Get.groups in MiSeq SOP

Commands in mothur
1

Hello all,

I’m following the MiSeq SOP nd all is going well until I get to the assessing error rates of the SOP. When I enter the get.groups command, the following appears:

_mothur > get.groups(count=stability.trim.contigs.good.unique.good.filter.unique.precluster.denovo.vsearch.pick.pick.count_table, fasta=stability.trim.contigs.good.unique.good.filter.unique.precluster.pick.pick.fasta, groups=Mock)

[NOTE]: The count file should contain only unique names, so mothur assumes your fasta, list and taxonomy files also contain only uniques.

Mock is not a valid group, and will be disregarded.
You provided no valid groups. I will run the command using all the groups in your file.
Selected 116011 sequences from your fasta file.
Selected 662924 sequences from your count file._

Could anyone explain to me what the problem is here?

Kind Regards,
Aidan.

2

Did you sequence a mock community as part of your sequencing run? If not, you’ll have to skip this part of the SOP because you don’t have reference data to compare to. If you did sequence a mock community, what name did you give it in your data?

3

Hello,

I am encountering the same problem. I am using the subset data provided on the wiki MiSeq SOP page. https://www.mothur.org/wiki/MiSeq_SOP. This example data contains mock community fastq files called : Mock_S280_L001_R1_001.fastq and Mock_S280_L001_R2_001.fastq

I am using mothur version 1.39.5_1. My code and output are below:


_mothur > get.groups(count=stability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table, fasta=stability.trim.contigs.good.unique.good.filter.unique.precluster.pick.pick.fasta, groups=Mock)

[NOTE]: The count file should contain only unique names, so mothur assumes your fasta, list and taxonomy files also contain only uniques.

Mock is not a valid group, and will be disregarded.
You provided no valid groups. I will run the command using all the groups in your file.
Selected 1342 sequences from your fasta file.
Selected 117930 sequences from your count file._[/i]

Many thanks in advance for advice!

Sarah

4

That is strange, because I just downloaded the files and the mock community is definitely in there, under the name ‘Mock’.

What do you see if you run the following command?

mothur > count.seqs(count=stability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table)
5

My output is below:

mothur > count.seqs(count=stability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table)
count is not a valid parameter.
The valid parameters are: shared, name, group, processors, groups, seed, inputdir, and outputdir.
Using MiSeq_SOP/stability.trim.contigs.good.names as input file for the name parameter.

Using 8 processors.
[ERROR]: did not complete count.seqs.

6

Damn! Sorry, I always make that mistake. Can you run count.groups() using the same file?

7

No worries! Output is below:

mothur > count.groups(count=stability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table)

Unable to open stability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table. Trying default /usr/local/bin/mothur/stability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table
Unable to open /usr/local/bin/mothur/stability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table. Trying mothur’s location /usr/local/bin/mothurstability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table
Unable to open /usr/local/bin/mothurstability.trim.contigs.good.unique.good.filter.unique.precluster.denvo.vsearch.pick.pick.count_table

You have no current groupfile, countfile or sharedfile and one is required.
[ERROR]: did not complete count.groups.

8

It looks like you’re in the wrong directory for your count table.

9

I agree it appears that way at first. However, I see the file in my current working directory. I received the same error when I ran the command from one directory above.

Do you know what file(s) it is referring to when it says:

“You have no current groupfile, countfile or sharedfile and one is required?”

10

For that message, it just means that if the file you specify isn’t found then it looks for a previous one in the current files cache.

If you’re absolutely sure you’re working from the correct directory, the only other thing I can think of is that there might be spaces in the path name? Sometimes mothur has issues with those.