The program PAUP works with files that are written in the NEXUS format. The NEXUS format is organized in blocks.
The TAXA-, CHARACTERS- (these two blocks may be linked in a DATA block), ASSUMPTIONS-, SET-, TREES-, CODON-, and DISTANCE blocks are normally placed in input files (i.e. together with the alignment). Commands available in the PAUP block can be used to specify tree searches. These commands can be written in batch files, facilitating tree search. Within most commands various options are available that allow a refined phylogenetic analysis.
The order of commands needs to be in an appropriate order. The order of options assigned to a command is arbitrary.
Each command begins with a command-name and ends with a semicolon. Otherwise, the commands are completely free-format. A command may span any number of lines (returns) and whitespace (tabs and blanks) may be inserted will.
Anything enclosed in square brackets is ignored when executing the runfile. Square brackets may be nested. Input of PAUP commands is case-insensitive, so command names, option keywords, etc. may be entered in any combination of upper- and lower-case characters (the only exception pertains to the MATRIX command). In addition, PAUP allows abbreviation of command names and keywords to the shortest unambiguous truncation. Note that other NEXUS-conforming programs may not accept these abbreviations (David L. Swofford, Beta Documentation Cmd_ref_v2.pdf).
For a better understanding, the runfile shown below is annotated with explanations available in the PAUP manual (David L. Swofford, Beta Documentation Cmd_ref_v2.pdf). For all tree searches conducted with PAUP in the course of this study the following DATA block and runfile (PAUP block) was used. Commands are capitalized and in bold, options are only capitalized. This routine can be obtained from the author and used as is, when saved in text format.
A.3.1 The DATA block:
#NEXUS [specifies the NEXUS format]
BEGIN DATA; [The DATA block contains the data matrix and other associated information. All options of the TAXA and CHARACTERS blocks are permitted under the DATA block.]
DIMENSIONS [The DIMENSIONS command specifies the size of the data matrix.]
NTAX = X [number of taxa]
NCHAR = Y; [number of characters]
FORMAT [The FORMAT command is used to specify information pertaining to the format of the data file.]
DATATYPE = DNA [This subcommand specifies the class of data. If present, it must be the first subcommand in the FORMAT command.]
INTERLEAVE [specifies that the data matrix is written in interleaved format (in contrast to the sequential format)]
MISSING = N [N is specified to represent missing data. If MISSING is not specified, it defaults to '?'.]
GAP = -; [Assigns the hyphen as the gap character. Whitespace is illegal as a MISSING DATA or GAP symbol, as are the following: ( ) [] {} / \, ; := * ' "` <>^ ]
MATRIX [In its standard format, the MATRIX command contains a sequence of taxon names, each]
taxon_1 ACGT... [followed by a space and sequence. Taxon names must not include blanks, hyphens or any of]
taxon_2 ACCT... [the following: ( ) [] {} / \, ; := * ' "` <>^. The matrix itself may appear in a variety of forms.]
taxon_3 ACCC...
;
END; [terminates the DATA block]
Appendix
148
A.3.2 The PAUP block:
#NEXUS [specifies the NEXUS format]
[CD C:\path\folder;] [specifies the directory to which files are saved]
LOG
FILE = Alignment_name_NJJC.log [saves output to the specified file]
START = YES [logging to the specified file is initiated]
REPLACE = YES; [A preexisting file with this name is replaced without warning]
FACTORY; [restores default settings. There is a bug report for this function, however, it is believed that this bug does not affect this routine]
[
### replace: Alignment_name Ten files are generated by this routine. Use the editor's replacement function to replace preexisting alignment names with the current one
### replace: [NJ]1000 You may adjust the number of bootstrap replicates for each
### replace: [MP]1000 phylogenetic method separately with the editor's replace function
### replace: [ML]100 ML bootstrapping requires extensive computing time and may be excluded by parenthesizing the respective commands. Support values
based on the ML criterion might be obtained in Bayesian analyses.
]
OUTGROUP taxon taxa [transfers specified taxa to the outgroup]
/ONLY; [no other taxa than those specified are kept as outgroup]
SET [The SET command is used to set a variety of options whose scope
extends beyond single commands.]
INCREASE = AUTO [When maximal number of trees that can be stored is reached during a search, this setting allows the automatic increase of this limit.]
ROOT = OUTGROUP [The ROOT option is used to specify how unrooted trees are to be
rooted prior to output]
OUTROOT = MONOPHYL; [Shows the outgroup monophyletic to the ingroup]
[*** doing NJJC ***] [NJ = neighbor-joining, JC = Jukes-Cantor model of evolution]
SET
CRITERION = DISTANCE; [Sets the optimality criterion to distance]
DSET [Under the DSET command options for distance analysis can be set]
DISTANCE = JC [Sets the model of evolution to the Jukes-Cantor model]
OBJECTIVE = ME [Sets minimum evolution (ME) as the objective function]
SUBST = ALL [counts or estimates substitutions of all types (default)]
NEGBRLEN = SETZERO; [Negative branch lengths are allowed but set to zero when computing the
value of the objective function (default)]
NJ [ The NJ command specifies neighbor-joining method]
SHOWTREE = NO [Trees are not output to the display buffer]
BREAKTIES = RANDOM; [ties encountered during tree construction are broken randomly and not
according to the input order of the taxa]
SAVETREES
FILE = Alignment_name_NJJC.tre [saves trees currently in memory to the specified file]
BRLENS = YES [saves branch lengths]
ROOT = NO [roots trees using the rooting options currently in effect before they are saved]
REPLACE = YES; [A preexisting file with this name is replaced without warning]
Appendix
BOOTSTRAP [Bootstrap analysis is initiated]
NREPS = [NJ]1000 [specifies the number of bootstrap replicates, see headlines]
CUTOFFPCT = 0 [specify the minimum bootstrap partition frequency to be displayed in the partition frequency table. Partitions occurring at
a frequency below this value are not shown]
KEEPALL = YES [Groups occurring at frequencies less than CONLEVEL will also be retained in the bootstrap consensus as long as they are compatible with all groups that are already included in the consensus. Effectively, this forces
CONLEVEL=50, which is also the default value]
SEARCH = NJ; [uses the NJ method for tree calculation from each bootstrap replicate]
SAVETREES
FILE = Alignment_name_NJJCboo.tre [Saves Bootstrap consensus tree to the specified file]
FROM = 1 TO = 1 [due to a bug in the program the tree number of the bootstrap consensus tree has to be specified]
SAVEBOOTP = NODELABELS [Saves bootstrap proportions as internal node labels]
MAXDECIMALS = 0 [rounds bootstrap proportions to integer values]
ROOT = YES [see above]
REPLACE = YES; [see above]
LOG STOP; [stops saving output to file]
[*** doing MPn ***] [MPn = MP with gap characters treated as fifth character state]
LOG [see above]
FILE = Alignment_name_MPn.log START = YES REPLACE = YES;
SET
CRITERION = PARSIMONY [Sets the optimality criterion to parsimony]
MAXTREES = 10000 [limits the maximum number of trees kept in memory to 10000]
increase = NO; [prevents MAXTREES to be increased automatically, this mode is chosen because of the endless number of trees that can be
generated when almost identical sequences are included in the alignment, which would require extensive computing time]
PSET GAPMODE = NEWSTATE; [treats gap characters as a fifth base. This option is only
available under the parsimony optimality criterion in PAUP
version 4.0b10]
HSEARCH [employs heuristic algorithms for tree search]
START = STEPWISE [sequences are added stepwise to obtain the]
ADDSEQ = RANDOM [the order of added sequences is random]
NREPS = 100 [Specifies the number of random-addition-sequence replications to be performed for finding the starting tree for branch
swapping]
SWAP = TBR [Specifies the algorithm used by branch-swapping (=default)]
STATUS = NO; [number of trees examined is not output to a status window ]
SAVETREES [see above]
FILE = Alignment_name_MPn.tre
ROOT = YES BRLENS = YES REPLACE = YES;
Appendix
150
PSCORES/ [requests a listing of tree lengths and/or fit measures for one or more trees]
CI = YES [consistency index]
RI = YES [retention index]
RC = YES [rescaled consistency index]
HI = YES; [homoplasy indices]
[*** doing wMPn ***] [wMPn = weighted MP with gap characters treated as fifth character state]
REWEIGHT [Use the REWEIGHT command to assign weights to the characters based on their fit to the trees currently in memory]
INDEX = RC [Specifies the fit measure to use when calculating the new character weights (RC = rescaled consistency index (= default)]
BASEWT = 1000 [Specifies the maximum possible weight that a character can be assigned, corresponding to an index. Weights are scaled from 0 to this value]
FIT = MEAN; [Specify whether the new weights are based on the mean of the fit values for each character over all of the trees in memory]
HSEARCH [The same heuristic search is executed as the previous one except that
alignment positions are weighted]
START = STEPWISE ADDSEQ = RANDOM NREPS = 100 SWAP = TBR STATUS = NO;
SAVETREES [see above]
FILE = Alignment_name_wMPn.tre
ROOT = YES BRLENS = YES REPLACE = YES;
PSCORES/ [see above]
CI = YES RI = YES RC = YES HI = YES;
[*** doing wMPn bootstrapping ***] [bootstrapping is based on current settings, which are weighted parsimony and gaps treated as fifth character state]
SET
MAXTREES = 100 increase = no; [see above]
BOOTSTRAP
NREPS = [MP]1000 [see above]
CUTOFFPCT = 0
KEEPALL = YES
SEARCH = HEURISTIC;
SAVETREES
FILE = Alignment_name_wMPnboo.tre [see above]
FROM = 1 TO = 1
SAVEBOOTP = NODELABELS MAXDECIMALS = 0
ROOT = YES
REPLACE = YES;
Appendix
SET
MAXTREES = 100
INCREASE = AUTO; [resets the autoincrease function]
PSET
GAPMODE = MISSING; [resets gap characters as missing data]
LOG STOP; [see above]
[*** doing MLNJ ***] [MLNJ = ML with ML scores as estimated by ModelTest and starting tree search with a NJ tree]
LOG
FILE = Alignment_name_ML.log [see above]
START = YES REPLACE = YES;
SET
CRITERION = LIKELIHOOD; [Sets the optimality criterion to maximum-likelihood]
LSET [Sets options for maximum likelihood analysis. All options specified
here can be approximated by ModelTest]
WTS = IGNORE [Ignores the weights calculated during the MP tree search (if in doubt]
BASE =(0.2085 0.2427 0.2603) [Explicitly specifies base frequencies (frqA frqC frqG)]
NST = 6 [Specifies the number of substitution types]
RMAT=(1.2330 3.9662 2.1121 0.6401 3.9662) [Specifies the values for a six-parameter instantaneous rate matrix if NST = 6]
RATES = GAMMA [Uses a gamma distribution to compensate for among site rate variation]
SHAPE = 0.3516 [specifies the α- (or shape-) parameter of the gamma distribution, which
determines the shape of the gamma distribution.]
NCAT = 4 [Specifies the number of categories to divide the discrete approximation of the gamma distribution. This value is not provided by ModelTest but the given setting (4) corresponds to the default setting of PAUP]
PINVAR = 0; [specify the proportion of sites unable to accept substitutions;
the remaining sites are assumed to vary at the same rate]
HSEARCH
START = NJ [Starts the heuristic search with a NJ-tree]
SWAP = TBR [see above]
STATUS = NO; [see above]
SAVETREES
FILE = Alignment_name_MLNJ.tre [see above]
ROOT = YES BRLENS = YES REPLACE = YES;
BOOTSTRAP
NREPS = [ML]100 [see above]
CUTOFFPCT = 0
KEEPALL = YES
SEARCH = HEURISTIC;
SAVETREES
FILE = Alignment_name_MLboo.tre [see above]
FROM = 1 TO = 1 SAVEBOOTP = NODELABELS MAXDECIMALS = 0
ROOT = YES
REPLACE = YES;
LOG STOP; [see above]
Appendix
152