cluster.split

Commands in mothur
1

Hi

I used the cluster.split command and used label=0.03 following Your protocol (https://www.mothur.org/wiki/MiSeq_SOP). As result I get a an.unique_list.list where it says unique 0.01? Why do I get the list with 0.01 results, when I specificed the label= 0.03??

Thank you very much!

Kristin

2

Can you upgrade to v1.39.5 and try again?

pat

3

Hello,

I have this same issue, both using v1.38.1 and 1.39.5. I’ve followed the MiSeq SOP, and right before dist.seqs my fasta looks like this:

  Start End NBases Ambigs Polymer NumSeqs
Minimum: 1 503 250 0 3 1
2.5%-tile: 1 503 252 0 3 4312
25%-tile: 1 503 253 0 4 43114
Median:  1 503 253 0 5 86228
75%-tile: 1 503 253 0 5 129342
97.5%-tile: 1 503 254 0 6 168144
Maximum: 1 503 254 0 8 172455
Mean:  1 503 252.968 0 4.65556
# of Seqs: 172455

I’ve run dist.seqs and cluster with a cutoff at 0.03. I’ve also run dist.seqs with a cutoff at 0.20 and cluster at 0.03 or 0.20.

I’m currently trying to use cluster.split with splitmethod=classify and cutoff at 0.03. My shared file always gives me “unique” instead of 0.03. Are my OTUs actually clustered at 97%? It looks like my total OTUs in my shared file are equal to my total number of unique sequences…

Thank you,
Patric

4

are you using method=an or method=opti? You should be using method=opti. For why an gives you 0.01 when you give it 0.03 data see this FAQ:

https://mothur.org/wiki/Frequently_asked_questions#Why_does_the_cutoff_change_when_I_cluster_with_average_neighbor.3F

Pat