Classifying seqs and shared file query

Commands in mothur
1

Hi all,

I possibly have a very obvious/silly question or rambling here. I’ve tried looking on the forum but I can’t find what I’m looking for (possibly because the answer is so basic!) :expressionless: Although I did find this thread helpful Phylotype vs OTUs

But I’m wondering if there is a way in mothur to make six separate shared files each with a different level of taxonomic classification? That would be the phylotype binning? And I get my taxonomy file after this and I can edit my shared file then with the classification of each OTU? (As I want to make a biom file of my shared file and import into QIIME).

I did some analysis in QIIME with the OTU-based approach using my Mothur generated shared files. But my supervisor wants me to look at the results of the analysis based on phylum level. Can I do this with my OTU -based shared files or do I need to use a phylotype-based approach for this? I was wondering if I could manually edit my files down to Phylum level etc since say I know that say for OTU 1 it is classified as (Bacteria; Proteobacteria; Gammaproteobacteria; Vibrionales; Vibrionaceae; Photobacterium)? Or would you cluster.split to say Phylum level then make shared and then classify?

I appreciate any help you could offer,

Ciara
NUI Galway

2

mothur shared files can hold multiple levels of clustering/phylotyping within a single shared file, so you don’t necessarily need to split them out.

If you classified your data using SILVA or RDP, then do

mothur > phylotype(taxonomy=current, count=current)
mothur > make.shared(list=current, count=current, label=1-2-3-4-5)

You’ll get all the levels of clustering within the single file (1 = genus, 5 = phylum). I think if you use Greengenes you’ll need 1 -> 6, since it includes species level classifications.

mothur can handle the multiple levels fine - in all the alpha/beta calculators your commands will be performed on each level independently, or you can specify which level to use with the label command.

If you need to use a different program then you’ll need to split them out properly. You can use the filter.shared() command, e.g.

mothur > filter.shared(shared=current, label=2)

Or just run the original make.shared() with a single label specified.

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