Changing Temperature of Calculations
To change the temperature at which calculations occur, choose "Temperature -> Set Temperature". A dialog will pop up showing the current temperature of calculations. This can be edited by changing the value in the text box and clicking "OK". If "Cancel" is selected, or an invalid temperature is entered, the temperature is not changed. Temperature changes should always be used with caution.
Changing the Maximum Pairing Distance
A maximum pairing distance can be specified by choosing "Force -> Maximum Pairing Distance" and clicking "Yes" for "Limit Distance Between Paired Bases", then entering the preferred distance. Nucleotides farther separated in sequence than the specified number will not be allowed to pair. The maximum distance constraint is not saved with other constraints.
Changing the Maximum Internal/Bulge Loop Size
The maximum size of bulge or internal loops can be by choosing "Maximum Loop -> Set Maximum Loop Size" on the menu. The default is 30, and this is sufficient for structure predictions.
Using Folding and Alignment Constraints
Reviewing and Resetting Constraints
To review current selections, choose "Force -> Show Current Constraints". To reset selections, select "Force -> Reset Current Constraints".
Note that neither of these options include SHAPE constraints.
Saving and Restoring Secondary Structure Prediction Constraints
Folding constraints can be saved for future use or restored from previous saves using the "Force -> Save Constraints" and "Force -> Restore Constraints" menu options, respectively. Constraint files are saved using the constraint file format. (See "Constraint File Format".)
As above, neither of these options include SHAPE constraints.
Helixes can be forced to occur in secondary structure prediction. After selecting the sequence to fold, choose the menu option "Force -> Base Pair". Under "Base 1", indicate the number of the 5 most base to be paired. Under "Base 2", indicate the 3 most base to be paired. Under "Helix Length", enter the number of base pairs long the forced helix must be.
For example, if 17 was indicated for the first base in pair, and 83 for the second base in pair, with a helix length of 5, a helix would occur with these base pairs: 17-83, 18-82, 19-81, 20-80, and 21-79.
Helixes can be specified as forbidden in secondary structure prediction. After selecting the sequence to fold, choose the menu option "Force -> Prohibit Base Pairs". Under "Base 1", indicate the number of the 5 most base of the forbidden helix. Under "Base 2", indicate the 3 most base of the forbidden helix. Under "Helix Length", enter the length of the helix that is to be forbidden.
For example, if 17 was indicated for the first base in pair, and 83 for the second base in pair, with a helix length of 5, the following base pairs would be prohibited: 17-83, 18-82, 19-81, 20-80, and 21-79.
Bases can be selected that should not pair in the secondary structure prediction. This can be done in one of two ways:
Bases can be selected that must be base paired. This is done with the "Force -> Double Stranded" menu. It is analogous to forcing a base to be single stranded.
Specifying Nucleotides Accessible to Chemical Modification
Nucleotides can be selected that are accessible to chemical modification. These nucleotides will then be single stranded. They can be located at the end of a helix, in a GU pair, or adjacent to a GU pair.
Select the "Force -> Chemical Modification" menu option. Into the "Base Number" box, enter the number indicating the position from the 5 end of a modified nucleotide. Click "OK" and the dialog will reset to take another nucleotide. Click "OK and Close" after all chemically modified nucleotides have been entered.
Specifying FMN Cleavage (U's in GU pairs)
Select the "Force -> FMN Cleavage" menu option. Into the "Base Number" box, enter the number indicating the position from the 5 end of a U in a GU pair. Click "OK" and the dialog will reset to take another nucleotide. Click "OK and Close" after all nucleotides have been entered.
See "SHAPE File Format" for details on the file type used for these constraints. Two methods exist to enter SHAPE data, as follows:
RNAstructure can output images from the "Draw" and "Dot Plot" modules as Postscript. Select one of the associated options to export these images.