Hej,
I have been running qiime2-2018.11 in a conda environment, now I installed qiime2-2019.4. I ran a very similar dataset succesfully (same illumina data, just different run) but with my new dataset I get stuck at demultiplexing.
Import works fine:
Imported rawfiles as MultiplexedPairedEndBarcodeInSequenceDirFmt to input.qza
But the following command:
qiime cutadapt demux-paired --i-seqs input.qza --m-forward-barcodes-file my-sample-metadata.tsv --m-forward-barcodes-column BarcodeSequence --p-error-rate 0.1 --o-per-sample-sequences demux_seqs.qza --o-untrimmed-sequences untrimmed.qza --verbose
Returns this error message:
Running external command line application. This may print messages to stdout and/or stderr.
The command being run is below. This command cannot be manually re-run as it will depend on temporary files that no longer exist.
Command: cutadapt --front file:/tmp/tmp4oqbe75g --error-rate 0.1 -o /tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.1.fastq.gz --untrimmed-output /tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/forward.fastq.gz -p /tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.2.fastq.gz --untrimmed-paired-output /tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/reverse.fastq.gz /tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/forward.fastq.gz /tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/reverse.fastq.gz
This is cutadapt 2.3 with Python 3.6.7
Command line parameters: --front file:/tmp/tmp4oqbe75g --error-rate 0.1 -o /tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.1.fastq.gz --untrimmed-output /tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/forward.fastq.gz -p /tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.2.fastq.gz --untrimmed-paired-output /tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/reverse.fastq.gz /tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/forward.fastq.gz /tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/reverse.fastq.gz
Processing reads on 1 core in paired-end mode …
Traceback (most recent call last):
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/xopen/init.py”, line 285, in _open_gz
return PipedGzipWriter(filename, mode, compresslevel, threads=threads)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/xopen/init.py”, line 117, in init
self.outfile = open(path, mode)
FileNotFoundError: [Errno 2] No such file or directory: ‘/tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/15054/055.1.fastq.gz’
During handling of the above exception, another exception occurred:
Traceback (most recent call last):
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/bin/cutadapt”, line 12, in
sys.exit(main())
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/cutadapt/main.py”, line 804, in main
stats = runner.run()
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/cutadapt/pipeline.py”, line 727, in run
(n, total1_bp, total2_bp) = self._pipeline.process_reads(progress=self.progress)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/cutadapt/pipeline.py”, line 327, in process_reads
if filter(read1, read2, matches1, matches2):
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/cutadapt/filters.py”, line 293, in call
self._demultiplexer1(read1, matches1)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/cutadapt/filters.py”, line 245, in call
mode=‘w’, qualities=self.qualities)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/dnaio/init.py”, line 83, in open
file1, fileformat=fileformat, mode=mode, qualities=qualities)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/dnaio/init.py”, line 112, in _open_single
file = xopen(file, mode + ‘b’)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/xopen/init.py”, line 337, in xopen
return _open_gz(filename, mode, compresslevel, threads)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/xopen/init.py”, line 287, in _open_gz
return buffered_writer(gzip.open(filename, mode, compresslevel=compresslevel))
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/gzip.py”, line 53, in open
binary_file = GzipFile(filename, gz_mode, compresslevel)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/gzip.py”, line 163, in init
fileobj = self.myfileobj = builtins.open(filename, mode or ‘rb’)
FileNotFoundError: [Errno 2] No such file or directory: ‘/tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/15054/055.1.fastq.gz’
Traceback (most recent call last):
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/q2cli/commands.py”, line 311, in call
results = action(**arguments)
File “</home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/decorator.py:decorator-gen-461>”, line 2, in demux_paired
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/qiime2/sdk/action.py”, line 231, in bound_callable
output_types, provenance)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/qiime2/sdk/action.py”, line 365, in callable_executor
output_views = self._callable(**view_args)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/q2_cutadapt/_demux.py”, line 193, in demux_paired
mux_fmt, batch_size)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/q2_cutadapt/_demux.py”, line 160, in _demux
run_command(cmd)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/site-packages/q2_cutadapt/_demux.py”, line 36, in run_command
subprocess.run(cmd, check=True)
File “/home/users/inga.martinek/miniconda3/envs/qiime2-2019.4/lib/python3.6/subprocess.py”, line 418, in run
output=stdout, stderr=stderr)
subprocess.CalledProcessError: Command ‘[‘cutadapt’, ‘–front’, ‘file:/tmp/tmp4oqbe75g’, ‘–error-rate’, ‘0.1’, ‘-o’, ‘/tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.1.fastq.gz’, ‘–untrimmed-output’, ‘/tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/forward.fastq.gz’, ‘-p’, ‘/tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.2.fastq.gz’, ‘–untrimmed-paired-output’, ‘/tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/reverse.fastq.gz’, ‘/tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/forward.fastq.gz’, ‘/tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/reverse.fastq.gz’]’ returned non-zero exit status 1.
Plugin error from cutadapt:
Command ‘[‘cutadapt’, ‘–front’, ‘file:/tmp/tmp4oqbe75g’, ‘–error-rate’, ‘0.1’, ‘-o’, ‘/tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.1.fastq.gz’, ‘–untrimmed-output’, ‘/tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/forward.fastq.gz’, ‘-p’, ‘/tmp/q2-CasavaOneEightSingleLanePerSampleDirFmt-qopsi_56/{name}.2.fastq.gz’, ‘–untrimmed-paired-output’, ‘/tmp/q2-MultiplexedPairedEndBarcodeInSequenceDirFmt-xqitwe7h/reverse.fastq.gz’, ‘/tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/forward.fastq.gz’, ‘/tmp/qiime2-archive-0r_c2gpp/92a999ed-44a7-403f-a5f2-d58dc6d173d5/data/reverse.fastq.gz’]’ returned non-zero exit status 1.
See above for debug info.
Since the main part of the error, I think, is a missing in a temporary directory, I do not even know where to begin really.
I tried the different versions, uplodaded and zipped unzipped the files several times in different ways, still the same error message…
I am sure it is something silly, but I would be very grateful if you could have a look at it.