Alignment in QIIME 1.9.0

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Slave Trajanoski

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Apr 10, 2015, 3:08:24 PM4/10/15
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QIIME 1.9.0 is using the latest GreenGenes DB and by default aligns the representative sequences aginst 85*.fasta. The issue that I have is with this setup the reads align with very low identity as in contrast to the previous core_set used before. I tried same dataset once against core_set and once against 85 and realized enormous difference, like 90% identity vs 50%. Is there explanation for this behavior and some solution to it?

John Chase

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Apr 10, 2015, 6:33:09 PM4/10/15
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Hi, 

I have forwarded your question to a QIIME developer who will be able to answer this question better than I can.

John 

On Fri, Apr 10, 2015 at 12:08 PM, Slave Trajanoski <straj...@gmail.com> wrote:
QIIME 1.9.0 is using the latest GreenGenes DB and by default aligns the representative sequences aginst 85*.fasta. The issue that I have is with this setup the reads align with very low identity as in contrast to the previous core_set used before. I tried same dataset once against core_set and once against 85 and realized enormous difference, like 90% identity vs 50%. Is there explanation for this behavior and some solution to it?

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Greg Caporaso

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Apr 13, 2015, 11:55:10 AM4/13/15
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Hello,
Can you provide me with one of the sequences that you observed these differences with (e.g., aligning at 90% against the core set and 50% against the 85% OTUs)? Also, can you provide the commands that you used for both of those alignments.

Thanks!
Greg

Slave Trajanoski

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Apr 14, 2015, 5:47:04 AM4/14/15
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Hi,

i've attached only several sequences from the dataset in question including log files generated for them on my computer with the parallel_align_seqs_pynast.py script. Here are the two commands that I've used for the alignment:

parallel_align_seqs_pynast.py -i ../otus_flx_denovo/rep_set/seqs_chim_filt_rep_set.fasta -o pynast_aligned_seqs/coreset -e 170 -p 50 -t /media/sf_data/projects2015/core_set_aligned.fasta.imputed.fasta -T --jobs_to_start 10
parallel_align_seqs_pynast.py -i ../otus_flx_denovo/rep_set/seqs_chim_filt_rep_set.fasta -o pynast_aligned_seqs/70 -e 170 -p 50 -t /media/sf_data/projects2014/gg_13_8_otus/rep_set_aligned/85_otus.fasta -T --jobs_to_start 10
As you can see I had to lower the percent identity parameter so that the reads pass thruogh and don't get filtered.

Thanks for the help.
Slave
alignment.tar.gz

Greg Caporaso

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Apr 14, 2015, 4:23:56 PM4/14/15
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Hi Slave,
I'm working on this issue now, and can confirm the issue. I think this may be a bug that we'll need to deal with - can you tell me what primers you did your sequencing with? I'm thinking it's 27F/338R, is that right? 

Greg

Greg Caporaso

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Apr 14, 2015, 7:04:54 PM4/14/15
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I'm still working on this, but will need a little more time. I'm keeping notes on progress and work-arounds as I go here:

More soon...
Greg

Slave Trajanoski

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Apr 15, 2015, 2:53:22 AM4/15/15
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Hi Greg,

thanks for your quick response, it helps me to consider how to finish my current project and sorry for my late reply, but it is the time difference. Primers used were 27F/534R (V1-V3).

all the best,
Slave

Greg Caporaso

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Apr 15, 2015, 9:11:43 AM4/15/15
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Hi Slave, No problem, we're used to time zone delays. The work-around that I describe at the link in my previous post should let you complete your analysis. Good luck, and thanks again for your help with this. 

Greg

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Carly

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Jun 2, 2015, 9:09:34 PM6/2/15
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Hello,

I am getting an error when I run the pip install --upgrade qiime-default-reference

The pip.log is attached. I tried running the align_seqs.py again even with the error thinking it might be fine (you never know, right?) and most of my sequences are landing in the failure file.

Thanks,

Carly
pip.log

Yoshiki Vázquez Baeza

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Jun 3, 2015, 2:20:11 PM6/3/15
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Hi Carly,

It seems like you don't have permissions to write to that folder. If you have sudo access to this machine, you can prepend the command with sudo and it should work. But before you try that, you may want to consider waiting for the next version of MacQIIME (Jeff Werner said it wouldn't be too long until it came out).

Thanks!

Yoshiki.
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