MrBayes 3.1 Manual - molecularevolution.org

MrBayes 3.1 Manual - molecularevolution.org
MrBayes 3.1 Manual
Draft 5/26/2005
Fredrik Ronquist1,3, John P. Huelsenbeck2,4, and Paul van der Mark1,5
1
School of Computational Science
Florida State University
Tallahassee, FL 32306-4120, U.S.A.
2
Division of Biological Sciences
University of California at San Diego
La Jolla, CA 92093, USA
3
4
[email protected]
[email protected]
5
[email protected]
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Contents
Contents......................................................................................................................... 2
1. Introduction............................................................................................................... 4
1.1. Conventions Used in this Manual ......................................................................... 4
1.2. Acquiring and Installing MrBayes ........................................................................ 4
1.3. Getting Started ..................................................................................................... 6
1.4. Changing the Size of the MrBayes Window ......................................................... 6
1.5. Getting Help......................................................................................................... 7
1.6. Reporting and Fixing Bugs ................................................................................... 7
1.7. License and Warranty........................................................................................... 8
2. Tutorial: A Simple Analysis...................................................................................... 8
2.1. Quick Start Version .............................................................................................. 8
2.2. Getting Data into MrBayes ................................................................................... 9
2.3. Specifying a Model ............................................................................................ 11
2.4. Setting the Priors ................................................................................................ 13
2.5. Checking the Model ........................................................................................... 15
2.6. Setting up the Analysis ....................................................................................... 16
2.7. Running the Analysis ......................................................................................... 19
2.8. When to Stop the Analysis ................................................................................. 21
2.9. Summarizing Samples of Substitution Model Parameters ................................... 22
2.10. Summarizing Samples of Trees and Branch Lengths ........................................ 24
3. Analyzing a Partitioned Data Set ........................................................................... 27
3.1. Getting Mixed Data into MrBayes ...................................................................... 27
3.2. Dividing the Data into Partitions......................................................................... 28
3.3. Specifying a Partitioned Model........................................................................... 29
3.4. Running the Analysis ......................................................................................... 30
4. Evolutionary Models Implemented in MrBayes 3 ................................................. 31
4.1. Nucleotide Models ............................................................................................. 31
4.1.1. Simple Nucleotide Models............................................................................... 31
4.1.2. The Doublet Model.......................................................................................... 33
4.1.3. Codon Models ................................................................................................. 35
4.2. Amino-acid Models............................................................................................ 36
4.2.1 Fixed Rate Models............................................................................................ 37
4.2.2. Estimating the Fixed Rate Model..................................................................... 37
4.2.3. Variable Rate Models ...................................................................................... 37
4.3. Restriction Site (Binary) Model .......................................................................... 39
4.4. Standard Discrete (Morphology) Model.............................................................. 40
4.5. Parsimony Model ............................................................................................... 42
4.6. Rate Variation Across Sites ................................................................................ 43
4.6.1. Gamma-distributed Rate Model ....................................................................... 43
4.6.2. Autocorrelated Gamma Model......................................................................... 43
4.6.3. Proportion of Invariable Sites .......................................................................... 44
4.6.4. Partitioned (Site Specific) Rate Model............................................................. 45
4.6.5. Inferring Site Rates.......................................................................................... 45
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4.7. Rate Variation Across the Tree: The Covarion Model......................................... 46
4.8. Topology and Branch Length Models................................................................. 47
4.8.1. Unconstrained and Constrained Topology ....................................................... 47
4.8.2. Non-clock (Standard) Trees ............................................................................. 48
4.8.3. Strict Clock Trees............................................................................................ 48
4.8.4. Relaxed Clock Trees........................................................................................ 49
4.9. Partitioned Models ............................................................................................. 49
4.10. Ancestral State Reconstruction ......................................................................... 50
5. Frequently Asked Questions................................................................................... 51
6. Differences Between Version 2 and Version 3........................................................ 57
7. Advanced Topics ..................................................................................................... 59
7.1. Compiling MrBayes ........................................................................................... 59
7.1.1. Compiling with GNU Make............................................................................. 59
7.1.2. Compiling with Code Warrior or Visual Studio ............................................... 61
7.2. Compiling and Running the Parallel Version of MrBayes................................... 61
7.2.1. The Parallel Macintosh Version....................................................................... 61
7.2.2. The MPI Version for Unix Clusters ................................................................. 62
7.3. Working with the Source Code........................................................................... 62
8. Acknowledgements.................................................................................................. 63
9. References................................................................................................................ 63
Appendix: Evolutionary models and proposals in MrBayes
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1. Introduction
MrBayes 3 is a program for the Bayesian inference of phylogeny. The program has a
command-line interface and should run on a variety of computer platforms, including
clusters of Macintosh and UNIX computers. Note that the computer should be reasonably
fast and should have a lot of RAM memory (depending on the size of the data matrix, the
program may require hundreds of megabytes of memory). The program is optimized for
speed and not for minimizing memory requirements.
This manual explains how to use the program. After this section, which introduces the
program, we will first walk you through a simple analysis (section 2 of the manual),
which will get you started, and a more complex analysis that uses more of the program’s
capabilities (section 3). We then briefly describe the models implemented in the program
(section 4), answer some frequently asked questions (section 5), and discuss the
differences between versions 2 and 3 of the program (section 6). Finally, we give more
detailed instructions on how to compile the program and how to run the parallel versions
of it (section 7). Section 7 also contains brief information for developers interested in
tweaking MrBayes code or contributing to the MrBayes project. The manual ends with a
series of diagrams giving a graphical overview of all the models and proposal
mechanisms implemented in the program (Appendix). For more detailed information
about commands and options in MrBayes, see the command reference that can either be
downloaded from the program web site or generated from the program itself (see section
1.4 Getting Help below). All the information in the command reference is also available
on-line when using the program.
The manual assumes that you are familiar with the basic concepts of Bayesian
phylogenetics. If you are new to the subject, we recommend the recent reviews by Holder
and Lewis (2003), Lewis (2001) and Huelsenbeck et al. (2001, 2002). It is also
worthwhile to study the early papers introducing Bayesian phylogenetic methods (Li
1996; Mau, 1996; Rannala and Yang, 1996; Mau and Newton, 1997; Rannala and Yang,
1997; Larget and Simon, 1999; Mau, Newton and Larget, 1999; Newton, Mau and
Larget, 1999). The basic MCMC techniques are described in Metropolis et al. (1953) and
Hastings (1970). The Metropolis-coupled MCMC used by MrBayes was introduced by
Geyer (1991).
1.1. Conventions Used in this Manual
What you see on the screen and what is in the input file is in plain typewriter
font. What you type is in bold typewriter font.
1.2. Acquiring and Installing MrBayes
MrBayes 3 is distributed without charge by download from the MrBayes web site,
http://mrbayes.net. If someone has given you a copy of MrBayes 3, we strongly suggest
that you download the most recent version from this site. The site also gives information
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about the MrBayes users email list and describes how you can report bugs or contribute
to the project.
MrBayes 3 is a plain-vanilla program that uses a command line interface and therefore
behaves virtually the same on all platforms - Macintosh, Windows and Unix. There is a
separate download package for each platform. The Macintosh package contains two
versions of the program: the standard serial version, named MrBayes3.1 (program icon
one copy of Reverend Bayes’s portrait), and a version for running the program in parallel
on clusters of Macintoshes, named MrBayes3.1p (program icon four portraits of
Reverend Bayes). For more information on the parallel Macintosh version of MrBayes,
which requires the installation of POOCH, see section 7 of this user manual. The
Windows package only contains the serial version of the program and is ready to run
after unzipping, just like the Macintosh serial version.
If you decide to run the program under Unix/Linux, then you need to compile the
program from the source code. In the latter case, simply unpack the file
mrbayes3.1_src.tar.gz by typing gunzip MrBayes_3.1_src.tar.gz
and then tar –xf MrBayes_3.1_src.tar. The gunzip command unzips the
compressed file and the tar –xf command extracts all of the files from the .tar
archive that resulted from the unzip operation (note that the .gz suffix is dropped in the
unzip operation). You then need to compile the program. We have included a “Makefile”
that contains compiler instructions producing the serial version of the program. You
simply type make to compile the program according to these instructions. A typical
compile session would look like this:
ronquistg5:mrbayes>ls
mrbayes3.1_src.tar.gz
ronquistg5:mrbayes>gunzip MrBayes_3.1_src.tar.gz
ronquistg5:mrbayes>ls
mrbayes3.1_src.tar
ronquistg5:mrbayes>tar -xf MrBayes_3.1_src.tar
ronquistg5:mrbayes>make
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized
-c
gcc -DUNIX_VERSION -O3 -Wall -Wno-uninitialized -lm
mbmath.o mcmc.o model.o plot.o sump.o sumt.o
-o mb
ronquistg5:mrbayes>
-o mb.o mb.c
-o mcmc.o mcmc.c
-o bayes.o bayes.c
-o command.o command.c
-o mbmath.o mbmath.c
-o model.o model.c
-o plot.o plot.c
-o sump.o sump.c
-o sumt.o sumt.c
mb.o bayes.o command.o
The compilation usually stops for several minutes at the mcmc.c file; this is perfectly
normal. This is the largest source file and optimization of the code takes quite a while.
We assume as the default C compiler gcc, which is installed on most systems. If you do
not have gcc installed on your machine, or you want to produce the MPI version or some
other special version of the program, you have to change the compiler information in the
Makefile as described in section 7 of this manual. The executable serial version of the
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program is called “mb”. To execute the program, simply type ./mb in the directory
where you compiled the program. The ./ prefix is needed to tell Unix that you want to
run the local copy of mb in your working directory. If you run MrBayes often, you will
probably want to add the program to your “path”; refer to your Unix manual or your local
Unix expert for more information on this.
All three packages of MrBayes come with example data files. These are intended to show
various types of analyses you can perform with the program, and you can use them as
templates for your own analyses. Two of the files, primates.nex and cynmix.nex, will be
used in the tutorial sections of this manual (sections 2 and 3).
1.3. Getting Started
Start MrBayes by double-clicking the application icon (or typing ./mb) and you will see
the information below:
MrBayes v3.1
(Bayesian Analysis of Phylogeny)
by
Fredrik Ronquist and John P. Huelsenbeck
School of Computational Science
Florida State University
[email protected]
Section of Ecology, Behavior and Evolution
Division of Biological Sciences
University of California, San Diego
[email protected]
Distributed under the GNU General Public License
Type "help" or "help <command>" for information
on the commands that are available.
MrBayes >
The order of the authors is randomized each time you start the program, so don’t be
surprised if the order differs from the one above. Note the MrBayes > prompt at the
bottom, which tells you that MrBayes is ready for your commands.
1.4. Changing the Size of the MrBayes Window
Some MrBayes commands will output a lot of information and write fairly long lines, so
you may want to change the size of the MrBayes window to make it easier to read the
output. On Macintosh and Unix machines, you should be able to increase the window
size simply by dragging the margins. On a Windows machine, you cannot increase the
size of the window beyond the preset value by simply dragging the margins but (on
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Windows XP, 2000 and NT) you can change both the size of the screen buffer and the
console window by right-clicking on the blue title bar of the MrBayes window and then
selecting “Properties” in the menu that appears. Make sure the “Layout” tab is selected in
the window that appears, and then set the Screen Buffer Size and Window Size to the
desired values.
1.5. Getting Help
At the MrBayes > prompt, type help to see a list of the commands available in
MrBayes. Most commands allow you to set values (options) for different parameters. If
you type help <command>, where <command> is any of the listed commands, you
will see the help information for that command as well as a description of the available
options. For most commands, you will also see a list of the current settings at the end.
Try, for instance, help lset or help mcmc. The lset settings table looks like this:
Default model settings:
Parameter
Options
Current Setting
-----------------------------------------------------------------Nucmodel
4by4/Doublet/Codon
4by4
Nst
1/2/6
1
Code
Universal/Vertmt/Mycoplasma/
Yeast/Ciliates/Metmt
Universal
Ploidy
Haploid/Diploid
Diploid
Rates
Equal/Gamma/Propinv/Invgamma/Adgamma Equal
Ngammacat
<number>
4
Nbetacat
<number>
5
Omegavar
Equal/Ny98/M3
Equal
Covarion
No/Yes
No
Coding
All/Variable/Noabsencesites/
Nopresencesites
All
Parsmodel
No/Yes
No
------------------------------------------------------------------
Note that MrBayes 3 supports abbreviation of commands and options, so in many cases it
is sufficient to type the first few letters of a command or option instead of the full name.
A complete list of commands and options is given in the command reference, which can
be downloaded from the program web site (www.mrbayes.net). You can also produce an
ASCII text version of the command reference at any time by giving the command
manual to MrBayes. Further help is available in a set of hyperlinked html pages
produced by Jeff Bates and available on the MrBayes web site. Finally, you can get in
touch with other MrBayes users and developers through the mrbayes-users email list
(subscription information at www.mrbayes.net).
1.6. Reporting and Fixing Bugs
If you find a bug in MrBayes, we are very grateful if you tell us about it using the bug
reporting functions of SourceForge, as explained on the MrBayes web site
(www.mrbayes.net). When you submit a bug, make sure that you upload a data file with
the data set and sequence of commands that produced the error. If the bug occurs during a
MCMC analysis (after issuing the “mcmc” command), you can help us greatly by making
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sure the bug can be reproduced reliably using a fixed seed and swapseed for the mcmc
command, and ideally also with a small data set. The Tracker software at SourceForge
will make sure that you get email notification when the bug has been fixed in the source
code on the MrBayes CVS repository at SourceForge. Note, however, that there may be
some time before new executables containing the bug fix will be released.
Advanced users may be interested in fixing bugs themselves in the source code. Refer to
section 7 of this manual for information on how to contribute bug fixes, improved
algorithms or expanded functionality to other users of MrBayes.
1.7. License and Warranty
MrBayes is free software; you can redistribute it and/or modify it under the terms of the
GNU General Public License as published by the Free Software Foundation; either
version 2 of the License, or (at your option) any later version.
The program is distributed in the hope that it will be useful, but WITHOUT ANY
WARRANTY; without even the implied warranty of MERCHANTABILITY or
FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public License for
more details (http://www.gnu.org/copyleft/gpl.html).
2. Tutorial: A Simple Analysis
This section is a tutorial based on the primates.nex data file. It will guide you
through a basic Bayesian MCMC analysis of phylogeny, explaining the most important
features of the program. There are two versions of the tutorial. You will first find a
Quick-Start version for impatient users who want to get an analysis started immediately.
The rest of the section contains a much more detailed description of the same analysis.
2.1. Quick Start Version
There are four steps to a typical Bayesian phylogenetic analysis using MrBayes:
1.
2.
3.
4.
Read the Nexus data file
Set the evolutionary model
Run the analysis
Summarize the samples
In more detail, each of these steps is performed as described in the following paragraphs.
1. At the MrBayes > prompt, type execute primates.nex. This will bring the
data into the program. The data file (primates.nex) must be in the same directory as
the MrBayes program. If you run into problems, refer to section 2.2 for possible
solutions. If you are running your own data file, beware that it may contain some
MrBayes commands that can change the behavior of the program; delete those commands
or put them in square brackets to follow this tutorial.
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2. At the MrBayes > prompt, type lset nst=6 rates=invgamma. This sets the
evolutionary model to the GTR model with gamma-distributed rate variation across sites
and a proportion of invariable sites. If your data are not DNA or RNA, if you want to
invoke a different model, or if you want to use non-default priors, refer to the rest of this
manual, particularly sections 3 to 5 and the Appendix.
3.1. At the MrBayes > prompt, type mcmc ngen=10000 samplefreq=10. This
will ensure that you get at least 1,000 samples from the posterior probability distribution.
For larger data sets you probably want to run the analysis longer and sample less
frequently (the default sampling frequency is every 100th generation). You can find the
predicted remaining time to completion of the analysis in the last column printed to
screen.
3.2. If the standard deviation of split frequencies is below 0.01 after 100,000 generations,
stop the run by answering no when the program asks “Continue the analysis?
(yes/no)”. Otherwise, keep adding generations until the value falls below 0.01.
4.1. Summarize the parameter values by typing sump burnin=250 (or whatever value
corresponds to 25 % of your samples). The program will output a table with summaries
of the samples of the substitution model parameters, including the mean, mode, and 95 %
credibility interval of each parameter. Make sure that the potential scale reduction factor
(PSRF) is reasonably close to 1.0 for all parameters; if not, you need to run the analysis
longer.
4.2. Summarize the trees by typing sumt burnin=250 (or whatever value
corresponds to 25 % of your samples). The program will output a cladogram with the
posterior probabilities for each split and a phylogram with mean branch lengths. The trees
will also be printed to a file that can be read by tree drawing programs such as TreeView,
MacClade, and Mesquite.
It does not have to be more complicated than this; however, as you get more proficient
you will probably want to know more about what is happening behind the scenes. The
rest of this section explains each of the steps in more detail and introduces you to all the
implicit assumptions you are making and the machinery that MrBayes uses in order to
perform your analysis.
2.2. Getting Data into MrBayes
To get data into MrBayes, you need a so-called Nexus file that contains aligned
nucleotide or amino acid sequences, morphological ("standard") data, restriction site
(binary) data, or any mix of these four data types. The Nexus file that we will use for this
tutorial, primates.nex, contains 12 mitochondrial DNA sequences of primates.
A Nexus file is a simple text (ASCII) file that begins with #NEXUS on the first line. The
rest of the file is divided into different blocks. The primates.nex file looks like this:
#NEXUS
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begin data;
dimensions ntax=12 nchar=898;
format datatype=dna interleave=no gap=-;
matrix
Saimiri_sciureus AAGCTTCATAGGAGC ... ACTATCCCTAAGCTT
Tarsius_syrichta AAGCTTCACCGGCGC ... ATTATGCCTAAGCTT
Lemur_catta
AAGCTTCACCGGCGC ... ACTATCTATTAGCTT
Macaca_fuscata
AAGCTTCACCGGCGC ... CCTAACGCTAAGCTT
M_mulatta
AAGCTTCACCGGCGC ... CCTAACACTAAGCTT
M_fascicularis
AAGCTTTACAGGTGC ... CCTAACACTAAGCTT
M_sylvanus
AAGCTTTTCCGGCGC ... CCTAACATTAAGCTT
Homo_sapiens
AAGCTTTTCTGGCGC ... GCTCTCCCTAAGCTT
Pan
AAGCTTCTCCGGCGC ... GCTCTCCCTAAGCTT
Gorilla
AAGCTTCTCCGGTGC ... ACTCTCCCTAAGCTT
Pongo
AAGCTTCACCGGCGC ... ACTCTCACTAAGCTT
Hylobates
AAGTTTCATTGGAGC ... ACTCTCCCTAAGCTT
;
end;
The file contains only one block, a DATA block. The DATA block is initialized with
begin data; followed by the dimensions statement, the format statement, and
the matrix statement. The dimensions statement must contain ntax, the number of
taxa, and nchar, the number of characters in each aligned sequence. The format
statement must specify the type of data, for instance datatype=DNA (or RNA or
Protein or Standard or Restriction). The format statement may also contain gap=- (or
whatever symbol is used for a gap in your alignment), missing=? (or whatever symbol
is used for missing data in your file), interleave=yes when the data matrix is
interleaved sequences, and match=. (or whatever symbol is used for matching
characters in the alignment). The format statement is followed by the matrix
statement, usually started by the word matrix on a separate line, followed by the
aligned sequences. Each sequence begins with the sequence name separated from the
sequence itself by at least one space. The data block is completed by an end; statement.
Note that the begin, dimensions, format, and end statements all end in a
semicolon. That semicolon is essential and must not be left out. Note that, although it
occupies many lines in the file, the matrix description is also a regular statement, a
matrix statement, which ends with a semicolon just as any other statement. Our
example file contains sequences in non-interleaved (block) format. Non-interleaved is the
default but you can put interleave=no in the format statement if you want to be
sure.
The Nexus data file can be generated by a program such as MacClade or Mesquite. Note,
however, that MrBayes does not support the full Nexus standard, so you may have to do a
little editing of the file for MrBayes to process it properly. In particular, MrBayes uses a
fixed set of symbols for each data type and does not support the “symbols” and “equate”
options of the “format” command. The supported symbols are {A, C, G, T, R, Y, M, K,
S, W, H, B, V, D, N} for DNA data, {A, C, G, U, R, Y, M, K, S, W, H, B, V, D, N} for
RNA data, {A, R, N, D, C, Q, E, G, H, I, L, K, M, F, P, S, T, W, Y, V, X} for protein
data, {0, 1} for restriction (binary) data, and {0, 1, 2, 3, 4, 5, 6, 5, 7, 8, 9} for standard
(morphology) data. In addition to the standard one-letter ambiguity symbols for DNA and
RNA listed above, ambiguity can also be expressed using the Nexus parenthesis or curly
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braces notation. For instance, a taxon polymorphic for states 2 and 3 can be coded as
(23), (2,3), {23}, or {2,3} and a taxon with either amino acid A or F can be coded as
(AF), (A,F), {AF} or {A,F}. Like most other statistical phylogenetics programs,
MrBayes effectively treats polymorphism and uncertainty the same way (as uncertainty),
so it does not matter whether you use parentheses or curly brackets. If you have other
symbols in your matrix than the ones supported by MrBayes, you need to replace them
before processing the data block in MrBayes. You also need to remove the “Equate” and
“Symbols” statements in the “Format” line if they are included. Unlike the Nexus
standard, MrBayes supports data blocks that contain mixed data types as described in
section 3 of this manual.
To put the data into MrBayes type execute <filename> at the MrBayes >
prompt, where <filename> is the name of the input file. To process our example file, type
execute primates.nex or simply exe primates.nex to save some typing.
Note that the input file must be located in the same folder (directory) as the MrBayes
application (or else you will have to give the path to the file) and the name of the input
file should not have blank spaces. If everything proceeds normally, MrBayes will
acknowledge that it has read the data in the DATA block of the Nexus file by outputting
the following information:
Executing file "primates.nex"
DOS line termination
Longest line length = 915
Parsing file
Expecting NEXUS formatted file
Reading data block
Allocated matrix
Matrix has 12 taxa and 898 characters
Data is Dna
Data matrix is not interleaved
Gaps coded as Setting default partition (does not divide up characters).
Taxon 1 -> Tarsius_syrichta
Taxon 2 -> Lemur_catta
Taxon 3 -> Homo_sapiens
Taxon 4 -> Pan
Taxon 5 -> Gorilla
Taxon 6 -> Pongo
Taxon 7 -> Hylobates
Taxon 8 -> Macaca_fuscata
Taxon 9 -> M_mulatta
Taxon 10 -> M_fascicularis
Taxon 11 -> M_sylvanus
Taxon 12 -> Saimiri_sciureus
Setting output file names to "primates.nex.<run<i>.p/run<i>.t>"
Successfully read matrix
Exiting data block
Reached end of file
2.3. Specifying a Model
All of the commands are entered at the MrBayes > prompt. At a minimum two
commands, lset and prset, are required to specify the evolutionary model that will be
used in the analysis. Usually, it is also a good idea to check the model settings prior to the
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analysis using the showmodel command. In general, lset is used to define the
structure of the model and prset is used to define the prior probability distributions on
the parameters of the model. In the following, we will specify a GTR + I + _ model (a
General Time Reversible model with a proportion of invariable sites and a gammashaped distribution of rates across sites) for the evolution of the mitochondrial sequences
and we will check all of the relevant priors. If you are unfamiliar with stochastic models
of molecular evolution, we suggest that you consult a general text, such as Felsenstein
(2004).
In general, a good start is to type help lset. Ignore the help information for now and
concentrate on the table at the bottom of the output, which specifies the current settings.
It should look like this:
Model settings for partition 1:
Parameter
Options
Current Setting
-----------------------------------------------------------------Nucmodel
4by4/Doublet/Codon
4by4
Nst
1/2/6
1
Code
Universal/Vertmt/Mycoplasma/
Yeast/Ciliates/Metmt
Universal
Ploidy
Haploid/Diploid
Diploid
Rates
Equal/Gamma/Propinv/Invgamma/Adgamma Equal
Ngammacat
<number>
4
Nbetacat
<number>
5
Omegavar
Equal/Ny98/M3
Equal
Covarion
No/Yes
No
Coding
All/Variable/Noabsencesites/
Nopresencesites
All
Parsmodel
No/Yes
No
------------------------------------------------------------------
First, note that the table is headed by Model settings for partition 1. By
default, MrBayes divides the data into one partition for each type of data you have in
your DATA block. If you have only one type of data, all data will be in a single partition
by default. How to change the partitioning of the data will be explained in section 3 of the
manual.
The Nucmodel setting allows you to specify the general type of DNA model. The
Doublet option is for the analysis of paired stem regions of ribosomal DNA and the
Codon option is for analyzing the DNA sequence in terms of its codons. We will analyze
the data using a standard nucleotide substitution model, in which case the default 4by4
option is appropriate, so we will leave Nucmodel at its default setting.
The general structure of the substitution model is determined by the Nst setting. By
default, all substitutions have the same rate (Nst=1), corresponding to the F81 model (or
the JC model if the stationary state frequencies are forced to be equal using the prset
command, see below). We want the GTR model (Nst=6 instead of the F81 model so we
type lset nst=6. MrBayes should acknowledge that it has changed the model
settings.
MrBayes 3.1 Manual
5/26/2005
13
The Code setting is only relevant if the Nucmodel is set to Codon. The Ploidy
setting is also irrelevant for us. However, we need to change the Rates setting from the
default Equal (no rate variation across sites) to Invgamma (gamma-shaped rate
variation with a proportion of invariable sites). Do this by typing lset
rates=invgamma. Again, MrBayes will acknowledge that it has changed the settings.
We could have changed both lset settings at once if we had typed lset nst=6
rates=invgamma in a single line.
We will leave the Ngammacat setting (the number of discrete categories used to
approximate the gamma distribution) at the default of 4. In most cases, four rate
categories are sufficient. It is possible to increase the accuracy of the likelihood
calculations by increasing the number of rate categories. However, the time it will take to
complete the analysis will increase in direct proportion to the number of rate categories
you use, and the effects on the results will be negligible in most cases.
Of the remaining settings, it is only Covarion and Parsmodel that are relevant for
single nucleotide models. We will use neither the parsimony model nor the covariotide
model for our data, so we will leave these settings at their default values. If you type
help lset now to verify that the model is correctly set, the table should look like this:
Model settings for partition 1:
Parameter
Options
Current Setting
-----------------------------------------------------------------Nucmodel
4by4/Doublet/Codon
4by4
Nst
1/2/6
6
Code
Universal/Vertmt/Mycoplasma/
Yeast/Ciliates/Metmt
Universal
Ploidy
Haploid/Diploid
Diploid
Rates
Equal/Gamma/Propinv/Invgamma/Adgamma Invgamma
Ngammacat
<number>
4
Nbetacat
<number>
5
Omegavar
Equal/Ny98/M3
Equal
Covarion
No/Yes
No
Coding
All/Variable/Noabsencesites/
Nopresencesites
All
Parsmodel
No/Yes
No
------------------------------------------------------------------
2.4. Setting the Priors
We now need to set the priors for our model. There are six types of parameters in the
model: the topology, the branch lengths, the four stationary frequencies of the
nucleotides, the six different nucleotide substitution rates, the proportion of invariable
sites, and the shape parameter of the gamma distribution of rate variation. The default
priors in MrBayes work well for most analyses, and we will not change any of them here
so if you are impatient, you can skip this step and continue with “Checking the Model”.
However, it is good practice to go through all the priors and make sure you understand
the default settings and the available options. Start reviewing this information by typing
help prset to obtain a list of the default settings for the parameters in your model.
The table at the end of the help information reads:
MrBayes 3.1 Manual
5/26/2005
14
Model settings for partition 1:
Parameter
Options
Current Setting
-----------------------------------------------------------------Tratiopr
Beta/Fixed
Beta(1.0,1.0)
Revmatpr
Dirichlet/Fixed
Dirichlet(1.0,1.0,1.0,1.0,1.0,1.0)
Aamodelpr
Fixed/Mixed
Fixed(Poisson)
Aarevmatpr
Dirichlet/Fixed
Dirichlet(1.0,1.0,...)
Omegapr
Dirichlet/Fixed
Dirichlet(1.0,1.0)
Ny98omega1pr
Beta/Fixed
Beta(1.0,1.0)
Ny98omega3pr
Uniform/Exponential/Fixed
Exponential(1.0)
M3omegapr
Exponential/Fixed
Exponential
Codoncatfreqs
Dirichlet/Fixed
Dirichlet(1.0,1.0,1.0)
Statefreqpr
Dirichlet/Fixed
Dirichlet(1.0,1.0,1.0,1.0)
Ratepr
Fixed/Variable=Dirichlet
Fixed
Shapepr
Uniform/Exponential/Fixed
Uniform(0.0,50.0)
Ratecorrpr
Uniform/Fixed
Uniform(-1.0,1.0)
Pinvarpr
Uniform/Fixed
Uniform(0.0,1.0)
Covswitchpr
Uniform/Exponential/Fixed
Uniform(0.0,100.0)
Symmetricbetapr Uniform/Exponential/Fixed
Fixed(Infinity)
Topologypr
Uniform/Constraints
Uniform
Brlenspr
Unconstrained/Clock
Unconstrained:Exp(10.0)
Speciationpr
Uniform/Exponential/Fixed
Uniform(0.0,10.0)
Extinctionpr
Uniform/Exponential/Fixed
Uniform(0.0,10.0)
Sampleprob
<number>
1.00
Thetapr
Uniform/Exponential/Fixed
Uniform(0.0,10.0)
Growthpr
Uniform/Exponential/
Fixed/Normal
Fixed(0.0)
------------------------------------------------------------------
We need to focus on Revmatpr (for the six substitution rates of the GTR rate matrix),
Statefreqpr (for the stationary nucleotide frequencies of the GTR rate matrix),
Shapepr (for the shape parameter of the gamma distribution of rate variation),
Pinvarpr (for the proportion of invariable sites), Topologypr (for the topology), and
Brlenspr (for the branch lengths).
The default prior probability density is a flat Dirichlet (all values are 1.0) for both
Revmatpr and Statefreqpr. This is appropriate if we want estimate these
parameters from the data assuming no prior knowledge about their values. It is possible
to fix the rates and nucleotide frequencies but this is generally not recommended.
However, it is occasionally necessary to fix the nucleotide frequencies to be equal, for
instance in specifying the JC and SYM models. This would be achieved by typing
prset statefreqpr=fixed(equal).
If we wanted to specify a prior that put more emphasis on equal nucleotide frequencies
than the default flat Dirichlet prior, we could for instance use prset statefreqpr
= Dirichlet(10,10,10,10) or, for even more emphasis on equal frequencies,
prset statefreqpr=Dirichlet(100,100,100,100). The sum of the
numbers in the Dirichlet distribution determines how focused the distribution is, and the
balance between the numbers determines the expected proportion of each nucleotide (in
the order A, C, G, and T). Usually, there is a connection between the parameters in the
Dirichlet distribution and observations. Thus, you can think of a Dirichlet
(150,100,90,140) distribution as one arising from observing 150 A’s, 100 C’s, 90 G’s and
140 T’s in some set of sequences. If that set of sequences were independent of your set of
MrBayes 3.1 Manual
5/26/2005
15
sequences but clearly relevant to the analysis of your sequences, it might be reasonable to
use those numbers as a prior in your analysis.
In our analysis, we will be cautious and leave the prior on state frequencies at its default
setting. If you have changed the setting according to the suggestions above, you need to
change it back by typing prset statefreqpr=Dirichlet(1,1,1,1) or prs
st = Dir(1,1,1,1) if you want to save some typing. Similarly, we will leave the
prior on the substitution rates at the default flat Dirichlet(1,1,1,1,1,1) distribution.
The Shapepr parameter determines the prior for the α (shape) parameter of the gamma
distribution of rate variation. We will leave it at its default setting, a uniform distribution
spanning a wide range of α values. The prior for the proportion of invariable sites is set
with Pinvarpr. The default setting is a uniform distribution between 0 and 1, an
appropriate setting if we don’t want to assume any prior knowledge about the proportion
of invariable sites.
For topology, the default Uniform setting for the Topologypr parameter puts equal
probability on all distinct, fully resolved topologies. The alternative is to constrain some
nodes in the tree to always be present but we will not attempt that in this analysis.
The Brlenspr parameter can either be set to unconstrained or clock-constrained. For
trees without a molecular clock (unconstrained) the branch length prior can be set either
to exponential or uniform. The default exponential prior with parameter 10.0 should work
well for most analyses. It has an expectation of 1/10 = 0.1 but allows a wide range of
branch length values (theoretically from 0 to infinity). Because the likelihood values vary
much more rapidly for short branches than for long branches, an exponential prior on
branch lengths is closer to being uninformative than a uniform prior.
2.5. Checking the Model
To check the model before we start the analysis, type showmodel. This will give an
overview of the model settings. In our case, the output will be as follows:
Model settings:
Datatype
Nucmodel
Nst
Covarion
# States
Rates
= DNA
= 4by4
= 6
Substitution rates, expressed as proportions
of the rate sum, have a Dirichlet prior
(1.00,1.00,1.00,1.00,1.00,1.00)
= No
= 4
State frequencies have a Dirichlet prior
(1.00,1.00,1.00,1.00)
= Invgamma
Gamma shape parameter is uniformly distributed on the interval (0.05,50.00).
Proportion of invariable sites is uniformly distributed on the interval (0.00,1.00).
Gamma distribution is approximated using 4 categories.
MrBayes 3.1 Manual
5/26/2005
Active parameters:
Parameters
-----------------Revmat
1
Statefreq
2
Shape
3
Pinvar
4
Topology
5
Brlens
6
-----------------1 -2 -3 -4 -5 -6 --
Parameter
Prior
Parameter
Prior
Parameter
Prior
Parameter
Prior
Parameter
Prior
Parameter
Prior
=
=
=
=
=
=
=
=
=
=
=
=
Revmat
Dirichlet(1.00,1.00,1.00,1.00,1.00,1.00)
Statefreq
Dirichlet
Shape
Uniform(0.05,50.00)
Pinvar
Uniform(0.00,1.00)
Topology
All topologies equally probable a priori
Brlens
Branch lengths are Unconstrained:Exponential(10.0)
Note that we have six types of parameters in our model. All of these parameters will be
estimated during the analysis (see section 5 for information on how to fix parameter
values).
2.6. Setting up the Analysis
The analysis is started by issuing the mcmc command. However, before doing this, we
recommend that you review the run settings by typing help mcmc. The help mcmc
command will produce the following table at the bottom of the output:
Parameter
Options
Current Setting
----------------------------------------------------Seed
<number>
1115403472
Swapseed
<number>
1115403472
Ngen
<number>
1000000
Nruns
<number>
2
Nchains
<number>
4
Temp
<number>
0.200000
Reweight
<number>,<number>
0.00 v 0.00 ^
Swapfreq
<number>
1
Nswaps
<number>
1
Samplefreq
<number>
100
Printfreq
<number>
100
Printall
Yes/No
Yes
Printmax
<number>
8
Mcmcdiagn
Yes/No
Yes
Diagnfreq
<number>
1000
Minpartfreq
<number>
0.10
Allchains
Yes/No
No
Allcomps
Yes/No
No
Relburnin
Yes/No
Yes
Burnin
<number>
0
Burninfrac
<number>
0.25
Stoprule
Yes/No
No
Stopval
<number>
0.01
16
MrBayes 3.1 Manual
Filename
Startingtree
Nperts
Savebrlens
Ordertaxa
<name>
Random/User
<number>
Yes/No
Yes/No
5/26/2005
17
primates.nex.<p/t>
Random
0
Yes
No
---------------------------------------------------------------------------
The Seed is simply the seed for the random number generator, and Swapseed is the
seed for the separate random number generator used to generate the chain swapping
sequence (see below). Unless they are set to user-specified values, these seeds are
generated from the system clock, so your values are likely to be different from the ones in
the screen dump above. The Ngen setting is the number of generations for which the
analysis will be run. It is useful to run a small number of generations first to make sure
the analysis is correctly set up and to get an idea of how long it will take to complete a
longer analysis. We will start with 10,000 generations. To change the Ngen setting
without starting the analysis we use the mcmcp command, which is equivalent to mcmc
except that it does not start the analysis. Type mcmcp ngen=10000 to set the number
of generations to 10,000. You can type help mcmc to confirm that the setting was
changed appropriately.
By default, MrBayes will run two simultaneous, completely independent analyses starting
from different random trees (Nruns = 2). Running more than one analysis
simultaneously is very helpful in determining when you have a good sample from the
posterior probability distribution, so we suggest that you leave this setting as is. The idea
is to start each run from different randomly chosen trees. In the early phases of the run,
the two runs will sample very different trees but when they have reached convergence
(when they produce a good sample from the posterior probability distribution), the two
tree samples should be very similar.
To make sure that MrBayes compares tree samples from the different runs, make sure
that Mcmcdiagn is set to yes and that Diagnfreq is set to some reasonable value,
such as every 1000th generation. MrBayes will now calculate various run diagnostics
every Diagnfreq generation and print them to a file with the name
<Filename>.mcmc. The most important diagnostic, a measure of the similarity of the
tree samples in the different runs, will also be printed to screen every Diagnfreq
generation. Every time the diagnostics are calculated, either a fixed number of samples
(burnin) or a percentage of samples (burninfrac) from the beginning of the chain is
discarded. The relburnin setting determines whether a fixed burnin
(relburnin=no) or a burnin percentage (relburnin=yes) is used. If you don’t
change any settings, MrBayes will by default discard the first 25 % samples from the cold
chain (relburnin=yes and burninfrac=0.25).
By default, MrBayes uses Metropolis coupling to improve the MCMC sampling of the
target distribution. The Swapfreq, Nswaps, Nchains, and Temp settings together
control the Metropolis coupling behavior. When Nchains is set to 1, no heating is used.
When Nchains is set to a value n larger than 1, then n - 1 heated chains are used. By
MrBayes 3.1 Manual
5/26/2005
18
default, Nchains is set to 4, meaning that MrBayes will use 3 heated chains and one
"cold" chain. In our experience, heating is essential for problems with more than about 50
taxa, whereas smaller problems often can be analyzed successfully without heating.
Adding more than three heated chains may be helpful in analyzing large and difficult data
sets. The time complexity of the analysis is directly proportional to the number of chains
used (unless MrBayes runs out of physical RAM memory, in which case the analysis will
suddenly become much slower), but the cold and heated chains can be distributed among
processors in a cluster of computers using the MPI version of the program (see below),
greatly speeding up the calculations.
MrBayes uses an incremental heating scheme, in which chain i is heated by raising its
posterior probability to the power 1/ (1 + iλ), where λ is the temperature controlled by the
Temp parameter. The effect of the heating is to flatten out the posterior probability, such
that the heated chains more easily find isolated peaks in the posterior distribution and can
help the cold chain move more rapidly between these peaks. Every Swapfreq
generation, two chains are picked at random and an attempt is made to swap their states.
For many analyses, the default settings should work nicely. If you are running many more
than three heated chains, however, you may want to increase the number of swaps
(Nswaps) that are tried each time the chain stops for swapping.
The Samplefreq setting determines how often the chain is sampled. By default, the
chain is sampled every 100th generation, and this works well for most analyses.
However, our analysis is so small that we are likely to get convergence quickly, so it
makes sense to sample the chain more frequently, say every 10th generation (this will
ensure that we get at least 1,000 samples when the number of generations is set to
10,000). To change the sampling frequency, type mcmcp samplefreq=10.
When the chain is sampled, the current values of the model parameters are printed to file.
The substitution model parameters are printed to a .p file (in our case, there will be one
file for each independent analysis, and they will be called primates.nex.run1.p
and primates.nex.run2.p). The .p files are tab delimited text files that can be
imported into most statistics and graphing programs. The topology and branch lengths are
printed to a .t file (in our case, there will be two files called
primates.nex.run1.t and primates.nex.run2.t). The .t files are Nexus
tree files that can be imported into programs like PAUP* and TreeView. The root of the
.p and .t file names can be altered using the Filename setting.
The Printfreq parameter controls the frequency with which the state of the chains is
printed to screen. Leave Printfreq at the default value (print to screen every 100th
generation).
The default behavior of MrBayes is to save trees with branch lengths to the .t file. Since
this is what we want, we leave this setting as is. If you are running a large analysis (many
taxa) and are not interested in branch lengths, you can save a considerable amount of disk
space by not saving branch lengths.
MrBayes 3.1 Manual
5/26/2005
19
The Startingtree parameter can be used to feed the chain(s) with a user-specified
starting tree. The default behavior is to start each chain with a different random tree; this
is recommended for general use.
2.7. Running the Analysis
Finally, we are ready to start the analysis. Type mcmc. MrBayes will first print
information about the model and then list the proposal mechanisms that will be used in
sampling from the posterior distribution. In our case, the proposals are the following:
The MCMC sampler will use the following moves:
With prob. Chain will change
4.17 %
param. 1 (revmat) with Dirichlet proposal
4.17 %
param. 2 (state frequencies) with Dirichlet proposal
4.17 %
param. 3 (gamma shape) with multiplier
4.17 %
param. 4 (prop. invariants) with beta proposal
20.83 %
param. 5 (topology and branch lengths) with LOCAL
62.50 %
param. 5 (topology and branch lengths) with extending TBR
Note that MrBayes will spend most of its effort changing topology and branch lengths. In
our experience, topology and branch lengths are the most difficult parameters to integrate
over and we therefore let MrBayes spend a large proportion of its time proposing new
values for those parameters. The proposal probabilities can be changed with the props
command but be warned that inappropriate changes of proposal probabilities may destroy
any hopes of achieving convergence.
After the initial log likelihoods, MrBayes will print the state of the chains every 100th
generation, like this:
Chain results:
1 -- (-7812.831) (-7523.685) [-7485.569]
7618.595) (-7776.608) (-7836.826)
100 -- (-6771.532) (-6857.529) (-6766.678)
6944.449) (-6784.126) (-6991.307) -- 0:01:39
200 -- (-6321.464) [-6179.561] (-6338.168)
6715.840) [-6265.329] (-6599.698) -- 0:01:38
300 -- (-6201.285) (-6084.899) (-6139.200)
6359.134) (-6106.834) (-6515.348) -- 0:01:37
400 -- (-6015.429) [-5924.177] (-6074.274)
6215.415) [-5980.096] (-6036.624) -- 0:01:12
500 -- (-5963.069) [-5851.415] (-6019.301)
6072.446) (-5925.549) (-5963.427) -- 0:01:16
600 -- (-5931.545) [-5802.472] (-5997.276)
6035.097) (-5904.327) [-5896.520] -- 0:01:18
700 -- (-5852.405) [-5782.934] (-5966.224)
6008.142) (-5876.956) [-5838.392] -- 0:01:19
800 -- (-5844.426) [-5754.416] (-5963.531)
5902.784) (-5790.765) [-5789.326] -- 0:01:20
900 -- (-5814.885) [-5741.101] (-5926.666)
5871.573) (-5783.012) [-5755.608] -- 0:01:20
1000 -- (-5811.949) [-5737.884] (-5888.234)
5851.693) (-5784.437) [-5749.264] -- 0:01:21
(-7700.309) * [-7832.045] ([-6682.527] * [-6506.277] ((-6272.242) * (-6339.400) ([-6049.061] * [-6073.056] ((-5967.236) * (-6002.941) ((-5948.768) * [-5925.423] ((-5927.204) * (-5899.590) ((-5880.975) * (-5890.220) ((-5860.182) * (-5883.474) ((-5855.916) * (-5870.806) ((-5819.793) * (-5867.377) (-
Average standard deviation of split frequencies: 0.073946
MrBayes 3.1 Manual
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20
1100 -- (-5784.586) [-5730.069] (-5880.476) (-5798.728) * (-5839.268) (5836.074) (-5779.940) [-5739.170] -- 0:01:20
...
9000 -- (-5737.756) [-5728.228] (-5735.787) (-5722.263) * [-5730.926] (5740.655) (-5730.045) (-5729.487) -- 0:00:09
Average standard deviation of split frequencies: 0.000000
9100 -- (-5732.093) (-5733.215) (-5735.816)
5741.841) (-5735.730) [-5726.010] -- 0:00:08
9200 -- (-5731.412) (-5728.251) (-5729.846)
5740.270) (-5742.079) [-5729.764] -- 0:00:07
9300 -- (-5739.416) (-5734.909) [-5723.707]
5728.550) [-5730.645] (-5726.709) -- 0:00:06
9400 -- [-5720.648] (-5732.631) (-5732.691)
5720.959] (-5739.326) (-5720.832) -- 0:00:05
9500 -- (-5730.870) (-5729.773) (-5729.010)
5720.702] (-5737.968) (-5726.367) -- 0:00:04
9600 -- (-5736.460) (-5734.016) (-5737.829)
5718.892] (-5736.313) (-5727.928) -- 0:00:03
9700 -- (-5740.378) (-5733.312) (-5735.204)
5724.006] (-5736.798) (-5722.292) -- 0:00:02
9800 -- (-5739.019) (-5735.541) (-5731.738)
5722.901) (-5734.184) [-5722.141] -- 0:00:01
9900 -- (-5743.556) [-5724.246] (-5736.484)
5732.476) (-5728.482) (-5725.045) -- 0:00:00
10000 -- (-5738.106) [-5723.380] (-5726.322)
5732.410) [-5728.515] (-5725.418) -- 0:00:00
[-5720.361] * (-5737.343) ([-5720.670] * (-5731.852) ((-5720.408) * (-5728.016) ((-5724.562) * (-5728.739) [[-5727.405] * (-5727.274) [[-5727.732] * (-5728.393) [[-5724.199] * (-5728.102) [[-5728.466] * (-5723.404) ((-5731.803) * [-5727.082] ((-5727.599) * (-5728.776) (-
Average standard deviation of split frequencies: 0.000000
Continue with analysis? (yes/no):
If you have the terminal window wide enough, each generation of the chain will print on
a single line like this (we have decreased the font size to show the layout):
Chain results:
1
100
200
300
400
500
600
700
800
900
1000
------------
(-7812.831)
(-6771.532)
(-6321.464)
(-6201.285)
(-6015.429)
(-5963.069)
(-5931.545)
(-5852.405)
(-5844.426)
(-5814.885)
(-5811.949)
(-7523.685)
(-6857.529)
[-6179.561]
(-6084.899)
[-5924.177]
[-5851.415]
[-5802.472]
[-5782.934]
[-5754.416]
[-5741.101]
[-5737.884]
[-7485.569]
(-6766.678)
(-6338.168)
(-6139.200)
(-6074.274)
(-6019.301)
(-5997.276)
(-5966.224)
(-5963.531)
(-5926.666)
(-5888.234)
(-7700.309)
[-6682.527]
(-6272.242)
[-6049.061]
(-5967.236)
(-5948.768)
(-5927.204)
(-5880.975)
(-5860.182)
(-5855.916)
(-5819.793)
*
*
*
*
*
*
*
*
*
*
*
[-7832.045]
[-6506.277]
(-6339.400)
[-6073.056]
(-6002.941)
[-5925.423]
(-5899.590)
(-5890.220)
(-5883.474)
(-5870.806)
(-5867.377)
(-7618.595)
(-6944.449)
(-6715.840)
(-6359.134)
(-6215.415)
(-6072.446)
(-6035.097)
(-6008.142)
(-5902.784)
(-5871.573)
(-5851.693)
(-7776.608)
(-6784.126)
[-6265.329]
(-6106.834)
[-5980.096]
(-5925.549)
(-5904.327)
(-5876.956)
(-5790.765)
(-5783.012)
(-5784.437)
(-7836.826)
(-6991.307)
(-6599.698)
(-6515.348)
(-6036.624)
(-5963.427)
[-5896.520]
[-5838.392]
[-5789.326]
[-5755.608]
[-5749.264]
-----------
0:01:39
0:01:38
0:01:37
0:01:12
0:01:16
0:01:18
0:01:19
0:01:20
0:01:20
0:01:21
Average standard deviation of split frequencies: 0.073946
1100 -- (-5784.586) [-5730.069] (-5880.476) (-5798.728) * (-5839.268) (-5836.074) (-5779.940) [-5739.170] -- 0:01:20
...
9000 -- (-5737.756) [-5728.228] (-5735.787) (-5722.263) * [-5730.926] (-5740.655) (-5730.045) (-5729.487) -- 0:00:09
Average standard deviation of split frequencies: 0.000000
9100
9200
9300
9400
9500
9600
9700
9800
9900
10000
-----------
(-5732.093)
(-5731.412)
(-5739.416)
[-5720.648]
(-5730.870)
(-5736.460)
(-5740.378)
(-5739.019)
(-5743.556)
(-5738.106)
(-5733.215)
(-5728.251)
(-5734.909)
(-5732.631)
(-5729.773)
(-5734.016)
(-5733.312)
(-5735.541)
[-5724.246]
[-5723.380]
(-5735.816)
(-5729.846)
[-5723.707]
(-5732.691)
(-5729.010)
(-5737.829)
(-5735.204)
(-5731.738)
(-5736.484)
(-5726.322)
[-5720.361]
[-5720.670]
(-5720.408)
(-5724.562)
[-5727.405]
[-5727.732]
[-5724.199]
[-5728.466]
(-5731.803)
(-5727.599)
*
*
*
*
*
*
*
*
*
*
Average standard deviation of split frequencies: 0.000000
(-5737.343)
(-5731.852)
(-5728.016)
(-5728.739)
(-5727.274)
(-5728.393)
(-5728.102)
(-5723.404)
[-5727.082]
(-5728.776)
(-5741.841)
(-5740.270)
(-5728.550)
[-5720.959]
[-5720.702]
[-5718.892]
[-5724.006]
(-5722.901)
(-5732.476)
(-5732.410)
(-5735.730)
(-5742.079)
[-5730.645]
(-5739.326)
(-5737.968)
(-5736.313)
(-5736.798)
(-5734.184)
(-5728.482)
[-5728.515]
[-5726.010]
[-5729.764]
(-5726.709)
(-5720.832)
(-5726.367)
(-5727.928)
(-5722.292)
[-5722.141]
(-5725.045)
(-5725.418)
-----------
0:00:08
0:00:07
0:00:06
0:00:05
0:00:04
0:00:03
0:00:02
0:00:01
0:00:00
0:00:00
MrBayes 3.1 Manual
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21
Continue with analysis? (yes/no):
The first column lists the generation number. The following four columns with negative
numbers each correspond to one chain in the first run. Each column corresponds to one
physical location in computer memory, and the chains actually shift positions in the
columns as the run proceeds. The numbers are the log likelihood values of the chains.
The chain that is currently the cold chain has its value surrounded by square brackets,
whereas the heated chains have their values surrounded by parentheses. When two chains
successfully change states, they trade column positions (places in computer memory). If
the Metropolis coupling works well, the cold chain should move around among the
columns; this means that the cold chain successfully swaps states with the heated chains.
If the cold chain gets stuck in one of the columns, then the heated chains are not
successfully contributing states to the cold chain, and the Metropolis coupling is
inefficient. The analysis may then have to be run longer or the temperature difference
between chains may have to be lowered.
The star column separates the two different runs. The last column gives the time left to
completion of the specified number of generations. This analysis approximately takes 1
second per 100 generations. Because different moves are used in each generation, the
exact time varies somewhat for each set of 100 generations, and the predicted time to
completion will be unstable in the beginning of the run. After a while, the predictions will
become more accurate and the time will decrease predictably.
2.8. When to Stop the Analysis
At the end of the run, MrBayes asks whether or not you want to continue with the
analysis. Before answering that question, examine the average standard deviation of split
frequencies. As the two runs converge onto the stationary distribution, we expect the
average standard deviation of split frequencies to approach zero, reflecting the fact that
the two tree samples become increasingly similar. In our case, the average standard
deviation is about 0.07 after 1,000 generations and then drops to less than 0.000001
towards the end of the run. Your values can differ slightly because of stochastic effects.
Given the extremely low value of the average standard deviation at the end of the run,
there appears to be no need to continue the analysis beyond 10,000 generations so when
MrBayes asks “Continue with analysis? (yes/no):”, stop the analysis by
typing no.
Although we recommend using a convergence diagnostic, such as the standard deviation
of split frequencies, to determine run length, we want to mention that there are simpler
but less powerful methods of determining when to stop the analysis. Arguably the
simplest technique is to examine the log likelihood values (or, more exactly, the log
probability of the data given the parameter values) of the cold chain, that is, the values
printed to screen within square brackets. In the beginning of the run, the values typically
increase rapidly (the absolute values decrease, since these are negative numbers). This
phase of the run is referred to as the “burn-in” and the samples from this phase are
typically discarded. Once the likelihood of the cold chain stops to increase and starts to
randomly fluctuate within a more or less stable range, the run may have reached
MrBayes 3.1 Manual
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22
stationarity, that is, it is producing a good sample from the posterior probability
distribution. At stationarity, we also expect different, independent runs to sample similar
likelihood values. Trends in likelihood values can be deceiving though; you’re more
likely to detect problems with convergence by comparing split frequencies than by
looking at likelihood trends.
When you stop the analysis, MrBayes will print several types of information useful in
optimizing the analysis. This is primarily of interest if you have difficulties in obtaining
convergence. Since we apparently have a good sample from the posterior distribution
already, we will ignore this information for now. We will return to the subject of
optimizing the MCMC analysis in section 5 of the manual.
2.9. Summarizing Samples of Substitution Model Parameters
During the run, samples of the substitution model parameters have been written to the .p
files every samplefreq generation. These files are tab-delimited text files that look
something like this:
[ID: 5848203808]
Gen LnL
1 -7559.137
10 -6585.519
...
9990 -5728.935
10000 -5722.857
TL
2.044
2.181
r(A<->C)
0.166667
0.090180
...
...
...
pi(G)
0.250000
0.169513
pi(T)
0.250000
0.247418
alpha
0.500000
0.569271
pinvar
0.000000
0.036162
2.527
2.642
0.051106
0.051106
...
...
0.076213
0.078383
0.262107
0.246791
0.932771
0.721217
0.194004
0.185486
The first number, in square brackets, is a randomly generated ID number that lets you
identify the analysis from which the samples come. The next line contains the column
headers, and is followed by the sampled values. From left to right, the columns contain:
(1) the generation number (Gen); (2) the log likelihood of the cold chain (LnL); (3) the
total tree length (the sum of all branch lengths, TL); (4) the six GTR rate parameters
(r(A<->C), r(A<->G) etc); (5) the four stationary nucleotide frequencies (pi(A),
pi(C) etc); (6) the shape parameter of the gamma distribution of rate variation
(alpha); and (7) the proportion of invariable sites (pinvar). If you use a different
model for your data set, the .p files will of course be different.
MrBayes provides the sump command to summarize the sampled parameter values.
Before using it, we need to decide on the burn-in. Since the convergence diagnostic we
used previously to determine when to stop the analysis discarded the first 25 % of the
samples and indicated that convergence had been reached after 10,000 generations, it
makes sense to discard 25 % of the samples obtained during the first 10,000 generations.
Since we sampled every 10th generation, there are 1,000 samples (1,001 to be exact, since
the first generation is always sampled) and 25 % translates to 250 samples. Thus,
summarize the information in the .p file by typing sump burnin=250. By default,
sump will summarize the information in the .p file generated most recently, but the
filename can be changed if necessary.
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23
The sump command will first generate a plot of the generation versus the log probability
of the data (the log likelihood values). If we are at stationarity, this plot should look like
‘white noise’, that is, there should be no tendency of increase or decrease over time. The
plot should look something like this:
+------------------------------------------------------------+ -5716.56
|
1
1
|
|
2
1
12
1
1 2|
|
1
2
1
1
1
2 |
|
1
22
2
1
1
2
2 |
| 2
2
2 21
2
1
1 2
|
|
22
2 1
1
1 1
12
1
|
|2
1 **1
2 2
2 * 2 2
1211
1 |
| 11
1
*
*
2
1
2
|
| 2 2 1
1 1 12
112
2
1
2 1|
|1
2
1 *2 2 2
2 1
2
2 22 1
|
|
1
1
2
2
2
1
2
|
|
1
2
2
1
|
|
21
11
|
|
2 1
2
|
|
1
2
|
+------+-----+-----+-----+-----+-----+-----+-----+-----+-----+ -5733.79
^
^
2500
10000
If you see an obvious trend in your plot, either increasing or decreasing, you probably
need to run the analysis longer to get an adequate sample from the posterior probability
distribution.
At the bottom of the sump output, there is a table summarizing the samples of the
parameter values:
Model parameter summaries over the runs sampled in files
"primates.nex.run1.p" and "primates.nex.run2.p":
(Summaries are based on a total of 1502 samples from 2 runs)
(Each run produced 1001 samples of which 751 samples were included)
95% Cred. Interval
---------------------Parameter
Mean
Variance
Lower
Upper
Median
PSRF *
----------------------------------------------------------------------------------------TL
2.885314
0.071446
2.408000
3.464000
2.861000
1.004
r(A<->C)
0.045110
0.000070
0.031406
0.061424
0.044756
0.999
r(A<->G)
0.477554
0.002421
0.387141
0.569168
0.473521
1.040
r(A<->T)
0.038541
0.000060
0.024204
0.053787
0.038402
0.999
r(C<->G)
0.033765
0.000195
0.010836
0.064049
0.032475
1.033
r(C<->T)
0.386144
0.001735
0.311504
0.459011
0.386897
1.012
r(G<->T)
0.018885
0.000133
0.001217
0.045924
0.016801
1.048
pi(A)
0.355301
0.000162
0.329769
0.377153
0.356808
1.092
pi(C)
0.320831
0.000131
0.299953
0.341757
0.321312
1.152
pi(G)
0.081154
0.000048
0.068816
0.098442
0.081723
1.014
pi(T)
0.242714
0.000104
0.226573
0.264684
0.242113
1.000
alpha
0.714405
0.037305
0.419210
1.147286
0.673545
1.007
pinvar
0.185901
0.004887
0.025889
0.307468
0.188027
1.009
----------------------------------------------------------------------------------------* Convergence diagnostic (PSRF = Potential scale reduction factor [Gelman
and Rubin, 1992], uncorrected) should approach 1 as runs converge. The
values may be unreliable if you have a small number of samples. PSRF should
only be used as a rough guide to convergence since all the assumptions
that allow one to interpret it as a scale reduction factor are not met in
the phylogenetic context.
MrBayes 3.1 Manual
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24
For each parameter, the table lists the mean and variance of the sampled values, the lower
and upper boundaries of the 95 % credibility interval, and the median of the sampled
values. The parameters are the same as those listed in the .p files: the total tree length
(TL), the six reversible substitution rates (r(A<->C), r(A<->G), etc), the four
stationary state frequencies (pi(A), pi(C), etc), the shape of the gamma distribution of
rate variation across sites (alpha), and the proportion of invariable sites (pinvar).
Note that the six rate parameters of the GTR model are given as proportions of the rate
sum (the Dirichlet parameterization). This parameterization has some advantages in the
Bayesian context; in particular, it allows convenient formulation of priors. If you want to
scale the rates relative to the G-T rate, just divide all rate proportions by the G-T rate
proportion.
The last column in the table contains a convergence diagnostic, the Potential Scale
Reduction Factor (PSRF). If we have a good sample from the posterior probability
distribution, these values should be close to 1.0. If you have a small number of samples,
there may be some spread in these values, indicating that you may need to sample the
analysis more often or run it longer. In our case, we can probably easily obtain more
accurate estimates of some parameters by running the analysis slightly longer.
2.10. Summarizing Samples of Trees and Branch Lengths
Trees and branch lengths are printed to the .t files. These files are Nexus-formatted tree
files with a structure like this:
#NEXUS
[ID: 5848203808]
begin trees;
translate
1 Lemur_catta,
2 Homo_sapiens,
3 Pan,
4 Gorilla,
5 Pongo,
6 Hylobates,
7 Macaca_fuscata,
8 M_mulatta,
9 M_fascicularis,
10 M_sylvanus,
11 Saimiri_sciureus,
12 Tarsius_syrichta;
tree rep.1 =
((((((2:0.100000,(8:0.100000,7:0.000748):0.197415):0.100000,(5:0.100000,6:0.100
000):0.100000):0.100000,(9:0.100000,(10:0.100000,3:0.100000):0.100000):0.100000
):0.100000,12:0.100000):0.100000,11:0.100000):0.100000,4:0.100000,1:0.100000);
...
tree rep.10000 =
((((6:0.126475,((4:0.079300,(3:0.049876,2:0.066929):0.040517):0.103191,5:0.1382
20):0.105224):0.147212,(10:0.059671,((8:0.035939,7:0.012332):0.037992,9:0.06947
7):0.060859):0.221336):0.149243,11:0.627561):0.180115,12:0.525606,1:0.339476);
end;
To summarize the tree and branch length information, type sumt burnin=250. The
sumt and sump commands each have separate burn-in settings so it is necessary to give
MrBayes 3.1 Manual
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25
the burn-in here again. Otherwise, many of the settings in MrBayes are persistent and
need not be repeated every time a command is executed. To make sure the settings for a
particular command are correct, you can always use help <command> before issuing
the command.
The sumt command will output, among other things, summary statistics for the taxon
bipartitions, a tree with clade credibility (posterior probability) values, and a phylogram
(if branch lengths have been saved). The summary statistics (see below) describes each
partition in the “dot-star” format (dots for the taxa that are on one side of the partition and
stars for the taxa on the other side; for instance, the first partition (ID 1) is the terminal
branch leading to taxon 8 since it has a star in the 8th position and a dot in all other
positions). Then it gives the number of times the partition was sampled (#obs), the
probability of the partition (Probab.), the standard deviation of the partition frequency
(Stdev(s)), the mean (Mean(v)) and variance (Var(v)) of the branch length, the
Potential Scale Reduction Factor (PSRF), and finally the number of runs in which the
partition was sampled (Nruns). In our analysis, there is overwhelming support for a
single tree, so all partitions have a posterior probability of 1.0.
Summary statistics for taxon bipartitions:
ID -- Partition
#obs
Probab. Stdev(s)
Mean(v)
Var(v)
PSRF Nruns
-----------------------------------------------------------------------------1 -- ....*....... 1502 1.000000 0.000000 0.058828 0.000235 1.002
2
2 -- ..***....... 1502 1.000000 0.000000 0.082206 0.000426 1.000
2
3 -- ..**........ 1502 1.000000 0.000000 0.030594 0.000152 0.999
2
4 -- ..*......... 1502 1.000000 0.000000 0.049234 0.000104 1.002
2
5 -- ...*........ 1502 1.000000 0.000000 0.063670 0.000207 1.001
2
6 -- ..********** 1502 1.000000 0.000000 0.271433 0.003850 0.999
2
7 -- .*********** 1502 1.000000 0.000000 0.472390 0.007137 1.010
2
8 -- ..*********. 1502 1.000000 0.000000 0.113412 0.002208 0.999
2
9 -- ......*..... 1502 1.000000 0.000000 0.165887 0.000861 1.017
2
10 -- ..*****..... 1502 1.000000 0.000000 0.122957 0.001186 1.009
2
11 -- ........*... 1502 1.000000 0.000000 0.022199 0.000039 1.003
2
12 -- .......*.... 1502 1.000000 0.000000 0.018330 0.000042 1.002
2
13 -- ..........*. 1502 1.000000 0.000000 0.070973 0.000449 1.043
2
14 -- .*.......... 1502 1.000000 0.000000 0.329111 0.003588 1.008
2
15 -- ...........* 1502 1.000000 0.000000 0.429012 0.004224 1.003
2
16 -- .........*.. 1502 1.000000 0.000000 0.056534 0.000137 1.009
2
17 -- .......****. 1502 1.000000 0.000000 0.248962 0.002162 1.005
2
18 -- .....*...... 1502 1.000000 0.000000 0.139867 0.000708 1.001
2
19 -- .......**... 1502 1.000000 0.000000 0.035640 0.000107 1.020
2
20 -- ..****...... 1501 0.999334 0.000942 0.054652 0.000435 1.022
2
21 -- .......***.. 1501 0.999334 0.000942 0.049466 0.000355 1.010
2
------------------------------------------------------------------------------
The clade credibility tree (upper tree, next page) gives the probability of each partition or
clade in the tree, and the phylogram (lower tree, next page) gives the branch lengths
measured in expected substitutions per site.
MrBayes 3.1 Manual
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26
Clade credibility values:
/--------------------------------------------------------(1)
|
|--------------------------------------------------------|
|
/-------|
/--100--+
|
|
\-------|
/--100--+
|
|
\---------------|
/---100--+
+
|
\-----------------------|
/--100--+
|
|
\--------------------------------|
|
|
|
/-------(8)
|
/--100--+
/--100--+
|
|
|
|
\-------|
|
|
/--100--+
|
|
|
|
\---------------(10)
\--100--+
\-------100------+
|
\-----------------------|
\------------------------------------------------(12)
Tarsius_syrichta
Lemur_catta (2)
Homo_sapiens (3)
Pan (4)
Gorilla (5)
Pongo (6)
Hylobates (7)
Macaca_fuscata
M_mulatta (9)
M_fascicularis
M_sylvanus (11)
Saimiri_sciureus
Phylogram:
/-------------------------------------- Tarsius_syrichta (1)
|
|--------------------------- Lemur_catta (2)
|
|
/---- Homo_sapiens (3)
|
/--+
|
| \----- Pan (4)
|
/-----+
|
|
\----- Gorilla (5)
|
/----+
+
|
\----------- Pongo (6)
|
/--------+
|
|
\-------------- Hylobates (7)
|
|
|
|
/-- Macaca_fuscata
(8)
|
/--------+
/--+
|
|
|
| \-- M_mulatta (9)
|
|
|
/--+
|
|
|
| \----- M_fascicularis
(10)
\--------------------+
\--------------------+
|
\----- M_sylvanus (11)
|
\---------------------------------- Saimiri_sciureus
(12)
|--------| 0.100 expected changes per site
MrBayes 3.1 Manual
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27
In the background, the sumt command creates three additional files. The first is a
.parts file, which contains the list of taxon bipartitions, their posterior probability (the
proportion of sampled trees containing them), and the branch lengths associated with
them (if branch lengths have been saved). The branch length values are based only on
those trees containing the relevant bipartition. The second generated file has the suffix
.con and includes two consensus trees. The first one has both the posterior probability
of clades (as interior node labels) and the branch lengths (if they have been saved) in its
description. A graphical representation of this tree can be generated in Rod Page’s
program TreeView. The second tree only contains the branch lengths and it can be
imported into a wide range of tree-drawing programs such as MacClade and Mesquite.
The third file generated by the sumt command is the .trprobs file, which contains the
trees that were found during the MCMC search, sorted by posterior probability.
3. Analyzing a Partitioned Data Set
MrBayes handles a wide variety of data types and models, as well as any mix of these
models. In this example we will look at how to set up a simple analysis of a combined
data set, consisting of data from four genes and morphology for 30 taxa of gall wasps and
outgroups. A similar approach can be used, e.g., to set up a partitioned analysis of
molecular data coming from different genes. The data set for this tutorial is found in the
file cynmix.nex.
3.1. Getting Mixed Data into MrBayes
First, open up the Nexus data file in a text editor. The DATA block of the Nexus file
should look familiar but there are some differences compared to the primates.nex
file in the format statement:
Format datatype=mixed(Standard:1-166,DNA:167-3246) interleave=yes gap=missing=?;
First, the datatype is specified as datatype=mixed(Standard:1-166,
DNA:167-3246). This means that the matrix contains standard (morphology)
characters in columns 1-166 and DNA characters in the remaining columns. The mixed
datatype is an extension to the Nexus ‘standard’. This extension was originated by
MrBayes 3 and may not be compatible with other phylogenetics programs.
Second, the matrix is interleaved. It is often convenient to specify mixed data in
interleaved format, with each block consisting of a natural subset of the matrix, such as
the morphological data or one of the gene regions.
Third, the Nexus file contains a MrBayes block at the end, after the data block. The
MrBayes block begins with begin mrbayes; and ends with the end; statement. The block
contains various instructions to the MrBayes program; for the purposes of the tutorial,
comment out these instructions by adding an opening square bracket before the begin
statement ([begin mrbayes;) and a closing square bracket after the end statement
(end;]). MrBayes will skip everything within square brackets (nested square brackets
MrBayes 3.1 Manual
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28
are allowed), so this will ensure that the entire existing MrBayes block is ignored when
the data file is processed.
3.2. Dividing the Data into Partitions
By default, MrBayes partitions the data according to data type. There are only two data
types in the matrix, so this model will include only a morphology (standard) and a DNA
partition. To divide the DNA partition into gene regions, it is convenient to first specify
character sets. In principle, this can be done from the command line but it is more
convenient to do it in a MrBayes block in the data file. In your text editor, add the
following new MrBayes block after the existing block that you just commented out (note
that each line must be terminated by a semicolon):
begin mrbayes;
charset morphology = 1-166;
charset COI = 167-1244;
charset EF1a = 1245-1611;
charset LWRh = 1612-2092;
charset 28S = 2093-3246;
The charset command simply associates a name with a set of characters. For instance,
the character set COI is defined above to include characters 167 to 1244. The next step is
to define a partition of the data according to genes and morphology. This is accomplished
with the line (add it after the lines above):
partition favored = 5: morphology, COI, EF1a, LWRh, 28S;
The elements of the partition command are: (1) the name of the partitioning scheme
(favored); (2) an equal sign (=); (3) the number of character divisions in the scheme
(5); (4) a colon (:); and (5) a list of the characters in each division, separated by commas.
The list of characters can simply be an enumeration of the character numbers (the above
line is equivalent to partition favored = 5: 1-166, 167-1244, 12451611, 1612-2092, 2093-3246;) but it is often more convenient to use
predefined character sets like we did above. The final step is to tell MrBayes that we
want to work with this partitioning of the data instead of the default partitioning. We do
this using the set command:
set partition = favored;
Finally, we need to add an end statement to close the MrBayes block. The entire block
should now look like this:
begin mrbayes;
charset morphology = 1-166;
charset COI = 167-1244;
charset EF1a = 1245-1611;
charset LWRh = 1612-2092;
charset 28S = 2093-3246;
partition favored = 5: morphology, COI, EF1a, LWRh, 28S;
set partition = favored;
end;
MrBayes 3.1 Manual
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29
When we read this block into MrBayes, we will get a partitioned model with the first
character division being morphology, the second division being the COI gene, etc. Save
the data file, exit your text editor, and finally launch MrBayes and type execute
cynmix.nex to read in your data and set up the partitioning scheme.
3.3. Specifying a Partitioned Model
Before starting to specify the partitioned model, it is useful to examine the default model.
Type showmodel and you should get this table as part of the output:
Active parameters:
Partition(s)
Parameters
1 2 3 4 5
-----------------------------Statefreq
1 2 2 2 2
Topology
3 3 3 3 3
Brlens
4 4 4 4 4
------------------------------
There is a lot of other useful information in the output of showmodel but this table is
the key to the partitioned model. We can see that there are five partitions in the model
and four active (free) parameters. There are two stationary state frequency parameters,
one for the morphological data (parameter 1) and one for the DNA data (parameter 2).
Then there is also a topology parameter (3) and a set of branch length parameters (4).
Both the topology and branch lengths are the same for all partitions.
Now, assume we want a separate GTR + Γ + I model for each gene partition. All the
parameters should be estimated separately for the individual genes. Assume further that
we want the overall evolutionary rate to be (potentially) different across partitions, and
that we want to assume gamma-shaped rate variation for the morphological data. We can
obtain this model by using lset and prset with the applyto mechanism, which
allows us to apply the settings to specific partitions. For instance, to apply a GTR + _ + I
model to the molecular partitions, we type lset applyto=(2,3,4,5) nst=6
rates=invgamma. This will produce the following table when showmodel is
invoked:
Active parameters:
Partition(s)
Parameters
1 2 3 4 5
-----------------------------Revmat
. 1 1 1 1
Statefreq
2 3 3 3 3
Shape
. 4 4 4 4
Pinvar
. 5 5 5 5
Topology
6 6 6 6 6
Brlens
7 7 7 7 7
------------------------------
As you can see, all molecular partitions now evolve under the correct model but all
parameters (statefreq, revmat, shape, pinvar) are shared across partitions. To
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30
unlink them such that each partition has its own set of parameters, type: unlink
statefreq=(all) revmat=(all) shape=(all) pinvar=(all). Gammashaped rate variation for the morphological data is enforced with lset applyto=(1)
rates=gamma. The trickiest part is to allow the overall rate to be different across
partitions. This is achieved using the ratepr parameter of the prset command. By
default, ratepr is set to fixed, meaning that all partitions have the same overall rate.
By changing this to variable, the rates are allowed to vary under a flat Dirichlet prior
(see the help info for prset if you want to modify this prior). To allow all our partitions
to evolve under different rates, type prset applyto=(all) ratepr=variable.
The model is now essentially complete but there is one final thing to consider. Typically
morphological data matrices do not include all types of characters. Specifically,
morphological data matrices do not usually include any constant (invariable) characters.
Sometimes, autapomorphies are not included either, and the matrix is restricted to
parsimony-informative characters. For MrBayes to calculate the probability of the data
correctly, we need to inform it of this ascertainment (coding) bias. By default, MrBayes
assumes that standard data sets include all variable characters but no constant characters.
If necessary, one can change this setting using lset coding. We will leave the
coding setting at the default, though. Now, showmodel should produce this table:
Active parameters:
Partition(s)
Parameters
1 2 3 4 5
-----------------------------Revmat
. 1 2 3 4
Statefreq
5 6 7 8 9
Shape
10 11 12 13 14
Pinvar
. 15 16 17 18
Ratemultiplier 19 19 19 19 19
Topology
20 20 20 20 20
Brlens
21 21 21 21 21
------------------------------
3.4. Running the Analysis
When the model has been completely specified, we can proceed with the analysis
essentially as described above in the previous tutorial for the primates.nex data set.
However, in the case of the cynmix.nex dataset, the analysis will have to be run longer
before there is any hope of adequate convergence to the stationary distribution.
When looking at the parameter samples from a partitioned analysis, it is useful to know
that the names of the parameters are followed by the character division (partition) number
in curly braces. For instance, pi(A){3} is the stationary frequency of nucleotide A in
character division 3, which is the EF1a division in the above analysis.
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4. Evolutionary Models Implemented in MrBayes 3
MrBayes implements a wide variety of evolutionary models for nucleotide, amino acid,
restriction site (binary), and standard discrete data. In addition, there are several different
ways of modeling the process generating phylogenetic trees. An overview of all the
models is given in diagrammatic form in the Appendix. Here, we provide a brief
description of each model with some comments on their implementation in MrBayes and
advice on how you can apply them successfully to your data.
4.1. Nucleotide Models
MrBayes implements a large number of DNA substitution models. These models are of
three different structures. The "4by4" models are the usual simple models of nucleotide
evolution. The “Doublet” model is intended for stem regions of ribosomal DNA, where
nucleotides evolve in pairs. Finally, the “Codon” models group the nucleotides in triplets
and model evolution based on these. The type of nucleotide model is set in MrBayes with
lset nucmodel; for instance, if you want to use the doublet model, the command is
lset nucmodel=doublet. The default setting is 4by4.
4.1.1. Simple Nucleotide Models
There has been more work based on the simple four by four nucleotide models than on
any other type of evolutionary model for molecular data. MrBayes 3 implements three
main types of models; you select among those by setting the number of substitution types
using lset nst to 1, 2, or 6. The widely used General Time Reversible (GTR) model
has six substitution types (lset nst=6), one for each pair of nucleotides. The
instantaneous rate matrix for the GTR model is (note that we order the rows and columns
alphabetically - A, C, G, T - unlike some other authors)
[A]

[A]
−

Q = [C] π A rAC

[G] π A rAG
[T] π A rAT
[C]
π C rAC
−
π C rCG
π C rCT
[G]
π G rAG
π G rCG
−
π G rGT
[T] 
π T rAT 

π T rCT  .

π T rGT 
− 
The GTR model (Tavaré, 1986) has four stationary state frequencies (πA, πC, πG, πT) and
six rate parameters (rAC, rAG, rAT, rCG, rCT, rGT). In total, there are eight free parameters,
since one of the stationary state frequencies and one of the substitution rates are
determined by the others. By default, MrBayes uses a “flat” Dirichlet distribution (with
all distribution parameters set to 1) as the prior for both the stationary state frequencies
and the substitution rates. This is a reasonable uninformative prior that should work well
for most analyses.
During the analysis, both the stationary state frequencies and substitution rates are
estimated. If you want to fix the stationary state frequencies or substitution rates, you can
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do that by using the prset command. For instance, assume that we want to fix the
stationary state frequencies to be equal, converting the GTR model to the so-called SYM
model. This is achieved by prset statefreqpr=fixed(0.25,0.25,0.25,
0.25) or, more conveniently, prset statefreqpr=fixed(equal). The
substitution rate matrix now becomes
 −
0.25r
AC
Q=
0.25rAG

0.25rAT
0.25rAC
−
0.25rCG
0.25rCT
0.25rAG
0.25rCG
−
0.25rGT
0.25rAT 
0.25rCT 

0.25rGT 

− 
and the stationary state frequencies are no longer estimated during the analysis.
Similarly, it is possible to fix the substitution rates of the GTR model using prset
revmatpr=fixed. Assume, for instance, that we want to fix the substitution rates to
be (rAC = 0.11, rAG = 0.22, rAT = 0.12, rCG = 0.14, rCT = 0.35, rGT = 0.06). Then the correct
statement would be prset revmatpr=fixed(0.11,0.22,0.12,0.14,0.35,
0.06). Note the order of the rates. The substitution rates can be given either as
percentages of the rate sum, as here, or they can be scaled to the rGT rate. The former
representation is the Dirichlet parameterization used internally by MrBayes. By default,
MrBayes reports substitution rates in the Dirichlet format but you can request conversion
of sampled rates to the scaled format using the report command if you prefer this
representation. One disadvantage with the scaled format is that the posterior distribution
tends to be strongly skewed such that the arithmetic mean of the sampled values is
considerably higher than the mode and the median. Therefore, consider using the median
instead of the mean when reporting a posterior distribution sampled in the scaled format.
Before using the GTR model for some of your data, we recommend that you make sure
there are at least a few possible substitutions of each type. For instance, if there is not a
single GT substitution in your data, it will be difficult to estimate the GT substitution
rate. In such cases, you should consider either the HKY model (nst=2) or the F81
model (nst=1) instead of the GTR model. The HKY model (Hasegawa, Kishino and
Yano, 1985) has different rates for transitions (rti) and transversions (rtv):
 −
π r
Q =  A tv
π A rti

π A rtv
π C rtv π G rti π T rtv 
−
π G rtv π T rti 
.
π C rtv
−
π T rtv 

π C rti π G rtv
− 
The HKY model is often parameterized in terms of the ratio between the transition and
transversion rates, κ = rti/rtv, and this is the default format used by MrBayes when
reporting samples from the posterior distribution. Internally, however, MrBayes uses the
Dirichlet parameterization in which the transition and transversion rates are expressed as
percentages of the (unscaled) rate sum. If you prefer, you can have MrBayes report the
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sampled values using the Dirichlet format instead of the ratio format by using the
command report tratio=dirichlet. The caveats described above for the GTscaled substitution rates also apply to the transition / transversion rate ratio. In the .p
files, the ratio will be referred to as kappa and the transition and transversion rate
proportions will be referred to as ti and tv. The setting of the report tratio
option will determine whether you will see a single kappa column or the two ti and tv
columns.
As with the GTR model, you can fix both the stationary state frequencies and the
transition / transversion rate ratio of the HKY model. If you fix the stationary state
frequencies to be equal, you will get the K2P model (Kimura, 1980).
Finally, MrBayes implements the F81 model (Felsenstein, 1981), which assumes that all
substitution rates are equal but stationary state frequencies are not, that is
 − πC
π
−
Q= A
π A π C

π A π C
πG πT 
πG πT 
.
− πT 

πG − 
If the stationary state frequencies are fixed to be equal using prset
statefreqpr=fixed(equal), you will get the simplest of all nucleotide
substitution models, the JC model (Jukes and Cantor, 1969).
A large number of other subsets of the GTR model are often used in Maximum
Likelihood inference. Why do we not allow more substitution model types in MrBayes?
One of the most important advantages of the Bayesian approach is that it allows you to
integrate out uncertainty concerning the model parameters you have little information
about. Thus, Bayesian inference is relatively robust to slight over-parameterization of
your model. In addition, the MCMC sampling procedure is typically efficient in dealing
with complex multi-parametric models. For these reasons, it is less important in the
Bayesian context to find the simplest possible model that can reasonably represent your
data. If you use a model-testing procedure and it suggests a four by four nucleotide model
not implemented in MrBayes, then you should obtain good results using the next more
complex model available in the program.
4.1.2. The Doublet Model
The doublet model of MrBayes is intended for stem regions of ribosomal sequences,
where nucleotides pair with each other to form doublets. The nucleotide pairing results in
strong correlation of substitutions across sites – when there is a substitution at one site it
is typically accompanied by a compensatory substitution at the paired site. If the
correlation between paired sites is not accounted for, parametric statistical methods will
overestimate the confidence we should have in the best tree(s). Incidentally, the same is
true for parsimony and the non-parametric bootstrap.
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There are various ways to model the evolution of nucleotide doublets. One method is to
focus on the common doublets, A-T and C-G in particular. MrBayes uses a more
complex model, originally formulated by Schoniger and von Haeseler (1994), where all
doublets are taken into account. The central idea in this model is that one common
doublet is converted into another through a two-step process. In the first step, one of the
nucleotides is substituted with another according to a standard four by four model of
nucleotide change. In the second step, the matching nucleotide is changed according to
the same standard four by four model. Thus, in this model there is no momentary change
from one doublet to another doublet; this always occurs through an intermediate, rare
doublet.
Assume that we are using a GTR model for the single nucleotide substitutions, that i and j
are two different doublets, that dij is the minimum number of nucleotides that must be
changed when going between i and j, and that mn is the pair of nucleotides that change
when going between i and j when dij = 1. Then the elements qij of the instantaneous rate
matrix Q of the doublet model can be expressed as follows
 0
qij = 
π j rmn
if
if
d ij > 1
d ij = 1
for the case when i differs from j; the diagonal elements (i = j) are determined, as usual,
to balance the rows in the instantaneous rate matrix to sum to zero. This gives the
instantaneous rate matrix (only 7 rows and columns out of 16 shown):

[AA]

[AC]

[AG]
Q = qij = [AT]

[CA]
 [CC]

 ...
 [TT]

[AA]
−
[AC]
π AC rAC
[AG]
π AG rAG
[AT]
π AT rAT
[CA]
π CA rAC
[CC]
0
...
...
π AA rAC
−
π AA rAG π AC rCG
π AA rAT π AC rCT
π AA rAC
0
0
π AC rAC
...
...
0
0
π AG rCG
−
π AG rGT
0
0
...
0
π AT rCT
π AT rGT
−
0
0
...
π AT rAT
0
0
0
−
π CA rAC
π CC rAC
0
0
π CC rAC
−
...
0
...
...
...
...
...
...
0
...
...
[TT] 
0 

0 

0 
π TT rAT 

0 
0 

... 
− 
Instead of using the GTR model, we could of course have used the HKY or F81 model,
resulting in obvious modifications of the rmn values. Use lset nst to change among
those options; for instance, use lset nst=6 to choose the GTR model. The default
model is the F81 model (lset nst=1).
Before the doublet model can be used, it is necessary to specify all the nucleotide pairs in
the alignment. This is done using the pairs command, most conveniently in a MrBayes
block in a data file. For instance, pairs 1:10, 2:9; would pair nucleotide 1 with 10
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and 2 with 9. See the kim.nex data file for an example of an analysis using the doublet
model.
4.1.3. Codon Models
The codon models implemented in MrBayes are based on the first formulations of such
models (Goldman and Yang, 1994; Muse and Gaut, 1994). The approach is similar to that
used in the doublet model. A codon can change to another only through steps of one
nucleotide change at a time. These steps are modeled using a standard four by four
nucleotide model. There is one additional complication though: some nucleotide changes
are synonymous while others lead to changes in amino acids and thus may be subject to
selection (a factor modifying the substitution rate). Assume that i and j are different
codons, that a(i) is the amino acid coded for by codon i, that dij is the minimum number
of nucleotide changes involved in changing between them, and that ω is the ratio of the
non-synonymous to the synonymous substitution rate. The off-diagonal elements of the
instantaneous rate matrix can now be defined as
 0
if

qij =  π j rmn if
π r ω if
 j mn
d ij > 1
d ij = 1 and a (i ) = a ( j )
d ij = 1 and a (i ) ≠ a ( j )
with the diagonal elements being defined to balance the rows of the instantaneous rate
matrix, as usual. The single-nucleotide substitution rates can be modeled using the GTR,
the F81, or the JC model, as before. Use lset nst to change among those options; for
instance, use lset nst=6 to choose the GTR model. The default model is the F81
model (lset nst=1).
Invoking the codon model is easy; just make sure that the aligned DNA or RNA
sequences start with a nucleotide in codon position 1 and that they end with a nucleotide
in codon position 3. Also, make sure that the sequences do not contain any stop codons.
To figure out whether a codon is a stop codon and whether a particular single-nucleotide
change involves an amino acid change, MrBayes uses one of several genetic codes. By
default, MrBayes uses the universal code but you can select other codes by using the
lset code command. Note that the codon models are computationally demanding.
Whereas the computations for the simple four by four models need to deal with only 16
Q-matrix and transition probability elements (4x4), the codon model computations need
to process more than 3,600 Q-matrix and transition probability elements resulting in these
runs being roughly 200 times slower. The codon models also require much more memory
than the four by four models, about 16 times as much.
The simplest codon model, described above, assumes that all amino acid sites are subject
to the same level of selection (the same ω factor). However, MrBayes also implements
models accommodating variation in selection across sites. This allows you to detect
positively selected amino-acid sites using a full hierarchical Bayes analysis (that is, an
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analysis that does not fix tree topology, branch lengths or any substitution model
parameters but instead integrates out uncertainty in all these factors).
The ω variation models work much like the model accommodating rate variation across
sites according to a gamma distribution. The likelihood of each site is calculated under
several different ω values and then the values are summed to give the site likelihood. A
difference is that the stationary frequency of each omega category is estimated, instead of
being fixed as in the case of the gamma distribution. There are two variants implemented
in MrBayes, and they are invoked using the lset omegavar option. In the Ny98
model (Nielsen and Yang, 1998), there are three classes with potentially different ω
values: 0 < ω1 < 1, ω2 = 1, and ω3 > 1. The M3 model also has three classes of ω values
but these values are less constrained in that they only have to be ordered ω1 < ω2 < ω3. If,
for instance, you would like to invoke the M3 model, use the command lset
omegavar=M3.
When you use a model with variation in selection pressure across sites, you probably
want to infer the positively selected sites. If you select report possel=yes before
you start your analysis, MrBayes will calculate the probability of each site being in a
positively selected omega class. For the M3 model, for instance, the likelihood of the site
is calculated under each of the three categories, taking the category frequencies into
account, and then the likelihoods are summed to yield the total likelihood of the site.
Finally, the proportion of this sum originating from categories that are positively selected
(those that have an ω value larger than 1); this is the posterior probability of the site being
positively selected.
Once the probabilities of each site being positively selected are printed to file, they can be
summarized using the standard sump command. When interpreting the output from the
Ny98 model, it is helpful to know that pi(-), pi(N) and pi(+) are the frequencies of
the negatively selected, neutral and positively selected categories, respectively, and
omega(-), omega(N) and omega(+) are the corresponding ω values. The M3 model
parameters are instead labeled pi(1), pi(2) and pi(3) for the category frequencies
and omega(1), omega(2) and omega(3) for the ω values. The probability of a
codon being positively selected is labeled by the site numbers in the original alignment.
Thus pr+(16,17,18) is the probability of the codon corresponding to the original
nucleotide alignment sites 16, 17, and 18 being in a positively selected omega category.
4.2. Amino-acid Models
MrBayes implements a large number of amino-acid models. They fall in two distinct
categories: the fixed-rate models and the variable-rate models. The former have both the
stationary state frequencies and the substitution rates fixed, whereas one or both of these
are estimated in the latter.
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4.2.1 Fixed Rate Models
The Poisson model (Bishop and Friday, 1987) is the simplest of the fixed rate models. It
assumes equal stationary state frequencies and equal substitution rates; thus, it is
analogous to the JC model for nucleotide characters. The rest of the fixed-rate models
have unequal but fixed stationary state frequencies and substitution rates reflecting
estimates of protein evolution based on some large training set of proteins. These models
include the Dayhoff model (Dayhoff, Schwartz and Orcutt, 1978), the Mtrev model
(Adachi and Hasegawa, 1996), the Mtmam model (Cao et al., 1998; Yang, Nielsen, and
Hasegawa, 1998), the WAG model (Wheland and Goldman, 2001), the Rtrev model
(Dimmic et al., 2002), the Cprev model (Adachi et al., 2000), the Vt model (Muller and
Vingron, 2000) and the Blosum62 model (Henikoff and Henikoff, 1992). Each model is
appropriate for a particular type of proteins. For instance, if you are analyzing mammal
mitochondrial proteins, you might want to use the Mtmam model. Invoke that model by
typing prset aamodelpr=fixed(mtmam).
4.2.2. Estimating the Fixed Rate Model
MrBayes allows a convenient way of estimating the fixed-rate model for your amino acid
data instead of specifying it prior to the analysis. If you choose this option, MrBayes will
allow the MCMC sampler to explore all of the fixed-rate models listed above, including
the Poisson model, by regularly proposing new models. When the MCMC procedure has
converged, each model will contribute to your results in proportion to its posterior
probability. For instance, if you are analyzing mammal mitochondrial proteins, it is likely
that the Mtmam model will contribute most to the posterior distribution but it is possible
that some other models, for instance the Mtrev model, will contribute significantly too. A
nice feature of the MCMC model estimation is that the extra computational cost is
negligible.
To allow so-called model jumping between fixed-rate amino acid models, simply set the
prior for the amino acid model to mixed, prset aamodelpr=mixed, prior to
analysis. During the run, MrBayes will print the index of the current model to the .p
file(s) in the aamodel column. The indices of the models are as follows: 0 = Poisson, 1
= Jones, 2 = Dayhoff, 3 = Mtrev, 4 = Mtmam, 5 = Wag, 6 = Rtrev, 7 = Cprev, 8 = Vt, 9 =
Blosum. When you use the sump command, you will get a table with the names of the
amino acid models and their posterior probabilities.
4.2.3. Variable Rate Models
There are two variable-rate models implemented in MrBayes for amino acid data. The
Equalin model is a “glorified” F81 model in that it allows the stationary state frequencies
of all amino acids to be different but assumes the same substitution rate. Thus, the
instantaneous rate matrix for this model is
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
[A]

[R]

[N]
Q = qij = [D]

[C]
[Q]

 ...
[V]

[A]
−
πA
πA
πA
πA
πA
...
πA
[R]
πR
−
πR
πR
πR
πR
...
πR
[N]
πN
πN
−
πN
πN
πN
...
πN
[D]
πD
πD
πD
−
πD
πD
...
πD
38
[C]
πC
πC
πC
πC
−
πC
...
πC
[Q]
πQ
πQ
πQ
πQ
πQ
−
...
πQ
...
...
...
...
...
...
...
...
...
[V]
πV 

πV 

πV 
πV 

πV 
πV 

... 
− 
and it has 19 free parameters (20 stationary state frequencies minus one because the
frequencies sum to 1) that will be estimated from data.
The other variable-rate model is the “glorified” GTR model, which allows all stationary
state frequencies and substitution rates to vary. Thus, the instantaneous rate matrix for
this model is

[A]

[R]

[N]
Q = qij = [D]

[C]
[Q]

 ...
[V]

[A]
−
π A rAR
π A rAN
π A rAD
π A rAC
π A rAQ
...
π A rAV
[R]
[N]
[D]
[C]
π R rAR π N rAN π D rAD π C rAC
−
π N rRN π D rRD π C rRC
π R rRN
−
π D rND π C rNC
π R rRD π N rND
−
π C rDC
π R rRC π N rNC π D rDC
−
π R rRQ π N rNQ π D rDQ π C rCQ
...
...
...
...
π R rRV π N rNV π D rDV π C rCV
[Q]
...
π Q rAQ
π Q rRQ
π Q rNQ
π Q rDQ
π Q rCQ
−
...
π Q rQV
...
...
...
...
...
...
...
...
[V] 
π V rAV 

π V rRV 

π V rNV 
π V rDV 

π V rCV 
π V rQV 

... 
− 
and the model has 19 free stationary state frequency parameters and 189 free substitution
rate parameters. The Bayesian MCMC approach is good at handling uncertainty in multiparameter models, so the GTR model may be used successfully with moderate-size data
sets, but the model is so parameter-rich that you need a fairly sizable data set to be able to
estimate all parameters with reasonable precision.
The GTR model can be used to express a user-derived fixed rate model other than those
already implemented in MrBayes. Simply use the prset command to fix the stationary
state frequencies and substitution rates of the GTR model to the desired values. You need
to set two options, prset aarevmatpr=fixed(<190 comma-separated
values>) and prset statefreqpr=fixed(<20 comma-separated
values>). Once the values are fixed prior to analysis, the MCMC procedure will not
change them and they will remain the same throughout the analysis.
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4.3. Restriction Site (Binary) Model
MrBayes implements a simple F81-like model for restriction sites and other binary data.
The instantaneous rate matrix for this model is simply
[0] [1]

Q = qij = [0] − π 1 


 [1] π 0 − 
Any asymmetry in the rate of 0 to 1 and 1 to 0 transitions is expressed in terms of the
stationary state frequencies. Thus, if the stationary frequencies are π0 = 0.25 and π1 =
0.75, then the rate of 0 to 1 transitions is 3 times as high as the rate of transitions in the
other direction (π1 / π0 = 3).
A problem with some binary data sets, notably restriction sites, is that there is an
ascertainment (coding) bias such that certain characters will always be missing from the
observed data. It is impossible, for instance, to detect restriction sites that are absent in all
of the studied taxa. MrBayes corrects for this bias by calculating likelihoods conditional
on the unobservable characters being absent (Felsenstein, 1992). The ascertainment
(coding) bias is selected using lset coding. There are five options: (1) there is no
bias, all types of characters could, in principle, be observed (lset coding=all); (2)
characters that are absent (state 0) in all taxa cannot be observed (lset coding=
noabsencesites); (3) characters that are present (state 1) in all taxa cannot be
observed (lset coding=nopresencesites); (4) characters that are constant
(either state 0 or 1) in all taxa cannot be observed (lset coding=variable); and
(5) only characters that are parsimony-informative have been scored (lset
coding=informative). For restriction sites it is typically true that all-absence sites
cannot be observed, so the correct coding bias option is noabsencesites.
The binary model is useful for a number of character types other than restriction sites. For
instance, the model can be used for gap characters. The presence and absence of gaps
must be coded consistently for all characters; let us assume here that absence of a gap is
coded as 0 and presence as 1. Since the detection of gaps is typically contingent on
observing some sequence length variation, neither all-absence nor all-presence characters
can be observed. Thus, the correct ascertainment bias for gap characters is variable.
The parameters π0 and π1 would represent the rate at which insertions and deletions
occur, respectively (assuming that state 0 denotes absence of a gap).
The binary model can also be used for ecological, morphological, or other binary
characters of arbitrary origin. However, if the binary model is applied to more than one
character, then there is an implicit assumption that the state labels are not arbitrary for
those characters. That is, the 0 state in one character must somehow be comparable to the
0 state in the others. For instance, 0 could mean absence (or presence) of a particular type
of feature, such as a wing vein, a restriction site, or a gap in a DNA sequence. It is not
appropriate to apply the default binary model to a set of characters where the state labels
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are arbitrary, as is true of most morphological characters. Thus, we can possibly estimate
the rate of loss versus gain of wing veins over a set of consistently coded wing venation
characters, but we cannot compare the rate of loss of antennal articles to the rate at which
a yellow patch evolves into a green patch. If state labels are truly arbitrary, then the
stationary state frequencies of the binary model must be fixed to be equal, such that the
estimation of model parameters becomes independent of the labeling of character states.
An alternative is to consider the standard model, which provides more sophisticated ways
of dealing with arbitrary state labels.
When is the correction for ascertainment bias important? This is strongly dependent on
the size of the tree (the sum of the branch lengths on the tree). The larger the tree, the less
important the correction for ascertainment bias becomes. In our experience, when there
are more than 20-30 taxa, even the most severe bias (only informative characters
observed) is associated with an insignificant correction of the likelihood values.
4.4. Standard Discrete (Morphology) Model
The model used by MrBayes for “standard” discrete data is based on the ideas originally
presented by Lewis (2001). Essentially, the model is analogous to a JC model except that
it has a variable number of states, from 2 to 10. For instance, a three-state standard
character would be evolving according to the instantaneous rate matrix
[0] [1] [2]

[0] − 1 1 
.
Q=
 [1] 1 − 1 


[2] 1 1 − 
Because all rates are the same, we can maintain the essential property of standard
characters, namely that state labels are arbitrary. Thus, the standard model assures that
you will get the same results regardless of the way in which you label the states.
In morphology-based parsimony analyses, one sometimes distinguishes between ordered
and unordered characters. In ordered characters, evolution between some states is
assumed to go through intermediate states. MrBayes implements a stochastic model for
such characters. For a three-state character assumed to be ordered (by convention in the
sequence 0-1-2), the instantaneous rate matrix is
[0] [1] [2]

[0] − 1 0 
.
Q=
 [1] 1 − 1 


[2] 0 1 − 
Note that the instantaneous rate of going between the two end states is 0, that is, a
transition from 0 to 2 or from 2 to 0 has to go through state 1. By default, MrBayes treats
all standard characters as unordered. To change this, use the ctype command. For
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instance, if you want to treat characters 3 and 4 as ordered you need to include the
statement ctype ordered: 3 4; in your MrBayes block (or enter it using the
command line, if you prefer).
The number of states of each standard character is determined by MrBayes simply by
looking at the state codes in your matrix. Thus, a three-state model will be used for a
three-state character and a six-state model for a six-state character. MrBayes does not
check that all state codess are used, it simply finds the largest state code in the matrix for
each character. Thus, make sure that you use the state codes 0, 1, and 2 for a three-state
character and state codes 0, 1, 2, 3, 4, and 5 for a six-state character.
Because state labels are arbitrary in the standard model, we cannot estimate unequal
stationary state frequencies or substitution rates (recall that the stationary state
frequencies are an important factor in determining the latter). However, it is still possible
to allow the state frequencies (rates) to vary over sites according to some appropriate
distribution. MrBayes uses a symmetric Dirichlet distribution for this purpose. For binary
data, the analogy of the Dirichlet distribution is called the beta distribution; MrBayes uses
Dirichlet and beta interchangeably for the distribution depending on context. The
approach is similar to the one used to allow rate variation across sites according to a
gamma distribution: we calculate the likelihood of a site assuming different discrete
categories of asymmetry and then we sum the values to obtain the site likelihood.
The symmetric Dirichlet distribution has one parameter that determines its shape, just
like the alpha parameter determines the shape of the gamma distribution. The larger the
parameter of the symmetric Dirichlet, the less transition rate (stationary frequency)
asymmetry there is across sites. By default, the parameter is fixed to infinity (prset
symdirihyperpr=fixed(infinity)); this corresponds to the standard
assumption of no transition rate asymmetry across sites: the rate of going from 0 to 1 is
equal to the rate of going from 1 to 0 for all characters. The prior is called a hyperprior
because it concerns a distribution that controls the distributions of other model
parameters (stationary state frequencies in this case). If you want to allow transition rate
(stationary frequency) asymmetry in standard data, then simply select another
(hyper)prior. For instance, you can fix the parameter to 1.0, which would result in a
uniform prior on the proportions of the state frequencies.
In practice, MrBayes uses a discrete approximation of the Dirichlet distribution for binary
characters; five categories are used by default (change this with lset nbetacat). For
instance, assume that we fix the hyperprior to 1.0 and then evaluate the likelihood of one
binary character using five discrete beta categories. MrBayes would then calculate the
likelihood of the character assuming that the stationary state frequencies of the two states
were 0.1:0.9, 0.3:0.7, 0.5:0.5, 0.7:0.3 and 0.1:0.9. The five category likelihoods would
then be multiplied by 0.20 (there is a probability of 0.20 of being in each of the
categories) and then summed up to give the total likelihood of the character. For multistate characters, MrBayes does not use the discrete approximation; instead, it uses the
MCMC procedure to explore different stationary state frequency proportions.
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4.5. Parsimony Model
MrBayes implements an incredibly parameter-rich model, first described by Tuffley and
Steel (1997). It orders trees in terms of their maximum likelihood in the same way as the
parsimony method would order them in terms of their parsimony score; hence we call it
the parsimony model. The model is also referred to as the No-Common-Mechanism
model because it treats each branch length for each character as a separate, completely
independent parameter. In principle, a Bayesian MCMC analysis using the parsimony
model should integrate out the branch lengths but MrBayes 3 uses a simpler approach, in
which the branch lengths are fixed to their maximum likelihood values (infinity if there is
a change on the branch and zero otherwise). This type of approach, where some
parameters are fixed prior to the Bayesian analysis according to some non-Bayesian
estimate, is typically referred to as an empirical Bayes method. Future versions of
MrBayes may implement the true (hierarchical) Bayesian approach to the parsimony
model but we expect the results to be very similar under both approaches.
The parsimony model is much less parsimonious with respect to parameters than any
other model implemented in MrBayes. Consider, for instance, an analysis of 1,000
characters and 100 taxa. The parsimony model would have about 200,000 free parameters
(200 branches times 1,000 characters). A more typical GTR + Γ + I model would have
only little more than 200 parameters, about 1,000 times fewer parameters. In this sense,
the standard stochastic models are much more parsimonious than the parsimony model.
Several problems are associated with the excessive number of parameters. Statistical
inconcistency is perhaps the best known of these but, more fundamentally, a model like
the parsimony model does not offer much in terms of generalities that can be inferred
about the evolutionary process.
The Goldman (1993) model is another example of a parameter-rich stochastic model that
orders trees in the same way as the parsimony method. In this model, the branch lengths
are the same but the ancestral states are estimated for all characters and all nodes in the
tree. For an analysis of 100 taxa and 1,000 characters, this results in approximately
100,000 free parameters. The Goldman model is actually very similar to the NoCommon-Mechanism model; it makes little difference if the character-specific and treesection-specific parameters are introduced at the nodes or at the branches. The Goldman
model is not implemented in MrBayes.
We would like to emphasize that we do not recommend the use of the parsimony model.
We have included it in MrBayes only to allow users to explore its properties and contrast
it with the other models implemented in the program. The parsimony model is not the
default model used for standard (morphological) data in MrBayes. The default standard
data model is described above and is similar to the models used for nucleotide and amino
acid data. By default, MrBayes does not use the parsimony model at all; you have to
invoke it using lset parsmodel=yes.
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4.6. Rate Variation Across Sites
By default, MrBayes assumes that all characters evolve at the same rate (measured in
expected changes per site over the tree). There are four ways in which you can allow rate
variation across sites. The simplest method is to assume that rates vary over sites
according to a gamma distribution (Yang, 1993). The gamma model can be combined
with spatial autocorrelation between the rates at adjacent sites, the autocorrelated gamma
model. A completely different approach to rate variation across sites is to allow a
proportion of sites to be invariable. This model can be combined with the gamma model
but not with the autocorrelated gamma model. Finally, it is possible to divide characters
into groups evolving at different rates, the partitioned or site specific rate model.
4.6.1. Gamma-distributed Rate Model
The commonly used model of gamma-shaped rate variation across sites is invoked using
lset rates=gamma. The shape of the gamma distribution is determined by the socalled α (alpha) parameter. When this parameter is small (below 1), the distribution takes
on an L-shaped form with a few sites evolving rapidly while most sites are conserved.
Conversely, when α is above 1 the distribution becomes similar to a normal distribution
with less and less variation in rates across sites as α becomes larger.
In practice, the gamma distribution is approximated using a small number of discrete rate
categories (Yang, 1994). By default, four rate categories are used; you can change this
setting by using lset ngammacat. For instance, if you want to use eight discrete rate
categories the appropriate command is lset ngammacat=8. The computational
complexity is proportional to the number of categories used. An analysis with four
discrete gamma categories is four times as slow as an analysis with no rate variation
across sites and twice as fast as one with eight categories.
The shape parameter (α) of the gamma distribution is similar to the shape parameter of
the Dirichlet distribution of stationary state frequencies used for standard data (see above)
in that it controls the distribution of another model parameter (the site rates). Therefore,
the prior probability distribution used for the shape parameter can be referred to as a
hyperprior. The default prior used in MrBayes is a uniform distribution on the interval
(0.05,50). The sampled values of the shape parameter are found under the column
heading alpha in the .p file(s).
4.6.2. Autocorrelated Gamma Model
In this model, rates vary across sites according to an autocorrelated gamma model where
the rate at each site depends to some extent on the rates at adjacent sites (Yang, 1995).
The spatial autocorrelation is measured by the ρ (rho) parameter, which ranges from -1
(negative autocorrelation, that is, adjacent sites tend to have wildly different rates) to 1
(adjacent sites have very similar rates). The default prior probability for rho is a uniform
distribution covering the entire interval (-1,1).
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In the worst case, a small symmetric tree, the extra computational complexity incurred by
invoking the auto correlated gamma model instead of the gamma model is comparable to
a doubling of the number of taxa in the analysis. In more typical cases, moderate to large
data sets, the additional computational cost is negligible and equivalent to adding a
single taxon. As with the gamma model, the autocorrelated gamma distribution is
approximated with a number of discrete rate categories determined by lset
ngammacat.
As described by Yang (1995), protein-coding sequences tend to have a three-position
offset in their autocorrelation. That is, the first codon position sites tend to have rates that
are correlated with the adjacent first-position sites, second-position site rates are
correlated with adjacent second-position rates, etc. You can take this effect into account
by partitioning your sites into the three codon positions and then applying a separate
autocorrelated gamma model to each of the categories.
You might want to invoke an autocorrelated gamma model with the same correlation
coefficient for a dataset consisting of several concatenated genes. If so, it is necessary to
inform MrBayes about the break points between different genes. Otherwise, the rates at
the first site of each gene except the first one will be erroneously compared to the rates at
the last site in the preceding gene. The command databreaks is provided for this
purpose. For instance, if there are only two genes in your data set, the first with 960 sites,
you would specify the break between them with the statement databreaks 960. Note
that you specify the break by giving the last sequence site before the break. The
databreaks command is only needed when you invoke a single autocorrelated gamma
model for a multigene dataset. The databreaks command cannot be used to partition a
data set.
4.6.3. Proportion of Invariable Sites
A completely different approach to rate variation is to allow a proportion of sites to be
invariable. This model is invoked using lset rates=propinv. The proportion of
invariable sites is referred to as pinvar; it can vary from 0 (no invariable sites) to 1 (all
sites are invariable). The default prior is a uniform distribution on the interval (0,1);
change it using prset pinvarpr.
The proportion of invariable sites model can be combined with the gamma model using
lset rates=invgamma. Although this model is slightly better than the simple
gamma model for many data sets, it sometimes results in a bimodal or ridge-like posterior
probability distribution. In particular, it is not uncommon to see two peaks in the
posterior, one with a low proportion of invariable sites and significant rate variation in
the gamma distribution (low alpha value) and the other with a high proportion of
invariable sites and moderate amounts of rate variation in the gamma distribution
(moderately high alpha value). If you have a posterior of this kind, you should not be
surprised if Metropolis-coupling results in rapid (instantaneous) shifts from one mode to
the other during the stationary phase of the analysis. The reason for this is that different
chains are likely to explore different peaks in the posterior, and successful swapping
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involving the cold chain is likely to result in mode-jumping. Also, you should consider
presenting the entire distribution of the sampled alpha and pinvar parameters since
simple point estimates of each parameter would be misleading.
4.6.4. Partitioned (Site Specific) Rate Model
For protein-coding nucleotide sequences, a site-specific rate model is often used,
allowing each codon position (first, second and third codon position sites) to have its own
rate. This results in a model with three rates, two of which are free to vary (since the
average rate is 1.0 by definition). More generally, we might have different character
divisions (separate genes, morphology, etc) which potentially evolve at very different
rates.
In MrBayes 3, we provide a general mechanism for setting up these models based on
partitioning the data set and then unlinking parameters across the partitions. Assume for
instance that we want to set up a site specific rate model for a data set with one sequence.
We first set up the codon site partitioning scheme using the following lines in a MrBayes
block:
charset pos1 = 1-.\3;
charset pos2 = 2-.\3;
charset pos3 = 3-.\3;
partition by_codon = 3: pos1, pos2, pos3;
set partition = by_codon;
The character sets are first defined using the dot sign (.) to mark the last character in the
data set and the \3 sequence to include every third character in the specified range. Then
a partitioning scheme called by_codon is defined using the previously named character
sets. Finally, the partitioning scheme called by_codon is invoked using the set
command.
When we process these commands in MrBayes using the execute command, the
characters are divided into three sets corresponding to the codon positions. By default,
however, all model parameters including the rate will be shared across partitions. To
allow the rates to differ across partitions, we need to change the prior for rates using
prset. Specifically, prset ratepr=variable invokes partition-specific rates.
The partition-specific rate parameter is referred to as ratemult, and the individual rates
are labeled m{1}, m{2}, etc for the rate (multiplier) of character division 1, 2, etc. See
below for more information on how to set up partitioned models.
4.6.5. Inferring Site Rates
When you are allowing rate variation across sites, you may be interested in inferring the
rates at each individual site. By default, the site rates are not sampled during a MCMC
run. You need to request the sampling of these values using report
siterates=yes. The rates will be referred to as r(<site number>). For
instance, r(45) is the inferred rate at site (character) 45 of your data set.
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4.7. Rate Variation Across the Tree: The Covarion Model
For both nucleotide sequence and amino-acid data, MrBayes allows rates to change
across the tree under a covarion-like model (Tuffley and Steel, 1998; Huelsenbeck, 2002;
see also Galtier, 2001). Specifically, the covarion-like model assumes that a site is either
“on” or “off”. When it is on, it evolves under a standard four-by-four nucleotide or 20 by
20 amino acid model but when it is off, it does not change at all. The switching between
on and off states is controlled by two rate parameters, s01 (from off to on) and s10 (from
on to off). The instantaneous rate matrix of the nucleotide variant (also referred to as the
covariotide model), assuming a GTR model for the on state, is
[A off ] [C off ] [G off ] [Toff ] [A on ]
[C on ]
[G on ]
[Ton ] 

[A ]
−
0
0
0
s01
0
0
0 
 off

0
−
0
0
0
s01
0
0 
[C off ]


0
0
−
0
0
0
s01
0 
[G off ]
Q =  [Toff ]
0
0
0
−
0
0
0
s01  ,


s10
0
0
0
−
π C rAC k π G rAG k π T rAT k 
 [A on ]
 [C on ]
0
s10
0
0
π A rAC k
−
π G rCG k π T rCT k 


0
0
s10
0
π A rAG k π C rCG k
−
π T rGT k 
 [G on ]
 [T ]
0
0
0
s10 π A rAT k π C rCT k π G rGT k
− 
 on
where k is a scaling constant determined by the proportion of time the sites spend in the
on state. The matrix can be simplified into
R
Q= 1
 R3
R2 
,
kR4 
where each R element is a four by four matrix: R1 contains the rates in the off state (all
rates are 0), R2 and R3 describe the switching process (the diagonal elements are either s01
or s10), and R4 is the chosen model for the evolution in the on state.
The covarion-like model can be described as a general case of the proportion of
invariable sites model (Huelsenbeck, 2002). As the switching rates go to zero, the
proportion of these rates represented by the switch to the off state (s10) becomes identical
to the proportion of invariable sites. When the switching rates are zero, there is no
exchange between the on and off states and the characters in the off state remain off
throughout the tree; in other words, they are invariable sites.
Note that the covarion-like model implemented in MrBayes differs from the original
covarion model in that sites switch completely independently of each other between the
on and off states. To invoke the covarion-like model, simply use lset
covarion=yes and then choose the desired nucleotide or amino acid model using the
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other lset and the prset options. The covarion-like model can be combined with the
gamma model of rate variation across sites.
4.8. Topology and Branch Length Models
The topology and branch length models in MrBayes are set using the prset
topologypr options, which deal with the tree topology prior, and the prset
brlenspr options, which deal with the branch lengths.
4.8.1. Unconstrained and Constrained Topology
There are two choices for the prior probability distribution on topology: uniform or
constrained. By default, topologies are not constrained in the prior (prset
topologypr=uniform), resulting in equal prior probability being associated with all
possible labeled trees (unless a different topology prior is induced by the branch length
model, see below). There are two instances in which you might want to constrain the
topology: (1) when you want to contrast a hypothesis of monophyly for a group with the
more general hypothesis with no topological constraints; and (2) when you want to infer
ancestral states for a particular node in the tree. In both cases, you specify the
constraint(s) first by listing the taxa that should form a monophyletic group. For instance,
if you wanted to constrain taxa 4, 5 and 6 to be monophyletic, you would use
constraint my_constraint -1 = 4 5 6;
This defines a constraint called “my_constraint” forcing taxa 4, 5 and 6 to form a
monophyletic group in all trees that are sampled from the chain. In future versions of
MrBayes, the value following the name of the constraint (-1 here), will give the relative
probability of trees having the constrained partition. A negative number will force the
constraint to always be present in the sampled trees; a positive number will specify how
many times more likely the trees with the constraint are compared to the trees not having
it. In version 3.1, however, MrBayes ignores this value and always treats the constraint as
absolute.
When you define constraints, make sure that you have the outgroup selected correctly. By
default, MrBayes uses the first taxon in the data matrix as the outgroup. You can change
this by using the outgroup command. For instance, if you want taxon number 7 called
“My_taxon” to be the outgroup, either use outgroup 7 or outgroup My_taxon.
MrBayes 3.1 only allows a single taxon as the outgroup.
Before the constraints take effect, you have to invoke them by using prset
topologypr=constraints(<comma-separated list of
constraints>). For instance, to enforce the constraint my_constraint defined above,
use prset topologypr=constraints(my_constraint).
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4.8.2. Non-clock (Standard) Trees
If you do not want to enforce a molecular clock, you choose an unconstrained branch
length prior. Actually, you do not have to do anything because unconstrained branch
lengths are the default. You can associate unconstrained branch lengths with either a
uniform prior from 0 to some arbitrary value or an exponential prior. The default is an
exponential distribution with parameter 10 (Exponential(10)); it has an expectation
of 0.1 (= 1/10) but (in principle) it allows branch lengths to vary from 0 to infinity. The
exponential distribution apparently puts a lot more probability on short branches than on
long branches. However, because transition (substitution) probabilities change rapidly at
small branch lengths but only very slowly at long branch lengths, the exponential prior is
actually closer to an uninformative prior than the uniform distribution. We advise against
using a uniform prior on branch lengths because of the large prior probability it puts on
long branches and their close-to-random substitution probabilities.
To change the prior on unconstrained branch lengths you use prset brlenspr. For
instance, assume you wanted to use an exponential prior with parameter 1 instead of the
default prior. This prior is set by typing prset brlenspr=unconstrained:
exponential(1).
4.8.3. Strict Clock Trees
MrBayes implements three strict clock models: the simple (uniform) model, the birthdeath model, and the coalescence model. The birth-death model and coalescence models
both have additional parameters describing the tree-generating process, whereas the
simple model does not.
In the birth-death model (see Yang and Rannala, 1997, for a Bayesian implementation),
trees are generated according to a birth-death model with a speciation and an extinction
rate. The model, as implemented in MrBayes, can also be associated with a sampling
probability of terminal lineages. The priors for these three parameters are set using the
speciationpr, extinctionpr, and sampleprob parameters of the prset
command.
In the coalescence model, the tree generating process is looked at from the opposite
perspective, backward in time. Instead of lineages branching, this model sees them as
coalescing into fewer and fewer ancestral lineages. This process occurs at a rate
determined by the θ (theta) parameter; it is also affected by the ploidy of the data
(haploid or diploid). The prior for the theta parameter is set using prset thetapr.
The ploidy level is set using lset ploidy.
The simple clock model is invoked by prset brlenspr=clock:uniform. There is
only one additional parameter in the simple clock model, namely the total tree height.
The prior for this parameter is set by prset treeheightpr. The default prior is an
exponential distribution with parameter 1.0 (Exponential(1.0)).
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4.8.4. Relaxed Clock Trees
Relaxed clock models and functions for dating are not implemented in MrBayes 3.1.
According to current plans, they will be available in version 3.2.
4.9. Partitioned Models
MrBayes provides great flexibility in setting up partitioned models. By default, the
characters are divided into partitions based on the data type. If there is only one data type
in the matrix, then all characters will be in a single partition. The default partitioning
scheme is called default. For information on how to set up a file with mixed data
types, see the example file cynmix.nex and the tutorial in section 3 of this manual.
You can easily set up partitioning schemes that divide the characters up further than the
default partition does using the partition command. The most convenient way of
partitioning data is to define character sets first using the charset command. For
instance, assume that you have concatenated nucleotide sequences from three genes in
your data set of length 1962, 1050, and 2082 sites, respectively. Then you create
character sets for those three genes using
charset gene1 = 1-1962;
charset gene2 = 1963-3012;
charset gene3 = 3013-.;
in a MrBayes block. Note the use of the dot (.) as a synonym of the last site. You can also
use the “backslash n” sequence to include every nth character in the preceding range of
characters (see the description of the site specific rate model above). Once the character
sets are defined, the partitioning scheme based on the genes is defined with the
partition command and selected using the set command:
partition by_gene = 3: gene1, gene2, gene3;
set partition=by_gene;
Here, by_gene is the name we chose for the partitioning scheme. The name is followed
by an equal sign, the number of partitions, and then a comma-separated list of characters
to include in each partition. Note that MrBayes requires the partitioning scheme to
include all characters. Say, for instance, that you wanted to run an analysis with only
gene 1 and gene 2. Then define a two-partition scheme and exclude the characters
represented by gene 3:
partition gene1&2 = 2: gene1, gene2 gene3;
exclude gene3;
set partition=gene1&2;
If the only purpose of the partition gene1&2 is to allow exclusion of gene 3, then gene 3
can of course be included in either of the two partitions before being excluded.
Once the partitions have been correctly set up, MrBayes allows you to set models for
individual partitions using the lset applyto and prset applyto mechanism. For
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instance, assume that we have two partitions, a standard data partition (partition 1) and a
nucleotide partition (partition 2), and want to apply a GTR model to the nucleotide data,
gamma-shaped rate variation to both partitions, and allow the partition rates to be
different. Then we would use the commands
lset applyto=(2) nst=6;
lset applyto=(all) rates=gamma;
prset applyto=(all) ratepr=variable;
By default, all model parameters that are identical and have the same prior probability
distribution associated with them, are linked across partitions (they are assumed to be one
and the same parameter). To unlink parameters, use the unlink command. For instance,
assume that we want to unlink the shape parameter across the partitions discussed above
(after all, why should the standard data and the molecular data have the same distribution
of rates across sites?). This would be achieved using
unlink shape=(all);
If you unlink parameters by mistake, they can be linked again using the link command.
All of the commands mentioned above and given as they would appear in a MrBayes
block in a Nexus file can of course be entered from the command line as well (without
the trailing semicolon). However, it is often more convenient to have them in either your
data file or in a separate Nexus file that you process after you have read in your data.
MrBayes will keep the data set in memory until you read in a new data block, so you can
have your different MrBayes blocks pertaining to the same data file distributed over as
many separate Nexus files as you like.
We recommend that, before you run your analysis, you check the current model settings
using the showmodel command. This command will list all the active parameters and
how they are linked across partitions, as well as the priors associated with each
parameter.
Finally, we want to give you a warning. Even though MrBayes allows you to easily set up
extremely complex and parameter-rich models, and the Bayesian MCMC approach is
good at handling such models, think carefully about the parameters you introduce in your
model. There should be at least some reasonable chance of estimating the parameters
based on your data. For instance, a common mistake is to use a separate GTR model for a
partition with so few substitutions that there is not a single observation for several rate
categories. Making sure there are at least some observations allowing you to estimate
each parameter is good practice. Over-parameterized models often result in problems
with convergence in addition to the excessive variance seen in the parameter estimates.
4.10. Ancestral State Reconstruction
MrBayes allows you to infer ancestral states at ancestral nodes using the full hierarchical
Bayesian approach (integrating out uncertainty concerning topology and other model
parameters). The basic approach is described by Huelsenbeck and Bollback (2001) as
well as in a recent review (Ronquist, 2004). You first need to constrain the node you want
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to infer ancestral states for using a constraint definition and the
topologypr=constraints(...) command as described above for constrained
topology models. Then ancestral state reconstruction is requested using report
ancstates=yes.
The probability of each state will be printed to the .p file(s) under the heading
p(<state_code>){character_number}. For instance, the probability of an A at
site 215 in a nucleotide data set would be found under the heading p(A){215}. If you
constrain several nodes in your data set, the node number will be given as well. If you
had constrained two nodes, the probabilities of the above character would be
distinguished as p(A){[email protected]} and p(A){[email protected]}. However, if you are interested in
inferring ancestral states at two or more different nodes, we recommend running separate
analyses, each constraining a single node. The reason is that when you focus on one node,
you probably want to integrate over uncertainty in the rest of the tree, including the
potential uncertainty concerning the presence of the other node(s).
Often, there is interest in mapping only one or a few characters onto trees inferred using
largely other types of data. For instance, a behavioral or ecological trait may be mapped
onto trees based on molecular data. To do this type of analysis in MrBayes, you would set
up a mixed data set including both the character to be mapped and the data used to infer
the phylogeny, with the character to be mapped in a separate data partition. How to do
this is explained in the tutorial given in section 3 of this manual as well as in the
description of partitioned models above. Typically, you also want to assume that the
evolutionary rate for the mapped character is proportional to that of the other data (rather
than identical). This is achieved by setting up a partitioned rate model using prset
ratepr=variable. Then you need to set up a constraint for the node of interest, as
described above. Finally, you request that ancestral states are inferred for the partition
with the mapped character (there is no need to wade through ancestral state probabilities
for the other partition(s)). For instance, if the character to be mapped is in partition 2,
request ancestral state sampling using prset applyto=(2) ancstates=yes.
Now only the ancestral states for the character of interest will be printed to the .p file(s).
The sampled values can be summarized as usual with the sump command.
5. Frequently Asked Questions
How do I cite the program?
If you want to cite the program, we suggest you use the two papers published in
Bioinformatics (Huelsenbeck and Ronquist, 2003; Ronquist and Huelsenbeck, 2005). If
you are using the MPI version of the program, you may also want to cite Altekar et al.
(2004).
How do I run MrBayes in batch mode?
When you become more familiar with MrBayes, you will undoubtedly want to run it in
batch mode instead of typing all commands at the prompt. This is done by adding a
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MRBAYES block to a Nexus file, either the same file containing the DATA block or a
separate Nexus file. The MRBAYES block simply contains the commands as you would
have given them from the command line, with the difference that each command line is
ended with a semi-colon. For instance, a MRBAYES block that performs three single-run
analyses of the data set primates.nex under the GTR + _ model and stores each result in a
separate file is given below:
begin mrbayes;
set autoclose=yes nowarn=yes;
execute primates.nex;
lset nst=6 rates=gamma;
mcmc nruns=1 ngen=10000 samplefreq=10 file=primates.nex1;
mcmc file=primates.nex2;
mcmc file=primates.nex3;
end;
Since this file contains the “execute” command, it must be in a file separate from the
primates.nex file. You start the analysis simply by typing execute
<filename>, where filename is the name of the file containing the MRBAYES block.
The set command is needed to change the behavior of MrBayes such that it is
appropriate for batch mode. When autoclose = yes, MrBayes will finish the
MCMC analysis without asking you whether you want to add more generations. When
nowarn = yes, MrBayes will overwrite existing files without warning you, so make
sure that your batch file does not inadvertently cause the deletion of previous result files
that should be saved for future reference.
The UNIX version of MrBayes can execute batch files in the background from the
command prompt. Just type mb <file> > log.txt & at the UNIX prompt, where
<file> is the name of your Nexus batch file, to have MrBayes run in the background,
logging its output to the file log.txt. If you want MrBayes to process more than one
file, just list the files one after the other with space between them, before the output
redirection sign (>). When MrBayes is run in this way, it will quit automatically when it
has processed all files; it will also terminate with an error signal if it encounters an error.
Alternatively, the UNIX version of MrBayes can also be run in batch mode using input
redirection. For that you need a text file containing the commands exactly as you would
have typed them from the command line. For instance, assume that your data set is in
primates.nex and that you want to perform the same analyses specified above. Then
type mb < batch.txt > log.txt & with the batch.txt file containing this
text:
set autoclose=yes nowarn=yes
execute primates.nex
lset nst=6 rates=gamma
mcmc ngen=10000 savebrlens=yes file=primates.nex1
mcmc file=primates.nex2
mcmc file=primates.nex3
quit
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The quit command forces MrBayes to terminate. With previous versions of MrBayes
we have had problems with infinite loops when the quit command is not included at the
end of the file. This problem should have been solved in version 3.1.
How are gaps and missing characters treated?
MrBayes uses the same method as most maximum likelihood programs: it treats gaps and
missing characters as missing data. Thus, gaps and missing characters will not contribute
any phylogenetic information. There is no way in which you can treat gaps as a fifth state
in MrBayes (but see below for information on how you can use gap information in your
analysis).
How do I use gap information in my analysis?
Often, insertion and deletion events contain phylogenetically useful information.
Although MrBayes 3 is not able to do statistical multiple sequence alignment, treating the
insertion-deletion process under a realistic stochastic model, there is nevertheless a way
of using some of the information in the indel events in your MrBayes analysis: Code the
indel events as binary characters (presence/absence of the gap) and include them as a
separate binary (restriction) data partition in your analysis. See more information on this
possibility in the section on the binary model in this manual.
What do I do when it is difficult to get convergence?
There are several things you can do to improve the efficiency of your analysis. The
simplest is to just increase the length of the run. However, the computational cost of
doing so may be prohibitive. A better way is then to try improving the mixing behavior of
the chain. First, examine the acceptance rates of the proposal mechanisms used in your
analysis (output at the end of the run). The Metropolis proposals used by MrBayes work
best when their acceptance rate is neither too low nor too high. A rough guide is to try to
get them within the range of 10 % to 70 %. Rates outside this range are not necessarily a
big problem but they typically mean that the analysis is inefficient. If the rate is too high,
you can make the proposal bolder by changing tuning parameters (see Appendix) using
the props command. Be warned, however, that changing tuning parameters of proposals
and proposal probabilities may destroy any hope of getting convergence. For instance,
you need at least one move changing each parameter in your model.
The next step is to examine the heating parameters if you are using Metropolis-coupled
MCMC. If acceptance rates for the swaps between adjacent chains (the values close to
the diagonal in the swap statistics matrix) are low, then it might be a good idea to
decrease the temperature to make the cold and heated chains more similar to each other
so that they can change states more easily. The efficiency of the Metropolis coupling can
also be improved by increasing the number of parallel chain.
A good way of improving convergence is to start the analysis from a good tree instead of
starting it from a randomly chosen tree. First define a good tree, with or without branch
lengths, using the command usertree. Then start the chains from this tree using
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mcmcp startingtree=user. A disadvantage with starting the analysis from a good
tree is that it is more difficult to detect problems with convergence using independent
runs. A compromise is to start each chain from a slightly perturbed version of a good tree.
MrBayes can introduce random perturbations of a starting tree; this is requested using
mcmcp nperts=<integer_value>.
How do I run MrBayes in parallel?
See section 7 of this manual.
The likelihood values first increase and then drop. What is the problem?
Several users have observed that likelihood values can sometimes increase in the early
phase of a run and then decrease to a stable value; one user referred to the phenomenon
as “burn-out”. Actually, this type of behavior can be seen with certain types of data sets
and models and is part of the normal burn-in. However, it does indicate a problem with
the model. Typically, the problem is due to over-parameterization, a poor prior, or a
combination of these factors.
In MrBayes, the starting value for most parameters is an arbitrarily chosen value that is
likely to be close to the maximum likelihood estimate (MLE) of the parameter. The MLE
value typically also corresponds to the mode (peak) in the posterior probability
distribution. In most cases, you expect the bulk of the probability mass in the posterior
probability distribution to be in the region close to the MLE. However, it is possible that
there is a region in parameter space with only moderate height (lower likelihood values)
but considerably larger probability mass than the MLE region. It is like comparing the
mass of a tower to the mass of a huge office complex. Even if the tower is considerably
higher, its mass is going to be only a fraction of the mass of the office complex.
A typical situation in which this can occur is if you: (1) use a uniform prior on branch
lengths, which puts considerable prior probability on long branches; (2) have data that are
relatively uninformative about branch lengths; and (3) have a model, such as the gamma
model, with a low but not insignificant probability associated with long branches for
weak data. In such cases, the MLE region at short branch lengths can have considerably
smaller probability mass than the less likely but much larger region at longer branch
lengths.
Unless you feed MrBayes with your own starting tree, the run will start with all branch
lengths set to 0.1. This is close to the MLE region, and in the early phase of the run you
will see the likelihood values climb as the topology is improved by branch
rearrangements while the branch lengths remain small. Eventually, however, the long
branch length region will attract the chain through its high probability mass and you will
see the branch lengths increase and the likelihood values decrease to a stable region.
There are basically two ways of fixing the “burn-out” problem. One is to change your
priors so that they put more probability in the MLE region. An obvious step is to change
a uniform prior on branch lengths to an exponential prior; as explained above in the
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section on branch length priors, an exponential prior is more uninformative than the
uniform prior anyway. The other possibility is to simplify your model. For instance,
assume equal rates over sites instead of a gamma model, or choose a substitution model
with fewer free parameters.
How can I test models using Bayes factors?
The Bayesian approach provides a convenient way of comparing models through the
calculation of Bayes factors, which can be interpreted as indicators of the strength of the
evidence in favor of the best of two models. The Bayes factor values are typically
interpreted according to recommendations developed by Kass and Raftery (1995).
Unlike a hierarchical likelihood ratio test, the models compared with Bayes factors need
not be hierarchically nested. A Bayes factor is calculated simply as the ratio of the
marginal likelihoods of the two models being compared. The logarithm of the Bayes
factor is the difference in the logarithms of the marginal model likelihoods.
The marginal likelihood of a model is difficult to estimate accurately but a rough estimate
may be obtained easily as the harmonic mean of the likelihood values of the MCMC
samples (Newton and Raftery, 1994). MrBayes calculates this estimator when you
summarize your samples with the command sump. In the output from the sump
command, you will find the following table (it might look a little different depending on
how many simultaneous runs you have performed; this table is for two runs):
Estimated marginal likelihoods for runs sampled in files
"replicase.nex.run1.p", "replicase.nex.run2.p", etc:
(Use the harmonic mean for Bayes factor comparisons of models)
Run
Arithmetic mean
Harmonic mean
-------------------------------------1
-5883.41
-5892.10
2
-5883.82
-5892.81
-------------------------------------TOTAL
-5883.60
-5892.52
--------------------------------------
For instance, assume we want to compare a GTR model with an HKY model. Then
simply run two separate analyses, one under each model, and estimate the logarithm of
the marginal likelihoods for the two models (using only samples from the stationary
phase of the runs). Then simply take the difference between the logarithms of the
harmonic means and find the corresponding interpretation in the table of Kass and
Raftery (1995; to use this table, you actually have to calculate twice the difference in the
logarithm of the model likelihoods). The same approach can be used to compare any pair
of models you are interested in. For instance, one model might have a group constrained
to be monophyletic while the other is unconstrained, or one model can have gammashaped rate variation while the other assumes equal rates across sites. As stated above,
models need not be hierarchically nested. An interesting property of the Bayes factor
comparisons is that it can favor either the more complex model or the simpler model, so
they need not be corrected for the number of parameters in the models being compared.
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Additional discussion of Bayesian model testing, with several examples, is found in
Nylander et al. (2004).
Can I do model-jumping in MrBayes?
Bayesian MCMC model jumping provides a convenient alternative to model selection
prior to the analysis. In model jumping, the MCMC sampler explores different models
and weights the results according to the posterior probability of each model.
The only model jumping implemented in MrBayes 3 is the estimation of fixed-rate
amino-acid substitution models (see the section on those models in this manual). General
model jumping across models of different dimensionality will be implemented in version
4 of MrBayes.
ModelTest suggests a model for my data. How do I implement it in MrBayes?
A model selection procedure, such as that implemented in ModelTest and MrModelTest,
often suggests a quite specific model for your analysis, including estimates of all
parameters. This suggestion is often based on several simplifications; for instance, you
might have fixed the topology when comparing models and you might have used a small
set of the possible models. In the Bayesian approach, there is only a moderate
computational penalty associated with estimating parameters rather than fixing them prior
to analysis. The Bayesian approach is typically also good at handling multi-parameter
models. Therefore, we recommend that you take the general type of model suggested by
your model selection method and then estimate all of the parameters in that model in
MrBayes. If the suggested model is not implemented in MrBayes, use the next more
complex model available in the program. If, for some reason, you feel that you really
need to fix model parameters in MrBayes to specific values, you can do that using
prset <parameter_prior_name> = fixed (<value or commaseparated values>). For instance, if you want to fix the shape parameter of the
gamma model to 0.12, use prset shapepr=fixed(0.05).
How many data partitions can I have in MrBayes?
MrBayes 3 allows 150 partitions. If you need more partitions, simply change the variable
MAX_NUM_DIVS in the source file mb.h and recompile the program.
Does MrBayes run faster on a dual-processor machine?
No. The Windows and Mac versions of MrBayes 3.1 are not multithreaded so they will
not take advantage of more than one processor on a single machine. However, you should
be able to run two copies of MrBayes without noticeable decrease in performance on a
dual-processor machine (provided you have enough RAM for both analyses).
How much memory is required?
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You can calculate the amount of memory needed to store the conditional likelihoods for
an analysis roughly as 2 * (# taxa) * (# states in the Q matrix) * (# gamma categories) * 4
bytes (for the single-precision float version of the code; double the memory requirement
for the double-precision code). The program will need slightly more memory for various
book-keeping purposes but the bulk of the memory required for an analysis is typically
occupied by the conditional likelihoods.
How do I fix the tree topology during an analysis?
In principle, one can fix a tree topology by specifying constraints for all of the nodes in
the tree. However, we do not recommend doing this because it is computationally very
inefficient. A better way is to set the proposal probability of all topology moves to 0
using the props command. Then you need to switch on one proposal that changes
branch lengths but not topology by increasing its proposal probability from 0 to some
reasonable positive value (like 5). The “node slider” is, in our experience, the best of
these proposals.
6. Differences Between Version 2 and Version 3
We have discontinued the development of version 2 of MrBayes and recommend all
users to switch over to version 3. With the release of version 3.1, virtually all models
implemented in version 2 are available in version 3 (plus many more). The only
exception is the time-irreversible model of nucleotide evolution, which is still not
implemented in version 3. If you are interested in seeing this model reappear, let us
know.
An important difference between versions 2 and 3 is found in the way models are
defined. MrBayes 3 by default estimates most parameters, there is no need to specify
estimate for any parameter. Some things that were previously set using lset are now
set using prset. This is true for the amino acid model and for the site specific rates (see
section 4 of this manual for more information on the site specific rate model in version
3). Thus, site-specifc rates, for instance, can no longer be invoked using
rates=sitespec.
In more detail, the changes are as follows, with emphasis on the features in version 2 that
are implemented differently in version 3 (commands and options in version 2 listed
alphabetically):
calibration. Calibration of clock and relaxed-clock trees is not (yet) implemented in
MrBayes 3.
constraint. The format of the constraint now includes a probability value (constraint
<name> <probability> = <list_of_taxa>). The probability value is ignored
by version 3.1 of the program but it must be included in the constraint definition. The
probability value will be used by future versions of the program.
lset aamodel. The amino-acid model is now set using prset aamodelpr.
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lset ancfile. Ancestral states are now written to the .p file(s).
lset basefreq. Whether this parameters is estimated or fixed to a particular value in
version 3 is controlled by setting the prior with prset statefreqpr. By default, all
parameters are estimated. There is no basefreq option in the lset command of
version 3 of MrBayes.
lset clock. Whether the tree is a clock or a non-clock tree is now set using prset
brlenspr.
lset enforcecal. Calibrations are not implemented (yet) in MrBayes 3.
lset enforcecodon. The type of nucleotide model is now set using lset
nucmodel=4by4/doublet/codon.
lset enforcecon. Now set using prset topologypr = constraints
(<list_of_constraints>).
lset inferanc. Now set using report ancstates.
lset inferpossel. Now set using report possel.
lset inferrates. This is now set with report siterates.
lset ncat (number of categories used to approximate the gamma distribution of site rates).
Now set using lset ngammacat to distinguish it from nbetacat (the number of
categories used to approximate the beta distribution of rate / stationary state asymmetry
across sites in the standard model).
lset nonrevmat. Time-irreversible models are not implemented in MrBayes 3.
lset omega. Whether this parameters is estimated or fixed to a particular value in version
3 is controlled by setting the prior with prset omegapr. By default, all parameters are
estimated. There is no omega option in the lset command of version 3 of MrBayes.
lset rates. The site specific rate models (sitespec, ssgamma, ssadgamma) are now
set using the more general partitioning model. See section 3 of this manual as well as the
discussion of partition models in section 4.
lset revmat. Whether this parameters is estimated or fixed to a particular value in version
3 is controlled by setting the prior with prset revmatpr. By default, all parameters
are estimated. There is no revmat option in the lset command of version 3 of
MrBayes.
lset seqerror. The model of sequencing error is no longer implemented in MrBayes 3.
The model typically results in small corrections in partition support values but it conflicts
with important algorithmic short-cuts implemented in version 3.
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lset shape. Whether this parameters is estimated or fixed to a particular value in version 3
is controlled by setting the prior with prset shapepr. By default, all parameters are
estimated. There is no shape option in the lset command of version 3 of MrBayes.
lset sitepartition. The partition of sites is now set using set partition.
lset tratio. Whether this parameters is estimated or fixed to a particular value in version 3
is controlled by setting the prior with prset tratiopr. By default, all parameters are
estimated. There is no tratio option in the lset command of version 3 of MrBayes.
prset basefreqpr. Now set using prset statefreqpr.
prset brlenpr. Now set using prset brlenspr. The options are now more
complicated as well, since the prior includes information both about the general type of
the branch lengths (clock, non-clock) and the specific shape of the prior (for example,
uniform or exponential).
prset siteratepr. Now set using prset ratepr. The options are fixed or
variable (Dirichlet). The parameters of the Dirichlet can be set to reflect various
types of prior information concerning the site rates.
prset qmatpr. Now set using prset revmatpr and prset aarevmatpr for
nucleotide and amino-acid substitution rates, respectively.
set. This command only controlled the autoclose option in MrBayes 2. In version 3 it
controls a number of different things, including the currently selected partition.
shownodes. This command is no longer included in MrBayes 3. Use showtree to
display the user tree.
7. Advanced Topics
7.1. Compiling MrBayes
Compiling the MrBayes executable from the source code can be done on several different
compilers targeting all the common operating systems: Macintosh, Windows, and Unix.
The easiest way to build MrBayes is to use the included Makefile with a make tool.
One can also compile MrBayes with the Metroworks Codewarrior and Microsoft Visual
Studio suites.
7.1.1. Compiling with GNU Make
In the header of the Makefile, you can define a number of variables:
ARCHITECTURE. This variable defines the architecture you are targeting. Setting this
variable is mandatory. For example, for a Unix environment you would use
ARCHITECTURE = unix. Other options are windows and mac.
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MPI. Set this variable to yes if you want to compile the parallel (MPI) version of
MrBayes. This variable is set to no by default, meaning that a serial version of MrBayes
will be built. See below for more information on how to compile and run the MPI version
of MrBayes.
CC. This variable defines which compiler to use. For example, gcc for the GNU
compiler or icc for the Intel C compiler. The default setting is the GNU compiler.
DEBUG. Set this variable to yes if you want to compile a debug version of MrBayes.
This adds the appropriate flag for the GNU gdb debugger.
OPTFLAGS. Sets the optimization flags for the compiler. This option is ignored if
DEBUG is set to yes. The default is set to -O2, which yields good results for every
platform. It is, however, possible to perform some tuning with this variable. We give a
few possibilities below for some common processor types, assuming you are using gcc
version 3. See the gcc manual for further information on optimization flags.
Intel x86. Some compiler flags for gcc under unix and for gcc/cygwin under
windows:
-march=X, with X one of pentium4, athlon-xp or opteron. If you have one
of these processors this will generate instructions specifically tailored for that
processor.
-mfpmath=sse attempts to use the SSE extension for numerical calculations. This
flag is only effective in combination with the above mentioned -march flag. This
flag can provide a big performance gain. However, using this flag in combination
with other optimization flags might yield numerically incorrect code. For example,
one can set
-mfpmath=sse,386 but this flag leads to incorrect results when used in
combination with -march=pentium4.
-fomit-frame-pointer saves some function overhead.
-O3 instead of -O2 turns on even more optimization flags. However, it does not
always produce faster code than -O2.
Mac G4 and G5. Some compiler flags for gcc for OS X:
-fast. This flag is specific for the gcc version delivered by Apple. It turns on a set
of compiler flags which tries to optimize for maximum performance. This is the
recommended setting if you have a G5 processor and this version of gcc.
-O2 or -O3.
-mcpu=X, with X one of G4 or G5.
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Setting -mcpu or -fast on the Mac results in gcc enabling a number of different
flags. Read the gcc manual carefully if you want to experiment with other flags.
7.1.2. Compiling with Code Warrior or Visual Studio
We provide MrBayes project files for both Metrowerks Code Warrior and Microsoft
Visual Studio in the source code package. All the relevant flags are set in these files, so
you should be able to compile the code without any further modifications.
7.2. Compiling and Running the Parallel Version of MrBayes
Metropolis coupling or heating is well suited for parallelization. MrBayes 3 takes
advantage of this and uses MPI to distribute heated and cold chains among available
processors (Altekar et al., 2004). There are two MPI versions of MrBayes. The first is the
parallel version for Macintosh computers distributed as part of the Macintosh package. It
is intended for use on clusters of Macintosh computers and runs under POOCH, which
must be installed first. The second MPI version of MrBayes is intended for use on
clusters running UNIX and must be compiled from the source code.
7.2.1. The Parallel Macintosh Version
There are several options available for running jobs in parallel on clusters of Macintosh
computers. For example, in OS X, you could configure your machine to run jobs using
mpich or lam-mpi and then compile the regular Unix MPI version of the program as
described in the next section. However, the simplest method is to use Dean Daugger's
program Pooch (available at www.daugerresearch.com/pooch/whatis.html) to control the
jobs. The Pooch web site gives a good description of the steps required to run a job in
parallel. The steps are as follows:
1. Configure a network of Macintosh computers. You have probably already done
this step! You simply need more than one computer hooked to the internet.
2. Buy and install a copy of Pooch for each computer you intend to run MrBayes on.
3. Start Pooch on all of the computers of your cluster. If you set Pooch to
automatically start on login, then this has already been done.
4. Select 'New Job' from Pooch on one of the computers.
5. Select the nodes (computers) you want to participate in the parallel job.
6. Drag the 'pMrBayes_3.1' application and the nexus file you wish to run to Pooch's
Job Window.
7. Click the 'Launch Job' button.
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From this point on, MrBayes behaves just like the serial version of the program.
7.2.2. The MPI Version for Unix Clusters
The MPI version for Unix clusters, including Xserve clusters, has to be compiled before
you can run it. To tell the compiler that you want the MPI version, you need to change a
line in the top section (the configuration section) of the Makefile. The line originally
reads:
MPI = no
Change this to:
MPI = yes
If your system is set up correctly, among other things you need to have mpicc with the
relevant libraries installed, you should now be able to compile the MPI version of
MrBayes. A typical make session would look as follows, after the Makefile has been
appropriately edited:
[[email protected] ~/mpi_mbdev]$ make
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c bayes.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c command.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c mbmath.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c mcmc.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c model.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c plot.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c sump.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized -c sumt.c
mpicc -DUNIX_VERSION -DMPI_ENABLED -O3 -Wall -Wno-uninitialized bayes.o
command.o mbmath.o mcmc.o model.o plot.o sump.o sumt.o -lm -o mb
and produces an MPI-enabled version of MrBayes called mb. Make sure that the mpicc
compiler is invoked and that the MPI_ENABLED flag is set. It is perfectly normal if the
build process stops for a few minutes on the mcmc.c file; this is the largest source file
and it takes the compiler some time to optimize the code. How you run the resulting
executable depends on the MPI implementation on your cluster. At FSU we typically run
MrBayes using LAM/MPI. First, the LAM virtual machine is set up as usual. Then the
parallel MrBayes job is started with a line such as
$mpirun –np 4 mb batch.nex > log.txt &
to have MrBayes process the file batch.nex and run all analyses on four processors
(-np 4), saving screen output to the file log.txt. If you keep both a serial and a
parallel version of MrBayes on your system, make sure you are using the parallel version
with your mpirun command.
7.3. Working with the Source Code
MrBayes 3 is written entirely in ANSI C. If you are interested in investigating or working
with the source code, you can download the latest (bleeding edge) version from the
MrBayes 3.1 Manual
5/26/2005
63
MrBayes CVS repository at SourceForge. You can access the CVS repository from the
MrBayes home page at SourceForge (www.mrbayes.sourceforge.net). SourceForge gives
detailed instructions for anonymous access to the CVS repository on their documentation
pages.
If you are interested in contributing code with bug fixes, the best way is to send a diff
with respect to the most recent file versions in the CVS repository to Paul van der Mark
([email protected]), and we will include your fixes in the main development branch
as soon as possible. If you would like to add functionality to MrBayes or improve some
of the algorithms, please contact Paul for directions before you start any extensive work
on your project to make sure your additions will be compatible with other ongoing
development activities. You should also consider whether you want to work with version
3 or version 4 of the program. We are currently shifting our focus to the development of
MrBayes 4. Unlike version 3, which is written in C, this version will be written in C++
and our goal is to provide a cleaner, faster, and more extensively documented
implementation of Bayesian MCMC phylogenetic analysis. This means, among other
things, that the code will be better organized, and all important sections will be
documented using Doxygen (www.doxygen.org) for easy access to other developers. You
are welcome to examine this project as it develops by downloading the source code,
doxygen documentation, or programming style directives from the MrBayes CVS
repository at SourceForge.
8. Acknowledgements
We would like to acknowledge the invaluable help we have received from students,
colleagues and numerous users of MrBayes; they are too many to name them all here.
Often, we have been overwhelmed by the generosity with which people have shared
ideas, bug fixes and other valuable tips with us. This feedback alone makes all the hours
we have put into developing MrBayes worthwhile. Thank you, all of you!
9. References
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and a model of amino acid substitution for proteins encoded by chloroplast DNA.
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Altekar, G., S. Dwarkadas, J. P. Huelsenbeck, and F. Ronquist. 2004. Parallel
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Bishop, M.J., and A.E. Friday. 1987. Tetropad relationships: the molecular evidence. Pp.
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Felsenstein, J. 1992. Phylogenies from restriction sites: A maximum-likelihood approach.
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Hasegawa, M., H. Kishino, and T. Yano. 1985. Dating the human-ape split by a
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Henikoff, S., and J. G. Henikoff. 1992. Amino acid substitution matrices from protein
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Holder M. and P. O. Lewis. 2003. Phylogeny estimation: Traditional and Bayesian
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Huelsenbeck, J. P., F. Ronquist, R. Nielsen and J. P. Bollback. 2001. Bayesian inference
of phylogeny and its impact on evolutionary biology. Science 294: 2310-2314.
Huelsenbeck, J. P. H., B. Larget, R. E. Miller, and F. Ronquist. 2002. Potential
applications and pitfalls of Bayesian inference of phylogeny. Systematic Biology
51:673-688.
Huelsenbeck, J. P. 2002. Testing a covariotide model of DNA sub- stitution. Molecular
Biology and Evolution 19(5):698-707.
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Bioinformatics 17:754-755.
Huelsenbeck, J. P., and J. P. Bollback. 2001. Empirical and hierarchical Bayesian
estimation of ancestral states. Systematic Biology 50:351-366.
Jones, D.T., W. R. Taylor, and J. M. Thornton. 1992. The rapid generation of mutation
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Restriction
0-1
Data Type
State Frequencies
equal/estimated (SymmDir)
prset symdirihyperpr
fixed/estimated (Dirichlet)
prset statefreqpr
State Frequencies
(Substitution Rates)
F81/HKY/GTR
lset nst=1/2/6
Substitution Rates
equal/gamma
lset rates
equal/gamma
lset rates
Across-Site
Rate Variation
all/variable/informative
lset coding
all/variable/
nopresencesites/noabsencesites
lset coding
Coding Bias
Models supported by MrBayes 3 (simplified)
Standard
0-9
Model Type
fixed/est. (Dirichlet)
prset statefreqpr
Misc.
equal/gamma/
propinv/invgamma/
adgamma
lset rates
Across-Site
Rate Variation
yes/no
lset covarion
Across-Tree
Rate Variation
equal/Ny98/M3
lset omegavar
Across-Site
Omega Variation
F81/HKY/GTR
lset nst=1/2/6
F81/HKY/GTR
lset nst=1/2/6
equal/gamma/
propinv/invgamma
lset rates
unordered/ordered
ctype
Data Type
4by4
lset nucmodel
fixed/est. (Dirichlet)
(over 16 states)
prset statefreqpr
DNA
ACGT
doublet
lset nucmodel
codon
lset nucmodel
fixed/est. (Dirichlet)
(over 61 states)
prset statefreqpr
Data Type
GTR
prset aamodelpr
Model Type
fixed/est. (Dirichlet)
prset statefreqpr
fixed/est. (Dirichlet)
prset statefreqpr
State Frequencies
fixed to equal
fixed/est. (Dirichlet)
prset statefreqpr
Substitution Rates
Models supported by MrBayes 3 (simplified)
Protein
A-Y
Equalin
prset aamodelpr
fixed/mixed
Non-clock
Tree Type
Unconstrained
prset brlenspr
Brlens Type
Uniform
prset brlenspr
Exponential/Uniform
prset brlenspr
Brlens Prior
Across-Tree
Rate Variation
page 2
Across-Site
Rate Variation
yes/no
lset covarion
yes/no
lset covarion
yes/no
lset covarion
equal/gamma/
propinv/invgamma/
adgamma
lset rates
equal/gamma/
propinv/invgamma/
adgamma
lset rates
equal/gamma/
propinv/invgamma/
adgamma
lset rates
Brlens Variation
Across Partitions
equal/proportional
prset ratepr
Theta, Ploidy
Growth
same/unlinked
link topology
unlink topology
Topology Variation
Across Partitions
unlinked
unlink brlens
Speciation
Extinction
Sampleprob
Treeheight
Additional
parameters
see prset
(lset for ploidy)
fixed/mixed
Poisson/Jones/
Dayhoff/Mtrev/
Mtmam/Wag/
Rtrev/Cprev/
Vt/Blossum/
mixed
prset aamodelpr
Clock
Clock
prset brlenspr
Coalescence
prset brlenspr
Birth-Death
prset brlenspr
The most common proposal types used by MrBayes 3
Sliding Window Proposal
Dirichlet proposal
δ
α large
α small
x
New values are picked uniformly from a sliding window
of size δ centered on x.
Tuning parameter: δ
Bolder proposals: increase δ
More modest proposals: decrease δ
Works best when the effect on the probability of the
data is similar throughout the parameter range
x
New values are picked from a Dirichlet (or Beta) distribution
centered on x.
Tuning parameter: α
Bolder proposals: decrease α
More modest proposals: increase α
Works well for proportions, such as revmat and statefreqs.
Multiplier Proposal
Node Slider
a
b
x
x/a x
ax
New values are picked from the equivalent of a
sliding window on the log-transformed x axis.
Tuning parameter: λ = 2 ln a
Bolder proposals: increase λ
More modest proposals: decrease λ
Two adjacent branches a and b are chosen at random
The length of a + b is changed using a multiplier with tuning
paremeter λ
The node x is randomly inserted on a + b according to a
uniform distribution
Bolder proposals: increase λ
More modest proposals: decrease λ
Works well when changes in small values of x have
a larger effect on the probability of data than
changes in large values of x. Example: branch lengths.
The boldness of the proposal depends heavily on the uniform
reinsertion of x, so changing λ may have limited effect
LOCAL
Extending TBR
A
a
b
c
A
x
a
y
B
B
Three internal branches - a, b, and c - are chosen at random.
Their total length is changed using a multiplier with tuning
paremeter λ.
One of the subtrees A or B is picked at random.
It is randomly reinserted on a + b + c according to a uniform distribution
Bolder proposals: increase λ
More modest proposals: decrease λ
Changing λ has little effect on the boldness of the proposal
An internal branch a is chosen at random
The length of a is changed using a multiplier with tuning
paremeter λ
The node x is moved, with one of the adjacent branches, in subtree A,
one node at a time, each time the probability of moving one more
branch is p (the extension probability).
The node y is moved similarly in subtree B.
Bolder proposals: increase p
More modest proposals: decrease p
Changing λ has little effect on the boldness of the proposal.
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