This help page contains usage instructions and documentation. Helpful explanations of melting concepts can also be found in the glossary. For general info about this website, conditions of use and litterature references, see the about page.
You must specify a DNA sequence in one of three ways (remember to click the radio button):
A stitch profile describes the conformations of a DNA at a given temperature. Therefore, you must select one of two possibilities (remember to click the radio button):
Futhermore, two parameters are needed that control the search by the probability peak finding algorithm that produces a stitch profile. You can change the two parameters, the maximum depth and the probability cutoff, or you can simply accept the default values: Dmax=3 and pc=0.01. The peak finding algorithm and the roles of the two parameters are further described in the introductionary article Tøstesen (2005).
Here is a tip about the probability cutoff: If you want to do more versions of a stitch profile, the only difference being their probability cutoff values pc, then you can first submit the lowest pc value to obtain one of the plots. From the results page with that plot, you can then submit another pc-value (which should be greater than the original value). This redraws the plot using the new cutoff, which is faster than going back to the input page to submit a new calculation, and the result is identical. The cutoff simply determines what stitches to include and what to exclude from the plot. In this way, you can also adjust the visual appearence of the stitch profile to make it most informative.
The helicity is a function of the temperature. In principle, any interval of this function can be plotted, but the interesting part is the melting range of temperatures where the function drops from 1 (everything is helix) to 0 (everything is melted). Usually, the melting range is somewhere within 40°C - 110°C, depending on the sequence and the salt concentration. The server can locate the melting range automatically, if you indicate an interval of helicities (a range on the y-axis instead of the x-axis). You have two choices (remember to click the radio button):
At present, a calculation of differentiated melting curves, i.e. -dθ/dT as a function of T (a curve with peaks), is not available.
Limitations: Max length of DNA sequence is 48kbp. This calculation will complete in ca. 3min. User should use much smaller sequence to get a good plot: ex. a sequence of length 50kbp can make (480/50) = 9 positions in a plot before going over maximum of 480kbp.
A probability profile depends on the temperature. A plot of a single probability profile describes the structure at a specific temperature only. In order to get structural information at different temperatures in the melting range, you may plot several probability profiles. The curves will be plotted in the same diagram. You have two choices (remember to click the radio button):
The probability level of a temperature profile is a value p between 0 and 1. The plot shows the temperatures at which the different regions of the sequence are basepaired with probability p. If, for example, p=0.5 then the regions are melted and basepaired with equal probabilities at the temperatures shown in the temperature profile. (This special case of a temperature profile is also called a Tm profile or stability map.) Regions that are more thermally stable have higher temperatures in the profile, and regions that are more thermally unstable have lower temperatures in the profile. A lower p-value gives higher temperatures and a higher p-value gives lower temperatures. The server calculates a temperature profile by first calculating a set of probability profiles at temperatures between 40°C - 110°C, and then interpolating between them. You can use the default 0.5 value, or you can type another value between 0 and 1.
Here you can change some thermodynamic and algorithmic settings, but the default settings are recommended.
Loop Entropy Factor: For the calculation of the classical melting curves, probability profiles and temperature profiles, you can choose between two versions of the DNA melting algorithm described by Tøstesen et al. (2003) using different methods for handling the loop entropy factor: