The binding of ethidium bromide with DNA: interaction with single- and double-stranded structures

The pH-induced helix-coil transition of DNA and its complexes with EtBr is carried out at acidic pH in a wide interval of change of concentration ratio of EtBr/DNA. The binding isotherms of EtBr on double and single-stranded DNA at pH = 7.0 and pH = 3.0 (t = 25(o)C) are obtained by absorption and fluorimetric methods. Binding constants (K) and number of bases (n), corresponding to one binding site were determined. Non fluorescent "strong" complex with ds-DNA at pH = 7.0 and t = 25(o)C as well as "strong" and "weak" complexes with ss-DNA at pH = 3.0 and t = 25(o)C are revealed.     

West's problem on equivariant hyperspaces and Banach-Mazur compacta

Let be a compact Lie group, a metric -space, and the hyperspace of all nonempty compact subsets of endowed with the Hausdorff metric topology and with the induced action of . We prove that the following three assertions are equivalent: (a) is locally continuum-connected (resp., connected and locally continuum-connected); (b) is a -ANR (resp., a -AR); (c) is an ANR (resp., an AR). This is applied to show that is an ANR (resp., an AR) for each compact (resp., connected) Lie group . If is a finite group, then is a Hilbert cube whenever is a nondegenerate Peano continuum. Let be the hyperspace of all centrally symmetric, compact, convex bodies , , for which the ordinary Euclidean unit ball is the ellipsoid of minimal volume containing , and let be the complement of the unique -fixed point in . We prove that: (1) for each closed subgroup , is a Hilbert cube manifold; (2) for each closed subgroup acting non-transitively on , the -orbit space and the -fixed point set are Hilbert cubes. As an application we establish new topological models for tha Banach-Mazur compacta and prove that and have the same -homotopy type.

     

The equivariant homotopy type of <Emphasis Type="Italic">G</Emphasis>-<Emphasis Type="Italic">ANR</Emphasis>’s for compact group actions

We prove that if G is a compact Hausdorff group then every G-ANR has the G-homotopy type of a G-CW complex. This is applied to extend the James–Segal G-homotopy equivalence theorem to the case of arbitrary compact group actions. The first author was supported in part by grant U42563-F from CONACYT (Mexico).     

Differential pulse voltammetric studies of ethidium bromide binding to DNA

The interaction of ethidium bromide (EtBr) with calf thymus DNA is investigated electrochemically with the use of differential pulse voltammetry (DPV) at two different ionic strengths of a solution (0.154 M and 0.02 M [Na+], pH 7.0). It is revealed that EtBr binds with DNA in more than one way. The appropriate values of constants (K) and number site sizes (n) of EtBr binding to DNA are determined. The values of binding constants are equal to 1.9 x 10(6) and 5.6 x 10(5) M(-1), and number site sizes to 9 and 3.6 for strong interactions at ionic strengths of solutions 0.02 and 0.154 M Na+ at 28 degrees C, respectively. For a weaker interaction, these parameters are equal to 7 x 10(4) and 8 x 10(4) M(-1) and 1.5 and 1 at the mentioned ionic strengths of solutions, respectively. Thus, EtBr interacts with DNA in more than one way--intercalative and electrostatic at low ionic strength, and semi-intercalative and electrostatic at a higher strength of the solution. These results are in good accordance with the ones obtained by spectroscopic (absorption and fluorimetric) methods.     

Precise mass measurement of a double-stranded 500 base-pair (309 kDa) polymerase chain reaction product by negative ion electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry

Electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICRMS) has been used to determine the mass of a double-stranded 500 base-pair (bp) polymerase chain reaction (PCR) product with an average theoretical mass of the blunt-ended (i.e. unadenylated) species of 308 859.35 Da. The PCR product was generated from the linearized bacteriophage Lambda genome which is a double-stranded template. Utilization of ethanol precipitation in tandem with a rapid microdialysis step to purify and desalt the PCR product was crucial to obtain a precise mass measurement. The PCR product (0.8 pmol/μL) was electrosprayed from a solution containing 75% acetonitrile, 25 mM piperidine, and 25 mM imidazole and was infused at a rate of 200 nL/min. The average molecular mass and the corresponding precision were determined using the charge-states ranging from 172 to 235 net negative charges. The experimental mass and corresponding precision (reported as the 95% confidence interval of the mean) was 309 406 +/- 27 Da (87 ppm). The mass accuracy was compromised due to the fact that the PCR generates multiple products when using Taq polymerase due to the non-template directed 3'-adenylation. This results in a mixture of three PCR products with nearly identical mass (i.e. blunt-ended, mono-adenylated and di-adenylated) with unknown relative abundances that were not resolved in the spectrum. Thus, the experimental mass will be a weighted average of the three species which, under our experimental conditions, reflects a nearly equal concentration of the mono- and di-adenylated species. This report demonstrates that precise mass measurements of PCR products up to 309 kDa (500 bp) can be routinely obtained by ESI-FTICR requiring low femtomole amounts. Copyright 1999 John Wiley & Sons, Ltd.     

Molecular interactions between benzimide trichloride (Hoechst 33258) and DNA in dimethyl sulfoxide aqueous solutions, according to spectroscopy data

Interaction between benzimide (Hoechst 33258, H33258) and calf thymus DNA in aqueous dimethyl sulfoxide is investigated by means of UV-Vis and fluorescence spectroscopy at a constant ratio (r) of the number of H33258 molecules and DNA base pairs. Melting curves of the DNA-H33258 complex are obtained from the temperature dependences of the normalized optical density and fluorescence intensity, and the melting temperatures of the complex are determined. It is shown that adding dimethyl sulfoxide (DMSO) lowers the complex’s melting temperature. It is concluded that a long wavelength shift of the fluorescence spectra occurs when the temperature is raised.