Passing primer info


Is there a specific way the primer info is passed to mothur that is also reflected in the header line of the fasta file? I’m getting NULL output from trim.seqs() and the header file from our sequencer output looks different than the one used by mothur in the stool example. Can anybody help me understand the specific file formats used by mothur for inputting fasta and oligos files? I copied the format of stool.oligos, but it didn’t help 'cos the primer ID on the fasta file is right at the beginning and I am not sure if that’s a requirement of the mothur parser?

Please point me to any important mothur docs that talk about preparing input files for mothur. Thanks much in advance,


If you look at the wiki page for trim.seqs it should be in there. You need to supply an oligos file with the barcode and primer information.


Okay, I just checked the scrap file produced by the trim.seqs function and it looks like all of my input short sequences have been rejected. Is that due to short lengths (e.g., 46, 47, 48, 65…etc.) or could it be due to improperly created oligos file? My oligos file has the exact format required by the trim.seqs documentation. How may I debug the situation? Any clues please? TiA,



There’s a code in the scrap file. If you look at the sequence names you should see something like this…


This means that the sequence was chucked because of length. If you see this…


It was chucked because it couldn’t find the barcode and the forward primer. If you look at the trim.seqs page you should see a description of this coding system.



I too am new to this a struggling quite a lot. I carried out the Costello eg. no problems but when applying this to my own samples I can not get past step one!

I have 18 .sff files (from 18 samples), how can I get these into the correct formatt so it possible to analyse the samples in the same way as the Costello e.g? Even just converting the sff file to a fasta file (untrimmed) and then trying to do the trim.seqs command as in the Costello e.g returns an error saying the file is blank and then mothur fails to respond and I have to close the window.

Im running this on a windows 7 OS with 4gb RAM, intel CORE i5 processor.

Any help is greatly appriciated, depite days of trying I seem to be getting nowhere!


Have you looked at the sffinfo command,