Denaturation, of DNA thermal

The interaction of tilorone hydrochloride with native DNA stabilizes the double helical structure of the macromolecule towards thermal denaturation. The effect of tilorone hydrochloride on the thermal denaturation of DNA s from various sources having different base composition has been studied28. At a drug to DNA-P molar ratio of 0.21, the Arm increased with increasing AT content of the DNA. This observation indicates that tilorone hydrochloride perferentially binds to the dAT portions of the DNA molecule. This is confirmed by the strong effect of tilorone hydrochloride on the thermal transition temperature of poly d (A-T), ATm = 29 °C. 

The effect of tilorone hydrochloride on the thermal denaturation of DNA s from various sources having different base composition is shown in Fig. 22. 

Thermal denaturation of DNA (UV melting curve). A solution of double-stranded DNA was placed in a spectrophotometer and the change in absorbance at 260 nm was measured as the temperature of the sample... 

Wada, A., Yabuki, S., Husimi, Y, 1980. Fine strncture in the thermal denaturation of DNA high temperature-resolution spectrophotometric stndies. CRC Crit. Rev. Biochem. 9 87-144. 

Fig. 1.15 Thermal denaturation of DNA (E. coli). Relative absorbance at 260 nm vs. temperature at various concentrations of KCI (given in the graph in units of mol L Y Adapted from Marmur et al. [41] with permission from Elsevier.

Maximization of immunoreactivity of heat stable adducts by thermal denaturation of DNA... 

Maximization of immunoreactivity of heat stable adducts by thermal denaturation of DNA 421 Maximization of immunoreactivity of heat labile adducts by digestion of DNA 421 Competitive ELISA 423 Analysis of data 425... 

B. Kessler, Interactions in vitro between gibberellins and DNA by optical rotatory profile of the thermal denaturation of DNA-gibberellin complexes, Biochim. Biophys. Acta 232, 611-613(1971). 

The effects of surface-active compounds on the thermal denaturation of DNA have been studied [90]. Anionic surfactants interact only weakly with DNA with the exception of N-lauroyl prolylprolylglycine and deoxycholic acid. Myristyl trimethylammonium chloride causes precipitation of DNA at the 1 mmoll level probably by interaction with the phosphate residues of the DNA. The conclusion of this preliminary study was that the structure of the surfactant rather than its surface activity was the determining factor for interaction and that typical surfactants were unlikely significantly to affect DNA structure. 

Raukas, E. Thermal denaturation of DNA complexes with protamine peptides. Eesti NSV Teaduste Akad. Toimetised, Biol. Seer. 15, 342—346 (1966). 

One of results of the binding of such compounds onto DNA chains is that the so called melting-temperature of the biopolymer is markedly increased. We have found that the stabilizing influence of spermine and spermidine against thermal denaturation of DNA and of t-RNA is, qualitatively, very similar. The results of equilibrium dialysis measurements have shown that also the free energy of binding of... 

The hybridization phase entails thermal denaturation of double-stranded DNA and incubation of the probe with the denatured DNA at a temperature 25°C below the melt temperature. Unless one is interested in partial homology, lower temperatures should be avoided. However, addition of formamide promotes the hybridization, thereby permitting the use of a lower temperature, if it should be required, to prevent structural modification. 

If your laboratory does not have access to a spectrophotometer with a thermoprogrammer, use the quick cool method described in steps 3 through 6 below to obtain a melting curve for your DNA. This method does not observe a true equilibrium between native and denatured ( melted ) DNA and is appreciably less accurate than the method described in step 1, but it will suffice to illustrate the principles underlying the thermal separation of DNA strands. 

Thermal denaturation of calf-thymus DNA has also been studied by SERS-spectro-scopy The examinations of the SER scattering of thermally denatured DNA indicates that the nucleic bases bands are sensitive to the termal transition from the helical double stranded structure to the disordered single stranded structure. In this thermally destabilized DNA the strands are open and the corresponding bases can easily re-orientate therfore becoming available for direct interaction with the surface. Generally at every adsorption potential there is a sensitive increase of the intensity of the Raman bands of the nucleic bases in the SERS spectra. In summary, one may say that SERS is useful to determine the structural changes of DNA under the action of physical or chemical disturbances. 

Apilux, A., Tabata, M., Chailapakul, O. (2007). Electrochemical behaviors of native and thermally denatured fish DNA in the presence of cytosine derivatives and porphyrin by cyclic voltammetry using boron-doped diamond electrode. Bioelectrochemistry 70, 435-439. 

The most widely used DNA polymerase in PCR is from Thermus acquaticus, the Taq polymerase. This enzyme has an optimal temperature for polymerization in the range of 70 to 75°C. It extends DNA chains at a rate of about 2 kb per minute. It is fairly resistant to the continual cycles of heating and cooling required for PCR. The half-life for thermal denaturation of Taq polymerase is 1.6h at 95°C. When very high denaturation temperatures are needed, as in the PCR amplification of very G+C-rich DNA, more thermal-stable polymerase such as the enzyme from Thermococcus litoralis with a half-life of 1.8h at 100°C or the enzyme from Pyrococcus furiosis with a half-life of 8 h at 100°C, can be employed. However, these enzymes are not as processive as Taq polymerase, which makes it more difficult to amplify long templates. 

Fritzsche (1966) has studied the helix-coil transition of calf thymus DNA in D2O. A plot of the ratio of absorbances at 1662 and 1682 cm during an examination of the thermal denaturation of the DNA gave a T of89.6 C for a 10% DNA solution. 

M. A. Rix-Montel, H. Grassi, and D. Vasilescu, Experimental Studies of Thermal Denaturation of the Na-DNA System with Respect to Manning s Model, Biophys. Chem. 2, 278-289 (1974). 

Fig. 9. Linear sweep voltammetry (LSV) at the HMDE of samples of double-helical and thermally denatured DNAs at the concentration of 0.1 mg/ml. Medium 0.1 M sodium phosphate, pH 7.1. Voltage scan rate of 1.0 V/s, waiting time at the initial potential Uj was 60 s. (A) Voltammograms upper curves, samples of thermally denatured DNA, lower curves, samples of double-helical DNA initial potentials Uj are indicated in the lower parts of individual panels. (B) Dependence of LSV peak III height of the sample of double-helical (o) and thermally denatured (d) DNAs on initial potential Ej. In this article the value of x (see top curve at LFj = -0.4 V in the panel A) was taken to represent the height of LSV peak III.

Closer examination of thermal denaturation reveals some of the chemical factors that determine protein and nucleic acid stability. For example, the thermal stability of DNA increases with the number of C-G base pairs in the sequence because each C-G base pair has three hydrogen bonds (1), whereris each T-A base pair has only two (2). More energy is required to unravel a double hehx that has a higher proportion of hydrogen bonding interactions per base pair. 

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