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RNALFOLD(1) User Commands RNALFOLD(1) NAME RNALfold - manual page for RNALfold 2.7.2 SYNOPSIS RNALfold [OPTION]... DESCRIPTION RNALfold 2.7.2 calculate locally stable secondary structures of RNAs Compute locally stable RNA secondary structure with a maximal base pair span. For a sequence of length n and a base pair span of L the algorithm uses only O(n+L*L) memory and O(n*L*L) CPU time. Thus it is practical to "scan" very large genomes for short RNA structures. Output consists of a list of secondary structure components of size <= L, one entry per line. Each output line contains the predicted local structure its energy in kcal/mol and the starting position of the local structure. -h, --help Print help and exit --detailed-help Print help, including all details and hidden options, and exit --full-help Print help, including hidden options, and exit -V, --version Print version and exit -v, --verbose Be verbose. (default=off) Lower the log level setting such that even INFO messages are passed through. I/O Options: Command line options for input and output (pre-)processing -i, --infile=filename Read a file instead of reading from stdin The default behavior of RNALfold is to read input from stdin. Using this parameter the user can specify an input file name where data is read from. -o, --outfile[=filename] Print output to file instead of stdout. This option may be used to write all output to output files rather than printing to stdout. The number of output files created for batch input (multiple sequences) depends on three conditions: (i) In case an optional filename is given as parameter argument, a single file with the specified filename will be written into. If the op- tional argument is omitted, (ii) FASTA input or an active --auto-id switch will write to multiple files that follow the naming scheme "prefix.lfold". Here, "prefix" is taken from the sequence id as specified in the FASTA header. Lastly, (iii) single-line sequence input without FASTA header will be written to a single file "RNAL- fold_output.lfold". In case an output file already exists, any out- put of the program will be appended to it. Since the filename argu- ment is optional, it must immediately follow the short option flag to not be mistaken as new parameter to the program. For instance \'-ornafold.out\' will write to a file "rnafold.out". Note: Any special characters in the filename will be replaced by the filename delimiter, hence there is no way to pass an entire directory path through this option yet. (See also the "--filename-delim" parameter) --noconv Do not automatically substitute nucleotide "T" with "U". (default=off) --auto-id Automatically generate an ID for each sequence. (default=off) The default mode of RNALfold is to automatically determine an ID from the input sequence data if the input file format allows to do that. Sequence IDs are usually given in the FASTA header of input sequences. If this flag is active, RNALfold ignores any IDs re- trieved from the input and automatically generates an ID for each sequence. This ID consists of a prefix and an increasing number. This flag can also be used to add a FASTA header to the output even if the input has none. --id-prefix=STRING Prefix for automatically generated IDs (as used in output file names). (default=`sequence') If this parameter is set, each sequence will be prefixed with the provided string. Hence, the output files will obey the following naming scheme: "prefix_xxxx.lfold" where xxxx is the sequence num- ber. Note: Setting this parameter implies --auto-id. --id-delim=CHAR Change the delimiter between prefix and increasing number for auto- matically generated IDs (as used in output file names). (default=`_') This parameter can be used to change the default delimiter "_" be- tween the prefix string and the increasing number for automatically generated ID. --id-digits=INT Specify the number of digits of the counter in automatically gener- ated alignment IDs. (default=`4') When alignments IDs are automatically generated, they receive an in- creasing number, starting with 1. This number will always be left-padded by leading zeros, such that the number takes up a cer- tain width. Using this parameter, the width can be specified to the users need. We allow numbers in the range [1:18]. This option im- plies --auto-id. --id-start=LONG Specify the first number in automatically generated IDs. (default=`1') When sequence IDs are automatically generated, they receive an in- creasing number, usually starting with 1. Using this parameter, the first number can be specified to the users requirements. Note: nega- tive numbers are not allowed. Note: Setting this parameter implies to ignore any IDs retrieved from the input data, i.e. it activates the --auto-id flag. --filename-delim=CHAR Change the delimiting character used in sanitized filenames. (default=`ID-delimiter') This parameter can be used to change the delimiting character used while sanitizing filenames, i.e. replacing invalid characters. Note, that the default delimiter ALWAYS is the first character of the "ID delimiter" as supplied through the --id-delim option. If the delim- iter is a whitespace character or empty, invalid characters will be simply removed rather than substituted. Currently, we regard the following characters as illegal for use in filenames: backslash '\', slash '/', question mark '?', percent sign '%', asterisk '*', colon ':', pipe symbol '|', double quote '"', triangular brackets '<' and '>'. --filename-full Use full FASTA header to create filenames. (default=off) This parameter can be used to deactivate the default behavior of limiting output filenames to the first word of the sequence ID. Con- sider the following example: An input with FASTA header '>NM_0001 Homo Sapiens some gene' usually produces output files with the pre- fix "NM_0001" without the additional data available in the FASTA header, e.g. "NM_0001_ss.ps" for secondary structure plots. With this flag set, no truncation of the output filenames is done, i.e. output filenames receive the full FASTA header data as prefixes. Note, however, that invalid characters (such as whitespace) will be substituted by a delimiting character or simply removed, (see also the parameter option --filename-delim). --log-level=level Set log level threshold. (default=`2') By default, any log messages are filtered such that only warnings (level 2) or errors (level 3) are printed. This setting allows for specifying the log level threshold, where higher values result in fewer information. Log-level 5 turns off all messages, even errors and other critical information. --log-file[=filename] Print log messages to a file instead of stderr. (default=`RNAL- fold.log') --log-time Include time stamp in log messages. (default=off) --log-call Include file and line of log calling function. (default=off) Algorithms: Select additional algorithms which should be included in the calcu- lations. The Minimum free energy (MFE) and a structure representa- tive are calculated in any case. -L, --span=INT Set the maximum distance between any two pairing nucleotides. (default=`150') This option specifies the window length L and therefore the upper limit for the distance between the bases i and j of any pair (i, j), i.e. (j - i + 1) <= L. -z, --zscore[=DOUBLE] Limit the output to predictions with a Z-score below a threshold. (default=`-2') This option activates z-score regression using a trained SVM. Any predicted structure that exceeds the specified threshold will be om- mited from the output. Since the Z-score threshold is given as a negative number, it must immediately preceed the short option to not be mistaken as a separate argument, e.g. -z-2.9 sets the threshold to a value of -2.9 --zscore-pre-filter Apply the z-score filtering in the forward recursions. (default=off) The default mode of z-score filtering considers the entire structure space to decide whether or not a locally optimal structure at any position i is reported or not. When using this post-filtering step, however, alternative locally optimal structures starting at i with higher energy but lower z-score can be easily missed. The pre-filter option restricts the structure space already in the forward recur- sions, such that only optimal solution among those candidates that satisfy the z-score threshold are considered. Therefore, good results according to the z-score threshold criterion are less likely to be superseded by re- sults with better energy but worse z-score. Note, that activating this switch results in higher computation time which scales linear in the window length. --zscore-report-subsumed Report subsumed structures if their z-score is less than that of the enclosing structure. (default=off) In default mode, RNALfold only reports locally optimal structures if they are no constituents of another, larger structure with less free energy. In z-score mode, however, such a larger structure may have a higher z-score, thus may be less informative than the smaller sub- structure. Using this switch activates reporting both, the smaller and the larger structure if the z-score of the smaller is lower than that of the larger. -b, --backtrack-global Backtrack a global MFE structure. (default=off) Instead of just reporting the locally stable secondary structure a global MFE structure can be constructed that only consists of lo- cally optimal substructures. This switch activates a post-processing step that takes the locally optimal structures to generate the global MFE structure which constitutes the MFE value reported in the last line. The respective global MFE structure is printed just after the inut sequence part on the last line, preceding the global MFE score. Note, that this option implies -o/--outfile since the lo- cally optimal structures must be read after the regular prediction step! Also note, that using this option in combination with -z/--zs- core implies --zscore-pre-filter to ensure proper construction of the global MFE structure! -g, --gquad Incoorporate G-Quadruplex formation into the structure prediction algorithm. (default=off) Structure Constraints: Command line options to interact with the structure constraints fea- ture of this program --shape=filename Use SHAPE reactivity data to guide structure predictions. --shapeMethod=method Select SHAPE reactivity data incorporation strategy. (default=`D') The following methods can be used to convert SHAPE reactivities into pseudo energy contributions. 'D': Convert by using the linear equation according to Deigan et al 2009. Derived pseudo energy terms will be applied for every nucleotide in- volved in a stacked pair. This method is recognized by a capital 'D' in the provided parameter, i.e.: --shapeMethod="D" is the default setting. The slope 'm' and the intercept 'b' can be set to a non-de- fault value if necessary, otherwise m=1.8 and b=-0.6. To alter these parameters, e.g. m=1.9 and b=-0.7, use a parameter string like this: --shapeMethod="Dm1.9b-0.7". You may also provide only one of the two parameters like: --shapeMethod="Dm1.9" or --shapeMethod="Db-0.7". 'Z': Convert SHAPE reactivities to pseudo energies according to Zarringhalam et al 2012. SHAPE reactivities will be converted to pairing proba- bilities by using linear mapping. Aberration from the observed pair- ing probabilities will be penalized during the folding recursion. The magnitude of the penalties can affected by adjusting the factor beta (e.g. --shapeMethod="Zb0.8"). 'W': Apply a given vector of perturbation energies to unpaired nu- cleotides according to Washietl et al 2012. Perturbation vectors can be calcu- lated by using RNApvmin. --shapeConversion=method Select method for SHAPE reactivity conversion. (default=`O') This parameter is useful when dealing with the SHAPE incorporation according to Zarringhalam et al. The following methods can be used to convert SHAPE reactivities into the probability for a certain nu- cleotide to be unpaired. 'M': Use linear mapping according to Zarringhalam et al. 'C': Use a cutoff-approach to divide into paired and unpaired nucleotides (e.g. "C0.25") 'S': Skip the normalizing step since the input data already represents probabilities for being unpaired rather than raw reactiv- ity values 'L': Use a linear model to convert the reactivity into a probability for being unpaired (e.g. "Ls0.68i0.2" to use a slope of 0.68 and an intercept of 0.2) 'O': Use a linear model to convert the log of the reactivity into a probability for being unpaired (e.g. "Os1.6i-2.29" to use a slope of 1.6 and an intercept of -2.29) --commands=filename Read additional commands from file Commands include hard and soft constraints, but also structure mo- tifs in hairpin and internal loops that need to be treeted differ- ently. Furthermore, commands can be set for unstructured and struc- tured domains. Energy Parameters: Energy parameter sets can be adapted or loaded from user-provided input files -T, --temp=DOUBLE Rescale energy parameters to a temperature of temp C. Default is 37C. (default=`37.0') -P, --paramFile=paramfile Read energy parameters from paramfile, instead of using the default parameter set. Different sets of energy parameters for RNA and DNA should accompany your distribution. See the RNAlib documentation for details on the file format. The placeholder file name 'DNA' can be used to load DNA parameters without the need to actually specify any input file. -4, --noTetra Do not include special tabulated stabilizing energies for tri-, tetra- and hexaloop hairpins. (default=off) Mostly for testing. --salt=DOUBLE Set salt concentration in molar (M). Default is 1.021M. -m, --modifications[=STRING] Allow for modified bases within the RNA sequence string. (default=`7I6P9D') Treat modified bases within the RNA sequence differently, i.e. use corresponding energy corrections and/or pairing partner rules if available. For that, the modified bases in the input sequence must be marked by their corresponding one-letter code. If no additional arguments are supplied, all available corrections are performed. Otherwise, the user may limit the modifications to a particular sub- set of modifications, resp. one-letter codes, e.g. -mP6 will only correct for pseudouridine and m6A bases. Currently supported one-letter codes and energy corrections are: '7': 7-deaza-adenonsine (7DA) 'I': Inosine '6': N6-methyladenosine (m6A) 'P': Pseudouridine '9': Purine (a.k.a. nebularine) 'D': Dihydrouridine --mod-file=STRING Use additional modified base data from JSON file. Model Details: Tweak the energy model and pairing rules additionally using the fol- lowing parameters -d, --dangles=INT How to treat "dangling end" energies for bases adjacent to helices in free ends and multi-loops. (default=`2') With -d1 only unpaired bases can participate in at most one dangling end. With -d2 this check is ignored, dangling energies will be added for the bases adjacent to a helix on both sides in any case; this is the default for mfe and partition function folding (-p). The option -d0 ignores dangling ends altogether (mostly for debug- ging). With -d3 mfe folding will allow coaxial stacking of adjacent helices in multi-loops. At the moment the implementation will not allow coaxial stacking of the two enclosed pairs in a loop of degree 3 and works only for mfe folding. Note that with -d1 and -d3 only the MFE computations will be using this setting while partition function uses -d2 setting, i.e. dan- gling ends will be treated differently. --noLP Produce structures without lonely pairs (helices of length 1). (default=off) For partition function folding this only disallows pairs that can only occur isolated. Other pairs may still occasionally occur as he- lices of length 1. --noGU Do not allow GU pairs. (default=off) --noClosingGU Do not allow GU pairs at the end of helices. (default=off) --nsp=STRING Allow other pairs in addition to the usual AU,GC,and GU pairs. Its argument is a comma separated list of additionally allowed pairs. If the first character is a "-" then AB will imply that AB and BA are allowed pairs, e.g. --nsp="-GA" will allow GA and AG pairs. Nonstandard pairs are given 0 stacking energy. --energyModel=INT Set energy model. Rarely used option to fold sequences from the artificial ABCD... al- phabet, where A pairs B, C-D etc. Use the energy parameters for GC (--energyModel 1) or AU (--energyModel 2) pairs. --helical-rise=FLOAT Set the helical rise of the helix in units of Angstrom. (default=`2.8') Use with caution! This value will be re-set automatically to 3.4 in case DNA parameters are loaded via -P DNA and no further value is provided. --backbone-length=FLOAT Set the average backbone length for looped regions in units of Angstrom. (default=`6.0') Use with caution! This value will be re-set automatically to 6.76 in case DNA parameters are loaded via -P DNA and no further value is provided. REFERENCES If you use this program in your work you might want to cite: R. Lorenz, S.H. Bernhart, C. Hoener zu Siederdissen, H. Tafer, C. Flamm, P.F. Stadler and I.L. Hofacker (2011), "ViennaRNA Package 2.0", Algorithms for Molecular Biology: 6:26 I.L. Hofacker, W. Fontana, P.F. Stadler, S. Bonhoeffer, M. Tacker, P. Schuster (1994), "Fast Folding and Comparison of RNA Secondary Structures", Monatshefte f. Chemie: 125, pp 167-188 R. Lorenz, I.L. Hofacker, P.F. Stadler (2016), "RNA folding with hard and soft constraints", Algorithms for Molecular Biology 11:1 pp 1-13 I.L. Hofacker, B. Priwitzer, and P.F. Stadler (2004), "Prediction of Lo- cally Stable RNA Secondary Structures for Genome-Wide Surveys", Bioinfor- matics: 20, pp 186-190 The energy parameters are taken from: D.H. Mathews, M.D. Disney, D. Matthew, J.L. Childs, S.J. Schroeder, J. Su- san, M. Zuker, D.H. Turner (2004), "Incorporating chemical modification constraints into a dynamic programming algorithm for prediction of RNA sec- ondary structure", Proc. Natl. Acad. Sci. USA: 101, pp 7287-7292 D.H Turner, D.H. Mathews (2009), "NNDB: The nearest neighbor parameter database for predicting stability of nucleic acid secondary structure", Nu- cleic Acids Research: 38, pp 280-282 AUTHOR Ivo L Hofacker, Peter F Stadler, Ronny Lorenz REPORTING BUGS If in doubt our program is right, nature is at fault. Comments should be sent to rna@tbi.univie.ac.at. SEE ALSO RNAplfold(1) RNALalifold(1) RNALfold 2.7.2 December 2025 RNALFOLD(1)
NAME | SYNOPSIS | DESCRIPTION | REFERENCES | AUTHOR | REPORTING BUGS | SEE ALSO
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