Dear all,
after Illumina sequencing of 16S rRNA regions V345 (500 bp) I was trying to follow to the protocol of schloss et al. In chapter pre.cluster it is explained that for each 100 bp a difference of 1 should be allowed. First I run the protocol with diff=2 - everything was working out. Then I realized that for my dataset, I have to allow diff=5. but for this command I got a error message - see below:
problem is, that I only got the unique sequences and I need the label 0.03.
mothur > pre.cluster(fasta=merged.5.trim.trim.contigs.trim.unique.good.filter.unique.fasta, name=merged.5.trim.trim.contigs.trim.unique.good.filter.names, group=mergegroupsgood, diffs=5)
Output File Names: merged.final.rdp.an.sabund merged.final.rdp.an.rabund merged.final.rdp.an.list
mothur > classify.otu(list=merged.final.rdp.an.list, name=merged.final.rdp.names, taxonomy=merged.final.rdp.wang.taxonomy, label=0.01-0.03-0.05)
reftaxonomy is not required, but if given will keep the rankIDs in the summary file static.
Your file does not include the label 0.01. I will use unique.
Your file does not include the label 0.03. I will use unique.
Your file does not include the label 0.05. I will use unique.
unique 91214
Output File Names:
merged.final.rdp.an.unique.cons.taxonomy
merged.final.rdp.an.unique.cons.tax.summary
mothur > make.shared(list=merged.final.rdp.an.list, group=merged.final.rdp.groups, label=0.01-0.03-0.05)
unique
Output File Names:
merged.final.rdp.an.shared
merged.final.rdp.an.AAns.rabund
merged.final.rdp.an.AAps.rabund
merged.final.rdp.an.ABns.rabund
merged.final.rdp.an.ABps.rabund
merged.final.rdp.an.AFns.rabund
…
Do anybody has a clue what could be the problem?
Many thanks in advance!