Hello,
I have generated a .sharedotus file, using the venn command. I now have a list of OTUs which are shared between all of my samples, which is great because I want to identify a “core” microbiome. However, is there a command that can be used to attach the respective classifications to these OTUs?
Thanks in advance,
Jo
Thank you for your quick reply.
Could you specify for me what file I need to add for the template and taxonomy parameters of the classify.seqs command? I get some pretty alarming error messages when progressing onto classify.otus after this command, so would be great if you could confirm this.
Thanks,
Jo
So if you are doing something similar to Pat’s example analysis, http://www.mothur.org/wiki/Schloss_SOP, the classify.seqs command would look something like:
classify.seqs(fasta=GQY1XT001.shhh.trim.unique.good.filter.unique.precluster.pick.fasta, name=GQY1XT001.shhh.trim.unique.good.filter.unique.precluster.pick.names, group=GQY1XT001.shhh.good.pick.groups, template=trainset9_032012.pds.fasta, taxonomy=trainset9_032012.pds.tax, cutoff=80, processors=2)
The training sets can be found here: http://www.mothur.org/wiki/Silva_reference_files
You want to classify your OTUs using the final.taxonomy file, which has the contaminants removed. If you continue along the SOP, the classify.otu command should look something like:
classify.otu(list=final.an.list, taxonomy=final.taxonomy, name=final.names)
or
classify.otu(list=final.an.unique_list, taxonomy=final.taxonomy, count=final.count_table)
What are the errors you are getting?
Thanks for your reply, and this did confirm what I was entering previously.
I get long clusters of error messages which include:
[ERROR]: Otu1278 is missing the final ‘;’, ignoring.
[ERROR]:Bacteria(100);unclassified(100);unclassified(100);unclassified(100);unclassified(100);unclassified(100); is missing the final ‘;’, ignoring.
[ERROR]: 1 is already in your taxonomy file, names must be unique.
Not sure if I’m missing something here…
Cheers,
Jo
It sounds like you are giving it the wrong kind of input file. Mothur is expecting sequences in the final.taxonomy file to look like this:
GQY1XT001C296C Bacteria(100);“Bacteroidetes”(100);“Bacteroidia”(95);“Bacteroidales”(95);“Porphyromonadaceae”(91);unclassified;
I think you are giving it a *cons.taxonomy file. Something like:
Otu01 29578 Bacteria(100);“Bacteroidetes”(100);“Bacteroidia”(100);“Bacteroidales”(100);“Porphyromonadaceae”(100);unclassified(100);
The first example classifies a sequence. The second example is a OTU classification.
Thank you very much for your help - the command is running now. I was indeed using a .cons.taxonomy file, instead of a .taxonomy file.
Jo
Apologies for another post, but I am getting myself tied in a knot over this!
After generating a .sharedotus file, is it possible to take this file directly forward and assign taxonomy to the shared otus? Or, would I need to somehow generate a group file for the shared otus?
Thanks,
Jo
Perhaps I should clarify what I would like to do here…
I have three gut lumen samples, of which there are 200 shared OTUs between all samples. I would like to generate a list of these 200 shared OTUs, and then assign taxonomy to them - therefore, giving me an idea of the bacteria which may be shared in this environment.
As yet, I can’t seem to find a way to do this. Sorry if I’m missing something obvious!
Cheers,
Jo
Have you run the classify.otu command for your whole data set?
Do you now have the consensus taxonomy of your OTUs?
As you know the OTU numbers of your shared OTUs can you just look at those OTUs in the consensus taxonomy (e.g. open the consensus taxonomy in excel) and there you will find the information you need.
Best,
Antti Rissanen
Dear Antti,
Thank you for your reply. Yes, I have the consensus taxonomy for my whole dataset and I know that there are 262 shared OTUs between all samples. However, how can I identify which OTU numbers are the ones which are shared?
Cheers,
Jo
Hi, I have not used venn-command but according to this, http://www.mothur.org/wiki/Venn#sharedotus, you should have the OTU labels that are shared among groups in the sharedsobs.sharedotus - file.
Antti Rissanen
Thanks so much - much appreciated!
I’m having the same issue as above - I have generated a .sharedotus file, and it contains hundreds of OTU’s - and this is just for one comparison I’m doing. It seems like there has to be a better way to convert the list of OTU’s to a taxonomy, since I already have all of the information and just need to do the mapping…is there?
You can do this with the following:
mothur > get.sharedseqs(shared=yourSharedFile, uniquegroups=group1-group2, label=yourLabel, output=accnos) - list OTU names in overlap in venn diagram. You can use various settings with unique groups and sharedgroups to find the different parts of the venn diagram.
mothur > get.otulabels(constaxonomy=yourConstaxonomyfile, accnos=accnosFileFromGetSharedSeqs) - select those Otus from constaxonomy file