Frequently asked questions
From mothur
If you have any questions, please feel free to email us or to use the discussion tab. If you have an answer, feel free to contribute!
How do I cite mothur?
Schloss, P.D., et al., Introducing mothur: Open-source, platform-independent, community-supported software for describing and comparing microbial communities. Appl Environ Microbiol, 2009. 75(23):7537-41.
The original tools:
Schloss, PD & Handelsman, J. (2006) Introducing SONS, a tool for OTU-based comparisons of membership and structure between microbial communities. Applied and Environmental Microbiology. 72:6773-9.
Schloss, PD & Handelsman, J. (2006). Introducing TreeClimber, a test to compare community structures. Applied and Environmental Microbiology. 72: 2379-84.
Schloss, PD & Handelsman, J. (2005). Introducing DOTUR, a computer program for defining operational taxonomic units and estimating species richness. Applied and Environmental Microbiology. 71:1501-6.
Schloss, PD, Larget, BR, & Handelsman J. (2004). Integration of microbial ecology and statistics: a test to compare gene libraries. Applied and Environmental Microbiology. 70:5485-92.
Why does the command window open and close quickly when I double click on the mothur?
This is because you have not given mothur the input files and you will get an error message quickly followed by the screen closing. Please see the section on how to run mothur from a command prompt.
Why doesn’t mothur do…?
If you would like to see something added to mothur, please let us know! It may take me a while to get around to implementing the feature, but we are generally reasonable and welcome everyone's expertise that would like to contribute source code.
Do you do workshops?
Yes! Please see our workshops page for more information.
Why is data missing for some distance levels?
Perhaps the second most commonly asked question is why there isn't a line for distance 0.XX. If you notice the previous example the distances jump from 0.003 to 0.006. Where are 0.004 and 0.005? mothur only outputs data if the clustering has been updated for a distance. So if you don't have data at your favorite distance, that means that nothing changed between the previous distance and the next one. Therefore if you want OTU data for a distance of 0.005 in this case, you would use the data from 0.003. But... many of the commands that use a label option are smart (e.g. read.otu). So, if you say you want data at the 0.03 cutoff, then you can set label=0.03 and mothur will figure out what data to give you.
Aren't the 'unique' and '0.00' distance levels the same?
Perhaps the most commonly asked question is why the cluster command produces data for both the "unique" and "0.00" lines. Aren't they the same? No. The "unique" line represents data for the situation where all of the sequences in an OTU are identical; the "0.00" line represents data for the situation where all of the sequences in an OTU have pairwise distances less than 0.0049. We made the decision that because there is error in everything, we should round these distances as well and not apply a hard cutoff at 0.01, 0.02, etc. But... if you don't buy into this and want a hard cutoff you can set the hard=T option in read.dist and cluster.
Mothur crashes when I read my distance file
There are two common causes for this, file size and format.
File Size:
- The read.dist and cluster commands load your distance matrix into RAM, and your distance file is most likely too large to fit in RAM. There are two options to help with this. The first is to use a cutoff. By using a cutoff mothur will only load distances that are below the cutoff. If that is still not enough, there is a command called hcluster, http://www.mothur.org/wiki/Hcluster which is memory light. A word of caution with hcluster, the average neighbor implementation is very slow, so if your analysis permits, I would recommend using furthest or nearest, by default mothur uses furthest. You may also be able to reduce the size of the original distance matrix by using the commands outline in the Costello Stool Analysis http://www.mothur.org/wiki/Costello_stool_analysis.
Wrong Format:
- This error can be caused by trying to read a column formatted distance matrix using the phylip parameter. By default, the dist.seqs command generates a column formatted distance matrix. To make a phylip formatted matrix set the dist.seqs command parameter output to lt.
Mothur can't find my input files
The most common cause of mothur not finding a file is because you double clicked on the mothur executable to run mothur. This will open a terminal or command prompt window in your home directory. Mothur will then look for the input files in the home directory instead of in the directory where mothur's executable is located. If this is the cause, you either put the input files in your home directory, give complete file names, or open a terminal or command prompt window, cd into the directory where the mothur executable is located and run mothur by using "./mothur" for mac or "mothur" for windows.
