An error was encountered while running DADA2

Technical Support
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1

Hi,

I run DADA2 and I encountered an error after approximately 15 minutes. My data is single-end reads generated using Illumina Miseq.

Here is the command for importing my data:
qiime tools import
–type ‘SampleData[SequencesWithQuality]’
–input-path manifest.txt
–output-path single-end-demux.qza
–input-format SingleEndFastqManifestPhred33V2

Here is the command to run DADA2:
qiime dada2 denoise-single
–i-demultiplexed-seqs single-end-demux.qza
–p-trim-left 0
–p-trunc-len 220
–o-representative-sequences rep-seqs-dada2.qza
–o-table table-dada2.qza
–o-denoising-stats stats-dada2.qza

Here is the content of the log file:

Running external command line application(s). This may print messages to stdout and/or stderr.
The command(s) being run are below. These commands cannot be manually re-run as they will depend on temporary files that no longer exist.

Command: run_dada_single.R /tmp/qiime2-archive-io9g5_0i/0cd39a3f-20f8-474f-b762-c2c68b6bf449/data /tmp/tmpsa8_flhq/output.tsv.biom /tmp/tmpsa8_flhq/track.tsv /tmp/tmpsa8_flhq 220 0 2.0 2 Inf consensus 1.0 1 1000000 NULL 16

R version 3.5.1 (2018-07-02)
Loading required package: Rcpp
DADA2: 1.10.0 / Rcpp: 1.0.3 / RcppParallel: 4.4.4

  1. Filtering …
  2. Learning Error Rates
    226886220 total bases in 1031301 reads from 14 samples will be used for learning the error rates.
    Traceback (most recent call last):
    File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/q2_dada2/_denoise.py”, line 177, in _denoise_single
    run_commands([cmd])
    File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/q2_dada2/_denoise.py”, line 36, in run_commands
    subprocess.run(cmd, check=True)
    File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/subprocess.py”, line 418, in run
    output=stdout, stderr=stderr)
    subprocess.CalledProcessError: Command ‘[‘run_dada_single.R’, ‘/tmp/qiime2-archive-io9g5_0i/0cd39a3f-20f8-474f-b762-c2c68b6bf449/data’, ‘/tmp/tmpsa8_flhq/output.tsv.biom’, ‘/tmp/tmpsa8_flhq/track.tsv’, ‘/tmp/tmpsa8_flhq’, ‘220’, ‘0’, ‘2.0’, ‘2’, ‘Inf’, ‘consensus’, ‘1.0’, ‘1’, ‘1000000’, ‘NULL’, ‘16’]’ died with <Signals.SIGKILL: 9>.

During handling of the above exception, another exception occurred:

Traceback (most recent call last):
File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/q2cli/commands.py”, line 328, in call
results = action(**arguments)
File “</home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/decorator.py:decorator-gen-453>”, line 2, in denoise_single
File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/qiime2/sdk/action.py”, line 245, in bound_callable
output_types, provenance)
File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/qiime2/sdk/action.py”, line 390, in callable_executor
output_views = self._callable(**view_args)
File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/q2_dada2/_denoise.py”, line 212, in denoise_single
band_size=‘16’)
File “/home/qiime2/miniconda/envs/qiime2-2020.2/lib/python3.6/site-packages/q2_dada2/_denoise.py”, line 188, in _denoise_single
" and stderr to learn more." % e.returncode)
Exception: An error was encountered while running DADA2 in R (return code -9), please inspect stdout and stderr to learn more.

Sorry the file is not authorized to be uploaded.

Please let me know if I need to provide any other information.

Thanks.

3

Would someone please help me figure it out.

4

Hi @ptalebic!

Thanks for sharing all of that info - very helpful!

I immediately noticed this:

This is a signal that computers use to tell a process or command to stop work immediately - it is usually initiated by someone like a sysadmin, or a cluster queuing system. Can you tell us a little bit more about your computation environment? Are you running on an institutional cluster? Are you submitting jobs through a queuing system?

Keep us posted!

:qiime2:

PS - I just want to remind you about the QIIME 2 Forum Code of Conduct, in particular, this section:

https://forum.qiime2.org/faq#patience

Sending multiple messages just bogs down our moderator queue, slowing down the time in which we can respond to you, and others. Thanks!

6

Dear Matthew,

Thank you for your reply and forgive me for Not being patient.

7

I am not submitting jobs through a queuing system. Just running the command on my laptop and I don't run it on an institutional cluster.

8

@ptalebic, could you please tell us more about your computing environment?

A quick search of the forum shows that errors like this are often caused by out-of-memory problems. If your sample(s) are large, and your system has limited memory, or you’re using a virtualbox or similar container that isn’t provisioned with enough memory, that may be your issue.

If you have access to a system with more resources, or are able to provision your virtual environment with more memory, you may have better luck.

1 Like
9

Dear Chris,

I am using virtualbox and I dedicated 1024 for memory as recommended. I have above 10 million reads and the file is about 1gb.

How much memory would you suggest I devote to my guest machine?

Best,

12

@ptalebic, this isn’t an issue I’ve dealt with personally before, but I’m sure you can figure it out by trial and error. Remember, if you’re running 64-bit Windows 10, that the host OS needs at least 2GB of dedicated RAM to work smoothly.

Virtualbox allocates memory in MB (1024 MB is ~1GB of memory), and your Box’s 1GB of RAM is probably not enough for DADA2 to run smoothly - try taking a look at how much memory your system has available, and bumping up your VirtualBox allocation without starving the host machine, and seeing how it goes.

1 Like
14

Thanks for the reply.

15

Of course! Let us know how it works out. :nerd_face:

16

Hi Chris,

I allocated much more memory (3000 MB) and it worked. Thank you for your guidance.

1 Like
17

Woohoo! Glad to hear it!
:partying_face::squid:

18

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