multiple_split_libraries.py :: FastqParseError :: incorrect value for phred_offset

[Daniel Hwang]

Hi,

I am trying to run the Qiime script `multiple_split_libraries_fastq.py` on a directory of fastq files I created from a pandaseq run.

Here is my call to run `multiple_split_libraries_fastq.py`:

multiple_split_libraries_fastq.py -i ../pandatest/pandaseq_results2/panda_ea_util_contig -o split_output --demultiplexing_method sampleid_by_file

Error message:

Traceback (most recent call last):
  File "/cbcb/project2-scratch/nolson/miniconda2/bin/multiple_split_libraries_fastq.py", line 4, in <module>
    __import__('pkg_resources').run_script('qiime==1.9.1', 'multiple_split_libraries_fastq.py')
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/setuptools-20.3-py2.7.egg/pkg_resources/__init__.py", line 726, in run_script
    
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/setuptools-20.3-py2.7.egg/pkg_resources/__init__.py", line 1484, in run_script
    
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/qiime-1.9.1-py2.7.egg-info/scripts/multiple_split_libraries_fastq.py", line 219, in <module>
    main()
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/qiime-1.9.1-py2.7.egg-info/scripts/multiple_split_libraries_fastq.py", line 216, in main
    close_logger_on_success=True)
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/qiime/workflow/util.py", line 122, in call_commands_serially
    raise WorkflowError(msg)
qiime.workflow.util.WorkflowError: 
*** ERROR RAISED DURING STEP: split_libraries_fastq.py
Command run was:
 split_libraries_fastq.py  -i /cbcb/project2-scratch/nolson/pipe16S/pandatest/pandaseq_results2/panda_ea_util_contig/2-D10_S142_L001_pandaseq.fastq --sample_ids 2-D10 -o /cbcb/project2-scratch/nolson/pipe16S/split_libraries/split_output  --barcode_type 'not-barcoded'
Command returned exit status: 1
Stdout:
Stderr
Traceback (most recent call last):
  File "/cbcb/project2-scratch/nolson/miniconda2/bin/split_libraries_fastq.py", line 4, in <module>
    __import__('pkg_resources').run_script('qiime==1.9.1', 'split_libraries_fastq.py')
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/setuptools-20.3-py2.7.egg/pkg_resources/__init__.py", line 726, in run_script
    
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/setuptools-20.3-py2.7.egg/pkg_resources/__init__.py", line 1484, in run_script
    
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/qiime-1.9.1-py2.7.egg-info/scripts/split_libraries_fastq.py", line 365, in <module>
    main()
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/qiime-1.9.1-py2.7.egg-info/scripts/split_libraries_fastq.py", line 344, in main
    for fasta_header, sequence, quality, seq_id in seq_generator:
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/qiime/split_libraries_fastq.py", line 239, in process_fastq_single_end_read_file_no_barcode
    phred_offset=phred_offset):
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/qiime/split_libraries_fastq.py", line 317, in process_fastq_single_end_read_file
    parse_fastq(fastq_read_f, strict=False, phred_offset=phred_offset)):
  File "/cbcb/project2-scratch/nolson/miniconda2/lib/python2.7/site-packages/skbio/parse/sequences/fastq.py", line 174, in parse_fastq
    seqid)
skbio.parse.sequences._exception.FastqParseError: Failed qual conversion for seq id: M01556:61:000000000-ANR13:1:1101:13462:1621:142. This may be because you passed an incorrect value for phred_offset.

 

So it seems that the issue was with the `phred_offset` not being correctly set (as I saw in another question and answer thread).

Since `multiple_split_libraries_fastq.py` executes the `split_libraries_fastq.py` I am able to adjust the `phred_offset` flag in there. However, I am unable to set a `phred_offset` from the `multiple_split_libraries_fastq.py` command.

Is there a way to set the `phred_offset` flag from the `multiple_split_libraries.py` command?

[Daniel Hwang]

Just wanted to answer my own (boneheaded) question.

`multiple_split_libraries_fastq.py` has a parameter to accept in things like `phred_offset` through a parameter file.

-p, --parameter_fp

Path to the parameter file, which specifies changes to the default behavior of split_libraries_fastq.py. Seehttp://www.qiime.org/documentation/file_formats.html#qiime-parameters [default: split_libraries_fastq.py defaults will be used]

[justink]

Hey thanks for following up with the answer!

[Daniel Hwang]

Not a problem! I've really appreciated the community here as I have just started using QIIME at university!