I am trying to cluster my sequences into OTUs using the average neighbor algorithm and it won’t cluster below the unique level.
I keep getting the message “cutoff changed to 0”. If I set the cutoff to something (e.g., cutoff=0.10) it still gives me the error message.
The furthest neighbor method works fine and clusters the OTUs from unique to 0.18.
I have a column formatted distance matrix that was generated using ESPRIT.
Thanks for your help,
Thanks for your reply.
I donâ€™t think this error is happening because I have large distances between groups. When I calculated the distances in ESPRIT I used a kmer distance cutoff of 0.5, (which roughly corresponds to a pairwise distance of ~0.18) to speed up the computation. This gets rid of the problem you mention about non-overlapping sequences and infinite distances.
BTW, I tried setting the cutoff to 0.20 and I got the same error message as before.
The reason I had to use ESPRIT was because I have 6000 ITS-LSU sequences that are ~ 1000 bp in length. These sequences were trimmed using phred/phrap and manually edited for errors. Because they are too divergent to align them reliably (they span the Ascomycota), I wanted to generate a distance matrix based on pairwise alignments and it was too many sequences for Mothur to handle. So I only used the ESPRIT kmer distance computation followed by the needledist computation to generate the distance matrix on our university computer cluster. I didnâ€™t use their hcluster algorithm.
Thanks for your help.
Hmm. I understand where you’re coming from. That is weird. Could you send us the distance matrix and names file to take a look for you? Have you tried the paiwise.seqs command which does essentially what esprit is doing without the kmer optimization?
I tried the pairwise.seqs option on my laptop with 4 processors and it was taking a really long time, which is why I used esprit. I didn’t try to run it on the cluster though.
My distance matrix is 50 mb when zipped, which is too large to email. How can I get it to you?
can you post it to the wiki and email me the link?
I made a google doc for the distance matrix and shared it with you, and I emailed you the .names file.