# Classify.seq could not be classified **URL:** \ **Category:** mothur bugs\ **Created:** [March 26, 2021, 10:51am UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937 "2021-03-26T10:51:35Z")\ **Posts on this page:** 8\ **Page:** 1
**Author:** ![Nicky](https://avatars.discourse-cdn.com/v4/letter/n/f0a364/32.png) [@Nicky](https://forum.mothur.org/u/Nicky)\ **Post date:** [March 26, 2021, 10:51am UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/1 "2021-03-26T10:51:36Z")
Hi I am trying to run a classify.seq command using the silva database as a reference however I am getting a warning saying [WARNING]: M05510\_34\_000000000-JD7WL\_1\_2119\_20097\_23995 could not be classified. You can use the remove.lineage command with taxon=unknown; to remove such sequences, this is for all my bacteria sequences and when I quit the command it says “all sequences have been removed”. I was using this version of mothur before mothur v.1.41.3 and then changed to this one (mothur v.1.44.3) but I’m still getting the same issue. Please kindly assist. ---
**Author:** ![pschloss](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/pschloss/32/4_2.png) [@pschloss](https://forum.mothur.org/u/pschloss)\ **Post date:** [March 29, 2021, 1:20pm UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/2 "2021-03-29T13:20:20Z")
Hi, Can you post one of the sequences so I can take a look at it? Thanks, Pat ---
**Author:** ![Nicky](https://avatars.discourse-cdn.com/v4/letter/n/f0a364/32.png) [@Nicky](https://forum.mothur.org/u/Nicky)\ **Post date:** [March 29, 2021, 3:28pm UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/3 "2021-03-29T15:28:22Z")
Hello Pat. I was advised to let it run until it finishes, which took a while but I did manage to get a taxonomy file at the end. Apparently, it’s normal for it to do that. kind regards Nicky ---
**Author:** ![Nicky](https://avatars.discourse-cdn.com/v4/letter/n/f0a364/32.png) [@Nicky](https://forum.mothur.org/u/Nicky)\ **Post date:** [April 6, 2021, 12:10pm UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/4 "2021-04-06T12:10:51Z")
Hie Pat I wanted to ask if you could help, when I run my dist.seqs command it says that it is blank and this might be that there are are no distances below my cutoff. I used 0.03. Kind regards Nicky ---
**Author:** ![pschloss](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/pschloss/32/4_2.png) [@pschloss](https://forum.mothur.org/u/pschloss)\ **Post date:** [April 8, 2021, 5:52pm UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/5 "2021-04-08T17:52:06Z")
What are you sequencing? Can you post your commands? ---
**Author:** ![Nicky](https://avatars.discourse-cdn.com/v4/letter/n/f0a364/32.png) [@Nicky](https://forum.mothur.org/u/Nicky)\ **Post date:** [April 8, 2021, 6:06pm UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/6 "2021-04-08T18:06:46Z")
I’m sequencing bacteria, here are my commands #make.contigs(file=bacteria.files) #summary.seqs(fasta=bacteria.trim.contigs.fasta, processors=24) #trim.seqs(fasta=bacteria.trim.contigs.fasta, maxambig=0, minlength=350, maxlength=550) #summary.seqs(fasta=bacteria.trim.contigs.trim.fasta) #unique.seqs(fasta=bacteria.trim.contigs.trim.fasta) #summary.seqs(fasta=bacteria.trim.contigs.trim.unique.fasta, name=bacteria.trim.contigs.trim.names) #count.seqs(name=bacteria.trim.contigs.trim.names, group=bacteria.contigs.groups) #summary.seqs(fasta=bacteria.trim.contigs.trim.unique.fasta, name=bacteria.trim.contigs.trim.names) #chimera.vsearch(fasta=bacteria.trim.contigs.trim.unique.fasta, count=bacteria.trim.contigs.trim.count\_table, dereplicate=t) #remove.seqs(fasta=bacteria.trim.contigs.trim.unique.fasta,accnos=bacteria.trim.contigs.trim.unique.denovo.vsearch.accnos) #summary.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.fasta, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.count\_table, processors=24) #classify.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.fasta, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.count\_table, reference=silva.nr\_v138\_1.align, taxonomy=silva.nr\_v138\_1.tax, cutoff=80) #remove.lineage(fasta=bacteria.trim.contigs.trim.unique.pick.fasta, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.count\_table, taxonomy=bacteria.trim.contigs.trim.unique.pick.nr\_v138\_1.wang.taxonomy, taxon=Chloroplast-Mitochondria-unknown-Archaea-Eukaryota) #summary.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.fasta, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.count\_table, processors=24) #summary.tax(taxonomy=bacteria.trim.contigs.trim.unique.pick.nr\_v138\_1.wang.pick.taxonomy, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.count\_table) #align.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.fasta, reference=silva.nr\_v138\_1.align, flip=T) #summary.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.fasta, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.count\_table) #summary.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.align,count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.count\_table, processors=24) #screen.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.align, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.count\_table, start=11895 end=28464) #summary.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.good.align, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.good.count\_table) #filter.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.good.align, vertical=T, trump=.) #count.groups(count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.good.count\_table) #sub.sample(fasta= bacteria.trim.contigs.trim.unique.pick.pick.good.filter.fasta, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.good.count\_table, size=24804, persample=t) #count.groups(count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.good.subsample.count\_table) #unique.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.good.filter.subsample.fasta, count=bacteria.trim.contigs.trim.denovo.vsearch.pick.pick.good.subsample.count\_table) #classify.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.good.filter.subsample.unique.fasta, #count=bacteria.trim.contigs.trim.unique.pick.pick.good.filter.subsample.count\_table, reference=silva.nr\_v138\_1.align, taxonomy=silva.nr\_v138\_1.tax, cutoff=80) #dist.seqs(fasta=bacteria.trim.contigs.trim.unique.pick.pick.good.filter.subsample.unique.fasta, cutoff=0.03) ---
**Author:** ![pschloss](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/pschloss/32/4_2.png) [@pschloss](https://forum.mothur.org/u/pschloss)\ **Post date:** [April 8, 2021, 6:41pm UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/7 "2021-04-08T18:41:29Z")
I assume 16S? What region? Can you include the output from running `summary.seqs` at each of the steps you have it listed above? ---
**Author:** ![system](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/system/32/2_2.png) [@system](https://forum.mothur.org/u/system)\ **Post date:** [April 18, 2021, 6:41pm UTC](https://forum.mothur.org/t/classify-seq-could-not-be-classified/20937/8 "2021-04-18T18:41:57Z")
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