I am an undergrad working on a soil microbial community project and I am having an issue with the mothur > unique.seqs.(fasta=stability.trim.contigs.good.fasta) command. I have been following the mothur MiSeq SOP. The primers I have are for the 16s rRNA gene for bacteria and archaea, and then the 18s rRNA gene for Eukaryota and our in-house sequencing facility ran them all together. I ran the following command: mothur > screen.seqs(fasta=stability.trim.contigs.fasta, group=stability.contigs.groups, maxambig=0, maxlength=600). I used 600 as my max length because that is about the maximum length of one of the primers I am using. When I run the summary.seqs command I still have sequences showing up in the file that was created from the screen.seqs command. I have attached an image the summary.seqs that I ran.
I then run the unique.seqs command on the file that was created from the screen.seqs command. I then ran a summary.seqs command after I have ran the unique.seqs command, I receive an error saying that the file created after unique.seqs is empty. Has anybody ever received this error. Any information about combatting it would be much appreciated.