonfluorescent species (the quantum yield is a factor of about 50 less than the AT-bound species) (Manzini et al, 1983(Manzini et al, , 1985bWilson et al, 1989Wilson et al, , 1990Tanious et al, 1994). The GC-bound species is populated only at very low P/D ratios.…”

Section: Depolarization Due To Energy Transfermentioning

confidence: 99%

“…The quantum yield increases by more than 50 between the bound and unbound form. A large number of thermodynamic and spectroscopic studies have been performed on the DNA-DAPI complex (Kapuscinski & Szer, 1979;Masotti et al, 1981Masotti et al, , 1982Manzini et al, 1983Manzini et al, , 1985aWilson et al, 1985aWilson et al, , 1989Szabo et al, 1986;Barcellona et al, 1986;Kubista et al, 1987). It is known that, in addition to binding to AT sites, DAPI can also bind to GC pairs, although with less affinity and without fluorescence enhancement.…”

mentioning

confidence: 99%

Torsional Dynamics and Orientation of DNA−DAPI Complexes

Barcellona

Gratton

1996

Biochemistry

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The flexibility of calf thymus DNA and several polynucleotides was measured using the anisotropy decay of DAPI bound to DNA, a minor groove probe. DNA torsional dynamics were analyzed using the Schurr model [Allison, S. A., & Schurr, J. M. (1979) Chem. Phys. 41, 35-44] in the infinite polymer length approximation. Time-resolved fluorescence depolarization was measured using a frequencydoubled mode-locked dye laser and frequency-domain acquisition methods. At very high P/D ratios, the anisotropy decay is dominated by DNA torsional dynamics. The recovered values of the torsional elastic constant were in good agreement with literature values obtained using other DNA probes. The exact knowledge of the angle between the probe emission dipole transition moment and the helix axis is critical for the determination of the polymer elastic constant. At low P/D ratios, energy transfer between dye molecules strongly contributes to the anisotropy decay. We have developed a statistical model that describes the anisotropy decay, when the correct geometrical factors are included. At low P/D ratios the anisotropy decay is dominated by fluorescence homotransfer. In this regime, it is possible to determine the orientation of the dye molecule with respect to the polymer with accuracy. The values obtained for the distance and orientation of the DAPI molecules in solution using the fluorescence measurements are in excellent agreement with those from the crystal structure of the oligonucleotides-DAPI complex by Dickerson's group [Larsen T.

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Interaction of diamidino-2-phenylindole (DAPI) with natural and synthetic nucleic acids

Cited by 152 publications

References 32 publications

4',6-Diamidino-2-phenylindole (DAPI) interacts with rare structures of GC polymers

4',6-Diamidino-2-phenylindole (DAPI) interacts with rare structures of GC polymers

Polarized fluorescence correlation spectroscopy of DNA‐DAPI complexes

Torsional Dynamics and Orientation of DNA−DAPI Complexes

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