RNAstructure 6.4 (12/8/2021)
- Added a new --stemloop option to calculate hairpin stemloop probabilities to ProbScan.
- Added a new --DMS mode to Rsample.
- Added --alphabet option to use alternative folding alphabets to AllSub, bifold, bipartition, and DuplexFold.
- Revised scorer so that it ignores nucleotide identities. This enables it to work on sequences with extended alphabets.
- Fixed a problem with m6A thermodynamic parameter files. Files were in dos text, and the dangling end free energies were not being correctly read on Linux.
RNAstructure 6.3 (4/5/2021)
- Added the m6A nearest neighbor parameters. These can be accessed using --alphabet m6A in most command line programs. Use M or 6 to represent m6A in sequences.
- Added the command line program SymmetryTester, which checks for proper symmetries in data tables.
- Improved the documentation about data tables.
- Biased selection of loop nucleotides and helix base pairs added to design.
- Added the capability to design to use extended alphabets (i.e. nucleotides in addition to A, C, G, U/T).
- Extended alphabets (--alphabet flag) added to Rsample.
- Local calculations were added for EDcalculator.
- Fixed historical numbering in CT files for long sequences (>16,000 nucleotides) on some machines.
- Added the --ct option to ETEcalculator. This allows the program to read a set of structures instead of generating them.
- Fixed the manual entry for orega to include all parameters and the objective function. Revised the objective function so that it is the one reported in our prior publication.
- Updated the documentation for building Multifind.
RNAstructure 6.2 (11/27/2019)
- The OREGA program was added as a command line program. This is a genetic algorithm that evolves an input sequence to have a longer end-to-end distance.
- The ETEcalculator program was added as a command line program to estimate the end-to-end distance of an input sequence.
- The TurboHomology command line program was added. This program does homology modeling for RNA secondary structure. It adds a new sequence to an existing multiple sequence alignment and estimates the secondary structure of the new sequence.
- ThreshKnot was added as an option to ProbKnot.
- The partition function was updated to work in the log(partition function) space. Previously, a scaling factor was used to prevent overflows or underflows. The log space is more robust to restraints and constraints.
- Fixed a problem with the JAVA GUI that prevented it from running on some Linux machines.
- Fixed a bug in Fold where it could miss the lowest free energy structure on rare occassions (for a window size set too large).
RNAstructure 6.1 (10/08/2018)
- The efn2 program now outputs folding free energy error estimates.
- Added capability to fold (and bifold) to predict structures with folding free energy change > 0 kcal/mol. If the lowest free energy structure with pairs has folding free energy change > 0 kcal/mol, the predicted lowest free energy structure will be a structure without pairs (with free energy change of 0 kcal/mol). Suboptimal structures can have pairs and folding free energy change > 0 kcal/mol.
- Updated the DNA enthalpy change parameters for tri-loops and tetra-loops. The new values are based on experimental data in the literature. This fixes a bug that sometimes prevented DNA structure prediction at temperatures other than 37 degrees C.
- Added the ability to read files from STDIN and write to STDOUT when passing "-" (hyphen) for the filename. This currently works for several programs including Fold and partition.
- Updated interfaces for Fold, partition, ct2dot, and dot2ct to include the "--quiet" flag to suppress progress or other verbose output.
- TurboFold can now accept multi-sequence FASTA files as input. The output CT file names are generated from the sequence names in the FASTA file.
- In StructureEditor, added ability to move number-labels on nucleobases by clicking and dragging with the mouse.
RNAstructure 6.0.1 Minor Update (02/12/2018)
- When using --MFE in Fold, the energy of the minimum free energy structure is now shown in the output CT file.
- Improved RNAstructure repository infrastructure, including Makefile organization, regression testing, and GUI testing.
- Updated the OligoWalk Text interface. The new version uses our standard command-line parser and both text and html formatted report output are available.
- Improved the Python Interface. It now supports Python 3.x. The pre-compiled extension is now provided in the binary distributions of RNAstructure. It should also be easier to compile the interface on all supported operating systems. Also updated and improved Python documentation in the manual.
- Added two new options to draw: The ability to specify a range of structures to draw (e.g. 3-7) and the ability to limit the total number of structures drawn.
- Fixed a bug in the Java GUI that prevented the Stochastic and OligoWalk tools from running when START was clicked.
- Fixed a minor bug in the energy calculation of flush coaxial stacking in exerior loops (ergexterior). The bug caused very small energy differences in rare circumstances.
RNAstructure 6.0 (09/21/2017)
- RNAstructure now can predict secondary structures for
sequences of any size alphabet. For example, an inosine
could be included along with RNA A, C, G, and U. To
accomodate this, a new data table format was implemented for
storing thermodynamic parameters. At this time, we have no
complete set of nearest neighbor rules for alphabets other
than standard DNA or standard RNA.
- TurboFold is upgraded to TurboFold II, which now predicts
alignments in addition to conserved RNA structures for a set
of input RNA homologs. TurboFold also can use Rsample to
make use of SHAPE mapping data for one or more homologs.
- Rsample is added. This program takes a
sequence and SHAPE mapping data and outputs a partition
function save file, which can then be used to model the
structure of a sequence that populates multiple
- CycleFold added. This program takes an RNA sequence predicts
both canonical and non-canonical base pairs or base pairing
- EDcalculator added. This program takes a sequence and a desired
secondary structure and calculates the ensemble defect for
that sequence to fold to the specific structure.
- StructureEditor added. This JAVA program
edits structure drawings and outputs publication-quality
drawings. This also can edit the underlying base pairing
structure and sequence.
- The –x option was added to ShapeKnots for incorporating
experimentally-determined pairing bonuses.
- Most RNAstructure programs that accept CT files as input
can now also accept Dot-Bracket
(dbn) files. The type of structure input file is
auto-detected, so no additional action is necessary on the
- RemovePseudoknots now has a much faster
pseudoknot analysis method in the -m
(Maximize Basepairs) mode.
- efn2 can now calculate the energies of
- Added validate program which helps in
automated input file validate for multiple file types
including CT, DotBracket, SEQ, FASTA, etc.
- Java GUI Enhancements:
- The native library is now compiled with SMP modules so
calculations are now performed in "parallel mode" (i.e.
using multiple threads).
- Improved Sequence "Speak" sound files (• Removed clicks
and gaps at beginning and end of files • Increased tempo
for shorter sounds • Added alert sound for invalid
characters • softened beginning of "C" to sound less like
- Added new menu "Structure Prediction" that combines
several structure analysis workflows for predicting single
structures, bimolecular structures, and structures common
to two or more sequences.
- Fixed a bug in dynalign_ii that resulted in a segfault
immediately after the progress indicator reached 100%
(i.e. during tracebacks).
- Fixed a bug where the coaxstack.dat (mismatch-mediated
coaxial stacking) table was assumed to be symmetric. This
can affect the energy output of efn2, Multilign,
oligoscreen, and OligoWalk.
- Fixed a bug where setting window size >0 in
AccessFold would lead to seg fault.
- Fixed a bug in Fold where using the --DMS option was not
working as expected because the underlying gamma
distribution parameters for modeling the pseudo free
energy change were not being correctly stored and read. A
new test case was added to check for bugs going forward.
- Fixed a bug where --DMS was an option for ShapeKnots,
but the DMS data was not being used.
- NAPSS, a program that uses NMR
experimental data to improve structure prediction, has
been updated to the version described in 10.1021/acs.biochem.5b00833.
(Previously an older version was distributed by mistake.)
The program has been updated further in conjunction with
other changes to the RNAstructure codebase.
- RemovePseudoknots no longer modifies structure labels
(previously it rewrote the structure energies, but did so
incorrectly). Users can use the efn2 program to more
accurately calculate the energies of structures with and
- The –x option in Fold and partition now
- The correct gamma distribution files used for DMS data
have now been added. (Previously these should have been
present, but were missing).
RNAstructure 5.8.1 Minor Update (06/06/2016)
- Fixed a bug with the Java GUI on Mac OS X not begin able
to set the DATAPATH variable.
- Released a new testing system for the Java GUI.
- Improved regression testing system for RNAstructure
- Example sequences are now mostly FASTA files instead
- Cleaned up whitespace and formatting in several RNA_class
- Java GUI Enhancements:
- Sequence Editor now allows copy/paste and does not
restrict keystrokes as before.
Text is color-coded to show invalid bases (red) and
lower-case bases (blue) to help highlight potential
- Users can now easily select Recent files
or Example files in prediction dialogs
- It is easier to run and monitor multiple concurrent
calculations because the progress indicator is now
displayed inside each parameter dialog (instead of as a
separate window disjointed from the main application).
- Some calculations (Fold, AllSub) can now be Canceled.
- Improved "Draw > Go to Structure.." dialog that
allows quick an easy scanning through available
- Added new backbone rendering styles for structure
drawing: "Render Circular Structure" and "Render Flat
- Improved About Window including function to extract
Example files (mostly useful for Mac App Bundle)
- Help menu can directly launch either local or online
- Java GUI now auto-detects the location of data_tables
and the native RNAstructure_GUI library, so setting these
on the command-line or environment are no longer
necessary. The jar can be executed directly. The GUI now
verifies the version of the native library to ensure
- Fixed detection of operating system to be compatible
with newer OS and Java versions.
- Deployment on Mac is now via DMG file for easier
- Fixed a bug that causes zooming issues in the structure
RNAstructure 5.8 (12/01/2015)
- Added design (to find a
sequence that will fold to a given structure).
- Added ProbScan (to
calculate loop and stack probabilities).
- Added functionality to draw
to annotate loop probabilities.
- Improved the end-to-end regression testing framework for
- Deprecated the native Windows GUI interface in favor of
the JAVA GUI on all platforms.
- Fixed a bug where efn2 was incorrectly calculating the
average asymmetry for some multibranch loops. Also, fixed a
bug that led to rounding errors. These changes can also
affect Multilign, OligoWalk, and ShapeKnots, which use efn2.
- Improved the Java GUI behavior when the native
RNAstructure library is not found or cannot be loaded. Now
an error message is shown and the application exits
gracefully. Previously it would crash.
- Improved the Java GUI behavior when the DATAPATH variable
is either not set or it points to an invalid location. Now
an informative message is shown at program startup.
Previously no message was shown until the user tried to
perform an action that required native access to the
DATAPATH, at which time a less informative message was
- Fixed bugs in the Java GUI:
- Printing structures and images did not work (only blank
pages were produced).
- The GUI would crash on startup if the splash window (or
any other program window) was still open when the main
application window was created.
- Typing a non-existant file name in the Open Sequence
window caused the program to freeze.
- Selecting a file with an upper- or mixed-case extension
resulted in a "File Not Found" error.
- The main menu became unresponsive when the toolbar was
moved to a floating window.
- The Java GUI DotPlotDialog window crashed when
reading in postscript legend colors under some
circumstances (such as when the colors were specified as
doubles instead of ints (e.g. 10.0 instead of 10).
RNAstructure 5.7 (12/4/2014)
- Added Dynalign II.
- Added AccessFold.
- Added phmm, a pairwise
Hidden Markov Model for RNA sequence alignment. This is in
the back ends to Dynalign, Dynalign II, PARTS, and
TurboFold. This is a new front end for sequence alignment.
- Added new help page that provides pathways
for using the command line interfaces.
- Added a flag (--simple) to efn2
so that the energies can be calculated for structures using
the same rules as the dynamic programming algorithms.
- Fixed a bug where specifying a constraint file (in any
program or OS) was removing prior unpairing constraints
specified in a sequence file.
- Fixed a bug in ShapeKnots. While filling in dangle or
double mismatches it created new non-canonical pairs, which
led to giving large positive energies to those structures.
- Improved ShapeKnots speed by removing boundaries checking
- Fixed a bug in the Windows GUI for bimolecular folding
where internal loops were intermittently being forbidden.
- Change the Macintosh compiler flags to match new Clang
compiler. Also, revised code to match the fact that OpenMP
is absent from Clang.
RNAstructure 5.6 (7/31/2013)
- Added Multifind to package (Linux and mac only).
- Added parallel processing (SMP) versions of bifold,
bipartition, efn2, Fold, MaxExpect, oligoscreen, partition,
and stochastic. These programs take advantage of multicore
- The Windows GUI now runs in parallel on multicore
computers for predicting lowest free energy structures,
calculating partition functions, calculating free energy
changes, oligoscreen calculations, and stochastic sampling.
- Fixed bug where command line programs were not recognizing
long flags without associated parameters. This also affected
the web servers.
- Fixed a bug with reading HMM alignment parameters for
Dynalign calculations in the Windows GUI. This was causing
the program to crash for Dynalign calculations.
- Fixed a bug where bootstraped restraints were not being
correctly calculated in Fold.
RNAstrucure 5.5 Update (5/16/2013)
- Uploaded a new version of the Windows 64-bit graphical
interface. This fixes problems with launching the program
from the start menu.
Online Documentation Update (4/16/2013)
- Added instructions for using Windows 8.
RNAstructure 5.5 (4/5/2013)
- Added ShapeKnots as text interface.
- Added GPU version of partition function (partition-cuda)
as text interface.
- Fixed minor memory access issues in NAPSS program.
- Fixed an error in Multilign where SHAPE intercept and
SHAPE slope weren't being set properly.
- Fixed bug in JAVA GUI where opening .seq files from disk
would sometimes fail.
- Updated Makefiles to build more efficiently and
standardized their formats.
- Simplified text interface parsing machinery and made
all flags case-insensitive.
- Cleaned up format, comments, and organization in all
text interfaces that use the standard parsing machinery.
- Updated dnacoaxstack.dat. Fixed problem where a subset
of mismatch-mediated coaxial stacks were missed in DNA.
- Fixed bug where not all special DNA tetraloops were
being read from disk.
- Updated default SHAPE slope and intercept for all
- Changed thresholds for SHAPE color annotations.
- Added support for knowledge-based free energy terms
for stacking restraints in Fold.
- Reduced the size of dot plot labels so they didn't run
together in certain instances.
- Made a small change to dot plot layout so dots at the
first nucleotide index always fall on the first grid
- Fixed a bug where sometimes plot bounds weren't set
right, so some dots weren't shown.
- Fixed a bug where sometimes the drawing windows
wouldn't resize properly when dragged.
- Added the ability to draw structures with their
backbone stretched out in a straight line.
- Improved drawing of structures so large structures are
drawn faster, and move and zoom more efficiently.
- Fixed a glitch in CircleCompare where sometimes the
probability annotation for the accepted structure was
being shown for all structures, not for each predicted
- Fixed a glitch in CircleCompare where probability
annotation was not always consistent in pairs.
- Added distinct flags to CircleCompare so the user can
annotation with either predicted or accepted structure
- Added the ability for structure drawing programs to
read probability information from dot plot text files as
- Added the ability for ProbabilityPlot to read dot data
from a standard dot plot file.
- Fixed mistake in TurboFold documentation where the
SHAPEslope and SHAPEintercept default values were
reversed. This reversal was only in the documentation.
Windows Text Interface Executables Update for
RNAstructure 5.4 (09/05/2012)
- Posted a new version of .zip files (both 32- and 64-bit),
which fixes an error with ProbabilityPlot, EnergyPlot, and
DynalignDotPlot where these programs would not work on all
Web Servers (08/28/2012)
- Improved CT file reading by making it more robust to
differences in format.
Mac OS X Executables Update for RNAstructure 5.4
- Fixed a bug where the Java GUI didn't always run properly.
Current executables run on Snow Leopard, Lion, and Mountain
Web Servers (08/13/2012)
- Fixed bug where pasted sequences without new lines at the
end sometimes caused calculation errors.
Web Servers (08/10/2012)
- Fixed bug in multiple sequence servers where a FASTA file
wasn't being uploaded and split into multiple sequences
- Changed wall time of run scripts from one hour to one day.
Web Servers (08/09/2012)
- Clarified error message that occurs when no sequence input
at all is given to a web server that takes sequence input.
- Fixed a bug in the Predict a Secondary Structure server
where the calculated partition function save file wasn't
being used for all calculations.
- Fixed a bug in the Multilign and TurboFold servers where
the proper error page wasn't being shown if too few or too
many sequences were given.
Web Servers (08/08/2012)
- Fixed broken links and style sheet references on page that
shows up if structures have no pairs.
RNAstructure 5.4 (08/02/2012)
- Added webservers for all programs.
- TurboKnot added to GUI as an option in TurboFold.
- Breakage of DNA pseudoknots, as well as pseudoknot
breakage options, added to JAVA GUI.
- Added 64-bit executables as downloads for Windows.
- Revised Windows GUI to conform with Windows 7 standards.
This also fixes issues with drawing dot plots from text
files and issues with showing oligonucleotide self-structure
from the OligoWalk interface.
- Fixed a bug in the Windows GUI, where pressing the F1 key
resulted in a "Failed to Launch Help" window in Windows XP
- Fixed command line bug in bifold interface where the
intramolecular flag did not always forbid intramolecular
pairs as it should have.
- Fixed command line bug in text interfaces where whole
numbers specified with zeros after a decimal point were
sometimes rejected as non-numeric. Negative numbers were
also sometimes rejected as non-numeric.
- Corrected the documentation for the Dynalign text
interface, where the default "DNA = false" should have been
written as "DNA = 0." Also, to set DNA folding, the user
should specify "DNA = 1."
- Fixed a typo in miscloop.dat and dnamiscloop.dat tables in
data_tables directory. The file stated "miscloop asym," but
now states "multiloop asym." This value is the multibranch
loop asymmetry penalty that is used by efn2. This did not
affect structure prediction or energy calculations, and the
change was made to simply clarify what thermodynamic
parameter this is.
- Corrected bug in JAVA GUI where the partition function
bimolecular module sometimes overwrote its second input
sequence file, rather than using the desired output name.
- Corrected bug in JAVA GUI where file chooser extension
sanity checks sometimes failed.
- Added an RNAstructure_java_utilities folder to facilitate
expansions in JAVA capabilities.
- Re-implemented internal workings of the JAVA GUI to aid in
organization and comprehension.
- Fixed a bug in Dynalign, where conserved 3' dangling ends
on helices at exterior loops were not allowed.
- Fixed a bug in extrapolation of free energies to
temperatures other than 37 degrees C.
- Fixed a dot plots bug where dots at the boundaries of
color ranges sometimes showed up as the wrong color or
- Reorganized Makefiles to build more efficiently and
include explicit dependencies.
- Standardized Postscript and SVG structure drawing so their
default is always clockwise orientation.
- Added a redone JAVA drawing framework, which includes the
ability to create self-contained JAVA drawing programs for
structures and dot plots.
- Simplified the controls in the JAVA drawing framework so
only the structure currently drawn can be exported to
- Added items where necessary to the JAVA drawing framework
so structures and dot plots can be exported as both
Postscript and SVG.
- Fixed a bug in JAVA drawing framework where the default
for structures wasn't clockwise orientation.
- Removed "New Window" and "Arrange Icons" options, which
did nothing, from the menus of the Windows GUI.
- Added ability to switch between structures and zoom in the
drawing framework using the command key (on Mac) as well as
the control key.
- Fixed the description of MAP mode in the PARTS text
RNAstructure 5.3 (05/10/2011)
- Added TurboFold to the package.
- Fixed a bug where drawing suboptimal structures did not
always work in the JAVA GUI for Mac and Linux.
- Fixed a bug where bimolecular partition functions were
missing some states (single dangling ends and no dangling
ends) at helix ends.
- Added the ability to include SHAPE data when calculating
folding free energy changes with efn2 program.
- Added the ability to apply folding restraints for the
formation of specified base pairs.
- Corrected Windows and JAVA graphical interfaces for
OligoWalk to show the correct temperature of folding free
energy changes when a user adjusts the temperature of the
calculation. (Prior to this, free energies' changes were
always listed as being at 37 degrees C.)
- Added the Overview help pages to the manual.
- Corrected bug in JAVA GUI where the message bar at the
bottom of the screen didn't always update when a temperature
menu was clicked.
- Corrected bug in JAVA GUI in OligoScreen where file names
were being refreshed incorrectly. Also switched oligomer
type default to DNA, rather than RNA.
- Corrected bug in JAVA GUI of OligoWalk where
oligonucleotides were always RNA, even when DNA was
RNAstructure 5.2 (11/17/2010)
- Added Multilign to the package.
- Fixed an issue when breaking pseudoknots, but leaving the
maximum non-pseudoknotted pairs, where non-canonical pairs
in the input structure were mistakenly broken. This was also
an issue with the count of pseudoknotted pairs in the
- Added support for writing ct files for long sequences
- Removed GAIL rule from dnamiscloop.dat and dnamiscloop.dh.
- Made the internal loop asymmetry penalty a single constant
for DNA folding. This fixed an occassional bug with folding
by DNA rules.
- In Windows GUI only, changed default value for the minimum
helix length in ProbKnot from 2 to 3, which is the preferred
- Added color-annotation for single-stranded nucleotides in
- Added the ability to use DNA parameters to the Dynalign
command line interface.
- Added the ability to change the folding temperature in the
Dynalign command line interface.
- In the Windows GUI, set the default file choice to .ALI
when clicking the "Alignment File" button.
- In Dynalign, revised the model for using alignment data to
limit the extent of calculation. This can lead to a
different set of suboptimal structures than previous
versions. This also fixes a traceback error encountered in
- Added the EnsembleEnergy program to the command line
- Improved the font in the html help for the GUI.
RNAstructure 5.1 (06/30/2010)
- Changed the thermodynamic parameter for adding an
additional helix to a multibranch loop ("helix penalty" in
miscloop.dat). The value is now set to -0.6 kcal/mol, which
is the value determined by a linear regression fit to
experimental data. This value provides slighly higher
average accuracy for maximum expected accuracy structure
prediction (MaxExpect and ProbKnot programs). This change
means that predicted structures might be different than
those predicted by previous version of RNAstructure. Base
pair probabilities, however, should be highly similar to
predictions from previous versions of the program.
- Added ProbKnot program to text interfaces and GUIs.
- Added online help for text (command line interfaces).
- Restructured downloads so that the text interface binaries
now also come with complete repository, including help and
- Added circular drawings to GUIs for drawing structures
that contain pseudoknots.
- Added the ability to read FASTA-formatted sequence files.
- Updated online help for GUIs to better reflect the current
state of the software.
- Fixed problems with Dynalign dot plots.
- Fixed problem reading SHAPE data in JAVA GUI.
RNAstructure 5.03 (03/22/2010)
- Added the automatic generation of a title for new
sequences in the GUIs (both Windows and JAVA). This fixes a
problem where RNAstructure fails to recognize a sequence
because it contained no title.
- Added CircleCompare (comparision of two secondary
structures, including pseudoknots).
- Fixed the grayed "forbid unimolecular pairs" menu option
when performing bimolecular folding with the Windows GUI.
RNAstructure 5.02 (01/22/2010)
- Revised PARTS to make it more user friendly.
- Fixed problem where precompiled text interfaces could not
find the shared library on some machines and therefore not
RNAstructure 5.01 (12/30/2009)
- Added the ability to read FASTA-formatted sequences to
- Added the Dynalign Intel C++ project for Microsoft Visual
Studio to download.
- Added the Fold Intel C++ project for Microsoft Visual
Studio to download.
- Changed the default parameters for SHAPE data pseudo
energies to match best practices.
- Cosmetic changes made to a number of the text interfaces.
- Fixed a problem where text interfaces on Linux and Mac
could not find the shared library and therefore did not run.
- 09/30/2009: New major update of RNAstructure. This version
has a graphical user interface in JAVA for use on Mac and
Linux machines. Text interfaces are now available for DOS,
Linux, and Mac OS X. A class library is available for
programmers. All source code is available for download.
- 10/03/2009: New Windows text interface binaries posted.
This fixes the problem that manifests with the error "This
application has failed to start because the application
configuration is incorrect," which appeared on some