Hi,
The Qiime2 tutorials ran flawlessly in my mac. I’m trying to run an analysis on a large file (imported from Casava1.8 demultiplexed files) and am getting the following:
(qiime2-2017.2) dhcp80ffdd4e:A05_Microbiome apzlo$ qiime dada2 denoise-paired --i-demultiplexed-seqs /Users/a
pzlo/Qiime2analyses/A05_Microbiome/demux-paired-end.qza --o-table table --o-representative-sequences rep-seq
s --p-trim-left-f 10 --p-trim-left-r 10 --p-trunc-len-f 230 --p-trunc-len-r 230 --verbose
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_paired.R /var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/forward /var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/reverse /var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/output.tsv.biom /var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/filt_f /var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/filt_r 230 230 10 10 2.0 2 1 1000000
R version 3.3.2 (2016-10-31)
Loading required package: Rcpp
DADA2 R package version: 1.2.1
1) Filtering .............
2) Learning Error Rates
2a) Forward Reads
Initial error matrix unspecified. Error rates will be initialized to the maximum possible estimate from this data.
Initializing error rates to maximum possible estimate.
Sample 1 - 346911 reads in 55363 unique sequences.
Sample 2 - 205941 reads in 64178 unique sequences.
Sample 3 - 346990 reads in 83357 unique sequences.
Sample 4 - 212410 reads in 55848 unique sequences.
selfConsist step 2
selfConsist step 3
selfConsist step 4
Convergence after 4 rounds.
2b) Reverse Reads
Initial error matrix unspecified. Error rates will be initialized to the maximum possible estimate from this data.
Initializing error rates to maximum possible estimate.
Sample 1 - 346911 reads in 24488 unique sequences.
Sample 2 - 205941 reads in 53880 unique sequences.
Sample 3 - 346990 reads in 59239 unique sequences.
Sample 4 - 212410 reads in 38235 unique sequences.
selfConsist step 2
selfConsist step 3
selfConsist step 4
selfConsist step 5
selfConsist step 6
Convergence after 6 rounds.
3) Denoise remaining samples Error in open.connection(con, "rb") : cannot open the connection
Calls: derepFastq ... FastqStreamer -> FastqStreamer -> open -> open.connection
In addition: Warning message:
In open.connection(con, "rb") :
cannot open file '/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/filt_f/16_S11_L001_R1_001.fastq.gz': No such file or directory
Execution halted
Traceback (most recent call last):
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/site-packages/q2_dada2-2017.2.0-py3.5.egg/q2_dada2/_denoise.py", line 154, in denoise_paired
run_commands([cmd])
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/site-packages/q2_dada2-2017.2.0-py3.5.egg/q2_dada2/_plot.py", line 26, in run_commands
subprocess.run(cmd, check=True)
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/subprocess.py", line 708, in run
output=stdout, stderr=stderr)
subprocess.CalledProcessError: Command '['run_dada_paired.R', '/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/forward', '/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/reverse', '/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/output.tsv.biom', '/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/filt_f', '/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o/filt_r', '230', '230', '10', '10', '2.0', '2', '1', '1000000']' returned non-zero exit status 1
During handling of the above exception, another exception occurred:
Traceback (most recent call last):
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/site-packages/q2cli-2017.2.0-py3.5.egg/q2cli/commands.py", line 217, in __call__
results = action(**arguments)
File "<decorator-gen-133>", line 2, in denoise_paired
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/site-packages/qiime2-2017.2.0-py3.5.egg/qiime2/sdk/action.py", line 171, in callable_wrapper
output_types, provenance)
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/site-packages/qiime2-2017.2.0-py3.5.egg/qiime2/sdk/action.py", line 248, in _callable_executor_
output_views = callable(**view_args)
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/site-packages/q2_dada2-2017.2.0-py3.5.egg/q2_dada2/_denoise.py", line 165, in denoise_paired
return _denoise_helper(biom_fp, hashed_feature_ids)
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/tempfile.py", line 808, in __exit__
self.cleanup()
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/tempfile.py", line 812, in cleanup
_shutil.rmtree(self.name)
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/shutil.py", line 488, in rmtree
return _rmtree_unsafe(path, onerror)
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/shutil.py", line 370, in _rmtree_unsafe
onerror(os.listdir, path, sys.exc_info())
File "/Users/apzlo/miniconda3/envs/qiime2-2017.2/lib/python3.5/shutil.py", line 368, in _rmtree_unsafe
names = os.listdir(path)
FileNotFoundError: [Errno 2] No such file or directory: '/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o'
Plugin error from dada2:
[Errno 2] No such file or directory:
'/var/folders/qr/vd5b6lqx18z8zzhc612cwk9r0000gn/T/tmphkndm05o'
See above for debug info.
I have no prior experience with dada2, I’m having trouble figuring where to start to solve this, I’d be happy to provide more info if needed.
Thank you very much,
Alejandro Pezzulo